1 32218221 We retrospectively assessed the characteristics and outcome of patients with R/R FLT3-mutated AML included in the Toulouse-Bordeaux DATAML registry. ('AML', 'Disease', (94, 97)) ('AML', 'Disease', 'MESH:D015470', (135, 138)) ('R/R', 'Var', (77, 80)) ('AML', 'Phenotype', 'HP:0004808', (94, 97)) ('AML', 'Phenotype', 'HP:0004808', (135, 138)) ('AML', 'Disease', (135, 138)) ('FLT3', 'Gene', '2322', (81, 85)) ('patients', 'Species', '9606', (63, 71)) ('AML', 'Disease', 'MESH:D015470', (94, 97)) ('FLT3', 'Gene', (81, 85)) 7 32218221 FLT3-activating mutations occur in approximately 30% of patients with acute myeloid leukemia (AML) and, as such, are among the most frequent mutations found in AML, either as in-frame internal tandem duplications (ITD) within the juxtamembrane region or as missense point mutations in the tyrosine kinase domain (TKD). ('patients', 'Species', '9606', (56, 64)) ('AML', 'Disease', (94, 97)) ('AML', 'Disease', 'MESH:D015470', (160, 163)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (70, 92)) ('AML', 'Phenotype', 'HP:0004808', (94, 97)) ('mutations', 'Var', (16, 25)) ('AML', 'Disease', (160, 163)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (76, 92)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (70, 92)) ('AML', 'Phenotype', 'HP:0004808', (160, 163)) ('missense point mutations', 'Var', (257, 281)) ('acute myeloid leukemia', 'Disease', (70, 92)) ('FLT3', 'Gene', '2322', (0, 4)) ('AML', 'Disease', 'MESH:D015470', (94, 97)) ('leukemia', 'Phenotype', 'HP:0001909', (84, 92)) ('FLT3', 'Gene', (0, 4)) 8 32218221 FLT3-ITD (but not FLT3-TKD) mutations confer a poor prognosis in AML, especially when NPM1 is not co-mutated and the allelic FLT3-ITD/wild-type ratio is high; such mutations are usually conserved at relapse and have therefore emerged as a relevant therapeutic target. ('AML', 'Disease', (65, 68)) ('AML', 'Phenotype', 'HP:0004808', (65, 68)) ('FLT3', 'Gene', '2322', (125, 129)) ('FLT3', 'Gene', (18, 22)) ('NPM1', 'Gene', '4869', (86, 90)) ('FLT3', 'Gene', (125, 129)) ('AML', 'Disease', 'MESH:D015470', (65, 68)) ('FLT3', 'Gene', '2322', (0, 4)) ('NPM1', 'Gene', (86, 90)) ('FLT3', 'Gene', '2322', (18, 22)) ('FLT3', 'Gene', (0, 4)) ('mutations', 'Var', (28, 37)) 13 32218221 In the ADMIRAL trial, overall survival (OS) was significantly improved in the gilteritinib arm compared to the control arm with a hazard ratio (HR) of 0.64 (95%CI: 0.49-0.83; p < 0.001). ('improved', 'PosReg', (62, 70)) ('overall survival', 'MPA', (22, 38)) ('gilteritinib', 'Var', (78, 90)) ('gilteritinib', 'Chemical', 'MESH:C000609080', (78, 90)) 16 32218221 The aim of our study was to describe the characteristics, treatments and outcome of R/R FLT3-mutated AML patients treated in a routine setting before the approval of second-generation FLT3 inhibitors in this indication. ('R/R', 'Var', (84, 87)) ('AML', 'Disease', 'MESH:D015470', (101, 104)) ('patients', 'Species', '9606', (105, 113)) ('FLT3', 'Gene', '2322', (184, 188)) ('FLT3', 'Gene', '2322', (88, 92)) ('FLT3', 'Gene', (184, 188)) ('AML', 'Phenotype', 'HP:0004808', (101, 104)) ('AML', 'Disease', (101, 104)) ('FLT3', 'Gene', (88, 92)) 31 32218221 Out of 3290 newly diagnosed AML patients included in the DATAML registry between 2000 and 2017, 1453 did not have a recorded status for FLT3 mutation and 364 were not selected to receive intensive chemotherapy as a first-line treatment. ('AML', 'Phenotype', 'HP:0004808', (28, 31)) ('patients', 'Species', '9606', (32, 40)) ('AML', 'Disease', 'MESH:D015470', (60, 63)) ('AML', 'Disease', (28, 31)) ('FLT3', 'Gene', '2322', (136, 140)) ('AML', 'Phenotype', 'HP:0004808', (60, 63)) ('AML', 'Disease', 'MESH:D015470', (28, 31)) ('AML', 'Disease', (60, 63)) ('FLT3', 'Gene', (136, 140)) ('mutation', 'Var', (141, 149)) 32 32218221 A total of 347 patients with FLT3-ITD (n = 317) or FLT3-TKD (n = 39) mutated AML fulfilled the inclusion criteria (Figure S1). ('patients', 'Species', '9606', (15, 23)) ('AML', 'Disease', 'MESH:D015470', (77, 80)) ('FLT3', 'Gene', '2322', (29, 33)) ('mutated', 'Var', (69, 76)) ('AML', 'Disease', (77, 80)) ('FLT3', 'Gene', (29, 33)) ('FLT3', 'Gene', '2322', (51, 55)) ('AML', 'Phenotype', 'HP:0004808', (77, 80)) ('FLT3', 'Gene', (51, 55)) 53 32218221 The characteristics of these 174 R/R FLT3-mutated AML patients are shown in Table 3. ('R/R', 'Var', (33, 36)) ('AML', 'Disease', 'MESH:D015470', (50, 53)) ('patients', 'Species', '9606', (54, 62)) ('FLT3', 'Gene', '2322', (37, 41)) ('AML', 'Phenotype', 'HP:0004808', (50, 53)) ('AML', 'Disease', (50, 53)) ('FLT3', 'Gene', (37, 41)) 74 32218221 Here, we examined the characteristics and outcome of R/R FLT3-mutated AML patients included in the Toulouse-Bordeaux DATAML registry, in order to evaluate the efficacy of standard treatments that appeared to have limited efficacy in the ADMIRAL study. ('AML', 'Disease', (70, 73)) ('R/R', 'Var', (53, 56)) ('AML', 'Phenotype', 'HP:0004808', (70, 73)) ('patients', 'Species', '9606', (74, 82)) ('FLT3', 'Gene', '2322', (57, 61)) ('AML', 'Disease', 'MESH:D015470', (120, 123)) ('AML', 'Disease', (120, 123)) ('FLT3', 'Gene', (57, 61)) ('AML', 'Disease', 'MESH:D015470', (70, 73)) ('AML', 'Phenotype', 'HP:0004808', (120, 123)) 85 32218221 In the ADMIRAL study, 30-day and 60-day mortality were 2.0% and 7.7% in the gilteritinib arm compared to 10.2% and 19.0% in the control arm or 6.1% and 14.9% in our study. ('mortality', 'Disease', 'MESH:D003643', (40, 49)) ('gilteritinib', 'Chemical', 'MESH:C000609080', (76, 88)) ('gilteritinib', 'Var', (76, 88)) ('mortality', 'Disease', (40, 49)) 86 32218221 Given the underperforming results of second-line therapeutic strategies, the treatment of R/R FLT3-mutated AML patients remains a major challenge. ('FLT3', 'Gene', '2322', (94, 98)) ('AML', 'Phenotype', 'HP:0004808', (107, 110)) ('AML', 'Disease', (107, 110)) ('FLT3', 'Gene', (94, 98)) ('R/R', 'Var', (90, 93)) ('AML', 'Disease', 'MESH:D015470', (107, 110)) ('patients', 'Species', '9606', (111, 119)) 89 32218221 Obviously, combinations of targeted therapies such as FLT3 inhibitors and BCL2 inhibitors, for example, are also promising options for R/R FLT3-mutated AML patients. ('patients', 'Species', '9606', (156, 164)) ('FLT3', 'Gene', '2322', (139, 143)) ('FLT3', 'Gene', '2322', (54, 58)) ('AML', 'Disease', 'MESH:D015470', (152, 155)) ('AML', 'Disease', (152, 155)) ('AML', 'Phenotype', 'HP:0004808', (152, 155)) ('FLT3', 'Gene', (139, 143)) ('FLT3', 'Gene', (54, 58)) ('BCL2', 'Gene', '596', (74, 78)) ('BCL2', 'Gene', (74, 78)) ('R/R', 'Var', (135, 138)) 106 31824191 Recent studies have implicated epigenetic changes such as abnormal DNA methylation, histone modification, and non-coding RNA in the pathogenesis of MDS. ('abnormal', 'Var', (58, 66)) ('MDS', 'Disease', (148, 151)) ('MDS', 'Disease', 'MESH:D009190', (148, 151)) ('MDS', 'Phenotype', 'HP:0002863', (148, 151)) ('DNA', 'Chemical', 'MESH:D000596', (67, 70)) ('non-coding RNA', 'Var', (110, 124)) ('histone modification', 'MPA', (84, 104)) 155 31824191 However, the de novo incidence of respiratory infection was 22.5% in Dec group as against 53% in the non-Dec group (P=0.006) (shown in Table 2 - Incidence of transplantation-related complications). ('respiratory infection', 'Disease', 'MESH:D012141', (34, 55)) ('Dec', 'Chemical', 'MESH:C014347', (105, 108)) ('respiratory infection', 'Phenotype', 'HP:0011947', (34, 55)) ('Dec', 'Var', (69, 72)) ('respiratory infection', 'Disease', (34, 55)) ('Dec', 'Chemical', 'MESH:C014347', (69, 72)) 161 31824191 Results of univariate analyses showed that recipients older than 40 years may be more susceptible to grade II to IV aGvHD (P=0.018) (Table 3 Univariate analysis). ('GvHD', 'Disease', 'MESH:D006086', (117, 121)) ('grade II', 'Var', (101, 109)) ('GvHD', 'Disease', (117, 121)) ('susceptible', 'Reg', (86, 97)) 168 31824191 However, there were only 2 patients in the Dec group and none of the patients in the non-Dec group developed no relapse death in the competing risk model; therefore, we cannot draw the conclusion that Dec increases NRM in higher-risk patients. ('Dec', 'Chemical', 'MESH:C014347', (201, 204)) ('death', 'Disease', 'MESH:D003643', (120, 125)) ('death', 'Disease', (120, 125)) ('Dec', 'Chemical', 'MESH:C014347', (43, 46)) ('NRM', 'Disease', (215, 218)) ('Dec', 'Chemical', 'MESH:C014347', (89, 92)) ('patients', 'Species', '9606', (27, 35)) ('Dec', 'Var', (201, 204)) ('patients', 'Species', '9606', (234, 242)) ('increases', 'PosReg', (205, 214)) ('patients', 'Species', '9606', (69, 77)) 180 31824191 In a study by Field et al, bridge therapy with 5-Aza prior to allo-HSCT achieved a lower relapse rate as compared to that with traditional chemotherapy (20% vs 32%). ('relapse', 'CPA', (89, 96)) ('5-Aza', 'Var', (47, 52)) ('5-Aza', 'Chemical', 'MESH:D001372', (47, 52)) ('lower', 'NegReg', (83, 88)) 188 31824191 Finally, Dec alleviates tissue damage by inhibiting the production of proinflammatory cytokines, such as interferon gamma and tumor necrosis factor beta. ('production of proinflammatory cytokines', 'MPA', (56, 95)) ('Dec', 'Chemical', 'MESH:C014347', (9, 12)) ('necrosis', 'Disease', (132, 140)) ('tumor', 'Phenotype', 'HP:0002664', (126, 131)) ('inhibiting', 'NegReg', (41, 51)) ('tumor', 'Disease', 'MESH:D009369', (126, 131)) ('interferon gamma', 'MPA', (105, 121)) ('alleviates', 'NegReg', (13, 23)) ('tissue damage', 'CPA', (24, 37)) ('necrosis', 'Disease', 'MESH:D009336', (132, 140)) ('Dec', 'Var', (9, 12)) ('tumor', 'Disease', (126, 131)) 192 31824191 In summary, Dec exerts its antitumor effects via direct cytotoxicity and strengthened tumor immunity, without sacrificing its GvHD protective effects. ('tumor', 'Phenotype', 'HP:0002664', (86, 91)) ('tumor', 'Phenotype', 'HP:0002664', (31, 36)) ('Dec', 'Chemical', 'MESH:C014347', (12, 15)) ('cytotoxicity', 'Disease', 'MESH:D064420', (56, 68)) ('tumor', 'Disease', (86, 91)) ('tumor', 'Disease', (31, 36)) ('strengthened', 'PosReg', (73, 85)) ('Dec', 'Var', (12, 15)) ('GvHD', 'Disease', (126, 130)) ('cytotoxicity', 'Disease', (56, 68)) ('tumor', 'Disease', 'MESH:D009369', (86, 91)) ('tumor', 'Disease', 'MESH:D009369', (31, 36)) ('GvHD', 'Disease', 'MESH:D006086', (126, 130)) 198 31757221 Most studies have suggested that m6A modification can affect the complexity of cancer progression by regulating biological functions related to cancer. ('complexity', 'MPA', (65, 75)) ('biological functions', 'MPA', (112, 132)) ('regulating', 'Reg', (101, 111)) ('modification', 'Var', (37, 49)) ('cancer', 'Phenotype', 'HP:0002664', (79, 85)) ('m6A', 'Protein', (33, 36)) ('cancer', 'Disease', 'MESH:D009369', (144, 150)) ('cancer', 'Disease', (144, 150)) ('affect', 'Reg', (54, 60)) ('cancer', 'Disease', (79, 85)) ('cancer', 'Disease', 'MESH:D009369', (79, 85)) ('cancer', 'Phenotype', 'HP:0002664', (144, 150)) 199 31757221 M6A modification of noncoding RNAs regulates the cleavage, transport, stability, and degradation of noncoding RNAs themselves. ('transport', 'MPA', (59, 68)) ('M6A', 'Gene', '56339', (0, 3)) ('stability', 'MPA', (70, 79)) ('M6A', 'Gene', (0, 3)) ('degradation', 'MPA', (85, 96)) ('cleavage', 'MPA', (49, 57)) ('modification', 'Var', (4, 16)) ('regulates', 'Reg', (35, 44)) 201 31757221 Additionally, it is becoming increasingly clear that m6A and noncoding RNAs potentially contribute to the clinical application of cancer treatment. ('contribute', 'Reg', (88, 98)) ('cancer', 'Disease', 'MESH:D009369', (130, 136)) ('cancer', 'Disease', (130, 136)) ('m6A', 'Var', (53, 56)) ('cancer', 'Phenotype', 'HP:0002664', (130, 136)) ('noncoding RNAs', 'Var', (61, 75)) 202 31757221 In this review, we summarize the effect of the interactions between m6A modifications and noncoding RNAs on the biological functions involved in cancer progression. ('cancer', 'Disease', (145, 151)) ('modifications', 'Var', (72, 85)) ('interactions', 'Interaction', (47, 59)) ('cancer', 'Phenotype', 'HP:0002664', (145, 151)) ('m6A', 'Gene', (68, 71)) ('cancer', 'Disease', 'MESH:D009369', (145, 151)) 203 31757221 In particular, we discuss the role of m6A and noncoding RNAs as possible potential biomarkers and therapeutic targets in the treatment of cancers. ('cancer', 'Phenotype', 'HP:0002664', (138, 144)) ('cancers', 'Disease', (138, 145)) ('cancers', 'Phenotype', 'HP:0002664', (138, 145)) ('noncoding RNAs', 'Var', (46, 60)) ('cancers', 'Disease', 'MESH:D009369', (138, 145)) 206 31757221 The mRNAs of 7676 mammalian genes were found to be modified by m6A. ('m6A', 'Var', (63, 66)) ('modified', 'Reg', (51, 59)) ('mRNAs', 'MPA', (4, 9)) ('mammalian', 'Species', '9606', (18, 27)) 208 31757221 M6A modifications occur via the m6A methyltransferases called "writers"; they are removed by the demethylases called "erasers" and are recognized by m6A-binding proteins called "readers". ('M6A', 'Gene', '56339', (0, 3)) ('m6A methyltransferases', 'Enzyme', (32, 54)) ('modifications', 'Var', (4, 17)) ('M6A', 'Gene', (0, 3)) 209 31757221 M6A modifications are quite prevalent, and the dynamic regulation of m6A modifications has been shown to be significantly related to gene expression. ('M6A', 'Gene', '56339', (0, 3)) ('modifications', 'Var', (4, 17)) ('M6A', 'Gene', (0, 3)) 215 31757221 Thus, concomitant targeting of m6A and noncoding RNAs may provide a synergistic effect in cancer therapy. ('cancer', 'Phenotype', 'HP:0002664', (90, 96)) ('m6A', 'Protein', (31, 34)) ('noncoding RNAs', 'Var', (39, 53)) ('cancer', 'Disease', (90, 96)) ('cancer', 'Disease', 'MESH:D009369', (90, 96)) 216 31757221 In this review, we summarize the interactive effects of m6A methylation and noncoding RNAs and describe the way their association influences biological functions in cancer and their possible uses in future clinical applications. ('cancer', 'Phenotype', 'HP:0002664', (165, 171)) ('methylation', 'Var', (60, 71)) ('cancer', 'Disease', (165, 171)) ('association', 'Interaction', (118, 129)) ('cancer', 'Disease', 'MESH:D009369', (165, 171)) ('biological functions', 'MPA', (141, 161)) ('influences', 'Reg', (130, 140)) ('m6A', 'Protein', (56, 59)) ('noncoding', 'Protein', (76, 85)) 222 31757221 Recently, an increasing number of studies have explored the control of mRNA metabolism by m6A modifications, showing dual characteristics of m6A modifications and the related regulators in cancers, such as leukemia, lung cancer, pancreatic carcinoma, glioblastoma, and hepatoma (Table 1). ('leukemia', 'Phenotype', 'HP:0001909', (206, 214)) ('glioblastoma', 'Disease', (251, 263)) ('modifications', 'Var', (145, 158)) ('glioblastoma', 'Phenotype', 'HP:0012174', (251, 263)) ('cancers', 'Disease', 'MESH:D009369', (189, 196)) ('lung cancer', 'Disease', 'MESH:D008175', (216, 227)) ('hepatoma', 'Disease', (269, 277)) ('leukemia', 'Disease', 'MESH:D007938', (206, 214)) ('lung cancer', 'Phenotype', 'HP:0100526', (216, 227)) ('leukemia', 'Disease', (206, 214)) ('pancreatic carcinoma', 'Disease', 'MESH:C562463', (229, 249)) ('pancreatic carcinoma', 'Disease', (229, 249)) ('modifications', 'Var', (94, 107)) ('cancer', 'Phenotype', 'HP:0002664', (221, 227)) ('cancers', 'Phenotype', 'HP:0002664', (189, 196)) ('cancers', 'Disease', (189, 196)) ('lung cancer', 'Disease', (216, 227)) ('cancer', 'Phenotype', 'HP:0002664', (189, 195)) ('hepatoma', 'Disease', 'MESH:D006528', (269, 277)) ('glioblastoma', 'Disease', 'MESH:D005909', (251, 263)) ('carcinoma', 'Phenotype', 'HP:0030731', (240, 249)) 223 31757221 M6A modifications not only affect miRNA, lncRNA, and circRNA cleavage, transport, stability, and degradation processes but also regulate biological cell functions by affecting noncoding RNA expression. ('transport', 'MPA', (71, 80)) ('affecting', 'Reg', (166, 175)) ('M6A', 'Gene', '56339', (0, 3)) ('biological cell functions', 'MPA', (137, 162)) ('miRNA', 'MPA', (34, 39)) ('M6A', 'Gene', (0, 3)) ('circRNA cleavage', 'MPA', (53, 69)) ('degradation processes', 'MPA', (97, 118)) ('lncRNA', 'MPA', (41, 47)) ('noncoding RNA expression', 'MPA', (176, 200)) ('regulate', 'Reg', (128, 136)) ('stability', 'MPA', (82, 91)) ('modifications', 'Var', (4, 17)) ('affect', 'Reg', (27, 33)) 226 31757221 Further experiments suggested that METTL3 can promote miRNA maturation, indicating that these m6A modifications enable DGCR8 to target pri-miRNAs and facilitate miRNA maturation (Fig. ('miRNA', 'MPA', (54, 59)) ('DGCR8', 'Gene', (119, 124)) ('METTL3', 'Gene', '56339', (35, 41)) ('miRNA maturation', 'CPA', (161, 177)) ('modifications', 'Var', (98, 111)) ('METTL3', 'Gene', (35, 41)) ('target pri-miRNAs', 'MPA', (128, 145)) ('facilitate', 'PosReg', (150, 160)) ('DGCR8', 'Gene', '54487', (119, 124)) 227 31757221 METTL3 was also found to indirectly regulate miRNA expression by methylating the hepatitis B virus X-interacting protein (HBXIP). ('HBXIP', 'Gene', '10542', (122, 127)) ('hepatitis B virus X-interacting protein', 'Gene', (81, 120)) ('METTL3', 'Gene', '56339', (0, 6)) ('HBXIP', 'Gene', (122, 127)) ('methylating', 'Var', (65, 76)) ('METTL3', 'Gene', (0, 6)) ('miRNA expression', 'MPA', (45, 61)) ('hepatitis', 'Phenotype', 'HP:0012115', (81, 90)) ('regulate', 'Reg', (36, 44)) ('hepatitis B virus X-interacting protein', 'Gene', '10542', (81, 120)) 229 31757221 METTL14 depletion leads to miR-126 downregulation and pri-miRNA accumulation. ('depletion', 'Var', (8, 17)) ('METTL14', 'Gene', '57721', (0, 7)) ('METTL14', 'Gene', (0, 7)) ('miR-126', 'Gene', '406913', (27, 34)) ('miR-126', 'Gene', (27, 34)) ('pri-miRNA accumulation', 'CPA', (54, 76)) ('downregulation', 'NegReg', (35, 49)) 232 31757221 In this study, Zang indicated the effect of cigarette smoking on m6A-modified miR-25-3p maturation in pancreatic ductal adenocarcinoma. ('carcinoma', 'Phenotype', 'HP:0030731', (125, 134)) ('miR-25-3p', 'Gene', (78, 87)) ('m6A-modified', 'Var', (65, 77)) ('pancreatic ductal adenocarcinoma', 'Disease', (102, 134)) ('pancreatic ductal adenocarcinoma', 'Phenotype', 'HP:0006725', (102, 134)) ('pancreatic ductal adenocarcinoma', 'Disease', 'MESH:D021441', (102, 134)) 234 31757221 This epigenetic modification causes an increase in the METTL3-dependent regulation of pri-miR-25-3p levels and enhances the processing of miR-25-3p. ('epigenetic modification', 'Var', (5, 28)) ('regulation of pri-miR-25-3p levels', 'MPA', (72, 106)) ('METTL3', 'Gene', '56339', (55, 61)) ('increase', 'PosReg', (39, 47)) ('processing of miR-25-3p', 'MPA', (124, 147)) ('METTL3', 'Gene', (55, 61)) ('enhances', 'PosReg', (111, 119)) 235 31757221 Then, the AKT-p70S6K signaling pathway is influenced after miR-25-3p affects the progression of its target: PH domain leucine-rich repeat protein phosphatase 2 (PHLPP2). ('PH domain leucine-rich repeat protein phosphatase 2', 'Gene', '23035', (108, 159)) ('AKT', 'Gene', (10, 13)) ('miR-25-3p', 'Var', (59, 68)) ('PHLPP2', 'Gene', (161, 167)) ('influenced', 'Reg', (42, 52)) ('p70S6K', 'Gene', '6198', (14, 20)) ('p70S6K', 'Gene', (14, 20)) ('AKT', 'Gene', '207', (10, 13)) ('PHLPP2', 'Gene', '23035', (161, 167)) ('affects', 'Reg', (69, 76)) 241 31757221 Furthermore, the regulation of miRNAs by m6A modifications also contributes to endocrine resistance in breast cancer cells. ('contributes', 'Reg', (64, 75)) ('endocrine resistance', 'MPA', (79, 99)) ('regulation', 'Reg', (17, 27)) ('modifications', 'Var', (45, 58)) ('breast cancer', 'Disease', 'MESH:D001943', (103, 116)) ('cancer', 'Phenotype', 'HP:0002664', (110, 116)) ('breast cancer', 'Disease', (103, 116)) ('miRNAs', 'Protein', (31, 37)) ('m6A', 'Protein', (41, 44)) ('breast cancer', 'Phenotype', 'HP:0003002', (103, 116)) 242 31757221 HNRNPA2/B1, as a "reader," can recognize m6A methylation. ('HNRNPA2/B1', 'Gene', '3181', (0, 10)) ('HNRNPA2/B1', 'Gene', (0, 10)) ('methylation', 'Var', (45, 56)) ('m6A', 'Var', (41, 44)) 244 31757221 HNRNPA2/B1 overexpression promotes increases in m6A-modified miRNA levels by enhancing the process by which pri-miRNAs develop into pre-miRNAs and mature miRNAs. ('increases', 'PosReg', (35, 44)) ('overexpression', 'Var', (11, 25)) ('HNRNPA2/B1', 'Gene', (0, 10)) ('m6A-modified miRNA levels', 'MPA', (48, 73)) ('HNRNPA2/B1', 'Gene', '3181', (0, 10)) ('enhancing', 'PosReg', (77, 86)) 245 31757221 Knocking down HNRNPA2/B1 can inhibit breast cancer cell proliferation and promote tumor cell apoptosis. ('breast cancer', 'Disease', (37, 50)) ('breast cancer', 'Phenotype', 'HP:0003002', (37, 50)) ('Knocking down', 'Var', (0, 13)) ('breast cancer', 'Disease', 'MESH:D001943', (37, 50)) ('promote', 'PosReg', (74, 81)) ('tumor', 'Disease', 'MESH:D009369', (82, 87)) ('inhibit', 'NegReg', (29, 36)) ('HNRNPA2/B1', 'Gene', (14, 24)) ('tumor', 'Phenotype', 'HP:0002664', (82, 87)) ('tumor', 'Disease', (82, 87)) ('cancer', 'Phenotype', 'HP:0002664', (44, 50)) ('HNRNPA2/B1', 'Gene', '3181', (14, 24)) 247 31757221 Then, m6A methylation results in TARBP2-bound transcripts intron retention and nuclear RNA decay. ('results in', 'Reg', (22, 32)) ('TARBP2', 'Gene', '6895', (33, 39)) ('methylation', 'Var', (10, 21)) ('TARBP2', 'Gene', (33, 39)) ('intron retention', 'CPA', (58, 74)) ('nuclear RNA decay', 'CPA', (79, 96)) ('m6A methylation', 'Var', (6, 21)) 250 31757221 Thus, TARBP2-mediated m6A modification may regulate miRNA processes, such as miRNA integration, maturation, and degradation; however, this hypothesis needs to be confirmed by further research. ('m6A', 'Protein', (22, 25)) ('TARBP2', 'Gene', (6, 12)) ('regulate', 'Reg', (43, 51)) ('TARBP2', 'Gene', '6895', (6, 12)) ('modification', 'Var', (26, 38)) ('miRNA processes', 'MPA', (52, 67)) ('miRNA integration', 'MPA', (77, 94)) ('degradation', 'CPA', (112, 123)) ('maturation', 'CPA', (96, 106)) 251 31757221 In summary, miRNA regulation by m6A modifications has an important effect on cancer progression. ('m6A', 'Protein', (32, 35)) ('miRNA regulation', 'MPA', (12, 28)) ('cancer', 'Disease', (77, 83)) ('cancer', 'Disease', 'MESH:D009369', (77, 83)) ('effect', 'Reg', (67, 73)) ('modifications', 'Var', (36, 49)) ('cancer', 'Phenotype', 'HP:0002664', (77, 83)) 255 31757221 In addition, knocking down METTL3 or RBM15 reduced the level of m6A modifications on specific transcripts such that the lncRNA X chromosome was inactivated. ('knocking down', 'Var', (13, 26)) ('RBM15', 'Gene', (37, 42)) ('METTL3', 'Gene', '56339', (27, 33)) ('m6A modifications on specific transcripts', 'MPA', (64, 105)) ('METTL3', 'Gene', (27, 33)) ('level', 'MPA', (55, 60)) ('reduced', 'NegReg', (43, 50)) ('RBM15', 'Gene', '64783', (37, 42)) 256 31757221 Furthermore, YTHDC1 recognized the m6A residues on XIST and prompted subsequent mediation of the regulation initiated by lncRNA-induced gene silencing. ('m6A', 'Var', (35, 38)) ('YTHDC1', 'Gene', '91746', (13, 19)) ('XIST', 'Gene', '7503', (51, 55)) ('XIST', 'Gene', (51, 55)) ('YTHDC1', 'Gene', (13, 19)) 258 31757221 ALKBH5 can induce high levels of FOXM1 transcripts by demethylating FOXM1 nascent transcripts. ('ALKBH5', 'Gene', (0, 6)) ('demethylating', 'Var', (54, 67)) ('nascent transcripts', 'MPA', (74, 93)) ('FOXM1', 'Gene', (33, 38)) ('FOXM1', 'Gene', '2305', (33, 38)) ('FOXM1', 'Gene', '2305', (68, 73)) ('FOXM1', 'Gene', (68, 73)) ('ALKBH5', 'Gene', '54890', (0, 6)) 262 31757221 MALAT1 contains a series of m6A modifications and is upregulated in neoplastic diseases. ('neoplastic diseases', 'Disease', (68, 87)) ('m6A', 'Var', (28, 31)) ('MALAT1', 'Gene', '378938', (0, 6)) ('MALAT1', 'Gene', (0, 6)) ('neoplastic diseases', 'Disease', 'MESH:D009386', (68, 87)) ('upregulated', 'PosReg', (53, 64)) 263 31757221 Studies have confirmed that MALAT1 undergoes structural changes due to m6A modifications, which regulate the interaction between RNAs and some special binding proteins. ('MALAT1', 'Gene', (28, 34)) ('binding', 'Interaction', (151, 158)) ('structural', 'MPA', (45, 55)) ('regulate', 'Reg', (96, 104)) ('m6A modifications', 'Var', (71, 88)) ('RNAs', 'Protein', (129, 133)) ('interaction', 'Interaction', (109, 120)) ('MALAT1', 'Gene', '378938', (28, 34)) 264 31757221 In addition, m6A modifications of MALAT1 can also affect its localization and activity in the nucleus. ('MALAT1', 'Gene', (34, 40)) ('affect', 'Reg', (50, 56)) ('MALAT1', 'Gene', '378938', (34, 40)) ('activity in the nucleus', 'MPA', (78, 101)) ('m6A modifications', 'Var', (13, 30)) ('localization', 'MPA', (61, 73)) 265 31757221 M6A modifications of MALAT1 promote the binding of HNRNPG, HNRNPC, or METTL16 to transcripts, which in turn regulates gene expression. ('MALAT1', 'Gene', '378938', (21, 27)) ('METTL16', 'Gene', (70, 77)) ('M6A', 'Gene', '56339', (0, 3)) ('MALAT1', 'Gene', (21, 27)) ('METTL16', 'Gene', '79066', (70, 77)) ('binding', 'Interaction', (40, 47)) ('M6A', 'Gene', (0, 3)) ('promote', 'PosReg', (28, 35)) ('HNRNPG', 'Gene', (51, 57)) ('HNRNPC', 'Gene', (59, 65)) ('HNRNPG', 'Gene', '27316', (51, 57)) ('regulates', 'Reg', (108, 117)) ('modifications', 'Var', (4, 17)) ('gene expression', 'MPA', (118, 133)) 266 31757221 It was revealed that altering the m6A modification level of lncRNA 1281 can significantly affect let-7 levels, thereby influencing ESC differentiation. ('affect', 'Reg', (90, 96)) ('ESC', 'CellLine', 'CVCL:M564', (131, 134)) ('m6A modification', 'Var', (34, 50)) ('let-7 levels', 'MPA', (97, 109)) ('lncRNA 1281', 'Gene', (60, 71)) ('ESC', 'Disease', (131, 134)) ('influencing', 'Reg', (119, 130)) 270 31757221 Among these regulators, METTL3 has an important role, as indicated by its specific performance when knocked down; that is, METTL3 downregulation reduces the expression of Olfr29-ps1 in MDSCs, indicating that m6A modifications induce Olfr29-ps1 formation and stability. ('METTL3', 'Gene', '56339', (123, 129)) ('METTL3', 'Gene', (123, 129)) ('expression', 'MPA', (157, 167)) ('ps1', 'Gene', (240, 243)) ('METTL3', 'Gene', (24, 30)) ('induce', 'PosReg', (226, 232)) ('stability', 'MPA', (258, 267)) ('downregulation reduces', 'NegReg', (130, 152)) ('ps1', 'Gene', '338399', (178, 181)) ('ps1', 'Gene', '338399', (240, 243)) ('ps1', 'Gene', (178, 181)) ('modifications', 'Var', (212, 225)) ('METTL3', 'Gene', '56339', (24, 30)) 271 31757221 Then, m6A modifications of Olfr29-ps1 prompt a functional interaction between Olfr29-ps1 and miR-214-3p by recruiting Olfr29-ps1 into the Ago2-related RNA complex. ('ps1', 'Gene', (34, 37)) ('ps1', 'Gene', '338399', (125, 128)) ('miR-214', 'Gene', (93, 100)) ('interaction', 'Interaction', (58, 69)) ('Ago2', 'Gene', '27161', (138, 142)) ('miR-214', 'Gene', '406996', (93, 100)) ('ps1', 'Gene', (125, 128)) ('ps1', 'Gene', '338399', (85, 88)) ('Ago2', 'Gene', (138, 142)) ('recruiting', 'PosReg', (107, 117)) ('ps1', 'Gene', '338399', (34, 37)) ('ps1', 'Gene', (85, 88)) ('modifications', 'Var', (10, 23)) ('m6A', 'Var', (6, 9)) 272 31757221 Finally, MyD88 is regulated by miR-214-3p, and knocking down MyD88 exerts an important effect on MDSC immunosuppression and differentiation. ('effect', 'Reg', (87, 93)) ('miR-214', 'Gene', '406996', (31, 38)) ('differentiation', 'CPA', (124, 139)) ('MyD88', 'Gene', (9, 14)) ('MyD88', 'Gene', '4615', (61, 66)) ('MyD88', 'Gene', '4615', (9, 14)) ('knocking down', 'Var', (47, 60)) ('MyD88', 'Gene', (61, 66)) ('miR-214', 'Gene', (31, 38)) ('MDSC immunosuppression', 'CPA', (97, 119)) 273 31757221 In conclusion, Olfr29-ps1 relies mainly on the m6A-modified Olfr29-ps1/miR-214-3p/MyD88 regulatory pathway to modulate MDSC immunosuppression and differentiation. ('ps1', 'Gene', '338399', (22, 25)) ('ps1', 'Gene', '338399', (67, 70)) ('m6A-modified', 'Var', (47, 59)) ('miR-214', 'Gene', (71, 78)) ('ps1', 'Gene', (67, 70)) ('differentiation', 'CPA', (146, 161)) ('ps1', 'Gene', (22, 25)) ('MyD88', 'Gene', (82, 87)) ('miR-214', 'Gene', '406996', (71, 78)) ('MyD88', 'Gene', '4615', (82, 87)) ('modulate', 'Reg', (110, 118)) 274 31757221 The study also revealed that circRNAs have m6A modifications, which are regulated by the demethylase FTO and the METTL3/14 methyltransferase complex. ('FTO', 'Gene', (101, 104)) ('METTL3', 'Gene', (113, 119)) ('m6A modifications', 'Var', (43, 60)) ('FTO', 'Gene', '79068', (101, 104)) ('modifications', 'Var', (47, 60)) ('METTL3', 'Gene', '56339', (113, 119)) 275 31757221 Further studies have shown that m6A modifications of circRNAs can promote the translation processes of circRNAs by mediating the eukaryotic translation initiation factor 4G2 (eIF4G2) and binding protein YTHDF3 (Fig. ('YTHDF3', 'Gene', (203, 209)) ('eIF4G2', 'Gene', '1982', (175, 181)) ('translation processes', 'MPA', (78, 99)) ('eukaryotic translation initiation factor 4G2', 'Gene', '1982', (129, 173)) ('eukaryotic translation initiation factor 4G2', 'Gene', (129, 173)) ('promote', 'PosReg', (66, 73)) ('eIF4G2', 'Gene', (175, 181)) ('m6A modifications', 'Var', (32, 49)) ('binding', 'Interaction', (187, 194)) ('YTHDF3', 'Gene', '253943', (203, 209)) 276 31757221 demonstrated that m6A modifications of human endogenous circRNAs exerted the important function of inhibiting innate immunity. ('m6A modifications', 'Var', (18, 35)) ('human', 'Species', '9606', (39, 44)) ('innate immunity', 'CPA', (110, 125)) ('inhibiting', 'NegReg', (99, 109)) 277 31757221 The authors also revealed that exogenous circRNAs could induce antigen-specific T and B cell activation, antibody production, and antitumor immunity in vivo, while the m6A modifications of these exogenous circRNAs could inhibit immunity activation. ('antigen-specific', 'CPA', (63, 79)) ('immunity activation', 'CPA', (228, 247)) ('tumor', 'Disease', (134, 139)) ('tumor', 'Disease', 'MESH:D009369', (134, 139)) ('induce', 'PosReg', (56, 62)) ('tumor', 'Phenotype', 'HP:0002664', (134, 139)) ('inhibit', 'NegReg', (220, 227)) ('antibody production', 'CPA', (105, 124)) ('m6A', 'Var', (168, 171)) ('modifications', 'Var', (172, 185)) 278 31757221 In addition, YTHDF2 was essential for inhibiting innate immunity by recognizing m6A. ('YTHDF2', 'Gene', '51441', (13, 19)) ('innate immunity', 'CPA', (49, 64)) ('inhibiting', 'NegReg', (38, 48)) ('YTHDF2', 'Gene', (13, 19)) ('m6A', 'Var', (80, 83)) 290 31757221 In addition, aberrant expression of mammalian HBXIP, a tumor protein, plays an important role in the occurrence and development of breast cancer. ('tumor', 'Disease', 'MESH:D009369', (55, 60)) ('tumor', 'Phenotype', 'HP:0002664', (55, 60)) ('aberrant expression', 'Var', (13, 32)) ('tumor', 'Disease', (55, 60)) ('cancer', 'Phenotype', 'HP:0002664', (138, 144)) ('mammalian', 'Species', '9606', (36, 45)) ('breast cancer', 'Disease', 'MESH:D001943', (131, 144)) ('HBXIP', 'Gene', (46, 51)) ('HBXIP', 'Gene', '10542', (46, 51)) ('breast cancer', 'Disease', (131, 144)) ('breast cancer', 'Phenotype', 'HP:0003002', (131, 144)) 307 31757221 Moreover, m6A-mediated degradation of GAS5 depends on the participation of YTHDF2. ('YTHDF2', 'Gene', (75, 81)) ('GAS5', 'Gene', (38, 42)) ('m6A-mediated', 'Var', (10, 22)) ('degradation', 'MPA', (23, 34)) ('YTHDF2', 'Gene', '51441', (75, 81)) ('GAS5', 'Gene', '60674', (38, 42)) 313 31757221 Most studies have shown that m6A modifications are very common in cancer. ('cancer', 'Disease', 'MESH:D009369', (66, 72)) ('cancer', 'Phenotype', 'HP:0002664', (66, 72)) ('m6A', 'Var', (29, 32)) ('cancer', 'Disease', (66, 72)) 315 31757221 Early detection of m6A upregulation in CTCs plays an important role in monitoring and preventing the occurrence and development of metastatic diseases, indicating that m6A RNA methylation in CTCs is a promising noninvasive diagnostic biomarker for cancer detection. ('cancer', 'Disease', (248, 254)) ('m6A', 'Var', (168, 171)) ('cancer', 'Disease', 'MESH:D009369', (248, 254)) ('cancer', 'Phenotype', 'HP:0002664', (248, 254)) 321 31757221 In addition, m6A modifications have the potential to be involved in combined tumor therapy. ('modifications', 'Var', (17, 30)) ('tumor', 'Disease', 'MESH:D009369', (77, 82)) ('m6A', 'Gene', (13, 16)) ('tumor', 'Phenotype', 'HP:0002664', (77, 82)) ('involved', 'Reg', (56, 64)) ('tumor', 'Disease', (77, 82)) 322 31757221 analyzed TCGA datasets and found that alterations in m6A regulatory genes were closely related to TP53 mutations in acute myeloid leukemia (AML) patients. ('patients', 'Species', '9606', (145, 153)) ('m6A regulatory genes', 'Gene', (53, 73)) ('mutations', 'Var', (103, 112)) ('leukemia', 'Phenotype', 'HP:0001909', (130, 138)) ('acute myeloid leukemia', 'Disease', (116, 138)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (122, 138)) ('AML', 'Disease', 'MESH:D015470', (140, 143)) ('AML', 'Phenotype', 'HP:0004808', (140, 143)) ('TP53', 'Gene', '7157', (98, 102)) ('alterations', 'Var', (38, 49)) ('related', 'Reg', (87, 94)) ('AML', 'Disease', (140, 143)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (116, 138)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (116, 138)) ('TP53', 'Gene', (98, 102)) 323 31757221 Furthermore, the analysis results showed that alterations in m6A regulatory genes reduced the survival rate of AML patients. ('AML', 'Disease', 'MESH:D015470', (111, 114)) ('patients', 'Species', '9606', (115, 123)) ('m6A regulatory genes', 'Gene', (61, 81)) ('AML', 'Disease', (111, 114)) ('AML', 'Phenotype', 'HP:0004808', (111, 114)) ('alterations', 'Var', (46, 57)) ('survival rate', 'CPA', (94, 107)) ('reduced', 'NegReg', (82, 89)) 326 31757221 found that YTHDF1 deletion significantly enhanced the therapeutic efficacy of PD-L1 checkpoint blockade, suggesting that YTHDF1, which plays a key role in m6A modifications, might be a potential therapeutic target in anticancer immunotherapy. ('cancer', 'Disease', 'MESH:D009369', (221, 227)) ('therapeutic efficacy', 'MPA', (54, 74)) ('cancer', 'Disease', (221, 227)) ('enhanced', 'PosReg', (41, 49)) ('cancer', 'Phenotype', 'HP:0002664', (221, 227)) ('YTHDF1', 'Gene', '54915', (11, 17)) ('PD-L1 checkpoint blockade', 'Disease', (78, 103)) ('YTHDF1', 'Gene', '54915', (121, 127)) ('PD-L1 checkpoint blockade', 'Disease', 'MESH:C536029', (78, 103)) ('YTHDF1', 'Gene', (121, 127)) ('deletion', 'Var', (18, 26)) ('YTHDF1', 'Gene', (11, 17)) 336 31757221 reported that R-2HG showed antitumor activity by blocking FTO to induce MYC degradation in AML patients with an IDH mutation. ('blocking', 'NegReg', (49, 57)) ('tumor', 'Phenotype', 'HP:0002664', (31, 36)) ('AML', 'Disease', 'MESH:D015470', (91, 94)) ('patients', 'Species', '9606', (95, 103)) ('mutation', 'Var', (116, 124)) ('tumor', 'Disease', (31, 36)) ('MYC', 'Gene', (72, 75)) ('AML', 'Disease', (91, 94)) ('FTO', 'Gene', (58, 61)) ('AML', 'Phenotype', 'HP:0004808', (91, 94)) ('IDH', 'Gene', (112, 115)) ('MYC', 'Gene', '4609', (72, 75)) ('tumor', 'Disease', 'MESH:D009369', (31, 36)) ('IDH', 'Gene', '3417', (112, 115)) ('FTO', 'Gene', '79068', (58, 61)) 337 31757221 TCGA data reveal high MYC levels and low FTO levels in tissue cells with an IDH mutation. ('MYC', 'Gene', '4609', (22, 25)) ('FTO', 'Gene', '79068', (41, 44)) ('IDH', 'Gene', (76, 79)) ('high', 'PosReg', (17, 21)) ('FTO', 'Gene', (41, 44)) ('IDH', 'Gene', '3417', (76, 79)) ('MYC', 'Gene', (22, 25)) ('mutation', 'Var', (80, 88)) 338 31757221 The use of R-2HG and MYC inhibitors could enhance cytotoxicity, which indicates that the combined utilization of R-2HG and MYC may be an effective treatment method for patients with leukemia characterized by an IDH mutation. ('mutation', 'Var', (215, 223)) ('enhance', 'PosReg', (42, 49)) ('leukemia', 'Phenotype', 'HP:0001909', (182, 190)) ('leukemia', 'Disease', 'MESH:D007938', (182, 190)) ('MYC', 'Gene', '4609', (123, 126)) ('cytotoxicity', 'Disease', 'MESH:D064420', (50, 62)) ('IDH', 'Gene', (211, 214)) ('leukemia', 'Disease', (182, 190)) ('MYC', 'Gene', '4609', (21, 24)) ('IDH', 'Gene', '3417', (211, 214)) ('patients', 'Species', '9606', (168, 176)) ('MYC', 'Gene', (123, 126)) ('MYC', 'Gene', (21, 24)) ('cytotoxicity', 'Disease', (50, 62)) 339 31757221 In summary, the analysis of m6A modifications could be used as a potential therapeutic target for cancer treatment. ('cancer', 'Phenotype', 'HP:0002664', (98, 104)) ('cancer', 'Disease', 'MESH:D009369', (98, 104)) ('cancer', 'Disease', (98, 104)) ('modifications', 'Var', (32, 45)) ('m6A', 'Protein', (28, 31)) 340 31757221 In addition to the clinical application of the m6A modifications described above, intervention through m6A modifications, and/or noncoding RNA regulatory mechanisms at certain carcinogenic sites may affect tumor proliferation. ('m6A', 'Gene', (103, 106)) ('tumor', 'Phenotype', 'HP:0002664', (206, 211)) ('modifications', 'Var', (51, 64)) ('modifications', 'Var', (107, 120)) ('tumor', 'Disease', (206, 211)) ('tumor', 'Disease', 'MESH:D009369', (206, 211)) ('carcinogenic', 'Disease', 'MESH:D063646', (176, 188)) ('carcinogenic', 'Disease', (176, 188)) ('affect', 'Reg', (199, 205)) 342 31757221 demonstrated that miRNA-6852 overexpression could induce G2/M phase arrest in cervical cancer cells. ('G2/M phase arrest', 'CPA', (57, 74)) ('miRNA-6852', 'Var', (18, 28)) ('overexpression', 'PosReg', (29, 43)) ('cervical cancer', 'Disease', 'MESH:D002583', (78, 93)) ('cancer', 'Phenotype', 'HP:0002664', (87, 93)) ('cervical cancer', 'Disease', (78, 93)) ('induce', 'Reg', (50, 56)) 348 31757221 Silencing lncRNA PVT1 expression can restore the inhibitory effect of miR-195 on FASN and thus inhibit the proliferation, migration, and invasion of osteosarcoma. ('invasion of osteosarcoma', 'Disease', (137, 161)) ('restore', 'PosReg', (37, 44)) ('miR-195', 'Gene', (70, 77)) ('FASN', 'Gene', (81, 85)) ('miR-195', 'Gene', '406971', (70, 77)) ('FASN', 'Gene', '2194', (81, 85)) ('PVT1', 'Gene', (17, 21)) ('proliferation', 'CPA', (107, 120)) ('inhibit', 'NegReg', (95, 102)) ('inhibitory effect', 'MPA', (49, 66)) ('invasion of osteosarcoma', 'Disease', 'MESH:D012516', (137, 161)) ('PVT1', 'Gene', '5820', (17, 21)) ('osteosarcoma', 'Phenotype', 'HP:0002669', (149, 161)) ('migration', 'CPA', (122, 131)) ('Silencing', 'Var', (0, 9)) 356 31757221 Another study reported that the upregulation of METTL3 caused an increase in miR-25-3p and that miR-25-3p could promote the expression of its target protein PHLPP2 to promote pancreatic ductal adenocarcinoma occurrence. ('increase', 'PosReg', (65, 73)) ('METTL3', 'Gene', '56339', (48, 54)) ('PHLPP2', 'Gene', (157, 163)) ('promote', 'PosReg', (112, 119)) ('upregulation', 'PosReg', (32, 44)) ('promote', 'PosReg', (167, 174)) ('METTL3', 'Gene', (48, 54)) ('miR-25-3p', 'Var', (96, 105)) ('carcinoma', 'Phenotype', 'HP:0030731', (198, 207)) ('pancreatic ductal adenocarcinoma', 'Disease', (175, 207)) ('miR-25-3p', 'MPA', (77, 86)) ('pancreatic ductal adenocarcinoma', 'Disease', 'MESH:D021441', (175, 207)) ('expression', 'MPA', (124, 134)) ('PHLPP2', 'Gene', '23035', (157, 163)) ('pancreatic ductal adenocarcinoma', 'Phenotype', 'HP:0006725', (175, 207)) 359 31757221 The m6A modification of MALAT1 could regulate gene expression. ('gene expression', 'MPA', (46, 61)) ('MALAT1', 'Gene', (24, 30)) ('regulate', 'Reg', (37, 45)) ('m6A modification', 'Var', (4, 20)) ('MALAT1', 'Gene', '378938', (24, 30)) 361 31757221 A recent study also revealed that altering the modification levels of m6A in lncRNA 1281 could significantly affect ESC differentiation. ('ESC', 'Disease', (116, 119)) ('m6A', 'Var', (70, 73)) ('lncRNA 1281', 'Gene', (77, 88)) ('affect', 'Reg', (109, 115)) ('ESC', 'CellLine', 'CVCL:M564', (116, 119)) 364 31757221 M6A modifications of noncoding RNAs have been demonstrated to control gene expression, as demonstrated by their ability to regulate the biological functions of cells in cancer, including proliferation, metastasis, stem cell differentiation, and homeostasis. ('homeostasis', 'CPA', (245, 256)) ('cancer', 'Disease', (169, 175)) ('stem cell differentiation', 'CPA', (214, 239)) ('M6A', 'Gene', '56339', (0, 3)) ('regulate', 'Reg', (123, 131)) ('M6A', 'Gene', (0, 3)) ('biological', 'MPA', (136, 146)) ('cancer', 'Phenotype', 'HP:0002664', (169, 175)) ('gene expression', 'MPA', (70, 85)) ('control', 'Reg', (62, 69)) ('proliferation', 'CPA', (187, 200)) ('metastasis', 'CPA', (202, 212)) ('modifications', 'Var', (4, 17)) ('cancer', 'Disease', 'MESH:D009369', (169, 175)) 366 31757221 The association of m6A modifications and noncoding RNAs provides a new direction for exploring the underlying regulatory mechanisms of gene expression in cancer. ('association', 'Interaction', (4, 15)) ('cancer', 'Disease', (154, 160)) ('m6A', 'Gene', (19, 22)) ('cancer', 'Disease', 'MESH:D009369', (154, 160)) ('modifications', 'Var', (23, 36)) ('cancer', 'Phenotype', 'HP:0002664', (154, 160)) 367 31757221 Furthermore, the clinical application of m6A modifications and noncoding RNAs includes their use as cancer indicators and targets for therapeutic interventions in cancer treatment. ('modifications', 'Var', (45, 58)) ('cancer', 'Phenotype', 'HP:0002664', (100, 106)) ('cancer', 'Phenotype', 'HP:0002664', (163, 169)) ('cancer', 'Disease', (100, 106)) ('cancer', 'Disease', 'MESH:D009369', (100, 106)) ('m6A modifications', 'Var', (41, 58)) ('cancer', 'Disease', (163, 169)) ('cancer', 'Disease', 'MESH:D009369', (163, 169)) 368 31757221 Further studies are needed to explore the mutual regulatory mechanisms between m6A modifications and noncoding RNAs in additional types of cancers and the effective therapeutic interventions of m6A modifications and noncoding RNAs for cancer patients. ('cancer', 'Disease', 'MESH:D009369', (139, 145)) ('patients', 'Species', '9606', (242, 250)) ('cancers', 'Disease', 'MESH:D009369', (139, 146)) ('cancer', 'Disease', (235, 241)) ('cancer', 'Disease', 'MESH:D009369', (235, 241)) ('cancers', 'Phenotype', 'HP:0002664', (139, 146)) ('cancer', 'Phenotype', 'HP:0002664', (139, 145)) ('cancers', 'Disease', (139, 146)) ('m6A', 'Gene', (79, 82)) ('cancer', 'Phenotype', 'HP:0002664', (235, 241)) ('modifications', 'Var', (83, 96)) ('cancer', 'Disease', (139, 145)) 371 31757221 This study was supported by the National Natural Science Foundation of China (81972663, 81560385), Key Scientific Research Projects of Institutions of Higher Education in Henan Province (19A310024), The Medical Scientific and Technological Research Project of Henan Province (201702027), Youth Innovation Fund Project of The First Affiliated Hospital of Zhengzhou University (YNQN2017035), the China Postdoctoral Science Foundation (2019T120648, 2017M610462), the National Natural Science Foundation of Henan Province (182300410342), and the Health Commission Technology Talents Overseas Training Project of Henan Province (2018140). ('182300410342', 'Var', (519, 531)) ('YNQN2017035', 'Var', (376, 387)) ('19A310024', 'Chemical', 'MESH:C039200', (187, 196)) ('2019T120648', 'Var', (433, 444)) ('201702027', 'Var', (276, 285)) ('YNQN2017035', 'Chemical', 'None', (376, 387)) 374 31700693 CX-5461 is a potent Pol I transcription inhibitor and stabilizer of the DNA G-quadruplex structure, which causes G2/M-phase arrest via the ATR (ataxia telangiectasia and Rad3-related protein)-mediated DNA damage response (DDR). ('ataxia', 'Phenotype', 'HP:0001251', (144, 150)) ('G2/M-phase arrest', 'Disease', (113, 130)) ('causes', 'Reg', (106, 112)) ('ataxia telangiectasia and Rad3-related protein', 'Gene', '545', (144, 190)) ('telangiectasia', 'Phenotype', 'HP:0001009', (151, 165)) ('CX-5461', 'Var', (0, 7)) 375 31700693 Therefore, we hypothesized that the ATR-selective inhibitor AZD6738 would synergistically enhance CX-5461-induced cell death via abolishment of the ATR-mediated DDR. ('CX-5461-induced', 'MPA', (98, 113)) ('ATR-mediated DDR', 'MPA', (148, 164)) ('enhance', 'PosReg', (90, 97)) ('abolishment', 'NegReg', (129, 140)) ('AZD6738', 'Chemical', 'MESH:C009217', (60, 67)) ('AZD6738', 'Var', (60, 67)) 377 31700693 Compared with monotherapy, combination treatment of AML cell lines with CX-5461 and AZD6738 for 48 h induced significantly increased apoptosis, as reflected by enhanced Annexin V positivity and substantially increased caspase 3 and PARP cleavage (Figs. ('CX-5461', 'Var', (72, 79)) ('PARP', 'Gene', (232, 236)) ('AML', 'Phenotype', 'HP:0004808', (52, 55)) ('apoptosis', 'CPA', (133, 142)) ('increased', 'PosReg', (208, 217)) ('caspase 3', 'Gene', (218, 227)) ('caspase 3', 'Gene', '836', (218, 227)) ('AZD6738', 'Var', (84, 91)) ('Annexin V', 'Gene', (169, 178)) ('Annexin V', 'Gene', '308', (169, 178)) ('PARP', 'Gene', '142', (232, 236)) ('enhanced', 'PosReg', (160, 168)) 379 31700693 Both monotherapy and combination therapy with AZD6738 downregulated CHK-1 protein expression. ('CHK-1', 'Gene', '1111', (68, 73)) ('downregulated', 'NegReg', (54, 67)) ('CHK-1', 'Gene', (68, 73)) ('AZD6738', 'Var', (46, 53)) 381 31700693 CX-5461 caused a time-dependent increase in phosphorylated CDK1 (p-CDK1), which was abolished by combination treatment (Fig. ('CDK1', 'Gene', '983', (67, 71)) ('increase', 'PosReg', (32, 40)) ('CDK1', 'Gene', (67, 71)) ('CDK1', 'Gene', (59, 63)) ('CDK1', 'Gene', '983', (59, 63)) ('phosphorylated', 'MPA', (44, 58)) ('CX-5461', 'Var', (0, 7)) 384 31700693 We previously demonstrated that the ATR inhibitor AZ20 causes downregulation of ribonucleotide reductase (RR, a key enzyme in the synthesis of dNDPs) subunits M1 (RRM1) and M2 (RRM2). ('AZ20', 'Var', (50, 54)) ('ribonucleotide', 'Chemical', 'MESH:D012265', (80, 94)) ('RRM1', 'Gene', '6240', (163, 167)) ('downregulation', 'NegReg', (62, 76)) ('RR', 'Gene', (106, 108)) ('RRM1', 'Gene', (163, 167)) ('RRM2', 'Gene', (177, 181)) ('RRM2', 'Gene', '6241', (177, 181)) 390 31700693 Therefore, ATR inhibition abolishes the G2/M cell cycle arrest and prevents RRM2 upregulation, decreasing dNTP pools and resulting in the accumulation of damaged DNA and cell death. ('prevents', 'NegReg', (67, 75)) ('decreasing', 'NegReg', (95, 105)) ('abolishes', 'NegReg', (26, 35)) ('RRM2', 'Gene', (76, 80)) ('accumulation', 'PosReg', (138, 150)) ('ATR', 'Gene', (11, 14)) ('upregulation', 'PosReg', (81, 93)) ('G2/M cell cycle arrest', 'CPA', (40, 62)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (45, 62)) ('dNTP', 'Chemical', 'MESH:C065818', (106, 110)) ('inhibition', 'Var', (15, 25)) ('dNTP pools', 'MPA', (106, 116)) ('damaged DNA', 'MPA', (154, 165)) 395 30472158 IKZF2 depletion in AML cells reduced colony formation, increased differentiation and apoptosis, and delayed leukemogenesis. ('AML', 'Disease', 'MESH:D015470', (19, 22)) ('differentiation', 'CPA', (65, 80)) ('increased', 'PosReg', (55, 64)) ('IKZF2', 'Gene', (0, 5)) ('delayed', 'NegReg', (100, 107)) ('colony formation', 'CPA', (37, 53)) ('apoptosis', 'CPA', (85, 94)) ('AML', 'Disease', (19, 22)) ('leukemogenesis', 'Disease', (108, 122)) ('reduced', 'NegReg', (29, 36)) ('depletion', 'Var', (6, 15)) 396 30472158 Gene expression, chromatin accessibility and direct IKZF2 binding in MLL-AF9 LSCs demonstrate that IKZF2 regulates a HOXA9 self-renewal gene expression program and inhibits a C/EBP-driven differentiation program. ('C/EBP', 'Gene', '12606', (175, 180)) ('inhibits', 'NegReg', (164, 172)) ('regulates', 'Reg', (105, 114)) ('C/EBP', 'Gene', (175, 180)) ('HOXA9', 'Gene', '15405', (117, 122)) ('IKZF2', 'Var', (99, 104)) ('HOXA9', 'Gene', (117, 122)) 402 30472158 Translocations of histone methyltransferases such as the mixed-lineage leukemia (MLL) gene give rise to one of the most aggressive subtypes of AML. ('leukemia', 'Phenotype', 'HP:0001909', (71, 79)) ('Translocations', 'Var', (0, 14)) ('mixed-lineage leukemia', 'Gene', (57, 79)) ('AML', 'Disease', 'MESH:D015470', (143, 146)) ('give rise to', 'Reg', (91, 103)) ('mixed-lineage leukemia', 'Gene', '214162', (57, 79)) ('AML', 'Disease', (143, 146)) ('MLL', 'Gene', (81, 84)) 413 30472158 To examine the role of IKZF2 in HSCs, we assessed different populations of the hematopoietic system in Ikzf2f/f and Ikzf2Delta/Delta littermates. ('Ikzf2f/f', 'Var', (103, 111)) ('HSC', 'Gene', '2523', (32, 35)) ('HSC', 'Gene', (32, 35)) 414 30472158 Ikzf2 deletion was confirmed by genotyping PCR in bone marrow cells and by qPCR in sorted Lin- Sca1+c-Kit+ (LSK) cells (Figure S1A,B). ('LSK', 'Gene', '16818', (108, 111)) ('LSK', 'Gene', (108, 111)) ('Sca1+c-Kit', 'Gene', (95, 105)) ('deletion', 'Var', (6, 14)) ('Sca1+c-Kit', 'Gene', '110454;16590', (95, 105)) ('Ikzf2', 'Gene', (0, 5)) 416 30472158 To assess the in vitro functional output of HSPCs, we sorted LSK cells from Ikzf2f/f and Ikzf2Delta/Delta mice and performed colony assays, which resulted in comparable number of myeloid progenitor colonies in primary and secondary plating (Figure 1A-B). ('mice', 'Species', '10090', (106, 110)) ('Ikzf2f/f', 'Var', (76, 84)) ('Ikzf2Delta/Delta', 'Var', (89, 105)) ('LSK', 'Gene', '16818', (61, 64)) ('LSK', 'Gene', (61, 64)) 417 30472158 Furthermore, transplantation of bone marrow cells from Ikzf2f/f and Ikzf2Delta/Delta mice resulted in equivalent chimerism in all hematopoietic cells analyzed at different time points post-transplantation in primary and secondary transplanted mice (Figure S1M-P). ('mice', 'Species', '10090', (85, 89)) ('chimerism', 'CPA', (113, 122)) ('Ikzf2f/f', 'Var', (55, 63)) ('Ikzf2Delta/Delta', 'Var', (68, 84)) ('mice', 'Species', '10090', (243, 247)) 421 30472158 LSK cells transduced with MLL-AF9 virus from Ikzf2f/f and Ikzf2Delta/Delta mice resulted in a significant block in colony formation in the Ikzf2Delta/Delta LSK cells (Figure 1C). ('colony formation', 'CPA', (115, 131)) ('LSK', 'Gene', '16818', (0, 3)) ('LSK', 'Gene', (0, 3)) ('Ikzf2f/f', 'Var', (45, 53)) ('mice', 'Species', '10090', (75, 79)) ('LSK', 'Gene', '16818', (156, 159)) ('LSK', 'Gene', (156, 159)) ('block', 'NegReg', (106, 111)) ('Ikzf2Delta/Delta', 'Var', (58, 74)) 425 30472158 Ikzf2 deletion was verified at 24hrs after 4-OHT treatment (Figure 2B). ('4-OHT', 'Chemical', 'MESH:C032278', (43, 48)) ('deletion', 'Var', (6, 14)) ('Ikzf2', 'Gene', (0, 5)) 427 30472158 Ikzf2 deletion also resulted in rapid increase in apoptosis 24hrs after 4-OHT treatment, based on Annexin-V and 7-AAD staining compared to control cells (Figure 2E). ('increase', 'PosReg', (38, 46)) ('Annexin-V', 'Gene', (98, 107)) ('Annexin-V', 'Gene', '11747', (98, 107)) ('deletion', 'Var', (6, 14)) ('4-OHT', 'Chemical', 'MESH:C032278', (72, 77)) ('apoptosis', 'CPA', (50, 59)) ('Ikzf2', 'Gene', (0, 5)) 428 30472158 Additionally, Ikzf2 deficiency led to a drastic increase in differentiation assessed by frequency and Median Fluorescence Intensity (MFI) of myeloid markers Mac-1, Gr-1, F4/80 and CD115 (Figure 2F-G and Figure S2A-B) and cellular morphology (Figure S2C) at 24hrs post treatment. ('Gr-1, F4', 'Gene', '546644', (164, 172)) ('Mac-1', 'Gene', (157, 162)) ('CD115', 'Gene', (180, 185)) ('Ikzf2', 'Gene', (14, 19)) ('deficiency', 'Var', (20, 30)) ('CD115', 'Gene', '12978', (180, 185)) ('cellular morphology', 'CPA', (221, 240)) ('differentiation', 'MPA', (60, 75)) ('Mac-1', 'Gene', '16409', (157, 162)) ('increase', 'PosReg', (48, 56)) 429 30472158 In agreement, leukemic mice treated in vivo with tamoxifen demonstrated delayed leukemogenesis in the Ikzf2f/f group compared to the wildtype group (44 days versus 22 days in 4-OHT treated Ikzf2f/f versus wildtype) (Figure 2H) indicating that IKZF2 plays an important role in maintaining MLL-AF9 leukemia. ('leukemic', 'Disease', 'MESH:D007938', (14, 22)) ('delayed', 'NegReg', (72, 79)) ('leukemia', 'Phenotype', 'HP:0001909', (296, 304)) ('4-OHT', 'Chemical', 'MESH:C032278', (175, 180)) ('MLL-AF9 leukemia', 'Disease', 'MESH:D007938', (288, 304)) ('leukemic', 'Disease', (14, 22)) ('MLL-AF9 leukemia', 'Disease', (288, 304)) ('Ikzf2f/f', 'Var', (102, 110)) ('tamoxifen', 'Chemical', 'MESH:D013629', (49, 58)) ('leukemogenesis', 'CPA', (80, 94)) ('mice', 'Species', '10090', (23, 27)) 435 30472158 IKZF2 was found to be highly expressed in the c-KitHigh cells, which are enriched for the LSC fraction both at the level of mRNA and protein, suggesting that IKZF2 could contribute to LSC function (Figure 3A-C and Figure S3A). ('LSC', 'Disease', (184, 187)) ('contribute', 'Reg', (170, 180)) ('IKZF2', 'Var', (158, 163)) ('c-Kit', 'Gene', (46, 51)) ('c-Kit', 'Gene', '16590', (46, 51)) 436 30472158 Interestingly, the frequency of the leukemic-GMPs (L-GMP; c-KitHigh FcgRIIb+ CD34+) and the expression of CD34 marker was modestly reduced in the primary Ikzf2Delta/Delta transplanted mice compared to control mice (Figure S3B-C). ('mice', 'Species', '10090', (209, 213)) ('L-GMP', 'Chemical', 'MESH:C026693', (51, 56)) ('S3B-C', 'Chemical', 'MESH:C027397', (222, 227)) ('leukemic', 'Disease', 'MESH:D007938', (36, 44)) ('CD34', 'Gene', (106, 110)) ('reduced', 'NegReg', (131, 138)) ('c-Kit', 'Gene', '16590', (58, 63)) ('c-Kit', 'Gene', (58, 63)) ('GMPs', 'Chemical', 'MESH:C009223', (45, 49)) ('mice', 'Species', '10090', (184, 188)) ('Ikzf2Delta/Delta', 'Var', (154, 170)) ('expression', 'MPA', (92, 102)) ('leukemic', 'Disease', (36, 44)) 443 30472158 Moreover, mice that were treated with tamoxifen in the primary transplant experiment did not have good deletion except for one mouse in which all secondarily transplanted mice from this donor no longer had Ikzf2 deletion at point of death, indicating a selective pressure for loss of Ikzf2 deletion in these leukemic mice (Figure S3E). ('mouse', 'Species', '10090', (127, 132)) ('Ikzf2', 'Gene', (206, 211)) ('deletion', 'Var', (290, 298)) ('mice', 'Species', '10090', (171, 175)) ('mice', 'Species', '10090', (10, 14)) ('donor', 'Species', '9606', (186, 191)) ('leukemic', 'Disease', (308, 316)) ('loss', 'NegReg', (276, 280)) ('deletion', 'Var', (212, 220)) ('mice', 'Species', '10090', (317, 321)) ('tamoxifen', 'Chemical', 'MESH:D013629', (38, 47)) ('death', 'Disease', 'MESH:D003643', (233, 238)) ('death', 'Disease', (233, 238)) ('Ikzf2', 'Gene', (284, 289)) ('leukemic', 'Disease', 'MESH:D007938', (308, 316)) 451 30472158 Similar to what was observed in mouse MLL-AF9 cells, IKZF2 depletion resulted in reduced colony formation, proliferation and increased apoptosis and differentiation (Figure 4D-H and S4D). ('apoptosis', 'CPA', (135, 144)) ('increased', 'PosReg', (125, 134)) ('IKZF2', 'Gene', (53, 58)) ('S4D', 'Chemical', 'MESH:C042345', (182, 185)) ('proliferation', 'CPA', (107, 120)) ('differentiation', 'CPA', (149, 164)) ('colony formation', 'CPA', (89, 105)) ('depletion', 'Var', (59, 68)) ('mouse', 'Species', '10090', (32, 37)) ('reduced', 'NegReg', (81, 88)) 453 30472158 Additionally, the role for IKZF2 in myeloid leukemia was not specific only to MOLM-13 cells as shRNA depletion in KCL-22 (BCR-ABL), Kasumi-1 (AML1-ETO) and NOMO1 (MLL-rearranged) also resulted in similar effects of reduced proliferation, increased apoptosis and differentiation (Figure S4E-Q). ('myeloid leukemia', 'Phenotype', 'HP:0012324', (36, 52)) ('leukemia', 'Phenotype', 'HP:0001909', (44, 52)) ('KCL-22', 'Gene', (114, 120)) ('MOLM-13', 'CellLine', 'CVCL:2119', (78, 85)) ('proliferation', 'CPA', (223, 236)) ('differentiation', 'CPA', (262, 277)) ('NOMO1', 'Gene', '23420', (156, 161)) ('increased', 'PosReg', (238, 247)) ('BCR-ABL', 'Gene', (122, 129)) ('reduced', 'NegReg', (215, 222)) ('BCR-ABL', 'Gene', '25', (122, 129)) ('NOMO1', 'Gene', (156, 161)) ('apoptosis', 'CPA', (248, 257)) ('depletion', 'Var', (101, 110)) ('leukemia', 'Disease', 'MESH:D007938', (44, 52)) ('leukemia', 'Disease', (44, 52)) 463 30472158 Interestingly, overlap of increased accessibility and increased RNA expression identified seven gene sets that include myeloid development, CCAAT/enhancer-binding proteinalpha (C/EBPalpha) network and myeloid cell maturation (Figure 5D) indicating the myeloid differentiation program is strongly turned on in the Ikzf2Delta/Delta LSCs. ('myeloid differentiation program', 'CPA', (252, 283)) ('CCAAT/enhancer-binding proteinalpha', 'Gene', (140, 175)) ('C/EBPalpha', 'Gene', (177, 187)) ('myeloid development', 'CPA', (119, 138)) ('turned', 'PosReg', (296, 302)) ('C/EBPalpha', 'Gene', '12606', (177, 187)) ('CCAAT/enhancer-binding proteinalpha', 'Gene', '12606', (140, 175)) ('Ikzf2Delta/Delta', 'Var', (313, 329)) 467 30472158 More specifically, we found that 14 genes shared IKZF2 and C/EBP motifs, that had increased gene expression (Log2FC>0.75, pval<0.05) coupled with increased accessibility (Log2FC>1, pval<0.05) after Ikzf2 deletion and had direct IKZF2 binding (Figure S5L and Table S4-5). ('C/EBP', 'Gene', '12606', (59, 64)) ('deletion', 'Var', (204, 212)) ('increased', 'PosReg', (146, 155)) ('gene expression', 'MPA', (92, 107)) ('C/EBP', 'Gene', (59, 64)) ('binding', 'Interaction', (234, 241)) ('Ikzf2', 'Gene', (198, 203)) ('increased', 'PosReg', (82, 91)) ('accessibility', 'MPA', (156, 169)) ('S4-5', 'Chemical', 'MESH:D013455', (264, 268)) 469 30472158 Utilizing the Cre-ER deletion system, we found that Ikzf2 deletion could be verified as early as 14hrs post 4-OHT treatment in the LSCs (Figure 6A). ('Ikzf2', 'Gene', (52, 57)) ('4-OHT', 'Chemical', 'MESH:C032278', (108, 113)) ('deletion', 'Var', (58, 66)) 470 30472158 Among the 14 genes tested that demonstrated increased accessibility and increased expression after Ikzf2 was deleted in the LSCs from the primary transplanted mice, we identified a similar rapid increase in mRNA levels of C3, Fpr2, S100a8 and S100a9 (Figure 6B-E and Figure S6A). ('increased', 'PosReg', (72, 81)) ('expression', 'MPA', (82, 92)) ('Fpr2', 'Gene', '14289', (226, 230)) ('mRNA levels', 'MPA', (207, 218)) ('S100a9', 'Gene', '20202', (243, 249)) ('S100a9', 'Gene', (243, 249)) ('mice', 'Species', '10090', (159, 163)) ('deleted', 'Var', (109, 116)) ('Fpr2', 'Gene', (226, 230)) ('Ikzf2', 'Gene', (99, 104)) ('S100a8', 'Gene', '20201', (232, 238)) ('increase', 'PosReg', (195, 203)) ('S100a8', 'Gene', (232, 238)) 471 30472158 In accordance with the C/EBPepsilon motif being the most accessible after Ikzf2 deletion, we found increased mRNA abundance of C/ebpepsilon among the C/ebp family members after acute IKZF2 depletion (Figure 6F and Figure S6C). ('C/ebpepsilon', 'Gene', (127, 139)) ('deletion', 'Var', (80, 88)) ('C/EBPepsilon', 'Gene', (23, 35)) ('C/ebp', 'Gene', '12606', (127, 132)) ('mRNA abundance', 'MPA', (109, 123)) ('C/EBPepsilon', 'Gene', '110794', (23, 35)) ('increased', 'PosReg', (99, 108)) ('C/ebp', 'Gene', (150, 155)) ('Ikzf2', 'Gene', (74, 79)) ('C/ebpepsilon', 'Gene', '110794', (127, 139)) ('C/ebp', 'Gene', (127, 132)) ('C/ebp', 'Gene', '12606', (150, 155)) 477 30472158 Similarly, this reduction of c-Myc and Hoxa9 gene expression was also observed after acute deletion at the mRNA levels at early time points and at protein levels after 24hrs post Ikzf2 deletion (Figure 6G-I). ('Hoxa9', 'Gene', '15405', (39, 44)) ('deletion', 'Var', (91, 99)) ('Hoxa9', 'Gene', (39, 44)) ('reduction', 'NegReg', (16, 25)) ('Myc', 'Gene', (31, 34)) ('Myc', 'Gene', '17869', (31, 34)) ('deletion', 'Var', (185, 193)) 480 30472158 Since genes containing HOXA9 motifs share IKZF2 binding and HOXA9 is rapidly lost after Ikzf2 deletion, we tested if reintroducing HOXA9 alone would reverse the cellular affects from depleting IKZF2. ('HOXA9', 'Gene', (23, 28)) ('deletion', 'Var', (94, 102)) ('HOXA9', 'Gene', (60, 65)) ('HOXA9', 'Gene', '15405', (131, 136)) ('HOXA9', 'Gene', '15405', (23, 28)) ('Ikzf2', 'Gene', (88, 93)) ('tested', 'Reg', (107, 113)) ('depleting', 'MPA', (183, 192)) ('HOXA9', 'Gene', (131, 136)) ('HOXA9', 'Gene', '15405', (60, 65)) 481 30472158 In line with HOXA9 being an important downstream mediator of IKZF2 function, forced expression of HOXA9 could partially rescue the differentiation, apoptosis and the reduction of colony formation caused by Ikzf2 deletion (Figure 7A-F). ('HOXA9', 'Gene', (98, 103)) ('HOXA9', 'Gene', '15405', (13, 18)) ('Ikzf2', 'Gene', (206, 211)) ('reduction', 'NegReg', (166, 175)) ('differentiation', 'CPA', (131, 146)) ('HOXA9', 'Gene', (13, 18)) ('deletion', 'Var', (212, 220)) ('colony formation', 'CPA', (179, 195)) ('HOXA9', 'Gene', '15405', (98, 103)) ('apoptosis', 'CPA', (148, 157)) ('rescue', 'PosReg', (120, 126)) 482 30472158 Similarly, CEBPE depletion could rescue the differentiation effects (Figure 7G-H) and reverse the activation of several co-regulated targets including S100a8 and S100a9 when IKZF2 is lost (Figure 7I-L). ('S100a9', 'Gene', '20202', (162, 168)) ('S100a9', 'Gene', (162, 168)) ('reverse', 'NegReg', (86, 93)) ('differentiation effects', 'CPA', (44, 67)) ('S100a8', 'Gene', '20201', (151, 157)) ('S100a8', 'Gene', (151, 157)) ('rescue', 'PosReg', (33, 39)) ('depletion', 'Var', (17, 26)) ('activation', 'MPA', (98, 108)) 484 30472158 Additionally, IKZF2 maintains accessibility of genes associated with the self-renewal program including genes with HOXA9 motifs in MLL-AF9 LSCs (Figure 7M). ('HOXA9', 'Gene', '15405', (115, 120)) ('accessibility', 'MPA', (30, 43)) ('HOXA9', 'Gene', (115, 120)) ('motifs', 'Var', (121, 127)) 487 30472158 Recently, post-transcriptional regulation in leukemia has been highlighted as a novel way for maintaining the LSC program. ('leukemia', 'Disease', 'MESH:D007938', (45, 53)) ('post-transcriptional regulation', 'Var', (10, 41)) ('leukemia', 'Disease', (45, 53)) ('leukemia', 'Phenotype', 'HP:0001909', (45, 53)) 491 30472158 Furthermore, we found that IKZF2 depletion results in reduced expression of MSI2 direct mRNA binding targets (Figure S5G). ('MSI2', 'Gene', (76, 80)) ('depletion', 'Var', (33, 42)) ('MSI2', 'Gene', '76626', (76, 80)) ('expression', 'MPA', (62, 72)) ('reduced', 'NegReg', (54, 61)) ('IKZF2', 'Gene', (27, 32)) 496 30472158 Similar to IKZF1, IKZF2 has also been found to interact with chromatin remodeling complexes that contain histone deacetylase (HDAC), the nucleosome-remodeling and histone deacetylase (NuRD) and SIN3 complexes which are involved in gene repression. ('SIN3', 'Gene', '20466', (194, 198)) ('SIN3', 'Gene', (194, 198)) ('IKZF1', 'Gene', (11, 16)) ('IKZF2', 'Var', (18, 23)) ('interact', 'Interaction', (47, 55)) ('IKZF1', 'Gene', '22778', (11, 16)) 500 30472158 The increased accessibility within the regions that contain both IKZF2 and C/EBP motifs in the Ikzf2Delta/Delta LSCs suggest that Ikzf2 deletion could lead to loss of the repressive complex recruitment or occupancy allowing transcription machinery to be recruited to the C/EBP motifs found in the myeloid genes to turn on transcription. ('C/EBP', 'Gene', (75, 80)) ('loss', 'NegReg', (159, 163)) ('repressive complex recruitment', 'MPA', (171, 201)) ('C/EBP', 'Gene', '12606', (271, 276)) ('C/EBP', 'Gene', (271, 276)) ('accessibility', 'MPA', (14, 27)) ('deletion', 'Var', (136, 144)) ('recruited', 'PosReg', (254, 263)) ('occupancy', 'MPA', (205, 214)) ('increased', 'PosReg', (4, 13)) ('Ikzf2', 'Gene', (130, 135)) ('C/EBP', 'Gene', '12606', (75, 80)) 501 30472158 Interestingly, acute deletion of Ikzf2 led to a rapid and significant increase in the transcription of myeloid genes regulated by C/EBPs and also the mRNA of C/ebpepsilon suggesting an autoregulatory mechanism found in C/EBPs. ('C/ebpepsilon', 'Gene', (158, 170)) ('C/EBP', 'Gene', '12606', (130, 135)) ('increase', 'PosReg', (70, 78)) ('C/EBP', 'Gene', (130, 135)) ('C/EBP', 'Gene', '12606', (219, 224)) ('C/EBP', 'Gene', (219, 224)) ('deletion', 'Var', (21, 29)) ('transcription', 'MPA', (86, 99)) ('Ikzf2', 'Gene', (33, 38)) ('mRNA', 'MPA', (150, 154)) ('myeloid genes', 'Gene', (103, 116)) ('C/ebpepsilon', 'Gene', '110794', (158, 170)) 505 30472158 Additionally, a recent study has identified a variant NuRD complex in ES cells that includes the WDR5, a component of the SET/MLL1 complex that contributes to H3K4 trimethlyation. ('WDR5', 'Gene', (97, 101)) ('variant', 'Var', (46, 53)) ('trimethlyation', 'MPA', (164, 178)) ('WDR5', 'Gene', '140858', (97, 101)) ('MLL1', 'Gene', '214162', (126, 130)) ('MLL1', 'Gene', (126, 130)) ('H3K4', 'Protein', (159, 163)) ('H3K4', 'Chemical', 'MESH:C024755', (159, 163)) 507 30472158 Furthermore, since IKZF2 knockdown results in similar phenotype in human cell lines with different translocations (Figure 4 and S4; MLL-AF9, BCR-ABL, AML1-ETO and PML-RAR) and in two different murine AML models, we speculate that IKZF2 could be working through transcriptional complexes, not limited to MLL rearranged complexes. ('BCR-ABL', 'Gene', '25', (141, 148)) ('murine', 'Species', '10090', (193, 199)) ('IKZF2', 'Gene', (19, 24)) ('AML', 'Disease', 'MESH:D015470', (200, 203)) ('AML', 'Disease', 'MESH:D015470', (150, 153)) ('PML-RAR', 'Gene', (163, 170)) ('AML', 'Disease', (200, 203)) ('AML', 'Disease', (150, 153)) ('human', 'Species', '9606', (67, 72)) ('PML-RAR', 'Gene', '84106', (163, 170)) ('BCR-ABL', 'Gene', (141, 148)) ('knockdown', 'Var', (25, 34)) 513 30472158 Despite the availability for drugs (i.e lenalidomide) that have specificity to degrade IKAROS members (IKZF1 and IKZF3), these proteins have an amino acid substitution in the critical region that precludes binding to these imids. ('IKZF3', 'Gene', (113, 118)) ('binding', 'Interaction', (206, 213)) ('IKZF3', 'Gene', '22780', (113, 118)) ('lenalidomide', 'Chemical', 'MESH:C467567', (40, 52)) ('IKAROS', 'Gene', '22778', (87, 93)) ('degrade', 'NegReg', (79, 86)) ('amino acid substitution', 'Var', (144, 167)) ('IKZF1', 'Gene', (103, 108)) ('IKZF1', 'Gene', '22778', (103, 108)) ('IKAROS', 'Gene', (87, 93)) 518 30472158 Bone marrow cells were extracted from 6 to 8 weeks old Ikzf2fl/fl or Ikzf2Delta/Delta mice. ('mice', 'Species', '10090', (86, 90)) ('Ikzf2Delta/Delta', 'Var', (69, 85)) ('Ikzf2fl/fl', 'Var', (55, 65)) 524 30472158 Mouse leukemic bone marrow cells from WT and Ikzf2f/f mice were grown in RPMI (Cellgro) medium containing 10% FBS, 6 ng/ml IL-3, 10 ng/ml SCF, 10 ng/ml IL-6 and 10 ng/ml GM-CSF. ('GM-CSF', 'Gene', (170, 176)) ('IL-6', 'Gene', (152, 156)) ('Ikzf2f/f', 'Var', (45, 53)) ('IL-6', 'Gene', '16193', (152, 156)) ('GM-CSF', 'Gene', '12981', (170, 176)) ('leukemic bone marrow', 'Disease', 'MESH:D001855', (6, 26)) ('leukemic bone marrow', 'Disease', (6, 26)) ('Mouse', 'Species', '10090', (0, 5)) ('mice', 'Species', '10090', (54, 58)) 543 30472158 Noncompetitive transplants were performed with 1 million BM cells from 6-8-wk-old Ikzf2fl/fl vavcre- or Ikzf2fl/fl vavcre+ mice, injected into lethally irradiated B6SJL congenic CD45.1 recipients. ('Ikzf2fl/fl', 'Var', (82, 92)) ('mice', 'Species', '10090', (123, 127)) ('CD45', 'Gene', (178, 182)) ('CD45', 'Gene', '19264', (178, 182)) 556 30472158 Murine leukemic cells were stained for Mac1-PB, Gr1-APC, F480-PE-Cy7, CD115-APC and c-Kit-APC-Cy7 and analyzed to assess differentiation status. ('c-Kit', 'Gene', '16590', (84, 89)) ('CD115', 'Gene', (70, 75)) ('Gr1', 'Gene', (48, 51)) ('leukemic', 'Disease', 'MESH:D007938', (7, 15)) ('F480-PE', 'Chemical', 'MESH:C082966', (57, 64)) ('APC', 'Chemical', 'MESH:C462103', (76, 79)) ('APC', 'Chemical', 'MESH:C462103', (90, 93)) ('APC', 'Chemical', 'MESH:C462103', (52, 55)) ('CD115', 'Gene', '12978', (70, 75)) ('Mac1', 'Gene', (39, 43)) ('F480-PE-Cy7', 'Var', (57, 68)) ('Mac1', 'Gene', '16409', (39, 43)) ('leukemic', 'Disease', (7, 15)) ('c-Kit', 'Gene', (84, 89)) ('Gr1', 'Gene', '546644', (48, 51)) 559 30472158 For deletion of Ikzf2, cells were treated with 10nM of 4-OHT Tamoxifen (Sigma-Aldrich) or ethanol. ('ethanol', 'Chemical', 'MESH:D000431', (90, 97)) ('4-OHT Tamoxifen', 'Chemical', 'MESH:C032278', (55, 70)) ('deletion', 'Var', (4, 12)) ('Ikzf2', 'Gene', (16, 21)) 565 30472158 10 000 sorted LSCs from Ikzf2fl/fl mice (n = 2) and Ikzf2Delta/Delta mice (n = 2) were processed for ATAC-sequencing. ('Ikzf2fl/fl', 'Var', (24, 34)) ('mice', 'Species', '10090', (69, 73)) ('Ikzf2Delta/Delta', 'Var', (52, 68)) ('mice', 'Species', '10090', (35, 39)) 569 30472158 MACS2 version 20160309 (URL https://github.com/taoliu/MACS) was used to call peaks from aligned reads, with the options extending the reads to both sides, assuming reads after removal of duplicates, and using a permissive p-value threshold (-p 1e-2 --nomodel --shift -100 --extsize 200 --keep-dup all). ('MACS2', 'Gene', '17357', (0, 5)) ('MACS2', 'Gene', (0, 5)) ('-p 1e-2', 'Var', (241, 248)) 573 30472158 Briefly, 250K MLL-AF9 leukemic cells harboring the Nras G12D (RN2 cells; kind gift from Dr. Vakoc, Cold Spring Harbor Laboratory) were transduced with retrovirus expressing FLAG-murine IKZF2 together with BFP. ('leukemic', 'Disease', (22, 30)) ('BFP', 'Gene', '22671', (205, 208)) ('IKZF2', 'Gene', (185, 190)) ('transduced', 'Reg', (135, 145)) ('murine', 'Species', '10090', (178, 184)) ('leukemic', 'Disease', 'MESH:D007938', (22, 30)) ('BFP', 'Gene', (205, 208)) ('G12D', 'Mutation', 'p.G12D', (56, 60)) ('Nras G12D', 'Var', (51, 60)) 581 30472158 Sorted c-KitHigh cells from Ikzf2 fl/fl mice (n = 2) and Ikzf2Delta/Delta mice (n = 2) were processed for RNA extraction using TRIZOL and RNeasy RNA extraction kit. ('Ikzf2 fl/fl', 'Var', (28, 39)) ('c-Kit', 'Gene', (7, 12)) ('c-Kit', 'Gene', '16590', (7, 12)) ('mice', 'Species', '10090', (40, 44)) ('mice', 'Species', '10090', (74, 78)) 594 29895953 Our results also reveal a correlation between amino acid variants in MICA external domains and ALL, AML, and MDS. ('amino acid variants', 'Var', (46, 65)) ('AML', 'Phenotype', 'HP:0004808', (100, 103)) ('MICA', 'Gene', (69, 73)) ('MICA', 'Gene', '100507436', (69, 73)) ('MDS', 'Disease', (109, 112)) ('MDS', 'Disease', 'MESH:D009190', (109, 112)) ('or', 'Gene', '18390', (27, 29)) ('AML', 'Disease', 'MESH:D015470', (100, 103)) ('MDS', 'Phenotype', 'HP:0002863', (109, 112)) ('AML', 'Disease', (100, 103)) 612 29895953 Interestingly, a methionine (ATG) to valine (GTG) change at position 129 of the alpha2-heavy chain domain categorized the MICA alleles into strong (MICA-129 met) and weak (MICA-129 val) binders of the NKG2D receptor. ('change', 'Var', (50, 56)) ('val', 'Chemical', 'MESH:D014633', (181, 184)) ('NKG2D receptor', 'Gene', (201, 215)) ('MICA', 'Gene', (172, 176)) ('MICA', 'Gene', (148, 152)) ('MICA', 'Gene', '100507436', (172, 176)) ('MICA', 'Gene', '100507436', (148, 152)) ('valine', 'Chemical', 'MESH:D014633', (37, 43)) ('or', 'Gene', '18390', (213, 215)) ('val', 'Chemical', 'MESH:D014633', (37, 40)) ('binders', 'Interaction', (186, 193)) ('NKG2D receptor', 'Gene', '22914', (201, 215)) ('methionine', 'Chemical', 'MESH:D008715', (17, 27)) ('MICA', 'Gene', (122, 126)) ('or', 'Gene', '18390', (111, 113)) ('MICA', 'Gene', '100507436', (122, 126)) ('GTG', 'Chemical', 'MESH:D006160', (45, 48)) 614 29895953 There are non-synonymous polymorphic sites covering exons 2-4, that encode non-conservative amino acid substitutions in MICA external domains. ('MICA', 'Gene', (120, 124)) ('MICA', 'Gene', '100507436', (120, 124)) ('substitutions', 'Var', (103, 116)) ('or', 'Gene', '18390', (30, 32)) 615 29895953 Codon 14 of MICA protein including MICA*002:01 showed strong odds ratios (1.8 ~ 2.7) and corrected P-value (Pc < 0.048). ('Codon', 'Var', (0, 5)) ('MICA', 'Gene', (35, 39)) ('MICA', 'Gene', '100507436', (35, 39)) ('MICA', 'Gene', (12, 16)) ('MICA', 'Gene', '100507436', (12, 16)) ('val', 'Chemical', 'MESH:D014633', (101, 104)) ('or', 'Gene', '18390', (90, 92)) 624 29895953 We can infer that, at least to some extent, the allelic repertoire of MICA represents an evolutionary record of past pathogen-driven selection and that the protective role of MICA*008:01 and A5.1 casts a 'selective' effect on the distribution of this allele. ('or', 'Gene', '18390', (105, 107)) ('MICA', 'Gene', (70, 74)) ('MICA', 'Gene', '100507436', (70, 74)) ('MICA', 'Gene', (175, 179)) ('MICA', 'Gene', '100507436', (175, 179)) ("'selective'", 'PosReg', (204, 215)) ('A5.1', 'Var', (191, 195)) 634 29563537 Using variables identified in multivariable models and co-occurring mutations in NPM1-mutated AML, we classified the patients into good-, intermediate- and poor-risk groups for complete remission (CR) attainment, disease-free (DFS) and overall survival (OS). ('AML', 'Disease', (94, 97)) ('OS', 'Chemical', '-', (254, 256)) ('disease-free', 'Disease', (213, 225)) ('AML', 'Phenotype', 'HP:0004808', (94, 97)) ('complete', 'Disease', (177, 185)) ('NPM1', 'Gene', (81, 85)) ('CR', 'Chemical', '-', (197, 199)) ('patients', 'Species', '9606', (117, 125)) ('overall survival', 'CPA', (236, 252)) ('mutations', 'Var', (68, 77)) ('AML', 'Disease', 'MESH:D015470', (94, 97)) ('NPM1', 'Gene', '4869', (81, 85)) 635 29563537 Whereas 81% of good-risk patients (comprising NPM1-mutated patients harboring mutations in chromatin remodeling, cohesin complex, methylation-related, spliceosome, and/or RAS pathway genes, FLT3-TKD, and/or patients without FLT3-ITD) achieved a CR, only 32% of poor-risk patients (with U2AF1, WT1 mutations and/or complex karyotype) did. ('FLT3', 'Gene', '2322', (190, 194)) ('NPM1', 'Gene', (46, 50)) ('spliceosome', 'Gene', (151, 162)) ('mutations', 'Var', (78, 87)) ('patients', 'Species', '9606', (59, 67)) ('FLT3', 'Gene', (224, 228)) ('chromatin remodeling', 'Gene', (91, 111)) ('U2AF1', 'Gene', (286, 291)) ('patients', 'Species', '9606', (25, 33)) ('FLT3', 'Gene', '2322', (224, 228)) ('patients', 'Species', '9606', (271, 279)) ('CR', 'Chemical', '-', (245, 247)) ('WT1', 'Gene', (293, 296)) ('U2AF1', 'Gene', '7307', (286, 291)) ('patients', 'Species', '9606', (207, 215)) ('achieved', 'PosReg', (234, 242)) ('NPM1', 'Gene', '4869', (46, 50)) ('cohesin complex', 'Gene', (113, 128)) ('FLT3', 'Gene', (190, 194)) ('WT1', 'Gene', '7490', (293, 296)) 637 29563537 Similarly, using NPM1 co-mutation patterns and SF1 mutation status, we identified patients with favorable DFS and OS 3-year rates of 46% and 45%, respectively. ('SF1', 'Gene', '7536', (47, 50)) ('OS', 'Chemical', '-', (114, 116)) ('patients', 'Species', '9606', (82, 90)) ('NPM1', 'Gene', (17, 21)) ('DFS', 'Disease', (106, 109)) ('NPM1', 'Gene', '4869', (17, 21)) ('mutation', 'Var', (51, 59)) ('SF1', 'Gene', (47, 50)) 644 29563537 Recently, large studies assessing biologic and prognostic significance of multiple gene mutations have increased our understanding of the molecular features of AML. ('AML', 'Disease', 'MESH:D015470', (160, 163)) ('increased', 'PosReg', (103, 112)) ('mutations', 'Var', (88, 97)) ('AML', 'Disease', (160, 163)) ('AML', 'Phenotype', 'HP:0004808', (160, 163)) 662 29563537 Gene mutations were assigned to functional groups similar to those previously described by the Cancer Genome Atlas Research Network as follows: chromatin remodeling (ASXL1, BCOR, BCORL1, EZH2 and SMARCA2), cohesin complex (RAD21, SMC1A, SMC3 and STAG2), kinases [AXL, FLT3 internal tandem duplications (FLT3-ITD), FLT3 tyrosine kinase domain mutations (FLT3-TKD), KIT and TYK2], methylation-related (DNMT3A, IDH1/2, and TET2), NPM1 (NPM1), RAS pathway (CBL, KRAS, NRAS and PTPN11), spliceosome (SF3B1, SRSF2, U2AF1 and ZRSR2), transcription factors (CEBPA, ETV6, GATA2, IKZF1, NOTCH1 and RUNX1) and tumor suppressors (PHF6, TP53 and WT1). ('FLT3', 'Gene', (268, 272)) ('ZRSR2', 'Gene', '8233', (519, 524)) ('NPM1', 'Gene', '4869', (427, 431)) ('PHF6', 'Gene', (618, 622)) ('tumor', 'Disease', (599, 604)) ('FLT3', 'Gene', '2322', (268, 272)) ('FLT3', 'Gene', (353, 357)) ('GATA2', 'Gene', (563, 568)) ('NPM1', 'Gene', '4869', (433, 437)) ('tumor', 'Disease', 'MESH:D009369', (599, 604)) ('FLT3', 'Gene', (314, 318)) ('FLT3', 'Gene', '2322', (353, 357)) ('NOTCH1', 'Gene', (577, 583)) ('DNMT3A', 'Gene', (400, 406)) ('FLT3', 'Gene', '2322', (314, 318)) ('U2AF1', 'Gene', (509, 514)) ('TYK2', 'Gene', (372, 376)) ('IDH1/2', 'Gene', (408, 414)) ('ZRSR2', 'Gene', (519, 524)) ('BCOR', 'Gene', '54880', (173, 177)) ('SRSF2', 'Gene', (502, 507)) ('PTPN11', 'Gene', '5781', (473, 479)) ('RUNX1', 'Gene', (588, 593)) ('RUNX1', 'Gene', '861', (588, 593)) ('CEBPA', 'Gene', '1050', (550, 555)) ('GATA2', 'Gene', '2624', (563, 568)) ('SF3B1', 'Gene', (495, 500)) ('STAG2', 'Gene', '10735', (246, 251)) ('RAD21', 'Gene', (223, 228)) ('CEBPA', 'Gene', (550, 555)) ('IKZF1', 'Gene', '10320', (570, 575)) ('ETV6', 'Gene', '2120', (557, 561)) ('SMC3', 'Gene', (237, 241)) ('NRAS', 'Gene', '4893', (464, 468)) ('SRSF2', 'Gene', '6427', (502, 507)) ('PTPN11', 'Gene', (473, 479)) ('BCORL1', 'Gene', '63035', (179, 185)) ('EZH2', 'Gene', '2146', (187, 191)) ('WT1', 'Gene', (633, 636)) ('IKZF1', 'Gene', (570, 575)) ('TET2', 'Gene', (420, 424)) ('WT1', 'Gene', '7490', (633, 636)) ('BCOR', 'Gene', (179, 183)) ('SMC3', 'Gene', '9126', (237, 241)) ('ASXL1', 'Gene', (166, 171)) ('PHF6', 'Gene', '84295', (618, 622)) ('TP53', 'Gene', (624, 628)) ('CBL', 'Gene', (453, 456)) ('STAG2', 'Gene', (246, 251)) ('RAD21', 'Gene', '5885', (223, 228)) ('SF3B1', 'Gene', '23451', (495, 500)) ('FLT3', 'Gene', (303, 307)) ('tumor', 'Phenotype', 'HP:0002664', (599, 604)) ('NPM1', 'Gene', (433, 437)) ('BCORL1', 'Gene', (179, 185)) ('EZH2', 'Gene', (187, 191)) ('ASXL1', 'Gene', '171023', (166, 171)) ('FLT3', 'Gene', '2322', (303, 307)) ('BCOR', 'Gene', '54880', (179, 183)) ('TP53', 'Gene', '7157', (624, 628)) ('U2AF1', 'Gene', '7307', (509, 514)) ('Cancer', 'Disease', (95, 101)) ('ETV6', 'Gene', (557, 561)) ('NRAS', 'Gene', (464, 468)) ('DNMT3A', 'Gene', '1788', (400, 406)) ('Cancer', 'Disease', 'MESH:D009369', (95, 101)) ('SMARCA2', 'Gene', (196, 203)) ('SMARCA2', 'Gene', '6595', (196, 203)) ('KRAS', 'Gene', (458, 462)) ('mutations', 'Var', (342, 351)) ('TET2', 'Gene', '54790', (420, 424)) ('SMC1A', 'Gene', (230, 235)) ('SMC1A', 'Gene', '8243', (230, 235)) ('IDH1/2', 'Gene', '3417;3418', (408, 414)) ('Cancer', 'Phenotype', 'HP:0002664', (95, 101)) ('NPM1', 'Gene', (427, 431)) ('NOTCH1', 'Gene', '4851', (577, 583)) ('CBL', 'Gene', '867', (453, 456)) ('TYK2', 'Gene', '7297', (372, 376)) ('BCOR', 'Gene', (173, 177)) ('KRAS', 'Gene', '3845', (458, 462)) 671 29563537 The most frequent were mutations in NPM1 (32% of patients), DNMT3A (27%), and TET2 (27%, Figure 1a). ('NPM1', 'Gene', (36, 40)) ('frequent', 'Reg', (9, 17)) ('DNMT3A', 'Gene', (60, 66)) ('TET2', 'Gene', '54790', (78, 82)) ('NPM1', 'Gene', '4869', (36, 40)) ('DNMT3A', 'Gene', '1788', (60, 66)) ('mutations', 'Var', (23, 32)) ('TET2', 'Gene', (78, 82)) ('patients', 'Species', '9606', (49, 57)) 672 29563537 Among previously defined functional groups, mutations involving methylation-related genes (found in 65% of patients), the spliceosome (38%) and NPM1 (32%) were predominant (Figure 1b, Supplementary Table S2). ('methylation-related genes', 'Gene', (64, 89)) ('NPM1', 'Gene', (144, 148)) ('mutations', 'Var', (44, 53)) ('NPM1', 'Gene', '4869', (144, 148)) ('patients', 'Species', '9606', (107, 115)) 676 29563537 In multivariable analysis of factors associated with the achievement of CR, mutated NPM1 (P<0.001) and the presence of a normal karyotype (P=0.03) were associated with higher probability of CR attainment (Table 2). ('NPM1', 'Gene', (84, 88)) ('CR', 'Chemical', '-', (72, 74)) ('CR attainment', 'CPA', (190, 203)) ('CR', 'Chemical', '-', (190, 192)) ('mutated', 'Var', (76, 83)) ('NPM1', 'Gene', '4869', (84, 88)) 677 29563537 Conversely, the presence of U2AF1 mutations (P=0.03) or WT1 mutations (P=0.01) were associated with lower CR rates. ('CR rates', 'CPA', (106, 114)) ('U2AF1', 'Gene', '7307', (28, 33)) ('lower', 'NegReg', (100, 105)) ('WT1', 'Gene', '7490', (56, 59)) ('WT1', 'Gene', (56, 59)) ('mutations', 'Var', (60, 69)) ('CR', 'Chemical', '-', (106, 108)) ('mutations', 'Var', (34, 43)) ('U2AF1', 'Gene', (28, 33)) 678 29563537 In multivariable analysis for DFS, the presence of NPM1 mutations (P<0.001) and SF1 mutations (P=0.04) associated with longer DFS, with 22% of NPM1-mutated patients and 40% of SF1-mutated patients being disease-free 3 years after diagnosis, compared with 8% of patients who did not harbor these mutations. ('patients', 'Species', '9606', (156, 164)) ('mutations', 'Var', (56, 65)) ('SF1', 'Gene', (176, 179)) ('NPM1', 'Gene', '4869', (51, 55)) ('NPM1', 'Gene', (143, 147)) ('DFS', 'MPA', (126, 129)) ('NPM1', 'Gene', (51, 55)) ('presence', 'Var', (39, 47)) ('mutations', 'Var', (84, 93)) ('SF1', 'Gene', (80, 83)) ('patients', 'Species', '9606', (188, 196)) ('NPM1', 'Gene', '4869', (143, 147)) ('SF1', 'Gene', '7536', (176, 179)) ('patients', 'Species', '9606', (261, 269)) ('SF1', 'Gene', '7536', (80, 83)) 679 29563537 The presence of FLT3-ITD (P<0.001), complex karyotype (P<0.001) and higher WBC counts (P=0.007) were associated with shorter DFS (Table 2). ('complex karyotype', 'Var', (36, 53)) ('FLT3', 'Gene', '2322', (16, 20)) ('higher', 'PosReg', (68, 74)) ('shorter', 'NegReg', (117, 124)) ('WBC counts', 'CPA', (75, 85)) ('FLT3', 'Gene', (16, 20)) ('DFS', 'MPA', (125, 128)) 680 29563537 In the multivariable analyses for OS, the presence of NPM1 mutations was the only feature associated with longer survival (P<0.001). ('NPM1', 'Gene', (54, 58)) ('longer', 'PosReg', (106, 112)) ('NPM1', 'Gene', '4869', (54, 58)) ('mutations', 'Var', (59, 68)) ('presence', 'Var', (42, 50)) ('OS', 'Chemical', '-', (34, 36)) 681 29563537 On the other hand, FLT3-ITD (P<0.001), BCOR mutations (P=0.003), TP53 mutation (P=0.003), t(9;11)(p22;q23) (P=0.03) and complex karyotype (P=0.002) were associated with poor OS (Table 2). ('TP53', 'Gene', '7157', (65, 69)) ('TP53', 'Gene', (65, 69)) ('complex karyotype', 'Var', (120, 137)) ('FLT3', 'Gene', '2322', (19, 23)) ('FLT3', 'Gene', (19, 23)) ('poor OS', 'Disease', (169, 176)) ('BCOR', 'Gene', (39, 43)) ('t(9;11)(p22;q23)', 'STRUCTURAL_ABNORMALITY', 'None', (90, 106)) ('OS', 'Chemical', '-', (174, 176)) ('BCOR', 'Gene', '54880', (39, 43)) ('t(9;11)(p22;q23', 'Var', (90, 105)) 682 29563537 We noted that CR rates of NPM1-mutated patients varied from 50% to 95% depending on the presence or absence of specific mutations coexisting with mutations in the NPM1 gene (Figure 1c). ('CR', 'Chemical', '-', (14, 16)) ('mutations', 'Var', (146, 155)) ('patients', 'Species', '9606', (39, 47)) ('NPM1', 'Gene', (26, 30)) ('NPM1', 'Gene', '4869', (163, 167)) ('NPM1', 'Gene', '4869', (26, 30)) ('NPM1', 'Gene', (163, 167)) 683 29563537 NPM1-mutated patients who additionally harbored a mutation in the chromatin remodeling gene or cohesin complex gene had CR rates of 89% and 95%, respectively. ('cohesin complex', 'Gene', (95, 110)) ('patients', 'Species', '9606', (13, 21)) ('NPM1', 'Gene', (0, 4)) ('CR', 'Chemical', '-', (120, 122)) ('NPM1', 'Gene', '4869', (0, 4)) ('mutation', 'Var', (50, 58)) 685 29563537 In contrast, CR rates of NPM1-mutated patients were reduced by the presence of tumor suppressor mutations (to 50%), kinase mutations other than FLT3-TKD (70%), the absence of any methylation-related mutation (65%), or the presence of a spliceosome mutation (69%). ('FLT3', 'Gene', (144, 148)) ('tumor', 'Phenotype', 'HP:0002664', (79, 84)) ('tumor', 'Disease', (79, 84)) ('reduced', 'NegReg', (52, 59)) ('absence', 'NegReg', (164, 171)) ('CR', 'Chemical', '-', (13, 15)) ('NPM1', 'Gene', (25, 29)) ('CR rates', 'CPA', (13, 21)) ('FLT3', 'Gene', '2322', (144, 148)) ('patients', 'Species', '9606', (38, 46)) ('tumor', 'Disease', 'MESH:D009369', (79, 84)) ('NPM1', 'Gene', '4869', (25, 29)) ('methylation-related', 'Protein', (179, 198)) ('mutations', 'Var', (123, 132)) ('mutations', 'Var', (96, 105)) 686 29563537 Interestingly, several of the mutations that enhanced the positive prognostic impact of NPM1 mutations had an adverse prognostic effect in patients with wild-type NPM1 (Figure 1d). ('NPM1', 'Gene', (88, 92)) ('mutations', 'Var', (93, 102)) ('patients', 'Species', '9606', (139, 147)) ('NPM1', 'Gene', '4869', (163, 167)) ('enhanced', 'PosReg', (45, 53)) ('NPM1', 'Gene', '4869', (88, 92)) ('mutations', 'Var', (30, 39)) ('positive prognostic', 'MPA', (58, 77)) ('NPM1', 'Gene', (163, 167)) 687 29563537 Specifically co-occurring mutations in chromatin remodeling genes, cohesin complex genes, FLT3-TKD or RAS pathway genes further lowered the overall CR rate of NPM1 wild-type patients from 44% to, respectively, 37%, 33%, 33% and 36%. ('NPM1', 'Gene', (159, 163)) ('chromatin remodeling genes', 'Gene', (39, 65)) ('NPM1', 'Gene', '4869', (159, 163)) ('CR', 'Chemical', '-', (148, 150)) ('FLT3', 'Gene', (90, 94)) ('RAS pathway genes', 'Gene', (102, 119)) ('patients', 'Species', '9606', (174, 182)) ('mutations', 'Var', (26, 35)) ('cohesin complex genes', 'Gene', (67, 88)) ('lowered', 'NegReg', (128, 135)) ('FLT3', 'Gene', '2322', (90, 94)) 688 29563537 Similarly, we assessed the prognostic impact of co-occurring mutations on DFS and OS. ('OS', 'Chemical', '-', (82, 84)) ('DFS', 'Disease', (74, 77)) ('mutations', 'Var', (61, 70)) 692 29563537 Of note, only IDH2 R140 mutations, but not IDH2 R172 mutations, co-occurred with NPM1. ('R140 mutations', 'Var', (19, 33)) ('co-occurred', 'Reg', (64, 75)) ('IDH2', 'Gene', (43, 47)) ('NPM1', 'Gene', (81, 85)) ('IDH2', 'Gene', (14, 18)) ('IDH2', 'Gene', '3418', (43, 47)) ('NPM1', 'Gene', '4869', (81, 85)) ('IDH2', 'Gene', '3418', (14, 18)) 693 29563537 Mutations in the ASXL1, IDH2, SF1, SMC1A, and SRSF2 genes are all early events, which have been previously implicated in clonal hematopoiesis. ('SMC1A', 'Gene', '8243', (35, 40)) ('hematopoiesis', 'Disease', 'MESH:C536227', (128, 141)) ('SF1', 'Gene', '7536', (30, 33)) ('SRSF2', 'Gene', '6427', (46, 51)) ('hematopoiesis', 'Disease', (128, 141)) ('IDH2', 'Gene', (24, 28)) ('Mutations', 'Var', (0, 9)) ('SF1', 'Gene', (30, 33)) ('SMC1A', 'Gene', (35, 40)) ('ASXL1', 'Gene', '171023', (17, 22)) ('SRSF2', 'Gene', (46, 51)) ('IDH2', 'Gene', '3418', (24, 28)) ('ASXL1', 'Gene', (17, 22)) 694 29563537 In our study, mutations in these genes frequently occurred at higher VAFs than the co-occurring NPM1 mutations (Figure 2c). ('NPM1', 'Gene', (96, 100)) ('NPM1', 'Gene', '4869', (96, 100)) ('mutations', 'Var', (14, 23)) ('occurred', 'Reg', (50, 58)) 695 29563537 In line with previous reports, this suggests that the mutation in NPM1 is acquired later during hematopoiesis, thereby possibly changing the fate of the leukemic clone with respect to its responsiveness to therapy. ('mutation', 'Var', (54, 62)) ('NPM1', 'Gene', '4869', (66, 70)) ('leukemic', 'Disease', (153, 161)) ('hematopoiesis', 'Disease', (96, 109)) ('hematopoiesis', 'Disease', 'MESH:C536227', (96, 109)) ('leukemic', 'Disease', 'MESH:D007938', (153, 161)) ('NPM1', 'Gene', (66, 70)) ('changing', 'Reg', (128, 136)) 697 29563537 The good-risk group included NPM1-mutated patients that simultaneously harbored mutations in chromatin remodeling genes, the cohesin complex, FLT3-TKD, methylation-related genes, spliceosome genes, RAS pathway genes, and/or patients that were NPM1 mutated and did not harbor FLT3-ITD mutations (n=127, 30% of the patient cohort). ('FLT3', 'Gene', (275, 279)) ('patient', 'Species', '9606', (224, 231)) ('NPM1', 'Gene', '4869', (29, 33)) ('patient', 'Species', '9606', (42, 49)) ('NPM1', 'Gene', '4869', (243, 247)) ('FLT3', 'Gene', '2322', (275, 279)) ('mutations', 'Var', (80, 89)) ('spliceosome genes', 'Gene', (179, 196)) ('methylation-related', 'Gene', (152, 171)) ('harbored', 'Reg', (71, 79)) ('NPM1', 'Gene', (29, 33)) ('NPM1', 'Gene', (243, 247)) ('chromatin remodeling genes', 'Gene', (93, 119)) ('patients', 'Species', '9606', (224, 232)) ('patients', 'Species', '9606', (42, 50)) ('mutated', 'Var', (248, 255)) ('FLT3', 'Gene', (142, 146)) ('RAS', 'Pathway', (198, 201)) ('patient', 'Species', '9606', (313, 320)) ('FLT3', 'Gene', '2322', (142, 146)) 698 29563537 The poor-risk group was characterized by patients harboring U2AF1 mutations, WT1 mutations and/or patients with typical complex karyotype or sole trisomy of chromosome 11 (n=95, 22% of patient cohort). ('mutations', 'Var', (66, 75)) ('WT1', 'Gene', (77, 80)) ('patient', 'Species', '9606', (98, 105)) ('patient', 'Species', '9606', (185, 192)) ('patients', 'Species', '9606', (98, 106)) ('patient', 'Species', '9606', (41, 48)) ('patients', 'Species', '9606', (41, 49)) ('U2AF1', 'Gene', '7307', (60, 65)) ('U2AF1', 'Gene', (60, 65)) ('mutations', 'Var', (81, 90)) ('WT1', 'Gene', '7490', (77, 80)) 705 29563537 The DFS good-risk group was comprised of NPM1-mutated patients that additionally harbored mutations in ASXL1, SF1, SMC1A or SRSF2, as well as SF1-mutated patients with wild-type NPM1. ('ASXL1', 'Gene', (103, 108)) ('mutations', 'Var', (90, 99)) ('SF1', 'Gene', (142, 145)) ('SRSF2', 'Gene', '6427', (124, 129)) ('SMC1A', 'Gene', '8243', (115, 120)) ('patients', 'Species', '9606', (54, 62)) ('SF1', 'Gene', (110, 113)) ('NPM1', 'Gene', '4869', (178, 182)) ('patients', 'Species', '9606', (154, 162)) ('SMC1A', 'Gene', (115, 120)) ('NPM1', 'Gene', (41, 45)) ('NPM1', 'Gene', (178, 182)) ('SF1', 'Gene', '7536', (110, 113)) ('SF1', 'Gene', '7536', (142, 145)) ('ASXL1', 'Gene', '171023', (103, 108)) ('NPM1', 'Gene', '4869', (41, 45)) ('SRSF2', 'Gene', (124, 129)) 707 29563537 The good-risk group for OS was defined by NPM1-mutated patients that also harbored mutations in chromatin remodeling genes, IDH2, SF1 and/or SRSF2. ('NPM1', 'Gene', '4869', (42, 46)) ('mutations', 'Var', (83, 92)) ('IDH2', 'Gene', '3418', (124, 128)) ('SRSF2', 'Gene', '6427', (141, 146)) ('OS', 'Chemical', '-', (24, 26)) ('SF1', 'Gene', '7536', (130, 133)) ('NPM1', 'Gene', (42, 46)) ('IDH2', 'Gene', (124, 128)) ('patients', 'Species', '9606', (55, 63)) ('SRSF2', 'Gene', (141, 146)) ('SF1', 'Gene', (130, 133)) 709 29563537 When specifically considering the genetic features associated with poor DFS and OS in both uni- and multivariable analyses [DFS: the presence of FLT3-ITD, RUNX1, TP53 or U2AF1 mutations and/or typical or atypical complex karyotype; OS: the presence of BCOR mutations, FLT3-ITD, TP53, U2AF1 or WT1 mutations, t(9;11), and/or typical complex karyotype], we further identified a patient subset with especially poor prognosis. ('FLT3', 'Gene', (268, 272)) ('mutations', 'Var', (257, 266)) ('U2AF1', 'Gene', (170, 175)) ('OS', 'Chemical', '-', (232, 234)) ('U2AF1', 'Gene', '7307', (284, 289)) ('FLT3', 'Gene', '2322', (268, 272)) ('RUNX1', 'Gene', (155, 160)) ('TP53', 'Gene', '7157', (278, 282)) ('RUNX1', 'Gene', '861', (155, 160)) ('OS', 'Chemical', '-', (80, 82)) ('TP53', 'Gene', (162, 166)) ('U2AF1', 'Gene', '7307', (170, 175)) ('U2AF1', 'Gene', (284, 289)) ('FLT3', 'Gene', (145, 149)) ('BCOR', 'Gene', '54880', (252, 256)) ('WT1', 'Gene', (293, 296)) ('FLT3', 'Gene', '2322', (145, 149)) ('TP53', 'Gene', (278, 282)) ('TP53', 'Gene', '7157', (162, 166)) ('BCOR', 'Gene', (252, 256)) ('WT1', 'Gene', '7490', (293, 296)) ('patient', 'Species', '9606', (376, 383)) 713 29563537 A recent, updated in 2017, version of the ELN recommendations contains a modified 3-group risk-stratification system based on the presence or absence of specific chromosome abnormalities and mutations in selected genes. ('ELN', 'Chemical', '-', (42, 45)) ('chromosome abnormalities', 'Phenotype', 'HP:0031411', (162, 186)) ('chromosome abnormalities', 'Disease', (162, 186)) ('mutations', 'Var', (191, 200)) ('genes', 'Gene', (213, 218)) ('chromosome abnormalities', 'Disease', 'MESH:D002869', (162, 186)) 715 29563537 As Figure 4 illustrates, the modified ELN Favorable-risk groups (i.e., without CBF-AML patients who are not included in our study) comprised almost four times as many patients for DFS and three times as many patients for OS than the respective good-risk groups for DFS and OS in our proposed genetic classification (Figure 3b and c). ('patients', 'Species', '9606', (208, 216)) ('ELN', 'Chemical', '-', (38, 41)) ('ELN', 'Gene', (38, 41)) ('CBF-AML', 'Disease', 'MESH:D015470', (79, 86)) ('AML', 'Phenotype', 'HP:0004808', (83, 86)) ('patients', 'Species', '9606', (167, 175)) ('OS', 'Chemical', '-', (273, 275)) ('OS', 'Chemical', '-', (221, 223)) ('DFS', 'Disease', (180, 183)) ('CBF-AML', 'Disease', (79, 86)) ('patients', 'Species', '9606', (87, 95)) ('modified', 'Var', (29, 37)) 717 29563537 The 3-year DFS rates were 25% for the modified ELN Favorable group, 9% for ELN Intermediate, and 3% for ELN Adverse-risk groups. ('ELN', 'Chemical', '-', (104, 107)) ('DFS', 'MPA', (11, 14)) ('modified', 'Var', (38, 46)) ('ELN', 'Chemical', '-', (75, 78)) ('ELN', 'Chemical', '-', (47, 50)) 721 29563537 The use of our model resulted in splitting the modified 2017 ELN Favorable-risk group into three: the good-risk group (comprising 21 patients for DFS and 38 for OS); the intermediate-risk group (comprising 57 and 62 patients, respectively) and the poor-risk group (comprising 21 patients for both DFS and OS). ('OS', 'Chemical', '-', (161, 163)) ('modified', 'Var', (47, 55)) ('patients', 'Species', '9606', (133, 141)) ('patients', 'Species', '9606', (216, 224)) ('OS', 'Chemical', '-', (305, 307)) ('2017 ELN', 'Gene', (56, 64)) ('patients', 'Species', '9606', (279, 287)) ('ELN', 'Chemical', '-', (61, 64)) 730 29563537 In line with previous studies, NPM1 mutations were the dominating marker in the older AML patient population. ('NPM1', 'Gene', '4869', (31, 35)) ('AML', 'Phenotype', 'HP:0004808', (86, 89)) ('AML', 'Disease', (86, 89)) ('mutations', 'Var', (36, 45)) ('patient', 'Species', '9606', (90, 97)) ('AML', 'Disease', 'MESH:D015470', (86, 89)) ('NPM1', 'Gene', (31, 35)) 731 29563537 NPM1 mutations were not only the most frequent, but also the only mutation whose presence is associated with a higher likelihood of achieving a CR by patients receiving standard induction chemotherapy. ('mutations', 'Var', (5, 14)) ('patients', 'Species', '9606', (150, 158)) ('NPM1', 'Gene', (0, 4)) ('NPM1', 'Gene', '4869', (0, 4)) ('CR', 'Chemical', '-', (144, 146)) 732 29563537 NPM1 mutations also confer longer DFS and OS. ('DFS', 'CPA', (34, 37)) ('OS', 'Chemical', '-', (42, 44)) ('mutations', 'Var', (5, 14)) ('longer', 'PosReg', (27, 33)) ('NPM1', 'Gene', (0, 4)) ('NPM1', 'Gene', '4869', (0, 4)) 733 29563537 However, we have shown that the presence or absence of additional mutations, such as mutations in the chromatin remodeling, cohesin complex, methylation-related and RAS pathway genes for CR achievement, mutations in the ASXL1, SF1, SMC1A and SRSF2 genes for DFS and ASXL1, IDH2, SF1 and SRSF2 mutations for OS, can influence the prognostic impact of NPM1 mutation. ('SF1', 'Gene', (227, 230)) ('NPM1', 'Gene', (350, 354)) ('SF1', 'Gene', '7536', (227, 230)) ('ASXL1', 'Gene', '171023', (220, 225)) ('SMC1A', 'Gene', (232, 237)) ('SF1', 'Gene', (279, 282)) ('ASXL1', 'Gene', '171023', (266, 271)) ('SMC1A', 'Gene', '8243', (232, 237)) ('prognostic impact', 'MPA', (329, 346)) ('SRSF2', 'Gene', '6427', (242, 247)) ('SF1', 'Gene', '7536', (279, 282)) ('SRSF2', 'Gene', (242, 247)) ('ASXL1', 'Gene', (220, 225)) ('IDH2', 'Gene', (273, 277)) ('influence', 'Reg', (315, 324)) ('SRSF2', 'Gene', '6427', (287, 292)) ('IDH2', 'Gene', '3418', (273, 277)) ('ASXL1', 'Gene', (266, 271)) ('NPM1', 'Gene', '4869', (350, 354)) ('mutations', 'Var', (203, 212)) ('SRSF2', 'Gene', (287, 292)) ('OS', 'Chemical', '-', (307, 309)) ('CR', 'Chemical', '-', (187, 189)) ('mutation', 'Var', (355, 363)) 736 29563537 We have shown that patients fit for chemotherapy who harbored NPM1 mutations together with such co-occurring mutations as those in chromatin remodeling, cohesin complex, FLT3-TKD, methylation-related, RAS pathway or spliceosome genes, or without FLT3-ITD constitute a good-risk group whose chance of achieving a CR was 81%. ('NPM1', 'Gene', '4869', (62, 66)) ('FLT3', 'Gene', '2322', (170, 174)) ('patients', 'Species', '9606', (19, 27)) ('FLT3', 'Gene', '2322', (246, 250)) ('mutations', 'Var', (67, 76)) ('FLT3', 'Gene', (170, 174)) ('NPM1', 'Gene', (62, 66)) ('FLT3', 'Gene', (246, 250)) ('CR', 'Chemical', '-', (312, 314)) 738 29563537 In contrast, patients with U2AF1 or WT1 mutations, sole trisomy 11 and/or a typical complex karyotype, of whom only 32% achieved a CR, should receive alternative treatment regimens, if possible. ('U2AF1', 'Gene', (27, 32)) ('CR', 'Chemical', '-', (131, 133)) ('U2AF1', 'Gene', '7307', (27, 32)) ('patients', 'Species', '9606', (13, 21)) ('mutations', 'Var', (40, 49)) ('WT1', 'Gene', '7490', (36, 39)) ('WT1', 'Gene', (36, 39)) 745 29563537 Again, those good-risk groups were driven by the presence of NPM1 mutations, in combination with other mutations. ('mutations', 'Var', (66, 75)) ('NPM1', 'Gene', '4869', (61, 65)) ('NPM1', 'Gene', (61, 65)) 752 29563537 However, we believe that our approach reflects the complex biology of the disease, in which the same cytogenetic or molecular marker, such as, for example mutated RUNX1, may be associated with worse DFS but not with CR attainment or OS. ('mutated', 'Var', (155, 162)) ('OS', 'Chemical', '-', (233, 235)) ('RUNX1', 'Gene', (163, 168)) ('RUNX1', 'Gene', '861', (163, 168)) ('DFS', 'Disease', (199, 202)) ('associated', 'Reg', (177, 187)) ('CR', 'Chemical', '-', (216, 218)) 755 29563537 We were able to identify combinations of mutations associated with favorable outcome in subsets of older AML patients. ('mutations', 'Var', (41, 50)) ('AML', 'Disease', 'MESH:D015470', (105, 108)) ('patients', 'Species', '9606', (109, 117)) ('AML', 'Phenotype', 'HP:0004808', (105, 108)) ('AML', 'Disease', (105, 108)) 758 27621679 IDH1/2 mutations define distinct subsets of cancers, including low-grade gliomas and secondary glioblastomas, chondrosarcomas, intrahepatic cholangiocarcinomas, and hematologic malignancies. ('gliomas', 'Phenotype', 'HP:0009733', (73, 80)) ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('glioblastomas', 'Disease', (95, 108)) ('chondrosarcomas', 'Disease', (110, 125)) ('intrahepatic cholangiocarcinomas', 'Disease', (127, 159)) ('IDH1/2', 'Gene', (0, 6)) ('hematologic malignancies', 'Disease', (165, 189)) ('glioblastomas', 'Disease', 'MESH:D005909', (95, 108)) ('cancers', 'Disease', 'MESH:D009369', (44, 51)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (165, 189)) ('intrahepatic cholangiocarcinomas', 'Disease', 'MESH:D018281', (127, 159)) ('mutations', 'Var', (7, 16)) ('gliomas', 'Disease', (73, 80)) ('carcinoma', 'Phenotype', 'HP:0030731', (149, 158)) ('chondrosarcoma', 'Phenotype', 'HP:0006765', (110, 124)) ('glioblastomas', 'Phenotype', 'HP:0012174', (95, 108)) ('chondrosarcomas', 'Phenotype', 'HP:0006765', (110, 125)) ('gliomas', 'Disease', 'MESH:D005910', (73, 80)) ('cancers', 'Phenotype', 'HP:0002664', (44, 51)) ('cancers', 'Disease', (44, 51)) ('glioblastoma', 'Phenotype', 'HP:0012174', (95, 107)) ('cholangiocarcinoma', 'Phenotype', 'HP:0030153', (140, 158)) ('chondrosarcomas', 'Disease', 'MESH:D002813', (110, 125)) ('glioma', 'Phenotype', 'HP:0009733', (73, 79)) ('cancer', 'Phenotype', 'HP:0002664', (44, 50)) 759 27621679 Somatic point mutations in IDH1/2 confer a gain-of-function in cancer cells, resulting in the accumulation and secretion in vast excess of an oncometabolite, the D-2-hydroxyglutarate (D-2HG). ('D-2-hydroxyglutarate', 'Chemical', 'MESH:C019417', (162, 182)) ('D-2-hydroxyglutarate', 'MPA', (162, 182)) ('gain-of-function', 'PosReg', (43, 59)) ('IDH1/2', 'Gene', '3417;3418', (27, 33)) ('cancer', 'Phenotype', 'HP:0002664', (63, 69)) ('point mutations', 'Var', (8, 23)) ('secretion', 'MPA', (111, 120)) ('cancer', 'Disease', 'MESH:D009369', (63, 69)) ('IDH1/2', 'Gene', (27, 33)) ('accumulation', 'MPA', (94, 106)) ('cancer', 'Disease', (63, 69)) 760 27621679 Indeed, high levels of D-2HG inhibit alpha-ketoglutarate-dependent dioxygenases, including histone and DNA demethylases, leading to histone and DNA hypermethylation and finally a block in cell differentiation. ('alpha-ketoglutarate-dependent dioxygenases', 'Enzyme', (37, 79)) ('DNA demethylases', 'Enzyme', (103, 119)) ('hypermethylation', 'MPA', (148, 164)) ('alpha-ketoglutarate', 'Chemical', 'MESH:D007656', (37, 56)) ('histone', 'MPA', (132, 139)) ('inhibit', 'NegReg', (29, 36)) ('D-2HG', 'Var', (23, 28)) ('histone', 'Enzyme', (91, 98)) ('cell differentiation', 'CPA', (188, 208)) ('block', 'NegReg', (179, 184)) 761 27621679 Furthermore, D-2HG is a biomarker suitable for the detection of IDH1/2 mutations at diagnosis and predictive of the clinical response. ('IDH1/2', 'Gene', '3417;3418', (64, 70)) ('mutations', 'Var', (71, 80)) ('IDH1/2', 'Gene', (64, 70)) 762 27621679 Finally, mutant-IDH1/2 enzymes inhibitors have entered clinical trials for patients with IDH1/2 mutations and represent a novel drug class for targeted therapy. ('IDH1/2', 'Gene', '3417;3418', (89, 95)) ('IDH1/2', 'Gene', (16, 22)) ('patients', 'Species', '9606', (75, 83)) ('IDH1/2', 'Gene', (89, 95)) ('IDH1/2', 'Gene', '3417;3418', (16, 22)) ('mutations', 'Var', (96, 105)) 764 27621679 Point mutations in IDH1/2 define distinct subsets of low-grade glioma and secondary glioblastoma (GBM), chondrosarcoma, intrahepatic cholangiocarcinomas, hematologic malignancies, as well as premalignant diseases and rare inherited metabolism disorders. ('chondrosarcoma', 'Disease', 'MESH:D002813', (104, 118)) ('chondrosarcoma', 'Disease', (104, 118)) ('glioma', 'Phenotype', 'HP:0009733', (63, 69)) ('intrahepatic cholangiocarcinomas', 'Disease', (120, 152)) ('inherited metabolism disorders', 'Disease', 'MESH:D008661', (222, 252)) ('glioblastoma', 'Disease', (84, 96)) ('carcinoma', 'Phenotype', 'HP:0030731', (142, 151)) ('glioblastoma', 'Phenotype', 'HP:0012174', (84, 96)) ('inherited metabolism disorders', 'Disease', (222, 252)) ('chondrosarcoma', 'Phenotype', 'HP:0006765', (104, 118)) ('hematologic malignancies', 'Disease', (154, 178)) ('intrahepatic cholangiocarcinomas', 'Disease', 'MESH:D018281', (120, 152)) ('Point mutations', 'Var', (0, 15)) ('premalignant diseases', 'Disease', (191, 212)) ('GBM', 'Phenotype', 'HP:0012174', (98, 101)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (154, 178)) ('IDH1/2', 'Gene', '3417;3418', (19, 25)) ('glioma', 'Disease', (63, 69)) ('cholangiocarcinoma', 'Phenotype', 'HP:0030153', (133, 151)) ('IDH1/2', 'Gene', (19, 25)) ('glioma', 'Disease', 'MESH:D005910', (63, 69)) ('glioblastoma', 'Disease', 'MESH:D005909', (84, 96)) 765 27621679 Multiple preclinical models have provided evidence for the oncogenic potential of IDH1/2 mutations, which alter epigenetic regulation, cancer cell differentiation, and metabolism. ('cancer', 'Disease', (135, 141)) ('IDH1/2', 'Gene', '3417;3418', (82, 88)) ('epigenetic regulation', 'MPA', (112, 133)) ('alter', 'Reg', (106, 111)) ('cancer', 'Phenotype', 'HP:0002664', (135, 141)) ('IDH1/2', 'Gene', (82, 88)) ('mutations', 'Var', (89, 98)) ('metabolism', 'MPA', (168, 178)) ('cancer', 'Disease', 'MESH:D009369', (135, 141)) 766 27621679 Depending on the associated genomic aberrations and the cellular context, the oncogenic potential of IDH1/2 mutations ranges from an initiating event - promoting transformation - to a secondary oncogenic event conferring selective advantage to cancer cells. ('mutations', 'Var', (108, 117)) ('IDH1/2', 'Gene', (101, 107)) ('cancer', 'Phenotype', 'HP:0002664', (244, 250)) ('IDH1/2', 'Gene', '3417;3418', (101, 107)) ('cancer', 'Disease', (244, 250)) ('cancer', 'Disease', 'MESH:D009369', (244, 250)) 767 27621679 In vitro and in vivo preclinical studies have demonstrated that inhibition of IDH1/2-mutant enzymes decreases intracellular D-2-hydroxyglutarate (D-2HG) levels, reverses epigenetic dysregulation, and releases the differentiation block. ('differentiation', 'MPA', (213, 228)) ('reverses', 'NegReg', (161, 169)) ('IDH1/2', 'Gene', '3417;3418', (78, 84)) ('D-2-hydroxyglutarate', 'Chemical', 'MESH:C019417', (124, 144)) ('IDH1/2', 'Gene', (78, 84)) ('inhibition', 'Var', (64, 74)) ('enzymes', 'Gene', (92, 99)) ('decreases', 'NegReg', (100, 109)) ('epigenetic dysregulation', 'MPA', (170, 194)) ('releases', 'PosReg', (200, 208)) 777 27621679 IDH1/2 mutations are heterozygous, missense mutations, leading to the substitution of the amino acids arginine 132 in IDH1 and arginine 172 or 140 in IDH2 (Table 1). ('IDH1', 'Gene', '3417', (0, 4)) ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('IDH1', 'Gene', (118, 122)) ('arginine', 'Chemical', 'MESH:D001120', (127, 135)) ('arginine', 'Chemical', 'MESH:D001120', (102, 110)) ('arginine 172 or 140', 'Var', (127, 146)) ('IDH1', 'Gene', '3417', (118, 122)) ('IDH2', 'Gene', (150, 154)) ('IDH1', 'Gene', (0, 4)) ('IDH1/2', 'Gene', (0, 6)) ('IDH2', 'Gene', '3418', (150, 154)) ('mutations', 'Var', (7, 16)) 778 27621679 Initial functional studies of IDH1/2-mutant cells revealed that IDH1/2 mutations decrease the ability of the mutant enzymes to convert isocitrate to alphaKG3 and that IDH1/2-mutant enzymes inhibit wild-type activity in a dominant-negative manner. ('IDH1/2', 'Gene', '3417;3418', (64, 70)) ('convert isocitrate', 'MPA', (127, 145)) ('IDH1/2', 'Gene', (167, 173)) ('decrease', 'NegReg', (81, 89)) ('IDH1/2', 'Gene', (30, 36)) ('IDH1/2', 'Gene', (64, 70)) ('isocitrate', 'Chemical', 'MESH:C034219', (135, 145)) ('mutations', 'Var', (71, 80)) ('IDH1/2', 'Gene', '3417;3418', (167, 173)) ('IDH1/2', 'Gene', '3417;3418', (30, 36)) 780 27621679 IDH1/2 mutant enzymes produce high levels of D-2HG in cells (50- to 100-fold higher than in normal tissues). ('mutant', 'Var', (7, 13)) ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('enzymes', 'Enzyme', (14, 21)) ('D-2HG', 'MPA', (45, 50)) ('IDH1/2', 'Gene', (0, 6)) 781 27621679 IDH1/2 mutants exclusively produce D-2HG. ('mutants', 'Var', (7, 14)) ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('IDH1/2', 'Gene', (0, 6)) ('D-2HG', 'MPA', (35, 40)) ('produce', 'Reg', (27, 34)) 784 27621679 In vitro ectopic expression of IDH1/2 mutants produces high D-2HG levels that inhibit alphaKG-dependent dioxygenases, including histone demethylases Jumonji 2 (JMJD2) and Jmj C domain-containing histone demethylase-1 (JHDM1), and DNA demethylases ten-eleven translocation 2 (TET2) proteins, resulting in the impairment of key steps in histone and DNA demethylation. ('JMJD2', 'Gene', '9682', (160, 165)) ('mutants', 'Var', (38, 45)) ('IDH1/2', 'Gene', '3417;3418', (31, 37)) ('TET2', 'Gene', '54790', (275, 279)) ('inhibit', 'NegReg', (78, 85)) ('key steps', 'MPA', (322, 331)) ('Jmj C', 'Var', (171, 176)) ('JMJD2', 'Gene', (160, 165)) ('DNA demethylation', 'MPA', (347, 364)) ('histone', 'MPA', (335, 342)) ('impairment', 'NegReg', (308, 318)) ('IDH1/2', 'Gene', (31, 37)) ('TET2', 'Gene', (275, 279)) ('alphaKG-dependent dioxygenases', 'Enzyme', (86, 116)) 785 27621679 The epigenetic deregulation induced by IDH1/2 mutant enzymes translates into histone and DNA hypermethylation in a passage-dependent manner. ('epigenetic deregulation', 'MPA', (4, 27)) ('IDH1/2', 'Gene', (39, 45)) ('DNA hypermethylation', 'MPA', (89, 109)) ('mutant', 'Var', (46, 52)) ('histone', 'MPA', (77, 84)) ('enzymes', 'Enzyme', (53, 60)) ('IDH1/2', 'Gene', '3417;3418', (39, 45)) 790 27621679 Consequently, as observed in TET2 mutants, increased expression of stem cell markers and impaired differentiation are seen in cells expressing IDH1/2 mutant enzymes. ('TET2', 'Gene', '54790', (29, 33)) ('mutant enzymes', 'Var', (150, 164)) ('IDH1/2', 'Gene', '3417;3418', (143, 149)) ('TET2', 'Gene', (29, 33)) ('increased', 'PosReg', (43, 52)) ('IDH1/2', 'Gene', (143, 149)) ('impaired', 'NegReg', (89, 97)) ('differentiation', 'CPA', (98, 113)) ('expression', 'MPA', (53, 63)) 791 27621679 Hypermethylation can also compromise the binding of methylation-sensitive insulator proteins, which may result in the loss of insulation between topological DNA domains and aberrant gene activation, as recently demonstrated in IDH1-mutant gliomasphere models. ('insulation', 'MPA', (126, 136)) ('activation', 'PosReg', (187, 197)) ('compromise', 'NegReg', (26, 36)) ('Hypermethylation', 'Var', (0, 16)) ('binding', 'Interaction', (41, 48)) ('IDH1', 'Gene', '3417', (227, 231)) ('gliomas', 'Disease', 'MESH:D005910', (239, 246)) ('loss', 'NegReg', (118, 122)) ('gliomas', 'Phenotype', 'HP:0009733', (239, 246)) ('gliomas', 'Disease', (239, 246)) ('IDH1', 'Gene', (227, 231)) ('glioma', 'Phenotype', 'HP:0009733', (239, 245)) 794 27621679 Mutations in D2HGDH and IDH2 are the molecular basis of this metabolic disorder, with a tendency to a more severe clinical phenotype in IDH2-mutant patients. ('patients', 'Species', '9606', (148, 156)) ('D2HGDH', 'Gene', (13, 19)) ('IDH2', 'Gene', (136, 140)) ('D2HGDH', 'Gene', '728294', (13, 19)) ('basis', 'Reg', (47, 52)) ('IDH2', 'Gene', (24, 28)) ('Mutations', 'Var', (0, 9)) ('IDH2', 'Gene', '3418', (136, 140)) ('IDH2', 'Gene', '3418', (24, 28)) ('metabolic disorder', 'Phenotype', 'HP:0001939', (61, 79)) 795 27621679 Interestingly, although D-2HG levels are excessively high in patients with D-2HGA, no cancers have been reported so far in this population, which suggests that D-2HG accumulation is not sufficient alone to induce cancer. ('cancer', 'Phenotype', 'HP:0002664', (213, 219)) ('high', 'PosReg', (53, 57)) ('cancer', 'Phenotype', 'HP:0002664', (86, 92)) ('patients', 'Species', '9606', (61, 69)) ('D-2HGA', 'Var', (75, 81)) ('cancer', 'Disease', 'MESH:D009369', (213, 219)) ('cancers', 'Phenotype', 'HP:0002664', (86, 93)) ('D-2HG', 'MPA', (24, 29)) ('cancer', 'Disease', (213, 219)) ('cancers', 'Disease', (86, 93)) ('cancer', 'Disease', 'MESH:D009369', (86, 92)) ('cancers', 'Disease', 'MESH:D009369', (86, 93)) ('cancer', 'Disease', (86, 92)) 796 27621679 Indeed, while expression of mutant IDH1/2 enzymes was associated with increased progenitor cell marker expression and impaired cell differentiation, such epigenetics effects were not sufficient alone to drive oncogenesis. ('impaired', 'NegReg', (118, 126)) ('cell differentiation', 'CPA', (127, 147)) ('mutant', 'Var', (28, 34)) ('increased', 'PosReg', (70, 79)) ('progenitor cell marker', 'CPA', (80, 102)) ('IDH1/2', 'Gene', '3417;3418', (35, 41)) ('enzymes', 'Gene', (42, 49)) ('IDH1/2', 'Gene', (35, 41)) 798 27621679 For example, excess of succinate is observed in case of mutations affecting genes coding for succinate dehydrogenase. ('mutations', 'Var', (56, 65)) ('succinate', 'Chemical', 'MESH:D019802', (93, 102)) ('succinate', 'MPA', (23, 32)) ('succinate', 'Chemical', 'MESH:D019802', (23, 32)) ('excess', 'PosReg', (13, 19)) 799 27621679 Similar to IDH1/2 mutations, such alterations have been linked with DNA hypermethylation, decreased expression of key genes involved in neuroendocrine differentiation, and a block in cellular differentiation. ('IDH1/2', 'Gene', '3417;3418', (11, 17)) ('DNA', 'MPA', (68, 71)) ('decreased', 'NegReg', (90, 99)) ('expression', 'MPA', (100, 110)) ('cellular differentiation', 'CPA', (183, 207)) ('IDH1/2', 'Gene', (11, 17)) ('alterations', 'Var', (34, 45)) ('block', 'NegReg', (174, 179)) ('mutations', 'Var', (18, 27)) 800 27621679 Few investigations have questioned metabolic changes associated with IDH1/2 mutations. ('metabolic', 'MPA', (35, 44)) ('IDH1/2', 'Gene', '3417;3418', (69, 75)) ('mutations', 'Var', (76, 85)) ('IDH1/2', 'Gene', (69, 75)) 801 27621679 Recent metabolic flux analyses have shown that IDH1 mutations render tumor cells more dependent on mitochondrial oxidative tricarboxylic acid metabolism and compromise the conversion of glutamine to citrate and fatty acids under hypoxia as compared to IDH1-wild type cells. ('mutations', 'Var', (52, 61)) ('tumor', 'Phenotype', 'HP:0002664', (69, 74)) ('hypoxia', 'Disease', (229, 236)) ('hypoxia', 'Disease', 'MESH:D000860', (229, 236)) ('glutamine', 'Chemical', 'MESH:D005973', (186, 195)) ('tumor', 'Disease', (69, 74)) ('IDH1', 'Gene', (47, 51)) ('more', 'PosReg', (81, 85)) ('dependent', 'MPA', (86, 95)) ('compromise', 'NegReg', (157, 167)) ('IDH1', 'Gene', (252, 256)) ('tricarboxylic acid', 'Chemical', 'MESH:D014233', (123, 141)) ('fatty acids', 'Chemical', 'MESH:D005227', (211, 222)) ('IDH1', 'Gene', '3417', (252, 256)) ('IDH1', 'Gene', '3417', (47, 51)) ('tumor', 'Disease', 'MESH:D009369', (69, 74)) ('citrate', 'Chemical', 'MESH:D019343', (199, 206)) 803 27621679 Of note, D-2HG inhibits both ATP synthase and mTOR signaling in glioma cells, resulting in growth arrest and cell death under conditions of glucose limitation. ('glioma', 'Phenotype', 'HP:0009733', (64, 70)) ('cell death', 'CPA', (109, 119)) ('ATP', 'Enzyme', (29, 32)) ('growth arrest', 'Disease', 'MESH:D006323', (91, 104)) ('mTOR', 'Gene', '2475', (46, 50)) ('growth arrest', 'Disease', (91, 104)) ('growth arrest', 'Phenotype', 'HP:0001510', (91, 104)) ('mTOR', 'Gene', (46, 50)) ('glioma', 'Disease', (64, 70)) ('inhibits', 'NegReg', (15, 23)) ('D-2HG', 'Var', (9, 14)) ('glucose', 'Chemical', 'MESH:D005947', (140, 147)) ('glucose limitation', 'Phenotype', 'HP:0040270', (140, 158)) ('glioma', 'Disease', 'MESH:D005910', (64, 70)) 805 27621679 Overall, IDH1/2 mutations are likely associated with wide metabolic reprogramming. ('IDH1/2', 'Gene', (9, 15)) ('mutations', 'Var', (16, 25)) ('IDH1/2', 'Gene', '3417;3418', (9, 15)) ('associated', 'Reg', (37, 47)) ('wide metabolic reprogramming', 'CPA', (53, 81)) 806 27621679 Further deciphering of alterations in cellular metabolism associated with IDH1/2 mutations should highlight novel opportunities for therapeutic intervention and drug development. ('cellular metabolism', 'MPA', (38, 57)) ('IDH1/2', 'Gene', (74, 80)) ('mutations', 'Var', (81, 90)) ('alterations', 'Reg', (23, 34)) ('IDH1/2', 'Gene', '3417;3418', (74, 80)) 807 27621679 First identified in colorectal cancer, IDH1/2 mutations affecting IDH1 (R132) or IDH2 (R140, R172) are found across a broad spectrum of cancer types (Table 1). ('IDH1/2', 'Gene', (39, 45)) ('R140', 'Var', (87, 91)) ('cancer', 'Disease', (136, 142)) ('cancer', 'Phenotype', 'HP:0002664', (136, 142)) ('IDH1', 'Gene', '3417', (66, 70)) ('colorectal cancer', 'Phenotype', 'HP:0003003', (20, 37)) ('mutations', 'Var', (46, 55)) ('IDH1', 'Gene', (39, 43)) ('cancer', 'Disease', (31, 37)) ('cancer', 'Phenotype', 'HP:0002664', (31, 37)) ('cancer', 'Disease', 'MESH:D009369', (136, 142)) ('IDH1', 'Gene', '3417', (39, 43)) ('colorectal cancer', 'Disease', 'MESH:D015179', (20, 37)) ('colorectal cancer', 'Disease', (20, 37)) ('IDH2', 'Gene', (81, 85)) ('cancer', 'Disease', 'MESH:D009369', (31, 37)) ('IDH2', 'Gene', '3418', (81, 85)) ('IDH1/2', 'Gene', '3417;3418', (39, 45)) ('IDH1', 'Gene', (66, 70)) 808 27621679 IDH1 mutations were described in 2008, in exome-sequencing studies of GBM (WHO grade IV astrocytoma). ('mutations', 'Var', (5, 14)) ('astrocytoma', 'Disease', 'MESH:D001254', (88, 99)) ('IDH1', 'Gene', (0, 4)) ('GBM', 'Phenotype', 'HP:0012174', (70, 73)) ('astrocytoma', 'Disease', (88, 99)) ('astrocytoma', 'Phenotype', 'HP:0009592', (88, 99)) ('IDH1', 'Gene', '3417', (0, 4)) 809 27621679 Subsequent studies have shown that IDH1/2 mutations occur in a mutually exclusive manner in ~80% of WHO grade II/III oligodendrogliomas, astrocytomas, and oligoastrocytomas and secondary GBM (ie, GBM that had progressed from lower grade gliomas). ('astrocytomas', 'Disease', (160, 172)) ('gliomas', 'Phenotype', 'HP:0009733', (128, 135)) ('III oligodendrogliomas', 'Disease', (113, 135)) ('glioma', 'Phenotype', 'HP:0009733', (237, 243)) ('III oligodendrogliomas', 'Disease', 'MESH:D009837', (113, 135)) ('GBM', 'Phenotype', 'HP:0012174', (196, 199)) ('secondary GBM', 'Disease', (177, 190)) ('gliomas', 'Phenotype', 'HP:0009733', (237, 244)) ('GBM', 'Phenotype', 'HP:0012174', (187, 190)) ('astrocytomas', 'Disease', 'MESH:D001254', (137, 149)) ('astrocytoma', 'Phenotype', 'HP:0009592', (137, 148)) ('astrocytomas', 'Disease', 'MESH:D001254', (160, 172)) ('astrocytoma', 'Phenotype', 'HP:0009592', (160, 171)) ('gliomas', 'Disease', (128, 135)) ('oligoastrocytomas', 'Disease', (155, 172)) ('IDH1/2', 'Gene', '3417;3418', (35, 41)) ('oligoastrocytomas', 'Disease', 'MESH:D001254', (155, 172)) ('gliomas', 'Disease', (237, 244)) ('IDH1/2', 'Gene', (35, 41)) ('gliomas', 'Disease', 'MESH:D005910', (128, 135)) ('astrocytomas', 'Disease', (137, 149)) ('glioma', 'Phenotype', 'HP:0009733', (128, 134)) ('gliomas', 'Disease', 'MESH:D005910', (237, 244)) ('mutations', 'Var', (42, 51)) 811 27621679 IDH1 R132H mutation represents 80% of all IDH mutations. ('IDH', 'Gene', (42, 45)) ('IDH', 'Gene', (0, 3)) ('R132H mutation', 'Var', (5, 19)) ('IDH', 'Gene', '3417', (42, 45)) ('IDH', 'Gene', '3417', (0, 3)) ('R132H', 'Mutation', 'rs121913500', (5, 10)) ('IDH1', 'Gene', (0, 4)) ('IDH1', 'Gene', '3417', (0, 4)) 812 27621679 Rarely, other mutations are found affecting either IDH1 at Arg132 (including R132S, R132C, R132G, and R132L substitutions) or IDH2 at Arg172 (R172K most frequently; Table 1). ('R132L', 'Mutation', 'rs121913500', (102, 107)) ('IDH1', 'Gene', '3417', (51, 55)) ('R132S', 'Mutation', 'rs121913499', (77, 82)) ('R132G', 'Var', (91, 96)) ('R172K', 'Mutation', 'rs121913503', (142, 147)) ('IDH2', 'Gene', (126, 130)) ('Arg132', 'Chemical', '-', (59, 65)) ('R132C', 'Var', (84, 89)) ('R132L', 'Var', (102, 107)) ('IDH2', 'Gene', '3418', (126, 130)) ('IDH1', 'Gene', (51, 55)) ('Arg172', 'Chemical', '-', (134, 140)) ('R132S', 'Var', (77, 82)) ('R132C', 'Mutation', 'rs121913499', (84, 89)) ('R132G', 'Mutation', 'rs121913499', (91, 96)) 813 27621679 IDH1 R132H mutation can be diagnosed by immunohistochemistry or sequencing, while other mutations can be identified only by sequencing. ('IDH1', 'Gene', (0, 4)) ('R132H', 'Mutation', 'rs121913500', (5, 10)) ('R132H', 'Var', (5, 10)) ('IDH1', 'Gene', '3417', (0, 4)) 817 27621679 Accordingly, the IDH1/2 mutation is often referred to as a "trunk" initiating event in the clonal evolutionary tree of IDH1/2-mutant gliomas. ('IDH1/2', 'Gene', '3417;3418', (119, 125)) ('glioma', 'Phenotype', 'HP:0009733', (133, 139)) ('IDH1/2', 'Gene', '3417;3418', (17, 23)) ('IDH1/2', 'Gene', (119, 125)) ('gliomas', 'Disease', 'MESH:D005910', (133, 140)) ('IDH1/2', 'Gene', (17, 23)) ('gliomas', 'Phenotype', 'HP:0009733', (133, 140)) ('mutation', 'Var', (24, 32)) ('gliomas', 'Disease', (133, 140)) 818 27621679 Secondary genetic alterations occurring during the evolution of IDH1/2-mutant gliomas are often referred as "lineage-defining events", as TP53 and ATRX mutations characterize tumors of astrocytic lineage, whereas hTERT promoter mutation and 1p/19q codeletion are associated with oligodendroglial tumors. ('IDH1/2', 'Gene', '3417;3418', (64, 70)) ('tumors', 'Disease', (175, 181)) ('mutations', 'Var', (152, 161)) ('IDH1/2', 'Gene', (64, 70)) ('tumors', 'Disease', 'MESH:D009369', (296, 302)) ('gliomas', 'Disease', 'MESH:D005910', (78, 85)) ('hTERT promoter', 'Gene', (213, 227)) ('oligodendroglial tumors', 'Disease', (279, 302)) ('glioma', 'Phenotype', 'HP:0009733', (78, 84)) ('tumors', 'Disease', 'MESH:D009369', (175, 181)) ('1p/19q codeletion', 'Var', (241, 258)) ('ATRX', 'Gene', (147, 151)) ('TP53', 'Gene', (138, 142)) ('gliomas', 'Phenotype', 'HP:0009733', (78, 85)) ('ATRX', 'Gene', '546', (147, 151)) ('oligodendroglial tumors', 'Disease', 'MESH:D009369', (279, 302)) ('tumors', 'Phenotype', 'HP:0002664', (296, 302)) ('tumors', 'Phenotype', 'HP:0002664', (175, 181)) ('tumor', 'Phenotype', 'HP:0002664', (296, 301)) ('TP53', 'Gene', '7157', (138, 142)) ('gliomas', 'Disease', (78, 85)) ('tumors', 'Disease', (296, 302)) ('associated', 'Reg', (263, 273)) ('tumor', 'Phenotype', 'HP:0002664', (175, 180)) 820 27621679 Importantly, IDH1/2 mutations have been associated with prognostic and predictive values as biomarkers in gliomas, and assessment of the IDH1/2 status is being implemented in routine clinical practice for patients with primary brain tumors. ('tumor', 'Phenotype', 'HP:0002664', (233, 238)) ('IDH1/2', 'Gene', '3417;3418', (137, 143)) ('primary brain tumors', 'Disease', 'MESH:D001932', (219, 239)) ('primary brain tumors', 'Disease', (219, 239)) ('IDH1/2', 'Gene', '3417;3418', (13, 19)) ('patients', 'Species', '9606', (205, 213)) ('tumors', 'Phenotype', 'HP:0002664', (233, 239)) ('glioma', 'Phenotype', 'HP:0009733', (106, 112)) ('IDH1/2', 'Gene', (137, 143)) ('IDH1/2', 'Gene', (13, 19)) ('brain tumors', 'Phenotype', 'HP:0030692', (227, 239)) ('brain tumor', 'Phenotype', 'HP:0030692', (227, 238)) ('gliomas', 'Disease', 'MESH:D005910', (106, 113)) ('gliomas', 'Phenotype', 'HP:0009733', (106, 113)) ('gliomas', 'Disease', (106, 113)) ('associated', 'Reg', (40, 50)) ('mutations', 'Var', (20, 29)) 822 27621679 Recent studies have demonstrated that IDH1/2 mutations are associated with younger age, better prognosis, and better response to treatment. ('IDH1/2', 'Gene', (38, 44)) ('mutations', 'Var', (45, 54)) ('IDH1/2', 'Gene', '3417;3418', (38, 44)) 824 27621679 Among IDH1/2-mutant gliomas, patients with oligodendroglial tumors harboring 1p19q codeletion have the more favorable prognosis. ('glioma', 'Phenotype', 'HP:0009733', (20, 26)) ('patients', 'Species', '9606', (29, 37)) ('1p19q codeletion', 'Var', (77, 93)) ('IDH1/2', 'Gene', '3417;3418', (6, 12)) ('oligodendroglial tumors', 'Disease', 'MESH:D009369', (43, 66)) ('oligodendroglial tumors', 'Disease', (43, 66)) ('tumor', 'Phenotype', 'HP:0002664', (60, 65)) ('tumors', 'Phenotype', 'HP:0002664', (60, 66)) ('IDH1/2', 'Gene', (6, 12)) ('gliomas', 'Disease', 'MESH:D005910', (20, 27)) ('gliomas', 'Phenotype', 'HP:0009733', (20, 27)) ('gliomas', 'Disease', (20, 27)) 825 27621679 The mechanisms underlying the relative chemo- and radiosensitive phenotypes associated with IDH1/2 mutations are not fully understood. ('mutations', 'Var', (99, 108)) ('IDH1/2', 'Gene', '3417;3418', (92, 98)) ('IDH1/2', 'Gene', (92, 98)) 826 27621679 Epigenetic silencing of the methyl-guanine methyl transferase (MGMT) - which encodes a DNA repair protein that counteracts the cytotoxic effect of alkylating agents - promoter gene by methylation is frequently observed in IDH1/2-mutant tumors and is associated with a partial inability of the tumor to repair the alkylating agent-induced DNA damage. ('IDH1/2', 'Gene', (222, 228)) ('tumor', 'Phenotype', 'HP:0002664', (293, 298)) ('inability of the tumor', 'Disease', 'MESH:D016388', (276, 298)) ('inability of the tumor', 'Disease', (276, 298)) ('tumors', 'Phenotype', 'HP:0002664', (236, 242)) ('tumor', 'Phenotype', 'HP:0002664', (236, 241)) ('IDH1/2', 'Gene', '3417;3418', (222, 228)) ('observed', 'Reg', (210, 218)) ('tumors', 'Disease', (236, 242)) ('Epigenetic', 'MPA', (0, 10)) ('tumors', 'Disease', 'MESH:D009369', (236, 242)) ('methylation', 'Var', (184, 195)) 827 27621679 IDH1 (R132) or IDH2 (R140 and R172) mutations are found in myeloid malignancies, that is, myelodysplastic syndromes (MDS), AML, and myeloproliferative neoplasms, and also in angioimmunoblastic T-cell lymphoma (AITL). ('MDS', 'Disease', (117, 120)) ('IDH2', 'Gene', (15, 19)) ('myeloproliferative neoplasms', 'Disease', (132, 160)) ('angioimmunoblastic T-cell lymphoma', 'Disease', (174, 208)) ('IDH2', 'Gene', '3418', (15, 19)) ('R140 and R172) mutations', 'Var', (21, 45)) ('myeloproliferative neoplasms', 'Disease', 'MESH:D009196', (132, 160)) ('IDH1', 'Gene', (0, 4)) ('cell lymphoma', 'Phenotype', 'HP:0012191', (195, 208)) ('lymphoma', 'Phenotype', 'HP:0002665', (200, 208)) ('neoplasms', 'Phenotype', 'HP:0002664', (151, 160)) ('found', 'Reg', (50, 55)) ('angioimmunoblastic T-cell lymphoma', 'Disease', 'MESH:D016399', (174, 208)) ('myeloid malignancies', 'Disease', 'MESH:D009369', (59, 79)) ('IDH1', 'Gene', '3417', (0, 4)) ('MDS', 'Phenotype', 'HP:0002863', (117, 120)) ('myeloid malignancies', 'Disease', (59, 79)) ('myelodysplastic syndromes', 'Phenotype', 'HP:0002863', (90, 115)) ('myelodysplastic syndromes', 'Disease', (90, 115)) ('MDS', 'Disease', 'MESH:D009190', (117, 120)) ('AML', 'Disease', 'MESH:D015470', (123, 126)) ('myeloproliferative neoplasms', 'Phenotype', 'HP:0005547', (132, 160)) ('mutations', 'Var', (36, 45)) ('myelodysplastic syndromes', 'Disease', 'MESH:D009190', (90, 115)) ('T-cell lymphoma', 'Phenotype', 'HP:0012190', (193, 208)) ('AML', 'Disease', (123, 126)) 828 27621679 In myeloid malignancies, they are considered as an initiating event in 19% of patients with IDH1 mutations and 34% of patients with IDH2 mutations. ('myeloid malignancies', 'Disease', (3, 23)) ('mutations', 'Var', (97, 106)) ('IDH2', 'Gene', '3418', (132, 136)) ('IDH1', 'Gene', (92, 96)) ('patients', 'Species', '9606', (78, 86)) ('IDH1', 'Gene', '3417', (92, 96)) ('myeloid malignancies', 'Disease', 'MESH:D009369', (3, 23)) ('patients', 'Species', '9606', (118, 126)) ('IDH2', 'Gene', (132, 136)) 829 27621679 However, IDH1 mutations are likely to be implicated in early stages of de novo AML as others are, that is, NPM1, DNMT3A, TET2, and ASXL1. ('AML', 'Disease', (79, 82)) ('ASXL1', 'Gene', (131, 136)) ('NPM1', 'Gene', '4869', (107, 111)) ('IDH1', 'Gene', (9, 13)) ('TET2', 'Gene', (121, 125)) ('DNMT3A', 'Gene', (113, 119)) ('DNMT3A', 'Gene', '1788', (113, 119)) ('IDH1', 'Gene', '3417', (9, 13)) ('AML', 'Disease', 'MESH:D015470', (79, 82)) ('implicated', 'Reg', (41, 51)) ('TET2', 'Gene', '54790', (121, 125)) ('mutations', 'Var', (14, 23)) ('NPM1', 'Gene', (107, 111)) ('ASXL1', 'Gene', '171023', (131, 136)) 830 27621679 In de novo AML, IDH1/2 mutations are associated with older age, normal karyotype, and NPM1 mutations. ('IDH1/2', 'Gene', (16, 22)) ('NPM1', 'Gene', '4869', (86, 90)) ('mutations', 'Var', (23, 32)) ('mutations', 'Var', (91, 100)) ('AML', 'Disease', 'MESH:D015470', (11, 14)) ('associated', 'Reg', (37, 47)) ('IDH1/2', 'Gene', '3417;3418', (16, 22)) ('NPM1', 'Gene', (86, 90)) ('AML', 'Disease', (11, 14)) 832 27621679 IDH1 mutations are found in 6%-16% of de novo AML and are associated with a poorer outcome in patients treated with intensive chemotherapy, even in patients with favorable prognosis as per European LeukemiaNet (ELN) classification. ('patients', 'Species', '9606', (148, 156)) ('IDH1', 'Gene', '3417', (0, 4)) ('associated', 'Reg', (58, 68)) ('patients', 'Species', '9606', (94, 102)) ('AML', 'Disease', (46, 49)) ('mutations', 'Var', (5, 14)) ('LeukemiaNet', 'Disease', (198, 209)) ('IDH1', 'Gene', (0, 4)) ('AML', 'Disease', 'MESH:D015470', (46, 49)) ('LeukemiaNet', 'Disease', 'None', (198, 209)) 833 27621679 IDH2 mutations are found in 8%-19% of de novo AML. ('mutations', 'Var', (5, 14)) ('AML', 'Disease', (46, 49)) ('IDH2', 'Gene', (0, 4)) ('AML', 'Disease', 'MESH:D015470', (46, 49)) ('IDH2', 'Gene', '3418', (0, 4)) 835 27621679 Depending on the mutational spectrum, IDH2 R140Q mutations confer favorable or no impact on overall survival. ('overall', 'MPA', (92, 99)) ('R140Q', 'Mutation', 'rs121913502', (43, 48)) ('IDH2', 'Gene', (38, 42)) ('IDH2', 'Gene', '3418', (38, 42)) ('R140Q mutations', 'Var', (43, 58)) 836 27621679 Conversely, patients with IDH2 R172K mutation have a worse prognosis, with lower complete remission rate, higher relapse rate, and lower overall survival. ('lower', 'NegReg', (75, 80)) ('patients', 'Species', '9606', (12, 20)) ('complete remission rate', 'CPA', (81, 104)) ('IDH2', 'Gene', (26, 30)) ('relapse', 'CPA', (113, 120)) ('higher', 'PosReg', (106, 112)) ('R172K', 'Mutation', 'rs121913503', (31, 36)) ('IDH2', 'Gene', '3418', (26, 30)) ('lower', 'NegReg', (131, 136)) ('overall', 'MPA', (137, 144)) ('R172K', 'Var', (31, 36)) 837 27621679 IDH1/2 mutations are found in 4% to 12% of MDS cases, with a higher incidence (up to 23%) in high-risk MDS. ('MDS', 'Phenotype', 'HP:0002863', (103, 106)) ('MDS', 'Disease', 'MESH:D009190', (103, 106)) ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('MDS', 'Disease', (103, 106)) ('MDS', 'Disease', (43, 46)) ('MDS', 'Disease', 'MESH:D009190', (43, 46)) ('MDS', 'Phenotype', 'HP:0002863', (43, 46)) ('IDH1/2', 'Gene', (0, 6)) ('mutations', 'Var', (7, 16)) 838 27621679 IDH1/2 mutations are associated with an older age, DNMT3A, ASXL1, SRSF2 mutations, and higher rate of transformation to AML. ('SRSF2', 'Gene', '6427', (66, 71)) ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('transformation', 'MPA', (102, 116)) ('associated', 'Reg', (21, 31)) ('AML', 'Disease', 'MESH:D015470', (120, 123)) ('ASXL1', 'Gene', '171023', (59, 64)) ('SRSF2', 'Gene', (66, 71)) ('DNMT3A', 'Gene', '1788', (51, 57)) ('DNMT3A', 'Gene', (51, 57)) ('AML', 'Disease', (120, 123)) ('IDH1/2', 'Gene', (0, 6)) ('ASXL1', 'Gene', (59, 64)) ('mutations', 'Var', (7, 16)) 839 27621679 In myeloproliferative neoplasms, the incidence of IDH1/2 mutations range between 2% and 4%, rising up to 31% after transformation to AML. ('IDH1/2', 'Gene', (50, 56)) ('myeloproliferative neoplasms', 'Disease', 'MESH:D009196', (3, 31)) ('neoplasms', 'Phenotype', 'HP:0002664', (22, 31)) ('AML', 'Disease', 'MESH:D015470', (133, 136)) ('rising', 'PosReg', (92, 98)) ('mutations', 'Var', (57, 66)) ('myeloproliferative neoplasms', 'Disease', (3, 31)) ('AML', 'Disease', (133, 136)) ('IDH1/2', 'Gene', '3417;3418', (50, 56)) ('myeloproliferative neoplasms', 'Phenotype', 'HP:0005547', (3, 31)) 840 27621679 They are associated with older age and SRSF2 mutations. ('SRSF2', 'Gene', '6427', (39, 44)) ('SRSF2', 'Gene', (39, 44)) ('mutations', 'Var', (45, 54)) ('associated', 'Reg', (9, 19)) 841 27621679 In patients with myelofibrosis, IDH1/2 mutations confer worse prognosis. ('myelofibrosis', 'Phenotype', 'HP:0011974', (17, 30)) ('IDH1/2', 'Gene', (32, 38)) ('mutations', 'Var', (39, 48)) ('patients', 'Species', '9606', (3, 11)) ('myelofibrosis', 'Disease', (17, 30)) ('IDH1/2', 'Gene', '3417;3418', (32, 38)) ('myelofibrosis', 'Disease', 'MESH:D055728', (17, 30)) 842 27621679 Besides myeloid neoplasms, IDH2 mutations (mostly R172K) are found in 20%-45% of patients with AITL although its prognostic value is not yet known. ('myeloid neoplasms', 'Disease', 'MESH:D007951', (8, 25)) ('R172K', 'Mutation', 'rs121913503', (50, 55)) ('neoplasms', 'Phenotype', 'HP:0002664', (16, 25)) ('patients', 'Species', '9606', (81, 89)) ('found', 'Reg', (61, 66)) ('R172K', 'Var', (50, 55)) ('IDH2', 'Gene', (27, 31)) ('myeloid neoplasms', 'Disease', (8, 25)) ('myeloid neoplasms', 'Phenotype', 'HP:0012324', (8, 25)) ('IDH2', 'Gene', '3418', (27, 31)) 843 27621679 IDH1/2 mutations have been detected in enchondromas and chondrosarcomas but rarely found in other mesenchymal tumors such as osteosarcomas. ('mesenchymal tumors', 'Disease', (98, 116)) ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('chondrosarcomas', 'Phenotype', 'HP:0006765', (56, 71)) ('enchondromas and chondrosarcomas', 'Disease', 'MESH:D002812', (39, 71)) ('tumor', 'Phenotype', 'HP:0002664', (110, 115)) ('osteosarcomas', 'Disease', (125, 138)) ('tumors', 'Phenotype', 'HP:0002664', (110, 116)) ('chondrosarcoma', 'Phenotype', 'HP:0006765', (56, 70)) ('osteosarcomas', 'Phenotype', 'HP:0002669', (125, 138)) ('enchondromas', 'Phenotype', 'HP:0030038', (39, 51)) ('IDH1/2', 'Gene', (0, 6)) ('osteosarcomas', 'Disease', 'MESH:D012516', (125, 138)) ('detected', 'Reg', (27, 35)) ('mesenchymal tumors', 'Disease', 'MESH:C535700', (98, 116)) ('mutations', 'Var', (7, 16)) 844 27621679 IDH1 mutations are the most frequent (40%-52%), and IDH2 mutations are present in 6%-11% of the cases. ('IDH2', 'Gene', '3418', (52, 56)) ('IDH1', 'Gene', (0, 4)) ('mutations', 'Var', (5, 14)) ('IDH2', 'Gene', (52, 56)) ('IDH1', 'Gene', '3417', (0, 4)) 845 27621679 IDH1/2 mutations occur in up to 25% of intrahepatic cholangiocarcinomas. ('IDH1/2', 'Gene', '3417;3418', (0, 6)) ('occur', 'Reg', (17, 22)) ('intrahepatic cholangiocarcinomas', 'Disease', (39, 71)) ('carcinoma', 'Phenotype', 'HP:0030731', (61, 70)) ('intrahepatic cholangiocarcinomas', 'Disease', 'MESH:D018281', (39, 71)) ('cholangiocarcinoma', 'Phenotype', 'HP:0030153', (52, 70)) ('IDH1/2', 'Gene', (0, 6)) ('mutations', 'Var', (7, 16)) 846 27621679 Again, IDH1 mutations are the most frequent (11%-24%) and IDH2 mutations are seen in 2%-6% of the cases. ('IDH1', 'Gene', '3417', (7, 11)) ('IDH2', 'Gene', '3418', (58, 62)) ('mutations', 'Var', (12, 21)) ('IDH2', 'Gene', (58, 62)) ('IDH1', 'Gene', (7, 11)) 847 27621679 Finally, sporadic cases of IDH1/2 mutations have been reported in other types of solid cancers: thyroid cancer, melanoma, prostate carcinoma, lung cancer, breast adenocarcinoma, colorectal cancer, esophageal cancer, and bladder cancer. ('esophageal cancer', 'Disease', 'MESH:D004938', (197, 214)) ('cancer', 'Phenotype', 'HP:0002664', (147, 153)) ('solid cancers', 'Disease', (81, 94)) ('colorectal cancer', 'Disease', (178, 195)) ('breast adenocarcinoma', 'Disease', 'MESH:D000230', (155, 176)) ('cancer', 'Phenotype', 'HP:0002664', (208, 214)) ('carcinoma', 'Phenotype', 'HP:0030731', (167, 176)) ('melanoma', 'Phenotype', 'HP:0002861', (112, 120)) ('lung cancer', 'Disease', (142, 153)) ('melanoma', 'Disease', (112, 120)) ('esophageal cancer', 'Disease', (197, 214)) ('prostate carcinoma', 'Disease', 'MESH:D011472', (122, 140)) ('breast adenocarcinoma', 'Disease', (155, 176)) ('cancer', 'Phenotype', 'HP:0002664', (104, 110)) ('prostate carcinoma', 'Phenotype', 'HP:0012125', (122, 140)) ('breast adenocarcinoma', 'Phenotype', 'HP:0003002', (155, 176)) ('cancers', 'Phenotype', 'HP:0002664', (87, 94)) ('thyroid cancer', 'Disease', (96, 110)) ('colorectal cancer', 'Phenotype', 'HP:0003003', (178, 195)) ('lung cancer', 'Disease', 'MESH:D008175', (142, 153)) ('IDH1/2', 'Gene', '3417;3418', (27, 33)) ('cancer', 'Phenotype', 'HP:0002664', (87, 93)) ('melanoma', 'Disease', 'MESH:D008545', (112, 120)) ('lung cancer', 'Phenotype', 'HP:0100526', (142, 153)) ('IDH1/2', 'Gene', (27, 33)) ('prostate carcinoma', 'Disease', (122, 140)) ('reported', 'Reg', (54, 62)) ('bladder cancer', 'Disease', 'MESH:D001749', (220, 234)) ('bladder cancer', 'Disease', (220, 234)) ('carcinoma', 'Phenotype', 'HP:0030731', (131, 140)) ('thyroid cancer', 'Disease', 'MESH:D013964', (96, 110)) ('cancer', 'Phenotype', 'HP:0002664', (189, 195)) ('bladder cancer', 'Phenotype', 'HP:0009725', (220, 234)) ('solid cancers', 'Disease', 'MESH:D009369', (81, 94)) ('colorectal cancer', 'Disease', 'MESH:D015179', (178, 195)) ('mutations', 'Var', (34, 43)) ('thyroid cancer', 'Phenotype', 'HP:0002890', (96, 110)) 848 27621679 D-2HG released in the serum and/or urine by cancer cells harboring IDH1/2 mutations is a biomarker for IDH1/2 mutations, presumably reflecting the neomorphic enzymatic activity of the mutant enzymes. ('mutations', 'Var', (110, 119)) ('IDH1/2', 'Gene', (67, 73)) ('cancer', 'Disease', (44, 50)) ('mutations', 'Var', (74, 83)) ('cancer', 'Disease', 'MESH:D009369', (44, 50)) ('IDH1/2', 'Gene', '3417;3418', (103, 109)) ('D-2HG', 'MPA', (0, 5)) ('IDH1/2', 'Gene', '3417;3418', (67, 73)) ('IDH1/2', 'Gene', (103, 109)) ('cancer', 'Phenotype', 'HP:0002664', (44, 50)) 850 27621679 At diagnosis, D-2HG is a strong predictive biomarker for the presence of IDH1/2 mutations in AML. ('IDH1/2', 'Gene', (73, 79)) ('mutations', 'Var', (80, 89)) ('AML', 'Disease', (93, 96)) ('IDH1/2', 'Gene', '3417;3418', (73, 79)) ('AML', 'Disease', 'MESH:D015470', (93, 96)) 851 27621679 High total 2HG concentration was highly predictive of the presence of an IDH1/2 mutation, although separation of the d- and l-enantiomers distinguished IDH1/2-mutant and -wild-type AML with greater specificity. ('IDH1/2', 'Gene', (73, 79)) ('IDH1/2', 'Gene', (152, 158)) ('predictive', 'Reg', (40, 50)) ('2HG concentration', 'MPA', (11, 28)) ('AML', 'Disease', 'MESH:D015470', (181, 184)) ('IDH1/2', 'Gene', '3417;3418', (73, 79)) ('IDH1/2', 'Gene', '3417;3418', (152, 158)) ('mutation', 'Var', (80, 88)) ('AML', 'Disease', (181, 184)) 852 27621679 Increased serum and/or urine D-2HG levels predict IDH1/2 mutation in AML and intrahepatic cholangiocarcinoma and may be used as predictive biomarker for tumor response/recurrence. ('intrahepatic cholangiocarcinoma', 'Disease', (77, 108)) ('mutation', 'Var', (57, 65)) ('tumor', 'Disease', 'MESH:D009369', (153, 158)) ('AML', 'Disease', (69, 72)) ('AML', 'Disease', 'MESH:D015470', (69, 72)) ('IDH1/2', 'Gene', (50, 56)) ('tumor', 'Phenotype', 'HP:0002664', (153, 158)) ('tumor', 'Disease', (153, 158)) ('intrahepatic cholangiocarcinoma', 'Disease', 'MESH:D018281', (77, 108)) ('carcinoma', 'Phenotype', 'HP:0030731', (99, 108)) ('IDH1/2', 'Gene', '3417;3418', (50, 56)) ('cholangiocarcinoma', 'Phenotype', 'HP:0030153', (90, 108)) 854 27621679 Failure to normalize D-2HG levels is associated with treatment failure, whereas elevated D-2HG levels at complete remission are associated with poorer outcome, suggesting that D-2HG is a biomarker predictive of clinical response to intensive chemotherapy in AML patients with IDH1/2 mutations. ('AML', 'Disease', (258, 261)) ('IDH1/2', 'Gene', (276, 282)) ('mutations', 'Var', (283, 292)) ('D-2HG', 'MPA', (21, 26)) ('IDH1/2', 'Gene', '3417;3418', (276, 282)) ('AML', 'Disease', 'MESH:D015470', (258, 261)) ('patients', 'Species', '9606', (262, 270)) 856 27621679 Nevertheless, recent studies suggested that the urinary levels of D-2HG may increase the sensitivity and specificity for IDH1/2 mutation detection in glioma patients. ('increase', 'PosReg', (76, 84)) ('IDH1/2', 'Gene', '3417;3418', (121, 127)) ('urinary', 'MPA', (48, 55)) ('patients', 'Species', '9606', (157, 165)) ('D-2HG', 'Var', (66, 71)) ('glioma', 'Disease', (150, 156)) ('IDH1/2', 'Gene', (121, 127)) ('mutation', 'Var', (128, 136)) ('glioma', 'Disease', 'MESH:D005910', (150, 156)) ('glioma', 'Phenotype', 'HP:0009733', (150, 156)) 860 27621679 The discovery of IDH1/2 mutations has resulted in a number of novel therapeutic approaches (Table 2), which either restore normal IDH1/2 function or block production or downstream effects of D-2HG. ('IDH1/2', 'Gene', '3417;3418', (17, 23)) ('resulted in', 'Reg', (38, 49)) ('IDH1/2', 'Gene', (130, 136)) ('block', 'NegReg', (149, 154)) ('IDH1/2', 'Gene', (17, 23)) ('restore', 'PosReg', (115, 122)) ('production', 'MPA', (155, 165)) ('mutations', 'Var', (24, 33)) ('IDH1/2', 'Gene', '3417;3418', (130, 136)) ('function', 'MPA', (137, 145)) 861 27621679 Hypomethylating agents (HMAs) may be of interest in the context of CpG island methylator phenotype induced by IDH1/2 mutations. ('IDH1/2', 'Gene', '3417;3418', (110, 116)) ('mutations', 'Var', (117, 126)) ('IDH1/2', 'Gene', (110, 116)) 864 27621679 These series suggested the lack of association between IDH1/2 mutations and efficacy of HMAs or showed a better response to DNMT inhibitors among patients with IDH1/2-mutant AML. ('IDH1/2', 'Gene', (160, 166)) ('DNMT', 'Gene', (124, 128)) ('HMAs', 'CPA', (88, 92)) ('AML', 'Disease', 'MESH:D015470', (174, 177)) ('DNMT', 'Gene', '1786', (124, 128)) ('IDH1/2', 'Gene', '3417;3418', (55, 61)) ('IDH1/2', 'Gene', '3417;3418', (160, 166)) ('AML', 'Disease', (174, 177)) ('patients', 'Species', '9606', (146, 154)) ('mutations', 'Var', (62, 71)) ('IDH1/2', 'Gene', (55, 61)) 867 27621679 Preclinical in vitro and in vivo studies have validated the proof of concept that targeted inhibition of IDH1/2 mutants resulted in normalization in a dose-dependent manner of 2-HG, reversal of histone and DNA hypermethylation, and release of cellular differentiation block. ('reversal', 'Reg', (182, 190)) ('IDH1/2', 'Gene', (105, 111)) ('2-HG', 'MPA', (176, 180)) ('cellular differentiation block', 'CPA', (243, 273)) ('inhibition', 'NegReg', (91, 101)) ('mutants', 'Var', (112, 119)) ('normalization', 'MPA', (132, 145)) ('IDH1/2', 'Gene', '3417;3418', (105, 111)) 868 27621679 AGI-5198 and AGI-6780 are selective inhibitors of mutant IDH1 and IDH2 enzymes, respectively. ('IDH2', 'Gene', '3418', (66, 70)) ('IDH1', 'Gene', (57, 61)) ('IDH1', 'Gene', '3417', (57, 61)) ('mutant', 'Var', (50, 56)) ('IDH2', 'Gene', (66, 70)) 869 27621679 They normalized 2HG, reversed histone and DNA hypermethylation, and induced differentiation of not only TF-1 erythroleukemia cells but also primary human AML cells harboring IDH1/2 mutations. ('IDH1/2', 'Gene', '3417;3418', (174, 180)) ('AML', 'Disease', 'MESH:D015470', (154, 157)) ('differentiation', 'CPA', (76, 91)) ('reversed', 'NegReg', (21, 29)) ('induced', 'Reg', (68, 75)) ('erythroleukemia', 'Disease', (109, 124)) ('IDH1/2', 'Gene', (174, 180)) ('human', 'Species', '9606', (148, 153)) ('AML', 'Disease', (154, 157)) ('mutations', 'Var', (181, 190)) ('histone', 'MPA', (30, 37)) ('DNA hypermethylation', 'MPA', (42, 62)) ('erythroleukemia', 'Disease', 'MESH:D004915', (109, 124)) ('normalized 2HG', 'MPA', (5, 19)) 872 27621679 Together, these studies indicate that differentiation therapy may be achievable in cancers with IDH1/2 mutations, thereby supporting the initiation of clinical trials (Table 2). ('cancers', 'Disease', 'MESH:D009369', (83, 90)) ('mutations', 'Var', (103, 112)) ('IDH1/2', 'Gene', '3417;3418', (96, 102)) ('cancer', 'Phenotype', 'HP:0002664', (83, 89)) ('IDH1/2', 'Gene', (96, 102)) ('cancers', 'Phenotype', 'HP:0002664', (83, 90)) ('cancers', 'Disease', (83, 90)) 874 27621679 Separate first-in-human, Phase I, dose-escalation studies of AG-120 and AG-221 are underway in patients with IDH1/2-mutated hematologic malignancies (NCT02074839 and NCT01915498). ('NCT02074839', 'Var', (150, 161)) ('hematologic malignancies', 'Disease', (124, 148)) ('IDH1/2', 'Gene', (109, 115)) ('patients', 'Species', '9606', (95, 103)) ('human', 'Species', '9606', (18, 23)) ('AG-221', 'Gene', (72, 78)) ('NCT01915498', 'Var', (166, 177)) ('AG-120', 'Gene', (61, 67)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (124, 148)) ('IDH1/2', 'Gene', '3417;3418', (109, 115)) 877 27621679 Recent studies have investigated vaccination-based immunotherapy to target IDH1 mutations. ('IDH1', 'Gene', '3417', (75, 79)) ('IDH1', 'Gene', (75, 79)) ('mutations', 'Var', (80, 89)) 878 27621679 In principle, IDH1/2 mutants are ideal tumor-specific neoantigens due to their uniform occurrence at specific codons and ubiquitous expression throughout all tumor cells. ('mutants', 'Var', (21, 28)) ('tumor', 'Disease', (158, 163)) ('tumor', 'Phenotype', 'HP:0002664', (158, 163)) ('tumor', 'Disease', 'MESH:D009369', (39, 44)) ('IDH1/2', 'Gene', '3417;3418', (14, 20)) ('tumor', 'Phenotype', 'HP:0002664', (39, 44)) ('tumor', 'Disease', 'MESH:D009369', (158, 163)) ('tumor', 'Disease', (39, 44)) ('IDH1/2', 'Gene', (14, 20)) 880 27621679 These preliminary results suggest that mutant IDH1-targeted immunotherapies can elicit potent antitumor immune responses. ('IDH1', 'Gene', (46, 50)) ('tumor', 'Disease', 'MESH:D009369', (98, 103)) ('IDH1', 'Gene', '3417', (46, 50)) ('tumor', 'Phenotype', 'HP:0002664', (98, 103)) ('elicit', 'Reg', (80, 86)) ('mutant', 'Var', (39, 45)) ('tumor', 'Disease', (98, 103)) 883 27621679 Interestingly, of the five patients who achieved complete remission with or without incomplete marrow recovery, three had IDH1/2 mutations. ('patients', 'Species', '9606', (27, 35)) ('mutations', 'Var', (129, 138)) ('IDH1/2', 'Gene', (122, 128)) ('IDH1/2', 'Gene', '3417;3418', (122, 128)) 885 27621679 The discovery of IDH1/2 mutations highlights the unique role of the "oncometabolite" D-2HG in oncogenesis. ('mutations', 'Var', (24, 33)) ('IDH1/2', 'Gene', (17, 23)) ('IDH1/2', 'Gene', '3417;3418', (17, 23)) 886 27621679 Relevant preclinical models and results of early Phase I trials in adults with hematologic malignancies demonstrate that targeting IDH1/2 mutant is a valid strategy. ('IDH1/2', 'Gene', (131, 137)) ('hematologic malignancies', 'Disease', (79, 103)) ('mutant', 'Var', (138, 144)) ('IDH1/2', 'Gene', '3417;3418', (131, 137)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (79, 103)) 887 26369982 Comprehensive quantitative proteomic profiling of the pharmacodynamic changes induced by MLN4924 in acute myeloid leukemia cells establishes rationale for its combination with azacitidine NEDDylation controls the ubiquitination and proteasomal degradation of proteins that are critical for cell survival, oncogenic transformation, and therapeutic sensitivity including p27, CDT1, IkappaBalpha, NRF-2, cyclin E, c-Myc, p53, and hypoxia-inducible factor-1alpha. ('NRF-2', 'Gene', (394, 399)) ('hypoxia-inducible factor-1alpha', 'Gene', '3091', (427, 458)) ('MLN4924', 'Var', (89, 96)) ('IkappaBalpha', 'Gene', (380, 392)) ('cyclin', 'MPA', (401, 407)) ('IkappaBalpha', 'Gene', '4792', (380, 392)) ('p53', 'Gene', '7157', (418, 421)) ('acute myeloid leukemia', 'Disease', (100, 122)) ('c-Myc', 'Gene', (411, 416)) ('ubiquitination', 'MPA', (213, 227)) ('leukemia', 'Phenotype', 'HP:0001909', (114, 122)) ('CDT1', 'Gene', '81620', (374, 378)) ('c-Myc', 'Gene', '4609', (411, 416)) ('NRF-2', 'Gene', '4780', (394, 399)) ('hypoxia-inducible factor-1alpha', 'Gene', (427, 458)) ('p53', 'Gene', (418, 421)) ('p27', 'Gene', '3429', (369, 372)) ('p27', 'Gene', (369, 372)) ('CDT1', 'Gene', (374, 378)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (100, 122)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (106, 122)) ('azacitidine', 'Chemical', 'MESH:D001374', (176, 187)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (100, 122)) ('proteasomal degradation', 'MPA', (232, 255)) 888 26369982 Aberrant NEDDylation has been implicated in malignant pathogenesis and drug resistance, thus providing strong rationale to develop strategies to disrupt this specific mechanism of protein turnover for cancer therapy. ('cancer', 'Disease', 'MESH:D009369', (201, 207)) ('NEDDylation', 'MPA', (9, 20)) ('Aberrant', 'Var', (0, 8)) ('cancer', 'Disease', (201, 207)) ('implicated', 'Reg', (30, 40)) ('cancer', 'Phenotype', 'HP:0002664', (201, 207)) ('drug resistance', 'Phenotype', 'HP:0020174', (71, 86)) 890 26369982 Despite its robust effects in preclinical models of acute myeloid leukemia (AML) and preliminary efficacy in patients with relapsed/refractory AML and high-risk myelodysplastic syndromes (MDS), the specific pharmacodynamic (PD) effects that mediate the anti-leukemic activity of MLN4924 have not been completely defined and no predictive biomarkers of clinical sensitivity to MLN4924 have been validated. ('AML', 'Disease', (76, 79)) ('patients', 'Species', '9606', (109, 117)) ('AML', 'Phenotype', 'HP:0004808', (76, 79)) ('PD', 'Disease', 'MESH:D010300', (224, 226)) ('MLN4924', 'Var', (279, 286)) ('AML', 'Disease', 'MESH:D015470', (143, 146)) ('acute myeloid leukemia', 'Disease', (52, 74)) ('AML', 'Disease', (143, 146)) ('MDS', 'Phenotype', 'HP:0002863', (188, 191)) ('AML', 'Phenotype', 'HP:0004808', (143, 146)) ('MLN4924', 'Chemical', 'MESH:C539933', (376, 383)) ('leukemic', 'Disease', 'MESH:D007938', (258, 266)) ('myelodysplastic syndromes', 'Phenotype', 'HP:0002863', (161, 186)) ('myelodysplastic syndromes', 'Disease', (161, 186)) ('leukemic', 'Disease', (258, 266)) ('MLN4924', 'Chemical', 'MESH:C539933', (279, 286)) ('MDS', 'Disease', 'MESH:D009190', (188, 191)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (52, 74)) ('myelodysplastic syndromes', 'Disease', 'MESH:D009190', (161, 186)) ('leukemia', 'Phenotype', 'HP:0001909', (66, 74)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (58, 74)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (52, 74)) ('MDS', 'Disease', (188, 191)) ('AML', 'Disease', 'MESH:D015470', (76, 79)) 893 26369982 We conducted comprehensive protein profiling of MV4-11 FLT3 ITD+ cells to determine the global impact of inhibiting NEDDylation with MLN4924 on the AML proteome. ('inhibiting', 'NegReg', (105, 115)) ('FLT3', 'Gene', (55, 59)) ('AML', 'Phenotype', 'HP:0004808', (148, 151)) ('NEDDylation', 'MPA', (116, 127)) ('AML', 'Disease', (148, 151)) ('MLN4924', 'Chemical', 'MESH:C539933', (133, 140)) ('MLN4924', 'Var', (133, 140)) ('AML', 'Disease', 'MESH:D015470', (148, 151)) ('FLT3', 'Gene', '2322', (55, 59)) 894 26369982 Cells were treated with MLN4924 (1 muM) for 24 hours and samples were processed for proteome quantification as previously described (Supplementary methods). ('MLN4924', 'Chemical', 'MESH:C539933', (24, 31)) ('muM', 'Gene', '56925', (35, 38)) ('MLN4924', 'Var', (24, 31)) ('muM', 'Gene', (35, 38)) 895 26369982 3,812 unique proteins were detected in vehicle and MLN4924-treated MV4-11 cells (Figure 1A). ('MLN4924', 'Chemical', 'MESH:C539933', (51, 58)) ('MLN4924-treated', 'Var', (51, 66)) ('proteins', 'Protein', (13, 21)) 896 26369982 Using a 2-fold PD change cut-off and a ratio of 1 unique peptide, 47 proteins were significantly upregulated following MLN4924 treatment (P< 0.05, Table 1). ('proteins', 'Protein', (69, 77)) ('PD', 'Disease', 'MESH:D010300', (15, 17)) ('MLN4924', 'Chemical', 'MESH:C539933', (119, 126)) ('MLN4924', 'Var', (119, 126)) ('upregulated', 'PosReg', (97, 108)) 897 26369982 The effects of MLN4924 treatment on the levels of selected proteins [NEDDylated cullins, free NEDD8, CDT1, CDKN1B (p27), WEE1, HMOX1, NQO1, RRM2, BRD2, and TXNRD1] were confirmed by immunoblotting (Figure 1B). ('CDT1', 'Gene', '81620', (101, 105)) ('CDKN1B', 'Gene', (107, 113)) ('CDT1', 'Gene', (101, 105)) ('MLN4924', 'Chemical', 'MESH:C539933', (15, 22)) ('TXNRD1', 'Gene', '7296', (156, 162)) ('NQO1', 'Gene', (134, 138)) ('NEDD8', 'Gene', '4738', (94, 99)) ('NEDD8', 'Gene', (94, 99)) ('HMOX1', 'Gene', '3162', (127, 132)) ('CDKN1B', 'Gene', '1027', (107, 113)) ('WEE1', 'Gene', (121, 125)) ('BRD2', 'Gene', '6046', (146, 150)) ('TXNRD1', 'Gene', (156, 162)) ('p27', 'Gene', '3429', (115, 118)) ('MLN4924', 'Var', (15, 22)) ('p27', 'Gene', (115, 118)) ('BRD2', 'Gene', (146, 150)) ('HMOX1', 'Gene', (127, 132)) ('NQO1', 'Gene', '1728', (134, 138)) ('WEE1', 'Gene', '7465', (121, 125)) 898 26369982 As expected, MLN4924 triggered increased levels of many established cullin-dependent substrates. ('MLN4924', 'Chemical', 'MESH:C539933', (13, 20)) ('MLN4924', 'Var', (13, 20)) ('increased', 'PosReg', (31, 40)) 900 26369982 Other notable factors whose levels were modulated by MLN4924 treatment included the cell cycle progression 1 CCPG1, the electron transport chain component NQO1, regulators of cellular redox status (GSR, GLUL, GCLM, TXNRD1, and HMOX1), the DNA helicase DNA2, the DNA replication factor ESCO2, the regulator of p53 stability KIAA0101 and heat shock proteins (HSBP1, DNAJB4). ('MLN4924', 'Chemical', 'MESH:C539933', (53, 60)) ('GCLM', 'Gene', (209, 213)) ('KIAA0101', 'Gene', (323, 331)) ('HMOX1', 'Gene', (227, 232)) ('DNAJB4', 'Gene', '11080', (364, 370)) ('TXNRD1', 'Gene', (215, 221)) ('shock', 'Phenotype', 'HP:0031273', (341, 346)) ('p53', 'Gene', '7157', (309, 312)) ('NQO1', 'Gene', '1728', (155, 159)) ('DNA helicase', 'Protein', (239, 251)) ('ESCO2', 'Gene', '157570', (285, 290)) ('heat shock proteins', 'Disease', (336, 355)) ('KIAA0101', 'Gene', '9768', (323, 331)) ('CCPG1', 'Gene', (109, 114)) ('NQO1', 'Gene', (155, 159)) ('GCLM', 'Gene', '2730', (209, 213)) ('p53', 'Gene', (309, 312)) ('DNAJB4', 'Gene', (364, 370)) ('GLUL', 'Gene', '2752', (203, 207)) ('GLUL', 'Gene', (203, 207)) ('heat shock proteins', 'Disease', 'MESH:D012769', (336, 355)) ('DNA2', 'Gene', (252, 256)) ('MLN4924', 'Var', (53, 60)) ('TXNRD1', 'Gene', '7296', (215, 221)) ('HMOX1', 'Gene', '3162', (227, 232)) ('HSBP1', 'Gene', (357, 362)) ('ESCO2', 'Gene', (285, 290)) ('CCPG1', 'Gene', '9236', (109, 114)) ('HSBP1', 'Gene', '3281', (357, 362)) ('DNA2', 'Gene', '1763', (252, 256)) 901 26369982 Several of these proteins such as CDT1 have been implicated as mediators of DNA re-replication and DNA damage-mediated apoptosis that are stimulated by MLN4924 treatment and responsible, at least in part, for the anti-neoplastic effects of this agent. ('MLN4924', 'Chemical', 'MESH:C539933', (152, 159)) ('CDT1', 'Gene', (34, 38)) ('MLN4924', 'Var', (152, 159)) ('CDT1', 'Gene', '81620', (34, 38)) ('stimulated', 'PosReg', (138, 148)) 902 26369982 Reactome network analysis of the 47 top proteins whose levels were increased 2 or more fold by MLN4924 treatment demonstrated that the affected proteins primarily clustered in the cell cycle, mitosis, and stress response pathways (Table S1). ('levels', 'MPA', (55, 61)) ('stress response', 'CPA', (205, 220)) ('MLN4924', 'Chemical', 'MESH:C539933', (95, 102)) ('cell cycle', 'CPA', (180, 190)) ('mitosis', 'Disease', (192, 199)) ('clustered', 'Reg', (163, 172)) ('increased', 'PosReg', (67, 76)) ('MLN4924', 'Var', (95, 102)) ('mitosis', 'Disease', 'None', (192, 199)) ('proteins', 'Protein', (144, 152)) 903 26369982 We also merged our proteomic profiling of MV4-11 AML cells with a comprehensive SILAC analysis performed in A375 melanoma cells similarly treated with MLN4924. ('AML', 'Disease', (49, 52)) ('MLN4924', 'Var', (151, 158)) ('A375', 'CellLine', 'CVCL:0132', (108, 112)) ('melanoma', 'Phenotype', 'HP:0002861', (113, 121)) ('melanoma', 'Disease', (113, 121)) ('melanoma', 'Disease', 'MESH:D008545', (113, 121)) ('AML', 'Disease', 'MESH:D015470', (49, 52)) ('MLN4924', 'Chemical', 'MESH:C539933', (151, 158)) ('AML', 'Phenotype', 'HP:0004808', (49, 52)) 904 26369982 We determined that 36% (17/47) of the proteins that were pharmacodynamically increased by MLN4924 were identical between the two analyses. ('MLN4924', 'Chemical', 'MESH:C539933', (90, 97)) ('MLN4924', 'Var', (90, 97)) ('increased', 'PosReg', (77, 86)) 906 26369982 It is possible that the spectrum of affected targets in individual tumors may be important in determining sensitivity to MLN4924 as patients with MDS/AML have responded better to this agent than those with other malignancies based on initial phase I studies. ('MLN4924', 'Chemical', 'MESH:C539933', (121, 128)) ('malignancies', 'Disease', 'MESH:D009369', (212, 224)) ('MDS', 'Phenotype', 'HP:0002863', (146, 149)) ('MDS', 'Disease', 'MESH:D009190', (146, 149)) ('AML', 'Disease', 'MESH:D015470', (150, 153)) ('tumors', 'Disease', 'MESH:D009369', (67, 73)) ('MLN4924', 'Var', (121, 128)) ('responded', 'MPA', (159, 168)) ('MDS', 'Disease', (146, 149)) ('malignancies', 'Disease', (212, 224)) ('tumor', 'Phenotype', 'HP:0002664', (67, 72)) ('AML', 'Disease', (150, 153)) ('AML', 'Phenotype', 'HP:0004808', (150, 153)) ('tumors', 'Phenotype', 'HP:0002664', (67, 73)) ('better', 'PosReg', (169, 175)) ('tumors', 'Disease', (67, 73)) ('patients', 'Species', '9606', (132, 140)) 907 26369982 We conducted an extensive literature search to identify potential genetic interactions and therapeutic implications of the proteins significantly affected by MLN4924 treatment in AML (Table S3). ('AML', 'Disease', (179, 182)) ('MLN4924 treatment', 'Var', (158, 175)) ('MLN4924', 'Chemical', 'MESH:C539933', (158, 165)) ('AML', 'Disease', 'MESH:D015470', (179, 182)) ('proteins', 'Protein', (123, 131)) ('affected', 'Reg', (146, 154)) ('AML', 'Phenotype', 'HP:0004808', (179, 182)) 908 26369982 Notably, several of the proteins modulated by MLN4924 in our study could represent biomarkers for patient stratification. ('MLN4924', 'Var', (46, 53)) ('proteins', 'Protein', (24, 32)) ('patient', 'Species', '9606', (98, 105)) ('MLN4924', 'Chemical', 'MESH:C539933', (46, 53)) 910 26369982 Considering this, it would be worthwhile to investigate whether patients with high basal CHD3 levels derive less benefit from treatment with MLN4924. ('MLN4924', 'Var', (141, 148)) ('patients', 'Species', '9606', (64, 72)) ('CHD3', 'Gene', (89, 93)) ('MLN4924', 'Chemical', 'MESH:C539933', (141, 148)) ('CHD3', 'Gene', '1107', (89, 93)) 911 26369982 We also detected drug-induced changes in 27 members of the RNA helicase family including the newly discovered DDX41, which has been found to be mutated in patients with AML and MDS (Table S4). ('mutated', 'Var', (144, 151)) ('MDS', 'Disease', (177, 180)) ('MDS', 'Disease', 'MESH:D009190', (177, 180)) ('MDS', 'Phenotype', 'HP:0002863', (177, 180)) ('AML', 'Disease', (169, 172)) ('DDX41', 'Gene', '51428', (110, 115)) ('AML', 'Phenotype', 'HP:0004808', (169, 172)) ('DDX41', 'Gene', (110, 115)) ('patients', 'Species', '9606', (155, 163)) ('RNA helicase', 'Protein', (59, 71)) ('AML', 'Disease', 'MESH:D015470', (169, 172)) 913 26369982 Notably, DDX24 was also upregulated (> 1.8-fold) in A375 melanoma cells following MLN4924 treatment. ('DDX24', 'Gene', '57062', (9, 14)) ('MLN4924', 'Chemical', 'MESH:C539933', (82, 89)) ('melanoma', 'Phenotype', 'HP:0002861', (57, 65)) ('melanoma', 'Disease', (57, 65)) ('A375', 'CellLine', 'CVCL:0132', (52, 56)) ('upregulated', 'PosReg', (24, 35)) ('melanoma', 'Disease', 'MESH:D008545', (57, 65)) ('MLN4924', 'Var', (82, 89)) ('DDX24', 'Gene', (9, 14)) 914 26369982 Although the impact of drug treatment on individual helicases fell below our set threshold of significance, the collective data suggest that MLN4924 may have a previously undefined class effect on RNA helicase function. ('MLN4924', 'Chemical', 'MESH:C539933', (141, 148)) ('RNA helicase', 'Protein', (197, 209)) ('MLN4924', 'Var', (141, 148)) ('function', 'MPA', (210, 218)) 915 26369982 Further investigation is required to assess whether NEDD8 plays a novel role in the regulation of RNA helicases and to determine how this may impact MLN4924 efficacy. ('impact', 'Reg', (142, 148)) ('MLN4924', 'Chemical', 'MESH:C539933', (149, 156)) ('regulation', 'MPA', (84, 94)) ('MLN4924 efficacy', 'Var', (149, 165)) ('NEDD8', 'Gene', '4738', (52, 57)) ('NEDD8', 'Gene', (52, 57)) ('RNA helicases', 'Protein', (98, 111)) 917 26369982 Notably, a number of the PD targets that were elevated in response to treatment with MLN4924 are directly actionable with existing approved and investigational drugs (Table S3). ('MLN4924', 'Var', (85, 92)) ('elevated', 'PosReg', (46, 54)) ('PD', 'Disease', 'MESH:D010300', (25, 27)) ('MLN4924', 'Chemical', 'MESH:C539933', (85, 92)) 918 26369982 We hypothesized that the ability of azacitidine (AZA) to antagonize ribonucleotide reductase (RRM2), a well-established mediator of resistance to many cytotoxic anticancer agents, may translate into increased sensitivity to MLN4924. ('sensitivity', 'MPA', (209, 220)) ('AZA', 'Chemical', 'MESH:D001374', (49, 52)) ('MLN4924', 'Var', (224, 231)) ('cancer', 'Phenotype', 'HP:0002664', (165, 171)) ('antagonize', 'NegReg', (57, 67)) ('azacitidine', 'Chemical', 'MESH:D001374', (36, 47)) ('RRM2', 'Gene', (94, 98)) ('cancer', 'Disease', (165, 171)) ('cancer', 'Disease', 'MESH:D009369', (165, 171)) ('MLN4924', 'Chemical', 'MESH:C539933', (224, 231)) ('increased', 'PosReg', (199, 208)) 919 26369982 To investigate this possibility, we first tested the ability of AZA to antagonize MLN4924-mediated RRM2 induction. ('antagonize', 'NegReg', (71, 81)) ('MLN4924', 'Chemical', 'MESH:C539933', (82, 89)) ('MLN4924-mediated', 'Var', (82, 98)) ('AZA', 'Chemical', 'MESH:D001374', (64, 67)) ('RRM2', 'Gene', (99, 103)) 922 26369982 The in vitro synergy we observed was recapitulated in a FLT3-ITD+ AML xenograft model where administration of the AZA/MLN4924 combination yielded significantly greater benefit than either monotherapy (P<0.05, Figure 1E). ('AML', 'Phenotype', 'HP:0004808', (66, 69)) ('AML', 'Disease', (66, 69)) ('benefit', 'PosReg', (168, 175)) ('AZA/MLN4924', 'Var', (114, 125)) ('AZA/MLN4924', 'Chemical', '-', (114, 125)) ('FLT3', 'Gene', (56, 60)) ('AML', 'Disease', 'MESH:D015470', (66, 69)) ('FLT3', 'Gene', '2322', (56, 60)) 924 26369982 These findings demonstrate proof of concept that proteomic profiling is a valuable approach to identify rational promising drug combinations and also support the investigation of a potential relationship between RRM2 expression and clinical response in the ongoing trial of MLN4924 plus AZA in elderly patients with AML (NCT0181426). ('RRM2', 'Gene', (212, 216)) ('AML', 'Disease', (316, 319)) ('AML', 'Disease', 'MESH:D015470', (316, 319)) ('MLN4924', 'Chemical', 'MESH:C539933', (274, 281)) ('MLN4924', 'Var', (274, 281)) ('AZA', 'Chemical', 'MESH:D001374', (287, 290)) ('patients', 'Species', '9606', (302, 310)) ('AML', 'Phenotype', 'HP:0004808', (316, 319)) 926 26369982 For example, the ability of MLN4924 to increase BRD2 levels may heighten the sensitivity of AML cells to BET inhibitors like OTX015. ('heighten', 'PosReg', (64, 72)) ('AML', 'Disease', 'MESH:D015470', (92, 95)) ('MLN4924', 'Var', (28, 35)) ('BET', 'Gene', '92737', (105, 108)) ('BRD2', 'Gene', (48, 52)) ('BRD2', 'Gene', '6046', (48, 52)) ('increase', 'PosReg', (39, 47)) ('BET', 'Gene', (105, 108)) ('AML', 'Phenotype', 'HP:0004808', (92, 95)) ('AML', 'Disease', (92, 95)) ('MLN4924', 'Chemical', 'MESH:C539933', (28, 35)) ('sensitivity', 'MPA', (77, 88)) 928 26369982 Considering that OTX015 and other BET inhibitors are currently in clinical trials, novel combination approaches with MLN4924 could be seamlessly incorporated into existing studies. ('BET', 'Gene', '92737', (34, 37)) ('BET', 'Gene', (34, 37)) ('MLN4924', 'Chemical', 'MESH:C539933', (117, 124)) ('MLN4924', 'Var', (117, 124)) 948 26120498 It showed that consolidation of first remission with 90Y-RIT was highly effective with no unexpected toxicities, producing a statistically significant longer time to progression in both PR and CR patients groups. ('toxicities', 'Disease', (101, 111)) ('longer', 'PosReg', (151, 157)) ('CR', 'Chemical', 'MESH:D002857', (193, 195)) ('patients', 'Species', '9606', (196, 204)) ('90Y-RIT', 'Var', (53, 60)) ('PR', 'Chemical', '-', (186, 188)) ('toxicities', 'Disease', 'MESH:D064420', (101, 111)) ('time', 'MPA', (158, 162)) 951 26120498 90Y-RIT also has been reported to be effective in patients with relapsed or refractory FL. ('90Y-RIT', 'Var', (0, 7)) ('relapsed', 'Disease', (64, 72)) ('patients', 'Species', '9606', (50, 58)) ('refractory FL', 'Disease', (76, 89)) 984 26120498 However, in the FIT study 8 patients who developed MDS/AML were treated with 90Y-RIT, suggesting a role played by 90Y-RIT in the risk of secondary MDS/AML, thus it is reasonable to consider monitoring these patients closely. ('patients', 'Species', '9606', (207, 215)) ('AML', 'Disease', (55, 58)) ('AML', 'Disease', 'MESH:D015470', (55, 58)) ('MDS', 'Disease', (51, 54)) ('AML', 'Disease', 'MESH:D015470', (151, 154)) ('90Y-RIT', 'Var', (114, 121)) ('AML', 'Disease', (151, 154)) ('patients', 'Species', '9606', (28, 36)) ('MDS', 'Disease', (147, 150)) ('MDS', 'Disease', 'MESH:D009190', (147, 150)) ('MDS', 'Disease', 'MESH:D009190', (51, 54)) 989 26120498 Hematologic toxicity occurring with FCR or with 90Y-RIT was clinically controllable and acceptable in a population composed mainly of patients with a history of prior treatment using rituximab plus chemotherapy. ('patients', 'Species', '9606', (134, 142)) ('FCR', 'Chemical', '-', (36, 39)) ('90Y-RIT', 'Var', (48, 55)) ('rituximab', 'Chemical', 'MESH:D000069283', (183, 192)) ('toxicity', 'Disease', 'MESH:D064420', (12, 20)) ('toxicity', 'Disease', (12, 20)) 991 26120498 90Y-RIT as consolidation appears to be best suited to patients with low burden of disease and may be more acceptable in those who are not candidates for high dose therapy/transplant approaches. ('low burden of disease', 'Disease', (68, 89)) ('patients', 'Species', '9606', (54, 62)) ('90Y-RIT', 'Var', (0, 7)) ('low burden of disease', 'Disease', 'MESH:D009800', (68, 89)) 992 26120498 The patients who were included in the current retrospective analysis had CD20+ histologically confirmed relapsed grade 1 or 2 follicular lymphoma and had received at least 1 prior treatment. ('follicular lymphoma', 'Disease', 'MESH:D008224', (126, 145)) ('follicular lymphoma', 'Disease', (126, 145)) ('lymphoma', 'Phenotype', 'HP:0002665', (137, 145)) ('patients', 'Species', '9606', (4, 12)) ('CD20+', 'Var', (73, 78)) 1002 26120498 Filgrastim was administered when the neutrophil count was less than 1x109/L and platelet support was planned for eventual episodes of bleeding and platelet count less than 15x109/L. ('bleeding', 'Disease', 'MESH:D006470', (134, 142)) ('less', 'Var', (58, 62)) ('bleeding', 'Disease', (134, 142)) 1013 33922062 Inhibiting ClpXP genetically or chemically impairs oxidative phosphorylation and is toxic to malignant cells with high ClpXP expression. ('oxidative phosphorylation', 'MPA', (51, 76)) ('Inhibiting', 'Var', (0, 10)) ('impairs', 'NegReg', (43, 50)) ('ClpX', 'Gene', '10845', (119, 123)) ('ClpX', 'Gene', (119, 123)) ('ClpX', 'Gene', '10845', (11, 15)) ('ClpX', 'Gene', (11, 15)) 1014 33922062 Likewise, hyperactivating the protease leads to increased degradation of ClpXP substrates and kills cancer cells. ('increased', 'PosReg', (48, 57)) ('hyperactivating', 'Var', (10, 25)) ('protease', 'Enzyme', (30, 38)) ('cancer', 'Phenotype', 'HP:0002664', (100, 106)) ('ClpX', 'Gene', (73, 77)) ('cancer', 'Disease', (100, 106)) ('cancer', 'Disease', 'MESH:D009369', (100, 106)) ('ClpX', 'Gene', '10845', (73, 77)) 1037 33922062 Furthermore, in response to stimuli such as increased ROS production, dissipation of mitochondrial membrane potential, heat shock, and loss of mitochondrial DNA (mtDNA), this enzyme performs central regulatory roles by complete or partial proteolysis of specific proteins, including OPA1 (optic atrophy 1), TIMM17A (translocase of inner mitochondrial membrane 17A) and ROMO1 (Reactive oxygen species modulator 1). ('ROMO1', 'Gene', '140823', (369, 374)) ('OPA1', 'Gene', (283, 287)) ('loss', 'Var', (135, 139)) ('optic atrophy 1', 'Gene', '4976', (289, 304)) ('optic atrophy', 'Phenotype', 'HP:0000648', (289, 302)) ('TIMM17A', 'Gene', '10440', (307, 314)) ('ROS', 'MPA', (54, 57)) ('translocase of inner mitochondrial membrane 17A', 'Gene', '10440', (316, 363)) ('translocase of inner mitochondrial membrane 17A', 'Gene', (316, 363)) ('increased', 'PosReg', (44, 53)) ('ROS', 'Chemical', 'MESH:D017382', (54, 57)) ('oxygen', 'Chemical', 'MESH:D010100', (385, 391)) ('TIMM17A', 'Gene', (307, 314)) ('increased ROS production', 'Phenotype', 'HP:0025464', (44, 68)) ('optic atrophy 1', 'Gene', (289, 304)) ('OPA1', 'Gene', '4976', (283, 287)) ('dissipation', 'MPA', (70, 81)) ('shock', 'Phenotype', 'HP:0031273', (124, 129)) ('ROMO1', 'Gene', (369, 374)) 1047 33922062 m-AAA also implements vital functions which are essential in neuronal cells and mutations in AFG3L2 and SPG7 result in neurodegenerative phenotypes. ('AAA', 'Gene', (2, 5)) ('AFG3L2', 'Gene', (93, 99)) ('mutations', 'Var', (80, 89)) ('neurodegenerative phenotypes', 'CPA', (119, 147)) ('SPG7', 'Gene', '6687', (104, 108)) ('result in', 'Reg', (109, 118)) ('AAA', 'Gene', '351', (2, 5)) ('AFG3L2', 'Gene', '10939', (93, 99)) ('SPG7', 'Gene', (104, 108)) 1078 33922062 For example, LonP1 knockdown in human lung fibroblasts and bladder cancer cells causes caspase-3-dependent cell death and suppresses cell proliferation. ('death', 'Disease', 'MESH:D003643', (112, 117)) ('death', 'Disease', (112, 117)) ('cell proliferation', 'CPA', (133, 151)) ('bladder cancer', 'Disease', 'MESH:D001749', (59, 73)) ('knockdown', 'Var', (19, 28)) ('caspase-3', 'Gene', '836', (87, 96)) ('bladder cancer', 'Phenotype', 'HP:0009725', (59, 73)) ('suppresses', 'NegReg', (122, 132)) ('cancer', 'Phenotype', 'HP:0002664', (67, 73)) ('caspase-3', 'Gene', (87, 96)) ('LonP1', 'Gene', (13, 18)) ('human', 'Species', '9606', (32, 37)) ('LonP1', 'Gene', '9361', (13, 18)) ('bladder cancer', 'Disease', (59, 73)) 1079 33922062 LonP1 knockdown also reduces cell proliferation in human mantle cell lymphoma and non-small-cell lung cancer (NSCLC) cell lines. ('human', 'Species', '9606', (51, 56)) ('NSCLC', 'Disease', 'MESH:D002289', (110, 115)) ('LonP1', 'Gene', (0, 5)) ('NSCLC', 'Disease', (110, 115)) ('small-cell lung cancer', 'Phenotype', 'HP:0030357', (86, 108)) ('non-small-cell lung cancer', 'Phenotype', 'HP:0030358', (82, 108)) ('lung cancer', 'Phenotype', 'HP:0100526', (97, 108)) ('mantle cell lymphoma', 'Disease', 'MESH:D020522', (57, 77)) ('NSCLC', 'Phenotype', 'HP:0030358', (110, 115)) ('mantle cell lymphoma', 'Disease', (57, 77)) ('cell proliferation', 'CPA', (29, 47)) ('lymphoma', 'Phenotype', 'HP:0002665', (69, 77)) ('cell lymphoma', 'Phenotype', 'HP:0012191', (64, 77)) ('LonP1', 'Gene', '9361', (0, 5)) ('reduces', 'NegReg', (21, 28)) ('cancer', 'Phenotype', 'HP:0002664', (102, 108)) ('non-small-cell lung cancer', 'Disease', (82, 108)) ('knockdown', 'Var', (6, 15)) ('non-small-cell lung cancer', 'Disease', 'MESH:D002289', (82, 108)) 1080 33922062 Similarly, siRNA knockdown of LonP1 reduces the viability of human malignant glioma cells and radically impairs glioma cell survival under hypoxic conditions. ('impairs glioma', 'Disease', 'MESH:D005910', (104, 118)) ('malignant glioma', 'Disease', (67, 83)) ('impairs glioma', 'Disease', (104, 118)) ('malignant glioma', 'Disease', 'MESH:D005910', (67, 83)) ('LonP1', 'Gene', (30, 35)) ('reduces', 'NegReg', (36, 43)) ('knockdown', 'Var', (17, 26)) ('LonP1', 'Gene', '9361', (30, 35)) ('human', 'Species', '9606', (61, 66)) ('glioma', 'Phenotype', 'HP:0009733', (112, 118)) ('glioma', 'Phenotype', 'HP:0009733', (77, 83)) 1081 33922062 Conversely, LonP1 overexpression promotes cancer cell proliferation, enhances colony formation, and more importantly, increases cellular resistance to apoptosis-inducing reagents. ('promotes', 'PosReg', (33, 41)) ('cancer', 'Phenotype', 'HP:0002664', (42, 48)) ('colony formation', 'CPA', (78, 94)) ('LonP1', 'Gene', (12, 17)) ('overexpression', 'Var', (18, 32)) ('cancer', 'Disease', (42, 48)) ('cellular', 'MPA', (128, 136)) ('enhances', 'PosReg', (69, 77)) ('increases', 'PosReg', (118, 127)) ('cancer', 'Disease', 'MESH:D009369', (42, 48)) ('LonP1', 'Gene', '9361', (12, 17)) 1083 33922062 CDDO, in the micromolar range, induces caspase-dependent apoptosis in several human cancer cells with high LonP1 expression, comprising colon carcinoma, B-cell lymphoma, breast ductal carcinoma, and liver hepatocellular carcinoma. ('cancer', 'Disease', 'MESH:D009369', (84, 90)) ('colon carcinoma', 'Disease', (136, 151)) ('carcinoma', 'Phenotype', 'HP:0030731', (184, 193)) ('liver hepatocellular carcinoma', 'Disease', 'MESH:D006528', (199, 229)) ('ductal carcinoma', 'Phenotype', 'HP:0030075', (177, 193)) ('induces', 'Reg', (31, 38)) ('carcinoma', 'Phenotype', 'HP:0030731', (142, 151)) ('lymphoma', 'Phenotype', 'HP:0002665', (160, 168)) ('LonP1', 'Gene', '9361', (107, 112)) ('high', 'Var', (102, 106)) ('cell lymphoma', 'Phenotype', 'HP:0012191', (155, 168)) ('colon carcinoma', 'Disease', 'MESH:D003110', (136, 151)) ('carcinoma', 'Phenotype', 'HP:0030731', (220, 229)) ('human', 'Species', '9606', (78, 83)) ('liver hepatocellular carcinoma', 'Disease', (199, 229)) ('cancer', 'Disease', (84, 90)) ('B-cell lymphoma', 'Phenotype', 'HP:0012191', (153, 168)) ('cancer', 'Phenotype', 'HP:0002664', (84, 90)) ('CDDO', 'Chemical', '-', (0, 4)) ('breast ductal carcinoma', 'Phenotype', 'HP:0003002', (170, 193)) ('caspase-dependent', 'MPA', (39, 56)) ('LonP1', 'Gene', (107, 112)) ('B-cell lymphoma, breast ductal carcinoma', 'Disease', 'MESH:D001943', (153, 193)) ('hepatocellular carcinoma', 'Phenotype', 'HP:0001402', (205, 229)) 1089 33922062 Inactivating mutations in other mitochondrial proteases leads to developmental, metabolic, and neurological disorders such as CODAS syndrome (CODASS), Perrault syndrome 3 (PRLTS3), optic atrophy 11 (OPA11), erythropoietic protoporphyria (EPP), spinocerebellar ataxia 28 (SCA28), and hereditary spastic paraplegia 7 (HSP7). ('ataxia', 'Phenotype', 'HP:0001251', (260, 266)) ('neurological disorders', 'Disease', 'MESH:D009422', (95, 117)) ('Perrault syndrome 3', 'Disease', (151, 170)) ('hereditary spastic paraplegia', 'Disease', 'MESH:D015419', (283, 312)) ('Inactivating mutations', 'Var', (0, 22)) ('leads to', 'Reg', (56, 64)) ('developmental', 'CPA', (65, 78)) ('metabolic', 'CPA', (80, 89)) ('OPA1', 'Gene', '4976', (199, 203)) ('optic atrophy 1', 'Gene', (181, 196)) ('paraplegia', 'Phenotype', 'HP:0010550', (302, 312)) ('spinocerebellar ataxia', 'Disease', (244, 266)) ('spinocerebellar ataxia', 'Disease', 'MESH:D020754', (244, 266)) ('spastic paraplegia', 'Phenotype', 'HP:0001258', (294, 312)) ('neurological disorders', 'Disease', (95, 117)) ('OPA1', 'Gene', (199, 203)) ('optic atrophy', 'Phenotype', 'HP:0000648', (181, 194)) ('erythropoietic protoporphyria', 'Disease', (207, 236)) ('hereditary spastic paraplegia', 'Disease', (283, 312)) ('CODAS syndrome', 'Disease', (126, 140)) ('optic atrophy 1', 'Gene', '4976', (181, 196)) 1090 33922062 Likewise, Lon is essential for survival in mammals, and homozygous deletion of LONP causes early embryonic lethality. ('deletion', 'Var', (67, 75)) ('LONP', 'Gene', '9361', (79, 83)) ('Lon', 'Gene', (10, 13)) ('LONP', 'Gene', (79, 83)) ('Lon', 'Gene', '9361', (10, 13)) ('embryonic lethality', 'Disease', 'MESH:D020964', (97, 116)) ('embryonic lethality', 'Disease', (97, 116)) ('causes', 'Reg', (84, 90)) 1093 33922062 In support of this hypothesis, ClpP -/- mice are viable, however, slightly smaller, infertile, and have acquired hearing loss. ('infertile', 'Disease', (84, 93)) ('hearing loss', 'Phenotype', 'HP:0000365', (113, 125)) ('mice', 'Species', '10090', (40, 44)) ('hearing loss', 'Disease', (113, 125)) ('infertile', 'Disease', 'MESH:D007247', (84, 93)) ('ClpP -/-', 'Var', (31, 39)) ('hearing loss', 'Disease', 'MESH:D034381', (113, 125)) 1096 33922062 Therefore, inhibiting ClpXP may have a therapeutic window in the treatment of malignancy, and we focus the remainder of this review on ClpXP as a potential therapeutic target for cancer therapy. ('ClpX', 'Gene', '10845', (135, 139)) ('malignancy', 'Disease', (78, 88)) ('ClpX', 'Gene', (135, 139)) ('inhibiting', 'Var', (11, 21)) ('ClpX', 'Gene', (22, 26)) ('cancer', 'Phenotype', 'HP:0002664', (179, 185)) ('ClpX', 'Gene', '10845', (22, 26)) ('cancer', 'Disease', 'MESH:D009369', (179, 185)) ('cancer', 'Disease', (179, 185)) ('malignancy', 'Disease', 'MESH:D009369', (78, 88)) 1099 33922062 In addition, optimal biogenesis and OXPHOS function are crucial for metastatic dissemination as shown with PGC-1alpha (peroxisome proliferator-activated receptor gamma coactivator 1-alpha) silencing in the breast cancer model. ('silencing', 'Var', (189, 198)) ('cancer', 'Phenotype', 'HP:0002664', (213, 219)) ('breast cancer', 'Disease', 'MESH:D001943', (206, 219)) ('PGC-1alpha', 'Gene', (107, 117)) ('breast cancer', 'Disease', (206, 219)) ('peroxisome proliferator-activated receptor gamma coactivator 1-alpha', 'Gene', '10891', (119, 187)) ('breast cancer', 'Phenotype', 'HP:0003002', (206, 219)) ('PGC-1alpha', 'Gene', '10891', (107, 117)) 1105 33922062 Genetic and chemical inhibition of ClpP reduces leukemia cell growth and viability and targets leukemic stem cells in vitro and in vivo (Figure 2). ('targets', 'Reg', (87, 94)) ('leukemia', 'Disease', (48, 56)) ('leukemia', 'Phenotype', 'HP:0001909', (48, 56)) ('leukemia', 'Disease', 'MESH:D007938', (48, 56)) ('inhibition', 'Var', (21, 31)) ('reduces', 'NegReg', (40, 47)) ('ClpP', 'Gene', (35, 39)) 1107 33922062 Interestingly, hyperactivation of ClpP also effectively targets AML cells by uncontrolled degradation of OXPHOS subunits. ('hyperactivation', 'Var', (15, 30)) ('AML', 'Disease', 'MESH:D015470', (64, 67)) ('ClpP', 'Gene', (34, 38)) ('AML', 'Phenotype', 'HP:0004808', (64, 67)) ('AML', 'Disease', (64, 67)) ('degradation of OXPHOS subunits', 'MPA', (90, 120)) 1108 33922062 Induction of constitutively active ClpP mutant (Y118A) and hyperactivation by imipridones increases degradation of respiratory chain proteins and metabolic enzymes resulting in impaired respiratory function. ('impaired respiratory function', 'Phenotype', 'HP:0002093', (177, 206)) ('Y118A', 'Mutation', 'p.Y118A', (48, 53)) ('degradation of respiratory chain proteins', 'MPA', (100, 141)) ('Y118A', 'Var', (48, 53)) ('impaired', 'NegReg', (177, 185)) ('increases', 'PosReg', (90, 99)) ('ClpP', 'Gene', (35, 39)) ('imipridones', 'Chemical', '-', (78, 89)) ('metabolic enzymes', 'MPA', (146, 163)) ('respiratory function', 'MPA', (186, 206)) 1110 33922062 ClpP hyperactivation induces apoptosis in leukemia and lymphoma cells preferentially over normal cells. ('apoptosis', 'CPA', (29, 38)) ('induces', 'Reg', (21, 28)) ('lymphoma', 'Disease', (55, 63)) ('leukemia', 'Disease', (42, 50)) ('leukemia', 'Phenotype', 'HP:0001909', (42, 50)) ('leukemia', 'Disease', 'MESH:D007938', (42, 50)) ('ClpP', 'Gene', (0, 4)) ('lymphoma', 'Disease', 'MESH:D008223', (55, 63)) ('hyperactivation', 'Var', (5, 20)) ('lymphoma', 'Phenotype', 'HP:0002665', (55, 63)) 1113 33922062 siRNA silencing of ClpP or ClpX in PC3 cells significantly suppresses proliferation and colony formation by decreasing the expression of cyclins A, B1, and D1 and inducing cell cycle arrest. ('cell cycle arrest', 'Phenotype', 'HP:0011018', (172, 189)) ('expression', 'MPA', (123, 133)) ('ClpP', 'Gene', (19, 23)) ('cyclins A, B1, and D1', 'Gene', '890;891;595', (137, 158)) ('arrest', 'Disease', 'MESH:D006323', (183, 189)) ('decreasing', 'NegReg', (108, 118)) ('proliferation', 'CPA', (70, 83)) ('colony formation', 'CPA', (88, 104)) ('ClpX', 'Gene', '10845', (27, 31)) ('ClpX', 'Gene', (27, 31)) ('inducing', 'Reg', (163, 171)) ('PC3', 'CellLine', 'CVCL:0035', (35, 38)) ('arrest', 'Disease', (183, 189)) ('suppresses', 'NegReg', (59, 69)) ('silencing', 'Var', (6, 15)) 1117 33922062 In addition, loss of ClpP suppresses cell growth, migration, and colony formation in the breast cancer cell lines MDA-MB-231 and ZR-75-1. ('breast cancer', 'Disease', 'MESH:D001943', (89, 102)) ('cancer', 'Phenotype', 'HP:0002664', (96, 102)) ('breast cancer', 'Disease', (89, 102)) ('migration', 'CPA', (50, 59)) ('suppresses', 'NegReg', (26, 36)) ('breast cancer', 'Phenotype', 'HP:0003002', (89, 102)) ('cell growth', 'CPA', (37, 48)) ('ClpP', 'Gene', (21, 25)) ('loss', 'Var', (13, 17)) ('MDA-MB-231', 'CellLine', 'CVCL:0062', (114, 124)) ('colony formation', 'CPA', (65, 81)) 1118 33922062 In contrast, in the breast adenocarcinoma cell line MCF-7, ClpP is highly expressed but the loss of ClpP had minimal effects on cell proliferation which suggests the importance of ClpP expression in cancer pathology may be cell-type specific. ('cancer', 'Phenotype', 'HP:0002664', (199, 205)) ('breast adenocarcinoma', 'Disease', 'MESH:D001943', (20, 41)) ('carcinoma', 'Phenotype', 'HP:0030731', (32, 41)) ('cancer', 'Disease', 'MESH:D009369', (199, 205)) ('MCF-7', 'CellLine', 'CVCL:0031', (52, 57)) ('cell proliferation', 'CPA', (128, 146)) ('ClpP', 'Gene', (100, 104)) ('breast adenocarcinoma', 'Phenotype', 'HP:0003002', (20, 41)) ('loss', 'Var', (92, 96)) ('cancer', 'Disease', (199, 205)) ('breast adenocarcinoma', 'Disease', (20, 41)) 1123 33922062 In cell culture models, A2-32-01 inhibits AML cell viability and reduces clonogenic growth (Figure 2). ('AML', 'Phenotype', 'HP:0004808', (42, 45)) ('AML', 'Disease', (42, 45)) ('reduces', 'NegReg', (65, 72)) ('clonogenic growth', 'CPA', (73, 90)) ('AML', 'Disease', 'MESH:D015470', (42, 45)) ('inhibits', 'NegReg', (33, 41)) ('A2-32-01', 'Var', (24, 32)) 1124 33922062 In vivo, daily treatment of A2-32-01 reduces leukemic growth without liver, muscle, or renal toxicity. ('A2-32-01', 'Var', (28, 36)) ('renal toxicity', 'Disease', 'MESH:D007674', (87, 101)) ('leukemic growth', 'Disease', (45, 60)) ('leukemic growth', 'Disease', 'MESH:D006130', (45, 60)) ('renal toxicity', 'Disease', (87, 101)) ('reduces', 'NegReg', (37, 44)) 1130 33922062 However, only the AV167 inhibits human ClpP peptidase activity. ('AV167', 'Var', (18, 23)) ('AV167', 'Chemical', '-', (18, 23)) ('inhibits', 'NegReg', (24, 32)) ('human ClpP peptidase', 'Enzyme', (33, 53)) ('human', 'Species', '9606', (33, 38)) ('activity', 'MPA', (54, 62)) 1131 33922062 To improve selectivity for human ClpP, the modified analogues TG42, TG43, and TG53 were generated by substituting a naphtofuran moiety at position-2. ('substituting', 'Var', (101, 113)) ('improve', 'PosReg', (3, 10)) ('TG42', 'Chemical', '-', (62, 66)) ('human', 'Species', '9606', (27, 32)) ('TG43', 'Chemical', '-', (68, 72)) ('TG53', 'Var', (78, 82)) ('naphtofuran', 'Chemical', '-', (116, 127)) 1155 33922062 AML, acute lymphoblastic leukemia (ALL) and breast cancer cells with inactive mutant ClpP (D190A) or ClpP knockout are resistant to ONC201 and ONC212, indicating the functional importance of ClpP for imipridones efficacy. ('cancer', 'Phenotype', 'HP:0002664', (51, 57)) ('acute lymphoblastic leukemia', 'Disease', (5, 33)) ('D190A', 'Mutation', 'p.D190A', (91, 96)) ('acute lymphoblastic leukemia', 'Disease', 'MESH:D054198', (5, 33)) ('knockout', 'Var', (106, 114)) ('AML', 'Disease', 'MESH:D015470', (0, 3)) ('acute lymphoblastic leukemia', 'Phenotype', 'HP:0006721', (5, 33)) ('AML', 'Phenotype', 'HP:0004808', (0, 3)) ('imipridones', 'Chemical', '-', (200, 211)) ('AML', 'Disease', (0, 3)) ('ONC212', 'CellLine', 'CVCL:0U35', (143, 149)) ('breast cancer', 'Phenotype', 'HP:0003002', (44, 57)) ('lymphoblastic leukemia', 'Phenotype', 'HP:0005526', (11, 33)) ('ClpP', 'Gene', (101, 105)) ('mutant', 'Var', (78, 84)) ('breast cancer', 'Disease', 'MESH:D001943', (44, 57)) ('leukemia', 'Phenotype', 'HP:0001909', (25, 33)) ('breast cancer', 'Disease', (44, 57)) ('ClpP', 'Gene', (85, 89)) ('ALL', 'Phenotype', 'HP:0006721', (35, 38)) 1160 33922062 Transient DRD2 knockdown also activates the integrated stress response. ('knockdown', 'Var', (15, 24)) ('DRD2', 'Gene', (10, 14)) ('integrated', 'CPA', (44, 54)) ('DRD2', 'Gene', '1813', (10, 14)) ('activates', 'PosReg', (30, 39)) 1164 33922062 Aberrant activities of ATP-dependent proteases are associated with pathologies ranging from neurodegenerative diseases to cancer. ('ATP', 'Chemical', 'MESH:D000255', (23, 26)) ('ATP-dependent proteases', 'Enzyme', (23, 46)) ('neurodegenerative diseases', 'Disease', 'MESH:D019636', (92, 118)) ('Aberrant', 'Var', (0, 8)) ('cancer', 'Disease', (122, 128)) ('cancer', 'Disease', 'MESH:D009369', (122, 128)) ('activities', 'MPA', (9, 19)) ('associated', 'Reg', (51, 61)) ('neurodegenerative diseases', 'Disease', (92, 118)) ('neurodegenerative diseases', 'Phenotype', 'HP:0002180', (92, 118)) ('cancer', 'Phenotype', 'HP:0002664', (122, 128)) 1166 33922062 Of all the mitochondrial proteases, ClpXP is unique in that humans, animals, and cells remain viable with mild phenotypes despite depletion or mutation of this protease. ('ClpX', 'Gene', (36, 40)) ('ClpX', 'Gene', '10845', (36, 40)) ('mutation', 'Var', (143, 151)) ('humans', 'Species', '9606', (60, 66)) ('depletion', 'MPA', (130, 139)) 1178 29551034 t(8;21) (AML) (MDS) AML 1 AML t(8;21) t(8;21)/RUNX1-RUNX1T1 2 , , (MFC) WT1 (MRD) , MDS 2 (t-MDS RAEB-2) , ,36 " 2 " 2015 4 :WBC 2.07x109/L,RBC 2.75x1012/L,HGB 94 g/L,PLT 36x109/L; 0.02 : , 0.075, 0.040, 0.480; AML-M2b :CD34 HLA-DR MPO CD117 CD33 CD13 CD15 ,CD19 CD56 :46,XX, t(8;21)(q22;q22)[16]/46,XX[4] RUNX1-RUNX1T1 41.3%,WT1 126.91% 112 (NGS) WT1 ( 8 p.His448As, 30%) :AML-M2 t(8;21)(q22;q22)/RUNX1-RUNX1T1 HAD(7+7+3) ( 2 mg m-2 d-1, , 1~7 ; 100 mg m-2 d-1, , 1~7 ; 40 mg m-2 d-1, , 1~3 ) 1 , (CRi) DA( 40 mg m-2 d-1, , 1~3 ; 1.5 g/m2, 12 h 1 , , 1~3 ) MA( 6 mg m-2 d-1, , 1~3 ; 1.5 g/m2, 12 h 1 , , 1~3 ) HA( 2.5 mg m-2 d-1, , 1~6 ; 100 mg m-2 d-1, , 1~6 )x2 MA( 6 mg m-2 d-1, , 1~3 ; 100 mg m-2 d-1, , 1~6 ) 2015 12 3 :HGB 80~118 g/L,WBC(0.94~4.20)x109/L,PLT(20~420)x109/L 2016 3 WT1 (>2%), 4.8%, 2 12%; MFC-MRD 0 1.1% 2017 3 " " : , 0.445, 0.175, ; 0.085; : , :45,XX,-7[6]/46,XX[1] RUNX1-RUNX1T1 0,WT1 4.87% AA ( 20 mg/d, , 1~6 ; 100 mg m-2 d-1, , 1~6 ) 1 2017 4 :WBC 3.7x109/L,RBC 2.66x1012/L,HGB 85 g/L,PLT 366x109/L; : 6%, 25%, 36%, 25%, 8% : (-), 0.410, 0.170, 0.030, 0.100, 0.090; :CD34 HLA-DR CD38dim CD117 CD13 CD33 CD7 9.47%, 30.42% :45,XX,-7[7]/46,XX[13] 7 CEP7 D7S486 (FISH) 64.6%, 0 RUNX1-RUNX1T1 0,WT1 27.51% 112 NGS WT1 : MDS(RAEB-2) (allo-HSCT) MDS/AML(t-MDS/AML) t-MDS/AML AML MDS 10%~20%, t-MDS/AML ,5 10%, ,t-MDS/AML AML t-MDS/AML , (APL) , (auto-HSCT) AML ,AML t(8;21) t-MDS/AML 7 ( 1) t(8;21)AML 2 , , t-MDS(RAEB-2) AML t-MDS/AML AML , , >=5%, ; , , t-MDS/AML , , ; , t-MDS/AML , , , AML t-MDS/AML ,MRD , , t(8;21) RUNX1-RUNX1T1 , WT1 , -7 , t-MDS , AML t-MDS/AML , , t-MDS/AML t-MDS/AML , , (5~10 ), ( 5 7 ), , t-MDS , , AML , 1 II(Topo II) t-AML/MDS 20%~30%, , 1~5 , AML, MDS , , 11q23/MLL 21q22/RUNX1 , t(15;17) inv(16)/t(16;16) , AML t-MDS/AML Topo II , Topo II , , 1 , , , -7 , t-MDS , <5% t-MDS t-AML , t-MDS t-AML t-MDS/AML MDS/AML Zeidan t-MDS MDS 19 46 , (P<0.005) t-AML AML CR , allo-HSCT t-MDS/AML , , AML t-AML/AML , , , t-MDS/AML " " " " MRD, , , , , ('AML', 'Disease', (51, 54)) ('RUNX1', 'Gene', '861', (86, 91)) ('AML', 'Disease', (2535, 2538)) ('AML', 'Disease', 'MESH:D015470', (2066, 2069)) ('45,XX,-7', 'STRUCTURAL_ABNORMALITY', 'None', (1340, 1348)) ('AML', 'Disease', (3120, 3123)) ('RUNX1', 'Gene', (1830, 1835)) ('AML', 'Disease', 'MESH:D015470', (3042, 3045)) ('AML', 'Disease', (1961, 1964)) ('RUNX1', 'Gene', '861', (1830, 1835)) ('CD117', 'Gene', (1683, 1688)) ('WT1', 'Gene', (1891, 1894)) ('CD56', 'Gene', (433, 437)) ('RUNX1', 'Gene', (623, 628)) ('AML', 'Disease', (9, 12)) ('AML MDS', 'Disease', 'MESH:D015470', (3449, 3462)) ('RUNX1', 'Gene', '861', (623, 628)) ('AML', 'Disease', (2348, 2351)) ('AML', 'Disease', 'MESH:D015470', (2137, 2140)) ('AML', 'Disease', (2150, 2153)) ('CD33', 'Gene', '945', (411, 415)) ('MDS 2', 'Gene', (164, 180)) ('AML', 'Disease', 'MESH:D015470', (2016, 2019)) ('AML', 'Disease', (3238, 3241)) ('CD34', 'Gene', '947', (1663, 1667)) ('AML AML', 'Disease', (2348, 2359)) ('AML', 'Disease', 'MESH:D015470', (2165, 2168)) ('WT1', 'Gene', '7490', (1891, 1894)) ('t(8;21)(q22;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (600, 616)) ('AML', 'Disease', (3617, 3620)) ('RUNX1T1', 'Gene', (2807, 2814)) ('AML', 'Disease', (2333, 2336)) ('AML AML', 'Disease', 'MESH:D015470', (2348, 2359)) ('WT1', 'Gene', '7490', (1851, 1854)) ('WT1', 'Gene', (1151, 1154)) ('AML', 'Disease', 'MESH:D015470', (2356, 2359)) ('CD13', 'Gene', (416, 420)) ('AML', 'Disease', (2943, 2946)) ('RUNX1T1', 'Gene', '862', (491, 498)) ('RUNX1', 'Gene', '861', (80, 85)) ('AML', 'Disease', 'MESH:D015470', (2862, 2865)) ('AML', 'Disease', (3432, 3435)) ('AML', 'Disease', (2639, 2642)) ('AML', 'Disease', (3703, 3706)) ('CD19', 'Gene', '930', (428, 432)) ('CD33', 'Gene', '945', (1694, 1698)) ('WT1', 'Gene', '7490', (1151, 1154)) ('CD117', 'Gene', '3815', (1683, 1688)) ('RUNX1', 'Gene', (491, 496)) ('AML AML', 'Disease', 'MESH:D015470', (2137, 2153)) ('CD34', 'Gene', '947', (389, 393)) ('RUNX1', 'Gene', (86, 91)) ('AML', 'Disease', 'MESH:D015470', (3120, 3123)) ('AML', 'Disease', (2225, 2228)) ('AML', 'Disease', (368, 371)) ('AML', 'Disease', (2020, 2023)) ('RUNX1', 'Gene', '861', (491, 496)) ('AML CR', 'Gene', (3534, 3546)) ('RUNX1', 'Gene', (2807, 2812)) ('RUNX1T1', 'Gene', (86, 93)) ('AML', 'Disease', 'MESH:D015470', (2248, 2251)) ('CD13', 'Gene', '290', (416, 420)) ('AML', 'Disease', 'MESH:D015470', (34, 37)) ('45,XX,-7', 'STRUCTURAL_ABNORMALITY', 'None', (1742, 1750)) ('RUNX1', 'Gene', '861', (2807, 2812)) ('CD15', 'Gene', (421, 425)) ('WT1', 'Gene', (510, 513)) ('AML AML MDS', 'Disease', 'MESH:D015470', (2016, 2027)) ('AML', 'Disease', 'MESH:D015470', (2271, 2274)) ('AML', 'Disease', (2216, 2219)) ('AML', 'Disease', (3222, 3225)) ('AML', 'Disease', 'MESH:D015470', (3594, 3597)) ('AML 2', 'Gene', (2271, 2292)) ('AML', 'Disease', (2875, 2878)) ('AML MDS', 'Disease', (3449, 3462)) ('AML', 'Disease', (3628, 3631)) ('WT1', 'Gene', (2826, 2829)) ('AML', 'Disease', (3079, 3082)) ('AML', 'Disease', (2921, 2924)) ('AML 2', 'Gene', '864', (2271, 2292)) ('WT1', 'Gene', '7490', (510, 513)) ('AML', 'Disease', (2722, 2725)) ('AML', 'Disease', 'MESH:D015470', (3617, 3620)) ('CD15', 'Gene', '2526', (421, 425)) ('WT1', 'Gene', (1851, 1854)) ('HAD', 'Disease', 'MESH:C535310', (638, 641)) ('CD19', 'Gene', (428, 432)) ('RUNX1T1', 'Gene', '862', (1367, 1374)) ('WT1', 'Gene', '7490', (2826, 2829)) ('AML', 'Disease', 'MESH:D015470', (3632, 3635)) ('AML', 'Disease', 'MESH:D015470', (3432, 3435)) ('RUNX1', 'Gene', (80, 85)) ('RUNX1', 'Gene', (3167, 3172)) ('CD38', 'Gene', '952', (1675, 1679)) ('AML', 'Disease', (3401, 3404)) ('AML', 'Disease', (3449, 3452)) ('AML', 'Disease', 'MESH:D015470', (1971, 1974)) ('AML', 'Disease', 'MESH:D015470', (2225, 2228)) ('RUNX1', 'Gene', '861', (3167, 3172)) ('AML', 'Disease', 'MESH:D015470', (51, 54)) ('AML', 'Disease', 'MESH:D015470', (2535, 2538)) ('AML', 'Disease', (3534, 3537)) ('RUNX1T1', 'Gene', (491, 498)) ('AML', 'Disease', (3463, 3466)) ('AML', 'Disease', 'MESH:D015470', (1961, 1964)) ('AML', 'Disease', (2066, 2069)) ('AML CR', 'Gene', '9163', (3534, 3546)) ('AML', 'Disease', (2708, 2711)) ('AML', 'Disease', 'MESH:D015470', (2875, 2878)) ('RUNX1', 'Gene', (1361, 1366)) ('CD117', 'Gene', (405, 410)) ('AML', 'Disease', 'MESH:D015470', (9, 12)) ('AML', 'Disease', (3042, 3045)) ('AML', 'Disease', 'MESH:D015470', (2722, 2725)) ('RUNX1', 'Gene', '861', (1361, 1366)) ('AML', 'Disease', 'MESH:D015470', (2348, 2351)) ('WT1', 'Gene', '7490', (1382, 1385)) ('AML', 'Disease', 'MESH:D015470', (2150, 2153)) ('RUNX1T1', 'Gene', '862', (623, 630)) ('RUNX1T1', 'Gene', '862', (1836, 1843)) ('AML AML MDS', 'Disease', (2016, 2027)) ('AML', 'Disease', 'MESH:D015470', (3238, 3241)) ('MDS 2', 'Gene', '259283', (164, 180)) ('WT1 , -7', 'Gene', '7490', (2826, 2843)) ('AML 1', 'Gene', '861', (34, 50)) ('AML', 'Disease', 'MESH:D015470', (2333, 2336)) ('AML', 'Disease', (2016, 2019)) ('AML', 'Disease', (2165, 2168)) ('p.His448A', 'Var', (570, 579)) ('AML', 'Disease', 'MESH:D015470', (2943, 2946)) ('AML', 'Disease', (593, 596)) ('AML', 'Disease', (2356, 2359)) ('AML', 'Disease', 'MESH:D015470', (2639, 2642)) ('AML', 'Disease', 'MESH:D015470', (3703, 3706)) ('AML', 'Disease', (2862, 2865)) ('APL', 'Disease', (2185, 2188)) ('p.His448A', 'SUBSTITUTION', 'None', (570, 579)) ('AML', 'Disease', 'MESH:D015470', (368, 371)) ('RUNX1T1', 'Gene', (1367, 1374)) ('AML', 'Disease', 'MESH:D015470', (2020, 2023)) ('CD117', 'Gene', '3815', (405, 410)) ('CD34', 'Gene', (1663, 1667)) ('AML', 'Disease', 'MESH:D015470', (3463, 3466)) ('AML', 'Disease', (3527, 3530)) ('HAD', 'Disease', (638, 641)) ('AML', 'Disease', 'MESH:D015470', (2708, 2711)) ('WT1', 'Gene', (552, 555)) ('AML', 'Disease', (2137, 2140)) ('CD13', 'Gene', (1689, 1693)) ('RUNX1T1', 'Gene', '862', (2807, 2814)) ('AML', 'Disease', (2248, 2251)) ('CD38', 'Gene', (1675, 1679)) ('WT1', 'Gene', (1382, 1385)) ('AML', 'Disease', (34, 37)) ('AML', 'Disease', 'MESH:D015470', (2216, 2219)) ('AML', 'Disease', 'MESH:D015470', (3222, 3225)) ('AML', 'Disease', 'MESH:D015470', (3628, 3631)) ('AML', 'Disease', (2271, 2274)) ('WT1', 'Gene', '7490', (552, 555)) ('AML', 'Disease', 'MESH:D015470', (3079, 3082)) ('AML', 'Disease', 'MESH:D015470', (2921, 2924)) ('AML AML', 'Disease', (2137, 2153)) ('RUNX1', 'Gene', (1836, 1841)) ('AML', 'Disease', (3594, 3597)) ('APL', 'Disease', 'MESH:D015473', (2185, 2188)) ('WT1', 'Gene', (128, 131)) ('RUNX1', 'Gene', '861', (1836, 1841)) ('CD56', 'Gene', '4684', (433, 437)) ('CD34', 'Gene', (389, 393)) ('RUNX1', 'Gene', (1367, 1372)) ('CD33', 'Gene', (411, 415)) ('AML', 'Disease', 'MESH:D015470', (593, 596)) ('RUNX1', 'Gene', '861', (1367, 1372)) ('CD13', 'Gene', '290', (1689, 1693)) ('WT1', 'Gene', '7490', (128, 131)) ('AML 1', 'Gene', (34, 50)) ('inv', 'Gene', (3186, 3189)) ('t(8;21)(q22;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (455, 471)) ('AML', 'Disease', (3632, 3635)) ('RUNX1', 'Gene', (485, 490)) ('RUNX1T1', 'Gene', '862', (86, 93)) ('AML', 'Disease', 'MESH:D015470', (3401, 3404)) ('AML', 'Disease', 'MESH:D015470', (3449, 3452)) ('RUNX1', 'Gene', '861', (485, 490)) ('RUNX1T1', 'Gene', (623, 630)) ('RUNX1T1', 'Gene', (1836, 1843)) ('RUNX1', 'Gene', (2801, 2806)) ('CD33', 'Gene', (1694, 1698)) ('RUNX1', 'Gene', (617, 622)) ('AML', 'Disease', 'MESH:D015470', (3527, 3530)) ('RUNX1', 'Gene', '861', (2801, 2806)) ('AML', 'Disease', 'MESH:D015470', (3534, 3537)) ('RUNX1', 'Gene', '861', (617, 622)) ('AML', 'Disease', (1971, 1974)) ('inv', 'Gene', '89782', (3186, 3189)) 1187 32270433 The inherited forms are rare and mainly include; Fanconi anemia, congenital keratosis, congenital pure red cell aplasia and Schwachman-Diamond syndrome as a result of genetic lesions leading to diminish HSC's ability to repair DNA. ('aplasia', 'Disease', 'MESH:C563261', (112, 119)) ('ability', 'MPA', (209, 216)) ('Fanconi anemia', 'Disease', 'MESH:D005199', (49, 63)) ('diminish', 'NegReg', (194, 202)) ('aplasia', 'Disease', (112, 119)) ('congenital keratosis', 'Disease', (65, 85)) ('genetic lesions', 'Var', (167, 182)) ('anemia', 'Phenotype', 'HP:0001903', (57, 63)) ('Schwachman-Diamond syndrome', 'Disease', (124, 151)) ('Fanconi anemia', 'Phenotype', 'HP:0001994', (49, 63)) ('diminish HSC', 'Phenotype', 'HP:0031851', (194, 206)) ('congenital keratosis', 'Disease', 'MESH:D007642', (65, 85)) ('pure red cell aplasia', 'Phenotype', 'HP:0012410', (98, 119)) ('Schwachman-Diamond syndrome', 'Disease', 'MESH:D029503', (124, 151)) ('Fanconi anemia', 'Disease', (49, 63)) ('HSC', 'Disease', (203, 206)) 1195 32270433 Genetic factors play an important role in the pathogenesis of AA with regard to uniparental disomy in 6p, somatic cell mutations, specific somatic mutation in HLA class I and class II genes, shorter telomerase with telomerase gene mutations and genetic susceptibility. ('uniparental disomy', 'Disease', 'MESH:D024182', (80, 98)) ('mutations', 'Var', (231, 240)) ('AA', 'Phenotype', 'HP:0001903', (62, 64)) ('mutation', 'Var', (147, 155)) ('uniparental disomy', 'Disease', (80, 98)) ('HLA', 'Gene', '3123', (159, 162)) ('HLA', 'Gene', (159, 162)) 1197 32270433 One of these alterations is copy number - the neutral loss of heterozygosity of chromosome arm 6 p (6p CN-LOH), which is the major histocompatibility complex (MHC) region reach in the multiple human leucocyte antigen (HLA) loci. ('HLA', 'Gene', '3123', (218, 221)) ('HLA', 'Gene', (218, 221)) ('alterations', 'Var', (13, 24)) ('loss of heterozygosity', 'NegReg', (54, 76)) ('human', 'Species', '9606', (193, 198)) 1198 32270433 The 6p CN-LOH is more frequent in AA compared to other bone marrow failure syndrome. ('bone marrow failure syndrome', 'Disease', (55, 83)) ('AA', 'Phenotype', 'HP:0001903', (34, 36)) ('bone marrow failure syndrome', 'Disease', 'MESH:D000080983', (55, 83)) ('frequent', 'Reg', (22, 30)) ('marrow failure', 'Phenotype', 'HP:0005528', (60, 74)) ('6p CN-LOH', 'Var', (4, 13)) ('bone marrow failure', 'Phenotype', 'HP:0005528', (55, 74)) 1206 32270433 Depending on the age of patients with acquired AA, different pathomechanisms are predominant, such as HLA mutation in children with a young onset of AA, whereas MDS-association mutation can be identified in the older population. ('children', 'Species', '9606', (118, 126)) ('HLA', 'Gene', '3123', (102, 105)) ('mutation', 'Var', (106, 114)) ('MDS', 'Phenotype', 'HP:0002863', (161, 164)) ('AA', 'Phenotype', 'HP:0001903', (149, 151)) ('HLA', 'Gene', (102, 105)) ('patients', 'Species', '9606', (24, 32)) ('MDS', 'Disease', (161, 164)) ('MDS', 'Disease', 'MESH:D009190', (161, 164)) ('AA', 'Phenotype', 'HP:0001903', (47, 49)) 1210 32270433 It should be emphasised that the presence of HLA risk alleles increases the risk of a more serious course of disease, clonal evolution and complications. ('HLA', 'Gene', (45, 48)) ('HLA', 'Gene', '3123', (45, 48)) ('presence', 'Var', (33, 41)) 1212 32270433 Because telomere shortening leads to cell proliferative arrest and apoptosis, bone marrow failures, including AA, are associated with inherited mutations in telomere repair or protection genes. ('telomere shortening', 'Phenotype', 'HP:0031413', (8, 27)) ('apoptosis', 'CPA', (67, 76)) ('telomere', 'Gene', (157, 165)) ('telomere', 'Var', (8, 16)) ('arrest', 'Disease', (56, 62)) ('bone marrow failures', 'Disease', (78, 98)) ('bone marrow failure', 'Phenotype', 'HP:0005528', (78, 97)) ('AA', 'Phenotype', 'HP:0001903', (110, 112)) ('bone marrow failures', 'Disease', 'MESH:D000080983', (78, 98)) ('marrow failure', 'Phenotype', 'HP:0005528', (83, 97)) ('bone marrow failures', 'Phenotype', 'HP:0005528', (78, 98)) ('associated', 'Reg', (118, 128)) ('mutations', 'Var', (144, 153)) ('arrest', 'Disease', 'MESH:D006323', (56, 62)) 1213 32270433 revealed that short telomeres result in chromosomal instability leading to short telomeres producing end-to-end chromosome fusion, nonreciprocal translocations, and aneuploidy in hematopoietic cells and preceding malignant evolution in human aplastic anemia. ('human', 'Species', '9606', (236, 241)) ('aneuploidy', 'Disease', (165, 175)) ('aplastic anemia', 'Disease', (242, 257)) ('short telomeres', 'Phenotype', 'HP:0031413', (14, 29)) ('anemia', 'Phenotype', 'HP:0001903', (251, 257)) ('producing', 'Reg', (91, 100)) ('chromosomal instability', 'MPA', (40, 63)) ('short telomeres', 'Phenotype', 'HP:0031413', (75, 90)) ('aplastic anemia', 'Phenotype', 'HP:0001915', (242, 257)) ('aneuploidy', 'Disease', 'MESH:D000782', (165, 175)) ('aplastic anemia', 'Disease', 'MESH:D000741', (242, 257)) ('short', 'Var', (75, 80)) ('chromosomal instability', 'Phenotype', 'HP:0040012', (40, 63)) ('end-to-end chromosome fusion', 'CPA', (101, 129)) 1216 32270433 Not only did AA patients have shorter telomere length compared to the health control, but also AA patients with somatic mutations had shorter telomere lengths, compared with patients without mutations. ('telomere length', 'MPA', (38, 53)) ('shorter telomere length', 'Phenotype', 'HP:0031413', (30, 53)) ('shorter telomere length', 'Phenotype', 'HP:0031413', (134, 157)) ('AA', 'Phenotype', 'HP:0001903', (13, 15)) ('shorter telomere lengths', 'Phenotype', 'HP:0031413', (134, 158)) ('telomere lengths', 'MPA', (142, 158)) ('patients', 'Species', '9606', (16, 24)) ('patients', 'Species', '9606', (98, 106)) ('patients', 'Species', '9606', (174, 182)) ('mutations', 'Var', (120, 129)) ('AA', 'Phenotype', 'HP:0001903', (95, 97)) ('shorter', 'NegReg', (30, 37)) ('shorter', 'NegReg', (134, 141)) 1217 32270433 Mutations in TERT, TERC and DKC1 are associated with critical telomere length loss of HSPCs in AA patients. ('DKC1', 'Gene', '1736', (28, 32)) ('critical telomere length', 'MPA', (53, 77)) ('TERT', 'Gene', (13, 17)) ('TERT', 'Gene', '7015', (13, 17)) ('loss of HSPCs', 'Disease', 'MESH:D014786', (78, 91)) ('TERC', 'Gene', '7012', (19, 23)) ('patients', 'Species', '9606', (98, 106)) ('Mutations', 'Var', (0, 9)) ('associated', 'Reg', (37, 47)) ('DKC1', 'Gene', (28, 32)) ('AA', 'Phenotype', 'HP:0001903', (95, 97)) ('loss of HSPCs', 'Disease', (78, 91)) ('TERC', 'Gene', (19, 23)) 1220 32270433 Although, heterozygous variants RTEL1 (regulator of telomere elongation helicase 1) were identified in bone marrow failure, which included AA (1-2%), they were likely benign or of uncertain significance. ('bone marrow failure', 'Phenotype', 'HP:0005528', (103, 122)) ('RTEL1', 'Gene', '51750', (32, 37)) ('RTEL1', 'Gene', (32, 37)) ('bone marrow failure', 'Disease', (103, 122)) ('regulator of telomere elongation helicase 1', 'Gene', '51750', (39, 82)) ('bone marrow failure', 'Disease', 'MESH:D000080983', (103, 122)) ('AA', 'Phenotype', 'HP:0001903', (139, 141)) ('marrow failure', 'Phenotype', 'HP:0005528', (108, 122)) ('variants', 'Var', (23, 31)) ('regulator of telomere elongation helicase 1', 'Gene', (39, 82)) 1222 32270433 showed that accelerated telomere attrition in the setting of a decreased hematopoietic stem and progenitor cells (HSPC) pool is characteristic of early myeloid oncogenesis, specifically for chromosome 7 loss, particularly in MDS/AML after treatment of AA, and provides a possible mechanism for the development of aneuploidy. ('accelerated', 'PosReg', (12, 23)) ('AML', 'Disease', 'MESH:D015470', (229, 232)) ('early myeloid oncogenesis', 'Disease', (146, 171)) ('aneuploidy', 'Disease', (313, 323)) ('chromosome', 'Var', (190, 200)) ('decreased', 'NegReg', (63, 72)) ('AML', 'Disease', (229, 232)) ('aneuploidy', 'Disease', 'MESH:D000782', (313, 323)) ('MDS', 'Disease', (225, 228)) ('MDS', 'Disease', 'MESH:D009190', (225, 228)) ('AML', 'Phenotype', 'HP:0004808', (229, 232)) ('MDS', 'Phenotype', 'HP:0002863', (225, 228)) ('telomere attrition', 'MPA', (24, 42)) ('AA', 'Phenotype', 'HP:0001903', (252, 254)) ('loss', 'NegReg', (203, 207)) 1224 32270433 The identification of the AA secondary to telomerase variants/mutations is very important due to, presumably, a less likely response to immunosuppressive therapy and the patient may be a candidate to upfront HLA-matched sibling or unrelated donor allogeneic bone marrow transplantation. ('variants/mutations', 'Var', (53, 71)) ('HLA', 'Gene', (208, 211)) ('telomerase', 'Gene', (42, 52)) ('patient', 'Species', '9606', (170, 177)) ('AA', 'Phenotype', 'HP:0001903', (26, 28)) ('HLA', 'Gene', '3123', (208, 211)) 1229 32270433 On the contrary, trisomy 8, commonly observed in other myeloid malignancies, with an occurrence of 1.3-6.7%, is associated with a favorable response to immunosuppressive therapy and lower risk of evolution to MDS or AML. ('myeloid malignancies', 'Disease', (55, 75)) ('AML', 'Disease', 'MESH:D015470', (216, 219)) ('MDS', 'Disease', (209, 212)) ('MDS', 'Disease', 'MESH:D009190', (209, 212)) ('MDS', 'Phenotype', 'HP:0002863', (209, 212)) ('myeloid malignancies', 'Disease', 'MESH:D009369', (55, 75)) ('AML', 'Disease', (216, 219)) ('trisomy 8', 'Var', (17, 26)) ('AML', 'Phenotype', 'HP:0004808', (216, 219)) 1230 32270433 Other cytogenetic abnormalities are less commonly detected, some of these are shared with myeloid malignancies, such as del(5q), whereas others, including trisomy 6, trisomy 15 are rarely seen in AML/MDS and their clinical relevance should be established. ('myeloid malignancies', 'Disease', (90, 110)) ('trisomy 15', 'Var', (166, 176)) ('AML', 'Phenotype', 'HP:0004808', (196, 199)) ('MDS', 'Disease', (200, 203)) ('MDS', 'Disease', 'MESH:D009190', (200, 203)) ('AML', 'Disease', 'MESH:D015470', (196, 199)) ('MDS', 'Phenotype', 'HP:0002863', (200, 203)) ('trisomy 6', 'Var', (155, 164)) ('AML', 'Disease', (196, 199)) ('myeloid malignancies', 'Disease', 'MESH:D009369', (90, 110)) 1234 32270433 Recent advances in genome analysis, including single nucleotide polymorphism (SNP-As) arrays, targeted new generation sequencing (NGS), whole exom-sequencing (WES) and NGS-based HLA typing, have revealed somatic mutations in patients with AA which confirm the association of clonal haematopoiesis in the pathogenesis of AA. ('patients', 'Species', '9606', (225, 233)) ('AA', 'Phenotype', 'HP:0001903', (320, 322)) ('HLA', 'Gene', '3123', (178, 181)) ('revealed', 'Reg', (195, 203)) ('HLA', 'Gene', (178, 181)) ('mutations', 'Var', (212, 221)) ('AA', 'Phenotype', 'HP:0001903', (239, 241)) 1239 32270433 reported that although some mutations are typical for MDS, they may reflect clonal hematopoiesis and may have potential to evolve from AA to secondary MDS but not in all cases. ('MDS', 'Disease', 'MESH:D009190', (151, 154)) ('hematopoiesis', 'Disease', (83, 96)) ('AA', 'Phenotype', 'HP:0001903', (135, 137)) ('MDS', 'Disease', (54, 57)) ('MDS', 'Disease', 'MESH:D009190', (54, 57)) ('hematopoiesis', 'Disease', 'MESH:C536227', (83, 96)) ('MDS', 'Phenotype', 'HP:0002863', (54, 57)) ('MDS', 'Phenotype', 'HP:0002863', (151, 154)) ('mutations', 'Var', (28, 37)) ('MDS', 'Disease', (151, 154)) ('reflect', 'Reg', (68, 75)) 1240 32270433 Some these mutations ale likely to predict secondary MDS. ('mutations', 'Var', (11, 20)) ('MDS', 'Disease', (53, 56)) ('MDS', 'Disease', 'MESH:D009190', (53, 56)) ('MDS', 'Phenotype', 'HP:0002863', (53, 56)) ('predict', 'Reg', (35, 42)) 1242 32270433 This group of mutations has been found to be at risk of inferior prognosis with a higher risk of evolution to MDS and AML. ('MDS', 'Disease', (110, 113)) ('MDS', 'Disease', 'MESH:D009190', (110, 113)) ('MDS', 'Phenotype', 'HP:0002863', (110, 113)) ('AML', 'Phenotype', 'HP:0004808', (118, 121)) ('AML', 'Disease', (118, 121)) ('mutations', 'Var', (14, 23)) ('AML', 'Disease', 'MESH:D015470', (118, 121)) 1243 32270433 DNMT3A mutation is involved in DNA methylation, ASXL1 belongs to the polycomb family influencing chromatin structure and function. ('involved', 'Reg', (19, 27)) ('ASXL1', 'Gene', '171023', (48, 53)) ('DNMT3A', 'Gene', (0, 6)) ('mutation', 'Var', (7, 15)) ('function', 'MPA', (121, 129)) ('DNMT3A', 'Gene', '1788', (0, 6)) ('chromatin', 'MPA', (97, 106)) ('ASXL1', 'Gene', (48, 53)) 1244 32270433 The second group of patients is younger and it is characterised by the most commonly mutated genes in AA include PIGA, BCOR, BCORL1, which are known to be relatively benign/indolent mutations. ('BCOR', 'Gene', (125, 129)) ('BCORL1', 'Gene', '63035', (125, 131)) ('AA', 'Phenotype', 'HP:0001903', (102, 104)) ('BCOR', 'Gene', '54880', (125, 129)) ('mutated', 'Var', (85, 92)) ('BCOR', 'Gene', (119, 123)) ('patients', 'Species', '9606', (20, 28)) ('PIGA', 'Gene', '5277', (113, 117)) ('BCORL1', 'Gene', (125, 131)) ('BCOR', 'Gene', '54880', (119, 123)) ('PIGA', 'Gene', (113, 117)) 1245 32270433 The PIGA gene plays a role in biosynthesis of glycosyl-phosphatidyl-inositol (GPI), the mutation inactivates its functions. ('mutation', 'Var', (88, 96)) ('PIGA', 'Gene', '5277', (4, 8)) ('inactivates', 'NegReg', (97, 108)) ('PIGA', 'Gene', (4, 8)) ('biosynthesis', 'MPA', (30, 42)) ('glycosyl-phosphatidyl-inositol', 'Chemical', 'MESH:D017261', (46, 76)) ('GPI', 'Chemical', 'MESH:D017261', (78, 81)) ('functions', 'MPA', (113, 122)) 1247 32270433 The second group of mutations is of great importance for clinicians because PIGA and BCOR may predict a favourable response to immunosuppressive therapy with improved progression-free survival and overall survival. ('improved', 'PosReg', (158, 166)) ('PIGA', 'Gene', '5277', (76, 80)) ('PIGA', 'Gene', (76, 80)) ('BCOR', 'Gene', (85, 89)) ('mutations', 'Var', (20, 29)) ('BCOR', 'Gene', '54880', (85, 89)) 1249 32270433 Although some mutations may drive clonal expression in other disorders such as TET and JAK2, the impact on clonal evolution in patients with AA is less prominent. ('AA', 'Phenotype', 'HP:0001903', (141, 143)) ('drive', 'Reg', (28, 33)) ('patients', 'Species', '9606', (127, 135)) ('JAK2', 'Gene', '3717', (87, 91)) ('clonal', 'MPA', (34, 40)) ('TET', 'Disease', (79, 82)) ('JAK2', 'Gene', (87, 91)) ('mutations', 'Var', (14, 23)) 1251 32270433 One of the most frequently reported somatic mutations, found in nearly half of AA patients, is PIGA, which results in clonal populations of cells lacking cell surface proteins linked to a glycosylphosphatidylinositol (GPI) anchor due to somatic loss-of-function mutations, one of a number of drivers of clonal evolution in AA and paroxysmal nocturnal hemoglobinuria (PNH). ('paroxysmal nocturnal hemoglobinuria', 'Disease', 'MESH:D006457', (330, 365)) ('paroxysmal nocturnal hemoglobinuria', 'Phenotype', 'HP:0004818', (330, 365)) ('PIGA', 'Gene', (95, 99)) ('PNH', 'Disease', (367, 370)) ('paroxysmal nocturnal hemoglobinuria', 'Disease', (330, 365)) ('glycosylphosphatidylinositol', 'Chemical', 'MESH:D017261', (188, 216)) ('patients', 'Species', '9606', (82, 90)) ('hemoglobinuria', 'Phenotype', 'HP:0003641', (351, 365)) ('PNH', 'Phenotype', 'HP:0004818', (367, 370)) ('AA', 'Phenotype', 'HP:0001903', (79, 81)) ('PNH', 'Disease', 'None', (367, 370)) ('lacking', 'NegReg', (146, 153)) ('mutations', 'Var', (44, 53)) ('AA', 'Phenotype', 'HP:0001903', (323, 325)) ('GPI', 'Chemical', 'MESH:D017261', (218, 221)) ('loss-of-function', 'NegReg', (245, 261)) ('PIGA', 'Gene', '5277', (95, 99)) ('mutations', 'Var', (262, 271)) 1255 32270433 Although at least one mutation was identified in 19% of patients with AA at the time of diagnosis, independently of the severity of the AA, these mutations included DNMT3A, PIGA, SRSF2 and CEBPA and there were no differences in the average number of mutations. ('patients', 'Species', '9606', (56, 64)) ('mutations', 'Var', (146, 155)) ('SRSF2', 'Gene', (179, 184)) ('AA', 'Phenotype', 'HP:0001903', (70, 72)) ('DNMT3A', 'Gene', (165, 171)) ('DNMT3A', 'Gene', '1788', (165, 171)) ('PIGA', 'Gene', '5277', (173, 177)) ('CEBPA', 'Gene', (189, 194)) ('PIGA', 'Gene', (173, 177)) ('CEBPA', 'Gene', '1050', (189, 194)) ('SRSF2', 'Gene', '6427', (179, 184)) ('AA', 'Phenotype', 'HP:0001903', (136, 138)) 1262 32270433 All these DAMPs lead to activation of the Nlrp3 inflammasome, that drives process of inflammaging in the all tissues including bone marrow. ('activation', 'PosReg', (24, 34)) ('Nlrp3', 'Gene', (42, 47)) ('DAMPs', 'Var', (10, 15)) ('Nlrp3', 'Gene', '114548', (42, 47)) 1267 32270433 The next-generation sequencing and array-based karyotyping obtained from 439 patients with aplastic anemia showed clonal hematopoiesis of stem cell with somatic mutations in 47% of patients with AA, mainly acquired mutations.The incidence of variant allele frequency in some genes has been reported to be up to 31% for ASXL1. ('AA', 'Phenotype', 'HP:0001903', (195, 197)) ('variant', 'Var', (242, 249)) ('hematopoiesis', 'Disease', (121, 134)) ('aplastic anemia', 'Disease', (91, 106)) ('anemia', 'Phenotype', 'HP:0001903', (100, 106)) ('ASXL1', 'Gene', '171023', (319, 324)) ('patients', 'Species', '9606', (77, 85)) ('aplastic anemia', 'Disease', 'MESH:D000741', (91, 106)) ('patients', 'Species', '9606', (181, 189)) ('aplastic anemia', 'Phenotype', 'HP:0001915', (91, 106)) ('hematopoiesis', 'Disease', 'MESH:C536227', (121, 134)) ('ASXL1', 'Gene', (319, 324)) 1271 32270433 Another study reports a significant negative correlation between the expression of miR-144-3p and ten-eleven translocation 2 (TET2) which cause reduced osteogenic capacity of bone marrow mesenchymal stem cells taken from patients with AA, this may be a therapeutic strategy in future. ('reduced', 'NegReg', (144, 151)) ('patients', 'Species', '9606', (221, 229)) ('osteogenic capacity', 'CPA', (152, 171)) ('miR-144-3p', 'Var', (83, 93)) ('TET2', 'Gene', (126, 130)) ('AA', 'Phenotype', 'HP:0001903', (235, 237)) 1272 32270433 Recent advances in understanding the molecular pathogenesis of AA, which tends to be a clonal disorder of hematopoietic stem cells and their progenitors with somatic cell mutations, including HLA-mediated immunity mutations, defective telomerase function, inherited and acquired cytogenetic aberrations, may allow, in some situations, for the prediction of the results of immunosuppressive therapy and a forecasting of an unfavourable outcome. ('HLA', 'Gene', '3123', (192, 195)) ('mutations', 'Var', (214, 223)) ('HLA', 'Gene', (192, 195)) ('AA', 'Phenotype', 'HP:0001903', (63, 65)) ('defective', 'Var', (225, 234)) ('allow', 'Reg', (308, 313)) 1275 31483066 Residual apoptotic activity of a tumorigenic p53 mutant improves cancer therapy responses Engineered p53 mutant mice are valuable tools for delineating p53 functions in tumor suppression and cancer therapy. ('apoptotic activity', 'MPA', (9, 27)) ('mutant', 'Var', (105, 111)) ('cancer', 'Disease', (65, 71)) ('p53', 'Gene', '22059', (45, 48)) ('cancer', 'Disease', (191, 197)) ('mutant', 'Var', (49, 55)) ('p53', 'Gene', (152, 155)) ('cancer', 'Phenotype', 'HP:0002664', (65, 71)) ('p53', 'Gene', '22059', (101, 104)) ('tumor', 'Disease', (169, 174)) ('tumor', 'Disease', (33, 38)) ('cancer', 'Phenotype', 'HP:0002664', (191, 197)) ('tumor', 'Disease', 'MESH:D009369', (169, 174)) ('tumor', 'Disease', 'MESH:D009369', (33, 38)) ('cancer', 'Disease', 'MESH:D009369', (65, 71)) ('mice', 'Species', '10090', (112, 116)) ('p53', 'Gene', (45, 48)) ('cancer', 'Disease', 'MESH:D009369', (191, 197)) ('p53', 'Gene', '22059', (152, 155)) ('tumor', 'Phenotype', 'HP:0002664', (169, 174)) ('tumor', 'Phenotype', 'HP:0002664', (33, 38)) ('p53', 'Gene', (101, 104)) ('improves', 'PosReg', (56, 64)) 1276 31483066 Here, we have introduced the R178E mutation into the Trp53 gene of mice to specifically ablate the cooperative nature of p53 DNA binding. ('cooperative', 'MPA', (99, 110)) ('R178E', 'Var', (29, 34)) ('p53', 'Protein', (121, 124)) ('ablate', 'NegReg', (88, 94)) ('Trp53', 'Gene', (53, 58)) ('R178E', 'Mutation', 'p.R178E', (29, 34)) ('mice', 'Species', '10090', (67, 71)) 1277 31483066 Trp53 R178E mice show no detectable target gene regulation and, at first sight, are largely indistinguishable from Trp53 -/- mice. ('R178E', 'Mutation', 'p.R178E', (6, 11)) ('R178E', 'Var', (6, 11)) ('mice', 'Species', '10090', (125, 129)) ('mice', 'Species', '10090', (12, 16)) ('Trp53', 'Gene', (0, 5)) 1278 31483066 Surprisingly, stabilization of p53R178E in Mdm2 -/- mice nevertheless triggers extensive apoptosis, indicative of residual wild-type activities. ('p53R178E', 'Var', (31, 39)) ('triggers', 'Reg', (70, 78)) ('apoptosis', 'CPA', (89, 98)) ('mice', 'Species', '10090', (52, 56)) 1279 31483066 Although this apoptotic activity suffices to trigger lethality of Trp53 R178E ;Mdm2 -/- embryos, it proves insufficient for suppression of spontaneous and oncogene-driven tumorigenesis. ('R178E ', 'Var', (72, 78)) ('Trp53', 'Gene', (66, 71)) ('tumor', 'Disease', 'MESH:D009369', (171, 176)) ('R178E', 'Mutation', 'p.R178E', (72, 77)) ('tumor', 'Phenotype', 'HP:0002664', (171, 176)) ('tumor', 'Disease', (171, 176)) 1280 31483066 Trp53 R178E mice develop tumors indistinguishably from Trp53 -/- mice and tumors retain and even stabilize the p53R178E protein, further attesting to the lack of significant tumor suppressor activity. ('tumor', 'Phenotype', 'HP:0002664', (25, 30)) ('tumors', 'Disease', (25, 31)) ('p53R178E', 'Var', (111, 119)) ('R178E', 'Var', (6, 11)) ('tumor', 'Disease', (74, 79)) ('tumor', 'Phenotype', 'HP:0002664', (174, 179)) ('tumors', 'Disease', 'MESH:D009369', (74, 80)) ('protein', 'Protein', (120, 127)) ('tumor', 'Disease', 'MESH:D009369', (74, 79)) ('tumors', 'Disease', 'MESH:D009369', (25, 31)) ('tumors', 'Disease', (74, 80)) ('R178E', 'Mutation', 'p.R178E', (114, 119)) ('mice', 'Species', '10090', (12, 16)) ('tumor', 'Disease', (25, 30)) ('mice', 'Species', '10090', (65, 69)) ('tumor', 'Phenotype', 'HP:0002664', (74, 79)) ('tumor', 'Disease', 'MESH:D009369', (25, 30)) ('tumors', 'Phenotype', 'HP:0002664', (74, 80)) ('tumor', 'Disease', (174, 179)) ('tumors', 'Phenotype', 'HP:0002664', (25, 31)) ('R178E', 'Mutation', 'p.R178E', (6, 11)) ('tumor', 'Disease', 'MESH:D009369', (174, 179)) ('Trp53', 'Gene', (0, 5)) 1281 31483066 However, Trp53 R178E tumors exhibit remarkably better chemotherapy responses than Trp53 -/- ones, resulting in enhanced eradication of p53-mutated tumor cells. ('tumor', 'Phenotype', 'HP:0002664', (147, 152)) ('tumor', 'Disease', (147, 152)) ('better', 'PosReg', (47, 53)) ('R178E', 'Var', (15, 20)) ('tumor', 'Disease', 'MESH:D009369', (21, 26)) ('tumor', 'Disease', (21, 26)) ('tumors', 'Disease', (21, 27)) ('eradication', 'MPA', (120, 131)) ('Trp53', 'Gene', (9, 14)) ('chemotherapy responses', 'CPA', (54, 76)) ('tumors', 'Disease', 'MESH:D009369', (21, 27)) ('enhanced', 'PosReg', (111, 119)) ('tumors', 'Phenotype', 'HP:0002664', (21, 27)) ('tumor', 'Disease', 'MESH:D009369', (147, 152)) ('R178E', 'Mutation', 'p.R178E', (15, 20)) ('tumor', 'Phenotype', 'HP:0002664', (21, 26)) 1282 31483066 Together, this provides genetic proof-of-principle evidence that a p53 mutant can be highly tumorigenic and yet retain apoptotic activity which provides a survival benefit in the context of cancer therapy. ('cancer', 'Disease', (190, 196)) ('cancer', 'Disease', 'MESH:D009369', (190, 196)) ('tumor', 'Disease', 'MESH:D009369', (92, 97)) ('p53', 'Gene', (67, 70)) ('tumor', 'Phenotype', 'HP:0002664', (92, 97)) ('mutant', 'Var', (71, 77)) ('apoptotic activity', 'CPA', (119, 137)) ('cancer', 'Phenotype', 'HP:0002664', (190, 196)) ('tumor', 'Disease', (92, 97)) ('survival benefit', 'CPA', (155, 171)) 1284 31483066 The presence of TP53 mutations correlates in many cancer types with enhanced metastasis and aggressiveness, reduced responses to chemotherapeutic drugs, and, thus, a poor prognosis (Robles et al, 2016; Sabapathy & Lane, 2018). ('aggressiveness', 'Disease', (92, 106)) ('enhanced', 'PosReg', (68, 76)) ('cancer', 'Disease', (50, 56)) ('cancer', 'Disease', 'MESH:D009369', (50, 56)) ('aggressiveness', 'Phenotype', 'HP:0000718', (92, 106)) ('reduced', 'NegReg', (108, 115)) ('metastasis', 'CPA', (77, 87)) ('aggressiveness', 'Disease', 'MESH:D001523', (92, 106)) ('cancer', 'Phenotype', 'HP:0002664', (50, 56)) ('TP53', 'Gene', (16, 20)) ('responses to chemotherapeutic drugs', 'MPA', (116, 151)) ('mutations', 'Var', (21, 30)) 1285 31483066 More than 85% of all amino acid positions were found to be mutated in cancer patients, generating a "rainbow" of > 2,000 distinct missense variants (Sabapathy & Lane, 2018). ('cancer', 'Phenotype', 'HP:0002664', (70, 76)) ('missense variants', 'Var', (130, 147)) ('patients', 'Species', '9606', (77, 85)) ('cancer', 'Disease', (70, 76)) ('cancer', 'Disease', 'MESH:D009369', (70, 76)) 1286 31483066 Mutations cluster in the central DNA binding domain (DBD), suggesting that tumorigenesis selects against p53's DNA binding function (Muller & Vousden, 2014; Stiewe & Haran, 2018). ('tumor', 'Phenotype', 'HP:0002664', (75, 80)) ('Mutations', 'Var', (0, 9)) ('tumor', 'Disease', (75, 80)) ('tumor', 'Disease', 'MESH:D009369', (75, 80)) 1287 31483066 Furthermore, missense mutants are unstable in normal unstressed cells, but become constitutively stabilized in tumors by Hsp90 which protects mutant p53 from degradation by Mdm2 and CHIP (Terzian et al, 2008; Alexandrova et al, 2015). ('missense', 'Var', (13, 21)) ('p53', 'Gene', (149, 152)) ('tumor', 'Phenotype', 'HP:0002664', (111, 116)) ('tumors', 'Disease', (111, 117)) ('mutant', 'Var', (142, 148)) ('tumors', 'Disease', 'MESH:D009369', (111, 117)) ('tumors', 'Phenotype', 'HP:0002664', (111, 117)) ('degradation', 'MPA', (158, 169)) ('CHIP', 'Chemical', 'MESH:C048067', (182, 186)) 1288 31483066 The preferential selection of missense mutants together with their excessive stabilization therefore points at additional mechanisms that promote tumor development beyond a mere loss of DNA binding activity: Missense mutants exhibit dominant-negative effects on remaining wild-type p53 and display neomorphic properties that:like an oncogene:actively drive tumor development to a metastatic and drug-resistant state (Freed-Pastor & Prives, 2012; Muller & Vousden, 2014; Kim & Lozano, 2018; Stiewe & Haran, 2018). ('Missense mutants', 'Var', (208, 224)) ('missense', 'Var', (30, 38)) ('tumor', 'Disease', (357, 362)) ('tumor', 'Phenotype', 'HP:0002664', (146, 151)) ('drug-resistant state', 'Phenotype', 'HP:0020174', (395, 415)) ('tumor', 'Disease', (146, 151)) ('promote', 'PosReg', (138, 145)) ('tumor', 'Disease', 'MESH:D009369', (357, 362)) ('drive', 'PosReg', (351, 356)) ('tumor', 'Phenotype', 'HP:0002664', (357, 362)) ('tumor', 'Disease', 'MESH:D009369', (146, 151)) 1289 31483066 Missense mutations therefore enhance tumor development and progression in three ways: the loss of wild-type-like DNA binding activity (loss of function, LOF), dominant-negative effects on wild-type p53, and the gain of new tumor-promoting oncogenic properties (gain of function, GOF) (Stiewe & Haran, 2018). ('enhance', 'PosReg', (29, 36)) ('tumor', 'Phenotype', 'HP:0002664', (223, 228)) ('tumor', 'Disease', (223, 228)) ('tumor', 'Disease', 'MESH:D009369', (37, 42)) ('loss', 'NegReg', (90, 94)) ('activity', 'MPA', (125, 133)) ('progression', 'CPA', (59, 70)) ('tumor', 'Phenotype', 'HP:0002664', (37, 42)) ('tumor', 'Disease', (37, 42)) ('tumor', 'Disease', 'MESH:D009369', (223, 228)) ('gain', 'PosReg', (211, 215)) ('DNA', 'Protein', (113, 116)) ('Missense mutations', 'Var', (0, 18)) 1290 31483066 Despite being no mutational hot-spots, cooperativity mutations at residues E180 and R181 are estimated to account for 34,000 cancer cases per year (Leroy et al, 2014). ('cooperativity', 'Var', (39, 52)) ('cancer', 'Phenotype', 'HP:0002664', (125, 131)) ('R181', 'Var', (84, 88)) ('account', 'Reg', (106, 113)) ('cancer', 'Disease', (125, 131)) ('cancer', 'Disease', 'MESH:D009369', (125, 131)) 1291 31483066 Of all mutations of the double salt bridge, the R181E mutant disrupts formation of the intermolecular salt bridge most effectively (Schlereth et al, 2010a). ('R181E', 'Var', (48, 53)) ('R181E', 'Mutation', 'p.R181E', (48, 53)) ('disrupts', 'NegReg', (61, 69)) ('formation of the intermolecular salt bridge', 'MPA', (70, 113)) 1292 31483066 Although R181E retains a native fold, it is entirely DNA binding deficient as assessed by electrophoretic mobility shift assays and genome-wide chromatin immunoprecipitation analysis (Dehner et al, 2005; Schlereth et al, 2010a, 2013). ('R181E', 'Var', (9, 14)) ('deficient', 'NegReg', (65, 74)) ('native fold', 'MPA', (25, 36)) ('DNA binding', 'Interaction', (53, 64)) ('R181E', 'Mutation', 'p.R181E', (9, 14)) 1294 31483066 Mutations engineered into the mouse Trp53 gene locus are valuable tools to delineate in vivo tumor suppressor functions in tumorigenesis and cancer therapy (Bieging et al, 2014; Mello & Attardi, 2018). ('tumor', 'Disease', (123, 128)) ('tumor', 'Phenotype', 'HP:0002664', (93, 98)) ('mouse', 'Species', '10090', (30, 35)) ('cancer', 'Phenotype', 'HP:0002664', (141, 147)) ('tumor', 'Disease', (93, 98)) ('tumor', 'Phenotype', 'HP:0002664', (123, 128)) ('Mutations', 'Var', (0, 9)) ('tumor', 'Disease', 'MESH:D009369', (123, 128)) ('Trp53', 'Gene', (36, 41)) ('cancer', 'Disease', 'MESH:D009369', (141, 147)) ('cancer', 'Disease', (141, 147)) ('tumor', 'Disease', 'MESH:D009369', (93, 98)) 1295 31483066 Besides mutations derived from cancer patients, especially non-naturally occurring mutations of post-translational modification sites (Sluss et al, 2004; Slee et al, 2010; Li et al, 2012) or functional domains (Toledo et al, 2006; Brady et al, 2011; Hamard et al, 2013; Simeonova et al, 2013) have yielded substantial mechanistic insight into the pathways required for tumor suppression. ('mutations', 'Var', (83, 92)) ('tumor', 'Disease', 'MESH:D009369', (369, 374)) ('cancer', 'Phenotype', 'HP:0002664', (31, 37)) ('tumor', 'Phenotype', 'HP:0002664', (369, 374)) ('mutations', 'Var', (8, 17)) ('tumor', 'Disease', (369, 374)) ('cancer', 'Disease', 'MESH:D009369', (31, 37)) ('patients', 'Species', '9606', (38, 46)) ('cancer', 'Disease', (31, 37)) ('functional', 'MPA', (191, 201)) 1296 31483066 To explore the relevance of DNA binding cooperativity for p53's anti-tumor activities, we therefore generated the "EE" mouse carrying the human R181E-equivalent R178E mutation at the endogenous Trp53 gene locus. ('tumor', 'Phenotype', 'HP:0002664', (69, 74)) ('R178E', 'Mutation', 'p.R178E', (161, 166)) ('R181E-equivalent', 'Var', (144, 160)) ('mouse', 'Species', '10090', (119, 124)) ('R181E', 'Mutation', 'p.R181E', (144, 149)) ('tumor', 'Disease', (69, 74)) ('human', 'Species', '9606', (138, 143)) ('R178E', 'Var', (161, 166)) ('Trp53', 'Gene', (194, 199)) ('tumor', 'Disease', 'MESH:D009369', (69, 74)) 1298 31483066 Surprisingly, the EE mutation:different from the p53-knock-out:does not rescue the embryonic lethality of the Mdm2 knock-out and triggers massive apoptotic cell death providing support for residual cytotoxic activities upon constitutive stabilization. ('apoptotic cell death', 'CPA', (146, 166)) ('triggers', 'Reg', (129, 137)) ('mutation', 'Var', (21, 29)) ('embryonic lethality', 'Disease', 'MESH:D020964', (83, 102)) ('Mdm2', 'Gene', (110, 114)) ('embryonic lethality', 'Disease', (83, 102)) 1300 31483066 Similar results were obtained with the human R181L cooperativity mutant, which has been recurrently identified in cancer patients. ('cancer', 'Disease', 'MESH:D009369', (114, 120)) ('cancer', 'Disease', (114, 120)) ('cancer', 'Phenotype', 'HP:0002664', (114, 120)) ('R181L', 'Var', (45, 50)) ('human', 'Species', '9606', (39, 44)) ('R181L', 'Mutation', 'p.R181L', (45, 50)) ('patients', 'Species', '9606', (121, 129)) 1301 31483066 Together, these findings highlight that mutant p53, in principle, can retain residual apoptotic activities that are insufficient to prevent tumorigenesis and not efficiently counter-selected during tumor evolution. ('tumor', 'Disease', 'MESH:D009369', (198, 203)) ('tumor', 'Phenotype', 'HP:0002664', (198, 203)) ('tumor', 'Disease', 'MESH:D009369', (140, 145)) ('apoptotic', 'CPA', (86, 95)) ('tumor', 'Phenotype', 'HP:0002664', (140, 145)) ('mutant', 'Var', (40, 46)) ('p53', 'Gene', (47, 50)) ('tumor', 'Disease', (198, 203)) ('tumor', 'Disease', (140, 145)) 1302 31483066 Stabilization of such a p53 mutant in combination with chemotherapy is capable to trigger mutant p53-mediated cytotoxicity resulting in improved anti-cancer responses and increased survival. ('mutant', 'Var', (28, 34)) ('p53', 'Gene', (24, 27)) ('cytotoxicity', 'Disease', 'MESH:D064420', (110, 122)) ('p53-mediated', 'Protein', (97, 109)) ('mutant', 'Var', (90, 96)) ('cancer', 'Disease', 'MESH:D009369', (150, 156)) ('survival', 'CPA', (181, 189)) ('cancer', 'Disease', (150, 156)) ('cytotoxicity', 'Disease', (110, 122)) ('increased', 'PosReg', (171, 180)) ('cancer', 'Phenotype', 'HP:0002664', (150, 156)) ('improved', 'PosReg', (136, 144)) 1303 31483066 We previously showed that the DNA binding cooperativity mutant p53R181E (EE) fails to bind p53 response elements in vitro and when exogenously expressed in p53-null cells (Schlereth et al, 2010a, 2013). ('bind', 'Interaction', (86, 90)) ('fails', 'NegReg', (77, 82)) ('R181E', 'Mutation', 'p.R181E', (66, 71)) ('p53R181E', 'Var', (63, 71)) 1304 31483066 To address how ablation of DNA binding cooperativity affects p53 functions in vivo, we generated a conditional knock-in mouse, carrying the R178E (EE) mutation in exon 5 of the endogenous mouse Trp53 gene locus (Fig EV1A-D). ('mouse', 'Species', '10090', (188, 193)) ('R178E', 'Mutation', 'p.R178E', (140, 145)) ('R178E', 'Var', (140, 145)) ('Trp53', 'Gene', (194, 199)) ('mouse', 'Species', '10090', (120, 125)) 1305 31483066 DNA binding deficiency of the EE mutation in the context of the mouse p53 protein was confirmed by electrophoretic mobility shift assays using nuclear extracts of homozygous p53EE/EE mouse embryonic fibroblasts (MEFs) and a high-affinity, consensus-like p53 response element (Fig EV1E). ('mutation', 'Var', (33, 41)) ('mouse', 'Species', '10090', (183, 188)) ('MEF', 'Gene', (212, 215)) ('MEF', 'Gene', '56501', (212, 215)) ('mouse', 'Species', '10090', (64, 69)) 1308 31483066 Only 3 peaks were identified in Nutlin-treated p53EE/EE MEFs that were, however, also called in p53-/- MEFs and therefore considered non-specific (Fig EV1F). ('MEF', 'Gene', '56501', (56, 59)) ('MEF', 'Gene', (103, 106)) ('MEF', 'Gene', (56, 59)) ('Nutlin', 'Chemical', 'MESH:C482204', (32, 38)) ('p53EE/EE', 'Var', (47, 55)) ('MEF', 'Gene', '56501', (103, 106)) 1309 31483066 Thus, the p53 binding pattern observed in p53EE/EE MEFs was indistinguishable from p53-/- MEFs, irrespective of Nutlin treatment, and therefore validated the p53EE mutant expressed from the endogenous Trp53 gene locus to be DNA binding deficient in cells. ('Nutlin', 'Chemical', 'MESH:C482204', (112, 118)) ('p53EE/EE', 'Var', (42, 50)) ('MEF', 'Gene', '56501', (90, 93)) ('binding', 'Interaction', (14, 21)) ('p53EE', 'Var', (158, 163)) ('MEF', 'Gene', (51, 54)) ('p53', 'Protein', (10, 13)) ('MEF', 'Gene', (90, 93)) ('MEF', 'Gene', '56501', (51, 54)) 1310 31483066 When global gene expression was profiled by RNA-seq, Nutlin exerted a significantly stronger effect on global gene expression in p53+/+ versus either p53EE/EE or p53-/- MEFs, while Nutlin effects on p53EE/EE and p53-/- cells showed no significant difference (Fig EV1H). ('stronger effect', 'PosReg', (84, 99)) ('Nutlin', 'Chemical', 'MESH:C482204', (53, 59)) ('Nutlin', 'Chemical', 'MESH:C482204', (181, 187)) ('p53+/+', 'Var', (129, 135)) ('global gene expression', 'MPA', (103, 125)) ('MEF', 'Gene', (169, 172)) ('MEF', 'Gene', '56501', (169, 172)) 1311 31483066 Furthermore, we observed in p53+/+ but not in p53EE/EE or p53-null MEFs a strong Nutlin-inducible expression of a p53 signature including both bona fide p53 pathway genes (MSigDB hallmarks_P53_pathway) and non-canonical targets previously identified to be critical mediators of tumor suppression (Figs 1C and EV1I) (Brady et al, 2011). ('Nutlin', 'Chemical', 'MESH:C482204', (81, 87)) ('tumor', 'Phenotype', 'HP:0002664', (278, 283)) ('p53', 'Gene', (114, 117)) ('tumor', 'Disease', (278, 283)) ('MEF', 'Gene', '56501', (67, 70)) ('MSigDB hallmarks', 'Disease', 'None', (172, 188)) ('P53', 'Gene', (189, 192)) ('p53', 'Gene', (153, 156)) ('P53', 'Gene', '22059', (189, 192)) ('MSigDB hallmarks', 'Disease', (172, 188)) ('p53+/+', 'Var', (28, 34)) ('tumor', 'Disease', 'MESH:D009369', (278, 283)) ('MEF', 'Gene', (67, 70)) ('Nutlin-inducible expression', 'MPA', (81, 108)) 1312 31483066 Gene set enrichment analysis (GSEA) showed a highly significant enrichment of a p53 target gene signature (MSigDB P53_downstream_pathway) in p53+/+ cells compared to either p53EE/EE or p53-/- MEFs, but no enrichment in p53EE/EE versus p53-/- MEFs (Fig 1D). ('SE', 'Disease', 'None', (31, 33)) ('p53 target', 'Gene', (80, 90)) ('MEF', 'Gene', (192, 195)) ('MEF', 'Gene', '56501', (192, 195)) ('P53', 'Gene', '22059', (114, 117)) ('P53', 'Gene', (114, 117)) ('MEF', 'Gene', (242, 245)) ('p53+/+', 'Var', (141, 147)) ('MEF', 'Gene', '56501', (242, 245)) 1313 31483066 The lack of p53 target gene activation in p53EE/EE MEFs was confirmed also under conditions of DNA damage induced with doxorubicin (Fig 1F). ('doxorubicin', 'Chemical', 'MESH:D004317', (119, 130)) ('MEF', 'Gene', (51, 54)) ('p53EE/EE', 'Var', (42, 50)) ('MEF', 'Gene', '56501', (51, 54)) 1314 31483066 Western blots revealed increased p53 expression in p53EE/EE versus p53+/+ MEFs, which was further augmented by Nutlin or doxorubicin:yet in the absence of detectable expression of the p53 targets p21 and Mdm2 (Fig 1G and H). ('p21', 'Gene', (196, 199)) ('MEF', 'Gene', (74, 77)) ('p53EE/EE', 'Var', (51, 59)) ('MEF', 'Gene', '56501', (74, 77)) ('p21', 'Gene', '12575', (196, 199)) ('p53', 'Gene', (33, 36)) ('doxorubicin', 'Chemical', 'MESH:D004317', (121, 132)) ('Nutlin', 'Chemical', 'MESH:C482204', (111, 117)) ('increased', 'PosReg', (23, 32)) ('expression', 'MPA', (37, 47)) 1316 31483066 In response to various types of stress, wild-type p53 elicits cell cycle arrest and senescence mediated by transcriptional activation of target genes, such as Cdkn1a/p21. ('senescence', 'CPA', (84, 94)) ('wild-type', 'Var', (40, 49)) ('Cdkn1a', 'Gene', (159, 165)) ('cell cycle arrest', 'CPA', (62, 79)) ('p21', 'Gene', (166, 169)) ('p53', 'Gene', (50, 53)) ('activation', 'PosReg', (123, 133)) ('elicits', 'Reg', (54, 61)) ('p21', 'Gene', '12575', (166, 169)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (62, 79)) ('Cdkn1a', 'Gene', '12575', (159, 165)) 1317 31483066 Consistent with the inability of p53EE to induce target genes, p53EE/EE and p53-null MEFs comparably failed to undergo cell cycle arrest in response to doxorubicin-triggered DNA damage (Fig 2A) or to enter senescence upon enforced expression of oncogenic Ras (Fig EV2A) or in vitro passaging (Figs 2B and EV2B). ('doxorubicin', 'Chemical', 'MESH:D004317', (152, 163)) ('cell cycle arrest', 'CPA', (119, 136)) ('failed', 'NegReg', (101, 107)) ('MEF', 'Gene', (85, 88)) ('p53EE/EE', 'Var', (63, 71)) ('enter', 'Reg', (200, 205)) ('MEF', 'Gene', '56501', (85, 88)) ('response to doxorubicin-triggered DNA damage', 'MPA', (140, 184)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (119, 136)) ('senescence', 'CPA', (206, 216)) 1318 31483066 While immortalization with adenoviral E1A.12S strongly sensitized p53+/+ MEFs to apoptosis, E1A-expressing p53EE/EE and p53-null MEFs remained refractory to apoptosis induction by genotoxic damage or Nutlin (Fig 2C). ('MEF', 'Gene', (129, 132)) ('MEF', 'Gene', '56501', (129, 132)) ('apoptosis', 'CPA', (81, 90)) ('p53EE/EE', 'Var', (107, 115)) ('sensitized', 'Reg', (55, 65)) ('Nutlin', 'Chemical', 'MESH:C482204', (200, 206)) ('genotoxic damage', 'Disease', (180, 196)) ('MEF', 'Gene', (73, 76)) ('MEF', 'Gene', '56501', (73, 76)) ('genotoxic damage', 'Disease', 'MESH:D009422', (180, 196)) 1320 31483066 Of note, we have previously reported a similar but more selective apoptosis defect in p53RR mice carrying the E177R (RR) cooperativity mutation (Fig EV2C) (Timofeev et al, 2013). ('E177R', 'Var', (110, 115)) ('defect', 'NegReg', (76, 82)) ('apoptosis', 'CPA', (66, 75)) ('E177R', 'Mutation', 'p.E177R', (110, 115)) ('mice', 'Species', '10090', (92, 96)) 1321 31483066 We therefore crossed p53EE/EE mice to p53RR/RR mice and obtained compound p53EE/RR animals that launched an apoptotic DNA damage response like p53+/+ animals in thymocytes ex vivo (Fig EV2D) and upon whole-body irradiation in vivo (Fig EV2E). ('mice', 'Species', '10090', (30, 34)) ('apoptotic DNA damage response', 'MPA', (108, 137)) ('launched', 'PosReg', (96, 104)) ('p53EE/RR', 'Var', (74, 82)) ('mice', 'Species', '10090', (47, 51)) 1322 31483066 Rescue of the apoptosis deficiency of p53EE/EE mice by the equally apoptosis-defective p53RR mutant proves that the p53EE loss-of-function phenotype is directly linked to the inability to form the salt bridge responsible for cooperative DNA binding and in turn further excludes global DBD misfolding or secondary local structural alterations at the DNA binding surface as an underlying cause. ('loss-of-function', 'NegReg', (122, 138)) ('p53EE', 'Gene', (116, 121)) ('mutant', 'Var', (93, 99)) ('inability', 'NegReg', (175, 184)) ('p53RR', 'Gene', (87, 92)) ('mice', 'Species', '10090', (47, 51)) 1324 31483066 None of these effects were recorded in p53EE/EE or p53-null mice (Figs 2F-I and EV2G and H), indicating a complete defect of p53EE regarding classical p53 effector functions in vivo. ('mice', 'Species', '10090', (60, 64)) ('defect', 'NegReg', (115, 121)) ('p53EE', 'Var', (125, 130)) 1325 31483066 When passaging p53EE/EE MEFs for longer time periods, we noted that:unlike p53-/- MEFs:the proliferation rate of p53EE/EE MEFs eventually declined and the cells started to express the senescence marker SA-beta-galactosidase (Fig EV3A and B). ('MEF', 'Gene', (82, 85)) ('beta-galactosidase', 'Gene', (205, 223)) ('beta-galactosidase', 'Gene', '12091', (205, 223)) ('MEF', 'Gene', '56501', (82, 85)) ('MEF', 'Gene', (122, 125)) ('MEF', 'Gene', '56501', (122, 125)) ('MEF', 'Gene', (24, 27)) ('MEF', 'Gene', '56501', (24, 27)) ('proliferation rate', 'CPA', (91, 109)) ('p53EE/EE', 'Var', (113, 121)) ('declined', 'NegReg', (138, 146)) 1326 31483066 This was accompanied by a progressive increase in p53EE protein levels, but without the increased expression in p53 target genes that was detectable in p53+/+ MEFs (Fig EV3C and D). ('increase', 'PosReg', (38, 46)) ('p53EE protein levels', 'MPA', (50, 70)) ('p53+/+', 'Var', (152, 158)) ('MEF', 'Gene', (159, 162)) ('MEF', 'Gene', '56501', (159, 162)) 1327 31483066 Senescent p53EE/EE MEFs exhibited strongly elevated levels of mitochondrial ROS (Fig EV3F). ('p53EE/EE', 'Var', (10, 18)) ('MEF', 'Gene', (19, 22)) ('elevated', 'PosReg', (43, 51)) ('MEF', 'Gene', '56501', (19, 22)) ('mitochondrial ROS', 'MPA', (62, 79)) ('levels', 'MPA', (52, 58)) 1328 31483066 Oxygen reduction from ambient to physiological levels promoted immortalization (Fig EV3G and H), implying ROS as the trigger of senescence in response to p53EE accumulation. ('promoted', 'PosReg', (54, 62)) ('Oxygen', 'Chemical', 'MESH:D010100', (0, 6)) ('immortalization', 'CPA', (63, 78)) ('p53EE', 'Var', (154, 159)) ('Oxygen reduction', 'Var', (0, 16)) 1329 31483066 Instead, we observed a somewhat reduced oxidative ATP production under basal conditions and significantly impaired spare respiratory capacity upon mitochondrial uncoupling, suggesting an inhibitory effect of p53EE on mitochondrial functions (Fig EV3I). ('p53EE', 'Var', (208, 213)) ('oxidative ATP production', 'MPA', (40, 64)) ('spare respiratory capacity', 'MPA', (115, 141)) ('reduced', 'NegReg', (32, 39)) ('impaired', 'NegReg', (106, 114)) ('mitochondrial', 'MPA', (217, 230)) ('ATP', 'Chemical', 'MESH:D000255', (50, 53)) 1330 31483066 In line, the mitochondrial DNA content of p53EE/EE MEFs was significantly decreased, especially at late passages (Fig EV3J). ('p53EE/EE', 'Var', (42, 50)) ('MEF', 'Gene', (51, 54)) ('mitochondrial DNA content', 'MPA', (13, 38)) ('decreased', 'NegReg', (74, 83)) ('MEF', 'Gene', '56501', (51, 54)) 1332 31483066 Basal and ROS-triggered expression of anti-oxidative Nrf2 target genes was diminished in p53EE/EE MEFs (Fig EV3K and L), strongly suggesting that p53EE inhibits Nrf2 activity similar as other p53 mutants (Walerych et al, 2016; Liu et al, 2017; Merkel et al, 2017). ('diminished', 'NegReg', (75, 85)) ('Nrf2', 'Gene', (53, 57)) ('Nrf2', 'Gene', '18024', (161, 165)) ('activity', 'MPA', (166, 174)) ('MEF', 'Gene', (98, 101)) ('anti-oxidative', 'MPA', (38, 52)) ('MEF', 'Gene', '56501', (98, 101)) ('inhibits', 'NegReg', (152, 160)) ('Nrf2', 'Gene', (161, 165)) ('p53EE', 'Var', (146, 151)) ('Nrf2', 'Gene', '18024', (53, 57)) 1333 31483066 We therefore postulate that the Nrf2-inhibitory activity of p53EE sensitizes to ROS-induced senescence. ('Nrf2', 'Gene', (32, 36)) ('ROS-induced senescence', 'MPA', (80, 102)) ('p53EE', 'Var', (60, 65)) ('Nrf2', 'Gene', '18024', (32, 36)) ('sensitizes', 'Reg', (66, 76)) 1334 31483066 Genetic ablation of Mdm2 in mice causes early embryonic lethality due to massive apoptosis initiated as early as E3.5, whereas simultaneous disruption of the Trp53 gene rescues the lethal phenotype (Jones et al, 1995; Montes de Oca Luna et al, 1995; Chavez-Reyes et al, 2003). ('Trp53', 'Gene', (158, 163)) ('disruption', 'Var', (140, 150)) ('mice', 'Species', '10090', (28, 32)) ('apoptosis', 'CPA', (81, 90)) ('Mdm2', 'Gene', (20, 24)) ('Lu', 'Chemical', 'MESH:D008187', (232, 234)) ('embryonic lethality', 'Disease', (46, 65)) ('embryonic lethality', 'Disease', 'MESH:D020964', (46, 65)) ('ablation', 'Var', (8, 16)) 1335 31483066 Embryonic lethality of Mdm2 knock-out animals is also rescued by p53 missense mutations such as Trp53 R246S and Trp53 R172H that mimic human hot-spot mutants p53R249S and p53R175H, respectively (Lee et al, 2012). ('p53', 'Gene', (65, 68)) ('Embryonic lethality', 'Disease', (0, 19)) ('p53R249S', 'Var', (158, 166)) ('R172H', 'Mutation', 'p.R172H', (118, 123)) ('Trp53', 'Gene', (96, 101)) ('R172H', 'Var', (118, 123)) ('Embryonic lethality', 'Disease', 'MESH:D020964', (0, 19)) ('human', 'Species', '9606', (135, 140)) ('p53R175H', 'Var', (171, 179)) ('p53R175H', 'Mutation', 'p.R53,175H', (171, 179)) ('R246S', 'Var', (102, 107)) ('Trp53', 'Gene', (112, 117)) ('R246S', 'Mutation', 'p.R246S', (102, 107)) ('rescued', 'PosReg', (54, 61)) 1336 31483066 Moreover, the apoptosis-deficient mutant p53R172P also rescued Mdm2 -/- embryos, but the newborn mice showed severe developmental defects and died soon due to ROS-dependent hematopoietic failure (Abbas et al, 2010). ('mice', 'Species', '10090', (97, 101)) ('hematopoietic failure', 'Disease', (173, 194)) ('developmental defects', 'Disease', 'MESH:D009436', (116, 137)) ('p53R172P', 'Var', (41, 49)) ('hematopoietic failure', 'Disease', 'MESH:D019337', (173, 194)) ('developmental defects', 'Disease', (116, 137)) 1338 31483066 Trp53 EE/EE;Mdm2 -/- embryos displayed severe developmental defects starting at days E9.5-10.5 (Fig 3C and D), and no living Trp53 EE/EE;Mdm2 -/- embryos were recovered after day E12.5 (Fig 3D). ('developmental defects', 'Disease', (46, 67)) ('EE/EE', 'Var', (6, 11)) ('developmental defects', 'Disease', 'MESH:D009436', (46, 67)) ('Trp53', 'Gene', (0, 5)) 1339 31483066 However, the lethality of Trp53 EE/EE;Mdm2 -/- embryos contrasted with the lack of any detectable cytotoxic activity of the Mdm2 inhibitor Nutlin in p53EE/EE MEFs (Fig 2C). ('lethality', 'CPA', (13, 22)) ('EE/EE', 'Var', (32, 37)) ('MEF', 'Gene', (158, 161)) ('MEF', 'Gene', '56501', (158, 161)) ('Nutlin', 'Chemical', 'MESH:C482204', (139, 145)) ('p53EE/EE', 'Var', (149, 157)) ('Trp53', 'Gene', (26, 31)) 1341 31483066 This prompted us to investigate whether accumulation of p53EE caused by loss of Mdm2 sensitizes cells to doxorubicin whose cytotoxicity involves ROS- and DNA damage-dependent mechanisms (Trachootham et al, 2009; Huang et al, 2011). ('cytotoxicity', 'Disease', (123, 135)) ('doxorubicin', 'Chemical', 'MESH:D004317', (105, 116)) ('loss', 'Var', (72, 76)) ('sensitizes', 'Reg', (85, 95)) ('cytotoxicity', 'Disease', 'MESH:D064420', (123, 135)) ('accumulation', 'PosReg', (40, 52)) ('p53EE', 'Var', (56, 61)) ('Mdm2', 'Gene', (80, 84)) 1342 31483066 Because of the embryonic lethality, we could not establish Trp53 EE/EE;Mdm2 -/- MEFs and used Trp53 -/-;Mdm2 -/- (DKO) MEFs ectopically expressing p53EE from a tetracycline-activated promoter instead (DKO-tetEE). ('embryonic lethality', 'Disease', 'MESH:D020964', (15, 34)) ('embryonic lethality', 'Disease', (15, 34)) ('MEF', 'Gene', (80, 83)) ('MEF', 'Gene', '56501', (80, 83)) ('MEF', 'Gene', (119, 122)) ('tetracycline', 'Chemical', 'MESH:D013754', (160, 172)) ('MEF', 'Gene', '56501', (119, 122)) ('p53EE', 'Var', (147, 152)) 1343 31483066 As expected, in the absence of tetracycline the DKO-tetEE MEFs were as resistant to doxorubicin as the parental DKO cells but showed significantly elevated levels of apoptosis in response to doxorubicin treatment following induction of p53EE expression (Fig 4A). ('doxorubicin', 'Chemical', 'MESH:D004317', (191, 202)) ('doxorubicin', 'Chemical', 'MESH:D004317', (84, 95)) ('response to doxorubicin treatment', 'MPA', (179, 212)) ('levels', 'MPA', (156, 162)) ('MEF', 'Gene', (58, 61)) ('MEF', 'Gene', '56501', (58, 61)) ('p53EE expression', 'Var', (236, 252)) ('elevated', 'PosReg', (147, 155)) ('tetracycline', 'Chemical', 'MESH:D013754', (31, 43)) ('apoptosis', 'MPA', (166, 175)) 1344 31483066 Next, we tested whether pharmacological inhibition of Mdm2 with Nutlin has a similar effect on E1A-MEFs with endogenous expression of p53EE. ('p53EE', 'Var', (134, 139)) ('MEF', 'Gene', (99, 102)) ('MEF', 'Gene', '56501', (99, 102)) ('tested', 'Reg', (9, 15)) ('Nutlin', 'Chemical', 'MESH:C482204', (64, 70)) 1345 31483066 While Nutlin or doxorubicin alone had no or minimal effects on p53EE/EE MEFs, combined treatment of p53EE/EE cells with Nutlin and doxorubicin caused significant, p53-dependent reduction in proliferation and survival (Fig 4C and D). ('Nutlin', 'Chemical', 'MESH:C482204', (120, 126)) ('doxorubicin', 'Chemical', 'MESH:D004317', (16, 27)) ('p53EE/EE', 'Var', (100, 108)) ('doxorubicin', 'Chemical', 'MESH:D004317', (131, 142)) ('reduction', 'NegReg', (177, 186)) ('Nutlin', 'Chemical', 'MESH:C482204', (6, 12)) ('proliferation', 'CPA', (190, 203)) ('survival', 'CPA', (208, 216)) ('MEF', 'Gene', (72, 75)) ('MEF', 'Gene', '56501', (72, 75)) 1348 31483066 The Hsp90 inhibitor ganetespib, which targets mutant p53 for proteasomal degradation, efficiently degraded p53EE in E1A-MEFs, highlighting a role for Hsp90 in p53EE stability (Fig 4H). ('MEF', 'Gene', (120, 123)) ('ganetespib', 'Chemical', 'MESH:C533237', (20, 30)) ('mutant', 'Var', (46, 52)) ('p53', 'Gene', (53, 56)) ('p53EE', 'Var', (107, 112)) ('MEF', 'Gene', '56501', (120, 123)) ('degraded', 'NegReg', (98, 106)) 1350 31483066 In line with the role of Mdm2 for ganetespib-induced mutant p53 degradation (Li et al, 2011), p53EE degradation by ganetespib was prevented when administered simultaneously with Nutlin (Fig 4H). ('Nutlin', 'Chemical', 'MESH:C482204', (178, 184)) ('ganetespib', 'Chemical', 'MESH:C533237', (34, 44)) ('p53', 'Gene', (60, 63)) ('degradation', 'MPA', (64, 75)) ('mutant', 'Var', (53, 59)) ('ganetespib', 'Chemical', 'MESH:C533237', (115, 125)) 1353 31483066 Remarkably, mitochondrial fractions of untreated p53EE/EE MEFs contained p53 at levels similarly high as p53+/+ cells following doxorubicin treatment. ('p53EE/EE', 'Var', (49, 57)) ('MEF', 'Gene', (58, 61)) ('MEF', 'Gene', '56501', (58, 61)) ('p53', 'MPA', (73, 76)) ('doxorubicin', 'Chemical', 'MESH:D004317', (128, 139)) 1355 31483066 The increased amount of p53EE at the mitochondria could therefore provide a plausible explanation for the sensitivity of p53EE/EE MEFs to apoptotic stimuli even in the absence of p53 target gene activation. ('MEF', 'Gene', '56501', (130, 133)) ('increased', 'PosReg', (4, 13)) ('p53EE/EE', 'Var', (121, 129)) ('MEF', 'Gene', (130, 133)) 1356 31483066 To test whether p53EE is also capable of inducing apoptosis in human cancer cells, we generated a human p53-null H1299 lung cancer cell line with stable Tet-inducible expression of the human p53R181E (p53EE) mutant. ('cancer', 'Disease', (124, 130)) ('human', 'Species', '9606', (185, 190)) ('lung cancer', 'Disease', (119, 130)) ('human', 'Species', '9606', (63, 68)) ('lung cancer', 'Phenotype', 'HP:0100526', (119, 130)) ('cancer', 'Phenotype', 'HP:0002664', (69, 75)) ('cancer', 'Phenotype', 'HP:0002664', (124, 130)) ('p53R181E', 'Var', (191, 199)) ('lung cancer', 'Disease', 'MESH:D008175', (119, 130)) ('cancer', 'Disease', 'MESH:D009369', (69, 75)) ('R181E', 'Mutation', 'p.R181E', (194, 199)) ('human', 'Species', '9606', (98, 103)) ('cancer', 'Disease', 'MESH:D009369', (124, 130)) ('cancer', 'Disease', (69, 75)) 1357 31483066 Overexpression of p53EE rendered H1299 cells, which express only barely detectable levels of Mdm2, sensitive to doxorubicin, and addition of Nutlin further augmented this effect (Fig 5A). ('Nutlin', 'Chemical', 'MESH:C482204', (141, 147)) ('doxorubicin', 'Chemical', 'MESH:D004317', (112, 123)) ('sensitive to doxorubicin', 'MPA', (99, 123)) ('p53EE', 'Var', (18, 23)) 1358 31483066 Cell death was inhibited by Q-VD-OPh, a blocker of caspase-dependent apoptosis, but not by the ferroptosis inhibitor ferrostatin as a control (Fig 5B). ('inhibited', 'NegReg', (15, 24)) ('ferrostatin', 'Chemical', 'MESH:C573944', (117, 128)) ('Cell death', 'CPA', (0, 10)) ('Q-VD-OPh', 'Var', (28, 36)) 1359 31483066 Annexin V staining revealed a p53-dependent increase in apoptotic cells under combined doxorubicin and Nutlin treatment, confirming the observed cell death as apoptotic (Fig 5C). ('Annexin V', 'Gene', '308', (0, 9)) ('Annexin V', 'Gene', (0, 9)) ('5C', 'Chemical', 'MESH:D002244', (174, 176)) ('Nutlin', 'Chemical', 'MESH:C482204', (103, 109)) ('p53-dependent', 'Var', (30, 43)) ('increase', 'PosReg', (44, 52)) ('apoptotic cells', 'CPA', (56, 71)) ('doxorubicin', 'Chemical', 'MESH:D004317', (87, 98)) 1360 31483066 Proximity ligation experiments demonstrated a specific co-localization of p53EE with the outer mitochondrial membrane protein Tom20 in untreated and, even more, in doxorubicin-treated cells (Fig 5E and F). ('co-localization', 'Interaction', (55, 70)) ('p53EE', 'Var', (74, 79)) ('doxorubicin', 'Chemical', 'MESH:D004317', (164, 175)) ('Tom20', 'Gene', (126, 131)) ('Tom20', 'Gene', '67952', (126, 131)) 1361 31483066 p53EE co-localization was also observed with the Bcl-2 family members Bcl-xL, Bcl-2, and Bak (Fig 5F), which were previously shown to specifically interact with wild-type but not mutant p53 (Mihara et al, 2003; Leu et al, 2004; Pietsch et al, 2008). ('Bcl-2', 'Gene', (49, 54)) ('mutant', 'Var', (179, 185)) ('Bcl-2', 'Gene', '12043', (49, 54)) ('Leu', 'Chemical', 'MESH:C038361', (211, 214)) ('Bak', 'Gene', (89, 92)) ('p53', 'Gene', (186, 189)) ('Bcl-2', 'Gene', (78, 83)) ('Bcl-2', 'Gene', '12043', (78, 83)) ('Bcl-xL', 'Gene', (70, 76)) ('Bcl-xL', 'Gene', '12048', (70, 76)) ('interact', 'Interaction', (147, 155)) ('Bak', 'Gene', '12018', (89, 92)) 1363 31483066 In support of a direct mitochondrial apoptotic priming activity, p53EE expression sensitized H1299 cells to mitochondrial depolarization by BID BH3 peptides (Fig 5G). ('sensitized', 'Reg', (82, 92)) ('mitochondrial depolarization', 'MPA', (108, 136)) ('BID', 'Gene', '637', (140, 143)) ('p53EE expression', 'Var', (65, 81)) ('BID', 'Gene', (140, 143)) 1364 31483066 The apoptotic activity of p53EE raised the question of whether p53 mutants in cancer patients can also retain apoptotic activity. ('p53', 'Gene', (63, 66)) ('apoptotic activity', 'CPA', (110, 128)) ('p53EE', 'Var', (26, 31)) ('cancer', 'Disease', (78, 84)) ('cancer', 'Disease', 'MESH:D009369', (78, 84)) ('mutants', 'Var', (67, 74)) ('patients', 'Species', '9606', (85, 93)) ('cancer', 'Phenotype', 'HP:0002664', (78, 84)) ('apoptotic activity', 'MPA', (4, 22)) 1365 31483066 Although p53EE (R181E) is not found in cancer patients, various other cooperativity mutations have been recurrently identified as somatic cancer mutations and account for an estimated 34,000 new cancer cases each year (Leroy et al, 2014). ('R181E', 'Mutation', 'p.R181E', (16, 21)) ('cancer', 'Disease', (138, 144)) ('patients', 'Species', '9606', (46, 54)) ('cancer', 'Disease', (39, 45)) ('cancer', 'Disease', 'MESH:D009369', (39, 45)) ('cancer', 'Disease', (195, 201)) ('cancer', 'Disease', 'MESH:D009369', (195, 201)) ('cancer', 'Phenotype', 'HP:0002664', (138, 144)) ('mutations', 'Var', (84, 93)) ('cancer', 'Phenotype', 'HP:0002664', (39, 45)) ('p53EE (R181E', 'Var', (9, 21)) ('cancer', 'Disease', 'MESH:D009369', (138, 144)) ('cancer', 'Phenotype', 'HP:0002664', (195, 201)) 1367 31483066 In particular, R181L was among the first p53 cancer mutants reported to be specifically deficient for binding and activating pro-apoptotic target genes while retaining regulation of other genes such as CDKN1A and MDM2 (Ludwig et al, 1996; Schlereth et al, 2010a). ('deficient', 'NegReg', (88, 97)) ('R181L', 'Var', (15, 20)) ('MDM2', 'Gene', '17246', (213, 217)) ('Lu', 'Chemical', 'MESH:D008187', (219, 221)) ('binding', 'Interaction', (102, 109)) ('R181L', 'Mutation', 'p.R181L', (15, 20)) ('cancer', 'Phenotype', 'HP:0002664', (45, 51)) ('CDKN1A', 'Gene', (202, 208)) ('MDM2', 'Gene', (213, 217)) ('cancer', 'Disease', 'MESH:D009369', (45, 51)) ('regulation', 'MPA', (168, 178)) ('activating', 'PosReg', (114, 124)) ('CDKN1A', 'Gene', '12575', (202, 208)) ('cancer', 'Disease', (45, 51)) ('p53', 'Gene', (41, 44)) 1369 31483066 However, R181L:like p53EE:effectively triggered apoptosis when combined with doxorubicin and Nutlin (Fig 5H). ('apoptosis', 'CPA', (48, 57)) ('triggered', 'Reg', (38, 47)) ('doxorubicin', 'Chemical', 'MESH:D004317', (77, 88)) ('Nutlin', 'Chemical', 'MESH:C482204', (93, 99)) ('R181L', 'Var', (9, 14)) ('R181L', 'Mutation', 'p.R181L', (9, 14)) 1370 31483066 In contrast, the R175P mutant, which also lacks the ability to transactivate pro-apoptotic target genes but belongs to the class of structural mutations (Rowan et al, 1996; Liu et al, 2004), failed to trigger apoptosis in the presence of doxorubicin and Nutlin (Fig 5H). ('Rowan', 'Species', '36599', (154, 159)) ('failed', 'NegReg', (191, 197)) ('Nutlin', 'Chemical', 'MESH:C482204', (254, 260)) ('apoptosis', 'CPA', (209, 218)) ('R175P', 'Var', (17, 22)) ('R175P', 'Mutation', 'p.R175P', (17, 22)) ('trigger', 'Reg', (201, 208)) ('doxorubicin', 'Chemical', 'MESH:D004317', (238, 249)) 1371 31483066 We conclude that, in principle, also cancer mutants can trigger apoptosis when stimulated sufficiently with Nutlin and doxorubicin in a manner depending on the identity of the individual mutant. ('cancer', 'Disease', 'MESH:D009369', (37, 43)) ('cancer', 'Disease', (37, 43)) ('apoptosis', 'CPA', (64, 73)) ('doxorubicin', 'Chemical', 'MESH:D004317', (119, 130)) ('Nutlin', 'Chemical', 'MESH:C482204', (108, 114)) ('cancer', 'Phenotype', 'HP:0002664', (37, 43)) ('mutants', 'Var', (44, 51)) ('trigger', 'Reg', (56, 63)) 1372 31483066 To investigate the role of DNA binding cooperativity for tumor suppression, we aged cohorts of EE mutant mice. ('tumor', 'Disease', 'MESH:D009369', (57, 62)) ('tumor', 'Phenotype', 'HP:0002664', (57, 62)) ('tumor', 'Disease', (57, 62)) ('mice', 'Species', '10090', (105, 109)) ('mutant', 'Var', (98, 104)) 1373 31483066 Surprisingly, despite evidence for residual apoptotic activity of p53EE (Figs 3, 4, 5), homozygous p53EE/EE and hemizygous p53EE/- mice developed tumors very rapidly resulting in a short median survival of 150 and 128 days, respectively (Fig 6A). ('p53EE/EE', 'Var', (99, 107)) ('tumors', 'Phenotype', 'HP:0002664', (146, 152)) ('tumor', 'Phenotype', 'HP:0002664', (146, 151)) ('tumors', 'Disease', 'MESH:D009369', (146, 152)) ('p53EE', 'Var', (66, 71)) ('mice', 'Species', '10090', (131, 135)) ('tumors', 'Disease', (146, 152)) 1375 31483066 The lack of a difference in survival between p53EE/+ and p53+/- mice, despite evidence for a dominant-negative activity of p53EE in overexpression studies (Schlereth et al, 2010a), is reminiscent of mice heterozygous for the hot-spot mutants R172H and R270H and consistent with the hypothesis that dominant-negative effects might require prior mutant p53 stabilization (Lang et al, 2004; Olive et al, 2004). ('p53', 'Gene', (351, 354)) ('R270H', 'Var', (252, 257)) ('mice', 'Species', '10090', (199, 203)) ('R172H', 'Mutation', 'p.R172H', (242, 247)) ('activity', 'MPA', (111, 119)) ('p53EE', 'Var', (123, 128)) ('R172H', 'Var', (242, 247)) ('R270H', 'Mutation', 'p.R270H', (252, 257)) ('mice', 'Species', '10090', (64, 68)) ('Olive', 'Species', '4146', (388, 393)) 1376 31483066 As previously reported for p53-/- animals, the tumor spectra of p53EE/EE and hemizygous p53EE/- were dominated by thymic lymphoma (Fig 6B; Appendix Table S1). ('thymic lymphoma', 'Disease', (114, 129)) ('tumor', 'Disease', 'MESH:D009369', (47, 52)) ('tumor', 'Phenotype', 'HP:0002664', (47, 52)) ('p53EE/EE', 'Var', (64, 72)) ('thymic lymphoma', 'Disease', 'MESH:D013953', (114, 129)) ('lymphoma', 'Phenotype', 'HP:0002665', (121, 129)) ('tumor', 'Disease', (47, 52)) 1378 31483066 The tumors of p53EE/+ were similar to those of p53+/- mice and mostly non-thymic lymphomas, sarcomas, and several types of carcinoma (Fig 6B, Appendix Table S2). ('sarcomas', 'Disease', (92, 100)) ('thymic lymphoma', 'Disease', 'MESH:D013953', (74, 89)) ('carcinoma', 'Disease', 'MESH:D002277', (123, 132)) ('lymphomas', 'Phenotype', 'HP:0002665', (81, 90)) ('carcinoma', 'Disease', (123, 132)) ('tumors', 'Disease', (4, 10)) ('p53EE/+', 'Var', (14, 21)) ('tumors', 'Disease', 'MESH:D009369', (4, 10)) ('tumors', 'Phenotype', 'HP:0002664', (4, 10)) ('carcinoma', 'Phenotype', 'HP:0030731', (123, 132)) ('lymphoma', 'Phenotype', 'HP:0002665', (81, 89)) ('sarcomas', 'Phenotype', 'HP:0100242', (92, 100)) ('sarcomas', 'Disease', 'MESH:D012509', (92, 100)) ('tumor', 'Phenotype', 'HP:0002664', (4, 9)) ('thymic lymphoma', 'Disease', (74, 89)) ('lymphomas', 'Disease', (81, 90)) ('lymphomas', 'Disease', 'MESH:D008223', (81, 90)) ('mice', 'Species', '10090', (54, 58)) 1379 31483066 Unlike what has been reported for the p53 hot-spot mutants R172H or R270H, we did not observe an increase in metastatic tumors in p53EE/EE and p53EE/- compared to p53-/- and p53+/- mice (Appendix Table S1), suggesting a lack of GOF properties of the EE mutant. ('p53EE/EE', 'Var', (130, 138)) ('tumors', 'Disease', (120, 126)) ('mice', 'Species', '10090', (181, 185)) ('tumors', 'Disease', 'MESH:D009369', (120, 126)) ('tumors', 'Phenotype', 'HP:0002664', (120, 126)) ('p53EE/-', 'Var', (143, 150)) ('R270H', 'Var', (68, 73)) ('tumor', 'Phenotype', 'HP:0002664', (120, 125)) ('R172H', 'Mutation', 'p.R172H', (59, 64)) ('R270H', 'Mutation', 'p.R270H', (68, 73)) 1380 31483066 Similar as described for tumors in R172H-mutant mice (Terzian et al, 2008), we noted varying levels of p53EE expression in spontaneous tumors arising in p53EE/EE or p53EE/- mice, with a high fraction (67%) of p53EE/EE tumors exhibiting p53EE stabilization comparable to p53-mutated human cancer samples (Figs 6C and EV4 and EV5). ('tumor', 'Phenotype', 'HP:0002664', (25, 30)) ('tumor', 'Phenotype', 'HP:0002664', (135, 140)) ('tumors', 'Disease', (25, 31)) ('cancer', 'Disease', (288, 294)) ('tumors', 'Disease', (135, 141)) ('cancer', 'Phenotype', 'HP:0002664', (288, 294)) ('mice', 'Species', '10090', (173, 177)) ('tumors', 'Disease', 'MESH:D009369', (25, 31)) ('p53EE', 'Gene', (103, 108)) ('tumors', 'Disease', 'MESH:D009369', (135, 141)) ('tumors', 'Phenotype', 'HP:0002664', (218, 224)) ('R172H', 'Mutation', 'p.R172H', (35, 40)) ('p53EE/EE', 'Var', (209, 217)) ('cancer', 'Disease', 'MESH:D009369', (288, 294)) ('tumor', 'Phenotype', 'HP:0002664', (218, 223)) ('tumors', 'Disease', (218, 224)) ('tumors', 'Phenotype', 'HP:0002664', (25, 31)) ('human', 'Species', '9606', (282, 287)) ('mice', 'Species', '10090', (48, 52)) ('tumors', 'Phenotype', 'HP:0002664', (135, 141)) ('tumors', 'Disease', 'MESH:D009369', (218, 224)) 1381 31483066 This indicates that the absence of a pro-metastatic GOF cannot be explained by a lack of constitutive p53EE stabilization and suggests that p53EE, similar to p53 mutants like R246S or G245S (Lee et al, 2012; Hanel et al, 2013), lacks the pro-metastatic GOF activity described for R172H and R270H (Olive et al, 2004). ('R270H', 'Var', (290, 295)) ('p53EE', 'Var', (140, 145)) ('R172H', 'Mutation', 'p.R172H', (280, 285)) ('G245S', 'Var', (184, 189)) ('G245S', 'Mutation', 'p.G245S', (184, 189)) ('R172H', 'Var', (280, 285)) ('Olive', 'Species', '4146', (297, 302)) ('R270H', 'Mutation', 'p.R270H', (290, 295)) ('pro-metastatic GOF activity', 'MPA', (238, 265)) ('R246S', 'Var', (175, 180)) ('R246S', 'Mutation', 'p.R246S', (175, 180)) ('lacks', 'NegReg', (228, 233)) 1382 31483066 To explore whether p53EE can counteract oncogene-induced tumorigenesis in a genetically more defined setting, we crossed p53EE animals with Emu-Myc mice which serve as a well-characterized model of Burkitt-like B-cell lymphoma (Adams et al, 1985). ('Burkitt-like B-cell lymphoma', 'Phenotype', 'HP:0030080', (198, 226)) ('lymphoma', 'Phenotype', 'HP:0002665', (218, 226)) ('tumor', 'Disease', 'MESH:D009369', (57, 62)) ('Myc', 'Gene', (144, 147)) ('Myc', 'Gene', '17869', (144, 147)) ('mice', 'Species', '10090', (148, 152)) ('tumor', 'Phenotype', 'HP:0002664', (57, 62)) ('B-cell lymphoma', 'Disease', 'MESH:D016393', (211, 226)) ('B-cell lymphoma', 'Disease', (211, 226)) ('p53EE', 'Var', (121, 126)) ('B-cell lymphoma', 'Phenotype', 'HP:0012191', (211, 226)) ('tumor', 'Disease', (57, 62)) 1383 31483066 Loss of p53 is known to strongly accelerate lymphoma development in this model (Schmitt et al, 1999). ('lymphoma', 'Disease', 'MESH:D008223', (44, 52)) ('lymphoma', 'Phenotype', 'HP:0002665', (44, 52)) ('p53', 'Gene', (8, 11)) ('accelerate', 'PosReg', (33, 43)) ('Loss', 'Var', (0, 4)) ('lymphoma', 'Disease', (44, 52)) 1385 31483066 Likewise, in a model of acute myeloid leukemia induced by co-expression of the AML1/ETO9a (AE9) fusion oncoprotein and oncogenic NrasG12D (Nras) (Zuber et al, 2009), we observed fast malignant transformation of p53EE/EE hematopoietic stem cells (HSCs) and disease progression. ('induced by', 'Reg', (47, 57)) ('leukemia', 'Phenotype', 'HP:0001909', (38, 46)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (24, 46)) ('malignant transformation', 'CPA', (183, 207)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (30, 46)) ('acute myeloid leukemia', 'Disease', (24, 46)) ('AML1', 'Gene', (79, 83)) ('p53EE/EE', 'Var', (211, 219)) ('AML1', 'Gene', '12394', (79, 83)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (24, 46)) 1386 31483066 Recipient mice transplanted with AE9+Nras-transduced p53EE/EE or p53-/- HSCs succumbed to AML with indistinguishably short latency and significantly earlier than mice transplanted with AE9+Nras-transduced p53+/+ HSCs (Fig 6E). ('AML', 'Disease', (90, 93)) ('mice', 'Species', '10090', (10, 14)) ('mice', 'Species', '10090', (162, 166)) ('earlier', 'PosReg', (149, 156)) ('AML', 'Disease', 'MESH:D015470', (90, 93)) ('p53EE/EE', 'Var', (53, 61)) ('p53-/- HSCs', 'Var', (65, 76)) 1387 31483066 Notably, distinct from heterogenous p53EE stabilization in spontaneous tumors (Figs 6C and EV5), p53EE was highly accumulated in all lymphomas from Emu-Myc;p53EE/+ mice and in all AE9+Nras;p53EE/EE AML samples (Fig 6F and G). ('AML', 'Disease', 'MESH:D015470', (198, 201)) ('mice', 'Species', '10090', (164, 168)) ('lymphoma', 'Phenotype', 'HP:0002665', (133, 141)) ('accumulated', 'PosReg', (114, 125)) ('AML', 'Disease', (198, 201)) ('tumor', 'Phenotype', 'HP:0002664', (71, 76)) ('lymphomas', 'Disease', (133, 142)) ('lymphomas', 'Disease', 'MESH:D008223', (133, 142)) ('tumors', 'Phenotype', 'HP:0002664', (71, 77)) ('p53EE', 'Var', (97, 102)) ('tumors', 'Disease', (71, 77)) ('Myc', 'Gene', (152, 155)) ('lymphomas', 'Phenotype', 'HP:0002665', (133, 142)) ('Myc', 'Gene', '17869', (152, 155)) ('tumors', 'Disease', 'MESH:D009369', (71, 77)) 1388 31483066 As stabilization of mutant p53 involves protection from Mdm2-mediated degradation (Terzian et al, 2008), the uniform p53EE accumulation might be explained by the exceptionally strong oncogenic signaling through enforced expression of Myc and mutant Nras which inhibits Mdm2 via p19Arf (Zindy et al, 1998). ('accumulation', 'PosReg', (123, 135)) ('inhibits', 'NegReg', (260, 268)) ('Myc', 'Gene', (234, 237)) ('p53EE', 'Gene', (117, 122)) ('Myc', 'Gene', '17869', (234, 237)) ('mutant', 'Var', (242, 248)) ('p19Arf', 'Gene', (278, 284)) ('Nras', 'Gene', (249, 253)) ('p19Arf', 'Gene', '12578', (278, 284)) ('mutant', 'Var', (20, 26)) ('p53', 'Gene', (27, 30)) 1390 31483066 The lack of detectable tumor suppressor activity implies that the residual apoptotic activities are not effectively counter-selected during tumorigenesis and can be retained by cancer mutants:the R181L cooperativity mutant being one example (Fig 5H). ('cancer', 'Disease', (177, 183)) ('cancer', 'Disease', 'MESH:D009369', (177, 183)) ('tumor', 'Phenotype', 'HP:0002664', (140, 145)) ('tumor', 'Disease', 'MESH:D009369', (23, 28)) ('tumor', 'Disease', 'MESH:D009369', (140, 145)) ('tumor', 'Phenotype', 'HP:0002664', (23, 28)) ('R181L', 'Var', (196, 201)) ('cancer', 'Phenotype', 'HP:0002664', (177, 183)) ('R181L', 'Mutation', 'p.R181L', (196, 201)) ('tumor', 'Disease', (23, 28)) ('tumor', 'Disease', (140, 145)) 1391 31483066 In fact, p53EE was even constitutively stabilized in various tumor types arising in p53EE/EE mice (Fig 6), indicating escape from Mdm2-mediated degradation. ('tumor', 'Disease', 'MESH:D009369', (61, 66)) ('mice', 'Species', '10090', (93, 97)) ('tumor', 'Phenotype', 'HP:0002664', (61, 66)) ('p53EE/EE', 'Var', (84, 92)) ('tumor', 'Disease', (61, 66)) 1392 31483066 We therefore explored whether this accumulation of p53EE suffices to sensitize tumor cells to cytotoxic stress and provide a therapeutic window for tumor treatment. ('tumor', 'Phenotype', 'HP:0002664', (148, 153)) ('tumor', 'Disease', (79, 84)) ('p53EE', 'Var', (51, 56)) ('tumor', 'Disease', (148, 153)) ('tumor', 'Disease', 'MESH:D009369', (79, 84)) ('sensitize', 'Reg', (69, 78)) ('tumor', 'Disease', 'MESH:D009369', (148, 153)) ('tumor', 'Phenotype', 'HP:0002664', (79, 84)) 1393 31483066 First, we investigated whether p53EE influences the response of Emu-Myc lymphoma cells to mafosfamide (MAF), a cyclophosphamide (CTX) analogue active in vitro. ('CTX', 'Gene', (129, 132)) ('Myc', 'Gene', (68, 71)) ('Myc', 'Gene', '17869', (68, 71)) ('cyclophosphamide', 'Chemical', 'MESH:D003520', (111, 127)) ('CTX', 'Gene', '57276', (129, 132)) ('lymphoma', 'Disease', (72, 80)) ('mafosfamide', 'Chemical', 'MESH:C048341', (90, 101)) ('p53EE', 'Var', (31, 36)) ('influences', 'Reg', (37, 47)) ('lymphoma', 'Disease', 'MESH:D008223', (72, 80)) ('lymphoma', 'Phenotype', 'HP:0002665', (72, 80)) 1394 31483066 We established lymphoma cell lines from Emu-Myc mice with different p53 genotypes (p53+/+, p53+/-, and p53+/EE). ('lymphoma', 'Phenotype', 'HP:0002665', (15, 23)) ('Myc', 'Gene', (44, 47)) ('Myc', 'Gene', '17869', (44, 47)) ('p53+/+', 'Var', (83, 89)) ('mice', 'Species', '10090', (48, 52)) ('p53+/EE', 'Var', (103, 110)) ('p53+/-', 'Var', (91, 97)) ('lymphoma', 'Disease', (15, 23)) ('lymphoma', 'Disease', 'MESH:D008223', (15, 23)) 1395 31483066 Lymphomas from p53+/EE mice (and p53+/- mice) showed the expected loss of the wild-type allele and strongly expressed p53EE as described above (Fig 6F). ('p53EE', 'Var', (118, 123)) ('loss', 'NegReg', (66, 70)) ('Lymphomas', 'Disease', 'MESH:D008223', (0, 9)) ('Lymphomas', 'Phenotype', 'HP:0002665', (0, 9)) ('Lymphomas', 'Disease', (0, 9)) ('mice', 'Species', '10090', (40, 44)) ('mice', 'Species', '10090', (23, 27)) 1396 31483066 In comparison with the rapid and strong response of p53+/+ lymphoma cells to MAF treatment, induction of apoptosis started in p53EE cells only after 6 h and gradually increased up to significantly elevated levels of 60-70% at 24 h, whereas p53-null lymphoma cells showed no response (Fig 7A). ('lymphoma', 'Disease', 'MESH:D008223', (249, 257)) ('elevated', 'PosReg', (197, 205)) ('lymphoma', 'Phenotype', 'HP:0002665', (249, 257)) ('p53EE', 'Var', (126, 131)) ('lymphoma', 'Disease', (59, 67)) ('lymphoma', 'Disease', 'MESH:D008223', (59, 67)) ('increased', 'PosReg', (167, 176)) ('lymphoma', 'Phenotype', 'HP:0002665', (59, 67)) ('lymphoma', 'Disease', (249, 257)) 1397 31483066 To test whether the increased sensitivity of p53EE lymphoma cells is directly p53EE-mediated, the lymphoma cells were transduced with a Tet-inducible, red fluorescence protein (RFP)-coupled shRNA to knockdown p53EE expression. ('lymphoma', 'Disease', (98, 106)) ('p53EE', 'Gene', (209, 214)) ('lymphoma', 'Disease', 'MESH:D008223', (98, 106)) ('lymphoma', 'Phenotype', 'HP:0002665', (98, 106)) ('lymphoma', 'Disease', (51, 59)) ('RFP', 'Gene', (177, 180)) ('lymphoma', 'Disease', 'MESH:D008223', (51, 59)) ('lymphoma', 'Phenotype', 'HP:0002665', (51, 59)) ('knockdown', 'Var', (199, 208)) ('RFP', 'Gene', '19720', (177, 180)) 1398 31483066 RFP-positive, i.e., p53 shRNA-expressing, lymphoma cells became significantly enriched under MAF treatment in a dose-dependent manner, identifying the enhanced cytotoxic response of p53EE lymphoma cells as directly p53EE-mediated (Fig 7B). ('lymphoma', 'Disease', (188, 196)) ('RFP', 'Gene', (0, 3)) ('enhanced', 'PosReg', (151, 159)) ('lymphoma', 'Disease', 'MESH:D008223', (188, 196)) ('lymphoma', 'Disease', (42, 50)) ('cytotoxic response', 'CPA', (160, 178)) ('lymphoma', 'Phenotype', 'HP:0002665', (188, 196)) ('p53EE', 'Var', (182, 187)) ('RFP', 'Gene', '19720', (0, 3)) ('lymphoma', 'Disease', 'MESH:D008223', (42, 50)) ('lymphoma', 'Phenotype', 'HP:0002665', (42, 50)) 1402 31483066 In line with previous studies (Schmitt et al, 1999, 2002b), p53-/- lymphomas rapidly relapsed in all mice, which resulted in a very modest median survival benefit of 19 days. ('lymphoma', 'Phenotype', 'HP:0002665', (67, 75)) ('p53-/-', 'Var', (60, 66)) ('lymphomas', 'Disease', (67, 76)) ('mice', 'Species', '10090', (101, 105)) ('lymphomas', 'Disease', 'MESH:D008223', (67, 76)) ('lymphomas', 'Phenotype', 'HP:0002665', (67, 76)) 1404 31483066 Importantly, the chemotherapy was also effective for treatment of p53EE lymphoma, provided a median survival benefit of 30 days, and:even more compelling:yielded a complete tumor-free remission in 36% of animals (Fig 7D and E). ('tumor', 'Disease', 'MESH:D009369', (173, 178)) ('tumor', 'Phenotype', 'HP:0002664', (173, 178)) ('tumor', 'Disease', (173, 178)) ('lymphoma', 'Disease', (72, 80)) ('p53EE', 'Var', (66, 71)) ('lymphoma', 'Disease', 'MESH:D008223', (72, 80)) ('lymphoma', 'Phenotype', 'HP:0002665', (72, 80)) 1407 31483066 In line with the delayed apoptotic response of p53EE lymphoma cells in vitro (Fig 7A), we observed only a slight, but significant, decline in tumor load at 24 h in p53EE compared to p53-/- lymphoma mice. ('lymphoma', 'Disease', (53, 61)) ('lymphoma', 'Disease', 'MESH:D008223', (53, 61)) ('tumor', 'Phenotype', 'HP:0002664', (142, 147)) ('lymphoma', 'Phenotype', 'HP:0002665', (53, 61)) ('tumor', 'Disease', (142, 147)) ('lymphoma', 'Disease', (189, 197)) ('lymphoma', 'Disease', 'MESH:D008223', (189, 197)) ('decline', 'NegReg', (131, 138)) ('p53EE', 'Var', (164, 169)) ('mice', 'Species', '10090', (198, 202)) ('lymphoma', 'Phenotype', 'HP:0002665', (189, 197)) ('tumor', 'Disease', 'MESH:D009369', (142, 147)) 1409 31483066 To validate in a second independent model that p53EE enhances the chemotherapy response in vivo, we transplanted syngeneic recipient mice with AE9/Nras-driven acute myeloid leukemia cells with different p53 genotypes and monitored disease progression by bioluminescence imaging (Fig 7G). ('mice', 'Species', '10090', (133, 137)) ('p53EE', 'Var', (47, 52)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (159, 181)) ('enhances', 'PosReg', (53, 61)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (165, 181)) ('chemotherapy response', 'CPA', (66, 87)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (159, 181)) ('leukemia', 'Phenotype', 'HP:0001909', (173, 181)) ('acute myeloid leukemia', 'Disease', (159, 181)) 1410 31483066 p53+/+ leukemia cells responded well to a standard chemotherapy protocol with cytarabine (AraC) combined with doxorubicin, which yielded a significant survival benefit (Fig 7G and H). ('p53+/+', 'Var', (0, 6)) ('survival', 'CPA', (151, 159)) ('cytarabine', 'Chemical', 'MESH:D003561', (78, 88)) ('leukemia', 'Disease', (7, 15)) ('AraC', 'Chemical', 'MESH:D003561', (90, 94)) ('leukemia', 'Disease', 'MESH:D007938', (7, 15)) ('leukemia', 'Phenotype', 'HP:0001909', (7, 15)) ('benefit', 'PosReg', (160, 167)) ('doxorubicin', 'Chemical', 'MESH:D004317', (110, 121)) 1411 31483066 p53-/- leukemia rapidly killed the transplanted animals irrespective of therapy. ('p53-/-', 'Var', (0, 6)) ('leukemia', 'Disease', 'MESH:D007938', (7, 15)) ('leukemia', 'Disease', (7, 15)) ('leukemia', 'Phenotype', 'HP:0001909', (7, 15)) 1412 31483066 In striking contrast, chemotherapy controlled progression of p53EE/EE leukemia remarkably well, which translated into a significantly improved survival (Fig 7G and H). ('improved', 'PosReg', (134, 142)) ('leukemia', 'Disease', (70, 78)) ('leukemia', 'Phenotype', 'HP:0001909', (70, 78)) ('survival', 'MPA', (143, 151)) ('leukemia', 'Disease', 'MESH:D007938', (70, 78)) ('p53EE/EE', 'Var', (61, 69)) 1413 31483066 Our results obtained with the R178E cooperativity mutation in different mouse cancer models illustrate that a p53 mutant can be as inefficient as a p53 knock-out allele in preventing tumor development and yet retain residual apoptotic activity. ('R178E', 'Mutation', 'p.R178E', (30, 35)) ('p53', 'Gene', (110, 113)) ('tumor', 'Disease', 'MESH:D009369', (183, 188)) ('mouse', 'Species', '10090', (72, 77)) ('apoptotic activity', 'CPA', (225, 243)) ('R178E', 'Var', (30, 35)) ('cancer', 'Disease', (78, 84)) ('mutant', 'Var', (114, 120)) ('cancer', 'Disease', 'MESH:D009369', (78, 84)) ('tumor', 'Phenotype', 'HP:0002664', (183, 188)) ('tumor', 'Disease', (183, 188)) ('cancer', 'Phenotype', 'HP:0002664', (78, 84)) 1414 31483066 Taken together, this indicates that residual apoptotic p53 functions on their own are unable to counteract tumorigenesis, are therefore not efficiently counter-selected, and can be retained by mutant p53 during tumor evolution. ('tumor', 'Phenotype', 'HP:0002664', (107, 112)) ('tumor', 'Disease', (107, 112)) ('tumor', 'Disease', 'MESH:D009369', (211, 216)) ('p53', 'Gene', (200, 203)) ('mutant', 'Var', (193, 199)) ('tumor', 'Phenotype', 'HP:0002664', (211, 216)) ('tumor', 'Disease', (211, 216)) ('tumor', 'Disease', 'MESH:D009369', (107, 112)) 1415 31483066 Superior chemotherapy responses in p53-mutated versus p53-deficient tumors challenge the current view that a p53 missense mutation invariably signals a worse prognosis than a gene deletion. ('p53-deficient tumors', 'Disease', 'MESH:D009369', (54, 74)) ('tumors', 'Phenotype', 'HP:0002664', (68, 74)) ('p53-deficient tumors', 'Disease', (54, 74)) ('p53', 'Gene', (109, 112)) ('missense mutation', 'Var', (113, 130)) ('tumor', 'Phenotype', 'HP:0002664', (68, 73)) 1416 31483066 p53WT is capable of inducing cell death through multiple pathways including apoptosis, necrosis, and ferroptosis. ('inducing', 'PosReg', (20, 28)) ('necrosis', 'Disease', (87, 95)) ('p53WT', 'Var', (0, 5)) ('ferroptosis', 'Disease', (101, 112)) ('necrosis', 'Disease', 'MESH:D009336', (87, 95)) ('apoptosis', 'CPA', (76, 85)) ('cell death', 'CPA', (29, 39)) 1417 31483066 The cytotoxic activity of p53EE is apoptotic in nature as shown by caspase cleavage (Figs 3E and 4F and H), phosphatidylserine externalization (Figs 4A and 5C and H), and blockage by caspase inhibitors, but not ferroptosis inhibitors (Fig 5B). ('5C', 'Chemical', 'MESH:D002244', (156, 158)) ('cytotoxic', 'CPA', (4, 13)) ('p53EE', 'Var', (26, 31)) ('4F', 'Chemical', 'MESH:C006011', (97, 99)) ('phosphatidylserine', 'Chemical', 'MESH:D010718', (108, 126)) ('apoptotic', 'CPA', (35, 44)) ('caspase cleavage', 'CPA', (67, 83)) ('phosphatidylserine externalization', 'MPA', (108, 142)) 1418 31483066 p53WT induces apoptosis by transcriptional upregulation of pro-apoptotic target genes such as PUMA, NOXA, and BAX in combination with non-transcriptional mechanisms mediated through protein-protein interactions with Bcl-2 family members, which lowers the threshold for engaging the mitochondrial apoptosis pathway (Mihara et al, 2003; Chipuk et al, 2004; Leu et al, 2004; Le Pen et al, 2016). ('mitochondrial apoptosis pathway', 'Pathway', (282, 313)) ('Leu', 'Chemical', 'MESH:C038361', (355, 358)) ('apoptosis', 'Gene', (14, 23)) ('p53WT', 'Var', (0, 5)) ('BAX', 'Gene', (110, 113)) ('upregulation', 'PosReg', (43, 55)) ('Bcl-2', 'Gene', (216, 221)) ('BAX', 'Gene', '12028', (110, 113)) ('Bcl-2', 'Gene', '12043', (216, 221)) 1419 31483066 The lack of detectable DNA binding and target gene regulation by p53EE suggested that its pro-apoptotic activity relies primarily on the non-transcriptional pathway, and we demonstrate that p53EE efficiently localizes to the mitochondria (Figs 4I and 5E and F), interacts with Bcl-2 family members (Fig 5F), and primes cells to mitochondrial depolarization by BH3 peptides (Fig 5G). ('p53EE', 'Var', (190, 195)) ('primes', 'PosReg', (312, 318)) ('p53EE', 'Var', (65, 70)) ('localizes', 'MPA', (208, 217)) ('mitochondrial depolarization', 'MPA', (328, 356)) ('Bcl-2', 'Gene', (277, 282)) ('interacts', 'Interaction', (262, 271)) ('Bcl-2', 'Gene', '12043', (277, 282)) 1420 31483066 In addition, we provide evidence that p53EE, similar to other p53 mutants, interferes with Nrf2 transcriptional activity, which is critical not only for ROS defense, but also for the respiratory function of mitochondria, their biogenesis, and integrity (Dinkova-Kostova & Abramov, 2015). ('transcriptional activity', 'MPA', (96, 120)) ('Nrf2', 'Gene', (91, 95)) ('p53EE', 'Var', (38, 43)) ('interferes', 'NegReg', (75, 85)) ('Nrf2', 'Gene', '18024', (91, 95)) 1424 31483066 However, deregulated p53 can cause embryonic lethality by multiple pathways including apoptosis, ferroptosis, or cell growth arrest (Jiang et al, 2015; Moyer et al, 2017). ('deregulated', 'Var', (9, 20)) ('cell growth arrest', 'CPA', (113, 131)) ('cause', 'Reg', (29, 34)) ('ferroptosis', 'CPA', (97, 108)) ('embryonic lethality', 'Disease', 'MESH:D020964', (35, 54)) ('growth arrest', 'Phenotype', 'HP:0001510', (118, 131)) ('embryonic lethality', 'Disease', (35, 54)) ('apoptosis', 'CPA', (86, 95)) ('p53', 'Gene', (21, 24)) 1425 31483066 In general, low p53 levels or reduced p53 DNA binding cooperativity primarily activates homeostatic survival functions, while only strongly elevated and prolonged expression levels shift the cellular response to apoptosis (Chen et al, 1996; Vousden & Lu, 2002; Vousden & Lane, 2007; Schlereth et al, 2013; Timofeev et al, 2013). ('low', 'Var', (12, 15)) ('p53 levels', 'MPA', (16, 26)) ('Lu', 'Chemical', 'MESH:D008187', (251, 253)) ('shift', 'Reg', (181, 186)) ('activates', 'PosReg', (78, 87)) ('p53 DNA', 'Protein', (38, 45)) ('reduced', 'NegReg', (30, 37)) ('homeostatic survival functions', 'MPA', (88, 118)) 1426 31483066 Last but not least, the charge inversion associated with the arginine to glutamic acid substitution in p53EE might affect protein-protein interactions that are responsible for its pro-apoptotic activity. ('protein-protein', 'Protein', (122, 137)) ('glutamic acid', 'Chemical', 'MESH:C030030', (73, 86)) ('affect', 'Reg', (115, 121)) ('arginine', 'Chemical', 'MESH:D001127', (61, 69)) ('charge inversion', 'Var', (24, 40)) ('p53EE', 'Gene', (103, 108)) ('arginine to glutamic acid', 'Var', (61, 86)) 1428 31483066 From a clinical perspective, it will be interesting to investigate which p53 mutants in cancer patients display residual apoptotic activity, as p53EE is not a naturally occurring mutation. ('cancer', 'Disease', (88, 94)) ('cancer', 'Disease', 'MESH:D009369', (88, 94)) ('patients', 'Species', '9606', (95, 103)) ('p53', 'Gene', (73, 76)) ('mutants', 'Var', (77, 84)) ('cancer', 'Phenotype', 'HP:0002664', (88, 94)) ('apoptotic activity', 'CPA', (121, 139)) 1429 31483066 Since protein-protein interactions with Bcl-2 family members that account for non-transcriptional apoptosis by p53WT (Mihara et al, 2003; Chipuk et al, 2004; Leu et al, 2004; Le Pen et al, 2016) are disrupted by many cancer-derived p53 mutations, such mutations are believed to be "dual hits" which simultaneously inactivate both DNA binding-dependent and non-transcriptional mechanisms of p53-triggered apoptosis (Mihara et al, 2003; Tomita et al, 2006). ('disrupted', 'Reg', (199, 208)) ('p53', 'Gene', (232, 235)) ('mutations', 'Var', (252, 261)) ('cancer', 'Disease', (217, 223)) ('mutations', 'Var', (236, 245)) ('Bcl-2', 'Gene', (40, 45)) ('inactivate', 'NegReg', (314, 324)) ('cancer', 'Disease', 'MESH:D009369', (217, 223)) ('Leu', 'Chemical', 'MESH:C038361', (158, 161)) ('Bcl-2', 'Gene', '12043', (40, 45)) ('cancer', 'Phenotype', 'HP:0002664', (217, 223)) ('protein-protein interactions', 'Protein', (6, 34)) 1430 31483066 However, this has only been shown for a small subset of mainly hot-spot mutants and cannot be directly extrapolated to the entire, functionally and structurally highly diverse, spectrum of > 2,000 distinct p53 mutations observed in cancer patients. ('p53', 'Gene', (206, 209)) ('mutants', 'Var', (72, 79)) ('cancer', 'Disease', 'MESH:D009369', (232, 238)) ('mutations', 'Var', (210, 219)) ('cancer', 'Disease', (232, 238)) ('patients', 'Species', '9606', (239, 247)) ('cancer', 'Phenotype', 'HP:0002664', (232, 238)) 1432 31483066 While the most common hot-spot mutants are thermodynamically destabilized and therefore structurally denatured, many non-hot-spot mutants retain the ability to regulate some bona fide p53 target genes like p21 (Ludwig et al, 1996; Rowan et al, 1996; Campomenosi et al, 2001). ('regulate', 'Reg', (160, 168)) ('p21', 'Gene', (206, 209)) ('mutants', 'Var', (31, 38)) ('p21', 'Gene', '12575', (206, 209)) ('Lu', 'Chemical', 'MESH:D008187', (211, 213)) ('mutants', 'Var', (130, 137)) ('Rowan', 'Species', '36599', (231, 236)) 1433 31483066 Several cooperativity mutants that together account for an estimated 34,000 cancer cases per year (Leroy et al, 2014) have selectively lost the ability to transactivate pro-apoptotic target genes (Ludwig et al, 1996; Schlereth et al, 2010a). ('transactivate', 'MPA', (155, 168)) ('Lu', 'Chemical', 'MESH:D008187', (197, 199)) ('cancer', 'Phenotype', 'HP:0002664', (76, 82)) ('mutants', 'Var', (22, 29)) ('lost', 'NegReg', (135, 139)) ('cancer', 'Disease', (76, 82)) ('cancer', 'Disease', 'MESH:D009369', (76, 82)) 1434 31483066 Here, we have examined the R181L mutant that has been identified as both a somatic and germline mutation in cancer patients. ('patients', 'Species', '9606', (115, 123)) ('cancer', 'Phenotype', 'HP:0002664', (108, 114)) ('R181L', 'Var', (27, 32)) ('R181L', 'Mutation', 'p.R181L', (27, 32)) ('cancer', 'Disease', (108, 114)) ('cancer', 'Disease', 'MESH:D009369', (108, 114)) 1435 31483066 Consistent with previous reports that R181L has a selective apoptosis defect (Ludwig et al, 1996; Schlereth et al, 2010a), we failed to detect apoptosis upon R181L overexpression (Fig 5H). ('defect', 'NegReg', (70, 76)) ('apoptosis', 'CPA', (60, 69)) ('R181L', 'Var', (38, 43)) ('R181L', 'Var', (158, 163)) ('R181L', 'Mutation', 'p.R181L', (38, 43)) ('R181L', 'Mutation', 'p.R181L', (158, 163)) ('Lu', 'Chemical', 'MESH:D008187', (78, 80)) 1436 31483066 However, just like with p53EE, genotoxic doxorubicin treatment revealed substantial pro-apoptotic activity, supporting the concept that a tumorigenic p53 mutant like R181L can retain residual apoptotic functions and support chemotherapy responses. ('apoptotic functions', 'CPA', (192, 211)) ('tumor', 'Disease', (138, 143)) ('p53', 'Gene', (150, 153)) ('pro-apoptotic', 'MPA', (84, 97)) ('doxorubicin', 'Chemical', 'MESH:D004317', (41, 52)) ('support', 'PosReg', (216, 223)) ('R181L', 'Mutation', 'p.R181L', (166, 171)) ('tumor', 'Disease', 'MESH:D009369', (138, 143)) ('R181L', 'Var', (166, 171)) ('chemotherapy responses', 'CPA', (224, 246)) ('tumor', 'Phenotype', 'HP:0002664', (138, 143)) 1437 31483066 Intriguingly, the R175P mutant, which resembles the R181L mutant in its selective loss of pro-apoptotic target gene regulation (Ludwig et al, 1996; Rowan et al, 1996; Liu et al, 2004), failed to enhance apoptosis under identical conditions of doxorubicin treatment (Fig 5H). ('pro-apoptotic target gene regulation', 'MPA', (90, 126)) ('enhance', 'PosReg', (195, 202)) ('Lu', 'Chemical', 'MESH:D008187', (128, 130)) ('doxorubicin', 'Chemical', 'MESH:D004317', (243, 254)) ('apoptosis', 'CPA', (203, 212)) ('Rowan', 'Species', '36599', (148, 153)) ('R175P', 'Var', (18, 23)) ('R175P', 'Mutation', 'p.R175P', (18, 23)) ('loss', 'NegReg', (82, 86)) ('R181L', 'Mutation', 'p.R181L', (52, 57)) 1438 31483066 A possible explanation is that R175 is crucial for the structural integrity of the L2-L3 loop which interacts not only with the minor groove of DNA response elements but also with Bcl-2 family members (Cho et al, 1994; Bullock et al, 2000; Mihara et al, 2003; Tomita et al, 2006; Hagn et al, 2010). ('Bcl-2', 'Gene', (180, 185)) ('R175', 'Var', (31, 35)) ('Bcl-2', 'Gene', '12043', (180, 185)) 1439 31483066 It is therefore tempting to speculate that in particular, cooperativity mutants of the H1 helix, which interfere with DNA binding but do not affect the interaction interface with DNA and Bcl-2 family members, have retained non-transcriptional apoptotic activity. ('non-transcriptional apoptotic activity', 'CPA', (223, 261)) ('mutants', 'Var', (72, 79)) ('binding', 'Interaction', (122, 129)) ('DNA', 'Protein', (118, 121)) ('Bcl-2', 'Gene', (187, 192)) ('Bcl-2', 'Gene', '12043', (187, 192)) 1440 31483066 A more detailed systematic analysis of the more than 2,000 different cancer mutants is needed to validate this hypothesis and possibly reveal further cancer mutants with a similar phenotype. ('cancer', 'Phenotype', 'HP:0002664', (150, 156)) ('cancer', 'Phenotype', 'HP:0002664', (69, 75)) ('cancer', 'Disease', 'MESH:D009369', (150, 156)) ('mutants', 'Var', (76, 83)) ('cancer', 'Disease', (150, 156)) ('cancer', 'Disease', 'MESH:D009369', (69, 75)) ('cancer', 'Disease', (69, 75)) 1441 31483066 Recent studies have revealed that tumors can be addicted to pro-survival GOF activities of mutant p53 and respond to mutant p53 depletion with tumor regression (Alexandrova et al, 2015). ('tumor', 'Disease', 'MESH:D009369', (143, 148)) ('tumors', 'Disease', (34, 40)) ('p53', 'Gene', (124, 127)) ('tumor', 'Disease', (34, 39)) ('tumor', 'Disease', (143, 148)) ('depletion', 'MPA', (128, 137)) ('tumors', 'Disease', 'MESH:D009369', (34, 40)) ('mutant', 'Var', (117, 123)) ('tumors', 'Phenotype', 'HP:0002664', (34, 40)) ('GOF', 'PosReg', (73, 76)) ('p53', 'Gene', (98, 101)) ('tumor', 'Disease', 'MESH:D009369', (34, 39)) ('activities', 'MPA', (77, 87)) ('mutant', 'Var', (91, 97)) ('tumor', 'Phenotype', 'HP:0002664', (34, 39)) ('tumor', 'Phenotype', 'HP:0002664', (143, 148)) 1442 31483066 Mutant p53-destabilizing drugs, such as the Hsp90 inhibitor ganetespib, are therefore considered a promising approach for treatment of p53-mutated tumors, and a first clinical trial combining chemotherapy with ganetespib for p53-mutated ovarian cancer patients has been initiated (Alexandrova et al, 2015; Bykov et al, 2018; Sabapathy & Lane, 2018). ('tumor', 'Phenotype', 'HP:0002664', (147, 152)) ('ovarian cancer', 'Phenotype', 'HP:0100615', (237, 251)) ('tumors', 'Disease', (147, 153)) ('ganetespib', 'Chemical', 'MESH:C533237', (60, 70)) ('patients', 'Species', '9606', (252, 260)) ('tumors', 'Phenotype', 'HP:0002664', (147, 153)) ('tumors', 'Disease', 'MESH:D009369', (147, 153)) ('ovarian cancer', 'Disease', 'MESH:D010051', (237, 251)) ('ganetespib', 'Chemical', 'MESH:C533237', (210, 220)) ('ovarian cancer', 'Disease', (237, 251)) ('Mutant', 'Var', (0, 6)) ('cancer', 'Phenotype', 'HP:0002664', (245, 251)) 1443 31483066 However, mouse models for the R246S (equivalent to human R249S) and humanized G245S mutants have shown that p53 mutations can be highly tumorigenic without exhibiting appreciable GOF activity (Lee et al, 2012; Hanel et al, 2013). ('human', 'Species', '9606', (68, 73)) ('p53', 'Gene', (108, 111)) ('mouse', 'Species', '10090', (9, 14)) ('R249S', 'Mutation', 'p.R249S', (57, 62)) ('tumor', 'Disease', 'MESH:D009369', (136, 141)) ('mutations', 'Var', (112, 121)) ('human', 'Species', '9606', (51, 56)) ('tumor', 'Phenotype', 'HP:0002664', (136, 141)) ('G245S', 'Mutation', 'p.G245S', (78, 83)) ('R246S', 'Mutation', 'p.R246S', (30, 35)) ('R246S', 'Var', (30, 35)) ('tumor', 'Disease', (136, 141)) 1444 31483066 The p53EE mutant does not significantly decrease the latency of tumor development or enhance the incidence of metastatic tumors compared to p53 knock-out mice (Fig 6A; Appendix Tables S1 and S2). ('decrease', 'NegReg', (40, 48)) ('tumor', 'Disease', (64, 69)) ('mice', 'Species', '10090', (154, 158)) ('tumor', 'Phenotype', 'HP:0002664', (121, 126)) ('tumors', 'Disease', (121, 127)) ('tumors', 'Phenotype', 'HP:0002664', (121, 127)) ('tumor', 'Disease', (121, 126)) ('p53EE', 'Var', (4, 9)) ('tumor', 'Disease', 'MESH:D009369', (121, 126)) ('tumors', 'Disease', 'MESH:D009369', (121, 127)) ('tumor', 'Disease', 'MESH:D009369', (64, 69)) ('tumor', 'Phenotype', 'HP:0002664', (64, 69)) ('enhance', 'PosReg', (85, 92)) 1445 31483066 There was a trend for increased incidence of non-lymphoid tumors in homozygous p53EE mice, but this was not statistically significant (Fig 6B), leading us to conclude that p53EE does not exhibit the strong GOF activities observed for some hot-spot mutants. ('mice', 'Species', '10090', (85, 89)) ('tumor', 'Phenotype', 'HP:0002664', (58, 63)) ('lymphoid tumors', 'Phenotype', 'HP:0002665', (49, 64)) ('tumors', 'Phenotype', 'HP:0002664', (58, 64)) ('non-lymphoid tumors', 'Disease', (45, 64)) ('p53EE', 'Var', (79, 84)) ('p53EE', 'Var', (172, 177)) ('increased', 'PosReg', (22, 31)) ('non-lymphoid tumors', 'Disease', 'MESH:D008228', (45, 64)) 1446 31483066 When tumors are not dependent on a mutant p53 GOF, its destabilization is expected to be therapeutically ineffective. ('p53', 'Gene', (42, 45)) ('tumors', 'Disease', (5, 11)) ('tumors', 'Disease', 'MESH:D009369', (5, 11)) ('mutant', 'Var', (35, 41)) ('tumor', 'Phenotype', 'HP:0002664', (5, 10)) ('tumors', 'Phenotype', 'HP:0002664', (5, 11)) 1447 31483066 In support of this, degradation of p53EE by ganetespib did not result in tumor cell death by itself and effectively counteracted the pro-apoptotic effect of combined doxorubicin and Nutlin treatment (Fig 4H). ('doxorubicin', 'Chemical', 'MESH:D004317', (166, 177)) ('tumor', 'Disease', 'MESH:D009369', (73, 78)) ('Nutlin', 'Chemical', 'MESH:C482204', (182, 188)) ('ganetespib', 'Chemical', 'MESH:C533237', (44, 54)) ('tumor', 'Phenotype', 'HP:0002664', (73, 78)) ('tumor', 'Disease', (73, 78)) ('p53EE', 'Var', (35, 40)) ('ganetespib', 'Gene', (44, 54)) 1448 31483066 Instead, our data suggest that tumors which contain a p53 mutant with residual apoptotic activity might require the opposite strategy. ('mutant', 'Var', (58, 64)) ('tumor', 'Phenotype', 'HP:0002664', (31, 36)) ('tumors', 'Phenotype', 'HP:0002664', (31, 37)) ('tumors', 'Disease', (31, 37)) ('p53', 'Gene', (54, 57)) ('tumors', 'Disease', 'MESH:D009369', (31, 37)) 1449 31483066 Chemotherapy responses of p53EE cells were enhanced in combination with Nutlin-3a or other Mdm2 inhibitors (Fig 4D and E). ('enhanced', 'PosReg', (43, 51)) ('Chemotherapy responses', 'CPA', (0, 22)) ('Nutlin', 'Chemical', 'MESH:C482204', (72, 78)) ('p53EE', 'Var', (26, 31)) 1450 31483066 Enforced overexpression of the human p53EE (R181E) or the cancer mutant R181L strongly sensitized human lung cancer cells to chemotherapy-induced apoptosis, and this was further boosted by Nutlin-3a (Fig 5H). ('lung cancer', 'Disease', 'MESH:D008175', (104, 115)) ('R181E', 'Mutation', 'p.R181E', (44, 49)) ('overexpression', 'PosReg', (9, 23)) ('cancer', 'Phenotype', 'HP:0002664', (58, 64)) ('cancer', 'Disease', 'MESH:D009369', (109, 115)) ('cancer', 'Disease', (109, 115)) ('lung cancer', 'Disease', (104, 115)) ('human', 'Species', '9606', (31, 36)) ('sensitized', 'Reg', (87, 97)) ('Nutlin', 'Chemical', 'MESH:C482204', (189, 195)) ('cancer', 'Disease', 'MESH:D009369', (58, 64)) ('lung cancer', 'Phenotype', 'HP:0100526', (104, 115)) ('R181L', 'Var', (72, 77)) ('cancer', 'Disease', (58, 64)) ('cancer', 'Phenotype', 'HP:0002664', (109, 115)) ('R181L', 'Mutation', 'p.R181L', (72, 77)) ('human', 'Species', '9606', (98, 103)) ('chemotherapy-induced apoptosis', 'CPA', (125, 155)) 1452 31483066 Mutant p53-stabilizing therapies with Mdm2 inhibitors might therefore support chemotherapeutics to trigger the residual cytotoxic activities that have not been efficiently counter-selected during tumorigenesis. ('tumor', 'Disease', (196, 201)) ('Mdm2', 'Gene', (38, 42)) ('tumor', 'Disease', 'MESH:D009369', (196, 201)) ('Mutant', 'Var', (0, 6)) ('p53-stabilizing', 'Gene', (7, 22)) ('tumor', 'Phenotype', 'HP:0002664', (196, 201)) 1453 31483066 In fact, the first clinical studies with Mdm2 inhibitors have observed clinical responses also in a few patients with p53 mutations (Andreeff et al, 2016). ('p53', 'Gene', (118, 121)) ('mutations', 'Var', (122, 131)) ('patients', 'Species', '9606', (104, 112)) 1456 31483066 Although it remains to be seen, which and how many p53 mutants have retained apoptotic activity during tumor evolution, our data call for more comprehensive investigations into the functional diversity of p53 mutations to make p53 mutation status more useful for clinical decision making. ('p53', 'Gene', (51, 54)) ('tumor', 'Phenotype', 'HP:0002664', (103, 108)) ('mutants', 'Var', (55, 62)) ('tumor', 'Disease', (103, 108)) ('p53', 'Gene', (205, 208)) ('apoptotic activity', 'CPA', (77, 95)) ('tumor', 'Disease', 'MESH:D009369', (103, 108)) 1457 31483066 For generation of the Trp53 LSL-R178E knock-in mouse, we used the targeting vector for the Trp53 LSL-R172H mouse, which was kindly provided by Tyler Jacks (Olive et al, 2004). ('R178E', 'Mutation', 'p.R178E', (32, 37)) ('mouse', 'Species', '10090', (107, 112)) ('mouse', 'Species', '10090', (47, 52)) ('Trp53', 'Gene', (91, 96)) ('LSL-R172H', 'Var', (97, 106)) ('Olive', 'Species', '4146', (156, 161)) ('R172H', 'Mutation', 'p.R172H', (101, 106)) 1458 31483066 The vector was modified by QuikChange Multi Site-Directed Mutagenesis (Stratagene) to carry only the mutation GCG->CTC in exon 5 of Trp53, resulting in a Arg->Glu substitution at codon 178 (Fig EV1A). ('in a', 'Reg', (149, 153)) ('Arg->Glu substitution at codon 178', 'Mutation', 'p.R178E', (154, 188)) ('mutation GCG->CTC', 'Var', (101, 118)) ('GCG->CTC', 'Var', (110, 118)) ('Trp53', 'Gene', (132, 137)) ('Arg->Glu', 'Var', (154, 162)) 1463 31483066 In 129S/Sv-Trp53 LSL-R178E mice, expression of the Trp53 gene is blocked by a transcriptional stop cassette, flanked by loxP sites (lox-stop-lox, LSL). ('LSL-R178E', 'Var', (17, 26)) ('mice', 'Species', '10090', (27, 31)) ('Trp53 gene', 'Gene', (51, 61)) ('R178E', 'Mutation', 'p.R178E', (21, 26)) ('expression', 'MPA', (33, 43)) ('blocked', 'NegReg', (65, 72)) 1464 31483066 The homozygous Trp53 LSL-R178E/LSL-R178E mice lack p53 protein expression. ('R178E', 'SUBSTITUTION', 'None', (35, 40)) ('R178E', 'SUBSTITUTION', 'None', (25, 30)) ('mice', 'Species', '10090', (41, 45)) ('protein', 'Protein', (55, 62)) ('p53 protein', 'Protein', (51, 62)) ('R178E', 'Mutation', 'p.R178E', (25, 30)) ('lack', 'NegReg', (46, 50)) ('R178E', 'Var', (25, 30)) ('Trp53', 'Gene', (15, 20)) ('R178E', 'Mutation', 'p.R178E', (35, 40)) ('R178E', 'Var', (35, 40)) 1465 31483066 Trp53 LSL-R178E/LSL-R178E mice, and embryonic fibroblasts isolated from these mice, were used as isogenic p53-null (Trp53 -/-) controls in all experiments. ('R178E', 'Mutation', 'p.R178E', (20, 25)) ('R178E', 'Mutation', 'p.R178E', (10, 15)) ('R178E', 'Var', (20, 25)) ('R178E', 'Var', (10, 15)) ('mice', 'Species', '10090', (26, 30)) ('R178E', 'SUBSTITUTION', 'None', (20, 25)) ('R178E', 'SUBSTITUTION', 'None', (10, 15)) ('mice', 'Species', '10090', (78, 82)) ('Trp53', 'Gene', (0, 5)) 1470 31483066 To study lethality of Mdm2 knock-out, we used heterozygous Mdm2 Delta7-9 (Mdm2) mice that lack exons 7-9. ('Delta7', 'Mutation', 'c.del7', (64, 70)) ('Mdm2', 'Gene', (59, 63)) ('mice', 'Species', '10090', (90, 94)) ('Delta7-9', 'Var', (64, 72)) 1475 31483066 Primary lymphomas were obtained from Emu-Myc mice with p53+/+, p53+/-, and p53+/EE genotypes, and 500,000 cells were transplanted into syngeneic recipients via tail vein as described in the literature (Schmitt et al, 2002a). ('mice', 'Species', '10090', (45, 49)) ('p53+/+', 'Var', (55, 61)) ('p53+/-', 'Var', (63, 69)) ('Primary lymphomas', 'Disease', 'MESH:D008223', (0, 17)) ('Myc', 'Gene', (41, 44)) ('Myc', 'Gene', '17869', (41, 44)) ('lymphomas', 'Phenotype', 'HP:0002665', (8, 17)) ('Primary lymphomas', 'Disease', (0, 17)) ('p53+/EE', 'Var', (75, 82)) ('lymphoma', 'Phenotype', 'HP:0002665', (8, 16)) 1492 31483066 Nutlin-3a (Sigma) was used at 10 muM, RG7112 (MedChemExpress) at 5 muM, RG7388 (MedChemExpress) at 8 muM, MI773 (Selleckchem) at 10 muM, and Mafosfamide (Santa Cruz) at 1-5 mug/ml as indicated. ('RG7388', 'Var', (72, 78)) ('Nutlin', 'Chemical', 'MESH:C482204', (0, 6)) ('RG7388', 'Chemical', 'MESH:C586849', (72, 78)) ('MI773', 'Var', (106, 111)) ('RG7112', 'Chemical', 'MESH:C579783', (38, 44)) ('Mafosfamide', 'Chemical', 'MESH:C048341', (141, 152)) 1495 31483066 For expression of mutant p53, the cDNA for murine or human p53 was cloned into pInducer20 using the Gateway System (Invitrogen). ('p53', 'Gene', (25, 28)) ('murine', 'Species', '10090', (43, 49)) ('mutant', 'Var', (18, 24)) ('human', 'Species', '9606', (53, 58)) 1496 31483066 MEFs were immortalized by retroviral transduction with pMSCVhygro-E1A.12S or pMSCVneo-E1A.12S (Timofeev et al, 2013). ('MEF', 'Gene', (0, 3)) ('MEF', 'Gene', '56501', (0, 3)) ('pMSCVhygro-E1A.12S', 'Var', (55, 73)) ('pMSCVneo-E1A.12S', 'Var', (77, 93)) 1503 31483066 Under these conditions, proliferation of p53EE/EE MEFs declined more rapidly than in Fig EV4A and eventually resulted in deterioration of cell cultures. ('p53EE/EE', 'Var', (41, 49)) ('MEF', 'Gene', (50, 53)) ('MEF', 'Gene', '56501', (50, 53)) ('resulted', 'Reg', (109, 117)) ('cell cultures', 'CPA', (138, 151)) ('declined', 'NegReg', (55, 63)) 1513 31483066 Other antibodies used for IHC were anti-p53 (NCL-p53-505, Leica Microsystems, 1:1,000) and anti-BrdU (BU1/75(ICR1), #OBT0030G, 1:100). ('NCL', 'Gene', '17975', (45, 48)) ('anti-BrdU', 'Var', (91, 100)) ('BU1/75', 'Var', (102, 108)) ('NCL', 'Gene', (45, 48)) ('anti-p53', 'Var', (35, 43)) 1515 31483066 The DPLA probe anti-rabbit minus binds to the p53 antibody, whereas the DPLA probe anti-mouse plus binds to the antibody against the probable interaction partner, respectively. ('rabbit', 'Species', '9986', (20, 26)) ('p53', 'Var', (46, 49)) ('binds', 'Interaction', (33, 38)) ('mouse', 'Species', '10090', (88, 93)) 1522 31483066 Emu-Myc;p53EE lymphoma cells were transduced with TtRMPVIR retroviral vectors (Zuber et al, 2011) expressing a p53-targeting (shp53.814, Dickins et al, 2005) or non-silencing control shRNA coupled to RFP. ('RFP', 'Gene', '19720', (200, 203)) ('Myc', 'Gene', (4, 7)) ('lymphoma', 'Disease', (14, 22)) ('Myc', 'Gene', '17869', (4, 7)) ('lymphoma', 'Disease', 'MESH:D008223', (14, 22)) ('lymphoma', 'Phenotype', 'HP:0002665', (14, 22)) ('RFP', 'Gene', (200, 203)) ('shp53.814', 'Var', (126, 135)) 1535 31483066 Crosslinking was reverted upon overnight incubation at 65 C in elution buffer supplemented with 200 mM NaCl followed by RNase A digestion at 37 C for 30 min, addition of 40 mM Tris-HCl pH 6.5 and 10 mM EDTA, proteinase K digestion for 2 h at 55 C, and inactivation of proteinase K at 99 C for 10 min. ('inactivation', 'Var', (252, 264)) ('NaCl', 'Chemical', 'MESH:D012965', (103, 107)) ('RNase A', 'Gene', '19752', (120, 127)) ('proteinase K', 'Enzyme', (268, 280)) ('Tris-HCl', 'Chemical', 'MESH:C014843', (176, 184)) ('EDTA', 'Chemical', 'MESH:D004492', (202, 206)) ('RNase A', 'Gene', (120, 127)) 1553 31632141 Midostaurin In Acute Myeloid Leukemia: An Evidence-Based Review And Patient Selection Fms-related-tyrosine kinase 3 (FLT3) mutations occur in approximately a third of acute myeloid leukemia (AML) patients and confer an adverse prognosis. ('Fms-related-tyrosine kinase 3', 'Gene', (86, 115)) ('Midostaurin', 'Chemical', 'MESH:C059539', (0, 11)) ('FLT3', 'Gene', (117, 121)) ('patients', 'Species', '9606', (196, 204)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (167, 189)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (173, 189)) ('FLT3', 'Gene', '2322', (117, 121)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (167, 189)) ('AML', 'Disease', 'MESH:D015470', (191, 194)) ('leukemia', 'Phenotype', 'HP:0001909', (181, 189)) ('AML', 'Disease', (191, 194)) ('Patient', 'Species', '9606', (68, 75)) ('occur', 'Reg', (133, 138)) ('Acute Myeloid Leukemia', 'Disease', 'MESH:D015470', (15, 37)) ('Myeloid Leukemia', 'Phenotype', 'HP:0012324', (21, 37)) ('Acute Myeloid Leukemia', 'Phenotype', 'HP:0004808', (15, 37)) ('Acute Myeloid Leukemia', 'Disease', (15, 37)) ('Leukemia', 'Phenotype', 'HP:0001909', (29, 37)) ('Fms-related-tyrosine kinase 3', 'Gene', '2322', (86, 115)) ('acute myeloid leukemia', 'Disease', (167, 189)) ('mutations', 'Var', (123, 132)) 1564 31632141 Traditionally, normal karyotype in AML conferred intermediate-risk disease. ('AML', 'Disease', (35, 38)) ('intermediate-risk disease', 'Disease', (49, 74)) ('conferred', 'Reg', (39, 48)) ('AML', 'Disease', 'MESH:D015470', (35, 38)) ('normal karyotype', 'Var', (15, 31)) 1565 31632141 However, with the advent of mutational analysis, it became evident that there were certain clonal mutations that could significantly alter the AML pathogenesis and prognosis. ('mutations', 'Var', (98, 107)) ('alter', 'Reg', (133, 138)) ('AML', 'Disease', (143, 146)) ('AML', 'Disease', 'MESH:D015470', (143, 146)) 1567 31632141 On the other hand, mutations in genes such as Fms-related-tyrosine kinase 3 (FLT3), runt-related transcription factor 1 (RUNX1), DNA methyl transferase 3 alpha (DNMT3A), tumor protein p53 (TP53) and others confer unfavorable risk. ('runt-related transcription factor 1', 'Gene', '861', (84, 119)) ('TP53', 'Gene', (189, 193)) ('p53', 'Gene', '7157', (184, 187)) ('runt-related transcription factor 1', 'Gene', (84, 119)) ('tumor', 'Disease', (170, 175)) ('FLT3', 'Gene', (77, 81)) ('DNMT3A', 'Gene', '1788', (161, 167)) ('tumor', 'Disease', 'MESH:D009369', (170, 175)) ('p53', 'Gene', (184, 187)) ('DNA methyl transferase 3 alpha', 'Gene', '1788', (129, 159)) ('FLT3', 'Gene', '2322', (77, 81)) ('DNA methyl transferase 3 alpha', 'Gene', (129, 159)) ('RUNX1', 'Gene', (121, 126)) ('TP53', 'Gene', '7157', (189, 193)) ('RUNX1', 'Gene', '861', (121, 126)) ('tumor', 'Phenotype', 'HP:0002664', (170, 175)) ('Fms-related-tyrosine kinase 3', 'Gene', '2322', (46, 75)) ('DNMT3A', 'Gene', (161, 167)) ('Fms-related-tyrosine kinase 3', 'Gene', (46, 75)) ('mutations', 'Var', (19, 28)) 1568 31632141 An internal tandem duplication (ITD) mutation in FLT3 gene on chromosome 13q12 is one of the most common mutations noted in AML, occurring in approximately a third of newly diagnosed adults with AML. ('internal tandem duplication', 'Var', (3, 30)) ('occurring', 'Reg', (129, 138)) ('FLT3', 'Gene', (49, 53)) ('AML', 'Disease', (124, 127)) ('mutation', 'Var', (37, 45)) ('AML', 'Disease', 'MESH:D015470', (195, 198)) ('AML', 'Disease', 'MESH:D015470', (124, 127)) ('FLT3', 'Gene', '2322', (49, 53)) ('AML', 'Disease', (195, 198)) 1572 31632141 Mutations in FLT3 commonly occur in patients with AML who have diploid cytogenetics indicating that the mutated FLT3 is the driver for leukemogenesis. ('mutated', 'Var', (104, 111)) ('AML', 'Disease', 'MESH:D015470', (50, 53)) ('patients', 'Species', '9606', (36, 44)) ('FLT3', 'Gene', '2322', (112, 116)) ('occur', 'Reg', (27, 32)) ('FLT3', 'Gene', '2322', (13, 17)) ('leukemogenesis', 'Disease', (135, 149)) ('AML', 'Disease', (50, 53)) ('Mutations', 'Var', (0, 9)) ('FLT3', 'Gene', (112, 116)) ('FLT3', 'Gene', (13, 17)) 1573 31632141 There are two major classes of activating FLT3 mutations reported in AML patients. ('FLT3', 'Gene', (42, 46)) ('AML', 'Disease', (69, 72)) ('mutations', 'Var', (47, 56)) ('activating', 'PosReg', (31, 41)) ('FLT3', 'Gene', '2322', (42, 46)) ('AML', 'Disease', 'MESH:D015470', (69, 72)) ('patients', 'Species', '9606', (73, 81)) 1575 31632141 FLT3-ITD mutations lead to constitutive activation of the FLT3-signaling cascade with subsequent stimulation of downstream signaling pathways including signal transducer and activator of transcription 5 (STAT5), phosphatidyl-inositol 3-kinase (PI3K) and protein kinase B (AKT) pathways. ('phosphatidyl-inositol', 'Chemical', 'MESH:D010716', (212, 233)) ('AKT', 'Gene', (272, 275)) ('STAT5', 'Gene', '6776', (204, 209)) ('FLT3', 'Gene', (58, 62)) ('mutations', 'Var', (9, 18)) ('stimulation', 'PosReg', (97, 108)) ('downstream signaling pathways', 'Pathway', (112, 141)) ('STAT5', 'Gene', (204, 209)) ('activation', 'PosReg', (40, 50)) ('FLT3', 'Gene', '2322', (58, 62)) ('FLT3', 'Gene', '2322', (0, 4)) ('AKT', 'Gene', '207', (272, 275)) ('FLT3', 'Gene', (0, 4)) ('signal transducer and activator of transcription 5', 'Gene', '6776', (152, 202)) 1576 31632141 The second class of FLT3 mutations occurs as point mutations, most commonly a substitution of tyrosine for aspartic acid at codon 835 (D835Y), in the tyrosine kinase domain (FLT3-TKD). ('tyrosine', 'Chemical', 'None', (94, 102)) ('mutations', 'Var', (25, 34)) ('D835Y', 'SUBSTITUTION', 'None', (135, 140)) ('tyrosine', 'Chemical', 'None', (150, 158)) ('FLT3', 'Gene', '2322', (174, 178)) ('tyrosine for aspartic acid at codon 835', 'Var', (94, 133)) ('D835Y', 'Var', (135, 140)) ('FLT3', 'Gene', '2322', (20, 24)) ('tyrosine for aspartic acid at codon 835', 'SUBSTITUTION', 'None', (94, 133)) ('FLT3', 'Gene', (20, 24)) ('FLT3', 'Gene', (174, 178)) 1578 31632141 Similar to FLT3-ITD, these point mutations lead to downstream activation of proliferative pathways. ('proliferative pathways', 'Pathway', (76, 98)) ('FLT3', 'Gene', (11, 15)) ('point mutations', 'Var', (27, 42)) ('activation', 'PosReg', (62, 72)) ('FLT3', 'Gene', '2322', (11, 15)) 1579 31632141 Patients with FLT3-ITD mutations have similar complete remission (CR) rates compared with non-FLT3 mutated patients, but inferior outcomes due to shorter CR duration, high relapse rates, and inferior overall survival when treated with induction therapy alone, without the addition of a TKI. ('FLT3', 'Gene', '2322', (94, 98)) ('FLT3', 'Gene', (14, 18)) ('relapse', 'MPA', (172, 179)) ('overall survival', 'MPA', (200, 216)) ('FLT3', 'Gene', (94, 98)) ('mutations', 'Var', (23, 32)) ('shorter', 'NegReg', (146, 153)) ('Patients', 'Species', '9606', (0, 8)) ('FLT3', 'Gene', '2322', (14, 18)) ('patients', 'Species', '9606', (107, 115)) ('inferior', 'NegReg', (191, 199)) 1581 31632141 AML patients with FLT3-ITD mutations have shorter CR durations, higher rates of recurrence, and inferior OS compared to patients without FLT3 mutations. ('mutations', 'Var', (27, 36)) ('FLT3', 'Gene', (18, 22)) ('shorter', 'NegReg', (42, 49)) ('FLT3', 'Gene', '2322', (137, 141)) ('FLT3', 'Gene', (137, 141)) ('patients', 'Species', '9606', (120, 128)) ('AML', 'Disease', 'MESH:D015470', (0, 3)) ('patients', 'Species', '9606', (4, 12)) ('CR durations', 'MPA', (50, 62)) ('FLT3', 'Gene', '2322', (18, 22)) ('AML', 'Disease', (0, 3)) 1587 31632141 Given the significance of FLT3 mutations in AML, there has been significant interest in developing and applying targeted therapies for FLT3-mutated AML patients in the induction, consolidation, and/or maintenance phases, to decrease the risk of relapse and improve OS, as well as in relapsed FLT3-mutated AML. ('FLT3', 'Gene', '2322', (135, 139)) ('mutations', 'Var', (31, 40)) ('AML', 'Disease', 'MESH:D015470', (305, 308)) ('AML', 'Disease', 'MESH:D015470', (44, 47)) ('FLT3', 'Gene', (292, 296)) ('relapse', 'CPA', (245, 252)) ('FLT3', 'Gene', (135, 139)) ('AML', 'Disease', (148, 151)) ('AML', 'Disease', (305, 308)) ('decrease', 'NegReg', (224, 232)) ('AML', 'Disease', (44, 47)) ('improve', 'PosReg', (257, 264)) ('FLT3', 'Gene', '2322', (26, 30)) ('FLT3', 'Gene', (26, 30)) ('AML', 'Disease', 'MESH:D015470', (148, 151)) ('FLT3', 'Gene', '2322', (292, 296)) ('patients', 'Species', '9606', (152, 160)) 1595 31632141 Of note, gilteritinib a potent second-generation FLT3-inhibitor was also recently approved as single-agent therapy for patients with relapsed/refractory FLT3 mutated (both ITD and TKD) AML based on improved OS and response rates compared to conventional cytotoxic or low-intensity therapies in a randomized Phase III setting. ('AML', 'Disease', (185, 188)) ('FLT3', 'Gene', '2322', (153, 157)) ('FLT3', 'Gene', (49, 53)) ('patients', 'Species', '9606', (119, 127)) ('gilteritinib', 'Chemical', 'None', (9, 21)) ('FLT3', 'Gene', (153, 157)) ('relapsed/refractory', 'Disease', (133, 152)) ('Ph', 'Chemical', 'MESH:C084701', (307, 309)) ('improved', 'PosReg', (198, 206)) ('mutated', 'Var', (158, 165)) ('AML', 'Disease', 'MESH:D015470', (185, 188)) ('FLT3', 'Gene', '2322', (49, 53)) 1609 31632141 Also, midostaurin demonstrated more potent synergism when combined with conventional anti-leukemic agents such as cytarabine, doxorubicin and idarubicin in inhibiting FLT3 mutated, compared with FLT3 wildtype leukemia cell lines in vitro. ('mutated', 'Var', (172, 179)) ('leukemia', 'Phenotype', 'HP:0001909', (209, 217)) ('synergism', 'MPA', (43, 52)) ('FLT3', 'Gene', '2322', (167, 171)) ('leukemia', 'Disease', 'MESH:D007938', (209, 217)) ('leukemia', 'Disease', (209, 217)) ('midostaurin', 'Chemical', 'MESH:C059539', (6, 17)) ('idarubicin', 'Chemical', 'MESH:D015255', (142, 152)) ('FLT3', 'Gene', '2322', (195, 199)) ('doxorubicin', 'Chemical', 'MESH:D004317', (126, 137)) ('FLT3', 'Gene', (167, 171)) ('cytarabine', 'Chemical', 'MESH:D003561', (114, 124)) ('FLT3', 'Gene', (195, 199)) ('inhibiting', 'NegReg', (156, 166)) 1614 31632141 To that end, a total of 20 patients with relapsed/refractory AML with an FLT3-ITD or FLT3-D835Y mutation received midostaurin at a dose of 75 mg orally for 3 times daily (TID). ('FLT3', 'Gene', (73, 77)) ('D835Y', 'SUBSTITUTION', 'None', (90, 95)) ('FLT3', 'Gene', '2322', (85, 89)) ('AML', 'Disease', 'MESH:D015470', (61, 64)) ('patients', 'Species', '9606', (27, 35)) ('midostaurin', 'Chemical', 'MESH:C059539', (114, 125)) ('D835Y', 'Var', (90, 95)) ('AML', 'Disease', (61, 64)) ('FLT3', 'Gene', '2322', (73, 77)) ('FLT3', 'Gene', (85, 89)) 1621 31632141 A subsequent Phase IIB trial treated 95 patients with relapsed/refractory AML and myelodysplastic syndrome (MDS) with mutant or wildtype FLT3 with midostaurin as a single agent. ('Ph', 'Chemical', 'MESH:C084701', (13, 15)) ('MDS', 'Disease', (108, 111)) ('myelodysplastic syndrome', 'Disease', 'MESH:D009190', (82, 106)) ('MDS', 'Disease', 'MESH:D009190', (108, 111)) ('AML', 'Disease', 'MESH:D015470', (74, 77)) ('patients', 'Species', '9606', (40, 48)) ('FLT3', 'Gene', '2322', (137, 141)) ('midostaurin', 'Chemical', 'MESH:C059539', (147, 158)) ('mutant', 'Var', (118, 124)) ('myelodysplastic syndrome', 'Phenotype', 'HP:0002863', (82, 106)) ('AML', 'Disease', (74, 77)) ('FLT3', 'Gene', (137, 141)) ('myelodysplastic syndrome', 'Disease', (82, 106)) 1630 31632141 In a Phase Ib study, midostaurin at different dosing schedules in combination with chemotherapy in 79 younger adults (<60 years of age) newly diagnosed AML patients with mutant or wild-type FLT3 demonstrated CR rates of 80% in the 50 mg twice a day dosing schedule cohort (40 patients). ('patients', 'Species', '9606', (156, 164)) ('patients', 'Species', '9606', (276, 284)) ('FLT3', 'Gene', '2322', (190, 194)) ('AML', 'Disease', (152, 155)) ('midostaurin', 'Chemical', 'MESH:C059539', (21, 32)) ('FLT3', 'Gene', (190, 194)) ('mutant', 'Var', (170, 176)) ('Ph', 'Chemical', 'MESH:C084701', (5, 7)) ('AML', 'Disease', 'MESH:D015470', (152, 155)) 1632 31632141 Interestingly, the median OS of FLT3-mutated patients was similar to that of the FLT3 wildtype patients, leading to the hypothesis that the addition of TKI midostaurin could potentially neutralize the adverse impact of the FLT3 mutation and improve the outcomes of these patients. ('FLT3', 'Gene', '2322', (32, 36)) ('FLT3', 'Gene', (223, 227)) ('mutation', 'Var', (228, 236)) ('outcomes', 'MPA', (253, 261)) ('patients', 'Species', '9606', (95, 103)) ('FLT3', 'Gene', (32, 36)) ('improve', 'PosReg', (241, 248)) ('neutralize', 'NegReg', (186, 196)) ('FLT3', 'Gene', '2322', (81, 85)) ('patients', 'Species', '9606', (45, 53)) ('patients', 'Species', '9606', (271, 279)) ('FLT3', 'Gene', '2322', (223, 227)) ('midostaurin', 'Chemical', 'MESH:C059539', (156, 167)) ('FLT3', 'Gene', (81, 85)) 1634 31632141 The tolerance improved significantly when patient received 3+7 alone on Days 1 to 7 and the midostaurin was introduced from Day 8 onwards, especially with the 50 mg twice a day dose. ('improved', 'PosReg', (14, 22)) ('to 7', 'Species', '1214577', (79, 83)) ('patient', 'Species', '9606', (42, 49)) ('tolerance', 'MPA', (4, 13)) ('50 mg', 'Var', (159, 164)) ('midostaurin', 'Chemical', 'MESH:C059539', (92, 103)) 1640 31632141 In order to ensure rapid FLT3 mutational testing and support enrollment to the trial across multiple sites in different countries, a large-scale cooperative effort established an efficient polymerase chain reaction-based FLT3 mutation assay with turnaround time of less than 48 hrs. ('FLT3', 'Gene', (221, 225)) ('mutation', 'Var', (226, 234)) ('FLT3', 'Gene', '2322', (25, 29)) ('FLT3', 'Gene', (25, 29)) ('FLT3', 'Gene', '2322', (221, 225)) 1642 31632141 Patients were stratified based on the type and frequency of FLT3 mutation into 3 groups: FLT3-TKD, high allelic ratio FLT3-ITD (>0.70) and low allelic ratio FLT3-ITD (=60 years with de novo CN-AML. ('AML', 'Phenotype', 'HP:0004808', (128, 131)) ('NPM1', 'Gene', (68, 72)) ('CN-AML', 'Disease', 'MESH:D015470', (125, 131)) ('CN-AML', 'Disease', (125, 131)) ('mutation', 'Var', (73, 81)) 2522 19822134 Fifty-six percent of patients had NPM1 mutations. ('patients', 'Species', '9606', (21, 29)) ('NPM1', 'Gene', (34, 38)) ('mutations', 'Var', (39, 48)) 2523 19822134 In multivariable analyses, NPM1 mutations remained independent predictors for higher CR rates (P<0.001), longer DFS (P=0.004), and longer OS (P<0.001), after adjustment for other prognostic clinical and molecular variables. ('CR rates', 'CPA', (85, 93)) ('DFS', 'MPA', (112, 115)) ('higher', 'PosReg', (78, 84)) ('NPM1', 'Gene', (27, 31)) ('mutations', 'Var', (32, 41)) ('CR', 'Chemical', '-', (85, 87)) 2529 32811837 Chlorpromazine eliminates acute myeloid leukemia cells by perturbing subcellular localization of FLT3-ITD and KIT-D816V Mutated receptor tyrosine kinases (MT-RTKs) such as internal tandem duplication of FMS-like tyrosine kinase 3 (FLT3 ITD) and a point mutation KIT D816V are driver mutations for acute myeloid leukemia (AML). ('Chlorpromazine', 'Chemical', 'MESH:D002746', (0, 14)) ('leukemia', 'Phenotype', 'HP:0001909', (311, 319)) ('perturbing', 'Reg', (58, 68)) ('D816V', 'Var', (114, 119)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (26, 48)) ('KIT', 'Gene', '3815', (262, 265)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (297, 319)) ('KIT', 'Gene', '3815', (110, 113)) ('tyrosine kinase', 'Gene', '7294', (212, 227)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (26, 48)) ('D816V', 'SUBSTITUTION', 'None', (266, 271)) ('FMS-like tyrosine kinase 3', 'Gene', (203, 229)) ('D816V', 'SUBSTITUTION', 'None', (114, 119)) ('acute myeloid leukemia', 'Disease', (297, 319)) ('FLT3', 'Gene', (97, 101)) ('tyrosine kinase', 'Gene', (137, 152)) ('FLT3', 'Gene', (231, 235)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (32, 48)) ('FLT3', 'Gene', '2322', (97, 101)) ('KIT', 'Gene', (262, 265)) ('tyrosine kinase', 'Gene', '7294', (137, 152)) ('FLT3', 'Gene', '2322', (231, 235)) ('KIT', 'Gene', (110, 113)) ('AML', 'Disease', 'MESH:D015470', (321, 324)) ('internal tandem duplication', 'Var', (172, 199)) ('acute myeloid leukemia', 'Disease', (26, 48)) ('leukemia', 'Phenotype', 'HP:0001909', (40, 48)) ('subcellular localization', 'MPA', (69, 93)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (297, 319)) ('AML', 'Phenotype', 'HP:0004808', (321, 324)) ('FMS-like tyrosine kinase 3', 'Gene', '2322', (203, 229)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (303, 319)) ('AML', 'Disease', (321, 324)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (303, 319)) ('D816V', 'Var', (266, 271)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (32, 48)) 2531 32811837 We here show that CALM knock down leads to severely impaired FLT3 ITD- or KIT D814V-dependent cell growth compared to marginal influence on wild-type FLT3- or KIT-mediated cell growth. ('impaired', 'NegReg', (52, 60)) ('D814V-dependent', 'Var', (78, 93)) ('D814V', 'Mutation', 'p.D814V', (78, 83)) ('KIT', 'Gene', (74, 77)) ('knock down', 'Var', (23, 33)) ('FLT3', 'Gene', (61, 65)) 2533 32811837 Mechanistically, CPZ reduces CALM protein at post transcriptional level and perturbs the intracellular localization of MT-RTKs, thereby blocking their signaling. ('MT-RTKs', 'Gene', (119, 126)) ('intracellular localization', 'MPA', (89, 115)) ('signaling', 'MPA', (151, 160)) ('perturbs', 'Reg', (76, 84)) ('CPZ', 'Chemical', 'MESH:D002746', (17, 20)) ('CALM protein at post transcriptional level', 'MPA', (29, 71)) ('CPZ', 'Var', (17, 20)) ('blocking', 'NegReg', (136, 144)) ('reduces', 'NegReg', (21, 28)) 2535 32811837 Receptor tyrosine kinase mutations are frequent and associated with poor prognosis in acute myeloid leukemia (AML). ('myeloid leukemia', 'Phenotype', 'HP:0012324', (92, 108)) ('mutations', 'Var', (25, 34)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (86, 108)) ('AML', 'Phenotype', 'HP:0004808', (110, 113)) ('AML', 'Disease', (110, 113)) ('leukemia', 'Phenotype', 'HP:0001909', (100, 108)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (86, 108)) ('AML', 'Disease', 'MESH:D015470', (110, 113)) ('Receptor tyrosine kinase', 'Gene', (0, 24)) ('acute myeloid leukemia', 'Disease', (86, 108)) ('Receptor tyrosine kinase', 'Gene', '5979', (0, 24)) 2536 32811837 Here the authors show that the antipsychotic drug chlorpromazine reduces AML cells viability by perturbing the intracellular localization of FLT3-ITD and KIT-D816V. ('FLT3-ITD', 'Gene', (141, 149)) ('D816V', 'Var', (158, 163)) ('AML', 'Disease', (73, 76)) ('chlorpromazine', 'Chemical', 'MESH:D002746', (50, 64)) ('AML', 'Phenotype', 'HP:0004808', (73, 76)) ('reduces', 'NegReg', (65, 72)) ('intracellular localization', 'MPA', (111, 137)) ('D816V', 'SUBSTITUTION', 'None', (158, 163)) ('perturbing', 'Reg', (96, 106)) ('AML', 'Disease', 'MESH:D015470', (73, 76)) 2538 32811837 These mutated RTKs (MT-RTKs) trigger the aberrant activation of signal pathways involved in cancer development and progression. ('cancer', 'Disease', (92, 98)) ('cancer', 'Disease', 'MESH:D009369', (92, 98)) ('signal pathways', 'Pathway', (64, 79)) ('activation', 'PosReg', (50, 60)) ('cancer', 'Phenotype', 'HP:0002664', (92, 98)) ('mutated', 'Var', (6, 13)) 2539 32811837 Although MT-RTKs are also carried by CCVs and follow the same route to early endosomes as was the case with WT-RTKs, most of MT-RTKs remain at endoplasmic reticulum (ER) or endolysosomes, where they transmit oncogenic signals persistently and cause malignant diseases. ('malignant diseases', 'Disease', (249, 267)) ('cause', 'Reg', (243, 248)) ('MT-RTKs', 'Var', (125, 132)) ('transmit', 'Reg', (199, 207)) ('malignant diseases', 'Disease', 'MESH:D009369', (249, 267)) 2541 32811837 Internal tandem duplication of FLT3 (FLT3 ITD) is detectable in one-third of AML patients and point mutations in the tyrosine kinase domain (TKD) in about 10% of AML patients, both of which are considered to be causative mutations of AML. ('AML', 'Disease', 'MESH:D015470', (77, 80)) ('tyrosine kinase', 'Gene', '7294', (117, 132)) ('AML', 'Phenotype', 'HP:0004808', (162, 165)) ('FLT3', 'Gene', (31, 35)) ('Internal tandem duplication', 'Var', (0, 27)) ('AML', 'Disease', (162, 165)) ('point mutations in', 'Var', (94, 112)) ('patients', 'Species', '9606', (81, 89)) ('AML', 'Disease', (77, 80)) ('AML', 'Phenotype', 'HP:0004808', (77, 80)) ('tyrosine kinase', 'Gene', (117, 132)) ('AML', 'Disease', 'MESH:D015470', (234, 237)) ('detectable', 'Reg', (50, 60)) ('AML', 'Phenotype', 'HP:0004808', (234, 237)) ('AML', 'Disease', 'MESH:D015470', (162, 165)) ('patients', 'Species', '9606', (166, 174)) ('AML', 'Disease', (234, 237)) 2542 32811837 In addition, FLT3 ITD has been shown to be a poor prognostic factor for AML, while the significance of TKD remains controversial. ('AML', 'Disease', 'MESH:D015470', (72, 75)) ('FLT3 ITD', 'Var', (13, 21)) ('AML', 'Disease', (72, 75)) ('AML', 'Phenotype', 'HP:0004808', (72, 75)) 2546 32811837 A point mutation in the KIT gene at amino acid 816 (KIT D816V) is an activating mutation that is found in about 30% of patients in core binding factor (CBF)-AML. ('core binding factor', 'Gene', (131, 150)) ('KIT', 'Gene', (52, 55)) ('CBF', 'Gene', (152, 155)) ('patients', 'Species', '9606', (119, 127)) ('D816V', 'Var', (56, 61)) ('AML', 'Disease', 'MESH:D015470', (157, 160)) ('AML', 'Phenotype', 'HP:0004808', (157, 160)) ('AML', 'Disease', (157, 160)) ('D816V', 'SUBSTITUTION', 'None', (56, 61)) ('CBF', 'Gene', '10153', (152, 155)) ('core binding factor', 'Gene', '10153', (131, 150)) 2547 32811837 Although AML with t(8;21)(q22;q22) is classified into a good prognostic group, the presence of KIT D816V has been shown to be a poor prognostic factor. ('AML', 'Disease', 'MESH:D015470', (9, 12)) ('D816V', 'Var', (99, 104)) ('D816V', 'SUBSTITUTION', 'None', (99, 104)) ('AML', 'Disease', (9, 12)) ('t(8;21)(q22;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (18, 34)) ('AML', 'Phenotype', 'HP:0004808', (9, 12)) 2548 32811837 CALM (clathrin assembly lymphoid myeloid leukemia) encoded by the PICALM gene was first identified as a component of the fusion gene CALM/AF10 resulting from the chromosomal translocation t(10;11) (p13;q14) in AML cells. ('AML', 'Disease', (210, 213)) ('leukemia', 'Phenotype', 'HP:0001909', (41, 49)) ('AF10', 'Gene', (138, 142)) ('PICALM', 'Gene', (66, 72)) ('clathrin assembly lymphoid myeloid leukemia', 'Gene', (6, 49)) ('AF10', 'Gene', '8028', (138, 142)) ('AML', 'Disease', 'MESH:D015470', (210, 213)) ('clathrin assembly lymphoid myeloid leukemia', 'Gene', '8301', (6, 49)) ('t(10;11) (p13;q14', 'Var', (188, 205)) ('AML', 'Phenotype', 'HP:0004808', (210, 213)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (33, 49)) 2553 32811837 These findings are largely consistent with the phenotypes of fit1 mice harboring a nonsense point mutation in the PICALM gene. ('fit1', 'Gene', '233489', (61, 65)) ('fit1', 'Gene', (61, 65)) ('mice', 'Species', '10090', (66, 70)) ('PICALM', 'Gene', (114, 120)) ('nonsense point mutation', 'Var', (83, 106)) 2559 32811837 For this purpose, we stably introduced FLT3 (FLT3 WT or FLT3 ITD) and KIT (KIT WT or KIT D814V that corresponds to human D816V in the murine gene) into Ba/F3 cells and generated Ba/F3 sublines as follows: Ba/F3-FLT3 WT, Ba/F3-FLT3 ITD, Ba/F3-KIT WT, and Ba/F3-KIT D814V. ('murine', 'Species', '10090', (134, 140)) ('D816V', 'Var', (121, 126)) ('human', 'Species', '9606', (115, 120)) ('D814V', 'Mutation', 'p.D814V', (89, 94)) ('FLT3', 'Gene', (39, 43)) ('D814V', 'Mutation', 'p.D814V', (264, 269)) ('D816V', 'SUBSTITUTION', 'None', (121, 126)) 2561 32811837 On the other hand, Ba/F3-FLT3 ITD/SCR and Ba/F3-KIT D814V/SCR cells proliferated under IL-3-, FL-, and SCF-deprived conditions (Fig. ('D814V', 'Var', (52, 57)) ('FL', 'Gene', '2323', (94, 96)) ('proliferated', 'CPA', (68, 80)) ('D814V', 'SUBSTITUTION', 'None', (52, 57)) ('FL', 'Gene', '2323', (25, 27)) 2565 32811837 As for this mechanism, autophosphorylation of FLT3 ITD and KIT D814V and phosphorylation of their downstream molecules (STAT5 for FLT3 and ERK1/2 and Akt for KIT) were suppressed by CALM KD in Ba/F3-FLT3 ITD and Ba/F3-KIT D814V cells (Supplementary Fig. ('Akt', 'Gene', '207', (150, 153)) ('D814V', 'Mutation', 'p.D814V', (222, 227)) ('autophosphorylation', 'MPA', (23, 42)) ('suppressed', 'NegReg', (168, 178)) ('ERK1/2', 'Gene', (139, 145)) ('FLT3', 'Gene', (46, 50)) ('ERK1/2', 'Gene', '5595;5594', (139, 145)) ('STAT5', 'Gene', '6776', (120, 125)) ('Akt', 'Gene', (150, 153)) ('D814V', 'Mutation', 'p.D814V', (63, 68)) ('phosphorylation', 'MPA', (73, 88)) ('STAT5', 'Gene', (120, 125)) ('D814V', 'Var', (63, 68)) 2566 32811837 Consistent with these in vitro findings, tumorigenic activities of Ba/F3-FLT3 ITD and Ba/F3-KIT D814V were severely suppressed by CALM KD in transplanted mice (Supplementary Fig. ('tumor', 'Phenotype', 'HP:0002664', (41, 46)) ('mice', 'Species', '10090', (154, 158)) ('suppressed', 'NegReg', (116, 126)) ('tumor', 'Disease', (41, 46)) ('tumor', 'Disease', 'MESH:D009369', (41, 46)) ('D814V', 'Mutation', 'p.D814V', (96, 101)) ('Ba/F3-KIT D814V', 'Var', (86, 101)) 2567 32811837 To examine whether these findings are applicable to AML cells, we KD CALM in an inducible manner (CLAM iKD) in MV4-11 (with FLT3 ITD), HMC-1 (with KIT D816V) and HL-60 (with WT-FLT3 and KIT) cells with a doxycycline (DOX) inducible system, in which CALM shRNA expression was induced by the addition of DOX into the culture medium. ('CLAM iKD', 'Disease', 'MESH:C548072', (98, 106)) ('CLAM iKD', 'Disease', (98, 106)) ('AML', 'Phenotype', 'HP:0004808', (52, 55)) ('HMC-1', 'CellLine', 'CVCL:0003', (135, 140)) ('AML', 'Disease', (52, 55)) ('doxycycline', 'Chemical', 'MESH:D004318', (204, 215)) ('MV4-11', 'Chemical', '-', (111, 117)) ('DOX', 'Chemical', 'MESH:D004318', (217, 220)) ('D816V', 'Var', (151, 156)) ('D816V', 'SUBSTITUTION', 'None', (151, 156)) ('DOX', 'Chemical', 'MESH:D004318', (302, 305)) ('HL-60', 'CellLine', 'CVCL:0002', (162, 167)) ('expression', 'Species', '29278', (260, 270)) ('AML', 'Disease', 'MESH:D015470', (52, 55)) 2572 32811837 These results indicate that CALM is required for oncogenic signals in AML cells with MT-RTKs. ('MT-RTKs', 'Var', (85, 92)) ('AML', 'Phenotype', 'HP:0004808', (70, 73)) ('AML', 'Disease', 'MESH:D015470', (70, 73)) ('AML', 'Disease', (70, 73)) 2575 32811837 Among them, CPZ induces clathrin misassemble on internal structures, thereby inhibiting CCV formation. ('CPZ', 'Chemical', 'MESH:D002746', (12, 15)) ('inhibiting', 'NegReg', (77, 87)) ('CPZ', 'Var', (12, 15)) ('misassemble', 'Reg', (33, 44)) ('clathrin', 'Protein', (24, 32)) ('induces', 'Reg', (16, 23)) ('CCV formation', 'CPA', (88, 101)) 2576 32811837 2a, CPZ dose-dependently suppressed the growth of Ba/F3-FLT3 ITD with IC50 6.940 muM and that of Ba/F3-KIT D814V with IC50 6.942 muM. ('IC50 6.940 muM', 'Var', (70, 84)) ('growth', 'MPA', (40, 46)) ('suppressed', 'NegReg', (25, 35)) ('CPZ', 'Chemical', 'MESH:D002746', (4, 7)) ('D814V', 'Mutation', 'p.D814V', (107, 112)) 2582 32811837 When these cell lines were treated with 7.0 muM CPZ for 72 h, CPZ drastically (more than 90%) inhibited the growth and survival of MV4-11 and HMC-1 but showed only marginal (0-40%) inhibitory effects on leukemia cell lines with FLT3 WT/KIT WT (Fig. ('survival', 'CPA', (119, 127)) ('leukemia', 'Disease', 'MESH:D007938', (203, 211)) ('leukemia', 'Disease', (203, 211)) ('growth', 'CPA', (108, 114)) ('leukemia', 'Phenotype', 'HP:0001909', (203, 211)) ('MV4-11', 'Chemical', '-', (131, 137)) ('CPZ', 'Chemical', 'MESH:D002746', (62, 65)) ('CPZ', 'Chemical', 'MESH:D002746', (48, 51)) ('CPZ', 'Var', (62, 65)) ('inhibited', 'NegReg', (94, 103)) ('HMC-1', 'CellLine', 'CVCL:0003', (142, 147)) 2583 32811837 We next examined the effects of CPZ on the growth of primary AML cells with FLT3 WT/KIT WT (n = 5), FLT3 ITD (n = 3), and KIT D816V (n = 1). ('D816V', 'Var', (126, 131)) ('CPZ', 'Chemical', 'MESH:D002746', (32, 35)) ('AML', 'Disease', 'MESH:D015470', (61, 64)) ('D816V', 'SUBSTITUTION', 'None', (126, 131)) ('AML', 'Disease', (61, 64)) ('AML', 'Phenotype', 'HP:0004808', (61, 64)) 2584 32811837 These cells were cultured in the presence of SCF, FL, and TPO with or without 7.0 muM CPZ for 72 h. As expected, CPZ inhibited the growth of AML cells harboring either FLT3 ITD (#1, #5, and #7) or KIT D816V (#10), while it showed no effect (#11 and #16) or limited effects (#12, #17, and #19) on AML cells with FLT3 WT/KIT WT (Fig. ('AML', 'Disease', (296, 299)) ('TPO', 'Gene', (58, 61)) ('AML', 'Disease', (141, 144)) ('D816V', 'SUBSTITUTION', 'None', (201, 206)) ('AML', 'Disease', 'MESH:D015470', (141, 144)) ('AML', 'Phenotype', 'HP:0004808', (141, 144)) ('CPZ', 'Chemical', 'MESH:D002746', (113, 116)) ('inhibited', 'NegReg', (117, 126)) ('CPZ', 'Var', (113, 116)) ('FL', 'Gene', '2323', (50, 52)) ('FL', 'Gene', '2323', (311, 313)) ('AML', 'Disease', 'MESH:D015470', (296, 299)) ('CPZ', 'Chemical', 'MESH:D002746', (86, 89)) ('growth', 'MPA', (131, 137)) ('FL', 'Gene', '2323', (168, 170)) ('D816V', 'Var', (201, 206)) ('TPO', 'Gene', '7173', (58, 61)) ('AML', 'Phenotype', 'HP:0004808', (296, 299)) 2588 32811837 This fraction (from Cases #3, #5) was also sensitive to CPZ, suggesting a possibility that CPZ may have a potential to eradicate AML initiating cells (Fig. ('CPZ', 'Chemical', 'MESH:D002746', (91, 94)) ('AML', 'Disease', 'MESH:D015470', (129, 132)) ('CPZ', 'Var', (91, 94)) ('eradicate', 'NegReg', (119, 128)) ('AML', 'Disease', (129, 132)) ('AML', 'Phenotype', 'HP:0004808', (129, 132)) ('CPZ', 'Chemical', 'MESH:D002746', (56, 59)) 2603 32811837 Of note, because CPZ reduced CALM protein levels in MV4-11 and HMC-1 cells (Fig. ('reduced', 'NegReg', (21, 28)) ('CALM protein levels', 'MPA', (29, 48)) ('MV4-11', 'Chemical', '-', (52, 58)) ('CPZ', 'Chemical', 'MESH:D002746', (17, 20)) ('CPZ', 'Var', (17, 20)) ('HMC-1', 'CellLine', 'CVCL:0003', (63, 68)) 2608 32811837 When CALM iKD MV4-11 cells were treated with DOX, the amount of CALM was effectively reduced by iKD (Fig. ('iKD', 'Var', (96, 99)) ('CALM iKD', 'Disease', 'None', (5, 13)) ('DOX', 'Chemical', 'MESH:D004318', (45, 48)) ('MV4-11', 'Chemical', '-', (14, 20)) ('CALM iKD', 'Disease', (5, 13)) 2614 32811837 Consistent with this observation, the amount of FLT3 ITD was severely reduced by CALM iKD or CPZ treatment in the isolated PDI-positive ER fraction compared with untreated cells (Fig. ('CALM iKD', 'Disease', (81, 89)) ('reduced', 'NegReg', (70, 77)) ('FLT3', 'Gene', (48, 52)) ('CPZ', 'Var', (93, 96)) ('PDI', 'Gene', '5034', (123, 126)) ('CALM iKD', 'Disease', 'None', (81, 89)) ('PDI', 'Gene', (123, 126)) ('CPZ', 'Chemical', 'MESH:D002746', (93, 96)) 2620 32811837 Although 86.5% of KIT D816V was co-localized with CALM, this co-localization was impaired by CALM iKD or CPZ treatment due to CALM depletion (to 7.1% by CALM iKD; to 12.2% by CPZ) (Fig. ('co-localized', 'Interaction', (32, 44)) ('CPZ', 'Chemical', 'MESH:D002746', (105, 108)) ('D816V', 'SUBSTITUTION', 'None', (22, 27)) ('CALM iKD', 'Disease', 'None', (153, 161)) ('CALM depletion', 'MPA', (126, 140)) ('CPZ', 'Chemical', 'MESH:D002746', (175, 178)) ('CALM iKD', 'Disease', (93, 101)) ('D816V', 'Var', (22, 27)) ('CALM iKD', 'Disease', (153, 161)) ('CALM iKD', 'Disease', 'None', (93, 101)) 2622 32811837 Without any treatment, KIT D816V was co-localized with LAMP1 (26.4%) (Fig. ('LAMP1', 'Gene', (55, 60)) ('D816V', 'SUBSTITUTION', 'None', (27, 32)) ('D816V', 'Var', (27, 32)) ('LAMP1', 'Gene', '3916', (55, 60)) 2624 32811837 In contrast, CALM iKD by DOX treatment or CPZ treatment obviously reduced co-localization of KIT D816V with LAMP1 (to 7.2% by CALM iKD; to 8% by CPZ), with EEA1 (to 6.3% by CALM iKD; to 6.1% by CPZ), and with PDI (to 8.4% by CALM iKD; to 8.3% by CPZ), GM130 (to 7.1% by CALM iKD; to 5.9% by CPZ) except for the co-localization with Rab11 (to 4.6% by CALM iKD; to 4.3% by CPZ) (Fig. ('reduced', 'NegReg', (66, 73)) ('Rab11', 'Gene', (332, 337)) ('LAMP1', 'Gene', '3916', (108, 113)) ('CALM iKD', 'Disease', 'None', (13, 21)) ('CPZ', 'Chemical', 'MESH:D002746', (291, 294)) ('PDI', 'Gene', '5034', (209, 212)) ('CALM iKD', 'Disease', 'None', (126, 134)) ('D816V', 'Var', (97, 102)) ('CALM iKD', 'Disease', (225, 233)) ('CPZ', 'Chemical', 'MESH:D002746', (145, 148)) ('GM130', 'Gene', '2801', (252, 257)) ('CPZ', 'Chemical', 'MESH:D002746', (194, 197)) ('GM130', 'Gene', (252, 257)) ('CALM iKD', 'Disease', 'None', (173, 181)) ('PDI', 'Gene', (209, 212)) ('CPZ', 'Chemical', 'MESH:D002746', (42, 45)) ('D816V', 'SUBSTITUTION', 'None', (97, 102)) ('CALM iKD', 'Disease', (270, 278)) ('co-localization', 'MPA', (74, 89)) ('LAMP1', 'Gene', (108, 113)) ('CALM iKD', 'Disease', 'None', (225, 233)) ('CPZ', 'Chemical', 'MESH:D002746', (246, 249)) ('DOX', 'Chemical', 'MESH:D004318', (25, 28)) ('CALM iKD', 'Disease', (350, 358)) ('Rab11', 'Gene', '8766', (332, 337)) ('EEA1', 'Gene', (156, 160)) ('CALM iKD', 'Disease', (13, 21)) ('CALM iKD', 'Disease', (126, 134)) ('CALM iKD', 'Disease', 'None', (270, 278)) ('CPZ', 'Chemical', 'MESH:D002746', (371, 374)) ('EEA1', 'Gene', '8411', (156, 160)) ('CALM iKD', 'Disease', 'None', (350, 358)) ('CALM iKD', 'Disease', (173, 181)) 2625 32811837 In accord with these findings, the amount of KIT D816V in the endolysosome fraction coimmunoprecipitated with LAMP1 was significantly reduced by CALM iKD or CPZ treatment compared with untreated HMC-1 cells (Fig. ('CALM iKD', 'Disease', 'None', (145, 153)) ('CPZ', 'Chemical', 'MESH:D002746', (157, 160)) ('D816V', 'SUBSTITUTION', 'None', (49, 54)) ('LAMP1', 'Gene', '3916', (110, 115)) ('D816V', 'Var', (49, 54)) ('CALM iKD', 'Disease', (145, 153)) ('reduced', 'NegReg', (134, 141)) ('HMC-1', 'CellLine', 'CVCL:0003', (195, 200)) ('CPZ', 'Var', (157, 160)) ('LAMP1', 'Gene', (110, 115)) 2627 32811837 We also found that phosphorylation of KIT D816V and Akt was inhibited by CPZ in a dose-dependent manner (Fig. ('phosphorylation', 'MPA', (19, 34)) ('D816V', 'Var', (42, 47)) ('Akt', 'Gene', (52, 55)) ('inhibited', 'NegReg', (60, 69)) ('D816V', 'SUBSTITUTION', 'None', (42, 47)) ('CPZ', 'Chemical', 'MESH:D002746', (73, 76)) ('Akt', 'Gene', '207', (52, 55)) ('KIT', 'Gene', (38, 41)) 2628 32811837 This result is largely consistent with a previous paper showing that localization at endolysosomes is critical for KIT D816V to activate Akt. ('D816V', 'Var', (119, 124)) ('D816V', 'SUBSTITUTION', 'None', (119, 124)) ('Akt', 'Gene', '207', (137, 140)) ('activate', 'PosReg', (128, 136)) ('Akt', 'Gene', (137, 140)) 2629 32811837 Together, these results indicate that CPZ reduces CALM protein and perturbs the intracellular localization of FLT ITD and KIT D816V, thereby inhibiting their oncogenic signals. ('oncogenic signals', 'CPA', (158, 175)) ('D816V', 'Var', (126, 131)) ('FLT', 'Gene', '2321', (110, 113)) ('CALM protein', 'MPA', (50, 62)) ('FLT', 'Gene', (110, 113)) ('reduces', 'NegReg', (42, 49)) ('D816V', 'SUBSTITUTION', 'None', (126, 131)) ('intracellular localization', 'MPA', (80, 106)) ('inhibiting', 'NegReg', (141, 151)) ('CPZ', 'Chemical', 'MESH:D002746', (38, 41)) ('KIT', 'Gene', (122, 125)) ('CPZ', 'Var', (38, 41)) ('perturbs', 'NegReg', (67, 75)) 2638 32811837 We analyzed the in vivo effect of CPZ on primary AML cells from 19 patients (Age 17-81, median 48, male/female 12/7), including AML cells with FLT3 WT/KIT WT (ten cases), FLT3 ITD (eight cases), and KIT D816V (one case) (Supplementary Table 1). ('AML', 'Disease', (49, 52)) ('AML', 'Disease', (128, 131)) ('patients', 'Species', '9606', (67, 75)) ('AML', 'Phenotype', 'HP:0004808', (128, 131)) ('D816V', 'Var', (203, 208)) ('CPZ', 'Chemical', 'MESH:D002746', (34, 37)) ('D816V', 'SUBSTITUTION', 'None', (203, 208)) ('AML', 'Disease', 'MESH:D015470', (49, 52)) ('AML', 'Disease', 'MESH:D015470', (128, 131)) ('AML', 'Phenotype', 'HP:0004808', (49, 52)) 2640 32811837 As seen in case #2, CPZ treatment effectively inhibited the growth of AML samples with MT-RTKs and statistically significant differences were observed in 6/8 cases with FLT3 ITD and 1/1 case with KIT D816V. ('FLT3 ITD', 'Var', (169, 177)) ('AML', 'Disease', (70, 73)) ('growth', 'MPA', (60, 66)) ('AML', 'Phenotype', 'HP:0004808', (70, 73)) ('D816V', 'Var', (200, 205)) ('CPZ', 'Chemical', 'MESH:D002746', (20, 23)) ('AML', 'Disease', 'MESH:D015470', (70, 73)) ('inhibited', 'NegReg', (46, 55)) ('D816V', 'SUBSTITUTION', 'None', (200, 205)) 2660 32811837 In this study we found that CALM protein is critical for the growth and survival of leukemia cells harboring MT-RTKs but not for WT-RTKs-dependent growth. ('leukemia', 'Phenotype', 'HP:0001909', (84, 92)) ('leukemia', 'Disease', (84, 92)) ('leukemia', 'Disease', 'MESH:D007938', (84, 92)) ('MT-RTKs', 'Var', (109, 116)) 2661 32811837 In accord with previous reports, we found that FLT3 ITD is predominantly localized at ER and KIT D816V at endolysosomes, where they activate themselves and their downstream molecules, STAT5 and Akt. ('Akt', 'Gene', '207', (194, 197)) ('STAT5', 'Gene', '6776', (184, 189)) ('KIT', 'Gene', (93, 96)) ('Akt', 'Gene', (194, 197)) ('STAT5', 'Gene', (184, 189)) ('D816V', 'Var', (97, 102)) ('activate', 'PosReg', (132, 140)) ('FLT3', 'Gene', (47, 51)) ('D816V', 'SUBSTITUTION', 'None', (97, 102)) 2664 32811837 Although we tried to determine the precise localization of FLT3 ITD and KIT D816V after CALM iKD or CPZ treatment, using several compartment markers such PDI (for ER), GM130 (for Golgi), EEA1 (for endosome), LAMP1 (for endolysosome), and Rab11 (for recycling endosome) in the immunofluorescence analyses, the co-localization pattern of these MT-RTKs didn't match those of any tested markers. ('GM130', 'Gene', '2801', (168, 173)) ('LAMP1', 'Gene', (208, 213)) ('CALM iKD', 'Disease', 'None', (88, 96)) ('EEA1', 'Gene', '8411', (187, 191)) ('PDI', 'Gene', (154, 157)) ('GM130', 'Gene', (168, 173)) ('LAMP1', 'Gene', '3916', (208, 213)) ('PDI', 'Gene', '5034', (154, 157)) ('Rab11', 'Gene', '8766', (238, 243)) ('D816V', 'Var', (76, 81)) ('EEA1', 'Gene', (187, 191)) ('CPZ', 'Chemical', 'MESH:D002746', (100, 103)) ('CALM iKD', 'Disease', (88, 96)) ('Rab11', 'Gene', (238, 243)) ('D816V', 'SUBSTITUTION', 'None', (76, 81)) 2669 32811837 CPZ also inhibits other receptors, including receptors for 5-HT, H1 histamine, alpha1 and alpha2 adrenaline, and M1 and M2 muscarinic acetylcholine receptors. ('acetylcholine', 'Chemical', 'MESH:D000109', (134, 147)) ('alpha2 adrenaline', 'Protein', (90, 107)) ('histamine', 'Chemical', 'MESH:D006632', (68, 77)) ('alpha1', 'Protein', (79, 85)) ('5-HT', 'Chemical', 'MESH:D012701', (59, 63)) ('5-HT', 'Protein', (59, 63)) ('CPZ', 'Chemical', 'MESH:D002746', (0, 3)) ('inhibits', 'NegReg', (9, 17)) ('CPZ', 'Var', (0, 3)) ('H1 histamine', 'Protein', (65, 77)) ('adrenaline', 'Chemical', 'MESH:D004837', (97, 107)) 2671 32811837 In this study, we utilized CPZ in experiments to test its properties as an inhibitor of CCV formation and to our surprise, we observed that CPZ inhibited the growth/survival of AML cells with MT-RTKs, while it showed minor effects on AML cells with WT-RTKs. ('MT-RTKs', 'Var', (192, 199)) ('AML', 'Disease', (177, 180)) ('AML', 'Phenotype', 'HP:0004808', (177, 180)) ('growth/survival', 'CPA', (158, 173)) ('AML', 'Disease', 'MESH:D015470', (177, 180)) ('inhibited', 'NegReg', (144, 153)) ('CPZ', 'Chemical', 'MESH:D002746', (140, 143)) ('AML', 'Disease', 'MESH:D015470', (234, 237)) ('CPZ', 'Chemical', 'MESH:D002746', (27, 30)) ('CPZ', 'Var', (140, 143)) ('AML', 'Phenotype', 'HP:0004808', (234, 237)) ('AML', 'Disease', (234, 237)) 2672 32811837 To clarify the mechanism underlying anti-AML activities of CPZ, we examined the effects of CPZ on the expression of CALM protein and found that CPZ reduced CALM protein at posttranscriptional levels. ('AML', 'Phenotype', 'HP:0004808', (41, 44)) ('expression', 'Species', '29278', (102, 112)) ('CALM protein', 'MPA', (156, 168)) ('CPZ', 'Chemical', 'MESH:D002746', (91, 94)) ('reduced', 'NegReg', (148, 155)) ('AML', 'Disease', 'MESH:D015470', (41, 44)) ('CPZ', 'Chemical', 'MESH:D002746', (144, 147)) ('CPZ', 'Chemical', 'MESH:D002746', (59, 62)) ('AML', 'Disease', (41, 44)) ('CPZ', 'Var', (144, 147)) 2673 32811837 Also, we found that the intracellular location of FLT3 ITD and KIT D816V was perturbed by CALM iKD or CPZ treatment in MV4-11 and HMC1 cells, respectively. ('MV4-11', 'Chemical', '-', (119, 125)) ('perturbed', 'Reg', (77, 86)) ('D816V', 'SUBSTITUTION', 'None', (67, 72)) ('HMC1', 'CellLine', 'CVCL:0003', (130, 134)) ('FLT3', 'Gene', (50, 54)) ('CPZ', 'Chemical', 'MESH:D002746', (102, 105)) ('intracellular location', 'MPA', (24, 46)) ('CALM iKD', 'Disease', (90, 98)) ('D816V', 'Var', (67, 72)) ('CALM iKD', 'Disease', 'None', (90, 98)) ('KIT', 'Gene', (63, 66)) 2674 32811837 In addition, we confirmed that a DR antagonist, THIO, and a 5-HTR antagonist, METHIO, showed only marginal growth inhibitory effects on MV4-11 and HMC-1 cells harboring MT-RTKs, while they were highly effective for HL-60, Kasumi, and KG-1 cells with WT-RTKs. ('5-HT', 'Chemical', 'MESH:D012701', (60, 64)) ('HMC-1', 'CellLine', 'CVCL:0003', (147, 152)) ('HTR', 'Gene', (62, 65)) ('METHIO', 'Chemical', 'MESH:D008719', (78, 84)) ('HL-60', 'CellLine', 'CVCL:0002', (215, 220)) ('MT-RTKs', 'Var', (169, 176)) ('growth', 'MPA', (107, 113)) ('THIO', 'Chemical', 'MESH:D013881', (48, 52)) ('THIO', 'Chemical', 'MESH:D013881', (80, 84)) ('MV4-11', 'Chemical', '-', (136, 142)) ('HTR', 'Gene', '7012', (62, 65)) 2676 32811837 However, CPZ still further suppressed the growth of CALM iKD MV4-11 and HMC-1 cells to some extent. ('HMC-1', 'CellLine', 'CVCL:0003', (72, 77)) ('CPZ', 'Chemical', 'MESH:D002746', (9, 12)) ('CPZ', 'Var', (9, 12)) ('suppressed', 'NegReg', (27, 37)) ('growth', 'MPA', (42, 48)) ('CALM iKD', 'Disease', (52, 60)) ('CALM iKD', 'Disease', 'None', (52, 60)) ('MV4-11', 'Chemical', '-', (61, 67)) 2678 32811837 This speculation is supported by the fact that METHIO and THIO partially inhibited the growth of MV4-11 and HMC-1 cells. ('METHIO', 'Chemical', 'MESH:D008719', (47, 53)) ('HMC-1', 'CellLine', 'CVCL:0003', (108, 113)) ('growth', 'CPA', (87, 93)) ('THIO', 'Var', (58, 62)) ('METHIO', 'Var', (47, 53)) ('inhibited', 'NegReg', (73, 82)) ('THIO', 'Chemical', 'MESH:D013881', (49, 53)) ('THIO', 'Chemical', 'MESH:D013881', (58, 62)) ('MV4-11', 'Chemical', '-', (97, 103)) 2685 32811837 Phenothiazine was first found to induce cell death in various leukemia cell lines, although its molecular mechanisms were not investigated. ('Phenothiazine', 'Chemical', 'MESH:C031637', (0, 13)) ('cell death', 'CPA', (40, 50)) ('leukemia', 'Disease', (62, 70)) ('leukemia', 'Phenotype', 'HP:0001909', (62, 70)) ('leukemia', 'Disease', 'MESH:D007938', (62, 70)) ('Phenothiazine', 'Var', (0, 13)) 2688 32811837 Both FLT3 ITD and KIT D816V confer poor prognosis on AML patients and a number of preclinical and clinical studies have focused on the development of inhibitors of these MT-RTKs. ('KIT', 'Gene', (18, 21)) ('D816V', 'SUBSTITUTION', 'None', (22, 27)) ('AML', 'Disease', 'MESH:D015470', (53, 56)) ('patients', 'Species', '9606', (57, 65)) ('AML', 'Disease', (53, 56)) ('FLT3', 'Gene', (5, 9)) ('AML', 'Phenotype', 'HP:0004808', (53, 56)) ('D816V', 'Var', (22, 27)) 2689 32811837 As for FLT3 inhibitors, 1st generation multikinase inhibitors, such as lestaurtinib, sorafenib, and midostaurin, have been investigated, however, clinical efficacies for AML with FLT3 mutations are still limited, and may contribute to adverse effects due to the inhibition of multiple other kinases. ('mul', 'Gene', (39, 42)) ('AML', 'Disease', 'MESH:D015470', (170, 173)) ('mul', 'Gene', '68729', (276, 279)) ('mutations', 'Var', (184, 193)) ('midostaurin', 'Chemical', 'MESH:C059539', (100, 111)) ('mul', 'Gene', '68729', (39, 42)) ('AML', 'Phenotype', 'HP:0004808', (170, 173)) ('AML', 'Disease', (170, 173)) ('sorafenib', 'Chemical', 'MESH:D000077157', (85, 94)) ('mul', 'Gene', (276, 279)) ('lestaurtinib', 'Chemical', 'MESH:C119379', (71, 83)) ('FLT3', 'Gene', (179, 183)) 2690 32811837 To overcome these problems, 2nd generation FLT3 inhibitors with selective FLT3 inhibitory activity such as gilteritinib, quizartinib, and crenolanib have been developed, and gilteritinib was recently approved for relapsed/refractory adult AML patients with mutated FLT3. ('gilteritinib', 'Chemical', 'MESH:C000609080', (174, 186)) ('AML', 'Disease', 'MESH:D015470', (239, 242)) ('patients', 'Species', '9606', (243, 251)) ('FLT3', 'Gene', (265, 269)) ('quizartinib', 'Chemical', 'MESH:C544967', (121, 132)) ('FLT3', 'Gene', (74, 78)) ('AML', 'Phenotype', 'HP:0004808', (239, 242)) ('gilteritinib', 'Chemical', 'MESH:C000609080', (107, 119)) ('AML', 'Disease', (239, 242)) ('relapsed/refractory', 'Disease', (213, 232)) ('crenolanib', 'Chemical', 'MESH:C577197', (138, 148)) ('mutated', 'Var', (257, 264)) 2691 32811837 As for KIT inhibitors, imatinib has been tested in AML with or without KIT D816V because KIT is autophosphorylated in AML cells regardless of the presence of KIT D816V. ('AML', 'Disease', (51, 54)) ('AML', 'Phenotype', 'HP:0004808', (51, 54)) ('D816V', 'SUBSTITUTION', 'None', (75, 80)) ('AML', 'Phenotype', 'HP:0004808', (118, 121)) ('D816V', 'Var', (162, 167)) ('AML', 'Disease', (118, 121)) ('imatinib', 'Chemical', 'MESH:D000068877', (23, 31)) ('AML', 'Disease', 'MESH:D015470', (51, 54)) ('D816V', 'SUBSTITUTION', 'None', (162, 167)) ('AML', 'Disease', 'MESH:D015470', (118, 121)) ('D816V', 'Var', (75, 80)) 2694 32811837 For example, T315I mutation in the BCR-ABL gene is known as a gatekeeper mutation against 1st and 2nd generation BCR-ABL inhibitors. ('T315I', 'Mutation', 'rs121913459', (13, 18)) ('gatekeeper', 'Species', '111938', (62, 72)) ('BCR-ABL', 'Gene', (35, 42)) ('T315I', 'Var', (13, 18)) 2695 32811837 Similarly, T790M mutation in the epidermal growth factor receptor (EGFR) gene causes resistance to 1st and 2nd generation EGFR inhibitors. ('epidermal growth factor receptor', 'Gene', '1956', (33, 65)) ('causes', 'Reg', (78, 84)) ('T790M', 'Mutation', 'rs121434569', (11, 16)) ('EGFR', 'Gene', '1956', (67, 71)) ('EGFR', 'Gene', (67, 71)) ('T790M', 'Var', (11, 16)) ('EGFR', 'Gene', '1956', (122, 126)) ('EGFR', 'Gene', (122, 126)) ('epidermal growth factor receptor', 'Gene', (33, 65)) 2696 32811837 Also, D835Y, D835V, and D835F mutations in the FLT3 gene confer resistance to quizartinib on AML cells with FLT3 ITD. ('D835V', 'Var', (13, 18)) ('AML', 'Phenotype', 'HP:0004808', (93, 96)) ('D835Y', 'Var', (6, 11)) ('AML', 'Disease', (93, 96)) ('quizartinib', 'Chemical', 'MESH:C544967', (78, 89)) ('D835V', 'Mutation', 'rs121909646', (13, 18)) ('D835F', 'Var', (24, 29)) ('FLT3', 'Gene', (47, 51)) ('resistance', 'MPA', (64, 74)) ('D835Y', 'Mutation', 'rs121913488', (6, 11)) ('AML', 'Disease', 'MESH:D015470', (93, 96)) ('D835F', 'SUBSTITUTION', 'None', (24, 29)) 2698 32811837 Together, these results indicate that kinase inhibition by ATP-competitive TKIs has a limit to elicit prominent and durable antitumor effects. ('tumor', 'Disease', 'MESH:D009369', (128, 133)) ('ATP', 'Chemical', 'MESH:D000255', (59, 62)) ('tumor', 'Phenotype', 'HP:0002664', (128, 133)) ('TKIs', 'Gene', (75, 79)) ('ATP-competitive', 'Var', (59, 74)) ('tumor', 'Disease', (128, 133)) ('kinase inhibition', 'MPA', (38, 55)) 2700 32811837 Importantly, CPZ exhibited antileukemic activities without inhibiting normal hematopoiesis at a concentration, which is attainable when it is used as an anti-psychiatric drug for psychopathic patients. ('antileukemic activities', 'CPA', (27, 50)) ('psychiatric', 'Disease', (158, 169)) ('psychopathic', 'Disease', 'MESH:D000987', (179, 191)) ('patients', 'Species', '9606', (192, 200)) ('psychopathic', 'Disease', (179, 191)) ('CPZ', 'Chemical', 'MESH:D002746', (13, 16)) ('CPZ', 'Var', (13, 16)) ('psychiatric', 'Disease', 'MESH:D001523', (158, 169)) 2701 32811837 CPZ also significantly reduced the number of AML initiating cells, proposing its potential to totally eradicate AML. ('reduced', 'NegReg', (23, 30)) ('AML', 'Phenotype', 'HP:0004808', (45, 48)) ('AML', 'Disease', 'MESH:D015470', (112, 115)) ('CPZ', 'Chemical', 'MESH:D002746', (0, 3)) ('AML', 'Disease', 'MESH:D015470', (45, 48)) ('AML', 'Phenotype', 'HP:0004808', (112, 115)) ('CPZ', 'Var', (0, 3)) ('AML', 'Disease', (112, 115)) ('AML', 'Disease', (45, 48)) 2717 32811837 These shRNA expression vectors were transduced into Ba/F3 clones each expressing FLT3 WT, FLT3 ITD, KIT WT, and KIT D814V by a retrovirus system. ('KIT D814V', 'Var', (112, 121)) ('D814V', 'Mutation', 'p.D814V', (116, 121)) ('FLT3 ITD', 'Var', (90, 98)) ('FLT3', 'Gene', (81, 85)) ('expression vectors', 'Species', '29278', (12, 30)) 2723 32811837 PCR primers for human CALM (Hs00200318_m1), human KIT (Hs00174029_m1), and human GAPDH (Hs02786624_g1) were purchased from applied biosystems (Thermo Fisher Scientific). ('GAPDH', 'Gene', (81, 86)) ('Hs00200318_m1', 'Var', (28, 41)) ('human', 'Species', '9606', (44, 49)) ('Hs00174029_m1', 'Var', (55, 68)) ('Hs02786624_g1', 'Var', (88, 101)) ('GAPDH', 'Gene', '2597', (81, 86)) ('human', 'Species', '9606', (16, 21)) ('human', 'Species', '9606', (75, 80)) 2751 32811837 Human KIT gene was initially amplified by 40 cycles of PCR with human KIT-D816V primers as follows: denaturation (95 C for 30 s), annealing, (61 C for 25 s), and extension (72 C for 25 s). ('denaturation', 'MPA', (100, 112)) ('human', 'Species', '9606', (64, 69)) ('Human', 'Species', '9606', (0, 5)) ('D816V', 'SUBSTITUTION', 'None', (74, 79)) ('61 C for', 'Var', (143, 152)) ('annealing', 'Var', (131, 140)) ('D816V', 'Var', (74, 79)) 2753 32811837 The sequences of the PCR primers used in this study were: Human FLT3-ITD forward: 5'-GCAATTTAGGTATGAAAGCCAGC-3' Human FLT3-ITD reverse: 5'-CTTTCAGCATTTTGACGGCAACC-3' Human KIT-D816V forward: 5'-CCTCCAACCTAATAGTGTATTCACAG-3' Human KIT-D816V reverse: 5'-ATGTGTGATATCCCTAGACAGGAT-3' For transplantation of Ba/F3 FLT3 ITD/Mock or/CALM KD cells (5 x 103 cells) were suspended in 100 mul PBS and injected via tail vein into 6-8-week old female BALB/C mice. ('D816V', 'Var', (234, 239)) ('D816V', 'Var', (176, 181)) ('Human', 'Species', '9606', (58, 63)) ('Human', 'Species', '9606', (224, 229)) ('mul', 'Gene', (378, 381)) ('Human', 'Species', '9606', (166, 171)) ('D816V', 'SUBSTITUTION', 'None', (234, 239)) ('mice', 'Species', '10090', (445, 449)) ('PBS', 'Chemical', 'MESH:D007854', (382, 385)) ('mul', 'Gene', '68729', (378, 381)) ('D816V', 'SUBSTITUTION', 'None', (176, 181)) ('Human', 'Species', '9606', (112, 117)) 2754 32811837 To transplant Ba/F3 KIT D814V/Mock and/CALM KD cells (each 2 x 106 cells/100 mul PBS) were subcutaneously injected into two sites in the posterior flank of 2.5 Gy irradiated 6-8-week old nude mice (BALB/cAJcl-nu/nu mice purchased from CLEA Japan, Inc., Tokyo, Japan). ('PBS', 'Chemical', 'MESH:D007854', (81, 84)) ('mice', 'Species', '10090', (192, 196)) ('mul', 'Gene', '68729', (77, 80)) ('nude mice', 'Species', '10090', (187, 196)) ('D814V', 'SUBSTITUTION', 'None', (24, 29)) ('mice', 'Species', '10090', (215, 219)) ('D814V', 'Var', (24, 29)) ('mul', 'Gene', (77, 80)) 2766 32811837 S.R., M.S., T.K, A.T. and H.T. ('M.S.', 'Var', (6, 10)) ('H.T', 'Disease', 'MESH:D000848', (26, 29)) ('H.T', 'Disease', (26, 29)) 2772 32321944 The stem-cell-targeted alpha-particle therapy using 211At-CXCR4 mAb for AML appears possible and requires further therapeutic studies. ('AML', 'Disease', 'MESH:D015470', (72, 75)) ('AML', 'Disease', (72, 75)) ('211At-CXCR4', 'Var', (52, 63)) ('AML', 'Phenotype', 'HP:0004808', (72, 75)) 2796 32321944 Table 1 summarizes the 211At-CXCR4 mAb and 125I-CXCR4 mAb uptakes in major organs and tumors up to 24 h after intravenous injection into the mice with tumor xenografts. ('tumor', 'Phenotype', 'HP:0002664', (86, 91)) ('tumor', 'Phenotype', 'HP:0002664', (151, 156)) ('tumor', 'Disease', (86, 91)) ('tumor', 'Disease', (151, 156)) ('tumors', 'Disease', (86, 92)) ('tumors', 'Disease', 'MESH:D009369', (86, 92)) ('tumor', 'Disease', 'MESH:D009369', (151, 156)) ('tumors', 'Phenotype', 'HP:0002664', (86, 92)) ('mice', 'Species', '10090', (141, 145)) ('tumor', 'Disease', 'MESH:D009369', (86, 91)) ('125I-CXCR4', 'Var', (43, 53)) 2797 32321944 Conversely, the thyroid gland and stomach showed significantly higher uptakes of 211At-CXCR4 mAb than 125I-CXCR4 mAb from 1 min to 24 h. The tumor showed gradually increased uptake of 211At-CXCR4 mAb, and the highest uptake was obtained at 6 h. 125I-CXCR4 mAb also showed the highest accumulation in the tumor at 6 h. Accumulation in other normal organs was almost similar for both 211At-CXCR4 mAb and 125I-CXCR4 mAb. ('tumor', 'Disease', 'MESH:D009369', (141, 146)) ('tumor', 'Phenotype', 'HP:0002664', (141, 146)) ('tumor', 'Disease', 'MESH:D009369', (304, 309)) ('tumor', 'Disease', (141, 146)) ('tumor', 'Phenotype', 'HP:0002664', (304, 309)) ('tumor', 'Disease', (304, 309)) ('211At-CXCR4 mAb', 'Var', (382, 397)) 2798 32321944 The localizations of 211At-CXCR4 mAb and 125I-CXCR4 mAb in the tumor showed no significant difference. ('tumor', 'Disease', (63, 68)) ('211At-CXCR4 mAb', 'Var', (21, 36)) ('tumor', 'Disease', 'MESH:D009369', (63, 68)) ('125I-CXCR4', 'Var', (41, 51)) ('tumor', 'Phenotype', 'HP:0002664', (63, 68)) 2799 32321944 Compared with the background accumulation such as that in muscle, 211At-CXCR4 mAb was highly accumulated in the tumors (Fig. ('211At-CXCR4', 'Var', (66, 77)) ('accumulated', 'PosReg', (93, 104)) ('tumors', 'Disease', (112, 118)) ('tumor', 'Phenotype', 'HP:0002664', (112, 117)) ('tumors', 'Disease', 'MESH:D009369', (112, 118)) ('tumors', 'Phenotype', 'HP:0002664', (112, 118)) 2802 32321944 The mean organ absorbed dose estimated with the 25 g mouse phantom was highest in the thyroid gland, followed by the stomach, lung, heart, and kidney for 211At-CXCR4 mAb. ('mouse', 'Species', '10090', (53, 58)) ('211At-CXCR4 mAb', 'Var', (154, 169)) ('organ absorbed dose', 'MPA', (9, 28)) 2807 32321944 The threefold difference in the tumor absorbed doses between 211At-CXCR4 mAb and 125I-CXCR4 mAb may be explained by the difference in the weights of the tumors. ('211At-CXCR4 mAb', 'Var', (61, 76)) ('tumor', 'Disease', 'MESH:D009369', (153, 158)) ('125I-CXCR4', 'Var', (81, 91)) ('tumor', 'Disease', 'MESH:D009369', (32, 37)) ('tumor', 'Phenotype', 'HP:0002664', (153, 158)) ('tumor', 'Disease', (153, 158)) ('tumor', 'Phenotype', 'HP:0002664', (32, 37)) ('tumors', 'Disease', (153, 159)) ('tumors', 'Phenotype', 'HP:0002664', (153, 159)) ('tumor', 'Disease', (32, 37)) ('tumors', 'Disease', 'MESH:D009369', (153, 159)) 2809 32321944 To examine the feasibility of alpha-particle therapy targeting CXCR4 in AML, a preclinical study was carried out with 211At-CXCR4 mAb and 125I-CXCR4 mAb in a human AML xenograft model. ('AML', 'Disease', 'MESH:D015470', (72, 75)) ('human', 'Species', '9606', (158, 163)) ('AML', 'Phenotype', 'HP:0004808', (72, 75)) ('AML', 'Disease', (72, 75)) ('AML', 'Disease', 'MESH:D015470', (164, 167)) ('211At-CXCR4', 'Var', (118, 129)) ('AML', 'Disease', (164, 167)) ('125I-CXCR4', 'Var', (138, 148)) ('AML', 'Phenotype', 'HP:0004808', (164, 167)) 2810 32321944 A biodistribution study revealed that 211At-CXCR4 mAb as well as 125I-CXCR4 mAb showed the highest tumor uptake and the highest tumor-to-muscle ratio at 6 h, which agreed with the physical half-life of 211At, although the precise biological behavior from 6 to 24 h was not evaluated in this study. ('211At-CXCR4', 'Var', (38, 49)) ('tumor', 'Disease', (99, 104)) ('tumor', 'Disease', 'MESH:D009369', (128, 133)) ('tumor', 'Phenotype', 'HP:0002664', (128, 133)) ('tumor', 'Disease', 'MESH:D009369', (99, 104)) ('tumor', 'Disease', (128, 133)) ('tumor', 'Phenotype', 'HP:0002664', (99, 104)) 2814 32321944 Although the highest %ID/g in the tumor was acquired at 6 h after the administration of 211At-CXCR4 mAb, it was still lower than those in the lung, heart, and kidneys. ('tumor', 'Disease', (34, 39)) ('tumor', 'Disease', 'MESH:D009369', (34, 39)) ('lower', 'NegReg', (118, 123)) ('tumor', 'Phenotype', 'HP:0002664', (34, 39)) ('211At-CXCR4 mAb', 'Var', (88, 103)) 2824 32321944 The biodistribution of a compound labeled with radioactive iodine, such as 123I and 125I, is assumed to be identical to that of an 211At-labeled compound. ('125I', 'Var', (84, 88)) ('123I', 'Var', (75, 79)) ('radioactive iodine', 'Chemical', '-', (47, 65)) 2831 32321944 Spatial and temporal changes in the distribution of 211At-CXCR4 mAb in the tumor should be accurately measured for accurate dosimetry. ('tumor', 'Phenotype', 'HP:0002664', (75, 80)) ('211At-CXCR4', 'Var', (52, 63)) ('tumor', 'Disease', (75, 80)) ('tumor', 'Disease', 'MESH:D009369', (75, 80)) 2840 32321944 This study suggests the possibility of stem-cell-targeted alpha-particle therapy using 211At-CXCR4 mAb for AML and supports further evaluation by therapeutic studies. ('211At-CXCR4', 'Var', (87, 98)) ('AML', 'Phenotype', 'HP:0004808', (107, 110)) ('AML', 'Disease', 'MESH:D015470', (107, 110)) ('AML', 'Disease', (107, 110)) 2880 32321944 Time-integrated activity was obtained on the basis of the tumor uptakes of 211At-CXCR4 mAb and 125I-CXCR4 mAb in the biodistribution study as mentioned above to integrate the activity in the tumor over time. ('tumor', 'Phenotype', 'HP:0002664', (58, 63)) ('tumor', 'Disease', (58, 63)) ('tumor', 'Disease', 'MESH:D009369', (191, 196)) ('tumor', 'Phenotype', 'HP:0002664', (191, 196)) ('tumor', 'Disease', 'MESH:D009369', (58, 63)) ('tumor', 'Disease', (191, 196)) ('211At-CXCR4', 'Var', (75, 86)) 2884 31500210 ABCB1 activity was observed in 58% of AML and was linked to low white blood cell count, high expression of CD34, absence of FLT3-ITD, and absence of mutant NPM1. ('activity', 'MPA', (6, 14)) ('NPM1', 'Gene', (156, 160)) ('AML', 'Disease', (38, 41)) ('white blood', 'CPA', (64, 75)) ('AML', 'Phenotype', 'HP:0004808', (38, 41)) ('FLT3', 'Gene', (124, 128)) ('low', 'NegReg', (60, 63)) ('absence', 'NegReg', (113, 120)) ('ABCB1', 'Gene', (0, 5)) ('NPM1', 'Gene', '4869', (156, 160)) ('mutant', 'Var', (149, 155)) ('expression', 'MPA', (93, 103)) ('absence', 'NegReg', (138, 145)) ('CD34', 'Gene', '947', (107, 111)) ('low white blood cell count', 'Phenotype', 'HP:0001882', (60, 86)) ('AML', 'Disease', 'MESH:D015470', (38, 41)) ('FLT3', 'Gene', '2322', (124, 128)) ('CD34', 'Gene', (107, 111)) 2889 31500210 Furthermore, ABCB1 was not directly linked to drug resistance, ABCB1 inhibition did not improve outcome of high ABCB1 AML patients and thus high ABCB1 may represent a passenger characteristic of high-risk AML. ('AML', 'Phenotype', 'HP:0004808', (118, 121)) ('AML', 'Phenotype', 'HP:0004808', (205, 208)) ('AML', 'Disease', (118, 121)) ('AML', 'Disease', (205, 208)) ('patients', 'Species', '9606', (122, 130)) ('drug resistance', 'Phenotype', 'HP:0020174', (46, 61)) ('high', 'Var', (107, 111)) ('AML', 'Disease', 'MESH:D015470', (118, 121)) ('AML', 'Disease', 'MESH:D015470', (205, 208)) 2901 31500210 Second generation specific ABCB1 inhibitor PSC833 failed to show clinical benefit in multiple clinical trials of AML. ('inhibitor PSC833', 'Var', (33, 49)) ('AML', 'Disease', 'MESH:D015470', (113, 116)) ('AML', 'Phenotype', 'HP:0004808', (113, 116)) ('PSC833', 'Var', (43, 49)) ('ABCB1', 'Gene', (27, 32)) ('AML', 'Disease', (113, 116)) 2904 31500210 Furthermore, we examined clinical outcome in high ABCB1 activity AML patients treated with the combination of the ABCB1 inhibitor quinine with standard chemotherapy and we investigated the role of ABCB1 in AML patients treated with GO plus chemotherapy. ('AML', 'Disease', (65, 68)) ('ABCB1', 'Gene', (50, 55)) ('AML', 'Phenotype', 'HP:0004808', (206, 209)) ('AML', 'Phenotype', 'HP:0004808', (65, 68)) ('quinine', 'Chemical', 'MESH:D011803', (130, 137)) ('high', 'Var', (45, 49)) ('AML', 'Disease', 'MESH:D015470', (206, 209)) ('AML', 'Disease', 'MESH:D015470', (65, 68)) ('patients', 'Species', '9606', (210, 218)) ('patients', 'Species', '9606', (69, 77)) ('ABCB1', 'Gene', (114, 119)) ('AML', 'Disease', (206, 209)) 2905 31500210 We used global gene expression to gain new insights into why high ABCB1 maybe associated to worse clinical outcome in AML. ('associated', 'Reg', (78, 88)) ('high', 'Var', (61, 65)) ('AML', 'Phenotype', 'HP:0004808', (118, 121)) ('AML', 'Disease', (118, 121)) ('ABCB1', 'Gene', (66, 71)) ('AML', 'Disease', 'MESH:D015470', (118, 121)) 2907 31500210 In our cohort of adult de novo AML, age at diagnosis was not higher in ABCB1 positive AML compared to ABCB1 negative AML (Figure S1A, Table 1), and no relation was observed between ABCB1 phenotype and cytogenetic classification (Figure S1B). ('AML', 'Phenotype', 'HP:0004808', (86, 89)) ('positive', 'Var', (77, 85)) ('AML', 'Disease', (117, 120)) ('AML', 'Disease', (86, 89)) ('ABCB1', 'Gene', (71, 76)) ('AML', 'Phenotype', 'HP:0004808', (31, 34)) ('AML', 'Phenotype', 'HP:0004808', (117, 120)) ('AML', 'Disease', 'MESH:D015470', (86, 89)) ('AML', 'Disease', 'MESH:D015470', (31, 34)) ('AML', 'Disease', 'MESH:D015470', (117, 120)) ('AML', 'Disease', (31, 34)) 2908 31500210 When compared among cytogenetic and molecular subtypes, ABCB1 activity is specifically low in AML with either mutant NPM1 or FLT3-ITD, and in AML with both markers. ('AML', 'Disease', (94, 97)) ('AML', 'Disease', (142, 145)) ('NPM1', 'Gene', '4869', (117, 121)) ('activity', 'MPA', (62, 70)) ('FLT3', 'Gene', (125, 129)) ('AML', 'Phenotype', 'HP:0004808', (142, 145)) ('AML', 'Phenotype', 'HP:0004808', (94, 97)) ('mutant', 'Var', (110, 116)) ('AML', 'Disease', 'MESH:D015470', (142, 145)) ('ABCB1', 'Gene', (56, 61)) ('low', 'NegReg', (87, 90)) ('AML', 'Disease', 'MESH:D015470', (94, 97)) ('NPM1', 'Gene', (117, 121)) ('FLT3', 'Gene', '2322', (125, 129)) 2910 31500210 When focused on the mutational status of CEBPA, NPM1, and FLT3, these analyses demonstrated that ABCB1 positivity is associated with the absence of both FLT3-ITD and NPM1 mutations (p = 0.006 and p < 0.001; Table 1). ('NPM1', 'Gene', '4869', (48, 52)) ('FLT3', 'Gene', '2322', (153, 157)) ('FLT3', 'Gene', (58, 62)) ('positivity', 'Var', (103, 113)) ('NPM1', 'Gene', '4869', (166, 170)) ('absence', 'NegReg', (137, 144)) ('FLT3', 'Gene', (153, 157)) ('CEBPA', 'Gene', '1050', (41, 46)) ('ABCB1', 'Gene', (97, 102)) ('NPM1', 'Gene', (48, 52)) ('mutations', 'Var', (171, 180)) ('CEBPA', 'Gene', (41, 46)) ('FLT3', 'Gene', '2322', (58, 62)) ('NPM1', 'Gene', (166, 170)) 2911 31500210 We found that positive ABCB1 activity was associated with high expression of CD34, specifically, in ABCB1 positive AML 85% of blasts express CD34 compared to ABCB1 negative AML in which only 12% of blasts express CD34 (p < 0.001; Table 1, Figure 1D). ('CD34', 'Gene', (141, 145)) ('AML', 'Disease', 'MESH:D015470', (115, 118)) ('positive', 'Var', (106, 114)) ('AML', 'Phenotype', 'HP:0004808', (173, 176)) ('AML', 'Phenotype', 'HP:0004808', (115, 118)) ('AML', 'Disease', (173, 176)) ('AML', 'Disease', (115, 118)) ('expression', 'MPA', (63, 73)) ('CD34', 'Gene', (77, 81)) ('activity', 'MPA', (29, 37)) ('CD34', 'Gene', '947', (77, 81)) ('CD34', 'Gene', '947', (141, 145)) ('CD34', 'Gene', '947', (213, 217)) ('ABCB1', 'Gene', (100, 105)) ('CD34', 'Gene', (213, 217)) ('AML', 'Disease', 'MESH:D015470', (173, 176)) ('ABCB1', 'Gene', (23, 28)) 2912 31500210 ABCB1 positivity was associated with low white blood cell count (WBC) median: 4.7 vs. 16.2 G/L found in ABCB1 negative AML (p = 0.011; Table 1). ('white blood cell count', 'MPA', (41, 63)) ('low', 'NegReg', (37, 40)) ('ABCB1', 'Gene', (0, 5)) ('positivity', 'Var', (6, 16)) ('AML', 'Disease', 'MESH:D015470', (119, 122)) ('low white blood cell count', 'Phenotype', 'HP:0001882', (37, 63)) ('AML', 'Phenotype', 'HP:0004808', (119, 122)) ('AML', 'Disease', (119, 122)) ('ABCB1', 'Gene', (104, 109)) 2914 31500210 Furthermore, negative ABCB1 activity occurred in AML carrying the NPM1 mutation. ('negative', 'NegReg', (13, 21)) ('AML', 'Disease', (49, 52)) ('ABCB1', 'Gene', (22, 27)) ('NPM1', 'Gene', '4869', (66, 70)) ('activity', 'MPA', (28, 36)) ('mutation', 'Var', (71, 79)) ('AML', 'Disease', 'MESH:D015470', (49, 52)) ('NPM1', 'Gene', (66, 70)) ('AML', 'Phenotype', 'HP:0004808', (49, 52)) 2916 31500210 Although not significant, lower CR rates (81%) were obtained in patients with positive ABCB1 AML compared to those with negative ABCB1 activity (91%) and ABCB1 activity was higher in non-responders vs. patients in CR (median V/E ratio: 2.2 vs. 1.5, p = 0.17, Figure 2A). ('AML', 'Phenotype', 'HP:0004808', (93, 96)) ('positive', 'Var', (78, 86)) ('AML', 'Disease', (93, 96)) ('patients', 'Species', '9606', (64, 72)) ('CR rates', 'CPA', (32, 40)) ('ABCB1', 'Gene', (87, 92)) ('patients', 'Species', '9606', (202, 210)) ('AML', 'Disease', 'MESH:D015470', (93, 96)) ('lower', 'NegReg', (26, 31)) 2918 31500210 In summary, high ABCB1 activity demonstrated an independent negative impact on event-free and overall survival of AML treated with standard chemotherapy. ('negative', 'NegReg', (60, 68)) ('AML', 'Disease', (114, 117)) ('high', 'Var', (12, 16)) ('AML', 'Phenotype', 'HP:0004808', (114, 117)) ('event-free', 'CPA', (79, 89)) ('overall survival', 'CPA', (94, 110)) ('ABCB1', 'Gene', (17, 22)) ('activity', 'MPA', (23, 31)) ('AML', 'Disease', 'MESH:D015470', (114, 117)) 2930 31500210 Negative ABCB1 AML were more frequently mutant NPM1 with low CD34 expression and high expression of HOX cluster genes. ('AML', 'Disease', 'MESH:D015470', (15, 18)) ('NPM1', 'Gene', (47, 51)) ('AML', 'Disease', (15, 18)) ('low', 'NegReg', (57, 60)) ('expression', 'MPA', (66, 76)) ('CD34', 'Gene', (61, 65)) ('expression', 'MPA', (86, 96)) ('NPM1', 'Gene', '4869', (47, 51)) ('CD34', 'Gene', '947', (61, 65)) ('AML', 'Phenotype', 'HP:0004808', (15, 18)) ('ABCB1', 'Gene', (9, 14)) ('mutant', 'Var', (40, 46)) 2940 31500210 Conversely, addition of GO showed no significant impact on survival in patients with high ABCB1 expression (Figure S2D-F). ('ABCB1', 'Gene', (90, 95)) ('patients', 'Species', '9606', (71, 79)) ('expression', 'MPA', (96, 106)) ('high', 'Var', (85, 89)) 2951 31500210 Similarly, disease relapse did not occur more frequently in positive ABCB1 AML. ('AML', 'Disease', 'MESH:D015470', (75, 78)) ('AML', 'Phenotype', 'HP:0004808', (75, 78)) ('AML', 'Disease', (75, 78)) ('ABCB1', 'Gene', (69, 74)) ('positive', 'Var', (60, 68)) 2954 31500210 Of note, we cannot rule out effects by ABCB1 single nucleotide polymorphism (SNP), even though mutations in ABCB1 in AML are rare . ('AML', 'Disease', (117, 120)) ('AML', 'Phenotype', 'HP:0004808', (117, 120)) ('single nucleotide polymorphism', 'Var', (45, 75)) ('AML', 'Disease', 'MESH:D015470', (117, 120)) ('ABCB1', 'Gene', (108, 113)) ('ABCB1', 'Gene', (39, 44)) 2957 31500210 Nevertheless, our larger study confirms the independent negative impact of high ABCB1 phenotype on AML clinical outcome in particular on EFS and OS. ('EFS', 'Disease', (137, 140)) ('high', 'Var', (75, 79)) ('AML', 'Disease', 'MESH:D015470', (99, 102)) ('AML', 'Phenotype', 'HP:0004808', (99, 102)) ('AML', 'Disease', (99, 102)) ('negative', 'NegReg', (56, 64)) ('ABCB1', 'Gene', (80, 85)) 2958 31500210 Furthermore, we confirmed the association of high ABCB1 phenotype with increased expression of CD34, a characteristic marker of the immature subset in AML blast cells. ('ABCB1', 'Gene', (50, 55)) ('AML', 'Disease', 'MESH:D015470', (151, 154)) ('CD34', 'Gene', (95, 99)) ('CD34', 'Gene', '947', (95, 99)) ('high', 'Var', (45, 49)) ('increased', 'PosReg', (71, 80)) ('AML', 'Disease', (151, 154)) ('AML', 'Phenotype', 'HP:0004808', (151, 154)) ('expression', 'MPA', (81, 91)) 2959 31500210 However, we newly identified that high ABCB1 activity was significantly associated with low WBC count and absence of FLT3-ITD and NPM1 mutations and this was not described in previous studies where mutational status of these genes was available. ('activity', 'MPA', (45, 53)) ('low WBC count', 'Phenotype', 'HP:0020060', (88, 101)) ('low', 'NegReg', (88, 91)) ('FLT3', 'Gene', (117, 121)) ('WBC count', 'CPA', (92, 101)) ('NPM1', 'Gene', (130, 134)) ('absence', 'NegReg', (106, 113)) ('mutations', 'Var', (135, 144)) ('NPM1', 'Gene', '4869', (130, 134)) ('ABCB1', 'Gene', (39, 44)) ('FLT3', 'Gene', '2322', (117, 121)) 2967 31500210 We then hypothesized that evaluation of ABCB1 co-expressed genes may uncover potential mechanisms explaining how high ABCB1 leads to treatment failure in AML. ('high', 'Var', (113, 117)) ('AML', 'Disease', 'MESH:D015470', (154, 157)) ('treatment', 'Disease', (133, 142)) ('AML', 'Disease', (154, 157)) ('AML', 'Phenotype', 'HP:0004808', (154, 157)) ('ABCB1', 'Gene', (118, 123)) 3009 31500210 The following are available online at , CONSORT diagram, Figure S1: ABCB1 phenotype in relation to age at diagnosis and cytogenetics, Figure S2: ABCB1 phenotype in relation to CD33 expression and clinical outcome in high ABCB1 AML treated with or without GO addition, and Figure S3: ABCB1 activity was determined by flow cytometry (MFC). ('AML', 'Phenotype', 'HP:0004808', (227, 230)) ('CO', 'Chemical', 'MESH:D002245', (40, 42)) ('AML', 'Disease', (227, 230)) ('CD33', 'Gene', '945', (176, 180)) ('CD33', 'Gene', (176, 180)) ('high', 'Var', (216, 220)) ('AML', 'Disease', 'MESH:D015470', (227, 230)) ('ABCB1', 'Gene', (221, 226)) 3023 31078321 Recently disclosed agents include the inhibitors, iCDK9, NVP-2, and MC180295, and a selective degrader, THAL-SNS-032 (Figure 2, Key Figure). ('CDK9', 'Gene', '1025', (51, 55)) ('MC180295', 'Var', (68, 76)) ('THAL-SNS-032', 'Chemical', '-', (104, 116)) ('CDK9', 'Gene', (51, 55)) ('degrader', 'NegReg', (94, 102)) 3032 31078321 BRD4 was first causally linked to cancer pathogenesis by the discovery that chimeric fusions of BRD4 to NUT (nuclear protein in testes) form oncogenes that drive a severe squamous cancer termed NUT midline carcinoma (NMC). ('cancer', 'Phenotype', 'HP:0002664', (34, 40)) ('drive', 'PosReg', (156, 161)) ('BRD4', 'Gene', (96, 100)) ('NUT midline carcinoma', 'Disease', (194, 215)) ('NUT', 'Gene', '256646', (194, 197)) ('NUT', 'Gene', (194, 197)) ('cancer', 'Disease', 'MESH:D009369', (34, 40)) ('NUT midline carcinoma', 'Disease', 'MESH:D009436', (194, 215)) ('NUT', 'Gene', '256646', (104, 107)) ('BRD4', 'Gene', (0, 4)) ('NUT', 'Gene', (104, 107)) ('cancer', 'Disease', (180, 186)) ('BRD4', 'Gene', '23476', (96, 100)) ('carcinoma', 'Phenotype', 'HP:0030731', (206, 215)) ('cancer', 'Phenotype', 'HP:0002664', (180, 186)) ('squamous cancer', 'Phenotype', 'HP:0002860', (171, 186)) ('chimeric fusions', 'Var', (76, 92)) ('squamous cancer', 'Disease', (171, 186)) ('BRD4', 'Gene', '23476', (0, 4)) ('cancer', 'Disease', (34, 40)) ('cancer', 'Disease', 'MESH:D009369', (180, 186)) ('squamous cancer', 'Disease', 'MESH:D002294', (171, 186)) 3035 31078321 Interestingly, wild-type BRD4 has also been identified as a non-oncogene addiction in a variety of human cancers. ('BRD4', 'Gene', '23476', (25, 29)) ('wild-type', 'Var', (15, 24)) ('cancers', 'Phenotype', 'HP:0002664', (105, 112)) ('cancers', 'Disease', (105, 112)) ('cancers', 'Disease', 'MESH:D009369', (105, 112)) ('BRD4', 'Gene', (25, 29)) ('cancer', 'Phenotype', 'HP:0002664', (105, 111)) ('human', 'Species', '9606', (99, 104)) 3036 31078321 Hematological malignancies, the first tumors shown to require wild-type BRD4, are hypersensitive to BET bromodomain inhibition, which has motivated a number of clinical trials that have reported preliminary signs of efficacy. ('BRD4', 'Gene', (72, 76)) ('tumor', 'Phenotype', 'HP:0002664', (38, 43)) ('tumors', 'Disease', (38, 44)) ('malignancies', 'Disease', 'MESH:D009369', (14, 26)) ('tumors', 'Phenotype', 'HP:0002664', (38, 44)) ('tumors', 'Disease', 'MESH:D009369', (38, 44)) ('BRD4', 'Gene', '23476', (72, 76)) ('malignancies', 'Disease', (14, 26)) ('Hematological malignancies', 'Phenotype', 'HP:0004377', (0, 26)) ('wild-type', 'Var', (62, 71)) ('hypersensitive', 'Disease', (82, 96)) ('hypersensitive', 'Disease', 'MESH:D004342', (82, 96)) ('bromodomain', 'Chemical', '-', (104, 115)) 3050 31078321 Recent application of the auxin-inducible degron system to selectively degrade BRD4 has elaborated on these results, demonstrating that like BET degradation, BRD4 degradation results in transcriptional collapse without affecting CDK9 occupancy. ('auxin', 'Chemical', 'MESH:D007210', (26, 31)) ('BRD4', 'Gene', '23476', (79, 83)) ('CDK9', 'Gene', (229, 233)) ('CDK9', 'Gene', '1025', (229, 233)) ('BRD4', 'Gene', (158, 162)) ('degradation', 'Var', (163, 174)) ('BRD4', 'Gene', (79, 83)) ('transcriptional collapse', 'MPA', (186, 210)) ('BRD4', 'Gene', '23476', (158, 162)) 3055 31078321 For example, genetic depletion of BRD2, which can act as an oncogene to drive B-cell malignancies, also diminishes growth of BRD4-dependent breast cancer and chronic lymphocytic leukemia cells, but not AML cells. ('BRD2', 'Gene', (34, 38)) ('cancer', 'Phenotype', 'HP:0002664', (147, 153)) ('BRD4', 'Gene', '23476', (125, 129)) ('chronic lymphocytic leukemia', 'Disease', 'MESH:D015451', (158, 186)) ('genetic depletion', 'Var', (13, 30)) ('breast cancer', 'Phenotype', 'HP:0003002', (140, 153)) ('chronic lymphocytic leukemia', 'Disease', (158, 186)) ('chronic lymphocytic leukemia', 'Phenotype', 'HP:0005550', (158, 186)) ('leukemia', 'Phenotype', 'HP:0001909', (178, 186)) ('breast cancer', 'Disease', 'MESH:D001943', (140, 153)) ('breast cancer', 'Disease', (140, 153)) ('BRD4', 'Gene', (125, 129)) ('malignancies', 'Disease', 'MESH:D009369', (85, 97)) ('growth', 'CPA', (115, 121)) ('BRD2', 'Gene', '6046', (34, 38)) ('diminishes', 'NegReg', (104, 114)) ('malignancies', 'Disease', (85, 97)) ('AML', 'Disease', 'MESH:D015470', (202, 205)) ('AML', 'Disease', (202, 205)) ('AML', 'Phenotype', 'HP:0004808', (202, 205)) 3068 31078321 While ENL is required for the survival of acute leukemia cells in vivo, normal hematopoietic stem and progenitors cells are insensitive to loss of ENL, suggesting ENL-targeted therapeutics might be able to elicit cancer-specific cell killing with a wide therapeutic window. ('acute leukemia', 'Disease', 'MESH:D015470', (42, 56)) ('acute leukemia', 'Disease', (42, 56)) ('acute leukemia', 'Phenotype', 'HP:0002488', (42, 56)) ('ENL', 'Gene', '4298', (147, 150)) ('cancer', 'Phenotype', 'HP:0002664', (213, 219)) ('ENL', 'Gene', '4298', (6, 9)) ('ENL', 'Gene', (147, 150)) ('leukemia', 'Phenotype', 'HP:0001909', (48, 56)) ('loss', 'Var', (139, 143)) ('ENL', 'Gene', '4298', (163, 166)) ('cancer', 'Disease', 'MESH:D009369', (213, 219)) ('ENL', 'Gene', (6, 9)) ('cancer', 'Disease', (213, 219)) ('ENL', 'Gene', (163, 166)) 3075 31078321 Coupling acute degradation of ENL to integrative transcriptional genomics, we identified a class of target genes - highly enriched for undruggable leukemic driver genes, including MYC and MYB - that are selectively downregulated by loss of ENL. ('downregulated', 'NegReg', (215, 228)) ('MYC', 'Gene', '4609', (180, 183)) ('ENL', 'Gene', (30, 33)) ('leukemic', 'Disease', (147, 155)) ('ENL', 'Gene', '4298', (240, 243)) ('MYC', 'Gene', (180, 183)) ('ENL', 'Gene', (240, 243)) ('MYB', 'Gene', '4602', (188, 191)) ('loss', 'Var', (232, 236)) ('leukemic', 'Disease', 'MESH:D007938', (147, 155)) ('MYB', 'Gene', (188, 191)) ('ENL', 'Gene', '4298', (30, 33)) 3080 31078321 The phenotypic effects of ENL degradation and BET bromodomain inhibition are similar in that they both affect promoter proximal pause release and preferentially suppress expression of tumorigenic transcription factors. ('degradation', 'Var', (30, 41)) ('ENL', 'Gene', (26, 29)) ('tumor', 'Disease', 'MESH:D009369', (184, 189)) ('bromodomain', 'Chemical', '-', (50, 61)) ('tumor', 'Phenotype', 'HP:0002664', (184, 189)) ('tumor', 'Disease', (184, 189)) ('suppress', 'NegReg', (161, 169)) ('promoter proximal pause release', 'MPA', (110, 141)) ('ENL', 'Gene', '4298', (26, 29)) ('expression', 'MPA', (170, 180)) ('affect', 'Reg', (103, 109)) 3089 31078321 Mutations to SMARCA4, which frequently occur in non-small-cell lung carcinomas (NSCLC), are synthetic lethal with loss of SMARCA2, which is otherwise non-essential for cell survival. ('SMARCA4', 'Gene', (13, 20)) ('NSCLC', 'Phenotype', 'HP:0030358', (80, 85)) ('lung carcinomas', 'Disease', (63, 78)) ('SMARCA2', 'Gene', (122, 129)) ('SMARCA4', 'Gene', '6597', (13, 20)) ('loss', 'NegReg', (114, 118)) ('carcinomas', 'Phenotype', 'HP:0030731', (68, 78)) ('NSCLC', 'Disease', (80, 85)) ('non-small-cell lung carcinomas', 'Phenotype', 'HP:0030358', (48, 78)) ('SMARCA2', 'Gene', '6595', (122, 129)) ('Mutations', 'Var', (0, 9)) ('lung carcinomas', 'Disease', 'MESH:D008175', (63, 78)) ('NSCLC', 'Disease', 'MESH:D002289', (80, 85)) ('carcinoma', 'Phenotype', 'HP:0030731', (68, 77)) ('small-cell lung carcinomas', 'Phenotype', 'HP:0030357', (52, 78)) 3100 31078321 dBRD9 (Figure 2), a PROTAC built via IMiD conjugation of BI-7273 is a potent and selective BRD9 degrader, which remarkably, does not degrade BRD7. ('BRD7', 'Gene', (141, 145)) ('BRD7', 'Gene', '29117', (141, 145)) ('BRD9', 'Gene', '65980', (1, 5)) ('IMiD', 'Chemical', '-', (37, 41)) ('BI-7273', 'Var', (57, 64)) ('degrader', 'NegReg', (96, 104)) ('BRD9', 'Gene', '65980', (91, 95)) ('BRD9', 'Gene', (1, 5)) ('BRD9', 'Gene', (91, 95)) ('BI-7273', 'Chemical', '-', (57, 64)) 3102 31078321 Together, these studies identified that sarcomas and rhabdoid tumors that are driven by mutations to BAF components are highly sensitive to loss of BRD9 via genetic disruption or exposure to dBRD9. ('rhabdoid tumors', 'Disease', (53, 68)) ('rhabdoid tumors', 'Disease', 'MESH:D018335', (53, 68)) ('tumor', 'Phenotype', 'HP:0002664', (62, 67)) ('BRD9', 'Gene', (148, 152)) ('tumors', 'Phenotype', 'HP:0002664', (62, 68)) ('BAF', 'Gene', '8815', (101, 104)) ('mutations', 'Var', (88, 97)) ('sarcomas', 'Disease', 'MESH:D012509', (40, 48)) ('BRD9', 'Gene', '65980', (192, 196)) ('sarcomas', 'Phenotype', 'HP:0100242', (40, 48)) ('BAF', 'Gene', (101, 104)) ('genetic disruption', 'Var', (157, 175)) ('loss', 'NegReg', (140, 144)) ('BRD9', 'Gene', (192, 196)) ('driven by', 'Reg', (78, 87)) ('BRD9', 'Gene', '65980', (148, 152)) ('sarcomas', 'Disease', (40, 48)) 3103 31078321 CREB binding protein (CBP) and p300 (adenovirus E1a associated 300 kDa protein) are two closely related histone acetyltransferases (HATs) that act as transcriptional co-activators by catalyzing the acylation of histone and non-histone lysine side chains (including H3K27ac, a modification found at promoters and putatively active enhancers). ('acetyl', 'Chemical', '-', (112, 118)) ('H3K27ac', 'Var', (265, 272)) ('adenovirus', 'Species', '10508', (37, 47)) ('lysine', 'Chemical', 'MESH:D008239', (235, 241)) ('CBP', 'Gene', (22, 25)) ('p300', 'Gene', (31, 35)) ('CBP', 'Gene', '1387', (22, 25)) ('acylation', 'MPA', (198, 207)) ('p300', 'Gene', '2033', (31, 35)) 3104 31078321 Owing to their synthetic lethal relationship, CBP and p300 have emerged as highly attractive cancer targets, particularly in certain lung cancers and lymphomas that frequently harbor mutations to these proteins. ('cancer', 'Disease', (93, 99)) ('cancer', 'Disease', (138, 144)) ('cancer', 'Disease', 'MESH:D009369', (93, 99)) ('lung cancers', 'Disease', 'MESH:D008175', (133, 145)) ('mutations', 'Var', (183, 192)) ('p300', 'Gene', '2033', (54, 58)) ('lymphomas', 'Phenotype', 'HP:0002665', (150, 159)) ('lung cancers', 'Phenotype', 'HP:0100526', (133, 145)) ('CBP', 'Gene', (46, 49)) ('cancer', 'Phenotype', 'HP:0002664', (93, 99)) ('cancer', 'Phenotype', 'HP:0002664', (138, 144)) ('CBP', 'Gene', '1387', (46, 49)) ('lymphomas', 'Disease', (150, 159)) ('cancers', 'Phenotype', 'HP:0002664', (138, 145)) ('lung cancers', 'Disease', (133, 145)) ('lymphomas', 'Disease', 'MESH:D008223', (150, 159)) ('cancer', 'Disease', 'MESH:D009369', (138, 144)) ('p300', 'Gene', (54, 58)) 3107 31078321 While none of the presently existing small-molecule inhibitors of CBP/p300 feature sufficient selectivity to exploit synthetic lethality between the two proteins, convergent chemical biology from separate groups using either A-485 or the CBP/p300 bromodomain inhibitor, GNE-272 (Figure 2), have revealed a selective dependency on CBP/p300 in androgen receptor-positive, but not androgen receptor-negative, prostate cancer growth. ('prostate cancer', 'Disease', (406, 421)) ('androgen receptor', 'Gene', '367', (378, 395)) ('cancer', 'Phenotype', 'HP:0002664', (415, 421)) ('androgen receptor', 'Gene', (342, 359)) ('bromodomain', 'Chemical', '-', (247, 258)) ('CBP/p300', 'Var', (330, 338)) ('prostate cancer', 'Disease', 'MESH:D011471', (406, 421)) ('prostate cancer', 'Phenotype', 'HP:0012125', (406, 421)) ('androgen receptor', 'Gene', (378, 395)) ('GNE-272', 'Chemical', '-', (270, 277)) ('androgen receptor', 'Gene', '367', (342, 359)) 3109 31078321 From both a therapeutic and basic biological perspective, it will be important to identify predictive biomarkers of sensitivity and mechanisms of transcriptional response to loss of CBP/p300-mediated lysine acetylation. ('lysine', 'Chemical', 'MESH:D008239', (200, 206)) ('acetyl', 'Chemical', '-', (207, 213)) ('CBP/p300-mediated', 'Gene', (182, 199)) ('lysine acetylation', 'MPA', (200, 218)) ('loss', 'Var', (174, 178)) 3110 31078321 Notably, CBP/p300 bromodomain inhibition has revealed that while the bromodomain is not required for chromatin localization, it is essential for CBP/p300-mediated H3K27 acetylation and enhancer activity. ('bromodomain', 'Chemical', '-', (69, 80)) ('acetyl', 'Chemical', '-', (169, 175)) ('enhancer', 'PosReg', (185, 193)) ('H3K27', 'Protein', (163, 168)) ('CBP/p300-mediated', 'Var', (145, 162)) ('bromodomain', 'Chemical', '-', (18, 29)) ('acetylation', 'MPA', (169, 180)) 3111 31078321 Moreover, CBP/p300 HAT inhibition coupled to quantitative proteomics have revealed that the repertoire of CBP/p300 substrates vastly exceeds a few histone and non-histone substrates; instead, CBP/p300 function through a so-called "acetyl-spray" mechanism, modifying thousands of sites at cis- regulatory elements. ('acetyl', 'Chemical', '-', (231, 237)) ('CBP/p300', 'Var', (192, 200)) ('modifying', 'Reg', (256, 265)) ('sites at cis- regulatory elements', 'MPA', (279, 312)) 3125 31078321 As comprehensively reviewed, EZH2 is implicated as an oncogene and non-oncogene dependency in a variety of cancers, owing to overexpression of EZH2, gain-of-function mutations to EZH2, and loss-of-function mutations to SWI/SNF complex subunits, which ordinarily oppose PRC2 function. ('EZH2', 'Gene', '2146', (143, 147)) ('mutations', 'Var', (206, 215)) ('cancers', 'Disease', (107, 114)) ('cancers', 'Disease', 'MESH:D009369', (107, 114)) ('EZH2', 'Gene', (179, 183)) ('EZH2', 'Gene', (143, 147)) ('mutations', 'Var', (166, 175)) ('EZH2', 'Gene', '2146', (179, 183)) ('cancer', 'Phenotype', 'HP:0002664', (107, 113)) ('loss-of-function', 'NegReg', (189, 205)) ('cancers', 'Phenotype', 'HP:0002664', (107, 114)) ('EZH2', 'Gene', '2146', (29, 33)) ('EZH2', 'Gene', (29, 33)) ('overexpression', 'PosReg', (125, 139)) ('SWI/SNF complex', 'Gene', (219, 234)) ('gain-of-function', 'PosReg', (149, 165)) 3127 31078321 Interestingly, EZH2 has also been reported to possess non-enzymatic, scaffolding function that is essential for the survival for SWI/SNF-mutant tumors. ('tumors', 'Disease', (144, 150)) ('tumors', 'Phenotype', 'HP:0002664', (144, 150)) ('tumors', 'Disease', 'MESH:D009369', (144, 150)) ('SWI/SNF-mutant', 'Var', (129, 143)) ('tumor', 'Phenotype', 'HP:0002664', (144, 149)) ('SWI/SNF-mutant', 'Gene', (129, 143)) ('EZH2', 'Gene', (15, 19)) ('EZH2', 'Gene', '2146', (15, 19)) 3128 31078321 In this context, genetic or chemical disruption of methyltransferase activity only partially exploits EZH2 dependency, presenting an attractive rationale to develop EZH2-selective PROTACs able to fully phenocopy genetic depletion. ('EZH2', 'Gene', (102, 106)) ('EZH2', 'Gene', '2146', (102, 106)) ('disruption', 'Var', (37, 47)) ('EZH2', 'Gene', (165, 169)) ('EZH2', 'Gene', '2146', (165, 169)) 3130 31078321 These structurally distinct small molecules, EED226 and A-395 (Figure 2), potently inhibit methyltransferase activity in vivo and excitingly, are able to overcome mutations to EZH2 that confer resistance to EZH2 HMT inhibitors. ('mutations', 'Var', (163, 172)) ('HMT', 'Gene', '3176', (212, 215)) ('resistance', 'MPA', (193, 203)) ('EED', 'Gene', '8726', (45, 48)) ('methyltransferase', 'Enzyme', (91, 108)) ('EZH2', 'Gene', '2146', (176, 180)) ('EZH2', 'Gene', '2146', (207, 211)) ('EED', 'Gene', (45, 48)) ('EZH2', 'Gene', (176, 180)) ('EZH2', 'Gene', (207, 211)) ('inhibit', 'NegReg', (83, 90)) ('HMT', 'Gene', (212, 215)) 3135 31078321 Small-molecule inhibitors of Menin-KMT2A are effective in several leukemic subtypes and are expected to enter clinical trials soon. ('Menin', 'Gene', (29, 34)) ('KMT2A', 'Gene', '4297', (35, 40)) ('leukemic', 'Disease', 'MESH:D007938', (66, 74)) ('Small-molecule', 'Var', (0, 14)) ('Menin', 'Gene', '4221', (29, 34)) ('leukemic', 'Disease', (66, 74)) ('KMT2A', 'Gene', (35, 40)) ('effective', 'Reg', (45, 54)) 3137 31078321 Additionally, the H3K36 HMT, NSD2, is known to be frequently translocated or mutated in various hematologic malignancies resulting in hyperactivity and increased sensitivity to loss of NSD2. ('HMT', 'Gene', (24, 27)) ('NSD2', 'Gene', '7468', (185, 189)) ('hyperactivity', 'Disease', 'MESH:D006948', (134, 147)) ('HMT', 'Gene', '3176', (24, 27)) ('increased', 'PosReg', (152, 161)) ('NSD2', 'Gene', (185, 189)) ('mutated', 'Var', (77, 84)) ('hyperactivity', 'Disease', (134, 147)) ('hyperactivity', 'Phenotype', 'HP:0000752', (134, 147)) ('sensitivity', 'MPA', (162, 173)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (96, 120)) ('NSD2', 'Gene', '7468', (29, 33)) ('NSD2', 'Gene', (29, 33)) ('hematologic malignancies', 'Disease', (96, 120)) 3165 31078321 After tagging a POI with FKBP12F36V, treatment with a small molecule, dTAG-13, induces association of the POI-FKBP12F36V to CRL4CRBN, resulting in degradation of the POI. ('dTAG-13', 'Chemical', '-', (70, 77)) ('CRBN', 'Gene', '51185', (128, 132)) ('association', 'Interaction', (87, 98)) ('CRBN', 'Gene', (128, 132)) ('POI-FKBP12F36V', 'Var', (106, 120)) ('degradation', 'MPA', (147, 158)) 3172 31078321 This concept was proposed based on the hypothesis that certain alterations present in a cancer cell may provoke a deficiency in, or additional need for, some function that does not affect physiological cell survival. ('cancer', 'Disease', (88, 94)) ('alterations', 'Var', (63, 74)) ('cancer', 'Disease', 'MESH:D009369', (88, 94)) ('function', 'MPA', (158, 166)) ('cancer', 'Phenotype', 'HP:0002664', (88, 94)) ('deficiency', 'NegReg', (114, 124)) 3174 31078321 Other non-oncogene dependencies may not be as easily defined by a single mutation, but by the combined action of many genetic, transcriptional, and/or post-transcriptional alterations that together, confer a cancer cell state expressing a unique vulnerability. ('confer', 'Reg', (199, 205)) ('cancer', 'Disease', (208, 214)) ('cancer', 'Phenotype', 'HP:0002664', (208, 214)) ('cancer', 'Disease', 'MESH:D009369', (208, 214)) ('alterations', 'Var', (172, 183)) 3181 31078321 A remarkable 20% of human cancers feature a mutation to a gene encoding a SWI/SNF subunit and many of these mutations present opportunities to exploit synthetic lethality for the development of anti-cancer therapeutics. ('human', 'Species', '9606', (20, 25)) ('cancers', 'Phenotype', 'HP:0002664', (26, 33)) ('mutation', 'Var', (44, 52)) ('cancer', 'Phenotype', 'HP:0002664', (199, 205)) ('cancer', 'Disease', (26, 32)) ('cancer', 'Disease', 'MESH:D009369', (26, 32)) ('cancers', 'Disease', 'MESH:D009369', (26, 33)) ('cancers', 'Disease', (26, 33)) ('cancer', 'Disease', 'MESH:D009369', (199, 205)) ('cancer', 'Phenotype', 'HP:0002664', (26, 32)) ('cancer', 'Disease', (199, 205)) 3184 31078321 Application of these small-molecule tools is opening new views into the function of co-regulators in transcriptional control and identifying underlying mechanisms by which these genes affect cancer-specific cell survival. ('cancer', 'Disease', (191, 197)) ('cancer', 'Disease', 'MESH:D009369', (191, 197)) ('affect', 'Reg', (184, 190)) ('genes', 'Var', (178, 183)) ('cancer', 'Phenotype', 'HP:0002664', (191, 197)) 3185 31417278 SOX30 methylation correlates with disease progression in patients with chronic myeloid leukemia Our previous study has reported that aberrant SOX30 methylation was associated with poor prognosis in AML, and it correlated with disease progression in MDS. ('AML', 'Disease', (199, 202)) ('aberrant', 'Var', (134, 142)) ('SOX30', 'Gene', (143, 148)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (80, 96)) ('SOX30', 'Gene', '11063', (1, 6)) ('methylation', 'MPA', (149, 160)) ('MDS', 'Disease', (250, 253)) ('MDS', 'Disease', 'MESH:D009190', (250, 253)) ('SOX30', 'Gene', (1, 6)) ('leukemia', 'Phenotype', 'HP:0001909', (88, 96)) ('chronic myeloid leukemia', 'Phenotype', 'HP:0005506', (72, 96)) ('chronic myeloid leukemia', 'Disease', 'MESH:D015464', (72, 96)) ('AML', 'Disease', 'MESH:D015470', (199, 202)) ('patients', 'Species', '9606', (58, 66)) ('chronic myeloid leukemia', 'Disease', (72, 96)) ('SOX30', 'Gene', '11063', (143, 148)) ('correlated with', 'Reg', (211, 226)) 3186 31417278 Herein, we further determined SOX30 methylation and its clinical significance in the other myeloid malignance - chronic myeloid leukemia (CML). ('chronic myeloid leukemia', 'Phenotype', 'HP:0005506', (112, 136)) ('chronic myeloid leukemia', 'Disease', 'MESH:D015464', (112, 136)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (120, 136)) ('chronic myeloid leukemia', 'Disease', (112, 136)) ('CML', 'Disease', 'MESH:D015464', (138, 141)) ('leukemia', 'Phenotype', 'HP:0001909', (128, 136)) ('SOX30', 'Gene', '11063', (30, 35)) ('SOX30', 'Gene', (30, 35)) ('methylation', 'Var', (36, 47)) ('CML', 'Disease', (138, 141)) 3188 31417278 SOX30 methylation was identified in 11% (10/95) CML patients. ('CML', 'Disease', 'MESH:D015464', (48, 51)) ('methylation', 'Var', (6, 17)) ('SOX30', 'Gene', '11063', (0, 5)) ('SOX30', 'Gene', (0, 5)) ('CML', 'Disease', (48, 51)) ('patients', 'Species', '9606', (52, 60)) 3189 31417278 SOX30 methylation was associated with lower hemoglobin and platelets (P=0.006 and 0.032, respectively). ('lower', 'NegReg', (38, 43)) ('lower hemoglobin', 'Phenotype', 'HP:0001903', (38, 54)) ('methylation', 'Var', (6, 17)) ('hemoglobin', 'MPA', (44, 54)) ('SOX30', 'Gene', '11063', (0, 5)) ('SOX30', 'Gene', (0, 5)) 3190 31417278 The frequency of SOX30 methylation in chronic phase (CP) stage occurred with lowest frequency (4/74, 5%), higher in accelerated phase (AP) stage (1/7, 14%), and the highest in blast crisis (BC) stage (12/31, 39%). ('blast crisis', 'Disease', (176, 188)) ('SOX30', 'Gene', (17, 22)) ('methylation', 'Var', (23, 34)) ('chronic phase', 'Disease', (38, 51)) ('higher', 'PosReg', (106, 112)) ('SOX30', 'Gene', '11063', (17, 22)) ('accelerated phase', 'Disease', (116, 133)) 3194 31417278 Our study revealed that SOX30 methylation correlated with disease progression in chronic myeloid leukemia. ('chronic myeloid leukemia', 'Disease', (81, 105)) ('methylation', 'Var', (30, 41)) ('correlated with', 'Reg', (42, 57)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (89, 105)) ('leukemia', 'Phenotype', 'HP:0001909', (97, 105)) ('chronic myeloid leukemia', 'Disease', 'MESH:D015464', (81, 105)) ('chronic myeloid leukemia', 'Phenotype', 'HP:0005506', (81, 105)) ('SOX30', 'Gene', '11063', (24, 29)) ('SOX30', 'Gene', (24, 29)) 3200 31417278 Recently, aberrant DNA methylation through the activation of tumor suppressor genes playing a role in the progression of CML has been aroused great attention. ('DNA methylation', 'MPA', (19, 34)) ('aberrant', 'Var', (10, 18)) ('activation', 'PosReg', (47, 57)) ('tumor', 'Disease', 'MESH:D009369', (61, 66)) ('CML', 'Disease', 'MESH:D015464', (121, 124)) ('tumor', 'Phenotype', 'HP:0002664', (61, 66)) ('tumor', 'Disease', (61, 66)) ('CML', 'Disease', (121, 124)) 3203 31417278 Recently, SOX30 has been identified as a key tumor suppressor gene mediated by promoter methylation in tumorigenesis including lung cancer, acute myeloid leukemia (AML), and myelodysplastic syndromes (MDS). ('tumor', 'Phenotype', 'HP:0002664', (103, 108)) ('lung cancer', 'Disease', (127, 138)) ('tumor', 'Disease', (45, 50)) ('MDS', 'Disease', 'MESH:D009190', (201, 204)) ('myelodysplastic syndromes', 'Phenotype', 'HP:0002863', (174, 199)) ('myelodysplastic syndromes', 'Disease', (174, 199)) ('tumor', 'Disease', 'MESH:D009369', (45, 50)) ('SOX30', 'Gene', (10, 15)) ('MDS', 'Disease', (201, 204)) ('myelodysplastic syndromes', 'Disease', 'MESH:D009190', (174, 199)) ('acute myeloid leukemia', 'Disease', (140, 162)) ('lung cancer', 'Disease', 'MESH:D008175', (127, 138)) ('tumor', 'Phenotype', 'HP:0002664', (45, 50)) ('lung cancer', 'Phenotype', 'HP:0100526', (127, 138)) ('tumor', 'Disease', (103, 108)) ('AML', 'Disease', 'MESH:D015470', (164, 167)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (146, 162)) ('AML', 'Disease', (164, 167)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (140, 162)) ('tumor', 'Disease', 'MESH:D009369', (103, 108)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (140, 162)) ('leukemia', 'Phenotype', 'HP:0001909', (154, 162)) ('cancer', 'Phenotype', 'HP:0002664', (132, 138)) ('SOX30', 'Gene', '11063', (10, 15)) ('promoter methylation', 'Var', (79, 99)) 3205 31417278 Moreover, aberrant SOX30 methylation was associated with poor prognosis in AML, whereas it correlated with disease progression in MDS. ('AML', 'Disease', 'MESH:D015470', (75, 78)) ('SOX30', 'Gene', '11063', (19, 24)) ('aberrant', 'Var', (10, 18)) ('AML', 'Disease', (75, 78)) ('methylation', 'Var', (25, 36)) ('SOX30', 'Gene', (19, 24)) ('MDS', 'Disease', (130, 133)) ('MDS', 'Disease', 'MESH:D009190', (130, 133)) ('correlated with', 'Reg', (91, 106)) 3206 31417278 In this study, we further determined SOX30 methylation and its clinical significance in CML. ('methylation', 'Var', (43, 54)) ('CML', 'Disease', (88, 91)) ('SOX30', 'Gene', '11063', (37, 42)) ('SOX30', 'Gene', (37, 42)) ('CML', 'Disease', 'MESH:D015464', (88, 91)) 3218 31417278 Our previous study has reported aberrant SOX30 methylation in AML and MDS, and SOX30 hypermethylation correlated with poor prognosis in AML, also associated with disease progression in MDS. ('hypermethylation', 'Var', (85, 101)) ('MDS', 'Disease', 'MESH:D009190', (185, 188)) ('SOX30', 'Gene', (79, 84)) ('SOX30', 'Gene', '11063', (41, 46)) ('SOX30', 'Gene', (41, 46)) ('SOX30', 'Gene', '11063', (79, 84)) ('MDS', 'Disease', (70, 73)) ('MDS', 'Disease', 'MESH:D009190', (70, 73)) ('AML', 'Disease', 'MESH:D015470', (62, 65)) ('AML', 'Disease', 'MESH:D015470', (136, 139)) ('associated with', 'Reg', (146, 161)) ('aberrant', 'Var', (32, 40)) ('AML', 'Disease', (62, 65)) ('methylation', 'MPA', (47, 58)) ('AML', 'Disease', (136, 139)) ('MDS', 'Disease', (185, 188)) 3221 31417278 In order to analyze the clinical significance in CML, we further compared the clinical and laboratory features between SOX30 methylated and non-methylated patients (Table 1). ('CML', 'Disease', 'MESH:D015464', (49, 52)) ('patients', 'Species', '9606', (155, 163)) ('methylated', 'Var', (125, 135)) ('SOX30', 'Gene', '11063', (119, 124)) ('CML', 'Disease', (49, 52)) ('SOX30', 'Gene', (119, 124)) 3222 31417278 However, SOX30 methylation was associated with lower hemoglobin and platelets (P=0.006 and 0.032, respectively). ('methylation', 'Var', (15, 26)) ('lower', 'NegReg', (47, 52)) ('lower hemoglobin', 'Phenotype', 'HP:0001903', (47, 63)) ('SOX30', 'Gene', '11063', (9, 14)) ('hemoglobin', 'MPA', (53, 63)) ('SOX30', 'Gene', (9, 14)) 3223 31417278 Among different clinical stages, the frequency of SOX30 methylation in chronic phase (CP) stage occurred with lowest frequency (4/74, 5%), higher in accelerated phase (AP) stage (1/7, 14%), and the highest in blast crisis (BC) stage (12/31, 39%). ('accelerated phase', 'Disease', (149, 166)) ('blast crisis', 'Disease', (209, 221)) ('SOX30', 'Gene', '11063', (50, 55)) ('chronic phase', 'Disease', (71, 84)) ('methylation', 'Var', (56, 67)) ('SOX30', 'Gene', (50, 55)) ('higher', 'PosReg', (139, 145)) 3224 31417278 For cytogenetics, t(9;22) with additional alterations patients showed the highest incidence of SOX30 methylation (4/11, 36%), whereas cases with t(9;22) patients presented the lowest incidence of SOX30 methylation (2/63, 3%). ('SOX30', 'Gene', '11063', (196, 201)) ('SOX30', 'Gene', (196, 201)) ('patients', 'Species', '9606', (153, 161)) ('alterations', 'Var', (42, 53)) ('patients', 'Species', '9606', (54, 62)) ('SOX30', 'Gene', '11063', (95, 100)) ('SOX30', 'Gene', (95, 100)) 3225 31417278 All the positive results indicated that SOX30 methylation was associated increased clinical stage and may play a role in disease progression in CML. ('clinical', 'Disease', (83, 91)) ('methylation', 'Var', (46, 57)) ('play', 'Reg', (106, 110)) ('role', 'Reg', (113, 117)) ('CML', 'Disease', 'MESH:D015464', (144, 147)) ('SOX30', 'Gene', '11063', (40, 45)) ('SOX30', 'Gene', (40, 45)) ('CML', 'Disease', (144, 147)) 3228 31417278 As is well known, DNA methylation in promoter-associated CpG islands plays a direct role in gene silencing that is one of the drivers of neoplastic transformation through the inactivation of critical tumor-suppressor pathways. ('tumor', 'Disease', 'MESH:D009369', (200, 205)) ('inactivation', 'NegReg', (175, 187)) ('DNA', 'Var', (18, 21)) ('tumor', 'Phenotype', 'HP:0002664', (200, 205)) ('gene', 'MPA', (92, 96)) ('tumor', 'Disease', (200, 205)) ('methylation', 'Var', (22, 33)) 3235 31417278 Mechanically, cytogenetic abnormalities and molecular alterations, such as double Ph chromosome, trisomy chromosome 8, i(17q), trisomy chromosome 19, t(3;21)(q26;q22), t(7;11)(p15;p15), p53 mutations, RAS mutation, and increased BCR-ABL transcript, are pathogenetically linked to the progression of CML. ('t(3;21)(q26;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (150, 166)) ('p53', 'Gene', (186, 189)) ('trisomy chromosome 19', 'Disease', (127, 148)) ('increased', 'PosReg', (219, 228)) ('mutations', 'Var', (190, 199)) ('BCR-ABL', 'Gene', '25', (229, 236)) ('t(7;11)(p15;p15)', 'STRUCTURAL_ABNORMALITY', 'None', (168, 184)) ('transcript', 'MPA', (237, 247)) ('linked', 'Reg', (270, 276)) ('t(3;21)(q26;q22', 'Var', (150, 165)) ('t(7;11)(p15;p15', 'Var', (168, 183)) ('CML', 'Disease', 'MESH:D015464', (299, 302)) ('RAS', 'Gene', (201, 204)) ('CML', 'Disease', (299, 302)) ('double Ph chromosome', 'Var', (75, 95)) ('mutation', 'Var', (205, 213)) ('trisomy chromosome 19', 'Disease', 'MESH:C538311', (127, 148)) ('BCR-ABL', 'Gene', (229, 236)) ('p53', 'Gene', '7157', (186, 189)) 3236 31417278 Recently, epigenetic changes especially aberrant DNA methylation associated with gene silencing are also identified to play crucial roles in the disease progression of CML. ('aberrant DNA methylation', 'Var', (40, 64)) ('CML', 'Disease', (168, 171)) ('roles', 'Reg', (132, 137)) ('CML', 'Disease', 'MESH:D015464', (168, 171)) 3237 31417278 For instance, SHP-1 hypermethylation was involved in the progression in CML through dysregulating BCR-ABL1, AKT, MAPK, MYC and JAK2/STAT5 signaling pathways. ('AKT', 'Gene', '207', (108, 111)) ('CML', 'Disease', (72, 75)) ('JAK2', 'Gene', '3717', (127, 131)) ('hypermethylation', 'Var', (20, 36)) ('STAT5', 'Gene', '6776', (132, 137)) ('MAPK', 'Pathway', (113, 117)) ('AKT', 'Gene', (108, 111)) ('involved', 'Reg', (41, 49)) ('MYC', 'Gene', '4609', (119, 122)) ('dysregulating', 'MPA', (84, 97)) ('BCR-ABL1', 'Gene', (98, 106)) ('JAK2', 'Gene', (127, 131)) ('SHP-1', 'Gene', (14, 19)) ('CML', 'Disease', 'MESH:D015464', (72, 75)) ('STAT5', 'Gene', (132, 137)) ('MYC', 'Gene', (119, 122)) ('SHP-1', 'Gene', '8431', (14, 19)) ('BCR-ABL1', 'Gene', '613;25', (98, 106)) 3238 31417278 In addition, our previous studies have revealed that hypermethylation of ID4 and DLX4 was related to disease progression in CML, and ID4 also had a direct role in affecting cell proliferation and apoptosis in K562 cell-line. ('affecting', 'Reg', (163, 172)) ('ID4', 'Gene', '3400', (73, 76)) ('ID4', 'Gene', (73, 76)) ('K562', 'CellLine', 'CVCL:0004', (209, 213)) ('DLX4', 'Gene', '1748', (81, 85)) ('ID4', 'Gene', (133, 136)) ('related to', 'Reg', (90, 100)) ('CML', 'Disease', 'MESH:D015464', (124, 127)) ('ID4', 'Gene', '3400', (133, 136)) ('apoptosis', 'CPA', (196, 205)) ('hypermethylation', 'Var', (53, 69)) ('DLX4', 'Gene', (81, 85)) ('cell proliferation', 'CPA', (173, 191)) ('CML', 'Disease', (124, 127)) 3239 31417278 Although we have proved that SOX30 methylation was associated with CML progression in clinics, the direct role of SOX30 in the pathogenesis of CML was not studied. ('CML', 'Disease', (67, 70)) ('SOX30', 'Gene', '11063', (114, 119)) ('CML', 'Disease', 'MESH:D015464', (143, 146)) ('SOX30', 'Gene', '11063', (29, 34)) ('SOX30', 'Gene', (114, 119)) ('associated', 'Reg', (51, 61)) ('SOX30', 'Gene', (29, 34)) ('CML', 'Disease', (143, 146)) ('methylation', 'Var', (35, 46)) ('CML', 'Disease', 'MESH:D015464', (67, 70)) 3249 31417278 In summary, SOX30 methylation correlated with disease progression in CML, and provided novel insights into CML biology acting as a potential therapeutic target against disease progression. ('correlated with', 'Reg', (30, 45)) ('disease', 'Disease', (46, 53)) ('SOX30', 'Gene', (12, 17)) ('SOX30', 'Gene', '11063', (12, 17)) ('CML', 'Disease', 'MESH:D015464', (69, 72)) ('CML', 'Disease', 'MESH:D015464', (107, 110)) ('methylation', 'Var', (18, 29)) ('CML', 'Disease', (69, 72)) ('CML', 'Disease', (107, 110)) 3297 30854206 Hematochezia resolved during hospitalization and was attributed to disseminated intravascular coagulation (DIC) secondary to APML. ('disseminated', 'Disease', (67, 79)) ('disseminated intravascular coagulation', 'Phenotype', 'HP:0005521', (67, 105)) ('Hematochezia', 'Disease', (0, 12)) ('intravascular coagulation', 'Disease', 'MESH:D004211', (80, 105)) ('APML', 'Phenotype', 'HP:0004836', (125, 129)) ('Hematochezia', 'Disease', 'MESH:D006471', (0, 12)) ('APML', 'Var', (125, 129)) ('Hematochezia', 'Phenotype', 'HP:0002573', (0, 12)) ('intravascular coagulation', 'Disease', (80, 105)) ('DIC', 'Phenotype', 'HP:0005521', (107, 110)) 3322 30259761 Dysregulation of protein kinases is implicated in various processes of carcinogenesis. ('Dysregulation', 'Var', (0, 13)) ('carcinogenesis', 'Disease', (71, 85)) ('implicated', 'Reg', (36, 46)) ('protein kinases', 'Enzyme', (17, 32)) ('carcinogenesis', 'Disease', 'MESH:D063646', (71, 85)) 3329 30259761 Phosphorylation results in a functional change of the target protein (substrate) by regulating signaling pathways by amplification (common) or cellular location, or by interactions with regulatory proteins. ('signaling pathways', 'Pathway', (95, 113)) ('Ph', 'Chemical', 'MESH:C084701', (0, 2)) ('change', 'Reg', (40, 46)) ('Phosphorylation', 'Var', (0, 15)) ('interactions', 'Interaction', (168, 180)) ('functional', 'MPA', (29, 39)) ('amplification', 'MPA', (117, 130)) ('regulating', 'Reg', (84, 94)) 3334 30259761 Phosphorylation of the target proteins by kinases is tightly regulated and any perturbation to this regulation may lead to a diseased state. ('Ph', 'Chemical', 'MESH:C084701', (0, 2)) ('Phosphorylation', 'MPA', (0, 15)) ('diseased state', 'Disease', (125, 139)) ('perturbation', 'Var', (79, 91)) ('lead to', 'Reg', (115, 122)) 3348 30259761 Since then, it has been approved for the treatment of rare hematologic malignancies and proto-oncogene c-Kit or tyrosine-protein kinase Kit (c-Kit) mutated gastrointestinal stromal tumors (GIST). ('Kit', 'Gene', '3815', (143, 146)) ('hematologic malignancies', 'Disease', (59, 83)) ('Kit', 'Gene', '3815', (136, 139)) ('gastrointestinal stromal tumor', 'Phenotype', 'HP:0100723', (156, 186)) ('tumor', 'Phenotype', 'HP:0002664', (181, 186)) ('tumors', 'Phenotype', 'HP:0002664', (181, 187)) ('tyrosine', 'Chemical', 'None', (112, 120)) ('Kit', 'Gene', (105, 108)) ('mutated', 'Var', (148, 155)) ('proto-oncogene c-Kit', 'Gene', '3815', (88, 108)) ('gastrointestinal stromal tumors', 'Disease', (156, 187)) ('Kit', 'Gene', '3815', (105, 108)) ('gastrointestinal stromal tumors', 'Phenotype', 'HP:0100723', (156, 187)) ('Kit', 'Gene', (136, 139)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (59, 83)) ('proto-oncogene c-Kit', 'Gene', (88, 108)) ('Kit', 'Gene', (143, 146)) ('gastrointestinal stromal tumors', 'Disease', 'MESH:D046152', (156, 187)) 3350 30259761 In the median follow up of 19 months, in the intention to treat analysis, imatinib group achieved significantly higher complete hematologic responses in 95.3 % [95% confidence interval (CI) 93.2-96.9] of patients compared to 27.3 % (95% CI 6.0-61.0) of patients in the interferon plus cytarabine group. ('patients', 'Species', '9606', (204, 212)) ('higher', 'PosReg', (112, 118)) ('patients', 'Species', '9606', (253, 261)) ('imatinib', 'Chemical', 'MESH:C097613', (74, 82)) ('imatinib', 'Var', (74, 82)) ('complete', 'MPA', (119, 127)) ('cytarabine', 'Chemical', 'MESH:D003561', (285, 295)) 3360 30259761 It was designed to overcome the imatinib resistance by binding to the kinase domain of imatinib-resistant mutants of Bcr-Abl and imatinib sensitive Bcr-Abl with higher affinity. ('imatinib', 'Chemical', 'MESH:C097613', (32, 40)) ('Bcr-Abl', 'Gene', (117, 124)) ('Bcr-Abl', 'Gene', '25', (117, 124)) ('imatinib', 'Chemical', 'MESH:C097613', (87, 95)) ('Bcr-Abl', 'Gene', (148, 155)) ('Bcr-Abl', 'Gene', '25', (148, 155)) ('imatinib', 'Chemical', 'MESH:C097613', (129, 137)) ('binding', 'Interaction', (55, 62)) ('mutants', 'Var', (106, 113)) 3361 30259761 In vitro nilotinib inhibited the kinase activity of most of the 15 common Bcr-Abl mutants except for T315I gatekeeper mutant enzyme. ('inhibited', 'NegReg', (19, 28)) ('Bcr-Abl', 'Gene', '25', (74, 81)) ('T315I', 'Var', (101, 106)) ('kinase activity', 'MPA', (33, 48)) ('nilotinib', 'Chemical', 'MESH:C498826', (9, 18)) ('T315I', 'SUBSTITUTION', 'None', (101, 106)) ('Bcr-Abl', 'Gene', (74, 81)) ('mutants', 'Var', (82, 89)) ('gatekeeper', 'Species', '111938', (107, 117)) 3381 30259761 In vitro, dasatinib inhibited the kinase activity of 14 of 15 Bcr-Abl mutant proteins except for the gatekeeper T315I mutant kinase. ('proteins', 'Protein', (77, 85)) ('Bcr-Abl', 'Gene', (62, 69)) ('T315I', 'SUBSTITUTION', 'None', (112, 117)) ('Bcr-Abl', 'Gene', '25', (62, 69)) ('gatekeeper', 'Species', '111938', (101, 111)) ('dasatinib', 'Chemical', 'MESH:C488369', (10, 19)) ('mutant', 'Var', (70, 76)) ('kinase activity', 'MPA', (34, 49)) ('inhibited', 'NegReg', (20, 29)) ('T315I', 'Var', (112, 117)) 3385 30259761 When dasatinib was compared with imatinib as first-line therapy in chronic phase CML, like nilotinib, dasatinib achieved better cytogenic responses and lesser transformation to accelerated or blast phase. ('dasatinib', 'Chemical', 'MESH:C488369', (102, 111)) ('nilotinib', 'Chemical', 'MESH:C498826', (91, 100)) ('dasatinib', 'Var', (102, 111)) ('CML', 'Disease', (81, 84)) ('blast phase', 'CPA', (192, 203)) ('better', 'PosReg', (121, 127)) ('imatinib', 'Chemical', 'MESH:C097613', (33, 41)) ('cytogenic responses', 'CPA', (128, 147)) ('dasatinib', 'Chemical', 'MESH:C488369', (5, 14)) 3389 30259761 Like dasatinib, it is a dual Src-Abl kinase inhibitor, more potent than imatinib in inhibiting Bcr-Abl and has activity against almost all imatinib-resistant Bcr-Abl except for T315I mutant kinase. ('imatinib', 'Chemical', 'MESH:C097613', (72, 80)) ('Bcr-Abl', 'Gene', '25', (158, 165)) ('c-Abl', 'Gene', (31, 36)) ('imatinib', 'Chemical', 'MESH:C097613', (139, 147)) ('activity', 'MPA', (111, 119)) ('c-Abl', 'Gene', '25', (31, 36)) ('Bcr-Abl', 'Gene', (95, 102)) ('dasatinib', 'Chemical', 'MESH:C488369', (5, 14)) ('inhibiting', 'NegReg', (84, 94)) ('T315I', 'Var', (177, 182)) ('Bcr-Abl', 'Gene', (158, 165)) ('Bcr-Abl', 'Gene', '25', (95, 102)) ('T315I', 'SUBSTITUTION', 'None', (177, 182)) 3398 30259761 Previous Bcr-Abl kinase inhibitors including second and third generation inhibitors were not able to bind to the T315I mutant kinase due to steric hindrance caused by the bulky isoleucine residue at position 315 in the T315I mutant kinase. ('T315I', 'SUBSTITUTION', 'None', (113, 118)) ('T315I', 'Var', (113, 118)) ('T315I', 'Var', (219, 224)) ('T315I', 'SUBSTITUTION', 'None', (219, 224)) ('Bcr-Abl', 'Gene', (9, 16)) ('Bcr-Abl', 'Gene', '25', (9, 16)) ('isoleucine', 'Chemical', 'MESH:C043801', (177, 187)) 3400 30259761 As expected it was active against all the mutant forms of Bcr-Abl in vitro. ('Bcr-Abl', 'Gene', (58, 65)) ('mutant', 'Var', (42, 48)) ('Bcr-Abl', 'Gene', '25', (58, 65)) 3401 30259761 Ponatinib was found to show response in patients in whom previous therapy had failed, in patients who harbour a T315I mutation, and in patients who are refractory to therapy with multiple TKIs in the absence of detectable Bcr-Abl mutations. ('Bcr-Abl', 'Gene', '25', (222, 229)) ('T315I', 'SUBSTITUTION', 'None', (112, 117)) ('patients', 'Species', '9606', (135, 143)) ('patients', 'Species', '9606', (40, 48)) ('Ponatinib', 'Chemical', 'MESH:C545373', (0, 9)) ('patients', 'Species', '9606', (89, 97)) ('Bcr-Abl', 'Gene', (222, 229)) ('T315I', 'Var', (112, 117)) 3403 30259761 Interestingly patients who eventually developed resistance to ponatinib did not have Bcr-Abl point mutations, they rather had compound mutations especially patients in blast phase, raising an important question as to whether ponatinib should be used as the front line to avoid the development of resistant clones. ('patients', 'Species', '9606', (156, 164)) ('ponatinib', 'Chemical', 'MESH:C545373', (225, 234)) ('had', 'Reg', (122, 125)) ('Bcr-Abl', 'Gene', '25', (85, 92)) ('Bcr-Abl', 'Gene', (85, 92)) ('patients', 'Species', '9606', (14, 22)) ('ponatinib', 'Chemical', 'MESH:C545373', (62, 71)) ('compound mutations', 'Var', (126, 144)) 3413 30259761 EGFR pathway is dysregulated in multiple cancers: EGFR is overexpressed in ~80% of non-small cell lung cancer (NSCLC) and mutated in 20% of NSCLC. ('NSCLC', 'Disease', (111, 116)) ('cancers', 'Phenotype', 'HP:0002664', (41, 48)) ('EGFR', 'Gene', (50, 54)) ('cancers', 'Disease', (41, 48)) ('overexpressed', 'PosReg', (58, 71)) ('NSCLC', 'Disease', 'MESH:D002289', (140, 145)) ('cancer', 'Phenotype', 'HP:0002664', (103, 109)) ('cancer', 'Phenotype', 'HP:0002664', (41, 47)) ('lung cancer', 'Phenotype', 'HP:0100526', (98, 109)) ('non-small cell lung cancer', 'Phenotype', 'HP:0030358', (83, 109)) ('small cell lung cancer', 'Phenotype', 'HP:0030357', (87, 109)) ('NSCLC', 'Disease', (140, 145)) ('EGFR', 'Gene', (0, 4)) ('EGFR', 'Gene', '1956', (50, 54)) ('non-small cell lung cancer', 'Disease', 'MESH:D002289', (83, 109)) ('cancers', 'Disease', 'MESH:D009369', (41, 48)) ('mutated', 'Var', (122, 129)) ('NSCLC', 'Disease', 'MESH:D002289', (111, 116)) ('non-small cell lung cancer', 'Disease', (83, 109)) ('EGFR', 'Gene', '1956', (0, 4)) 3420 30259761 Two landmark articles showed that patients who responded to gefitinib had mutated EGFR genes specifically exon 19 deletions or exon 21 (L858R) substitution mutation, thus these EGFR mutations became the predictive marker for EGFR TKI therapies. ('EGFR', 'Gene', (82, 86)) ('L858R', 'Var', (136, 141)) ('EGFR', 'Gene', '1956', (225, 229)) ('EGFR', 'Gene', '1956', (177, 181)) ('patients', 'Species', '9606', (34, 42)) ('EGFR', 'Gene', (225, 229)) ('EGFR', 'Gene', '1956', (82, 86)) ('L858R', 'SUBSTITUTION', 'None', (136, 141)) ('substitution mutation', 'Var', (143, 164)) ('EGFR', 'Gene', (177, 181)) ('gefitinib', 'Chemical', 'MESH:C419708', (60, 69)) 3421 30259761 Currently, gefitinib is approved as first-line therapy in patients with EGFR exon 19 deletions or exon 21 (L858R) substitution mutation-positive metastatic NSCLC. ('L858R', 'SUBSTITUTION', 'None', (107, 112)) ('substitution mutation-positive', 'Var', (114, 144)) ('gefitinib', 'Chemical', 'MESH:C419708', (11, 20)) ('deletions', 'Var', (85, 94)) ('EGFR', 'Gene', '1956', (72, 76)) ('L858R', 'Var', (107, 112)) ('EGFR', 'Gene', (72, 76)) ('NSCLC', 'Disease', (156, 161)) ('patients', 'Species', '9606', (58, 66)) ('NSCLC', 'Disease', 'MESH:D002289', (156, 161)) ('exon', 'Var', (77, 81)) 3427 30259761 Currently, erlotinib is approved as first-line therapy in patients with EGFR exon 19 deletions or exon 21 (L858R) substitution mutation-positive NSCLC. ('L858R', 'SUBSTITUTION', 'None', (107, 112)) ('deletions', 'Var', (85, 94)) ('NSCLC', 'Disease', (145, 150)) ('EGFR', 'Gene', '1956', (72, 76)) ('erlotinib', 'Chemical', 'MESH:C400278', (11, 20)) ('L858R', 'Var', (107, 112)) ('NSCLC', 'Disease', 'MESH:D002289', (145, 150)) ('EGFR', 'Gene', (72, 76)) ('patients', 'Species', '9606', (58, 66)) 3428 30259761 Two randomized controlled trials Erlotinib versus standard chemotherapy as first-line treatment for European patients with advanced EGFR mutation-positive Non-small-cell Lung Cancer (EURTAC) and Erlotinib vs Chemotherapy as first-line Treatment for Patients with advanced EGFR mutation-positive Non-small-cell lung cancer (OPTIMAL) compared erlotinib with the standard of care cisplatin plus docetaxel and erlotinib with gemcitabine plus carboplatin respectively in patients with EGFR mutated NSCLC. ('Erlotinib', 'Chemical', 'MESH:C400278', (33, 42)) ('cisplatin', 'Chemical', 'MESH:D002945', (377, 386)) ('Non-small-cell lung cancer', 'Disease', 'MESH:D002289', (295, 321)) ('Patients', 'Species', '9606', (249, 257)) ('patients', 'Species', '9606', (109, 117)) ('docetaxel', 'Chemical', 'MESH:C067311', (392, 401)) ('EGFR', 'Gene', (480, 484)) ('EGFR', 'Gene', '1956', (132, 136)) ('patients', 'Species', '9606', (466, 474)) ('EGFR', 'Gene', (272, 276)) ('Cancer', 'Phenotype', 'HP:0002664', (175, 181)) ('erlotinib', 'Chemical', 'MESH:C400278', (406, 415)) ('lung cancer', 'Phenotype', 'HP:0100526', (310, 321)) ('gemcitabine', 'Chemical', 'MESH:C056507', (421, 432)) ('carboplatin', 'Chemical', 'MESH:D016190', (438, 449)) ('Non-small-cell lung cancer', 'Phenotype', 'HP:0030358', (295, 321)) ('Erlotinib', 'Chemical', 'MESH:C400278', (195, 204)) ('Non-small-cell Lung Cancer', 'Phenotype', 'HP:0030358', (155, 181)) ('erlotinib', 'Chemical', 'MESH:C400278', (341, 350)) ('small-cell lung cancer', 'Phenotype', 'HP:0030357', (299, 321)) ('NSCLC', 'Disease', 'MESH:D002289', (493, 498)) ('EGFR', 'Gene', '1956', (480, 484)) ('cancer', 'Phenotype', 'HP:0002664', (315, 321)) ('mutated', 'Var', (485, 492)) ('EGFR', 'Gene', '1956', (272, 276)) ('Non-small-cell lung cancer', 'Disease', (295, 321)) ('Lung Cancer', 'Phenotype', 'HP:0100526', (170, 181)) ('small-cell Lung Cancer', 'Phenotype', 'HP:0030357', (159, 181)) ('EGFR', 'Gene', (132, 136)) ('NSCLC', 'Disease', (493, 498)) 3429 30259761 OPTIMAL study conducted in China showed that erlotinib significantly increased PFS when compared to chemotherapy (13.1 months vs 4.6 months); hazard ratio (HR) 0.16, (95% CI 0.10-0.26; p<0.0001) with a better side effect profile. ('erlotinib', 'Chemical', 'MESH:C400278', (45, 54)) ('erlotinib', 'Var', (45, 54)) ('increased', 'PosReg', (69, 78)) ('PFS', 'Disease', (79, 82)) 3431 30259761 In 2013, erlotinib was approved as first-line therapy for EGFR mutation harboring NSCLC. ('EGFR', 'Gene', '1956', (58, 62)) ('EGFR', 'Gene', (58, 62)) ('mutation', 'Var', (63, 71)) ('erlotinib', 'Chemical', 'MESH:C400278', (9, 18)) ('NSCLC', 'Disease', (82, 87)) ('NSCLC', 'Disease', 'MESH:D002289', (82, 87)) 3432 30259761 It is also approved as a second or third line therapy in EGFR mutated NSCLC patients who had failed previous chemotherapy. ('patients', 'Species', '9606', (76, 84)) ('NSCLC', 'Disease', (70, 75)) ('NSCLC', 'Disease', 'MESH:D002289', (70, 75)) ('EGFR', 'Gene', '1956', (57, 61)) ('EGFR', 'Gene', (57, 61)) ('mutated', 'Var', (62, 69)) 3440 30259761 In another randomized control trial LUX-LUNG 6 that compared afatinib to gemcitabine plus cisplatin, afatinib again proved to offer significantly longer PFS 11.0 months vs 5.6 months (HR-0.28; 95% CI 0.2-0.39; p < .0001) and objective response rate 66.9% vs 23.0%. ('cisplatin', 'Chemical', 'MESH:D002945', (90, 99)) ('afatinib', 'Chemical', 'MESH:C522924', (61, 69)) ('afatinib', 'Var', (101, 109)) ('gemcitabine', 'Chemical', 'MESH:C056507', (73, 84)) ('afatinib', 'Chemical', 'MESH:C522924', (101, 109)) ('longer', 'PosReg', (146, 152)) ('PFS', 'MPA', (153, 156)) 3442 30259761 These studies lead to the FDA approval of afatinib as a first-line agent in the treatment of advanced adenocarcinoma of the lung harboring EGFR exon 19 deletions or exon 21 (L858R) substitution mutations in 2013. ('carcinoma', 'Phenotype', 'HP:0030731', (107, 116)) ('L858R', 'SUBSTITUTION', 'None', (174, 179)) ('adenocarcinoma of the lung', 'Disease', (102, 128)) ('L858R', 'Var', (174, 179)) ('carcinoma of the lung', 'Phenotype', 'HP:0100526', (107, 128)) ('EGFR', 'Gene', '1956', (139, 143)) ('afatinib', 'Chemical', 'MESH:C522924', (42, 50)) ('deletions', 'Var', (152, 161)) ('EGFR', 'Gene', (139, 143)) ('adenocarcinoma of the lung', 'Disease', 'MESH:C538231', (102, 128)) 3451 30259761 Multiple investigations have revealed that a secondary mutation EGFR T790M lead to treatment failure in ~60% of the patients. ('lead to', 'Reg', (75, 82)) ('patients', 'Species', '9606', (116, 124)) ('T790M', 'SUBSTITUTION', 'None', (69, 74)) ('EGFR', 'Gene', '1956', (64, 68)) ('treatment failure', 'CPA', (83, 100)) ('EGFR', 'Gene', (64, 68)) ('T790M', 'Var', (69, 74)) 3452 30259761 This understanding has led to the development of third-generation EGFR TKIs osimertinib and other irreversible EGFR TKI selective for EGFR-TKI-sensitizing and T790M mutation to overcome resistance mediated by EGFR T790M. ('T790M', 'Var', (159, 164)) ('EGFR', 'Gene', (209, 213)) ('EGFR', 'Gene', (134, 138)) ('EGFR', 'Gene', '1956', (66, 70)) ('EGFR', 'Gene', '1956', (111, 115)) ('EGFR', 'Gene', (66, 70)) ('EGFR', 'Gene', '1956', (209, 213)) ('T790M', 'Var', (214, 219)) ('EGFR', 'Gene', (111, 115)) ('T790M', 'SUBSTITUTION', 'None', (159, 164)) ('osimertinib', 'Chemical', 'None', (76, 87)) ('EGFR', 'Gene', '1956', (134, 138)) ('T790M', 'SUBSTITUTION', 'None', (214, 219)) 3456 30259761 Osimertinib was approved by FDA in 2015 for EGFR T790M mutation-positive advanced NSCLC in patients who had progressed on or after TKI therapy. ('patients', 'Species', '9606', (91, 99)) ('mutation-positive', 'Reg', (55, 72)) ('Osimertinib', 'Chemical', 'None', (0, 11)) ('T790M', 'SUBSTITUTION', 'None', (49, 54)) ('EGFR', 'Gene', '1956', (44, 48)) ('EGFR', 'Gene', (44, 48)) ('NSCLC', 'Disease', (82, 87)) ('T790M', 'Var', (49, 54)) ('NSCLC', 'Disease', 'MESH:D002289', (82, 87)) 3485 30259761 Monoclonal antibody against VEGF-R significantly prolonged the PFS in patients with metastatic RCC. ('patients', 'Species', '9606', (70, 78)) ('VEGF', 'Gene', (28, 32)) ('PFS', 'MPA', (63, 66)) ('VEGF', 'Gene', '7422', (28, 32)) ('RCC', 'Disease', (95, 98)) ('Monoclonal antibody', 'Var', (0, 19)) ('RCC', 'Disease', 'MESH:D002292', (95, 98)) ('prolonged', 'PosReg', (49, 58)) 3502 30259761 In a randomized, double-blind, phase III trial in patients with loco-regional RCC who have undergone nephrectomy, sunitinib significantly increased the median duration of disease-free survival when compared to placebo; 6.8 years (95% CI 5.8 to not reached) in the sunitinib group vs 5.6 years (95% CI 3.8-6.6) in the placebo group; HR-0.76; (95% CI 0.59-0.98; P=0.03). ('sunitinib', 'Chemical', 'MESH:C473478', (114, 123)) ('increased', 'PosReg', (138, 147)) ('patients', 'Species', '9606', (50, 58)) ('RCC', 'Disease', (78, 81)) ('sunitinib', 'Var', (114, 123)) ('sunitinib', 'Chemical', 'MESH:C473478', (264, 273)) ('RCC', 'Disease', 'MESH:D002292', (78, 81)) 3504 30259761 In a prospective, placebo-controlled, randomised phase III clinical trial comparing the safety and efficacy of sunitinib vs placebo in advanced GIST patients who did not respond to imatinib, sunitinib significantly decreased the median time to tumour progression 27 3 weeks (95% CI 16 0-32 1) when compared to 6 4 weeks (4 4-10 0) in the placebo group; HR-0 33 (p<0 0001). ('decreased', 'NegReg', (215, 224)) ('patients', 'Species', '9606', (149, 157)) ('sunitinib', 'Chemical', 'MESH:C473478', (191, 200)) ('tumour', 'Phenotype', 'HP:0002664', (244, 250)) ('tumour', 'Disease', 'MESH:D009369', (244, 250)) ('sunitinib', 'Var', (191, 200)) ('sunitinib', 'Chemical', 'MESH:C473478', (111, 120)) ('imatinib', 'Chemical', 'MESH:C097613', (181, 189)) ('tumour', 'Disease', (244, 250)) 3517 30259761 Axitinib significantly increased the median PFS 6 7 months with axitinib vs 4 7 months with sorafenib; HR-0 665; (95% CI 0 544-0 812; one-sided p<0 0001). ('PFS', 'MPA', (44, 47)) ('axitinib', 'Chemical', 'MESH:C503983', (64, 72)) ('Axitinib', 'Chemical', 'MESH:C503983', (0, 8)) ('axitinib', 'Var', (64, 72)) ('increased', 'PosReg', (23, 32)) ('sorafenib', 'Chemical', 'MESH:C471405', (92, 101)) 3518 30259761 In updated efficacy, quality of life, and safety results, there was no survival advantage for using axitinib over sorafenib, although axitinib significantly increased the median investigator-assessed PFS of 8 3 months (95% CI 6 7-9 2) vs sorafenib of 5 7 months (4 7-6 5); HR-0 656 (95% CI 0 552-0 779; one-sided p<0 0001). ('axitinib', 'Chemical', 'MESH:C503983', (134, 142)) ('axitinib', 'Chemical', 'MESH:C503983', (100, 108)) ('sorafenib', 'Chemical', 'MESH:C471405', (238, 247)) ('sorafenib', 'Chemical', 'MESH:C471405', (114, 123)) ('increased', 'PosReg', (157, 166)) ('axitinib', 'Var', (134, 142)) ('PFS', 'MPA', (200, 203)) 3526 30259761 The rate of progression or death was 42% lower with cabozantinib vs everolimus; HR- 0.58; (95% CI 0.45-0.75; p<0.001). ('cabozantinib', 'Var', (52, 64)) ('cabozantinib', 'Chemical', 'MESH:C558660', (52, 64)) ('lower', 'NegReg', (41, 46)) ('everolimus', 'Chemical', 'MESH:C107135', (68, 78)) ('death', 'Disease', 'MESH:D003643', (27, 32)) ('death', 'Disease', (27, 32)) 3562 30259761 Interestingly, almost 50% of patients with melanoma have BRAF activation by V600E mutation and are susceptible to BRAF or MAPK/ERK kinase (MEK) inhibition. ('patients', 'Species', '9606', (29, 37)) ('V600E', 'Var', (76, 81)) ('BRAF', 'Gene', (57, 61)) ('melanoma', 'Disease', (43, 51)) ('ERK', 'Gene', '5594', (127, 130)) ('melanoma', 'Phenotype', 'HP:0002861', (43, 51)) ('melanoma', 'Disease', 'MESH:D008545', (43, 51)) ('V600E', 'SUBSTITUTION', 'None', (76, 81)) ('ERK', 'Gene', (127, 130)) ('MEK', 'Gene', (139, 142)) ('activation', 'PosReg', (62, 72)) ('MEK', 'Gene', '5609', (139, 142)) 3564 30259761 Vemurafenib is the first BRAF kinase inhibitor designed to inhibit the mutant BRAF V600E kinase in patients with advanced melanoma. ('V600E', 'Var', (83, 88)) ('Vemurafenib', 'Chemical', 'MESH:C551177', (0, 11)) ('BRAF', 'Gene', (78, 82)) ('V600E', 'SUBSTITUTION', 'None', (83, 88)) ('inhibit', 'NegReg', (59, 66)) ('patients', 'Species', '9606', (99, 107)) ('melanoma', 'Phenotype', 'HP:0002861', (122, 130)) ('melanoma', 'Disease', (122, 130)) ('melanoma', 'Disease', 'MESH:D008545', (122, 130)) 3565 30259761 Vemurafenib was approved by FDA in 2011 for the treatment of patients with BRAF V600E bearing metastatic melanoma. ('Vemurafenib', 'Chemical', 'MESH:C551177', (0, 11)) ('melanoma', 'Phenotype', 'HP:0002861', (105, 113)) ('melanoma', 'Disease', (105, 113)) ('patients', 'Species', '9606', (61, 69)) ('melanoma', 'Disease', 'MESH:D008545', (105, 113)) ('V600E', 'Var', (80, 85)) ('BRAF', 'Gene', (75, 79)) ('V600E', 'SUBSTITUTION', 'None', (80, 85)) 3566 30259761 A Study of Vemurafenib (RO5185426) in Comparison With Dacarbazine in Previously Untreated Patients With Metastatic Melanoma (BRIM 3) (BRIM3) is a landmark phase III randomized control trial that compared vemurafenib vs dacarbazine in patients with previously untreated melanoma with the BRAF V600E mutation. ('Patients', 'Species', '9606', (90, 98)) ('melanoma', 'Phenotype', 'HP:0002861', (269, 277)) ('melanoma', 'Disease', (269, 277)) ('Vemurafenib', 'Chemical', 'MESH:C551177', (11, 22)) ('Dacarbazine', 'Chemical', 'MESH:D003606', (54, 65)) ('melanoma', 'Disease', 'MESH:D008545', (269, 277)) ('RO5185426', 'Chemical', 'MESH:C542959', (24, 33)) ('V600E', 'Var', (292, 297)) ('BRAF', 'Gene', (287, 291)) ('V600E', 'SUBSTITUTION', 'None', (292, 297)) ('vemurafenib', 'Chemical', 'MESH:C551177', (204, 215)) ('dacarbazine', 'Chemical', 'MESH:D003606', (219, 230)) ('patients', 'Species', '9606', (234, 242)) ('Melanoma', 'Phenotype', 'HP:0002861', (115, 123)) ('Metastatic Melanoma', 'Disease', 'MESH:D008545', (104, 123)) ('Metastatic Melanoma', 'Disease', (104, 123)) 3572 30259761 Dabrafenib is the second BRAF kinase inhibitor designed to inhibit the mutant BRAF V600E kinase in patients with advanced melanoma. ('V600E', 'Var', (83, 88)) ('BRAF', 'Gene', (78, 82)) ('V600E', 'SUBSTITUTION', 'None', (83, 88)) ('inhibit', 'NegReg', (59, 66)) ('patients', 'Species', '9606', (99, 107)) ('melanoma', 'Phenotype', 'HP:0002861', (122, 130)) ('Dabrafenib', 'Chemical', 'MESH:C561627', (0, 10)) ('melanoma', 'Disease', (122, 130)) ('melanoma', 'Disease', 'MESH:D008545', (122, 130)) 3573 30259761 Dabrafenib was also approved in 2014 as a single agent for treatment of BRAF V600E mutation-positive unresectable or metastatic melanoma. ('melanoma', 'Disease', 'MESH:D008545', (128, 136)) ('BRAF', 'Gene', (72, 76)) ('melanoma', 'Phenotype', 'HP:0002861', (128, 136)) ('V600E', 'Var', (77, 82)) ('Dabrafenib', 'Chemical', 'MESH:C561627', (0, 10)) ('V600E', 'SUBSTITUTION', 'None', (77, 82)) ('melanoma', 'Disease', (128, 136)) 3574 30259761 In 2015, FDA approved dabrafenib in combination with trametinib to treat patients with unresectable or metastatic melanoma with a BRAF V600E mutation in patients who had received at least one platinum-based chemotherapy. ('melanoma', 'Disease', 'MESH:D008545', (114, 122)) ('patients', 'Species', '9606', (153, 161)) ('dabrafenib', 'Chemical', 'MESH:C561627', (22, 32)) ('V600E', 'Var', (135, 140)) ('BRAF', 'Gene', (130, 134)) ('V600E', 'SUBSTITUTION', 'None', (135, 140)) ('melanoma', 'Phenotype', 'HP:0002861', (114, 122)) ('platinum', 'Chemical', 'MESH:D010984', (192, 200)) ('patients', 'Species', '9606', (73, 81)) ('trametinib', 'Chemical', 'MESH:C560077', (53, 63)) ('melanoma', 'Disease', (114, 122)) 3575 30259761 An open-label, phase III randomized control trial compared dabrafenib to dacarbazine in patients with BRAFV600E mutant metastatic melanoma. ('patients', 'Species', '9606', (88, 96)) ('melanoma', 'Disease', 'MESH:D008545', (130, 138)) ('melanoma', 'Phenotype', 'HP:0002861', (130, 138)) ('melanoma', 'Disease', (130, 138)) ('dacarbazine', 'Chemical', 'MESH:D003606', (73, 84)) ('BRAFV600E', 'Var', (102, 111)) ('dabrafenib', 'Chemical', 'MESH:C561627', (59, 69)) 3578 30259761 The understanding of the implication BRAF mutation in melanoma prompted scientists to study its upstream and downstream signaling mediators. ('melanoma', 'Disease', 'MESH:D008545', (54, 62)) ('melanoma', 'Phenotype', 'HP:0002861', (54, 62)) ('BRAF', 'Gene', (37, 41)) ('melanoma', 'Disease', (54, 62)) ('mutation', 'Var', (42, 50)) 3581 30259761 Furthermore, BRAF mutation in cancer cells predicted response to MEK inhibition. ('MEK', 'Gene', (65, 68)) ('cancer', 'Disease', (30, 36)) ('cancer', 'Disease', 'MESH:D009369', (30, 36)) ('response', 'MPA', (53, 61)) ('MEK', 'Gene', '5609', (65, 68)) ('BRAF', 'Gene', (13, 17)) ('mutation', 'Var', (18, 26)) ('predicted', 'Reg', (43, 52)) ('cancer', 'Phenotype', 'HP:0002664', (30, 36)) 3584 30259761 This lead to the clinical trials testing the combination of BRAF kinase inhibitor plus MEK inhibitor in patients with BRAF mutated metastatic melanoma. ('melanoma', 'Disease', 'MESH:D008545', (142, 150)) ('MEK', 'Gene', (87, 90)) ('MEK', 'Gene', '5609', (87, 90)) ('mutated', 'Var', (123, 130)) ('patients', 'Species', '9606', (104, 112)) ('melanoma', 'Phenotype', 'HP:0002861', (142, 150)) ('melanoma', 'Disease', (142, 150)) 3585 30259761 Trametinib is a MEK inhibitor developed to target MEK in the treatment of BRAF mutated metastatic melanoma. ('Trametinib', 'Chemical', 'MESH:C560077', (0, 10)) ('mutated', 'Var', (79, 86)) ('BRAF', 'Gene', (74, 78)) ('melanoma', 'Disease', 'MESH:D008545', (98, 106)) ('melanoma', 'Phenotype', 'HP:0002861', (98, 106)) ('MEK', 'Gene', (16, 19)) ('melanoma', 'Disease', (98, 106)) ('MEK', 'Gene', (50, 53)) ('MEK', 'Gene', '5609', (16, 19)) ('MEK', 'Gene', '5609', (50, 53)) 3586 30259761 It is FDA approved for the treatment of BRAF V600E mutated metastatic melanoma as a monotherapy in 2013 as well as in combination with dabrafenib in 2014. ('melanoma', 'Disease', (70, 78)) ('melanoma', 'Disease', 'MESH:D008545', (70, 78)) ('V600E', 'Var', (45, 50)) ('V600E', 'SUBSTITUTION', 'None', (45, 50)) ('melanoma', 'Phenotype', 'HP:0002861', (70, 78)) ('dabrafenib', 'Chemical', 'MESH:C561627', (135, 145)) 3587 30259761 Currently, the combination is approved for first-line therapy for the treatment of BRAF V600E mutant metastatic melanoma. ('V600E', 'SUBSTITUTION', 'None', (88, 93)) ('melanoma', 'Disease', 'MESH:D008545', (112, 120)) ('melanoma', 'Phenotype', 'HP:0002861', (112, 120)) ('melanoma', 'Disease', (112, 120)) ('V600E', 'Var', (88, 93)) ('BRAF', 'Gene', (83, 87)) 3588 30259761 GSK1120212 vs Chemotherapy In Advanced or Metastatic BRAF V600E/K Mutation-positive Melanoma (METRIC) is a phase III randomized control trial that compared the efficacy of trametinib vs dacarbazine in patients with BRAF V600E mutant metastatic melanoma. ('melanoma', 'Disease', 'MESH:D008545', (244, 252)) ('patients', 'Species', '9606', (201, 209)) ('V600E', 'Var', (58, 63)) ('V600E', 'Var', (220, 225)) ('dacarbazine', 'Chemical', 'MESH:D003606', (186, 197)) ('V600E', 'SUBSTITUTION', 'None', (58, 63)) ('V600E', 'SUBSTITUTION', 'None', (220, 225)) ('trametinib', 'Chemical', 'MESH:C560077', (172, 182)) ('Melanoma', 'Phenotype', 'HP:0002861', (84, 92)) ('GSK1120212', 'Chemical', 'MESH:C560077', (0, 10)) ('Melanoma', 'Disease', 'MESH:D008545', (84, 92)) ('melanoma', 'Phenotype', 'HP:0002861', (244, 252)) ('melanoma', 'Disease', (244, 252)) ('Melanoma', 'Disease', (84, 92)) 3590 30259761 In 'A Study Comparing the Trametinib to Dabrafenib Monotherapy in Subjects With BRAF-mutant Melanoma (COMBI-d)', a double-blind, phase III randomized control trial, compared the combination of dabrafenib plus trametinib vs dabrafenib monotherapy in patients with BRAF V600E mutant metastatic melanoma. ('melanoma', 'Disease', 'MESH:D008545', (292, 300)) ('Melanoma', 'Disease', (92, 100)) ('V600E', 'Var', (268, 273)) ('BRAF', 'Gene', (263, 267)) ('patients', 'Species', '9606', (249, 257)) ('trametinib', 'Chemical', 'MESH:C560077', (209, 219)) ('dabrafenib', 'Chemical', 'MESH:C561627', (193, 203)) ('Dabrafenib', 'Chemical', 'MESH:C561627', (40, 50)) ('V600E', 'SUBSTITUTION', 'None', (268, 273)) ('Trametinib', 'Chemical', 'MESH:C560077', (26, 36)) ('dabrafenib', 'Chemical', 'MESH:C561627', (223, 233)) ('Melanoma', 'Phenotype', 'HP:0002861', (92, 100)) ('melanoma', 'Phenotype', 'HP:0002861', (292, 300)) ('Melanoma', 'Disease', 'MESH:D008545', (92, 100)) ('melanoma', 'Disease', (292, 300)) 3594 30259761 Similar results were observed in 'Dabrafenib Plus Trametinib vs Vemurafenib Alone in Unresectable or Metastatic BRAF V600E/K Cutaneous Melanoma (COMBI-v)', a double-blind, phase III randomized control trial, compared the combination of dabrafenib plus trametinib vs vemurafenib in patients with BRAF V600E mutant metastatic melanoma. ('trametinib', 'Chemical', 'MESH:C560077', (252, 262)) ('V600E', 'Var', (117, 122)) ('melanoma', 'Phenotype', 'HP:0002861', (324, 332)) ('dabrafenib', 'Chemical', 'MESH:C561627', (236, 246)) ('melanoma', 'Disease', (324, 332)) ('V600E', 'Var', (300, 305)) ('vemurafenib', 'Chemical', 'MESH:C551177', (266, 277)) ('V600E', 'SUBSTITUTION', 'None', (300, 305)) ('patients', 'Species', '9606', (281, 289)) ('melanoma', 'Disease', 'MESH:D008545', (324, 332)) ('V600E', 'SUBSTITUTION', 'None', (117, 122)) ('Melanoma', 'Phenotype', 'HP:0002861', (135, 143)) ('Cutaneous Melanoma', 'Disease', (125, 143)) ('Dabrafenib', 'Chemical', 'MESH:C561627', (34, 44)) ('Trametinib', 'Chemical', 'MESH:C560077', (50, 60)) ('Cutaneous Melanoma', 'Phenotype', 'HP:0012056', (125, 143)) ('Cutaneous Melanoma', 'Disease', 'MESH:C562393', (125, 143)) ('Vemurafenib', 'Chemical', 'MESH:C551177', (64, 75)) 3602 30259761 Cobimetinib is a MEK inhibitor that the FDA approved in 2015 for the treatment of patients with BRAFV600E mutant metastatic melanoma in combination with vemurafenib. ('melanoma', 'Disease', 'MESH:D008545', (124, 132)) ('melanoma', 'Phenotype', 'HP:0002861', (124, 132)) ('MEK', 'Gene', (17, 20)) ('melanoma', 'Disease', (124, 132)) ('mutant', 'Var', (106, 112)) ('vemurafenib', 'Chemical', 'MESH:C551177', (153, 164)) ('patients', 'Species', '9606', (82, 90)) ('MEK', 'Gene', '5609', (17, 20)) ('BRAFV600E', 'Gene', (96, 105)) ('Cobimetinib', 'Chemical', 'MESH:C574276', (0, 11)) 3608 30259761 In addition to pathogenic EGFR mutations, gene rearrangement involving ALK and echinoderm microtubule-associated protein-like 4 (EML4) resulting in EML4-ALK fusion gene also acts as driver mutation in adenocarcinoma of the lung (NSCLC). ('EML4', 'Gene', (148, 152)) ('gene rearrangement', 'Var', (42, 60)) ('NSCLC', 'Disease', (229, 234)) ('EML4', 'Gene', '27436', (148, 152)) ('EML4', 'Gene', (129, 133)) ('EGFR', 'Gene', (26, 30)) ('mutations', 'Var', (31, 40)) ('carcinoma', 'Phenotype', 'HP:0030731', (206, 215)) ('adenocarcinoma of the lung', 'Disease', (201, 227)) ('carcinoma of the lung', 'Phenotype', 'HP:0100526', (206, 227)) ('EML4', 'Gene', '27436', (129, 133)) ('echinoderm microtubule-associated protein-like 4', 'Gene', '27436', (79, 127)) ('ALK', 'Gene', '238', (71, 74)) ('ALK', 'Gene', '238', (153, 156)) ('adenocarcinoma of the lung', 'Disease', 'MESH:C538231', (201, 227)) ('ALK', 'Gene', (71, 74)) ('ALK', 'Gene', (153, 156)) ('echinoderm microtubule-associated protein-like 4', 'Gene', (79, 127)) ('NSCLC', 'Disease', 'MESH:D002289', (229, 234)) ('EGFR', 'Gene', '1956', (26, 30)) 3610 30259761 Originally developed to target c-met, crizotinib was also found to inhibit ALK tyrosine kinase activity and was tested on NCSLC patients with ALK fusion gene with significant benefits. ('crizotinib', 'Var', (38, 48)) ('inhibit', 'NegReg', (67, 74)) ('ALK', 'Gene', (75, 78)) ('ALK', 'Gene', '238', (142, 145)) ('c-met', 'Gene', (31, 36)) ('tyrosine kinase', 'Gene', '7294', (79, 94)) ('crizotinib', 'Chemical', 'MESH:C551994', (38, 48)) ('c-met', 'Gene', '4233', (31, 36)) ('ALK', 'Gene', '238', (75, 78)) ('tyrosine kinase', 'Gene', (79, 94)) ('ALK', 'Gene', (142, 145)) ('patients', 'Species', '9606', (128, 136)) 3611 30259761 AKL and c-ros oncogene (ROS1) tyrosine kinases have high sequence similarity and crizotinib has also been found to have beneficial effects on ROS-1 gene rearrangement-positive NSCLC. ('crizotinib', 'Chemical', 'MESH:C551994', (81, 91)) ('tyrosine kinase', 'Gene', (30, 45)) ('ROS-1', 'Gene', '6098', (142, 147)) ('NSCLC', 'Disease', (176, 181)) ('ROS1', 'Gene', (24, 28)) ('rearrangement-positive', 'Var', (153, 175)) ('ROS1', 'Gene', '6098', (24, 28)) ('NSCLC', 'Disease', 'MESH:D002289', (176, 181)) ('tyrosine kinase', 'Gene', '7294', (30, 45)) ('beneficial effects', 'PosReg', (120, 138)) ('ROS-1', 'Gene', (142, 147)) 3614 30259761 Crizotinib significantly increased the median PFS 7.7 months vs pemetrexed at 3.0 months; HR for progression or death with crizotinib was 0.49 (95% CI 0.37-0.64; p<0.001). ('Crizotinib', 'Chemical', 'MESH:C551994', (0, 10)) ('death', 'Disease', 'MESH:D003643', (112, 117)) ('death', 'Disease', (112, 117)) ('increased', 'PosReg', (25, 34)) ('crizotinib', 'Chemical', 'MESH:C551994', (123, 133)) ('Crizotinib', 'Var', (0, 10)) ('PFS', 'MPA', (46, 49)) 3617 30259761 Crizotinib significantly improved the median PFS of 10.9 months (95% CI 8.3-13.9) vs chemotherapy of 7.0 months (95% CI 6.8-8.2). ('improved', 'PosReg', (25, 33)) ('PFS', 'MPA', (45, 48)) ('Crizotinib', 'Var', (0, 10)) ('Crizotinib', 'Chemical', 'MESH:C551994', (0, 10)) 3625 30259761 Ceritinib significantly improved the median PFS vs chemotherapy 16 6 months (95% CI 12 6-27 2) in the ceritinib group vs 8 1 months (5 8-11 1) in the chemotherapy group; HR-0 55 (95% CI 0 42-0 73; p<0 00001). ('PFS', 'MPA', (44, 47)) ('ceritinib', 'Var', (102, 111)) ('improved', 'PosReg', (24, 32)) ('ceritinib', 'Chemical', 'MESH:C586847', (102, 111)) ('Ceritinib', 'Chemical', 'MESH:C586847', (0, 9)) 3639 30259761 It differs from the other ALK-ROS1 TKI by its ability to inhibit most known resistant mutations to other ALK TKIs and it crosses the blood-brain barrier. ('mutations', 'Var', (86, 95)) ('ALK', 'Gene', (26, 29)) ('ALK', 'Gene', (105, 108)) ('ROS1', 'Gene', (30, 34)) ('ROS1', 'Gene', '6098', (30, 34)) ('inhibit', 'NegReg', (57, 64)) ('ALK', 'Gene', '238', (105, 108)) ('ALK', 'Gene', '238', (26, 29)) 3642 30259761 The ORR was 2% in patients who had progressed on crizotinib only, 9% in patients who had progressed on crizotinib and chemotherapy, 17% in patients who had progressed on two ALK TKIs and chemotherapy and 7% in patients who had progressed on three prior TKIs and chemotherapy. ('crizotinib', 'Var', (103, 113)) ('patients', 'Species', '9606', (139, 147)) ('ALK', 'Gene', (174, 177)) ('crizotinib', 'Chemical', 'MESH:C551994', (49, 59)) ('crizotinib', 'Chemical', 'MESH:C551994', (103, 113)) ('patients', 'Species', '9606', (210, 218)) ('patients', 'Species', '9606', (18, 26)) ('ALK', 'Gene', '238', (174, 177)) ('patients', 'Species', '9606', (72, 80)) 3676 30259761 FLT3 mutations have been found 1-3% of patients with ALL, 5-10% of patients with myelodysplasia and 15-35% of patients with AML. ('patients', 'Species', '9606', (39, 47)) ('patients', 'Species', '9606', (67, 75)) ('AML', 'Disease', (124, 127)) ('FLT3', 'Gene', '2322', (0, 4)) ('myelodysplasia', 'Disease', 'MESH:D009190', (81, 95)) ('mutations', 'Var', (5, 14)) ('myelodysplasia', 'Disease', (81, 95)) ('AML', 'Disease', 'MESH:D015470', (124, 127)) ('FLT3', 'Gene', (0, 4)) ('patients', 'Species', '9606', (110, 118)) ('myelodysplasia', 'Phenotype', 'HP:0002863', (81, 95)) 3677 30259761 Internal tandem duplication (ITD) JM domain-coding sequence of the FLT3 gene (FLT3/ITD) mutation constitutes approximately two-third of FLT3 mutations in AML patients and tyrosine kinase domain (TKD) mutations mostly point mutations in codon D835 or deletions of codon I836 constitute the remaining one third.. ('mutation', 'Var', (88, 96)) ('deletions', 'Var', (250, 259)) ('tyrosine kinase', 'Gene', '7294', (171, 186)) ('mutations', 'Var', (141, 150)) ('AML', 'Disease', 'MESH:D015470', (154, 157)) ('FLT3', 'Gene', '2322', (67, 71)) ('FLT3', 'Gene', '2322', (78, 82)) ('point mutations', 'Var', (217, 232)) ('tyrosine kinase', 'Gene', (171, 186)) ('FLT3', 'Gene', '2322', (136, 140)) ('AML', 'Disease', (154, 157)) ('FLT3', 'Gene', (78, 82)) ('FLT3', 'Gene', (67, 71)) ('patients', 'Species', '9606', (158, 166)) ('FLT3', 'Gene', (136, 140)) 3681 30259761 It is FDA approved for the treatment of FLT3 mutation-positive AML patients in combination with chemotherapy, making this is as the first targeted therapy approved for the treatment of AML. ('AML', 'Disease', (185, 188)) ('AML', 'Disease', (63, 66)) ('patients', 'Species', '9606', (67, 75)) ('FLT3', 'Gene', '2322', (40, 44)) ('FLT3', 'Gene', (40, 44)) ('mutation-positive', 'Var', (45, 62)) ('AML', 'Disease', 'MESH:D015470', (185, 188)) ('AML', 'Disease', 'MESH:D015470', (63, 66)) 3696 30259761 Interestingly, the JAK2 mutation did not predict the response to ruxolitinib therapy. ('ruxolitinib', 'Chemical', 'MESH:C540383', (65, 76)) ('JAK2', 'Gene', '3717', (19, 23)) ('JAK2', 'Gene', (19, 23)) ('mutation', 'Var', (24, 32)) 3708 30259761 Class I PI3K has been implicated in many cancers particularly those with pathogenic mutations. ('cancer', 'Phenotype', 'HP:0002664', (41, 47)) ('implicated', 'Reg', (22, 32)) ('mutations', 'Var', (84, 93)) ('cancers', 'Phenotype', 'HP:0002664', (41, 48)) ('PI3K', 'Var', (8, 12)) ('cancers', 'Disease', 'MESH:D009369', (41, 48)) ('cancers', 'Disease', (41, 48)) 3709 30259761 PI3K acts downstream to many growth factors and acts upstream to AKT and mTOR. ('mTOR', 'Gene', '2475', (73, 77)) ('PI3K', 'Var', (0, 4)) ('mTOR', 'Gene', (73, 77)) ('AKT', 'Gene', (65, 68)) ('AKT', 'Gene', '207', (65, 68)) 3725 30259761 Many tumorigenic signals converge on the CKD4/6-cyclin D complex in the G1 phase of the cell cycle. ('CKD4/6-cyclin', 'Var', (41, 54)) ('tumor', 'Phenotype', 'HP:0002664', (5, 10)) ('tumor', 'Disease', 'MESH:D009369', (5, 10)) ('tumor', 'Disease', (5, 10)) 3727 30259761 Genotoxic events leading to DNA damage and chromosomal instability dysregulate the progression of the cell cycle from S phase to G2/M phase which is regulated by CDK1/2. ('chromosomal instability', 'Phenotype', 'HP:0040012', (43, 66)) ('CDK1/2', 'Gene', (162, 168)) ('CDK1/2', 'Gene', '51755', (162, 168)) ('dysregulate', 'Reg', (67, 78)) ('chromosomal instability', 'Var', (43, 66)) ('progression', 'CPA', (83, 94)) 3735 30259761 However, unlike conventional chemotherapies, neutropenia caused by CDK4/CDK6 inhibitors are rapidly reversible due to cytostatic rather than cytotoxic effects on the bone marrow. ('inhibitors', 'Var', (77, 87)) ('CDK4', 'Gene', (67, 71)) ('neutropenia', 'Disease', 'MESH:D009503', (45, 56)) ('CDK4', 'Gene', '1019', (67, 71)) ('neutropenia', 'Phenotype', 'HP:0001875', (45, 56)) ('CDK6', 'Gene', (72, 76)) ('neutropenia', 'Disease', (45, 56)) ('CDK6', 'Gene', '1021', (72, 76)) 3739 30259761 Common grade 3 or 4 adverse events that were reported were neutropenia (59.3% in the ribociclib group vs 0.9% in the placebo group) and leukopenia (21.0% vs 0.6%). ('neutropenia', 'Phenotype', 'HP:0001875', (59, 70)) ('neutropenia', 'Disease', 'MESH:D009503', (59, 70)) ('leukopenia', 'Phenotype', 'HP:0001882', (136, 146)) ('leukopenia', 'Disease', (136, 146)) ('neutropenia', 'Disease', (59, 70)) ('ribociclib', 'Var', (85, 95)) ('leukopenia', 'Disease', 'MESH:D007970', (136, 146)) 3748 30259761 For example, understanding of the role of EGFR T790M in the resistance to the EGFR TKIs in NSCLC, lead to the designing of osimertinib that particularly targeted the EGFR T790M gatekeeper mutant protein kinase and identification T315I gatekeeper mutant gene as a secondary mutation responsible resistance to first and second generation Bcr-Abl kinase inhibitors lead to the designing of ponatinib that targeted the T315I mutant kinase. ('T315I', 'SUBSTITUTION', 'None', (415, 420)) ('T790M', 'Var', (47, 52)) ('EGFR', 'Gene', '1956', (166, 170)) ('EGFR', 'Gene', '1956', (78, 82)) ('EGFR', 'Gene', (42, 46)) ('ponatinib', 'Chemical', 'MESH:C545373', (387, 396)) ('gatekeeper', 'Species', '111938', (235, 245)) ('T790M', 'Var', (171, 176)) ('T315I', 'SUBSTITUTION', 'None', (229, 234)) ('Bcr-Abl', 'Gene', (336, 343)) ('NSCLC', 'Disease', 'MESH:D002289', (91, 96)) ('T790M', 'SUBSTITUTION', 'None', (47, 52)) ('gatekeeper', 'Species', '111938', (177, 187)) ('T315I', 'Var', (415, 420)) ('EGFR', 'Gene', (166, 170)) ('osimertinib', 'Chemical', 'None', (123, 134)) ('EGFR', 'Gene', (78, 82)) ('NSCLC', 'Disease', (91, 96)) ('EGFR', 'Gene', '1956', (42, 46)) ('T790M', 'SUBSTITUTION', 'None', (171, 176)) ('T315I', 'Var', (229, 234)) ('Bcr-Abl', 'Gene', '25', (336, 343)) 3751 30259761 For example, the Bcr-Abl kinase inhibitor bosutinib is more specific to Bcr-Abl kinase when compared to other Bcr-Abl kinase inhibitors like dasatinib or nilotinib which also inhibit c-Kit and PDGFR. ('Bcr-Abl', 'Gene', (72, 79)) ('Bcr-Abl', 'Gene', '25', (17, 24)) ('Bcr-Abl', 'Gene', (110, 117)) ('nilotinib', 'Chemical', 'MESH:C498826', (154, 163)) ('Bcr-Abl', 'Gene', '25', (72, 79)) ('Bcr-Abl', 'Gene', '25', (110, 117)) ('dasatinib', 'Chemical', 'MESH:C488369', (141, 150)) ('PDGFR', 'Gene', (193, 198)) ('PDGFR', 'Gene', '5159', (193, 198)) ('bosutinib', 'Var', (42, 51)) ('bosutinib', 'Chemical', 'MESH:C471992', (42, 51)) ('inhibit', 'NegReg', (175, 182)) ('c-Kit', 'Protein', (183, 188)) ('Bcr-Abl', 'Gene', (17, 24)) 3761 28123529 Profiling of microRNAs in AML cells following overexpression or silencing of the VEGF gene Acute myeloid leukemia (AML) is a disease of the hematopoietic progenitor cells associated with heterogeneous clonal proliferation. ('AML', 'Disease', 'MESH:D015470', (115, 118)) ('VEGF', 'Gene', '7422', (81, 85)) ('AML', 'Disease', 'MESH:D015470', (26, 29)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (97, 113)) ('AML', 'Phenotype', 'HP:0004808', (115, 118)) ('leukemia', 'Phenotype', 'HP:0001909', (105, 113)) ('AML', 'Disease', (115, 118)) ('AML', 'Phenotype', 'HP:0004808', (26, 29)) ('AML', 'Disease', (26, 29)) ('VEGF', 'Gene', (81, 85)) ('Acute myeloid leukemia', 'Disease', (91, 113)) ('Acute myeloid leukemia', 'Phenotype', 'HP:0004808', (91, 113)) ('silencing', 'Var', (64, 73)) ('Acute myeloid leukemia', 'Disease', 'MESH:D015470', (91, 113)) 3765 28123529 The present study aimed to investigate the relationship between VEGF dysregulation and microRNA profiles in AML cells and patients. ('AML', 'Disease', (108, 111)) ('AML', 'Phenotype', 'HP:0004808', (108, 111)) ('VEGF', 'Gene', (64, 68)) ('patients', 'Species', '9606', (122, 130)) ('dysregulation', 'Var', (69, 82)) ('AML', 'Disease', 'MESH:D015470', (108, 111)) ('VEGF', 'Gene', '7422', (64, 68)) 3814 28123529 Genes that were screened using the microarray needed to meet the criteria of >=2-fold increase or decrease in expression in the cells with overexpression or knockdown of VEGF compared with the controls. ('increase', 'PosReg', (86, 94)) ('decrease', 'NegReg', (98, 106)) ('VEGF', 'Gene', (170, 174)) ('knockdown', 'Var', (157, 166)) ('expression', 'MPA', (110, 120)) ('VEGF', 'Gene', '7422', (170, 174)) 3818 28123529 In the present study, several microRNAs showed changes in their levels of expression following overexpression or silencing of the expression of the VEGF gene. ('VEGF', 'Gene', (148, 152)) ('changes', 'Reg', (47, 54)) ('overexpression', 'PosReg', (95, 109)) ('VEGF', 'Gene', '7422', (148, 152)) ('silencing', 'Var', (113, 122)) ('levels of expression', 'MPA', (64, 84)) 3829 28123529 It was observed that overexpression of VEGF had a limited impact on leukemia cells, but that silencing of VEGF significantly altered the microRNA profiles of leukemia cells. ('leukemia', 'Disease', (68, 76)) ('silencing', 'Var', (93, 102)) ('VEGF', 'Gene', '7422', (106, 110)) ('leukemia', 'Disease', 'MESH:D007938', (68, 76)) ('leukemia', 'Phenotype', 'HP:0001909', (68, 76)) ('leukemia', 'Disease', (158, 166)) ('altered', 'Reg', (125, 132)) ('leukemia', 'Disease', 'MESH:D007938', (158, 166)) ('leukemia', 'Phenotype', 'HP:0001909', (158, 166)) ('VEGF', 'Gene', (39, 43)) ('microRNA profiles', 'MPA', (137, 154)) ('VEGF', 'Gene', (106, 110)) ('VEGF', 'Gene', '7422', (39, 43)) 3840 28123529 The results of the present study suggested that knockdown of VEGF could induce large changes in the expression of microRNAs, likely because VEGF signaling has an important role in the development and progression of AML cells. ('VEGF', 'Gene', '7422', (61, 65)) ('AML', 'Disease', 'MESH:D015470', (215, 218)) ('VEGF', 'Gene', '7422', (140, 144)) ('changes', 'Reg', (85, 92)) ('AML', 'Disease', (215, 218)) ('expression', 'MPA', (100, 110)) ('AML', 'Phenotype', 'HP:0004808', (215, 218)) ('VEGF', 'Gene', (61, 65)) ('VEGF', 'Gene', (140, 144)) ('microRNAs', 'Protein', (114, 123)) ('knockdown', 'Var', (48, 57)) 3844 21323862 Human genetic disorders that develop due to mutations in telomerase subunits, the role of single nucleotide polymorphisms in genes encoding telomerase components and diseases as a result of telomerase regulation going awry are also discussed. ('genetic disorders', 'Disease', 'MESH:D030342', (6, 23)) ('Human', 'Species', '9606', (0, 5)) ('single nucleotide polymorphisms', 'Var', (90, 121)) ('mutations', 'Var', (44, 53)) ('genetic disorders', 'Disease', (6, 23)) 3859 21323862 studied multiple cell types to try to determine if there was some underlying mechanism behind the epigenetic regulation of telomerase in all cancer types. ('cancer', 'Phenotype', 'HP:0002664', (141, 147)) ('epigenetic', 'Var', (98, 108)) ('cancer', 'Disease', (141, 147)) ('cancer', 'Disease', 'MESH:D009369', (141, 147)) 3865 21323862 A recent study has indicated that subtelomeric methylation may influence telomerase activity, and specifically whether a tumour maintains its telomeres with telomerase or through the alternative lengthening of telomeres (ALT) pathway. ('tumour', 'Phenotype', 'HP:0002664', (121, 127)) ('telomerase', 'Enzyme', (73, 83)) ('tumour', 'Disease', 'MESH:D009369', (121, 127)) ('activity', 'MPA', (84, 92)) ('influence', 'Reg', (63, 72)) ('subtelomeric methylation', 'Var', (34, 58)) ('tumour', 'Disease', (121, 127)) ('telomeres', 'MPA', (142, 151)) 3888 21323862 Also, miRNA activities may be cell-type specific, and the disregulation of miR-138 may be important in telomerase activity in thyroid carcinomas, whereas other miRNAs and mechanisms may be more important in regulating telomerase in other cells and tissues. ('miR', 'Gene', (160, 163)) ('miR', 'Gene', '220972', (75, 78)) ('telomerase activity', 'MPA', (103, 122)) ('miR', 'Gene', (75, 78)) ('thyroid carcinomas', 'Disease', 'MESH:D013964', (126, 144)) ('important', 'Reg', (90, 99)) ('miR', 'Gene', '220972', (6, 9)) ('thyroid carcinomas', 'Phenotype', 'HP:0002890', (126, 144)) ('miR', 'Gene', (6, 9)) ('thyroid carcinomas', 'Disease', (126, 144)) ('carcinoma', 'Phenotype', 'HP:0030731', (134, 143)) ('disregulation', 'Var', (58, 71)) ('thyroid carcinoma', 'Phenotype', 'HP:0002890', (126, 143)) ('carcinomas', 'Phenotype', 'HP:0030731', (134, 144)) ('miR', 'Gene', '220972', (160, 163)) 3909 21323862 Using ChIP, they found that TTAGGG repeats were capable of pulling down both the N- and C-terminal domains of MLL in human haematopoietic cell lines and human ovarian surface epithelium. ('human', 'Species', '9606', (117, 122)) ('C-terminal domains', 'MPA', (88, 106)) ('TTAGGG repeats', 'Var', (28, 42)) ('MLL', 'Gene', (110, 113)) ('human', 'Species', '9606', (153, 158)) ('MLL', 'Gene', '4297', (110, 113)) ('pulling down', 'NegReg', (59, 71)) 3910 21323862 It was also shown that MLL binding positively correlated to H3K4 methylation and transcription of telomeres, and that knocking down MLL resulted in histone modifications at the telomeres. ('MLL', 'Gene', (23, 26)) ('H3K4', 'Protein', (60, 64)) ('resulted in', 'Reg', (136, 147)) ('histone modifications', 'MPA', (148, 169)) ('methylation', 'MPA', (65, 76)) ('correlated', 'Reg', (46, 56)) ('knocking down', 'Var', (118, 131)) ('MLL', 'Gene', (132, 135)) ('MLL', 'Gene', '4297', (132, 135)) ('transcription', 'MPA', (81, 94)) ('MLL', 'Gene', '4297', (23, 26)) 3917 21323862 Dysfunction due to mutations in the human genes essential for telomerase function (TERT and TERC) exhausts the proliferative capacity of cells with a high turnover rate, such as the haematopoietic system and other tissues. ('exhausts', 'NegReg', (98, 106)) ('TERC', 'Gene', (92, 96)) ('proliferative capacity of cells', 'CPA', (111, 142)) ('Dysfunction', 'Disease', (0, 11)) ('mutations', 'Var', (19, 28)) ('TERC', 'Gene', '7012', (92, 96)) ('human', 'Species', '9606', (36, 41)) 3920 21323862 The mutations in hTERC affect the function of telomerase RNA either by disrupting the secondary structure to prevent its folding into a stable conformation or by sequence changes that prevent its interaction with other telomerase complex components. ('prevent', 'NegReg', (184, 191)) ('disrupting', 'NegReg', (71, 81)) ('prevent', 'NegReg', (109, 116)) ('function', 'MPA', (34, 42)) ('hTERC', 'Gene', (17, 22)) ('interaction', 'Interaction', (196, 207)) ('secondary structure', 'MPA', (86, 105)) ('hTERC', 'Gene', '7012', (17, 22)) ('changes', 'Var', (171, 178)) ('affect', 'Reg', (23, 29)) ('mutations', 'Var', (4, 13)) ('telomerase', 'Protein', (46, 56)) ('folding into a stable conformation', 'MPA', (121, 155)) 3921 21323862 Two mutations (See Table 1) have been found in the template region of hTERC, which decrease telomerase activity to levels that are undetectable by the Telomeric Repeat Amplification Protocol (TRAP) assay. ('hTERC', 'Gene', '7012', (70, 75)) ('hTERC', 'Gene', (70, 75)) ('decrease', 'NegReg', (83, 91)) ('telomerase activity', 'MPA', (92, 111)) ('mutations', 'Var', (4, 13)) 3922 21323862 The mutations in hTERT can affect its reverse transcriptase function, DNA-binding ability and protein-protein interaction strength. ('hTERT', 'Gene', (17, 22)) ('reverse transcriptase', 'Enzyme', (38, 59)) ('protein-protein interaction', 'Interaction', (94, 121)) ('mutations', 'Var', (4, 13)) ('function', 'MPA', (60, 68)) ('DNA-binding', 'Interaction', (70, 81)) ('affect', 'Reg', (27, 33)) ('hTERT', 'Gene', '7015', (17, 22)) 3923 21323862 hTERT mutations show varying degrees of effect on telomerase activity, with both conserved and non-conserved amino acids being able to drastically reduce activity. ('hTERT', 'Gene', (0, 5)) ('telomerase', 'Enzyme', (50, 60)) ('reduce', 'NegReg', (147, 153)) ('hTERT', 'Gene', '7015', (0, 5)) ('activity', 'MPA', (61, 69)) ('activity', 'MPA', (154, 162)) ('mutations', 'Var', (6, 15)) 3924 21323862 Dyskerin interacts with hTR prior to assembly of the holoenzyme and mutations in Dyskerin 1 (DKC1) are shown to significantly impair this interaction. ('impair', 'NegReg', (126, 132)) ('hTR', 'Gene', '7012', (24, 27)) ('interacts', 'Interaction', (9, 18)) ('interaction', 'Interaction', (138, 149)) ('DKC1', 'Gene', '1736', (93, 97)) ('hTR', 'Gene', (24, 27)) ('mutations', 'Var', (68, 77)) ('DKC1', 'Gene', (93, 97)) 3925 21323862 hTERT mutations do not show a strict linkage between mutation and disease, but rather may show an increased risk of disease in individuals with hTERT mutations. ('disease', 'Disease', (116, 123)) ('hTERT', 'Gene', (144, 149)) ('hTERT', 'Gene', (0, 5)) ('mutations', 'Var', (150, 159)) ('hTERT', 'Gene', '7015', (0, 5)) ('hTERT', 'Gene', '7015', (144, 149)) 3931 21323862 Mutations in patients suffering from DC were initially discovered in the DKC1 gene (See Table 1) located on the X chromosome, which encodes dyskerin, a protein involved in stabilizing telomerase RNA and forming the enzyme complex. ('DKC1', 'Gene', '1736', (73, 77)) ('DKC1', 'Gene', (73, 77)) ('Mutations', 'Var', (0, 9)) ('patients', 'Species', '9606', (13, 21)) 3932 21323862 The most common mutation found in DKC1 is a C T nucleotide substitution, which results in the A353V missense mutation in the protein sequence. ('A353V', 'Mutation', 'rs121912288', (94, 99)) ('DKC1', 'Gene', (34, 38)) ('results in', 'Reg', (79, 89)) ('DKC1', 'Gene', '1736', (34, 38)) ('A353V missense', 'Var', (94, 108)) 3933 21323862 Approximately half of DC patients have a mutation in the genes of the telomerase holoenzyme complex, hTERT, hTERC or DKC1. ('hTERT', 'Gene', '7015', (101, 106)) ('DKC1', 'Gene', '1736', (117, 121)) ('hTERC', 'Gene', '7012', (108, 113)) ('mutation', 'Var', (41, 49)) ('hTERC', 'Gene', (108, 113)) ('patients', 'Species', '9606', (25, 33)) ('hTERT', 'Gene', (101, 106)) ('DKC1', 'Gene', (117, 121)) 3934 21323862 It is likely that the remaining 50% of patients have mutations in other telomere maintenance genes, such as the mutations found in TINF2, NHP2 and NOP10. ('TINF2', 'Gene', '26277', (131, 136)) ('patients', 'Species', '9606', (39, 47)) ('NOP10', 'Gene', (147, 152)) ('TINF2', 'Gene', (131, 136)) ('mutations', 'Var', (53, 62)) ('NOP10', 'Gene', '79954', (147, 152)) ('NHP2', 'Gene', '55651', (138, 142)) ('NHP2', 'Gene', (138, 142)) 3936 21323862 AA is another bone marrow failure disease associated with telomerase mutations. ('mutations', 'Var', (69, 78)) ('AA', 'Phenotype', 'HP:0001903', (0, 2)) ('associated', 'Reg', (42, 52)) ('telomerase', 'Protein', (58, 68)) ('bone marrow failure disease', 'Disease', 'MESH:D000080983', (14, 41)) ('bone marrow failure disease', 'Disease', (14, 41)) ('bone marrow failure', 'Phenotype', 'HP:0005528', (14, 33)) 3940 21323862 In acquired AA, approximately 2.5% of patients have mutations in hTERC and 3.5% have mutations in hTERT (See Table 1 for list). ('mutations', 'Var', (52, 61)) ('hTERC', 'Gene', '7012', (65, 70)) ('hTERC', 'Gene', (65, 70)) ('mutations', 'Var', (85, 94)) ('hTERT', 'Gene', '7015', (98, 103)) ('AA', 'Phenotype', 'HP:0001903', (12, 14)) ('patients', 'Species', '9606', (38, 46)) ('hTERT', 'Gene', (98, 103)) 3941 21323862 The majority of hTERC mutations cluster in the pseudoknot region, which is essential for hTERT binding and telomerase enzymatic activity. ('hTERT', 'Gene', (89, 94)) ('mutations', 'Var', (22, 31)) ('hTERC', 'Gene', (16, 21)) ('hTERT', 'Gene', '7015', (89, 94)) ('hTERC', 'Gene', '7012', (16, 21)) 3942 21323862 The identified hTERT mutations are spread throughout the gene and act by haploinsufficiency, not a dominant negative mechanism. ('haploinsufficiency', 'Disease', (73, 91)) ('hTERT', 'Gene', '7015', (15, 20)) ('hTERT', 'Gene', (15, 20)) ('haploinsufficiency', 'Disease', 'MESH:D058495', (73, 91)) ('mutations', 'Var', (21, 30)) 3947 21323862 Although the percentage of patients with telomerase mutations is lower in IPF than dyskeratosis congentia, the higher incidence rate makes pulmonary fibrosis the most common clinical manifestation of telomerase gene mutations. ('mutations', 'Var', (52, 61)) ('telomerase', 'Protein', (41, 51)) ('pulmonary fibrosis', 'Disease', (139, 157)) ('lower', 'NegReg', (65, 70)) ('patients', 'Species', '9606', (27, 35)) ('IPF than dyskeratosis congentia', 'Disease', (74, 105)) ('IPF than dyskeratosis congentia', 'Disease', 'MESH:D054990', (74, 105)) ('pulmonary fibrosis', 'Phenotype', 'HP:0002206', (139, 157)) ('mutations', 'Var', (216, 225)) ('clinical', 'Species', '191496', (174, 182)) ('pulmonary fibrosis', 'Disease', 'MESH:D011658', (139, 157)) 3948 21323862 There are fifteen known mutations in hTERT and only two in hTERC (See Table 1); moreover, the mutations do not appear to cluster in any region or domain of either gene. ('hTERC', 'Gene', '7012', (59, 64)) ('hTERC', 'Gene', (59, 64)) ('hTERT', 'Gene', '7015', (37, 42)) ('mutations', 'Var', (24, 33)) ('hTERT', 'Gene', (37, 42)) 3950 21323862 Due to AA patients with telomerase mutations often having a family history of AML and patients suffering from AA and DC having a predisposition to AML, researchers looked for and found telomerase mutations in AML patients. ('AML', 'Disease', (209, 212)) ('AA', 'Phenotype', 'HP:0001903', (7, 9)) ('AML', 'Disease', 'MESH:D015470', (78, 81)) ('patients', 'Species', '9606', (86, 94)) ('AML', 'Disease', 'MESH:D015470', (209, 212)) ('AA', 'Phenotype', 'HP:0001903', (110, 112)) ('telomerase', 'Gene', (24, 34)) ('AML', 'Disease', 'MESH:D015470', (147, 150)) ('AML', 'Phenotype', 'HP:0004808', (209, 212)) ('patients', 'Species', '9606', (213, 221)) ('patients', 'Species', '9606', (10, 18)) ('AML', 'Disease', (78, 81)) ('AML', 'Phenotype', 'HP:0004808', (78, 81)) ('AML', 'Phenotype', 'HP:0004808', (147, 150)) ('AML', 'Disease', (147, 150)) ('mutations', 'Var', (35, 44)) 3953 21323862 recently reported that 6.8% of patients with AML had mutations in hTERT and no mutations were found in hTERC. ('hTERT', 'Gene', '7015', (66, 71)) ('AML', 'Phenotype', 'HP:0004808', (45, 48)) ('hTERC', 'Gene', '7012', (103, 108)) ('hTERC', 'Gene', (103, 108)) ('mutations', 'Var', (53, 62)) ('hTERT', 'Gene', (66, 71)) ('patients', 'Species', '9606', (31, 39)) ('AML', 'Disease', 'MESH:D015470', (45, 48)) ('AML', 'Disease', (45, 48)) 3954 21323862 The A1062T hTERT mutation was found to be the most common variant between cases and controls and constituted 60% of all hTERT mutations found in AML patients. ('AML', 'Phenotype', 'HP:0004808', (145, 148)) ('patients', 'Species', '9606', (149, 157)) ('AML', 'Disease', (145, 148)) ('hTERT', 'Gene', (11, 16)) ('A1062T', 'SUBSTITUTION', 'None', (4, 10)) ('A1062T', 'Var', (4, 10)) ('hTERT', 'Gene', '7015', (120, 125)) ('AML', 'Disease', 'MESH:D015470', (145, 148)) ('hTERT', 'Gene', '7015', (11, 16)) ('hTERT', 'Gene', (120, 125)) 3956 21323862 The A1062T mutation has been found in multiple haematopoietic cancers and may serve as a surrogate marker for mutations throughout hTERT as a screening tool to identify other malignancies with telomerase as a predisposing factor. ('found', 'Reg', (29, 34)) ('hTERT', 'Gene', '7015', (131, 136)) ('malignancies', 'Disease', 'MESH:D009369', (175, 187)) ('A1062T', 'Mutation', 'rs35719940', (4, 10)) ('cancer', 'Phenotype', 'HP:0002664', (62, 68)) ('hTERT', 'Gene', (131, 136)) ('malignancies', 'Disease', (175, 187)) ('A1062T', 'Var', (4, 10)) ('cancers', 'Disease', 'MESH:D009369', (62, 69)) ('cancers', 'Phenotype', 'HP:0002664', (62, 69)) ('cancers', 'Disease', (62, 69)) 3957 21323862 Although mutations in telomerase have been found to have important implications in growth and development, naturally occurring variations in hTERT and hTERC remain to be fully understood. ('hTERT', 'Gene', '7015', (141, 146)) ('mutations', 'Var', (9, 18)) ('growth', 'CPA', (83, 89)) ('hTERC', 'Gene', (151, 156)) ('hTERC', 'Gene', '7012', (151, 156)) ('hTERT', 'Gene', (141, 146)) ('development', 'CPA', (94, 105)) ('implications', 'Reg', (67, 79)) ('telomerase', 'Gene', (22, 32)) 3959 21323862 In fact, defects in telomerase function contribute to disease pathogenesis particularly in bone-marrow failure syndromes. ('telomerase', 'Protein', (20, 30)) ('defects', 'Var', (9, 16)) ('contribute', 'Reg', (40, 50)) ('bone-marrow failure syndromes', 'Disease', (91, 120)) ('bone-marrow failure', 'Phenotype', 'HP:0005528', (91, 110)) ('bone-marrow failure syndromes', 'Disease', 'MESH:D000080983', (91, 120)) 3960 21323862 Studies have also identified an association between telomerase mutations in both hTERC and hTERT with familial myelodysplastic syndrome and AML as mentioned previously. ('mutations', 'Var', (63, 72)) ('myelodysplastic syndrome', 'Phenotype', 'HP:0002863', (111, 135)) ('familial myelodysplastic syndrome', 'Disease', 'MESH:D009190', (102, 135)) ('AML', 'Disease', 'MESH:D015470', (140, 143)) ('telomerase', 'Protein', (52, 62)) ('familial myelodysplastic syndrome', 'Disease', (102, 135)) ('hTERT', 'Gene', '7015', (91, 96)) ('AML', 'Disease', (140, 143)) ('hTERT', 'Gene', (91, 96)) ('AML', 'Phenotype', 'HP:0004808', (140, 143)) ('hTERC', 'Gene', '7012', (81, 86)) ('hTERC', 'Gene', (81, 86)) 3961 21323862 Variations in the genes required to form the active telomerase complex may contribute to cancer risk. ('contribute', 'Reg', (75, 85)) ('cancer', 'Disease', 'MESH:D009369', (89, 95)) ('cancer', 'Disease', (89, 95)) ('Variations', 'Var', (0, 10)) ('cancer', 'Phenotype', 'HP:0002664', (89, 95)) 3962 21323862 The correlation between telomerase function and cancer is most likely due to the fact that tumour formation can be driven by telomerase dysfunction. ('tumour', 'Disease', 'MESH:D009369', (91, 97)) ('cancer', 'Phenotype', 'HP:0002664', (48, 54)) ('tumour', 'Disease', (91, 97)) ('driven by', 'Reg', (115, 124)) ('telomerase', 'Protein', (125, 135)) ('cancer', 'Disease', (48, 54)) ('cancer', 'Disease', 'MESH:D009369', (48, 54)) ('correlation', 'Reg', (4, 15)) ('dysfunction', 'Var', (136, 147)) ('tumour', 'Phenotype', 'HP:0002664', (91, 97)) 3963 21323862 Immortalization of tumour cells can occur through hTERT gene rearrangement which may contribute to cancer risk if variations in telomerase promote gene rearrangement and telomerase activation. ('tumour', 'Disease', 'MESH:D009369', (19, 25)) ('gene rearrangement', 'MPA', (147, 165)) ('promote', 'PosReg', (139, 146)) ('cancer', 'Disease', 'MESH:D009369', (99, 105)) ('tumour', 'Disease', (19, 25)) ('contribute', 'Reg', (85, 95)) ('cancer', 'Disease', (99, 105)) ('hTERT', 'Gene', '7015', (50, 55)) ('telomerase', 'Protein', (128, 138)) ('activation', 'PosReg', (181, 191)) ('variations', 'Var', (114, 124)) ('telomerase', 'Protein', (170, 180)) ('cancer', 'Phenotype', 'HP:0002664', (99, 105)) ('tumour', 'Phenotype', 'HP:0002664', (19, 25)) ('hTERT', 'Gene', (50, 55)) 3965 21323862 Variations in hTERC and hTERT have been found to contribute to overall telomere length implying a potential role for these variations in cancer risk. ('cancer', 'Phenotype', 'HP:0002664', (137, 143)) ('hTERC', 'Gene', (14, 19)) ('hTERT', 'Gene', (24, 29)) ('hTERC', 'Gene', '7012', (14, 19)) ('telomere', 'MPA', (71, 79)) ('cancer', 'Disease', 'MESH:D009369', (137, 143)) ('Variations', 'Var', (0, 10)) ('cancer', 'Disease', (137, 143)) ('contribute', 'Reg', (49, 59)) ('hTERT', 'Gene', '7015', (24, 29)) 3967 21323862 There is conflicting data regarding the role of hTERT variations and increased risk of breast cancer. ('hTERT', 'Gene', (48, 53)) ('breast cancer', 'Disease', 'MESH:D001943', (87, 100)) ('cancer', 'Phenotype', 'HP:0002664', (94, 100)) ('breast cancer', 'Disease', (87, 100)) ('breast cancer', 'Phenotype', 'HP:0003002', (87, 100)) ('variations', 'Var', (54, 64)) ('hTERT', 'Gene', '7015', (48, 53)) 3968 21323862 Although some studies have found an association between breast cancer risk and telomerase SNP, the most compelling data involve studies performed on a total of 1656 breast cancer samples and 2019 matched controls. ('breast cancer', 'Disease', 'MESH:D001943', (165, 178)) ('cancer', 'Phenotype', 'HP:0002664', (172, 178)) ('breast cancer', 'Disease', (165, 178)) ('breast cancer', 'Phenotype', 'HP:0003002', (165, 178)) ('cancer', 'Phenotype', 'HP:0002664', (63, 69)) ('telomerase', 'Var', (79, 89)) ('breast cancer', 'Disease', 'MESH:D001943', (56, 69)) ('breast cancer', 'Phenotype', 'HP:0003002', (56, 69)) ('association', 'Interaction', (36, 47)) ('breast cancer', 'Disease', (56, 69)) 3969 21323862 These compelling, high power studies did not find an association between increased breast cancer risk and the SNPs of the telomerase promoter. ('cancer', 'Phenotype', 'HP:0002664', (90, 96)) ('breast cancer', 'Disease', 'MESH:D001943', (83, 96)) ('breast cancer', 'Disease', (83, 96)) ('breast cancer', 'Phenotype', 'HP:0003002', (83, 96)) ('SNPs', 'Var', (110, 114)) 3970 21323862 did not observe a strong association of the SNP rs401681 in the TERT-CLPTM1L locus with cancer risk, related to telomere length. ('cancer', 'Disease', (88, 94)) ('cancer', 'Disease', 'MESH:D009369', (88, 94)) ('rs401681', 'Mutation', 'rs401681', (48, 56)) ('CLPTM1L', 'Gene', '81037', (69, 76)) ('cancer', 'Phenotype', 'HP:0002664', (88, 94)) ('CLPTM1L', 'Gene', (69, 76)) ('SNP rs401681', 'Var', (44, 56)) 3972 21323862 For example, variation in the TERT-CLPTM1L genes was found to be associated with a decreased risk of squamous cell carcinoma of the head and neck. ('carcinoma', 'Phenotype', 'HP:0030731', (115, 124)) ('variation', 'Var', (13, 22)) ('squamous cell carcinoma', 'Phenotype', 'HP:0002860', (101, 124)) ('decreased', 'NegReg', (83, 92)) ('CLPTM1L', 'Gene', '81037', (35, 42)) ('squamous cell carcinoma', 'Disease', 'MESH:D002294', (101, 124)) ('CLPTM1L', 'Gene', (35, 42)) ('squamous cell carcinoma', 'Disease', (101, 124)) 3976 21323862 Other studies have linked cancer risk and telomerase SNP for several cancers including the following: bladder cancer, ovarian cancer and cervix cancer. ('linked cancer', 'Disease', 'MESH:D009369', (19, 32)) ('ovarian cancer', 'Disease', (118, 132)) ('cervix cancer', 'Disease', (137, 150)) ('cancer', 'Phenotype', 'HP:0002664', (26, 32)) ('ovarian cancer', 'Phenotype', 'HP:0100615', (118, 132)) ('linked cancer', 'Disease', (19, 32)) ('cancer', 'Phenotype', 'HP:0002664', (144, 150)) ('cancers', 'Disease', 'MESH:D009369', (69, 76)) ('telomerase', 'Var', (42, 52)) ('bladder cancer', 'Disease', (102, 116)) ('bladder cancer', 'Disease', 'MESH:D001749', (102, 116)) ('cancer', 'Phenotype', 'HP:0002664', (126, 132)) ('bladder cancer', 'Phenotype', 'HP:0009725', (102, 116)) ('cervix cancer', 'Phenotype', 'HP:0030079', (137, 150)) ('cervix cancer', 'Disease', 'MESH:D002583', (137, 150)) ('cancer', 'Phenotype', 'HP:0002664', (110, 116)) ('cancers', 'Phenotype', 'HP:0002664', (69, 76)) ('ovarian cancer', 'Disease', 'MESH:D010051', (118, 132)) ('cancers', 'Disease', (69, 76)) ('cancer', 'Phenotype', 'HP:0002664', (69, 75)) 3977 21323862 What is the functional consequence of a SNP/variant in hTERT or hTERC? ('hTERT', 'Gene', '7015', (55, 60)) ('hTERT', 'Gene', (55, 60)) ('hTERC?', 'Gene', (64, 70)) ('SNP/variant', 'Var', (40, 51)) ('hTERC?', 'Gene', '7012', (64, 70)) 3978 21323862 It is known that telomerase is tightly controlled and complex; mutations or haploinsufficiency of hTERT/hTERC drive telomere dysfunction and bone marrow failures that are detrimental to the organism's survival. ('mutations', 'Var', (63, 72)) ('haploinsufficiency', 'Disease', (76, 94)) ('telomere', 'Gene', (116, 124)) ('bone marrow failures', 'Disease', (141, 161)) ('bone marrow failures', 'Disease', 'MESH:D000080983', (141, 161)) ('hTERC', 'Gene', (104, 109)) ('hTERT', 'Gene', '7015', (98, 103)) ('hTERC', 'Gene', '7012', (104, 109)) ('haploinsufficiency', 'Disease', 'MESH:D058495', (76, 94)) ('bone marrow failures', 'Phenotype', 'HP:0005528', (141, 161)) ('bone marrow failure', 'Phenotype', 'HP:0005528', (141, 160)) ('hTERT', 'Gene', (98, 103)) 3979 21323862 More research is needed to determine whether a variant in the hTERT promoter could alter the ability of a repressor to bind and maintain its tight control over its transcription and ultimately activity. ('tight control', 'MPA', (141, 154)) ('transcription', 'MPA', (164, 177)) ('hTERT', 'Gene', (62, 67)) ('variant', 'Var', (47, 54)) ('activity', 'MPA', (193, 201)) ('bind', 'Interaction', (119, 123)) ('alter', 'Reg', (83, 88)) ('ability', 'MPA', (93, 100)) ('maintain', 'Reg', (128, 136)) ('hTERT', 'Gene', '7015', (62, 67)) 3981 21323862 Furthermore, could a SNP in hTERC slightly alter structure of the RNA which in turn affects telomerase activity and telomere maintenance? ('SNP', 'Var', (21, 24)) ('affects', 'Reg', (84, 91)) ('hTERC', 'Gene', (28, 33)) ('alter', 'Reg', (43, 48)) ('hTERC', 'Gene', '7012', (28, 33)) ('telomere', 'MPA', (116, 124)) ('structure', 'MPA', (49, 58)) ('telomerase activity', 'MPA', (92, 111)) 3994 31723796 HiDAC as induction therapy has been explored by several groups including the EORTC-GIMEMA, the ALLG (formerly ALSG), SWOG, and the German AMLCG:based on the hypothesis that HiDAC as induction therapy will induce a better initial tumor cell kill and circumvent the development of resistance. ('HiDAC', 'Var', (173, 178)) ('tumor', 'Disease', 'MESH:D009369', (229, 234)) ('AML', 'Phenotype', 'HP:0004808', (138, 141)) ('AML', 'Disease', (138, 141)) ('tumor', 'Phenotype', 'HP:0002664', (229, 234)) ('tumor', 'Disease', (229, 234)) ('AML', 'Disease', 'MESH:D015470', (138, 141)) ('better', 'PosReg', (214, 220)) 4013 31723796 The covariates used in the model included gender, AML etiology (primary, secondary, unknown), cytogenetic risk stratification (intermediate, adverse, unknown), year of diagnosis, age group (<=40, >40) and WCC at diagnosis category (<50, >=50 to <100, >=100). ('<50', 'Var', (232, 235)) ('AML', 'Disease', (50, 53)) ('AML', 'Phenotype', 'HP:0004808', (50, 53)) ('AML', 'Disease', 'MESH:D015470', (50, 53)) 4037 31723796 The 5-year OS of patients undergoing alloHSCT in CR1 was similar in both the HiDAC induction cohort and the HiDAC consolidation cohort (5-year OS; 61% vs 61%, P = ns). ('CR1', 'Gene', (49, 52)) ('alloHSCT', 'Var', (37, 45)) ('HiDAC', 'Disease', (77, 82)) ('CR1', 'Gene', '1378', (49, 52)) ('patients', 'Species', '9606', (17, 25)) 4049 31723796 The improved CR rate with HiDAC-based induction therapy compared to SDAC-based induction therapy was also noted in the EORTC-GIMEMA study, although other, mostly older, studies have shown no differences. ('CR', 'Gene', '1401', (13, 15)) ('improved', 'PosReg', (4, 12)) ('HiDAC-based', 'Var', (26, 37)) 4055 31723796 Perhaps once CR is obtained, 2 cycles of HiDAC as consolidation (HiDAC consolidation cohort) are more effective at curing the patient compared with 2 cycles of SDAC (HiDAC induction cohort); or perhaps that HiDAC-based induction therapy, as a more intensive regimen, is better able to achieve a morphological CR but the increased proportion includes mostly patients with intrinsically resistant disease which is destined to relapse; or perhaps there were other unmeasured or unknown risk factors that were imbalanced between the 2 cohorts, such as the molecular mutations of FLT3, NPM1, and others. ('FLT3', 'Gene', '2322', (575, 579)) ('CR', 'Gene', '1401', (309, 311)) ('NPM1', 'Gene', '4869', (581, 585)) ('patient', 'Species', '9606', (357, 364)) ('patient', 'Species', '9606', (126, 133)) ('FLT3', 'Gene', (575, 579)) ('patients', 'Species', '9606', (357, 365)) ('mutations', 'Var', (562, 571)) ('NPM1', 'Gene', (581, 585)) ('CR', 'Gene', '1401', (13, 15)) 4071 31723796 Most, if not all AMLs, will have one or more mutations identified which are known to be important drivers of the leukemia, and which provide considerable prognostic information. ('mutations', 'Var', (45, 54)) ('AML', 'Disease', 'MESH:D015470', (17, 20)) ('AML', 'Disease', (17, 20)) ('leukemia', 'Phenotype', 'HP:0001909', (113, 121)) ('AML', 'Phenotype', 'HP:0004808', (17, 20)) ('leukemia', 'Disease', (113, 121)) ('leukemia', 'Disease', 'MESH:D007938', (113, 121)) 4078 27479497 In mouse cancer models driven by insertional mutagenesis, transposon insertions in tumor cells are detected with PCR-based approaches to enrich for transposon-containing tumor DNA fragmented by enzymatic digestion or random shearing. ('cancer', 'Disease', (9, 15)) ('cancer', 'Disease', 'MESH:D009369', (9, 15)) ('tumor', 'Disease', 'MESH:D009369', (170, 175)) ('tumor', 'Phenotype', 'HP:0002664', (83, 88)) ('mouse', 'Species', '10090', (3, 8)) ('tumor', 'Disease', (83, 88)) ('tumor', 'Phenotype', 'HP:0002664', (170, 175)) ('cancer', 'Phenotype', 'HP:0002664', (9, 15)) ('transposon-containing', 'Var', (148, 169)) ('tumor', 'Disease', (170, 175)) ('tumor', 'Disease', 'MESH:D009369', (83, 88)) 4084 27479497 Using Ion Torrent sequencing technology, SBCapSeq identifies insertions in a mouse model of AML in a semiquantitative manner whereby read depth can be used to identify clonally selected events. ('AML', 'Disease', 'MESH:D015470', (92, 95)) ('mouse', 'Species', '10090', (77, 82)) ('ML', 'Phenotype', 'HP:0012324', (93, 95)) ('AML', 'Disease', (92, 95)) ('SBCapSeq', 'Chemical', '-', (41, 49)) ('insertions', 'Var', (61, 71)) 4086 27479497 We developed a bioinformatics pipeline to identify SB insertions in bulk tumor and single cells by mapping quality scores of sequence reads that contain both SB transposons and flanking genomic sequences that map uniquely to the mouse genome. ('insertions', 'Var', (54, 64)) ('SB', 'Chemical', '-', (51, 53)) ('tumor', 'Disease', 'MESH:D009369', (73, 78)) ('tumor', 'Phenotype', 'HP:0002664', (73, 78)) ('tumor', 'Disease', (73, 78)) ('mouse', 'Species', '10090', (229, 234)) ('SB', 'Chemical', '-', (158, 160)) 4087 27479497 Finally, we integrated data from whole-genome sequencing (WGS), SB capture hybridization and RNA-seq and found that SB insertions cause acute ML (AML) through overexpression of genes in the MAP kinase (MAPK) and JAK-STAT signaling pathways and identified cooperating mutagenic events in single tumor cells. ('tumor', 'Disease', (294, 299)) ('overexpression', 'PosReg', (159, 173)) ('insertions', 'Var', (119, 129)) ('cause', 'Reg', (130, 135)) ('AML', 'Disease', (146, 149)) ('ML', 'Phenotype', 'HP:0012324', (147, 149)) ('ML', 'Gene', '14025', (147, 149)) ('ML', 'Phenotype', 'HP:0012324', (142, 144)) ('ML', 'Gene', '14025', (142, 144)) ('SB', 'Chemical', '-', (116, 118)) ('SB', 'Chemical', '-', (64, 66)) ('tumor', 'Disease', 'MESH:D009369', (294, 299)) ('tumor', 'Phenotype', 'HP:0002664', (294, 299)) ('GS', 'Disease', 'MESH:D011125', (59, 61)) ('AML', 'Disease', 'MESH:D015470', (146, 149)) 4088 27479497 SB transposition in mice carrying an inducible SB transposase and the Actb-Cre transgene drives a rapidly fatal hematopoietic disease prominently affecting the spleen (Fig. ('SB transposition', 'Phenotype', 'HP:0011540', (0, 16)) ('transposition', 'Var', (3, 16)) ('hematopoietic disease', 'Phenotype', 'HP:0001871', (112, 133)) ('hematopoietic disease', 'Disease', (112, 133)) ('drives', 'PosReg', (89, 95)) ('mice', 'Species', '10090', (20, 24)) ('Actb', 'Gene', (70, 74)) ('Actb', 'Gene', '11461', (70, 74)) ('hematopoietic disease', 'Disease', 'MESH:D019337', (112, 133)) ('SB', 'Chemical', '-', (47, 49)) ('SB', 'Chemical', '-', (0, 2)) 4090 27479497 Trp 53R172H/+ mice died earlier than Trp53+/- and wild-type mice and accumulated Trp53 protein in their tumor cells (Supplementary Fig. ('accumulated', 'PosReg', (69, 80)) ('Trp53', 'Gene', '22059', (37, 42)) ('tumor', 'Phenotype', 'HP:0002664', (104, 109)) ('Trp 53R172H/+', 'Var', (0, 13)) ('Trp53', 'Gene', '22059', (81, 86)) ('mice', 'Species', '10090', (14, 18)) ('tumor', 'Disease', (104, 109)) ('Trp53', 'Gene', (37, 42)) ('Trp53', 'Gene', (81, 86)) ('mice', 'Species', '10090', (60, 64)) ('tumor', 'Disease', 'MESH:D009369', (104, 109)) 4091 27479497 1b -d), results that are consistent with previous studies showing that missense mutations in Trp53 abrogate the tumor suppressor functions of Trp53 and promote malignant transformation. ('missense mutations', 'Var', (71, 89)) ('Trp53', 'Gene', (93, 98)) ('tumor', 'Disease', 'MESH:D009369', (112, 117)) ('Trp53', 'Gene', (142, 147)) ('tumor', 'Phenotype', 'HP:0002664', (112, 117)) ('malignant transformation', 'CPA', (160, 184)) ('Trp53', 'Gene', '22059', (142, 147)) ('Trp53', 'Gene', '22059', (93, 98)) ('tumor', 'Disease', (112, 117)) ('abrogate', 'NegReg', (99, 107)) ('promote', 'PosReg', (152, 159)) 4101 27479497 Another 110 CCGs, including Flt3 and Runx1, have human orthologs with mutations identified by exome or WGS in at least one AML genome (P = 3.5 x 10-7, Bonferroni corrected). ('GS', 'Disease', 'MESH:D011125', (104, 106)) ('mutations', 'Var', (70, 79)) ('Flt3', 'Gene', (28, 32)) ('CCGs', 'Chemical', '-', (12, 16)) ('AML', 'Disease', 'MESH:D015470', (123, 126)) ('human', 'Species', '9606', (49, 54)) ('ML', 'Phenotype', 'HP:0012324', (124, 126)) ('Flt3', 'Gene', '2322', (28, 32)) ('AML', 'Disease', (123, 126)) 4104 27479497 Previous studies suggest that most CCGs identified by SB mutagenesis function during tumor progression rather than initiation. ('tumor', 'Disease', (85, 90)) ('function', 'Reg', (69, 77)) ('CCGs', 'Chemical', '-', (35, 39)) ('SB', 'Chemical', '-', (54, 56)) ('mutagenesis', 'Var', (57, 68)) ('tumor', 'Disease', 'MESH:D009369', (85, 90)) ('tumor', 'Phenotype', 'HP:0002664', (85, 90)) ('CCGs', 'Disease', (35, 39)) 4108 27479497 AML patients who had mutations in one or more trunk drivers showed significantly P < 0.03 lower survival than patients lacking mutations in these genes (Supplementary Fig. ('patients', 'Species', '9606', (110, 118)) ('lower', 'NegReg', (90, 95)) ('AML', 'Disease', 'MESH:D015470', (0, 3)) ('patients', 'Species', '9606', (4, 12)) ('ML', 'Phenotype', 'HP:0012324', (1, 3)) ('survival', 'MPA', (96, 104)) ('AML', 'Disease', (0, 3)) ('mutations', 'Var', (21, 30)) 4109 27479497 Pathway enrichment analysis using all 466 CCGs identified MAPK and JAK-STAT signaling as significantly P < 0.001 enriched pathways (Supplementary Table 2), similar to what is observed in human AML Ninety-seven percent of our CCGs had a mutation in at least one gene involved in MAPK, JAK-STAT or Jun N-terminal kinase (JNK)-p38 MAPK pathways (Fig. ('Jun N-terminal kinase', 'Gene', (296, 317)) ('p38', 'Gene', (324, 327)) ('mutation', 'Var', (236, 244)) ('CCGs', 'Chemical', '-', (225, 229)) ('p38', 'Gene', '26416', (324, 327)) ('AML', 'Disease', (193, 196)) ('human', 'Species', '9606', (187, 192)) ('Jun N-terminal kinase', 'Gene', '5599', (296, 317)) ('CCGs', 'Chemical', '-', (42, 46)) ('ML', 'Phenotype', 'HP:0012324', (194, 196)) ('AML', 'Disease', 'MESH:D015470', (193, 196)) 4110 27479497 Notably, mutations in human homologs of the Sleeping Beauty-myeloid leukemia (SB-ML) CCGs that mapped to JAK-STAT or MAPK signaling are also associated with decreased survival in human AML (Supplementary Fig. ('AML', 'Disease', (185, 188)) ('human', 'Species', '9606', (179, 184)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (60, 76)) ('human', 'Species', '9606', (22, 27)) ('leukemia', 'Phenotype', 'HP:0001909', (68, 76)) ('Beauty-myeloid leukemia', 'Disease', 'MESH:D007951', (53, 76)) ('survival', 'CPA', (167, 175)) ('mutations', 'Var', (9, 18)) ('ML', 'Phenotype', 'HP:0012324', (186, 188)) ('CCGs', 'Chemical', '-', (85, 89)) ('ML', 'Gene', '14025', (186, 188)) ('SB', 'Chemical', '-', (78, 80)) ('AML', 'Disease', 'MESH:D015470', (185, 188)) ('Beauty-myeloid leukemia', 'Disease', (53, 76)) ('decreased', 'NegReg', (157, 166)) ('ML', 'Phenotype', 'HP:0012324', (81, 83)) ('ML', 'Gene', '14025', (81, 83)) 4111 27479497 The most frequently mutated CCGs were Erg, Ets1 and Fli1, which encode ETS-related transcription factors and were collectively mutated in 80% of tumors. ('Fli1', 'Gene', (52, 56)) ('Erg', 'Gene', (38, 41)) ('tumor', 'Phenotype', 'HP:0002664', (145, 150)) ('mutated', 'Var', (20, 27)) ('tumors', 'Disease', (145, 151)) ('Fli1', 'Gene', '14247', (52, 56)) ('tumors', 'Phenotype', 'HP:0002664', (145, 151)) ('tumors', 'Disease', 'MESH:D009369', (145, 151)) ('CCGs', 'Chemical', '-', (28, 32)) ('CCGs', 'Gene', (28, 32)) ('Ets1', 'Gene', (43, 47)) 4112 27479497 We also identified statistically significant (P < 0.05) co-occurrences of SB insertions in trunk drivers such as Ghr and Jak2 and mutually exclusive insertions in trunk drivers such as Erg and Ets1. ('SB', 'Chemical', '-', (74, 76)) ('Jak2', 'Gene', (121, 125)) ('Jak2', 'Gene', '16452', (121, 125)) ('insertions', 'Var', (77, 87)) 4113 27479497 2c) indicate that cooperating oncogenic mutations drive leukemogenesis in this mouse model of human AML. ('mouse', 'Species', '10090', (79, 84)) ('leukemogenesis', 'Disease', (56, 70)) ('mutations', 'Var', (40, 49)) ('ML', 'Phenotype', 'HP:0012324', (101, 103)) ('AML', 'Disease', 'MESH:D015470', (100, 103)) ('human', 'Species', '9606', (94, 99)) ('AML', 'Disease', (100, 103)) 4114 27479497 On average, each SB-induced ML contains 17.2 insertional mutations in CCGs, reflecting the high degree of intra- and intertumor heterogeneity of these tumors. ('CCGs', 'Chemical', '-', (70, 74)) ('tumor', 'Phenotype', 'HP:0002664', (151, 156)) ('CCGs', 'Gene', (70, 74)) ('ML', 'Phenotype', 'HP:0012324', (28, 30)) ('insertional mutations', 'Var', (45, 66)) ('tumors', 'Disease', (151, 157)) ('tumor', 'Disease', (151, 156)) ('tumors', 'Disease', 'MESH:D009369', (151, 157)) ('tumor', 'Disease', 'MESH:D009369', (122, 127)) ('ML', 'Gene', '14025', (28, 30)) ('tumors', 'Phenotype', 'HP:0002664', (151, 157)) ('tumor', 'Phenotype', 'HP:0002664', (122, 127)) ('SB', 'Chemical', '-', (17, 19)) ('tumor', 'Disease', (122, 127)) ('tumor', 'Disease', 'MESH:D009369', (151, 156)) 4122 27479497 We also assessed the reproducibility of SBCapSeq to identify the same SB insertions using two independent sequencing runs for one tumor and found an overall high correlation between shared insertions and sequencing depth across experiments (Supplementary Fig. ('SB', 'Chemical', '-', (70, 72)) ('tumor', 'Disease', 'MESH:D009369', (130, 135)) ('SB', 'Chemical', '-', (40, 42)) ('insertions', 'Var', (73, 83)) ('SBCapSeq', 'Chemical', '-', (40, 48)) ('tumor', 'Phenotype', 'HP:0002664', (130, 135)) ('tumor', 'Disease', (130, 135)) 4129 27479497 4b), and detected 18 CCGs in the 10 resampled tumors with at least two independent insertions in a single tumor (possibly a major clone and a minor clone), as defined by read counts for individual insertions. ('tumor', 'Disease', 'MESH:D009369', (46, 51)) ('tumor', 'Phenotype', 'HP:0002664', (46, 51)) ('tumor', 'Disease', (46, 51)) ('tumors', 'Phenotype', 'HP:0002664', (46, 52)) ('CCGs', 'Chemical', '-', (21, 25)) ('tumor', 'Disease', 'MESH:D009369', (106, 111)) ('tumor', 'Phenotype', 'HP:0002664', (106, 111)) ('insertions', 'Var', (83, 93)) ('tumors', 'Disease', (46, 52)) ('tumor', 'Disease', (106, 111)) ('tumors', 'Disease', 'MESH:D009369', (46, 52)) 4134 27479497 Hierarchical clustering of insertions from splink 454, splink HiSeq and SBCapSeq shared by at least two platforms showed that in all but one sample, the data from SBCapSeq were closely related to the splink HiSeq output (Supplementary Fig. ('splink HiSeq output', 'MPA', (200, 219)) ('HiSeq', 'Chemical', '-', (62, 67)) ('insertions', 'Var', (27, 37)) ('SBCapSeq', 'Chemical', '-', (72, 80)) ('SBCapSeq', 'Chemical', '-', (163, 171)) ('HiSeq', 'Chemical', '-', (207, 212)) 4137 27479497 Notably, SBCapSeq yielded <0.01% cross-contamination of insertions in more than one sample, showing its higher sensitivity than Splink_HiSeq methods and potentially expanding the sequencing capability to single-cell sequencing. ('SBCapSeq', 'Gene', (9, 17)) ('HiSeq', 'Chemical', '-', (135, 140)) ('insertions', 'Var', (56, 66)) ('SBCapSeq', 'Chemical', '-', (9, 17)) ('sensitivity', 'MPA', (111, 122)) 4144 27479497 SBCapSeq detected all but three of the insertions identified by WGS in D5 with >=2 reads, at an average 947-fold increased read depth, and the read depth across the two platforms for the ML CCGs mutated in this tumor was highly correlated (Fig. ('CCGs', 'Chemical', '-', (190, 194)) ('tumor', 'Disease', 'MESH:D009369', (211, 216)) ('ML', 'Phenotype', 'HP:0012324', (187, 189)) ('tumor', 'Phenotype', 'HP:0002664', (211, 216)) ('tumor', 'Disease', (211, 216)) ('increased', 'PosReg', (113, 122)) ('insertions', 'Var', (39, 49)) ('ML', 'Gene', '14025', (187, 189)) ('GS', 'Disease', 'MESH:D011125', (65, 67)) ('mutated', 'Var', (195, 202)) ('SBCapSeq', 'Chemical', '-', (0, 8)) 4154 27479497 WGS showed that the D5 leukemia has a 'quiet' genome containing only 1,080 significant single-nucleotide variations (SNVs) and 160 significant insertions or deletions (indels), most of which map to introns and non-coding regions (Supplementary Fig. ('leukemia', 'Disease', (23, 31)) ('insertions', 'Var', (143, 153)) ('deletions', 'Var', (157, 166)) ('GS', 'Disease', 'MESH:D011125', (1, 3)) ('single-nucleotide', 'Var', (87, 104)) ('leukemia', 'Phenotype', 'HP:0001909', (23, 31)) ('leukemia', 'Disease', 'MESH:D007938', (23, 31)) 4155 27479497 Only five events occurred within exons, including a frameshift mutation in Olfr429; three nonsynonymous SNVs in Krt5, Tdrd6 and Tenm3 ; and one synonymous SNV in Lactb. ('Tenm3', 'Gene', '23965', (128, 133)) ('Krt5', 'Gene', (112, 116)) ('Krt5', 'Gene', '110308', (112, 116)) ('Tdrd6', 'Gene', (118, 123)) ('Olfr429', 'Gene', '258717', (75, 82)) ('Tenm3', 'Gene', (128, 133)) ('Tdrd6', 'Gene', '210510', (118, 123)) ('Olfr429', 'Gene', (75, 82)) ('frameshift mutation', 'Var', (52, 71)) 4160 27479497 Together, these data strongly suggest that SB mutagenesis is the predominant force driving tumor evolution. ('tumor', 'Disease', 'MESH:D009369', (91, 96)) ('mutagenesis', 'Var', (46, 57)) ('SB', 'Chemical', '-', (43, 45)) ('tumor', 'Phenotype', 'HP:0002664', (91, 96)) ('tumor', 'Disease', (91, 96)) 4162 27479497 Each single-cell genome was screened for the presence or absence of a subset of SB insertions that are highly represented in the D5 bulk tumor, including insertions in trunk drivers Ets1 and Notch1 and in Ankib1, Phf2, Kdm4c and Zfp704 (Supplementary Fig. ('insertions', 'Var', (154, 164)) ('tumor', 'Disease', (137, 142)) ('SB', 'Chemical', '-', (80, 82)) ('Zfp704', 'Gene', (229, 235)) ('Zfp704', 'Gene', '170753', (229, 235)) ('Phf2', 'Gene', '18676', (213, 217)) ('Ankib1', 'Gene', (205, 211)) ('Notch1', 'Gene', (191, 197)) ('Ets1', 'Gene', (182, 186)) ('Kdm4c', 'Gene', '76804', (219, 224)) ('Kdm4c', 'Gene', (219, 224)) ('tumor', 'Disease', 'MESH:D009369', (137, 142)) ('Ankib1', 'Gene', '70797', (205, 211)) ('tumor', 'Phenotype', 'HP:0002664', (137, 142)) ('Phf2', 'Gene', (213, 217)) 4164 27479497 This heterogeneity of SB insertion patterns was unexpected, given that insertions in these genes were represented by high read counts in bulk tumor DNA. ('tumor', 'Phenotype', 'HP:0002664', (142, 147)) ('tumor', 'Disease', (142, 147)) ('SB', 'Chemical', '-', (22, 24)) ('insertions', 'Var', (71, 81)) ('tumor', 'Disease', 'MESH:D009369', (142, 147)) 4166 27479497 A total of 3,819 unique insertions were identified in the 26 single-cell genomes, with a median of 147 unique insertions per cell; 16.3% (622/3,819) of the insertions mapped to the donor chromosome, whereas 83.7% (3,197/3,819) mapped to non-donor chromosomes. ('insertions', 'Var', (156, 166)) ('donor', 'Species', '9606', (241, 246)) ('donor', 'Species', '9606', (181, 186)) ('mapped', 'Reg', (167, 173)) 4169 27479497 Only 27% of these genes were found to have insertions in the bulk tumor DNA by WGS or SBCapSeq. ('tumor', 'Phenotype', 'HP:0002664', (66, 71)) ('GS', 'Disease', 'MESH:D011125', (80, 82)) ('tumor', 'Disease', (66, 71)) ('SBCapSeq', 'Chemical', '-', (86, 94)) ('tumor', 'Disease', 'MESH:D009369', (66, 71)) ('insertions', 'Var', (43, 53)) 4171 27479497 We also identified SB insertions in 58 ML CCGs in single cells not found in the bulk tumor by SBCapSeq or WGS, suggesting that these events were subclonal in this leukemia. ('tumor', 'Disease', (85, 90)) ('leukemia', 'Disease', 'MESH:D007938', (163, 171)) ('SB', 'Chemical', '-', (94, 96)) ('ML', 'Phenotype', 'HP:0012324', (39, 41)) ('SBCapSeq', 'Chemical', '-', (94, 102)) ('SB', 'Chemical', '-', (19, 21)) ('tumor', 'Disease', 'MESH:D009369', (85, 90)) ('leukemia', 'Disease', (163, 171)) ('leukemia', 'Phenotype', 'HP:0001909', (163, 171)) ('ML', 'Gene', '14025', (39, 41)) ('CCGs', 'Chemical', '-', (42, 46)) ('tumor', 'Phenotype', 'HP:0002664', (85, 90)) ('GS', 'Disease', 'MESH:D011125', (107, 109)) ('insertions', 'Var', (22, 32)) 4177 27479497 Insertions in several other loci, including Kdm4c, Phf2 and Ankib1, were also coincident with insertions in Erg and Ghr. ('Ankib1', 'Gene', '70797', (60, 66)) ('Phf2', 'Gene', '18676', (51, 55)) ('Erg', 'Gene', (108, 111)) ('Insertions', 'Var', (0, 10)) ('Kdm4c', 'Gene', '76804', (44, 49)) ('Kdm4c', 'Gene', (44, 49)) ('Ghr', 'Gene', (116, 119)) ('Ankib1', 'Gene', (60, 66)) ('insertions', 'Var', (94, 104)) ('Phf2', 'Gene', (51, 55)) 4179 27479497 A cluster of three Ter119+ cells at the right of the heat map had insertions in Ets1 and Notch1. ('Notch1', 'Gene', (89, 95)) ('Ter119', 'Gene', (19, 25)) ('insertions', 'Var', (66, 76)) ('Ter119', 'Gene', '104231', (19, 25)) ('Ets1', 'Gene', (80, 84)) 4180 27479497 Another Ter119+ cell (D5_SP_SC17) carried the Ets1 insertion but lacked the Notch1 insertion, whereas two Ter119+ cells (D5_SP_SC06 and D5_SP_SC09) had the Notch1 insertion but lacked the Ets1 insertion. ('Ter119', 'Gene', (8, 14)) ('Ter119', 'Gene', '104231', (106, 112)) ('lacked', 'NegReg', (65, 71)) ('insertion', 'Var', (51, 60)) ('Notch1 insertion', 'MPA', (76, 92)) ('Ter119', 'Gene', '104231', (8, 14)) ('Ter119', 'Gene', (106, 112)) ('Ets1', 'Gene', (46, 50)) 4181 27479497 When the analysis was expanded to include all insertions that mapped to ML CCGs and not just those shared in three or more cells, we found that most CCG insertions were unique to a single cell (Supplementary Fig. ('ML', 'Phenotype', 'HP:0012324', (72, 74)) ('ML', 'Gene', '14025', (72, 74)) ('CCGs', 'Chemical', '-', (75, 79)) ('CCG', 'Gene', (149, 152)) ('insertions', 'Var', (153, 163)) 4183 27479497 Mapped SB insertions in single cells and D5 bulk tumor cells identified by SBCapSeq and SB insertions identified in the ML population by splink 454 for the trunk drivers Erg, Ghr, Ets1, Notch1 and Akt1 showed activating driver events predicted to upregulate gene expression from the minimal promoter from the transposon (Supplementary Fig. ('tumor', 'Disease', 'MESH:D009369', (49, 54)) ('tumor', 'Phenotype', 'HP:0002664', (49, 54)) ('SB', 'Chemical', '-', (75, 77)) ('Notch1', 'Gene', (186, 192)) ('tumor', 'Disease', (49, 54)) ('SB', 'Chemical', '-', (88, 90)) ('upregulate', 'PosReg', (247, 257)) ('gene expression', 'MPA', (258, 273)) ('ML', 'Phenotype', 'HP:0012324', (120, 122)) ('SBCapSeq', 'Chemical', '-', (75, 83)) ('Akt1', 'Gene', (197, 201)) ('insertions', 'Var', (91, 101)) ('SB', 'Chemical', '-', (7, 9)) ('ML', 'Gene', '14025', (120, 122)) 4188 27479497 CCGs with SB insertions in the sense orientation relative to the gene showed increased expression in the tumor over normal spleen; the same was true of insertions private to the D5 tumor predicted to activate gene expression (Supplementary Table 6). ('insertions', 'Var', (13, 23)) ('tumor', 'Disease', (181, 186)) ('tumor', 'Disease', 'MESH:D009369', (105, 110)) ('expression', 'MPA', (87, 97)) ('CCGs', 'Chemical', '-', (0, 4)) ('tumor', 'Phenotype', 'HP:0002664', (105, 110)) ('tumor', 'Disease', 'MESH:D009369', (181, 186)) ('increased', 'PosReg', (77, 86)) ('tumor', 'Disease', (105, 110)) ('tumor', 'Phenotype', 'HP:0002664', (181, 186)) ('SB', 'Chemical', '-', (10, 12)) 4189 27479497 Insertions with high read counts led to greater changes in gene expression; however, insertions in CCGs and genes private to the D5 tumor oriented in the antisense direction rarely led to decreased gene expression in the tumor, indicating that they are passenger insertions and that this ML is largely an oncogene-driven disease. ('ML', 'Gene', '14025', (288, 290)) ('decreased', 'NegReg', (188, 197)) ('insertions', 'Var', (85, 95)) ('gene expression', 'MPA', (198, 213)) ('tumor', 'Disease', 'MESH:D009369', (132, 137)) ('tumor', 'Phenotype', 'HP:0002664', (221, 226)) ('tumor', 'Disease', (221, 226)) ('tumor', 'Phenotype', 'HP:0002664', (132, 137)) ('CCGs', 'Chemical', '-', (99, 103)) ('tumor', 'Disease', (132, 137)) ('CCGs', 'Gene', (99, 103)) ('ML', 'Phenotype', 'HP:0012324', (288, 290)) ('tumor', 'Disease', 'MESH:D009369', (221, 226)) 4190 27479497 Figure 7 shows all of the genes identified in the two leukemic subpopulations, anchored by insertions in Erg and Ghr, or Ets1 and Notch 1. ('leukemic', 'Disease', (54, 62)) ('Ghr', 'Gene', (113, 116)) ('Notch 1', 'Gene', (130, 137)) ('Erg', 'Gene', (105, 108)) ('Ets1', 'Gene', (121, 125)) ('leukemic', 'Disease', 'MESH:D007938', (54, 62)) ('insertions', 'Var', (91, 101)) 4203 27479497 In murine models, high Erg expression induces both T cell acute lymphoblastic leukemia (T-ALL) and AML, and high ERG expression in both human AML and early T cell precursor ALL is associated with poor clinical outcome. ('human', 'Species', '9606', (136, 141)) ('leukemia', 'Phenotype', 'HP:0001909', (78, 86)) ('acute lymphoblastic leukemia', 'Phenotype', 'HP:0006721', (58, 86)) ('AML', 'Disease', (142, 145)) ('ML', 'Phenotype', 'HP:0012324', (100, 102)) ('ERG', 'Gene', '13876', (113, 116)) ('AML', 'Disease', 'MESH:D015470', (99, 102)) ('induces', 'Reg', (38, 45)) ('lymphoblastic leukemia', 'Phenotype', 'HP:0005526', (64, 86)) ('lymphoblastic leukemia', 'Disease', (64, 86)) ('murine', 'Species', '10090', (3, 9)) ('AML', 'Disease', (99, 102)) ('lymphoblastic leukemia', 'Disease', 'MESH:D054198', (64, 86)) ('AML', 'Disease', 'MESH:D015470', (142, 145)) ('ERG', 'Gene', (113, 116)) ('ML', 'Phenotype', 'HP:0012324', (143, 145)) ('high Erg expression', 'Var', (18, 37)) 4208 27479497 Consistent with this, the subpopulation of leukemic cells containing Notch1 and Ets1 insertions in our data set expressed Ter119, a marker of late differentiation of erythroid cells. ('Ter119', 'Gene', '104231', (122, 128)) ('insertions', 'Var', (85, 95)) ('leukemic', 'Disease', (43, 51)) ('Notch1', 'Gene', (69, 75)) ('Ter119', 'Gene', (122, 128)) ('leukemic', 'Disease', 'MESH:D007938', (43, 51)) ('Ets1', 'Gene', (80, 84)) 4209 27479497 One unanticipated finding of our analysis was the large number of unique transposon insertions in every tumor cell. ('tumor', 'Phenotype', 'HP:0002664', (104, 109)) ('tumor', 'Disease', (104, 109)) ('transposon insertions', 'Var', (73, 94)) ('tumor', 'Disease', 'MESH:D009369', (104, 109)) 4213 27479497 Lineage tracing in adult mice showed several-hundred-fold larger myeloid than lymphoid output, which could explain why the myeloid compartment is particularly susceptible to the effects of SB mutagenesis. ('mutagenesis', 'Var', (192, 203)) ('mice', 'Species', '10090', (25, 29)) ('myeloid', 'CPA', (65, 72)) ('SB', 'Chemical', '-', (189, 191)) 4223 27479497 The resulting cohorts of mice were on mixed genetic backgrounds consisting of C57BL/6J, 129, C3H and FVB. ('mice', 'Species', '10090', (25, 29)) ('C57BL/6J', 'Var', (78, 86)) ('C3H', 'Var', (93, 96)) 4249 27479497 If they mapped with MAPQ >=30 and no secondary alignments, then they were considered to have a distinct IRDR (left or right). ('IRDR', 'Gene', (104, 108)) ('IRDR', 'Gene', '14685', (104, 108)) ('MAPQ >=30', 'Var', (20, 29)) 4265 27479497 Mapping of SB insertions to the mouse genome used a custom splink HiSeq bioinformatics workflow modeled to perform similarly to the splink 454 analysis pipeline (J.Y.N., K.M.M., N.A.J., N.G.C. ('SB', 'Chemical', '-', (11, 13)) ('HiSeq', 'Chemical', '-', (66, 71)) ('K.M.M.', 'Var', (170, 176)) ('J.Y.N.', 'Var', (162, 168)) ('mouse', 'Species', '10090', (32, 37)) 4290 27479497 To do this, we first identified SB insertions as being activating or inactivating: activating when the insertion orientation (defined with respect to the chromosome) is sense with the gene, and inactivating when the insertion orientation is antisense with the gene. ('SB', 'Chemical', '-', (32, 34)) ('activating', 'PosReg', (83, 93)) ('insertions', 'Var', (35, 45)) 4298 29097609 The mechanism of action as well as the safety and tolerability of drugs in pediatric patients, including compounds that target CDK4/CDK6 (palbociclib, ribociclib, abemaciclib), aurora kinases (AT9283 and MLN8237), Wee1 kinase (MK-1775), KSP (ispinesib) and tubulin (taxanes, vinca alkaloids), are presented. ('vinca alkaloids', 'Chemical', 'MESH:D014748', (275, 290)) ('MLN8237', 'Var', (204, 211)) ('MLN8237', 'Chemical', 'MESH:C550258', (204, 211)) ('palbociclib', 'Chemical', 'MESH:C500026', (138, 149)) ('CDK4', 'Gene', (127, 131)) ('AT9283', 'Chemical', 'MESH:C535237', (193, 199)) ('taxanes', 'Chemical', 'MESH:D043823', (266, 273)) ('patients', 'Species', '9606', (85, 93)) ('Wee1', 'Gene', (214, 218)) ('AT9283', 'Var', (193, 199)) ('ispinesib', 'Chemical', 'MESH:C508757', (242, 251)) ('CDK4', 'Gene', '1019', (127, 131)) ('CDK6', 'Gene', (132, 136)) ('Wee1', 'Gene', '7465', (214, 218)) ('CDK6', 'Gene', '1021', (132, 136)) ('KSP', 'Chemical', '-', (237, 240)) ('MK-1775', 'Chemical', 'MESH:C549567', (227, 234)) 4310 29097609 Pharmacological inhibition of CDKs typically results in cell cycle arrest, apoptosis, and transcriptional repression to provide the rationale for therapeutically targeting CDKs in cancer. ('cell cycle arrest', 'Phenotype', 'HP:0011018', (56, 73)) ('CDKs', 'Gene', '983;1017;1019;12567;1021;12571;1022;1025', (30, 34)) ('results in', 'Reg', (45, 55)) ('apoptosis', 'CPA', (75, 84)) ('CDKs', 'Gene', (30, 34)) ('arrest', 'Disease', 'MESH:D006323', (67, 73)) ('CDKs', 'Gene', '983;1017;1019;12567;1021;12571;1022;1025', (172, 176)) ('cancer', 'Phenotype', 'HP:0002664', (180, 186)) ('transcriptional repression', 'CPA', (90, 116)) ('CDKs', 'Gene', (172, 176)) ('cancer', 'Disease', 'MESH:D009369', (180, 186)) ('arrest', 'Disease', (67, 73)) ('cancer', 'Disease', (180, 186)) ('inhibition', 'Var', (16, 26)) 4312 29097609 CDK4/6 inhibitors have emerged as promising cell-cycle therapeutics. ('inhibitors', 'Var', (7, 17)) ('CDK4/6', 'Gene', (0, 6)) ('CDK4/6', 'Gene', '1019;1021', (0, 6)) 4319 29097609 High expression of D-type cyclins, genetic mutations or amplification of the CDK4 and CDK6 loci, or loss of the p16Ink4A inhibitory protein that regulates cyclin D/CDK4/6 complexes, are associated with unrestricted cancer cell growth. ('p16Ink4A', 'Gene', '1029', (112, 120)) ('expression', 'MPA', (5, 15)) ('cyclin', 'Gene', (26, 32)) ('CDK4', 'Gene', (77, 81)) ('CDK4', 'Gene', (164, 168)) ('loss', 'Var', (100, 104)) ('unrestricted cancer', 'Disease', 'MESH:D009369', (202, 221)) ('CDK4', 'Gene', '1019', (77, 81)) ('CDK4/6', 'Gene', (164, 170)) ('cyclin', 'Gene', '5111', (155, 161)) ('genetic mutations', 'Var', (35, 52)) ('CDK4', 'Gene', '1019', (164, 168)) ('CDK6', 'Gene', '1021', (86, 90)) ('cancer', 'Phenotype', 'HP:0002664', (215, 221)) ('p16Ink4A', 'Gene', (112, 120)) ('unrestricted cancer', 'Disease', (202, 221)) ('associated', 'Reg', (186, 196)) ('CDK6', 'Gene', (86, 90)) ('cyclin', 'Gene', (155, 161)) ('amplification', 'Var', (56, 69)) ('cyclin', 'Gene', '5111', (26, 32)) ('CDK4/6', 'Gene', '1019;1021', (164, 170)) 4320 29097609 In addition, deletion of RB1 occurs in many tumors and accelerates proliferation independently of cyclin D-CDK4/6 activity. ('tumors', 'Disease', (44, 50)) ('tumors', 'Disease', 'MESH:D009369', (44, 50)) ('tumors', 'Phenotype', 'HP:0002664', (44, 50)) ('RB1', 'Gene', '5925', (25, 28)) ('cyclin', 'Gene', '5111', (98, 104)) ('deletion', 'Var', (13, 21)) ('CDK4/6', 'Gene', '1019;1021', (107, 113)) ('proliferation', 'CPA', (67, 80)) ('cyclin', 'Gene', (98, 104)) ('tumor', 'Phenotype', 'HP:0002664', (44, 49)) ('accelerates', 'PosReg', (55, 66)) ('RB1', 'Gene', (25, 28)) ('CDK4/6', 'Gene', (107, 113)) 4322 29097609 Currently, three selective CDK4/6 inhibitors are approved: palbociclib (PD-0332991), ribociclib (LEE011), and abemaciclib (LY2835219). ('LY2835219', 'Var', (123, 132)) ('PD-0332991', 'Var', (72, 82)) ('palbociclib', 'Chemical', 'MESH:C500026', (59, 70)) ('LY2835219', 'Chemical', 'MESH:C000590451', (123, 132)) ('LEE011', 'Var', (97, 103)) ('CDK4/6', 'Gene', '1019;1021', (27, 33)) ('PD-0332991', 'Chemical', 'MESH:C500026', (72, 82)) ('CDK4/6', 'Gene', (27, 33)) 4328 29097609 In another investigation, palbociclib plus PLX4720, an inhibitor against v-raf murine sarcoma viral oncogene homolog B1 (BRAF) extended survival in pediatric malignant astrocytoma, relative to either monotherapy. ('astrocytoma', 'Disease', (168, 179)) ('sarcoma', 'Phenotype', 'HP:0100242', (86, 93)) ('v-raf murine sarcoma viral oncogene homolog B1', 'Gene', (73, 119)) ('astrocytoma', 'Phenotype', 'HP:0009592', (168, 179)) ('extended', 'PosReg', (127, 135)) ('PLX4720', 'Var', (43, 50)) ('palbociclib', 'Chemical', 'MESH:C500026', (26, 37)) ('PLX4720', 'Chemical', 'MESH:C528407', (43, 50)) ('survival', 'MPA', (136, 144)) ('astrocytoma', 'Disease', 'MESH:D001254', (168, 179)) ('v-raf murine sarcoma viral oncogene homolog B1', 'Gene', '673', (73, 119)) 4329 29097609 Response was specific for a subset of pediatric astrocytomas with BRAF (V600E) mutation and CDKN2A (encoding cyclin-dependent kinase inhibitor 2A) deficiency. ('CDKN2A', 'Gene', (92, 98)) ('cyclin-dependent kinase inhibitor 2A', 'Gene', '1029', (109, 145)) ('pediatric astrocytomas', 'Phenotype', 'HP:0009592', (38, 60)) ('deficiency', 'Var', (147, 157)) ('CDKN2A', 'Gene', '1029', (92, 98)) ('cyclin-dependent kinase inhibitor 2A', 'Gene', (109, 145)) ('astrocytoma', 'Phenotype', 'HP:0009592', (48, 59)) ('pediatric astrocytomas', 'Disease', 'MESH:D001254', (38, 60)) ('pediatric astrocytomas', 'Disease', (38, 60)) ('BRAF', 'Gene', (66, 70)) ('V600E', 'Mutation', 'rs113488022', (72, 77)) 4341 29097609 Ribociclib was active in vitro in leukemia cells and in vivo in mutant NOTCH1-driven T-ALL mouse models in combination therapy with corticosteroids and mTOR inhibitors. ('leukemia', 'Disease', (34, 42)) ('mTOR', 'Gene', (152, 156)) ('NOTCH1', 'Gene', (71, 77)) ('leukemia', 'Disease', 'MESH:D007938', (34, 42)) ('leukemia', 'Phenotype', 'HP:0001909', (34, 42)) ('mTOR', 'Gene', '56717', (152, 156)) ('mouse', 'Species', '10090', (91, 96)) ('mutant', 'Var', (64, 70)) ('NOTCH1', 'Gene', '18128', (71, 77)) 4362 29097609 Preclinical evaluation of SCH 727965 by the Pediatric Preclinical Testing Program (PTPP) against a panel of human cell lines and pediatric-derived xenografts and ALL, induced significant delays in event free survival distribution compared to control in 64% of solid tumor xenografts and in 43% of ALL xenografts. ('tumor', 'Disease', 'MESH:D009369', (266, 271)) ('tumor', 'Phenotype', 'HP:0002664', (266, 271)) ('human', 'Species', '9606', (108, 113)) ('delays', 'NegReg', (187, 193)) ('SCH 727965', 'Chemical', 'MESH:C553669', (26, 36)) ('SCH 727965', 'Var', (26, 36)) ('tumor', 'Disease', (266, 271)) ('event', 'MPA', (197, 202)) 4380 29097609 AT9283 is a multikinase inhibitor with high potency against Aurora A and Aurora B kinase activity. ('Aurora A', 'Gene', (60, 68)) ('AT9283', 'Var', (0, 6)) ('Aurora B', 'Gene', (73, 81)) ('Aurora B', 'Gene', '9212', (73, 81)) ('Aurora A', 'Gene', '6790', (60, 68)) ('AT9283', 'Chemical', 'MESH:C535237', (0, 6)) 4381 29097609 AT9283 also inhibits Janus kinase 2 (JAK2) and JAK3, FMS-like tyrosine kinase 3 (FLT3), and Abelson tyrosine kinase (ABLT315I) with lower efficacy. ('tyrosine', 'Chemical', 'MESH:D014443', (62, 70)) ('FMS-like tyrosine kinase 3', 'Gene', '2322', (53, 79)) ('Janus kinase 2', 'Gene', '3717', (21, 35)) ('AT9283', 'Chemical', 'MESH:C535237', (0, 6)) ('JAK2', 'Gene', (37, 41)) ('ABLT315I', 'Mutation', 'rs121913459', (117, 125)) ('AT9283', 'Var', (0, 6)) ('JAK3', 'Gene', (47, 51)) ('JAK3', 'Gene', '3718', (47, 51)) ('FLT3', 'Gene', '2322', (81, 85)) ('inhibits', 'NegReg', (12, 20)) ('FMS-like tyrosine kinase 3', 'Gene', (53, 79)) ('Janus kinase 2', 'Gene', (21, 35)) ('FLT3', 'Gene', (81, 85)) ('tyrosine', 'Chemical', 'MESH:D014443', (100, 108)) ('JAK2', 'Gene', '3717', (37, 41)) 4382 29097609 Selective inhibition of Aurora-A blocks Thr228 phosphorylation, promotes formation of monopolar spindles, cell cycle arrest at G2/M-phase and apoptosis. ('promotes', 'PosReg', (64, 72)) ('phosphorylation', 'MPA', (47, 62)) ('Thr228', 'Protein', (40, 46)) ('formation', 'MPA', (73, 82)) ('arrest', 'Disease', (117, 123)) ('Aurora-A', 'Gene', (24, 32)) ('arrest', 'Disease', 'MESH:D006323', (117, 123)) ('Thr228', 'Chemical', '-', (40, 46)) ('Aurora-A', 'Gene', '6790', (24, 32)) ('apoptosis', 'CPA', (142, 151)) ('inhibition', 'Var', (10, 20)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (106, 123)) ('blocks', 'NegReg', (33, 39)) 4383 29097609 Preclinical evidence of activity of AT9283 was demonstrated by growth suppression in imatinib-resistant BCR-ABL-positive leukemic cells and in mice engrafted with BaF3/BCR-ABL, human BCR-ABL(+) cell lines, or primary patient samples expressing BCR-ABL/T315I or glutamic acid 255 to lysine imatinib-resistant mutation. ('BCR-ABL', 'Gene', '25', (183, 190)) ('AT9283', 'Chemical', 'MESH:C535237', (36, 42)) ('human', 'Species', '9606', (177, 182)) ('glutamic acid 255 to lysine', 'Var', (261, 288)) ('suppression', 'NegReg', (70, 81)) ('activity', 'MPA', (24, 32)) ('BCR-ABL', 'Gene', (168, 175)) ('BCR-ABL', 'Gene', (104, 111)) ('BCR-ABL', 'Gene', '25', (244, 251)) ('BCR-ABL', 'Gene', (183, 190)) ('imatinib', 'Chemical', 'MESH:D000068877', (85, 93)) ('imatinib', 'Chemical', 'MESH:D000068877', (289, 297)) ('T315I', 'Mutation', 'rs121913459', (252, 257)) ('AT9283', 'Var', (36, 42)) ('mice', 'Species', '10090', (143, 147)) ('patient', 'Species', '9606', (217, 224)) ('leukemic', 'Disease', 'MESH:D007938', (121, 129)) ('BCR-ABL', 'Gene', '25', (168, 175)) ('growth', 'MPA', (63, 69)) ('BCR-ABL', 'Gene', '25', (104, 111)) ('BCR-ABL', 'Gene', (244, 251)) ('leukemic', 'Disease', (121, 129)) ('glutamic acid 255 to lysine', 'SUBSTITUTION', 'None', (261, 288)) 4384 29097609 Preclinical reports also indicate that AT9283 synergizes with dasatinib and enhances the repression of medulloblastoma survival and migration. ('medulloblastoma', 'Disease', (103, 118)) ('AT9283', 'Chemical', 'MESH:C535237', (39, 45)) ('dasatinib', 'Chemical', 'MESH:D000069439', (62, 71)) ('medulloblastoma', 'Disease', 'MESH:D008527', (103, 118)) ('medulloblastoma', 'Phenotype', 'HP:0002885', (103, 118)) ('AT9283', 'Var', (39, 45)) ('repression', 'MPA', (89, 99)) ('enhances', 'PosReg', (76, 84)) 4385 29097609 The first phase I/II trial with AT9283 in thirty-three pediatric patients with solid tumors (NCT00985868) led to a partial response in one patient diagnosed with central nervous system (CNS)-primitive neuroectodermal tumour (PNET) and disease stabilization in 38% of patients, with manageable hematological toxicity. ('patient', 'Species', '9606', (267, 274)) ('tumour', 'Phenotype', 'HP:0002664', (217, 223)) ('patients', 'Species', '9606', (267, 275)) ('hematological toxicity', 'Disease', 'MESH:D006402', (293, 315)) ('tumor', 'Phenotype', 'HP:0002664', (85, 90)) ('hematological toxicity', 'Disease', (293, 315)) ('primitive neuroectodermal tumour', 'Phenotype', 'HP:0030065', (191, 223)) ('solid tumors', 'Disease', (79, 91)) ('neuroectodermal tumour', 'Disease', 'MESH:D017599', (201, 223)) ('AT9283', 'Chemical', 'MESH:C535237', (32, 38)) ('patient', 'Species', '9606', (65, 72)) ('patients', 'Species', '9606', (65, 73)) ('solid tumors', 'Disease', 'MESH:D009369', (79, 91)) ('neuroectodermal tumour', 'Disease', (201, 223)) ('AT9283', 'Var', (32, 38)) ('neuroectodermal tumour', 'Phenotype', 'HP:0030061', (201, 223)) ('patient', 'Species', '9606', (139, 146)) ('tumors', 'Phenotype', 'HP:0002664', (85, 91)) 4390 29097609 It is likely that the evaluation of this agent in a cohort of pediatric patients with leukemia harboring the JAK, ABL, or FLT3 mutations might increase the potential of clinical responses. ('ABL', 'Gene', (114, 117)) ('FLT3', 'Gene', '2322', (122, 126)) ('leukemia', 'Disease', (86, 94)) ('leukemia', 'Phenotype', 'HP:0001909', (86, 94)) ('leukemia', 'Disease', 'MESH:D007938', (86, 94)) ('clinical responses', 'CPA', (169, 187)) ('mutations', 'Var', (127, 136)) ('increase', 'PosReg', (143, 151)) ('FLT3', 'Gene', (122, 126)) ('patients', 'Species', '9606', (72, 80)) 4391 29097609 Alisertib (MLN8237) inhibits Aurora-A and Aurora-B kinase with an IC50 of 1.2 nM and 396.5 nM, respectively. ('Aurora-B', 'Gene', '9212', (42, 50)) ('inhibits', 'NegReg', (20, 28)) ('Alisertib', 'Chemical', 'MESH:C550258', (0, 9)) ('MLN8237', 'Chemical', 'MESH:C550258', (11, 18)) ('MLN8237', 'Var', (11, 18)) ('Aurora-B', 'Gene', (42, 50)) ('Aurora-A', 'Gene', (29, 37)) ('Aurora-A', 'Gene', '6790', (29, 37)) 4393 29097609 In the PPTP in vitro and in vivo models of childhood cancer, MLN8237 treatment led to growth repression in neuroblastoma and Ewing sarcoma cell lines as well as maintained complete responses and complete responses in neuroblastoma and ALL xenografts, respectively. ('neuroblastoma', 'Disease', (107, 120)) ('Ewing sarcoma', 'Phenotype', 'HP:0012254', (125, 138)) ('Ewing sarcoma', 'Disease', 'MESH:C563168', (125, 138)) ('cancer', 'Disease', (53, 59)) ('cancer', 'Disease', 'MESH:D009369', (53, 59)) ('MLN8237', 'Chemical', 'MESH:C550258', (61, 68)) ('neuroblastoma', 'Phenotype', 'HP:0003006', (107, 120)) ('neuroblastoma', 'Phenotype', 'HP:0003006', (217, 230)) ('PPTP', 'Chemical', '-', (7, 11)) ('growth repression', 'MPA', (86, 103)) ('cancer', 'Phenotype', 'HP:0002664', (53, 59)) ('sarcoma', 'Phenotype', 'HP:0100242', (131, 138)) ('neuroblastoma', 'Disease', 'MESH:D009447', (217, 230)) ('Ewing sarcoma', 'Disease', (125, 138)) ('neuroblastoma', 'Disease', 'MESH:D009447', (107, 120)) ('neuroblastoma', 'Disease', (217, 230)) ('MLN8237 treatment', 'Var', (61, 78)) 4394 29097609 Preclinical studies established a correlation between alisertib sensitivity and decreased AURKA copy number, in addition to a steep dose-response relationship (1muM). ('AURKA', 'Gene', '6790', (90, 95)) ('alisertib', 'Chemical', 'MESH:C550258', (54, 63)) ('AURKA', 'Gene', (90, 95)) ('decreased', 'NegReg', (80, 89)) ('alisertib sensitivity', 'Var', (54, 75)) 4405 29097609 In comparison to normal cells that repair DNA damage during G1 arrest, deficiencies in the G1 checkpoint in cancer cells result in DNA repair at the G2/M checkpoint. ('deficiencies', 'Var', (71, 83)) ('arrest', 'Disease', 'MESH:D006323', (63, 69)) ('arrest', 'Disease', (63, 69)) ('cancer', 'Phenotype', 'HP:0002664', (108, 114)) ('G1 checkpoint', 'Gene', (91, 104)) ('DNA repair', 'MPA', (131, 141)) ('cancer', 'Disease', (108, 114)) ('cancer', 'Disease', 'MESH:D009369', (108, 114)) 4406 29097609 Ataxia-telangiectasia mutated (ATM) kinase and/or ataxia-telangiectasia-related (ATR) kinase mediate the repair of DNA double-strand breaks and DNA single-strand breaks, respectively. ('ataxia-telangiectasia', 'Disease', 'MESH:D001260', (50, 71)) ('ATR', 'Gene', (81, 84)) ('ATR', 'Gene', '545', (81, 84)) ('Ataxia-telangiectasia', 'Disease', 'MESH:D001260', (0, 21)) ('ATM', 'Gene', '472', (31, 34)) ('DNA', 'Var', (115, 118)) ('telangiectasia', 'Phenotype', 'HP:0001009', (57, 71)) ('ataxia-telangiectasia', 'Disease', (50, 71)) ('telangiectasia', 'Phenotype', 'HP:0001009', (7, 21)) ('Ataxia', 'Phenotype', 'HP:0001251', (0, 6)) ('ATM', 'Gene', (31, 34)) ('Ataxia-telangiectasia', 'Disease', (0, 21)) ('ataxia', 'Phenotype', 'HP:0001251', (50, 56)) 4408 29097609 Phosphorylation of tyrosine 15 on CDK1 by Wee1 inhibits CDK1/cyclin B function, resulting in cell cycle arrest at G2, entry into mitosis and DNA-damage repair. ('arrest', 'Disease', 'MESH:D006323', (104, 110)) ('cyclin', 'Gene', '5111', (61, 67)) ('Wee1', 'Gene', (42, 46)) ('Phosphorylation', 'Var', (0, 15)) ('cyclin', 'Gene', (61, 67)) ('DNA-damage repair', 'CPA', (141, 158)) ('mitosis', 'Disease', (129, 136)) ('arrest', 'Disease', (104, 110)) ('Wee1', 'Gene', '7465', (42, 46)) ('CDK1', 'Gene', (34, 38)) ('function', 'MPA', (70, 78)) ('mitosis', 'Disease', 'None', (129, 136)) ('CDK1', 'Gene', '983', (34, 38)) ('tyrosine', 'Chemical', 'MESH:D014443', (19, 27)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (93, 110)) ('CDK1', 'Gene', '983', (56, 60)) ('CDK1', 'Gene', (56, 60)) ('inhibits', 'NegReg', (47, 55)) 4410 29097609 Since Wee1 functionally interacts with critical regulators of mitosis, inhibition interrupts the DNA repair machinery leading to mitotic catastrophe and apoptotic cell death. ('Wee1', 'Gene', (6, 10)) ('apoptotic cell death', 'CPA', (153, 173)) ('inhibition', 'Var', (71, 81)) ('mitotic catastrophe', 'CPA', (129, 148)) ('Wee1', 'Gene', '7465', (6, 10)) ('interrupts', 'NegReg', (82, 92)) ('DNA', 'Protein', (97, 100)) ('interacts', 'Reg', (24, 33)) ('mitosis', 'Disease', (62, 69)) ('mitosis', 'Disease', 'None', (62, 69)) 4412 29097609 Inhibition of Wee1 could sensitize cancer cells to radiation therapy and chemotherapy, by disruption of the G2/M checkpoint. ('Wee1', 'Gene', (14, 18)) ('disruption', 'NegReg', (90, 100)) ('Wee1', 'Gene', '7465', (14, 18)) ('cancer', 'Phenotype', 'HP:0002664', (35, 41)) ('Inhibition', 'Var', (0, 10)) ('cancer', 'Disease', 'MESH:D009369', (35, 41)) ('sensitize', 'Reg', (25, 34)) ('cancer', 'Disease', (35, 41)) ('G2/M checkpoint', 'CPA', (108, 123)) 4413 29097609 AZD1775 is the most frequently tested Wee1 inhibitor in preclinical and clinical studies. ('Wee1', 'Gene', '7465', (38, 42)) ('Wee1', 'Gene', (38, 42)) ('AZD1775', 'Var', (0, 7)) ('AZD1775', 'Chemical', 'MESH:C549567', (0, 7)) 4414 29097609 Combining gamma radiation and AZD1775 in HGG cell lines and orthotopic HGG and DIPG xenograft tumors led to enhanced sensitivity than achieved by radiation treatment alone. ('sensitivity', 'MPA', (117, 128)) ('tumor', 'Phenotype', 'HP:0002664', (94, 99)) ('DIPG xenograft tumors', 'Disease', (79, 100)) ('enhanced', 'PosReg', (108, 116)) ('tumors', 'Phenotype', 'HP:0002664', (94, 100)) ('AZD1775', 'Var', (30, 37)) ('DIPG xenograft tumors', 'Disease', 'MESH:D000080443', (79, 100)) ('AZD1775', 'Chemical', 'MESH:C549567', (30, 37)) 4416 29097609 In addition, AZD1775 caused activation of cell division cycle protein 2 (CDC2; also known as CDK1) in an osteosarcoma xenograft and reduced tumor growth by 50% as monotherapy, and by approximately 70% when combined with gemcitabine. ('CDK1', 'Gene', '983', (93, 97)) ('AZD1775', 'Chemical', 'MESH:C549567', (13, 20)) ('gemcitabine', 'Chemical', 'MESH:C056507', (220, 231)) ('CDC2', 'Gene', (73, 77)) ('sarcoma', 'Phenotype', 'HP:0100242', (110, 117)) ('reduced', 'NegReg', (132, 139)) ('osteosarcoma xenograft', 'Disease', 'MESH:D012516', (105, 127)) ('CDC2', 'Gene', '983', (73, 77)) ('tumor', 'Disease', 'MESH:D009369', (140, 145)) ('activation', 'PosReg', (28, 38)) ('osteosarcoma', 'Phenotype', 'HP:0002669', (105, 117)) ('tumor', 'Phenotype', 'HP:0002664', (140, 145)) ('CDK1', 'Gene', (93, 97)) ('osteosarcoma xenograft', 'Disease', (105, 127)) ('AZD1775', 'Var', (13, 20)) ('tumor', 'Disease', (140, 145)) 4417 29097609 In hematological malignancies, AZD1775 plus panobinostat demonstrated synergistic antitumor effects in preclinical models of AML and when combined with CDK inhibitor (roscovitine), and cytarabine. ('AML', 'Disease', 'MESH:D015470', (125, 128)) ('tumor', 'Phenotype', 'HP:0002664', (86, 91)) ('AZD1775', 'Var', (31, 38)) ('cytarabine', 'Chemical', 'MESH:D003561', (185, 195)) ('tumor', 'Disease', (86, 91)) ('AML', 'Disease', (125, 128)) ('hematological malignancies', 'Phenotype', 'HP:0004377', (3, 29)) ('panobinostat', 'Gene', (44, 56)) ('hematological malignancies', 'Disease', (3, 29)) ('AZD1775', 'Chemical', 'MESH:C549567', (31, 38)) ('hematological malignancies', 'Disease', 'MESH:D019337', (3, 29)) ('panobinostat', 'Chemical', 'MESH:D000077767', (44, 56)) ('tumor', 'Disease', 'MESH:D009369', (86, 91)) ('roscovitine', 'Chemical', 'MESH:D000077546', (167, 178)) 4418 29097609 In the latter study, AZD1775 inhibited cytabarine-mediated S-phase arrest and prevented DNA repair. ('prevented', 'NegReg', (78, 87)) ('inhibited', 'NegReg', (29, 38)) ('AZD1775', 'Var', (21, 28)) ('AZD1775', 'Chemical', 'MESH:C549567', (21, 28)) ('arrest', 'Disease', 'MESH:D006323', (67, 73)) ('cytabarine', 'Chemical', '-', (39, 49)) ('arrest', 'Disease', (67, 73)) ('DNA repair', 'CPA', (88, 98)) ('cytabarine-mediated', 'MPA', (39, 58)) 4419 29097609 Further, MYC-driven tumors or CDKN2A mutation combined with TP53 mutation could show aberrations in the G1 checkpoint and become more reliant on the S- and G2- phase checkpoints. ('tumors', 'Disease', (20, 26)) ('MYC', 'Gene', '4609', (9, 12)) ('tumors', 'Disease', 'MESH:D009369', (20, 26)) ('tumors', 'Phenotype', 'HP:0002664', (20, 26)) ('mutation', 'Var', (37, 45)) ('CDKN2A', 'Gene', (30, 36)) ('MYC', 'Gene', (9, 12)) ('CDKN2A', 'Gene', '1029', (30, 36)) ('G1 checkpoint', 'CPA', (104, 117)) ('TP53', 'Gene', '7157', (60, 64)) ('mutation', 'Var', (65, 73)) ('tumor', 'Phenotype', 'HP:0002664', (20, 25)) ('TP53', 'Gene', (60, 64)) 4420 29097609 S- and G2- checkpoint abrogation caused by inhibition of Wee1 may selectively sensitize p53-deficient tumors. ('deficient tumors', 'Disease', 'MESH:D009369', (92, 108)) ('inhibition', 'Var', (43, 53)) ('tumors', 'Phenotype', 'HP:0002664', (102, 108)) ('G2- checkpoint abrogation', 'CPA', (7, 32)) ('p53', 'Gene', (88, 91)) ('p53', 'Gene', '7157', (88, 91)) ('Wee1', 'Gene', (57, 61)) ('tumor', 'Phenotype', 'HP:0002664', (102, 107)) ('Wee1', 'Gene', '7465', (57, 61)) ('deficient tumors', 'Disease', (92, 108)) 4421 29097609 Other studies also show that AZD1775 sensitizes AML cells to cytarabine and HGG cells to radiation irrespective of p53 mutation status. ('cytarabine', 'Chemical', 'MESH:D003561', (61, 71)) ('AML', 'Disease', (48, 51)) ('sensitizes', 'Reg', (37, 47)) ('p53', 'Gene', (115, 118)) ('p53', 'Gene', '7157', (115, 118)) ('AZD1775', 'Var', (29, 36)) ('AML', 'Disease', 'MESH:D015470', (48, 51)) ('AZD1775', 'Chemical', 'MESH:C549567', (29, 36)) 4422 29097609 These observations suggest that the requirement of p53 mutations for sensitivity with combination therapies using AZD1775 is context-dependent and may not be a critical consideration during development of novel therapies. ('p53', 'Gene', (51, 54)) ('p53', 'Gene', '7157', (51, 54)) ('mutations', 'Var', (55, 64)) ('AZD1775', 'Gene', (114, 121)) ('AZD1775', 'Chemical', 'MESH:C549567', (114, 121)) 4424 29097609 Patient recruitment for two pediatric phase 1 trials to study side effects and best dose of Wee1 kinase inhibitor AZD1775 when given together with local radiation therapy in treating newly diagnosed DIPG (NCT01922076); and with irinotecan hydrochloride in treating advanced solid tumors (NCT02095132) is ongoing. ('DIPG', 'Disease', (199, 203)) ('DIPG', 'Chemical', '-', (199, 203)) ('AZD1775', 'Gene', (114, 121)) ('irinotecan hydrochloride', 'Chemical', 'MESH:D000077146', (228, 252)) ('solid tumors', 'Disease', 'MESH:D009369', (274, 286)) ('tumor', 'Phenotype', 'HP:0002664', (280, 285)) ('Wee1', 'Gene', (92, 96)) ('AZD1775', 'Chemical', 'MESH:C549567', (114, 121)) ('NCT01922076);', 'Var', (205, 218)) ('tumors', 'Phenotype', 'HP:0002664', (280, 286)) ('Wee1', 'Gene', '7465', (92, 96)) ('solid tumors', 'Disease', (274, 286)) ('Patient', 'Species', '9606', (0, 7)) 4427 29097609 Inhibition of KSP leads to cell cycle arrest in mitosis with the formation of characteristic monoaster spindles. ('cell cycle arrest', 'Phenotype', 'HP:0011018', (27, 44)) ('KSP', 'Chemical', '-', (14, 17)) ('arrest in mitosis', 'Disease', (38, 55)) ('Inhibition', 'Var', (0, 10)) ('KSP', 'Gene', (14, 17)) ('arrest in mitosis', 'Disease', 'MESH:D006323', (38, 55)) 4441 29097609 Stabilizing factors include microtubule-associated proteins, MAPs (MAP4, XMAP215, XMAP230/XMAP4 and XMAP310) and destabilizing factors include (Stathmin1 and XKCM1). ('MAP4', 'Gene', '4134', (67, 71)) ('MAP4', 'Gene', (91, 95)) ('MAP4', 'Gene', '4134', (91, 95)) ('microtubule-associated proteins', 'Protein', (28, 59)) ('XMAP310', 'Var', (100, 107)) ('Stathmin1', 'Gene', '3925', (144, 153)) ('Stathmin1', 'Gene', (144, 153)) ('MAP4', 'Gene', (67, 71)) 4442 29097609 Consistent with the disruption of microtubules and the mitotic spindle, MTAs can arrest cell cycle progression in mitosis, resulting in cell death. ('mitosis', 'Disease', (114, 121)) ('mitosis', 'Disease', 'None', (114, 121)) ('MTAs', 'Var', (72, 76)) ('arrest', 'Disease', 'MESH:D006323', (81, 87)) ('arrest', 'Disease', (81, 87)) 4470 29097609 Ongoing trials for the same patient population are evaluating effects of combining brentuximab vedotin with doxorubicin (NCT01920932), dacarbazine (NCT02979522), and gemcitabine (NCT01780662). ('dacarbazine', 'Chemical', 'MESH:D003606', (135, 146)) ('patient', 'Species', '9606', (28, 35)) ('gemcitabine', 'Chemical', 'MESH:C056507', (166, 177)) ('NCT02979522', 'Var', (148, 159)) ('doxorubicin', 'Chemical', 'MESH:D004317', (108, 119)) ('NCT01920932', 'Var', (121, 132)) 4480 29097609 For example, investigations of antitumor activity for inhibitors against ATM (KU-55933, KU-59403; KuDOS Pharmaceuticals, AstraZeneca) and ATR (Schisandrin B, AZD6738) have advanced to early phase trials in adult patients with cancer. ('ATM', 'Gene', (73, 76)) ('tumor', 'Disease', 'MESH:D009369', (35, 40)) ('ATM', 'Gene', '472', (73, 76)) ('tumor', 'Phenotype', 'HP:0002664', (35, 40)) ('KU-55933', 'Var', (78, 86)) ('patients', 'Species', '9606', (212, 220)) ('tumor', 'Disease', (35, 40)) ('cancer', 'Disease', (226, 232)) ('cancer', 'Disease', 'MESH:D009369', (226, 232)) ('Schisandrin B', 'Chemical', 'MESH:C015499', (143, 156)) ('ATR', 'Gene', '545', (138, 141)) ('ATR', 'Gene', (138, 141)) ('AZD6738', 'Chemical', 'MESH:C000611951', (158, 165)) ('KU-59403', 'Var', (88, 96)) ('cancer', 'Phenotype', 'HP:0002664', (226, 232)) 4489 29097609 This was seen when MLN8237 (alisertib) failed to reach its primary endpoint for adult lymphomas and lung cancers during early phase studies in pediatric patients. ('lung cancers', 'Disease', (100, 112)) ('MLN8237', 'Chemical', 'MESH:C550258', (19, 26)) ('cancers', 'Phenotype', 'HP:0002664', (105, 112)) ('patients', 'Species', '9606', (153, 161)) ('lymphomas', 'Disease', (86, 95)) ('lymphomas', 'Disease', 'MESH:D008223', (86, 95)) ('MLN8237', 'Var', (19, 26)) ('lung cancer', 'Phenotype', 'HP:0100526', (100, 111)) ('lymphomas', 'Phenotype', 'HP:0002665', (86, 95)) ('lung cancers', 'Disease', 'MESH:D008175', (100, 112)) ('lung cancers', 'Phenotype', 'HP:0100526', (100, 112)) ('cancer', 'Phenotype', 'HP:0002664', (105, 111)) ('alisertib', 'Chemical', 'MESH:C550258', (28, 37)) ('lymphoma', 'Phenotype', 'HP:0002665', (86, 94)) 4505 28107369 On the other hand, Natural Killer cells (NK) are lymphocytes that have the potential to recognize and lyse cells with aberrant HLA-expression profiles, thereby mediating a GvL-effect of their own. ('mediating', 'Reg', (160, 169)) ('HLA', 'Gene', (127, 130)) ('HLA', 'Gene', '3123', (127, 130)) ('aberrant', 'Var', (118, 126)) 4513 28107369 However, HLA-C/KIR interactions appear to be even more complex, given that HLA-C antigens are divided into two ligand groups based on specific amino acid residues: Antigens in the C1 group bear an asparagine residue on position 80 and interact with inhibitory KIR2DL2 and KIR2DL3 as well as activating KIR2DS2, whereas C2 group ligands carry a lysine residue at position 80 and interact with inhibitory KIR2DL1 and activating KIR2DS1(. ('KIR2DL2', 'Gene', '3803', (260, 267)) ('KIR2DL2', 'Gene', (260, 267)) ('HLA-C', 'Gene', (75, 80)) ('KIR', 'Gene', '3811', (272, 275)) ('HLA-C', 'Gene', '3107', (75, 80)) ('HLA-C', 'Gene', (9, 14)) ('KIR', 'Gene', (272, 275)) ('HLA-C', 'Gene', '3107', (9, 14)) ('KIR', 'Gene', '3811', (302, 305)) ('KIR2DL1', 'Gene', (403, 410)) ('KIR', 'Gene', '3811', (403, 406)) ('KIR', 'Gene', '3811', (15, 18)) ('asparagine', 'Chemical', 'MESH:D001216', (197, 207)) ('KIR', 'Gene', '3811', (260, 263)) ('interact', 'Interaction', (235, 243)) ('KIR', 'Gene', (302, 305)) ('KIR', 'Gene', '3811', (426, 429)) ('KIR', 'Gene', (403, 406)) ('interact', 'Interaction', (378, 386)) ('activating', 'PosReg', (291, 301)) ('KIR', 'Gene', (15, 18)) ('KIR2DS1', 'Gene', (426, 433)) ('KIR2DL3', 'Gene', (272, 279)) ('lysine', 'Chemical', 'MESH:D008239', (344, 350)) ('KIR', 'Gene', (260, 263)) ('KIR2DS2', 'Gene', (302, 309)) ('KIR2DL3', 'Gene', '3804', (272, 279)) ('lysine', 'Var', (344, 350)) ('KIR', 'Gene', (426, 429)) ('KIR2DS1', 'Gene', '3806', (426, 433)) ('KIR2DS2', 'Gene', '100132285', (302, 309)) ('KIR2DL1', 'Gene', '3802', (403, 410)) 4549 28107369 Accordingly, HLA-C antigens with a lysine residue at this position possess the C2 epitope, which acts as ligand for the inhibitory KIR2DL1 as well as for the activating KIR2DS1. ('lysine', 'Chemical', 'MESH:D008239', (35, 41)) ('KIR2DS1', 'Gene', '3806', (169, 176)) ('HLA-C', 'Gene', (13, 18)) ('KIR2DL1', 'Gene', (131, 138)) ('C2 epitope', 'Disease', 'OMIM:217000', (79, 89)) ('C2 epitope', 'Disease', (79, 89)) ('lysine residue', 'Var', (35, 49)) ('KIR2DS1', 'Gene', (169, 176)) ('HLA-C', 'Gene', '3107', (13, 18)) ('KIR2DL1', 'Gene', '3802', (131, 138)) 4578 28107369 The protective effect of C1 could be observed only in the myeloid group (Table 5), where absence of C1 was associated with reduced OS (myeloid: HR = 1.51, CI = 1.15-1.99, p = 0.003; lymphoblastic: HR = 1.26, CI = 0.91-1.75, p = 0.16), reduced DFS (myeloid: HR = 1.31, CI = 1.01-1.70, p = 0.04; lymphoblastic: HR = 1.21, CI = 0.90-1.61, p = 0.21), and increased RI (myeloid: HR = 1.31, CI = 1.01-1.70, p = 0.04; lymphoblastic: HR = 1.21, CI = 0.90-1.61, p = 0.21). ('increased', 'PosReg', (351, 360)) ('DFS', 'MPA', (243, 246)) ('OS', 'Chemical', '-', (131, 133)) ('absence', 'Var', (89, 96)) ('reduced', 'NegReg', (235, 242)) ('reduced', 'NegReg', (123, 130)) 4585 28107369 Similarly, donor KIR2DS2 positivity resulted in higher DFS rates in the 9/10 matched patient group (10/10: 40.6% vs. 37.9%; 29.5% vs. 28.4%; 9/10: 51.6% vs. 21.1%; 24.6% vs. 15.8%; Fig 11 and Fig 12). ('patient', 'Species', '9606', (85, 92)) ('KIR2DS2', 'Gene', '100132285', (17, 24)) ('DFS rates', 'MPA', (55, 64)) ('positivity', 'Var', (25, 35)) ('KIR2DS2', 'Gene', (17, 24)) ('higher', 'PosReg', (48, 54)) ('donor', 'Species', '9606', (11, 16)) 4593 28107369 In the presence of a single HLA class I mismatch, TRM was increased in C1 negative patients who were transplanted with a KIR2DS1 (HR = 2.61, CI = 1.26-5.41, p = 0.001; Fig 17) positive donor. ('increased', 'PosReg', (58, 67)) ('KIR2DS1', 'Gene', '3806', (121, 128)) ('patients', 'Species', '9606', (83, 91)) ('KIR2DS1', 'Gene', (121, 128)) ('HLA', 'Gene', '3123', (28, 31)) ('TRM', 'MPA', (50, 53)) ('donor', 'Species', '9606', (185, 190)) ('mismatch', 'Var', (40, 48)) ('HLA', 'Gene', (28, 31)) 4597 28107369 It has been shown by Cooley et al, that C1-positive, but not C1-negative patients benefit from transplantation with donors who carry two or more B-motifs. ('transplantation', 'CPA', (95, 110)) ('donor', 'Species', '9606', (116, 121)) ('C1-positive', 'Var', (40, 51)) ('patients', 'Species', '9606', (73, 81)) ('benefit', 'PosReg', (82, 89)) 4606 28107369 Deficiency for the C1-ligand had significantly negative effects on all analysed survival endpoints (OS, DFS, RI, and TRM). ('negative', 'NegReg', (47, 55)) ('C1-ligand', 'Protein', (19, 28)) ('OS', 'Chemical', '-', (100, 102)) ('DFS', 'Disease', (104, 107)) ('Deficiency', 'Var', (0, 10)) 4624 28107369 Furthermore, again in the presence of a single HLA-class I mismatch, RI was significantly reduced in risk group patients who had received a graft from a KIR2DS1 -positive donor when compared to risk group patients who were transplanted with KIR2DS1-negative donor cells. ('patients', 'Species', '9606', (205, 213)) ('reduced', 'NegReg', (90, 97)) ('KIR2DS1', 'Gene', '3806', (153, 160)) ('KIR2DS1', 'Gene', (241, 248)) ('mismatch', 'Var', (59, 67)) ('KIR2DS1', 'Gene', (153, 160)) ('patients', 'Species', '9606', (112, 120)) ('HLA', 'Gene', '3123', (47, 50)) ('donor', 'Species', '9606', (171, 176)) ('HLA', 'Gene', (47, 50)) ('donor', 'Species', '9606', (258, 263)) ('KIR2DS1', 'Gene', '3806', (241, 248)) 4632 28107369 Taking advantage of a large multicentre study cohort of 1446 transplant pairs we were able to identify the 9/10 HLA-class I mismatched transplants as the decisive patient subgroup. ('HLA', 'Gene', (112, 115)) ('patient', 'Species', '9606', (163, 170)) ('HLA', 'Gene', '3123', (112, 115)) ('mismatched', 'Var', (124, 134)) 4649 28107369 The most important finding of our study, however, is the association of donor KIR2DS2 positivity with a better overall outcome after HSCT in the C1-negative risk patient group. ('donor', 'Species', '9606', (72, 77)) ('KIR2DS2', 'Gene', '100132285', (78, 85)) ('KIR2DS2', 'Gene', (78, 85)) ('positivity', 'Var', (86, 96)) ('better', 'PosReg', (104, 110)) ('patient', 'Species', '9606', (162, 169)) 4653 27248327 Expression and functional characterization of CD33 transcript variants in human acute myeloid leukemia With the demonstration of improved survival of some acute myeloid leukemia (AML) patients with the CD33 antibody-drug conjugate, gemtuzumab ozogamicin (GO), CD33 has been validated as a target for antigen-specific immunotherapy. ('AML', 'Disease', (179, 182)) ('acute myeloid leukemia', 'Disease', (80, 102)) ('rat', 'Species', '10116', (119, 122)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (86, 102)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (86, 102)) ('GO', 'Chemical', 'MESH:C406061', (255, 257)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (80, 102)) ('CD33', 'Gene', '945', (46, 50)) ('CD33', 'Gene', (46, 50)) ('leukemia', 'Phenotype', 'HP:0001909', (94, 102)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (80, 102)) ('acute myeloid leukemia', 'Disease', (155, 177)) ('CD33', 'Gene', '945', (260, 264)) ('CD33', 'Gene', '945', (202, 206)) ('gemtuzumab ozogamicin', 'Chemical', 'MESH:C406061', (232, 253)) ('CD33', 'Gene', (202, 206)) ('CD33', 'Gene', (260, 264)) ('patients', 'Species', '9606', (184, 192)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (155, 177)) ('variants', 'Var', (62, 70)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (161, 177)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (161, 177)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (155, 177)) ('AML', 'Disease', 'MESH:D015470', (179, 182)) ('leukemia', 'Phenotype', 'HP:0001909', (169, 177)) ('human', 'Species', '9606', (74, 79)) ('improved', 'PosReg', (129, 137)) ('AML', 'Phenotype', 'HP:0004808', (179, 182)) 4655 27248327 In primary AML specimens, we not only found full-length CD33 (CD33FL) and CD33 E2 but also corresponding variants containing an alternate exon 7 predicted to encode a CD33 protein lacking most of the intracellular domain (CD33E7a and, not previously described, CD33 E2,E7a) in almost all cases. ('CD33E7a', 'Chemical', '-', (222, 229)) ('CD33', 'Gene', '945', (167, 171)) ('variants', 'Var', (105, 113)) ('lacking', 'NegReg', (180, 187)) ('CD33', 'Gene', (167, 171)) ('CD33', 'Gene', '945', (56, 60)) ('CD33', 'Gene', (56, 60)) ('CD33', 'Gene', '945', (222, 226)) ('CD33', 'Gene', (222, 226)) ('CD33FL', 'Chemical', '-', (62, 68)) ('CD33', 'Gene', '945', (62, 66)) ('CD33', 'Gene', (62, 66)) ('CD33', 'Gene', '945', (261, 265)) ('CD33', 'Gene', (261, 265)) ('CD33', 'Gene', '945', (74, 78)) ('CD33', 'Gene', (74, 78)) ('AML', 'Disease', 'MESH:D015470', (11, 14)) ('AML', 'Phenotype', 'HP:0004808', (11, 14)) ('AML', 'Disease', (11, 14)) 4660 27248327 They identify CD33 variants that lack exon 2 and are not recognized by current CD33-directed therapeutics as potential target for future unconjugated or conjugated antibodies. ('CD33', 'Gene', '945', (14, 18)) ('CD33', 'Gene', '945', (79, 83)) ('CD33', 'Gene', (14, 18)) ('CD33', 'Gene', (79, 83)) ('variants', 'Var', (19, 27)) 4665 27248327 For CD33, the precise physiological functions have remained unclear, but recently, genome-wide association studies have identified variants of CD33 as a major risk factor for Alzheimer's disease, suggesting an important role in neurodegeneration. ("Alzheimer's disease", 'Disease', 'MESH:D000544', (175, 194)) ('risk factor', 'Reg', (159, 170)) ("Alzheimer's disease", 'Phenotype', 'HP:0002511', (175, 194)) ("Alzheimer's disease", 'Disease', (175, 194)) ('neurodegeneration', 'Phenotype', 'HP:0002180', (228, 245)) ('variants', 'Var', (131, 139)) ('neurodegeneration', 'Disease', (228, 245)) ('neurodegeneration', 'Disease', 'MESH:D019636', (228, 245)) ('CD33', 'Gene', '945', (143, 147)) ('CD33', 'Gene', (143, 147)) ('CD33', 'Gene', '945', (4, 8)) ('CD33', 'Gene', (4, 8)) 4673 27248327 UCSC genome browser and Ensembl databases contain 3 unique, validated CD33 transcript variants supported by at least one non-suspect mRNA sequence, NM_001772, NM_001082618 and NM_001177608. ('NM_001177608', 'Var', (176, 188)) ('NM_001082618', 'Var', (159, 171)) ('CD33', 'Gene', '945', (70, 74)) ('CD33', 'Gene', (70, 74)) ('NM_001772', 'Var', (148, 157)) 4674 27248327 NM_001772 represents full-length CD33 (CD33FL) consisting of 7 coding exons and a 364 amino acid protein. ('NM_001772', 'Var', (0, 9)) ('CD33', 'Gene', '945', (33, 37)) ('CD33FL', 'Chemical', '-', (39, 45)) ('CD33', 'Gene', '945', (39, 43)) ('CD33', 'Gene', (33, 37)) ('CD33', 'Gene', (39, 43)) 4675 27248327 NM_001082618 skips exon 2 resulting in CD33 with a deleted exon 2 (CD33 E2) and a 237 amino acid protein. ('NM_001082618', 'Var', (0, 12)) ('CD33', 'Gene', '945', (67, 71)) ('CD33', 'Gene', '945', (39, 43)) ('CD33', 'Gene', (67, 71)) ('CD33', 'Gene', (39, 43)) 4676 27248327 NM_001177608 uses an alternate exon 7, exon 7a (CD33E7a), which is predicted to encode an early translational stop, resulting in a 310 amino acid protein with truncation of the 54 c-terminal amino acids of the intracellular domain relative to CD33FL. ('NM_001177608', 'Var', (0, 12)) ('CD33E7a', 'Chemical', '-', (48, 55)) ('CD33FL', 'Chemical', '-', (243, 249)) ('truncation', 'MPA', (159, 169)) 4677 27248327 The existence of CD33 variants lacking exon 2 is relevant for therapeutic targeting of CD33 since the V-set domain, which is encoded by exon 2, contains immune-dominant epitope(s) recognized by all currently clinically exploited CD33-directed therapeutics. ('CD33', 'Gene', '945', (87, 91)) ('CD33', 'Gene', (87, 91)) ('CD33', 'Gene', '945', (229, 233)) ('CD33', 'Gene', (229, 233)) ('CD33', 'Gene', '945', (17, 21)) ('variants', 'Var', (22, 30)) ('CD33', 'Gene', (17, 21)) 4678 27248327 Moreover, our previous studies in cell lines engineered to express mutant forms of CD33 demonstrated that CD33 endocytosis is largely limited and determined by the intracellular domain of CD33 and that changes in this domain significantly impact CD33 internalization. ('CD33', 'Gene', '945', (83, 87)) ('CD33', 'Gene', (83, 87)) ('rat', 'Species', '10116', (95, 98)) ('internalization', 'MPA', (251, 266)) ('impact', 'Reg', (239, 245)) ('changes', 'Var', (202, 209)) ('CD33', 'Gene', '945', (188, 192)) ('mutant', 'Var', (67, 73)) ('CD33', 'Gene', '945', (246, 250)) ('CD33', 'Gene', (246, 250)) ('CD33', 'Gene', (188, 192)) ('limited', 'NegReg', (134, 141)) ('CD33', 'Gene', (106, 110)) ('CD33', 'Gene', '945', (106, 110)) 4679 27248327 It is therefore conceivable that transcript variants that lack most of the intracellular domain of CD33 differ in their endocytic properties from the wild-type protein. ('CD33', 'Gene', (99, 103)) ('variants', 'Var', (44, 52)) ('endocytic properties', 'MPA', (120, 140)) ('CD33', 'Gene', '945', (99, 103)) 4682 27248327 CD33 coding sequence splice junctions were filtered for junctions with 10% prevalence at a threshold of >= 10 reads per specimen, resulting in 8 observed splice junctions (E1/E2, E1/E3, E2/E3, E3/E4, E4/E5, E5/E6, E6/E7a, E6/E7b; Figure 1A). ('CD33', 'Gene', '945', (0, 4)) ('CD33', 'Gene', (0, 4)) ('E3/E4', 'Var', (193, 198)) ('E6/E7a', 'Var', (214, 220)) ('E1/E2', 'Var', (172, 177)) ('E5/E6', 'Var', (207, 212)) ('E2/E3', 'Var', (186, 191)) ('E6/E7b', 'Var', (222, 228)) ('E4/E5', 'Var', (200, 205)) ('E1/E3', 'Var', (179, 184)) 4683 27248327 These 8 splice junctions were consistent with the 3 known and well-supported protein coding CD33 transcripts (NM_001772, NM_001082618, and NM_001177608; Figure 1B). ('NM_001177608', 'Var', (139, 151)) ('NM_001082618', 'Var', (121, 133)) ('CD33', 'Gene', '945', (92, 96)) ('CD33', 'Gene', (92, 96)) ('NM_001772', 'Var', (110, 119)) 4684 27248327 Of the 8 junctions, 6 were shared by 2 or more known CD33 variants, whereas 2 were unique to one of two variants, NM_001082618 (E1/E3; specific for CD33 E2) and NM_001177608 (E6/E7a; specific for CD33E7a). ('CD33E7a', 'Chemical', '-', (196, 203)) ('variants', 'Var', (58, 66)) ('CD33', 'Gene', '945', (148, 152)) ('CD33', 'Gene', (148, 152)) ('NM_001082618', 'Var', (114, 126)) ('CD33', 'Gene', '945', (53, 57)) ('CD33', 'Gene', (53, 57)) ('NM_001177608', 'Var', (161, 173)) ('CD33', 'Gene', '945', (196, 200)) ('CD33', 'Gene', (196, 200)) 4685 27248327 The observed CD33 splice junctions in our RNAseq dataset also offered the possibility of a previously unknown fourth, distinct CD33 transcript that lacked exon 2 and used exon E7a instead of E7b (CD33 E2,E7a). ('exon', 'Protein', (155, 159)) ('CD33', 'Gene', (127, 131)) ('E7b', 'Var', (191, 194)) ('CD33', 'Gene', '945', (127, 131)) ('lacked', 'NegReg', (148, 154)) ('CD33', 'Gene', '945', (13, 17)) ('CD33', 'Gene', (13, 17)) ('CD33', 'Gene', '945', (196, 200)) ('CD33', 'Gene', (196, 200)) 4686 27248327 To further study CD33 variants in human AML and validate the findings obtained with RNAseq, we designed transcript-specific primers and used RT-PCR to assess expression of the CD33FL, CD33 E2, CD33E7a, and CD33 E2,E7a transcripts in 29 pre-treatment specimens from adults with AML. ('human', 'Species', '9606', (34, 39)) ('CD33FL', 'Chemical', '-', (176, 182)) ('CD33', 'Gene', '945', (176, 180)) ('CD33', 'Gene', (176, 180)) ('CD33', 'Gene', '945', (184, 188)) ('CD33', 'Gene', (184, 188)) ('CD33', 'Gene', '945', (206, 210)) ('CD33', 'Gene', (206, 210)) ('AML', 'Disease', 'MESH:D015470', (40, 43)) ('AML', 'Phenotype', 'HP:0004808', (40, 43)) ('AML', 'Disease', (40, 43)) ('CD33E7a', 'Chemical', '-', (193, 200)) ('AML', 'Disease', 'MESH:D015470', (277, 280)) ('AML', 'Phenotype', 'HP:0004808', (277, 280)) ('AML', 'Disease', (277, 280)) ('CD33', 'Gene', '945', (193, 197)) ('CD33', 'Gene', (193, 197)) ('E7a', 'Var', (214, 217)) ('CD33', 'Gene', '945', (17, 21)) ('CD33', 'Gene', (17, 21)) 4688 27248327 Three unique amplicons per CD33 transcript were subjected to Sanger sequencing to confirm the lack of exon 2 in CD33 E2, use of exon 7a in CD33E7a, and lack of exon 2 combined with use of exon 7a in CD33 E2,E7a; the latter sequence was subsequently deposited in GenBank (accession no. ('CD33', 'Gene', (27, 31)) ('CD33', 'Gene', (199, 203)) ('CD33E7a', 'Chemical', '-', (139, 146)) ('CD33', 'Gene', '945', (139, 143)) ('lack', 'NegReg', (94, 98)) ('CD33', 'Gene', (139, 143)) ('CD33', 'Gene', '945', (112, 116)) ('CD33', 'Gene', (112, 116)) ('lack', 'Var', (152, 156)) ('CD33', 'Gene', '945', (199, 203)) ('CD33', 'Gene', '945', (27, 31)) 4692 27248327 Having demonstrated the presence of CD33 E2, CD33E7a, and CD33 E2,E7a mRNA in primary human AML cells, we then studied the functional characteristics of encoded proteins. ('CD33', 'Gene', (45, 49)) ('E7a', 'Var', (66, 69)) ('CD33', 'Gene', '945', (58, 62)) ('AML', 'Disease', 'MESH:D015470', (92, 95)) ('CD33', 'Gene', (58, 62)) ('CD33E7a', 'Chemical', '-', (45, 52)) ('human', 'Species', '9606', (86, 91)) ('CD33', 'Gene', '945', (36, 40)) ('CD33', 'Gene', (36, 40)) ('AML', 'Phenotype', 'HP:0004808', (92, 95)) ('AML', 'Disease', (92, 95)) ('CD33', 'Gene', '945', (45, 49)) ('rat', 'Species', '10116', (14, 17)) 4693 27248327 Therefore, we generated cell line models of human acute leukemia in which we selectively expressed untagged or His-tagged CD33FL, CD33 E2, CD33E7a or CD33 E2,E7a via lentivirus-mediated gene transfer, using a series of human AML (KG-1a, ML- 1, and OCI-AML3) and acute lymphoblastic leukemia (ALL; RCH-ACV, REH, and RS4;11) cell lines. ('rat', 'Species', '10116', (18, 21)) ('human', 'Species', '9606', (44, 49)) ('CD33', 'Gene', '945', (130, 134)) ('acute lymphoblastic leukemia', 'Phenotype', 'HP:0006721', (262, 290)) ('AML', 'Phenotype', 'HP:0004808', (225, 228)) ('AML', 'Disease', (225, 228)) ('leukemia', 'Phenotype', 'HP:0001909', (56, 64)) ('lymphoblastic leukemia', 'Phenotype', 'HP:0005526', (268, 290)) ('ALL', 'Phenotype', 'HP:0006721', (292, 295)) ('leukemia', 'Disease', (56, 64)) ('leukemia', 'Disease', 'MESH:D007938', (56, 64)) ('CD33', 'Gene', '945', (150, 154)) ('CD33E7a', 'Chemical', '-', (139, 146)) ('CD33', 'Gene', (150, 154)) ('AML', 'Disease', 'MESH:D015470', (252, 255)) ('CD33FL', 'Chemical', '-', (122, 128)) ('AML', 'Phenotype', 'HP:0004808', (252, 255)) ('AML', 'Disease', (252, 255)) ('leukemia', 'Phenotype', 'HP:0001909', (282, 290)) ('acute leukemia', 'Phenotype', 'HP:0002488', (50, 64)) ('E7a', 'Var', (158, 161)) ('human', 'Species', '9606', (219, 224)) ('His', 'Chemical', 'MESH:C471213', (111, 114)) ('CD33', 'Gene', '945', (139, 143)) ('CD33', 'Gene', (139, 143)) ('leukemia', 'Disease', 'MESH:D007938', (282, 290)) ('acute lymphoblastic leukemia', 'Disease', (262, 290)) ('leukemia', 'Disease', (282, 290)) ('KG-1a, ML- 1, and OCI-AML3', 'Gene', '112744', (230, 256)) ('CD33', 'Gene', '945', (122, 126)) ('CD33', 'Gene', (130, 134)) ('CD33', 'Gene', (122, 126)) ('acute lymphoblastic leukemia', 'Disease', 'MESH:D054198', (262, 290)) ('AML', 'Disease', 'MESH:D015470', (225, 228)) 4694 27248327 We used tagged versions of the CD33 proteins to distinguish those from endogenously expressed CD33 in some of these cell lines and to be able to track the CD33 E2 and CD33 E2,E7a variants. ('CD33', 'Gene', '945', (155, 159)) ('CD33', 'Gene', '945', (31, 35)) ('CD33', 'Gene', (31, 35)) ('CD33', 'Gene', '945', (94, 98)) ('CD33', 'Gene', (155, 159)) ('CD33', 'Gene', (94, 98)) ('CD33', 'Gene', '945', (167, 171)) ('CD33', 'Gene', (167, 171)) ('E7a', 'Var', (175, 178)) 4695 27248327 While previous studies suggested that one CD33 antibody (clone Him3-4) specifically recognizes the C2- set Ig-like domain and could be used to detect CD33 variants that lack exon 2, in our experiments Him3- 4 only recognized CD33FL and CD33E7a but not CD33 E2 or CD33 E2,E7awhen expressed in virally-transduced ALL cell lines that are devoid of endogenous CD33 (Jurkat, RCH-ACV, REH, or RS4;11: Figure 3) and only recognized CD33FL but not CD33 E2 when expressed in virally-transduced HEK293T cells (Figure 4). ('ALL', 'Phenotype', 'HP:0006721', (311, 314)) ('CD33', 'Gene', '945', (42, 46)) ('CD33', 'Gene', (42, 46)) ('CD33', 'Gene', '945', (263, 267)) ('CD33FL', 'Chemical', '-', (225, 231)) ('CD33', 'Gene', (263, 267)) ('CD33', 'Gene', '945', (150, 154)) ('CD33', 'Gene', '945', (356, 360)) ('CD33', 'Gene', (150, 154)) ('CD33', 'Gene', (356, 360)) ('CD33', 'Gene', '945', (252, 256)) ('CD33', 'Gene', (252, 256)) ('CD33', 'Gene', '945', (236, 240)) ('CD33', 'Gene', (236, 240)) ('CD33FL', 'Chemical', '-', (425, 431)) ('CD33', 'Gene', '945', (425, 429)) ('CD33', 'Gene', (425, 429)) ('CD33', 'Gene', '945', (440, 444)) ('CD33', 'Gene', (440, 444)) ('variants', 'Var', (155, 163)) ('CD33', 'Gene', '945', (225, 229)) ('Jurkat', 'CellLine', 'CVCL:0065', (362, 368)) ('CD33', 'Gene', (225, 229)) ('HEK293T', 'CellLine', 'CVCL:0063', (485, 492)) ('CD33E7a', 'Chemical', '-', (236, 243)) 4696 27248327 As shown in Figure 5, engineered sublines expressed high amounts of CD33FL and CD33E7a in all cell lines. ('CD33FL', 'Chemical', '-', (68, 74)) ('CD33FL', 'Var', (68, 74)) ('CD33E7a', 'Var', (79, 86)) ('CD33E7a', 'Chemical', '-', (79, 86)) 4697 27248327 Lentivirus-mediated gene transfer also resulted in readily detectable cell surface display of the CD33 E2 and CD33 E2,E7a splice variants, although the level of expression varied considerably across cell line backgrounds. ('CD33', 'Gene', '945', (110, 114)) ('CD33', 'Gene', (110, 114)) ('cell surface display', 'MPA', (70, 90)) ('resulted in', 'Reg', (39, 50)) ('E7a', 'Var', (118, 121)) ('CD33', 'Gene', '945', (98, 102)) ('CD33', 'Gene', (98, 102)) 4702 27248327 Overall, our experiments in the panel of sublines of AML and ALL cells expressing His-tagged CD33 spice variants demonstrated that each variant is internalized when bound by a bivalent His antibody (Figure 5). ('bound', 'Interaction', (165, 170)) ('His', 'Chemical', 'MESH:C471213', (82, 85)) ('AML', 'Disease', 'MESH:D015470', (53, 56)) ('ALL', 'Phenotype', 'HP:0006721', (61, 64)) ('CD33', 'Gene', '945', (93, 97)) ('CD33', 'Gene', (93, 97)) ('variants', 'Var', (104, 112)) ('AML', 'Disease', (53, 56)) ('AML', 'Phenotype', 'HP:0004808', (53, 56)) ('rat', 'Species', '10116', (120, 123)) ('internalized', 'MPA', (147, 159)) ('His', 'Chemical', 'MESH:C471213', (185, 188)) 4709 27248327 In 24-hour assays, we found some differences between individual CD33 variants in several acute leukemia cell lines; however, while highly reproducible, these differences were not consistent across cell lines but, rather, appeared cell context specific. ('differences', 'Reg', (33, 44)) ('variants', 'Var', (69, 77)) ('rat', 'Species', '10116', (213, 216)) ('CD33', 'Gene', '945', (64, 68)) ('CD33', 'Gene', (64, 68)) ('leukemia', 'Disease', (95, 103)) ('acute leukemia', 'Phenotype', 'HP:0002488', (89, 103)) ('leukemia', 'Disease', 'MESH:D007938', (95, 103)) ('leukemia', 'Phenotype', 'HP:0001909', (95, 103)) 4722 27248327 One of these variants (CD33 E2) has previously been found in normal myeloid cells, microglial cells, T-lymphocytes, and natural killer (NK) cells, and has been implicated in influencing the susceptibility to Alzheimer's disease. ('variants', 'Var', (13, 21)) ('implicated', 'Reg', (160, 170)) ("Alzheimer's disease", 'Disease', 'MESH:D000544', (208, 227)) ("Alzheimer's disease", 'Phenotype', 'HP:0002511', (208, 227)) ('CD33', 'Gene', '945', (23, 27)) ('CD33', 'Gene', (23, 27)) ("Alzheimer's disease", 'Disease', (208, 227)) ('influencing', 'Reg', (174, 185)) 4731 27248327 Consistent with previous reports, we found that CD33 variants lacking exon 2 can be expressed on the cell surface, although our studies indicate that the efficiency with which this happens varies between different cell line backgrounds. ('CD33', 'Gene', '945', (48, 52)) ('CD33', 'Gene', (48, 52)) ('variants', 'Var', (53, 61)) 4732 27248327 Similar to wild-type CD33, our experiments in engineered acute leukemia cell lines document that the CD33 E2 variant is internalized, and could thus also serve as target for CD33-directed therapeutics that depend on intracellular delivery of a toxic payload. ('CD33', 'Gene', (21, 25)) ('engineered acute leukemia', 'Disease', 'MESH:D015470', (46, 71)) ('variant', 'Var', (109, 116)) ('E2 variant', 'Var', (106, 116)) ('CD33', 'Gene', (101, 105)) ('CD33', 'Gene', '945', (101, 105)) ('leukemia', 'Phenotype', 'HP:0001909', (63, 71)) ('acute leukemia', 'Phenotype', 'HP:0002488', (57, 71)) ('engineered acute leukemia', 'Disease', (46, 71)) ('CD33', 'Gene', '945', (174, 178)) ('CD33', 'Gene', '945', (21, 25)) ('CD33', 'Gene', (174, 178)) 4733 27248327 Our studies in lentivirally-transduced acute leukemia cell lines also show that the CD33 variants that contain exon 7a, and therefore lack almost the entire cytoplasmic tail of CD33, are internalized. ('internalized', 'MPA', (187, 199)) ('variants', 'Var', (89, 97)) ('lack', 'NegReg', (134, 138)) ('leukemia', 'Phenotype', 'HP:0001909', (45, 53)) ('leukemia', 'Disease', 'MESH:D007938', (45, 53)) ('leukemia', 'Disease', (45, 53)) ('CD33', 'Gene', '945', (84, 88)) ('acute leukemia', 'Phenotype', 'HP:0002488', (39, 53)) ('CD33', 'Gene', '945', (177, 181)) ('CD33', 'Gene', (177, 181)) ('CD33', 'Gene', (84, 88)) 4735 27248327 Introduction of point mutations in this domain, for example in the immunoreceptor tyrosine-based inhibitory motifs or clusters of lysine residues, reduced internalization of antibody/CD33 complexes in our earlier studies. ('point mutations', 'Var', (16, 31)) ('CD33', 'Gene', '945', (183, 187)) ('internalization of', 'MPA', (155, 173)) ('CD33', 'Gene', (183, 187)) ('lysine', 'Chemical', 'MESH:D008239', (130, 136)) ('tyrosine', 'Chemical', 'MESH:D014443', (82, 90)) ('reduced', 'NegReg', (147, 154)) 4736 27248327 Whether the cell membrane localization is influenced by the presence of the cytoplasmic tail of CD33, and whether the mechanistic principles for the uptake process differ between wild-type CD33 and variants that contain exon 7a, is currently unknown and will be subject of future investigations. ('CD33', 'Gene', '945', (189, 193)) ('CD33', 'Gene', (189, 193)) ('CD33', 'Gene', (96, 100)) ('cell membrane localization', 'MPA', (12, 38)) ('influenced', 'Reg', (42, 52)) ('variants', 'Var', (198, 206)) ('CD33', 'Gene', '945', (96, 100)) 4737 27248327 The cell context-specific differences we found with regard to modulation of individual CD33 variants would be consistent with such differences in membrane localization and/or internalization mechanisms. ('CD33', 'Gene', '945', (87, 91)) ('CD33', 'Gene', (87, 91)) ('internalization', 'MPA', (175, 190)) ('variants', 'Var', (92, 100)) ('differences', 'Reg', (131, 142)) 4738 27248327 Planned studies will also aim to identify differences between CD33 variants containing exon 7a with wild-type protein with regard to suppression of myeloid cell function - we hypothesize that such differences exist given the lack of immunoreceptor tyrosine-based inhibitory motifs when exon 7a is utilized. ('CD33', 'Gene', '945', (62, 66)) ('CD33', 'Gene', (62, 66)) ('variants', 'Var', (67, 75)) ('tyrosine', 'Chemical', 'MESH:D014443', (248, 256)) 4741 27248327 Furthermore, they identify CD33 variants that lack exon 2 and are not recognized by currently explored CD33-directed therapeutics as potential, hitherto unexploited, targets for immunotherapy with unconjugated or conjugated CD33 antibodies. ('CD33', 'Gene', '945', (103, 107)) ('CD33', 'Gene', (27, 31)) ('CD33', 'Gene', (224, 228)) ('CD33', 'Gene', (103, 107)) ('variants', 'Var', (32, 40)) ('CD33', 'Gene', '945', (224, 228)) ('CD33', 'Gene', '945', (27, 31)) 4750 27248327 Approximately 25 ng of RNA was then reverse transcribed for each PCR reaction with specific primers to amplify CD33FL, CD33 E2, CD33E7a, and CD33 E2,E7a (see Table 2 for primer sequences). ('CD33FL', 'Chemical', '-', (111, 117)) ('CD33', 'Gene', '945', (111, 115)) ('CD33', 'Gene', '945', (128, 132)) ('CD33', 'Gene', (128, 132)) ('E7a', 'Var', (149, 152)) ('CD33', 'Gene', '945', (141, 145)) ('CD33', 'Gene', (141, 145)) ('CD33', 'Gene', (111, 115)) ('CD33E7a', 'Chemical', '-', (128, 135)) ('CD33', 'Gene', '945', (119, 123)) ('CD33', 'Gene', (119, 123)) 4758 27248327 Expression of untagged CD33 variants on leukemia cell lines was quantified by flow cytometry using an unconjugated CD33 antibody (clone P67.6; BD Biosciences, San Jose, CA, USA) or unconjugated CD33 antibody followed by a biotin-anti-mouse secondary antibody and APC-streptavidin (both BD Biosciences). ('mouse', 'Species', '10090', (234, 239)) ('P67', 'Gene', (136, 139)) ('CD33', 'Gene', '945', (115, 119)) ('variants', 'Var', (28, 36)) ('P67', 'Gene', '945', (136, 139)) ('leukemia', 'Phenotype', 'HP:0001909', (40, 48)) ('CD33', 'Gene', (115, 119)) ('CD33', 'Gene', '945', (194, 198)) ('CD33', 'Gene', (194, 198)) ('CD33', 'Gene', '945', (23, 27)) ('biotin', 'Chemical', 'MESH:D001710', (222, 228)) ('CD33', 'Gene', (23, 27)) ('leukemia', 'Disease', (40, 48)) ('leukemia', 'Disease', 'MESH:D007938', (40, 48)) 4759 27248327 In some cases, a second CD33 antibody (clone HIM3-4) previously reported to recognize the C2-set Ig-like domain of CD33 was used to detect cell surface display of CD33 variants. ('CD33', 'Gene', (24, 28)) ('CD33', 'Gene', '945', (115, 119)) ('CD33', 'Gene', '945', (163, 167)) ('variants', 'Var', (168, 176)) ('CD33', 'Gene', (115, 119)) ('CD33', 'Gene', (163, 167)) ('CD33', 'Gene', '945', (24, 28)) 4785 25428027 Deregulation of canonical or noncanonical Wnt signaling pathway plays critical roles in the pathogenesis of various cancers including AML. ('Deregulation', 'Var', (0, 12)) ('canonical or noncanonical Wnt signaling pathway', 'Pathway', (16, 63)) ('AML', 'Disease', 'MESH:D015470', (134, 137)) ('cancers', 'Disease', 'MESH:D009369', (116, 123)) ('cancers', 'Phenotype', 'HP:0002664', (116, 123)) ('cancers', 'Disease', (116, 123)) ('AML', 'Disease', (134, 137)) ('AML', 'Phenotype', 'HP:0004808', (134, 137)) 4788 25428027 Deregulation of the canonical branch of Wnt signaling pathway by aberrant stabilization and constitutive activation of beta-catenin is linked to the initiation and progression of AML and other cancers. ('AML', 'Disease', (179, 182)) ('stabilization', 'MPA', (74, 87)) ('beta-catenin', 'Gene', '1499', (119, 131)) ('cancers', 'Disease', 'MESH:D009369', (193, 200)) ('cancers', 'Phenotype', 'HP:0002664', (193, 200)) ('Deregulation', 'Var', (0, 12)) ('canonical branch of Wnt signaling pathway', 'Pathway', (20, 61)) ('cancers', 'Disease', (193, 200)) ('aberrant', 'Var', (65, 73)) ('activation', 'PosReg', (105, 115)) ('linked', 'Reg', (135, 141)) ('beta-catenin', 'Gene', (119, 131)) ('AML', 'Disease', 'MESH:D015470', (179, 182)) ('AML', 'Phenotype', 'HP:0004808', (179, 182)) 4790 25428027 In both AML cell lines and primary samples, silencing genes associated with the canonical Wnt/beta-catenin pathway through methylation have been observed. ('AML', 'Phenotype', 'HP:0004808', (8, 11)) ('AML', 'Disease', (8, 11)) ('methylation', 'Var', (123, 134)) ('beta-catenin', 'Gene', (94, 106)) ('beta-catenin', 'Gene', '1499', (94, 106)) ('silencing', 'NegReg', (44, 53)) ('AML', 'Disease', 'MESH:D015470', (8, 11)) 4791 25428027 In addition, inhibition of the Wnt/beta-catenin pathway by small-molecules results in apoptosis of AML cells. ('beta-catenin', 'Gene', (35, 47)) ('AML', 'Disease', 'MESH:D015470', (99, 102)) ('beta-catenin', 'Gene', '1499', (35, 47)) ('small-molecules', 'Var', (59, 74)) ('AML', 'Phenotype', 'HP:0004808', (99, 102)) ('AML', 'Disease', (99, 102)) ('inhibition', 'NegReg', (13, 23)) ('apoptosis', 'CPA', (86, 95)) 4805 25428027 Primary antibodies for beta-catenin, p-PKC (Thr638), PKC, p-CaMKII (Thr286), CaMKII and beta-actin were purchased from Abcam. ('Thr286', 'Chemical', '-', (68, 74)) ('beta-actin', 'Gene', '728378', (88, 98)) ('beta-actin', 'Gene', (88, 98)) ('CaMKII', 'Gene', (77, 83)) ('Thr638', 'Var', (44, 50)) ('CaMKII', 'Gene', '818', (60, 66)) ('CaMKII', 'Gene', (60, 66)) ('beta-catenin', 'Gene', (23, 35)) ('CaMKII', 'Gene', '818', (77, 83)) ('PKC', 'Gene', (53, 56)) ('PKC', 'Gene', (39, 42)) ('Thr286', 'Var', (68, 74)) ('PKC', 'Gene', '112476', (39, 42)) ('Thr638', 'Chemical', '-', (44, 50)) ('PKC', 'Gene', '112476', (53, 56)) ('beta-catenin', 'Gene', '1499', (23, 35)) 4826 25428027 The negative control FOPFLASH construct contains mutated TCF/LEF binding sites. ('mutated', 'Var', (49, 56)) ('TCF/LEF', 'Gene', '3172', (57, 64)) ('TCF/LEF', 'Gene', (57, 64)) 4868 25428027 noted that silencing of beta-catenin, using a short hairpin RNA (shRNA) lentiviral approach, was the strikingly enhanced myeloid differentiation of the HL-60 cell line after ATRA induction, which was consistent with our results. ('enhanced', 'PosReg', (112, 120)) ('beta-catenin', 'Gene', (24, 36)) ('silencing', 'Var', (11, 20)) ('beta-catenin', 'Gene', '1499', (24, 36)) ('ATRA', 'Chemical', 'MESH:D014212', (174, 178)) ('myeloid differentiation of the', 'CPA', (121, 151)) ('HL-60', 'CellLine', 'CVCL:0002', (152, 157)) 4883 25428027 This result further suggested that deregulation of the Wnt signaling pathway was critical in the pathogenesis of AML. ('deregulation', 'Var', (35, 47)) ('AML', 'Disease', 'MESH:D015470', (113, 116)) ('Wnt signaling pathway', 'Pathway', (55, 76)) ('AML', 'Phenotype', 'HP:0004808', (113, 116)) ('AML', 'Disease', (113, 116)) 4886 25428027 Overexpression of beta-catenin is thought to be an independent adverse prognostic factor in AML. ('beta-catenin', 'Gene', (18, 30)) ('AML', 'Disease', 'MESH:D015470', (92, 95)) ('beta-catenin', 'Gene', '1499', (18, 30)) ('Overexpression', 'Var', (0, 14)) ('AML', 'Phenotype', 'HP:0004808', (92, 95)) ('AML', 'Disease', (92, 95)) 4888 25428027 In a recent study, aberrant methylation of canonical Wnt antagonists was detected in four AML cell lines and in up to 64% of AML marrow samples. ('AML', 'Disease', 'MESH:D015470', (125, 128)) ('AML', 'Phenotype', 'HP:0004808', (90, 93)) ('AML', 'Disease', (90, 93)) ('detected', 'Reg', (73, 81)) ('methylation', 'MPA', (28, 39)) ('AML', 'Disease', (125, 128)) ('AML', 'Phenotype', 'HP:0004808', (125, 128)) ('AML', 'Disease', 'MESH:D015470', (90, 93)) ('aberrant', 'Var', (19, 27)) 4901 25428027 Silencing Wnt5a, a non-canonical Wnt ligand, by methylation has been reported in some types of leukemia. ('reported', 'Reg', (69, 77)) ('Silencing', 'NegReg', (0, 9)) ('Wnt5a', 'Gene', (10, 15)) ('leukemia', 'Disease', (95, 103)) ('Wnt5a', 'Gene', '7474', (10, 15)) ('methylation', 'Var', (48, 59)) ('leukemia', 'Disease', 'MESH:D007938', (95, 103)) ('leukemia', 'Phenotype', 'HP:0001909', (95, 103)) 4912 25428027 reported that in adipocyte differentiation, the noncanonical Wnt signaling pathway inhibited the canonical Wnt signaling pathway, and BIM inhibited PKC that both can activate the Wnt/beta-catenin signaling pathway. ('BIM', 'Chemical', '-', (134, 137)) ('beta-catenin', 'Gene', (183, 195)) ('BIM', 'Var', (134, 137)) ('PKC', 'Gene', (148, 151)) ('inhibited', 'NegReg', (83, 92)) ('PKC', 'Gene', '112476', (148, 151)) ('canonical Wnt signaling pathway', 'Pathway', (97, 128)) ('beta-catenin', 'Gene', '1499', (183, 195)) ('noncanonical Wnt signaling pathway', 'Pathway', (48, 82)) ('activate', 'PosReg', (166, 174)) ('inhibited', 'NegReg', (138, 147)) ('adipocyte differentiation', 'CPA', (17, 42)) 4922 24895580 FLT3-ITD, NPM1, and DNMT3A mutations frequently occurred in AML patients and have been found conferred with myeloproliferative neoplasms in mouse model. ('mutations', 'Var', (27, 36)) ('myeloproliferative neoplasms', 'Disease', (108, 136)) ('occurred', 'Reg', (48, 56)) ('mouse', 'Species', '10090', (140, 145)) ('AML', 'Disease', 'MESH:D015470', (60, 63)) ('patients', 'Species', '9606', (64, 72)) ('myeloproliferative neoplasms', 'Phenotype', 'HP:0005547', (108, 136)) ('DNMT3A', 'Gene', (20, 26)) ('NPM1', 'Gene', (10, 14)) ('FLT3-ITD', 'Gene', (0, 8)) ('DNMT3A', 'Gene', '1788', (20, 26)) ('myeloproliferative neoplasms', 'Disease', 'MESH:D009196', (108, 136)) ('AML', 'Disease', (60, 63)) ('neoplasms', 'Phenotype', 'HP:0002664', (127, 136)) ('conferred with', 'Reg', (93, 107)) ('NPM1', 'Gene', '4869', (10, 14)) 4923 24895580 Therefore, we sought to search for mutations in JAK2V617F, FLT3-ITD, NPM1, and DNMT3A in 129 cases including 120 classic MPN cases and 9 MDS/MPN cases. ('MDS', 'Disease', 'MESH:D009190', (137, 140)) ('FLT3-ITD', 'Gene', (59, 67)) ('JAK2', 'Gene', '3717', (48, 52)) ('DNMT3A', 'Gene', (79, 85)) ('MDS', 'Disease', (137, 140)) ('DNMT3A', 'Gene', '1788', (79, 85)) ('NPM1', 'Gene', (69, 73)) ('JAK2', 'Gene', (48, 52)) ('NPM1', 'Gene', '4869', (69, 73)) ('mutations', 'Var', (35, 44)) 4928 24895580 Since the discovery of the JAK2V617F mutation in 2005, an increasing number of novel somatic and germline mutations have been described in MPN in recent years, including myeloproliferative leukemia virus (MPL), TET oncogene family member 2 (TET2), additional sex combs-like 1 (ASXL1), casitas B-lineage lymphoma proto-oncogene (CBL), isocitrate dehydrogenase (IDH), and IKAROS family zinc finger 1 (IKZF1). ('CBL', 'Gene', (328, 331)) ('mutation', 'Var', (37, 45)) ('ASXL1', 'Gene', (277, 282)) ('lymphoma', 'Phenotype', 'HP:0002665', (303, 311)) ('CBL', 'Gene', '867', (328, 331)) ('leukemia', 'Phenotype', 'HP:0001909', (189, 197)) ('TET2', 'Gene', (241, 245)) ('myeloproliferative leukemia', 'Phenotype', 'HP:0005547', (170, 197)) ('JAK2', 'Gene', '3717', (27, 31)) ('myeloproliferative leukemia virus', 'Species', '11973', (170, 203)) ('lymphoma', 'Disease', (303, 311)) ('lymphoma', 'Disease', 'MESH:D008223', (303, 311)) ('ASXL1', 'Gene', '171023', (277, 282)) ('MPL', 'Species', '11973', (205, 208)) ('B-lineage lymphoma', 'Phenotype', 'HP:0012191', (293, 311)) ('myeloproliferative leukemia virus', 'Disease', (170, 203)) ('mutations', 'Var', (106, 115)) ('JAK2', 'Gene', (27, 31)) ('TET2', 'Gene', '54790', (241, 245)) 4929 24895580 Several lines of evidence suggest that mutations in genes other than these mentioned above must be present in MPN patients, and the initiating genetic events responsible for the development of MPN are still not totally understood. ('patients', 'Species', '9606', (114, 122)) ('MPN', 'Disease', (110, 113)) ('mutations', 'Var', (39, 48)) 4932 24895580 FLT3-ITD (internal tandem duplication of FLT3) is the most prevalent mutation found in AML and has been identified in 20-30% of all AML patients. ('FLT3', 'Gene', (41, 45)) ('internal tandem duplication', 'Var', (10, 37)) ('AML', 'Disease', (87, 90)) ('identified', 'Reg', (104, 114)) ('AML', 'Disease', (132, 135)) ('patients', 'Species', '9606', (136, 144)) ('AML', 'Disease', 'MESH:D015470', (87, 90)) ('AML', 'Disease', 'MESH:D015470', (132, 135)) 4933 24895580 Studies suggest that AML patients with FLT3-ITD have significantly elevated peripheral white blood cell counts and increased bone marrow blasts at diagnosis. ('bone marrow blasts', 'CPA', (125, 143)) ('elevated peripheral white blood cell counts', 'Phenotype', 'HP:0001974', (67, 110)) ('elevated', 'PosReg', (67, 75)) ('increased', 'PosReg', (115, 124)) ('increased bone marrow', 'Phenotype', 'HP:0005528', (115, 136)) ('AML', 'Disease', 'MESH:D015470', (21, 24)) ('patients', 'Species', '9606', (25, 33)) ('peripheral', 'CPA', (76, 86)) ('AML', 'Disease', (21, 24)) ('FLT3-ITD', 'Var', (39, 47)) 4934 24895580 recently showed that knock-in of an ITD mutation into murine FLT3 conferred myeloproliferative disease in a mouse model, which indicated the potential involvement of FLT3-ITD in MPN. ('myeloproliferative disease', 'Disease', (76, 102)) ('mutation', 'Var', (40, 48)) ('conferred', 'Reg', (66, 75)) ('myeloproliferative disease', 'Disease', 'MESH:D009196', (76, 102)) ('murine', 'Species', '10090', (54, 60)) ('mouse', 'Species', '10090', (108, 113)) ('myeloproliferative disease', 'Phenotype', 'HP:0005547', (76, 102)) ('FLT3', 'Gene', (61, 65)) 4937 24895580 NPM1 mutations are known to be common in AML and are commonly associated with a diploid karyotype. ('associated', 'Reg', (62, 72)) ('mutations', 'Var', (5, 14)) ('AML', 'Disease', 'MESH:D015470', (41, 44)) ('NPM1', 'Gene', (0, 4)) ('NPM1', 'Gene', '4869', (0, 4)) ('AML', 'Disease', (41, 44)) 4938 24895580 demonstrated that NPM1 acts indeed as a haploinsufficient tumor suppressor gene in vivo, while some findings suggested that NPM1 mutation in AML is likely a gain-of-function one rather than simple haploinsufficiency. ('tumor', 'Phenotype', 'HP:0002664', (58, 63)) ('AML', 'Disease', (141, 144)) ('NPM1', 'Gene', '4869', (18, 22)) ('NPM1', 'Gene', '4869', (124, 128)) ('haploinsufficient tumor', 'Disease', 'MESH:D058495', (40, 63)) ('mutation', 'Var', (129, 137)) ('haploinsufficiency', 'Disease', (197, 215)) ('haploinsufficient tumor', 'Disease', (40, 63)) ('NPM1', 'Gene', (124, 128)) ('NPM1', 'Gene', (18, 22)) ('AML', 'Disease', 'MESH:D015470', (141, 144)) ('haploinsufficiency', 'Disease', 'MESH:D058495', (197, 215)) 4939 24895580 The most common NPM1 mutation in AML is a duplication of a TCTG tetra-nucleotide at positions 956 to 959 of the reference sequence (GenBank accession number NM_002520) and accounts for 75% to 80% of cases. ('NPM1', 'Gene', (16, 20)) ('AML', 'Disease', 'MESH:D015470', (33, 36)) ('mutation', 'Var', (21, 29)) ('AML', 'Disease', (33, 36)) ('NPM1', 'Gene', '4869', (16, 20)) ('tetra', 'Species', '42554', (64, 69)) 4940 24895580 One "conventional" knock-in model of NPM1 mutation demonstrated that NPM1 mutation can result in myeloproliferative disease but is insufficient for leukemogenesis. ('mutation', 'Var', (74, 82)) ('myeloproliferative disease', 'Disease', 'MESH:D009196', (97, 123)) ('myeloproliferative disease', 'Disease', (97, 123)) ('NPM1', 'Gene', (37, 41)) ('myeloproliferative disease', 'Phenotype', 'HP:0005547', (97, 123)) ('NPM1', 'Gene', '4869', (37, 41)) ('mutation', 'Var', (42, 50)) ('result in', 'Reg', (87, 96)) ('NPM1', 'Gene', (69, 73)) ('NPM1', 'Gene', '4869', (69, 73)) 4941 24895580 Nevertheless, the frequency of NPM1 mutation and its possible pathogenetic role in MPNs are rarely investigated until now. ('NPM1', 'Gene', '4869', (31, 35)) ('MPNs', 'Phenotype', 'HP:0005547', (83, 87)) ('MPNs', 'Disease', (83, 87)) ('NPM1', 'Gene', (31, 35)) ('mutation', 'Var', (36, 44)) 4942 24895580 Alterations of epigenetic markers are thought to play an important role in myeloid malignancies. ('myeloid malignancies', 'Disease', (75, 95)) ('Alterations', 'Var', (0, 11)) ('epigenetic markers', 'Protein', (15, 33)) ('myeloid malignancies', 'Disease', 'MESH:D009369', (75, 95)) 4944 24895580 More recently, a whole-genome sequencing study in AML uncovered recurrent mutations of DNMT3A in 22% of AML patients and DNMT3A mutations were associated with poor outcome. ('associated', 'Reg', (143, 153)) ('AML', 'Disease', (104, 107)) ('AML', 'Disease', 'MESH:D015470', (50, 53)) ('mutations', 'Var', (74, 83)) ('AML', 'Disease', (50, 53)) ('AML', 'Disease', 'MESH:D015470', (104, 107)) ('DNMT3A', 'Gene', (121, 127)) ('patients', 'Species', '9606', (108, 116)) ('DNMT3A', 'Gene', '1788', (121, 127)) ('DNMT3A', 'Gene', (87, 93)) ('DNMT3A', 'Gene', '1788', (87, 93)) 4945 24895580 Many of the reported DNMT3A mutations mainly occurred at codon R882 in exon 23, but the occurrence of DNMT3A mutations in MPN patients is not well clarified. ('DNMT3A', 'Gene', (102, 108)) ('DNMT3A', 'Gene', '1788', (102, 108)) ('DNMT3A', 'Gene', (21, 27)) ('DNMT3A', 'Gene', '1788', (21, 27)) ('patients', 'Species', '9606', (126, 134)) ('occurred', 'Reg', (45, 53)) ('mutations', 'Var', (28, 37)) 4947 24895580 Furthermore, some studies indicate that the mutagenesis of these genes may differ within different races and be associated with patient's age. ('mutagenesis', 'Var', (44, 55)) ('associated', 'Reg', (112, 122)) ('patient', 'Species', '9606', (128, 135)) 4956 24895580 The samples with mutated FLT3-ITD, mutated NPM1, or mutated DNMT3A were used as positive controls. ('mutated', 'Var', (35, 42)) ('NPM1', 'Gene', (43, 47)) ('DNMT3A', 'Gene', (60, 66)) ('DNMT3A', 'Gene', '1788', (60, 66)) ('NPM1', 'Gene', '4869', (43, 47)) ('FLT3-ITD', 'Gene', (25, 33)) ('mutated', 'Var', (17, 24)) 4967 24895580 In older patients, JAK2V617F mutation rate in PV patients was higher than in ET, MF, and MPN-u patients; however, this phenomenon was not seen in younger patients. ('mutation', 'Var', (29, 37)) ('patients', 'Species', '9606', (9, 17)) ('JAK2', 'Gene', (19, 23)) ('patients', 'Species', '9606', (95, 103)) ('patients', 'Species', '9606', (154, 162)) ('higher', 'PosReg', (62, 68)) ('patients', 'Species', '9606', (49, 57)) ('JAK2', 'Gene', '3717', (19, 23)) 4972 24895580 By sequence analysis of the DNMT3A gene, we found DNMT3A mutations in 2 patients. ('DNMT3A', 'Gene', (50, 56)) ('DNMT3A', 'Gene', '1788', (50, 56)) ('found', 'Reg', (44, 49)) ('DNMT3A', 'Gene', (28, 34)) ('DNMT3A', 'Gene', '1788', (28, 34)) ('mutations', 'Var', (57, 66)) ('patients', 'Species', '9606', (72, 80)) 4973 24895580 The two mutations were heterozygous and missense: one was a MF patient (c.2644C>T, p.R882C; JAK2V617F-positive); the other one was a MDS/MPN patient (c.2645G>A, p.R882H; JAK2V617F-negative) (Figure 1). ('JAK2', 'Gene', (170, 174)) ('c.2645G>A', 'Var', (150, 159)) ('JAK2', 'Gene', (92, 96)) ('c.2644C>T', 'Mutation', 'rs377577594', (72, 81)) ('p.R882C', 'Var', (83, 90)) ('patient', 'Species', '9606', (141, 148)) ('c.2645G>A', 'Mutation', 'rs147001633', (150, 159)) ('MDS', 'Disease', (133, 136)) ('patient', 'Species', '9606', (63, 70)) ('MDS', 'Disease', 'MESH:D009190', (133, 136)) ('JAK2', 'Gene', '3717', (170, 174)) ('c.2644C>T', 'Var', (72, 81)) ('p.R882H', 'Mutation', 'rs147001633', (161, 168)) ('JAK2', 'Gene', '3717', (92, 96)) ('p.R882C', 'Mutation', 'rs377577594', (83, 90)) 4980 24895580 However, our present study did not reveal any sequence variation in the 120 MPN patients and 9 MDS/MPN patients we studied, as shown in Figures 2(a) and 2(c), while Figures 2(b) and 2(d) showed the chromatograms of positive controls (samples from AML patients with FLT3-ITD or NPM1 mutation; Figure 2(b) showed the FLT3 internal tandem duplications mutation in exons 11 and 12; Figure 2(d) showed the insertion of a TCTG tetra nucleotide at positions 956 to 959 of the reference sequence.). ('patients', 'Species', '9606', (80, 88)) ('mutation', 'Var', (349, 357)) ('NPM1', 'Gene', (277, 281)) ('internal tandem duplications mutation', 'Var', (320, 357)) ('tetra', 'Species', '42554', (421, 426)) ('MDS', 'Disease', (95, 98)) ('NPM1', 'Gene', '4869', (277, 281)) ('AML', 'Disease', 'MESH:D015470', (247, 250)) ('MDS', 'Disease', 'MESH:D009190', (95, 98)) ('patients', 'Species', '9606', (251, 259)) ('patients', 'Species', '9606', (103, 111)) ('AML', 'Disease', (247, 250)) ('FLT3', 'Gene', (315, 319)) 4981 24895580 All these results suggested that the studied FLT3-ITD or NPM1 mutation points are unlikely the candidate factors for human MPN development. ('human', 'Species', '9606', (117, 122)) ('NPM1', 'Gene', (57, 61)) ('NPM1', 'Gene', '4869', (57, 61)) ('mutation points', 'Var', (62, 77)) ('FLT3-ITD', 'Gene', (45, 53)) 4984 24895580 However, none of the 120 MPN patients and 9 MDS/MPN patients displayed known FLT3-ITD mutation at the juxtamembrane (JM) coding sequences and NPM1 mutation. ('patients', 'Species', '9606', (29, 37)) ('NPM1', 'Gene', '4869', (142, 146)) ('MDS', 'Disease', (44, 47)) ('FLT3-ITD', 'Gene', (77, 85)) ('MDS', 'Disease', 'MESH:D009190', (44, 47)) ('mutation', 'Var', (86, 94)) ('NPM1', 'Gene', (142, 146)) ('patients', 'Species', '9606', (52, 60)) 4996 24895580 The majority of FLT3 mutations are ITDs in the JM domain encoded by exons 11 and 12 and were first reported in patients with AML in 1996. ('AML', 'Disease', 'MESH:D015470', (125, 128)) ('mutations', 'Var', (21, 30)) ('AML', 'Disease', (125, 128)) ('FLT3', 'Gene', (16, 20)) ('patients', 'Species', '9606', (111, 119)) 4997 24895580 detected the patients with various malignant hematologic diseases and found that FLT3-ITD mutation mainly occurred in AML patients and might be a strong prognostic factor. ('patients', 'Species', '9606', (13, 21)) ('FLT3-ITD', 'Gene', (81, 89)) ('AML', 'Disease', (118, 121)) ('malignant hematologic diseases', 'Disease', 'MESH:D019337', (35, 65)) ('patients', 'Species', '9606', (122, 130)) ('malignant hematologic diseases', 'Disease', (35, 65)) ('hematologic diseases', 'Phenotype', 'HP:0001871', (45, 65)) ('mutation', 'Var', (90, 98)) ('occurred', 'Reg', (106, 114)) ('AML', 'Disease', 'MESH:D015470', (118, 121)) 4998 24895580 In another study, FLT3 mutations were also observed in patients with MDS or CMML, but at a much lower frequency than AML, and did not predict poor outcome. ('MDS', 'Disease', (69, 72)) ('MDS', 'Disease', 'MESH:D009190', (69, 72)) ('AML', 'Disease', (117, 120)) ('FLT3', 'Gene', (18, 22)) ('observed', 'Reg', (43, 51)) ('mutations', 'Var', (23, 32)) ('CMML', 'Disease', (76, 80)) ('patients', 'Species', '9606', (55, 63)) ('AML', 'Disease', 'MESH:D015470', (117, 120)) ('CMML', 'Disease', 'MESH:D054429', (76, 80)) 5000 24895580 Several studies in animal models uncovered the importance of FLT3-ITD in MPN; four studies have indicated that FLT3-ITD could induce myeloproliferative disease using transgenic mouse models, respectively. ('induce', 'Reg', (126, 132)) ('FLT3-ITD', 'Var', (111, 119)) ('myeloproliferative disease', 'Phenotype', 'HP:0005547', (133, 159)) ('mouse', 'Species', '10090', (177, 182)) ('myeloproliferative disease', 'Disease', (133, 159)) ('myeloproliferative disease', 'Disease', 'MESH:D009196', (133, 159)) 5003 24895580 report that no FLT3 mutations were found in a cohort of patients with chronic myeloid disorders, while being in contrast to Lin's study that FLT3 mutations occur in approximately 10% of Philadelphia (Ph) chromosome-CMPD and CMPD/MDS. ('patients', 'Species', '9606', (56, 64)) ('CMPD', 'Disease', 'MESH:D055036', (215, 219)) ('mutations', 'Var', (146, 155)) ('FLT3', 'Gene', (141, 145)) ('MDS', 'Disease', (229, 232)) ('MDS', 'Disease', 'MESH:D009190', (229, 232)) ('CMPD', 'Disease', (224, 228)) ('chronic myeloid disorders', 'Disease', (70, 95)) ('CMPD', 'Disease', 'MESH:D055036', (224, 228)) ('chronic myeloid disorders', 'Disease', 'MESH:D015464', (70, 95)) ('chronic myeloid disorders', 'Phenotype', 'HP:0005506', (70, 95)) ('Philadelphia', 'Disease', (186, 198)) ('CMPD', 'Disease', (215, 219)) ('occur', 'Reg', (156, 161)) 5004 24895580 In a recent article, FLT3 mutation analysis was performed on 90 cases of JAK2-negative MPNs or MDS/MPNs and 62 cases of JAK2V617F-positive MPNs. ('FLT3', 'Gene', (21, 25)) ('MPNs', 'Phenotype', 'HP:0005547', (99, 103)) ('JAK2', 'Gene', '3717', (120, 124)) ('performed', 'Reg', (48, 57)) ('mutation', 'Var', (26, 34)) ('MPNs', 'Phenotype', 'HP:0005547', (87, 91)) ('JAK2', 'Gene', (120, 124)) ('MDS', 'Disease', (95, 98)) ('MDS', 'Disease', 'MESH:D009190', (95, 98)) ('JAK2', 'Gene', '3717', (73, 77)) ('MPNs', 'Phenotype', 'HP:0005547', (139, 143)) ('JAK2', 'Gene', (73, 77)) ('MPNs', 'Disease', (87, 91)) 5006 24895580 NPM1 mutations, first identified by the aberrant cytoplasmic localization of NPM1 protein, were found to be frequent events in AML. ('NPM1', 'Gene', (77, 81)) ('mutations', 'Var', (5, 14)) ('NPM1', 'Gene', '4869', (77, 81)) ('AML', 'Disease', 'MESH:D015470', (127, 130)) ('NPM1', 'Gene', (0, 4)) ('NPM1', 'Gene', '4869', (0, 4)) ('AML', 'Disease', (127, 130)) 5007 24895580 Some animal models bearing enforced human NPM1-mutant expression showed an expansion of hematopoietic cells and developed myeloproliferation, indicating a pathogenic role of mutant NPM1 protein in myeloid disorders. ('NPM1', 'Gene', '4869', (42, 46)) ('myeloproliferation', 'Disease', (122, 140)) ('NPM1', 'Gene', (181, 185)) ('myeloid disorders', 'Disease', 'MESH:D007951', (197, 214)) ('expansion', 'PosReg', (75, 84)) ('NPM1', 'Gene', (42, 46)) ('protein', 'Protein', (186, 193)) ('hematopoietic cells', 'CPA', (88, 107)) ('NPM1', 'Gene', '4869', (181, 185)) ('human', 'Species', '9606', (36, 41)) ('developed', 'CPA', (112, 121)) ('myeloid disorders', 'Disease', (197, 214)) ('mutant', 'Var', (174, 180)) 5008 24895580 Some studies showed that NPM1 mutation occurred with low frequencies in patients with MDS, while others found no mutations in MDS. ('MDS', 'Disease', (86, 89)) ('MDS', 'Disease', 'MESH:D009190', (86, 89)) ('mutation', 'Var', (30, 38)) ('patients', 'Species', '9606', (72, 80)) ('NPM1', 'Gene', (25, 29)) ('NPM1', 'Gene', '4869', (25, 29)) ('MDS', 'Disease', (126, 129)) ('MDS', 'Disease', 'MESH:D009190', (126, 129)) 5009 24895580 showed that NPM1 mutations occurred in 6/187 (3%) MDS/MPN patients and the 6 patients were all CMML patients, indicating that NPM1 mutation may be associated with a poor prognosis. ('NPM1', 'Gene', (12, 16)) ('patients', 'Species', '9606', (58, 66)) ('CMML', 'Disease', (95, 99)) ('NPM1', 'Gene', '4869', (12, 16)) ('NPM1', 'Gene', (126, 130)) ('MDS', 'Disease', (50, 53)) ('patients', 'Species', '9606', (77, 85)) ('patients', 'Species', '9606', (100, 108)) ('MDS', 'Disease', 'MESH:D009190', (50, 53)) ('CMML', 'Disease', 'MESH:D054429', (95, 99)) ('occurred', 'Reg', (27, 35)) ('NPM1', 'Gene', '4869', (126, 130)) ('mutations', 'Var', (17, 26)) ('mutation', 'Var', (131, 139)) 5010 24895580 found that NPM1 mutation was observed in 6/67 secondary AML (s-AML) patients with a history of MPN and concluded that the NPM1 mutations are not only a key factor in the initiation of de novo AML but may contribute to s-AML following MPN. ('contribute', 'Reg', (204, 214)) ('AML', 'Disease', (63, 66)) ('mutation', 'Var', (16, 24)) ('observed', 'Reg', (29, 37)) ('NPM1', 'Gene', '4869', (11, 15)) ('AML', 'Disease', 'MESH:D015470', (63, 66)) ('NPM1', 'Gene', (122, 126)) ('AML', 'Disease', (56, 59)) ('AML', 'Disease', 'MESH:D015470', (192, 195)) ('patients', 'Species', '9606', (68, 76)) ('AML', 'Disease', 'MESH:D015470', (220, 223)) ('AML', 'Disease', (192, 195)) ('NPM1', 'Gene', '4869', (122, 126)) ('mutations', 'Var', (127, 136)) ('NPM1', 'Gene', (11, 15)) ('AML', 'Disease', 'MESH:D015470', (56, 59)) ('AML', 'Disease', (220, 223)) 5011 24895580 However, relatively few data regarding the presence of NPM1 mutations are available for classic MPN cases. ('mutations', 'Var', (60, 69)) ('NPM1', 'Gene', (55, 59)) ('NPM1', 'Gene', '4869', (55, 59)) ('MPN', 'Disease', (96, 99)) 5014 24895580 Mutations in DNMT3A were shown to be one of the early initiating events in AML pathogenesis. ('AML', 'Disease', 'MESH:D015470', (75, 78)) ('DNMT3A', 'Gene', (13, 19)) ('DNMT3A', 'Gene', '1788', (13, 19)) ('AML', 'Disease', (75, 78)) ('Mutations', 'Var', (0, 9)) 5015 24895580 DNMT3A mutations were also noted in patients with MDS and s-AML. ('AML', 'Disease', 'MESH:D015470', (60, 63)) ('patients', 'Species', '9606', (36, 44)) ('MDS', 'Disease', (50, 53)) ('MDS', 'Disease', 'MESH:D009190', (50, 53)) ('DNMT3A', 'Gene', (0, 6)) ('AML', 'Disease', (60, 63)) ('DNMT3A', 'Gene', '1788', (0, 6)) ('mutations', 'Var', (7, 16)) 5016 24895580 found DNMT3A mutations in 8% of MDS patients, similar to the trend noted in AML. ('AML', 'Disease', (76, 79)) ('patients', 'Species', '9606', (36, 44)) ('MDS', 'Disease', (32, 35)) ('MDS', 'Disease', 'MESH:D009190', (32, 35)) ('DNMT3A', 'Gene', (6, 12)) ('DNMT3A', 'Gene', '1788', (6, 12)) ('mutations', 'Var', (13, 22)) ('AML', 'Disease', 'MESH:D015470', (76, 79)) 5017 24895580 The studies about DNMT3A mutation in MPN were limited and inconsistent. ('DNMT3A', 'Gene', '1788', (18, 24)) ('mutation', 'Var', (25, 33)) ('DNMT3A', 'Gene', (18, 24)) 5019 24895580 reported DNMT3A mutations in 7% PV, 15% MF, and 14.3% s-AML and indicated DNMT3A alterations occurred concurrently with JAK2. ('JAK2', 'Gene', '3717', (120, 124)) ('DNMT3A', 'Gene', (74, 80)) ('AML', 'Disease', (56, 59)) ('mutations', 'Var', (16, 25)) ('JAK2', 'Gene', (120, 124)) ('DNMT3A', 'Gene', '1788', (74, 80)) ('AML', 'Disease', 'MESH:D015470', (56, 59)) ('DNMT3A', 'Gene', (9, 15)) ('DNMT3A', 'Gene', '1788', (9, 15)) 5024 24895580 Therefore, DNMT3A mutation may not be a frequent characteristic of MPN, but could be a poor prognostic indicator, always concurrently with JAK2V617F mutation. ('JAK2', 'Gene', (139, 143)) ('DNMT3A', 'Gene', (11, 17)) ('mutation', 'Var', (18, 26)) ('DNMT3A', 'Gene', '1788', (11, 17)) ('JAK2', 'Gene', '3717', (139, 143)) 5025 24895580 However, larger cohort of patients are needed to determine the exact frequency of DNMT3A mutations in Chinese MPN patients and to clarify its role in the molecular pathogenesis of MPN. ('DNMT3A', 'Gene', (82, 88)) ('DNMT3A', 'Gene', '1788', (82, 88)) ('patients', 'Species', '9606', (26, 34)) ('mutations', 'Var', (89, 98)) ('patients', 'Species', '9606', (114, 122)) 5027 24895580 (1) DNMT3A alterations are involved in epigenetic regulation of gene transcription:aberrant DNA methylation. ('alterations', 'Var', (11, 22)) ('DNA methylation', 'MPA', (92, 107)) ('involved', 'Reg', (27, 35)) ('aberrant', 'Var', (83, 91)) ('DNMT3A', 'Gene', (4, 10)) ('DNMT3A', 'Gene', '1788', (4, 10)) 5028 24895580 FLT3 as well as JAK2V617F abnormity can activate tyrosine kinase and result in aberrant activation of tyrosine kinase signaling. ('tyrosine', 'Enzyme', (49, 57)) ('JAK2', 'Gene', '3717', (16, 20)) ('activation', 'PosReg', (88, 98)) ('JAK2', 'Gene', (16, 20)) ('tyrosine kinase signaling', 'MPA', (102, 127)) ('activate', 'PosReg', (40, 48)) ('FLT3', 'Gene', (0, 4)) ('abnormity', 'Var', (26, 35)) 5029 24895580 (3) NPM1 mutation in exon 12 results in loss of its nuclear localization signal; the altered protein concentrates in the cytoplasm, where it dimerizes to wild-type NPM1, blocking its activity in the nucleus. ('mutation', 'Var', (9, 17)) ('activity in the nucleus', 'MPA', (183, 206)) ('blocking', 'NegReg', (170, 178)) ('loss', 'NegReg', (40, 44)) ('NPM1', 'Gene', (4, 8)) ('nuclear localization signal', 'MPA', (52, 79)) ('NPM1', 'Gene', (164, 168)) ('NPM1', 'Gene', '4869', (164, 168)) ('NPM1', 'Gene', '4869', (4, 8)) 5030 24895580 So burgeoning insight into the role of these genes mutations in the pathogenesis of myeloid malignancies has prompted increased interest in development of novel targeted therapies. ('myeloid malignancies', 'Disease', 'MESH:D009369', (84, 104)) ('myeloid malignancies', 'Disease', (84, 104)) ('mutations', 'Var', (51, 60)) 5061 24124438 G-banding analysis in patient one, AML-M1 showed that all of 20 metaphases cells revealed both numerical and structural abnormalities, including 46, XX, t (9;22)(q34.1;q11) + dmin; 45, XX, t (9;22)(q34.1;q11), -15. ('t (9;22)(q34.1;q11', 'Var', (189, 207)) ('AML-M1', 'Disease', (35, 41)) ('AML-M1', 'Disease', 'MESH:D015470', (35, 41)) ('dmin', 'Chemical', '-', (175, 179)) ('structural abnormalities', 'Disease', (109, 133)) ('patient', 'Species', '9606', (22, 29)) ('structural abnormalities', 'Disease', 'MESH:C566527', (109, 133)) 5073 24124438 Generally, C-myc amplification results in an over expression of the myc protein, which is known to be a critical nuclear transcription factor. ('myc', 'Gene', '4609', (13, 16)) ('myc', 'Gene', '4609', (68, 71)) ('C-myc', 'Gene', '4609', (11, 16)) ('amplification', 'Var', (17, 30)) ('C-myc', 'Gene', (11, 16)) ('myc', 'Gene', (68, 71)) ('myc', 'Gene', (13, 16)) ('over', 'PosReg', (45, 49)) 5075 24124438 Amplification of the ETS1, FL11, SRPR, NFRKB, and KCNJ4 genes located at 11q23-24 distal to MLL was demonstrated in a patient with AML. ('AML', 'Disease', 'MESH:D015470', (131, 134)) ('MLL', 'Gene', (92, 95)) ('Amplification', 'Var', (0, 13)) ('MLL', 'Gene', '4297', (92, 95)) ('ETS1', 'Gene', '2113', (21, 25)) ('ETS1', 'Gene', (21, 25)) ('AML', 'Disease', (131, 134)) ('FL11', 'Gene', (27, 31)) ('SRPR', 'Gene', (33, 37)) ('KCNJ4', 'Gene', '3761', (50, 55)) ('KCNJ4', 'Gene', (50, 55)) ('NFRKB', 'Gene', (39, 44)) ('NFRKB', 'Gene', '4798', (39, 44)) ('SRPR', 'Gene', '6734', (33, 37)) ('patient', 'Species', '9606', (118, 125)) 5086 18841055 Herein we review recent discoveries of genetic and epigenetic alterations in CBF-AML that may represent novel prognostic markers and therapeutic targets and lead to improvement of the still disappointing clinical outcome of these patients. ('genetic', 'Var', (39, 46)) ('CBF-AML', 'Disease', 'MESH:D015470', (77, 84)) ('CBF-AML', 'Disease', (77, 84)) ('AML', 'Phenotype', 'HP:0004808', (81, 84)) ('patients', 'Species', '9606', (230, 238)) ('men', 'Species', '9606', (172, 175)) ('epigenetic alterations', 'Var', (51, 73)) 5087 18841055 Recent findings Several acquired gene mutations and gene-expression and microRNA-expression changes that occur in addition to t(8;21)(q22;q22) and inv(16)(p13q22)/t(16;16)(p13;q22), the cytogenetic hallmarks of CBF-AML, have been recently reported. ('CBF-AML', 'Disease', (211, 218)) ('CBF-AML', 'Disease', 'MESH:D015470', (211, 218)) ('t(8;21)(q22;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (126, 142)) ('inv(16)(p13q22)/t(16;16', 'Var', (147, 170)) ('AML', 'Phenotype', 'HP:0004808', (215, 218)) ('t(8;21)(q22;q22', 'Var', (126, 141)) ('t(16;16)(p13;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (163, 180)) 5088 18841055 Alterations that may represent cooperative events in CBF-AML leukemogenesis include mutations in the KIT, FLT3, JAK2 and RAS genes, haploinsufficiency of the putative tumor suppressor genes TLE1 and TLE4 in t(8;21)-positive patients with del(9q), MN1 overexpression in inv(16) patients, and epigenetic and posttranscriptional silencing of CEBPA. ('JAK2', 'Gene', '3717', (112, 116)) ('KIT', 'Gene', (101, 104)) ('patients', 'Species', '9606', (277, 285)) ('overexpression', 'PosReg', (251, 265)) ('CBF-AML leukemogenesis', 'Disease', 'MESH:D015470', (53, 75)) ('MN1', 'Gene', (247, 250)) ('TLE1', 'Gene', (190, 194)) ('CBF-AML leukemogenesis', 'Disease', (53, 75)) ('JAK2', 'Gene', (112, 116)) ('del(9q', 'Var', (238, 244)) ('AML', 'Phenotype', 'HP:0004808', (57, 60)) ('TLE4', 'Gene', (199, 203)) ('FLT3', 'Gene', (106, 110)) ('TLE4', 'Gene', '7091', (199, 203)) ('mutations', 'Var', (84, 93)) ('haploinsufficiency of the putative tumor', 'Disease', 'MESH:D058495', (132, 172)) ('patients', 'Species', '9606', (224, 232)) ('CEBPA', 'Gene', '1050', (339, 344)) ('TLE1', 'Gene', '7088', (190, 194)) ('haploinsufficiency of the putative tumor', 'Disease', (132, 172)) ('RAS', 'Gene', (121, 124)) ('CEBPA', 'Gene', (339, 344)) ('tumor', 'Phenotype', 'HP:0002664', (167, 172)) 5091 18841055 Core-binding factor (CBF) acute myeloid leukemia (AML) is characterized by the presence of t(8;21)(q22;q22) or inv(16)(p13q22)/t(16;16)(p13;q22). ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (26, 48)) ('inv(16)(p13q22)/t(16;16)(p13;q22', 'Var', (111, 143)) ('CBF', 'Gene', (21, 24)) ('AML', 'Disease', 'MESH:D015470', (50, 53)) ('t(8;21)(q22;q22', 'Var', (91, 106)) ('t(16;16)(p13;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (127, 144)) ('leukemia', 'Phenotype', 'HP:0001909', (40, 48)) ('AML', 'Phenotype', 'HP:0004808', (50, 53)) ('AML', 'Disease', (50, 53)) ('CBF', 'Gene', '10153', (21, 24)) ('t(8;21)(q22;q22)', 'STRUCTURAL_ABNORMALITY', 'None', (91, 107)) ('leukemia', 'Disease', (40, 48)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (32, 48)) ('leukemia', 'Disease', 'MESH:D007938', (40, 48)) 5093 18841055 Likewise, variants of inv(16)/t(16;16) and cryptic rearrangements leading to CBFB-MYH11 have been reported. ('CBFB-MYH11', 'Gene', (77, 87)) ('variants', 'Var', (10, 18)) ('inv(16)/t(16;16', 'Gene', (22, 37)) ('cryptic', 'Gene', '55997', (43, 50)) ('cryptic', 'Gene', (43, 50)) ('men', 'Species', '9606', (60, 63)) 5101 18841055 Thus, for overt leukemia to occur additional genetic, epigenetic alterations, or both are necessary. ('leukemia', 'Disease', 'MESH:D007938', (16, 24)) ('epigenetic alterations', 'Var', (54, 76)) ('leukemia', 'Phenotype', 'HP:0001909', (16, 24)) ('leukemia', 'Disease', (16, 24)) 5102 18841055 This is consistent with a multistep model of leukemogenesis, in which two classes of mutations cooperate: one affecting genes encoding transcription factors capable of impairing hematopoietic differentiation, such as RUNX1-RUNX1T1 and CBFB-MYH11 (class II mutations), and another involving genes such as KIT, FLT3, NRAS or KRAS, whose mutations lead to increased cell proliferation, survival (class I mutations), or both. ('survival', 'CPA', (383, 391)) ('FLT3', 'Gene', (309, 313)) ('hematopoietic differentiation', 'CPA', (178, 207)) ('KRAS', 'Gene', (323, 327)) ('cell proliferation', 'CPA', (363, 381)) ('increased', 'PosReg', (353, 362)) ('CBFB-MYH11', 'Gene', (235, 245)) ('KRAS', 'Gene', '3845', (323, 327)) ('mutations', 'Var', (85, 94)) ('NRAS', 'Gene', (315, 319)) ('NRAS', 'Gene', '4893', (315, 319)) ('impairing', 'NegReg', (168, 177)) 5103 18841055 Secondary chromosome aberrations are detected at diagnosis in approximately 40% of patients with inv(16)/t(16;16), and 70% of t(8;21) patients. ('patients', 'Species', '9606', (83, 91)) ('inv(16)/t(16;16', 'Var', (97, 112)) ('patients', 'Species', '9606', (134, 142)) ('Secondary chromosome aberrations', 'CPA', (0, 32)) 5105 18841055 One recent study suggested that secondary aberrations other than +8, +21 and +22 are more common in inv(16)-positive patients with rare, nontype A CBFB-MYH11 transcripts. ('common', 'Reg', (90, 96)) ('+22', 'Var', (77, 80)) ('+21', 'Var', (69, 72)) ('CBFB-MYH11', 'Gene', (147, 157)) ('patients', 'Species', '9606', (117, 125)) 5106 18841055 Secondary aberrations, such as +22 in inv(16)/t(16;16) patients, and -Y, -X and del(9q) in t(8;21) patients are less frequent in non-CBF-AML signifying their importance for CBF-AML leukemogenesis. ('CBF-AML', 'Disease', 'MESH:D015470', (173, 180)) ('CBF-AML leukemogenesis', 'Disease', (173, 195)) ('AML', 'Phenotype', 'HP:0004808', (177, 180)) ('del(9q', 'Var', (80, 86)) ('CBF-AML', 'Disease', (133, 140)) ('patients', 'Species', '9606', (99, 107)) ('CBF-AML', 'Disease', 'MESH:D015470', (133, 140)) ('AML', 'Phenotype', 'HP:0004808', (137, 140)) ('+22', 'Var', (31, 34)) ('patients', 'Species', '9606', (55, 63)) ('CBF-AML leukemogenesis', 'Disease', 'MESH:D015470', (173, 195)) ('CBF-AML', 'Disease', (173, 180)) 5107 18841055 In contrast, the TLE1 and TLE4 genes have recently been suggested as candidate tumor suppressors residing within the 2.4Mb region at 9q21.32-9q21.33 invariably lost in t(8;21) patients with del(9q). ('patients', 'Species', '9606', (176, 184)) ('tumor', 'Phenotype', 'HP:0002664', (79, 84)) ('tumor', 'Disease', (79, 84)) ('TLE1', 'Gene', '7088', (17, 21)) ('TLE4', 'Gene', (26, 30)) ('del(9q', 'Var', (190, 196)) ('TLE1', 'Gene', (17, 21)) ('tumor', 'Disease', 'MESH:D009369', (79, 84)) ('lost', 'NegReg', (160, 164)) ('TLE4', 'Gene', '7091', (26, 30)) 5109 18841055 Additionally, TLE1 and TLE4 expression was lower in t(8;21) patients with del(9q) compared with normal CD34+ cells or cells from t(8;21) patients without del(9q). ('TLE1', 'Gene', (14, 18)) ('patients', 'Species', '9606', (60, 68)) ('TLE4', 'Gene', (23, 27)) ('expression', 'MPA', (28, 38)) ('TLE1', 'Gene', '7088', (14, 18)) ('patients', 'Species', '9606', (137, 145)) ('CD34', 'Gene', '947', (103, 107)) ('TLE4', 'Gene', '7091', (23, 27)) ('del(9q', 'Var', (74, 80)) ('CD34', 'Gene', (103, 107)) ('lower', 'NegReg', (43, 48)) 5111 18841055 These data suggest that haploinsufficiency of TLE1 and TLE4 may cooperate with RUNX1-RUNX1T1 in the leukemogenesis of t(8;21) patients with del(9q). ('cooperate', 'Reg', (64, 73)) ('TLE4', 'Gene', '7091', (55, 59)) ('haploinsufficiency', 'Disease', 'MESH:D058495', (24, 42)) ('leukemogenesis', 'Disease', (100, 114)) ('TLE1', 'Gene', (46, 50)) ('patients', 'Species', '9606', (126, 134)) ('haploinsufficiency', 'Disease', (24, 42)) ('TLE4', 'Gene', (55, 59)) ('TLE1', 'Gene', '7088', (46, 50)) ('del(9q', 'Var', (140, 146)) 5112 18841055 Genetic events cooperating with CBF-AML-associated gene fusions may also include activating mutations in genes encoding receptor tyrosine kinases (RTKs) or small GTPases. ('fusions', 'Var', (56, 63)) ('CBF-AML', 'Disease', (32, 39)) ('CBF-AML', 'Disease', 'MESH:D015470', (32, 39)) ('tyrosine kinase', 'Gene', (129, 144)) ('AML', 'Phenotype', 'HP:0004808', (36, 39)) ('mutations', 'Var', (92, 101)) ('tyrosine kinase', 'Gene', '7294', (129, 144)) ('activating', 'PosReg', (81, 91)) 5114 18841055 In t(8;21) AML, KIT mutations occur mostly in exon 17 and confer adverse prognosis, whereas the prognostic significance of KIT mutations (both in exon 8 and 17) in inv(16) AML is less well established (Table 2 Table 2). ('AML', 'Phenotype', 'HP:0004808', (172, 175)) ('KIT', 'Gene', (16, 19)) ('AML', 'Disease', 'MESH:D015470', (172, 175)) ('AML', 'Disease', 'MESH:D015470', (11, 14)) ('AML', 'Phenotype', 'HP:0004808', (11, 14)) ('mutations', 'Var', (20, 29)) ('AML', 'Disease', (172, 175)) ('AML', 'Disease', (11, 14)) 5119 18841055 An internal tandem duplication (ITD) within the juxtamembrane domain of the FLT3 gene, that encodes an RTK protein, is relatively infrequent in CBF-AML, being detected in 2-9% of t(8;21) patients and 0-7% of inv(16) patients. ('patients', 'Species', '9606', (187, 195)) ('internal tandem duplication', 'Var', (3, 30)) ('detected', 'Reg', (159, 167)) ('AML', 'Phenotype', 'HP:0004808', (148, 151)) ('FLT3', 'Gene', (76, 80)) ('patients', 'Species', '9606', (216, 224)) ('CBF-AML', 'Disease', (144, 151)) ('CBF-AML', 'Disease', 'MESH:D015470', (144, 151)) 5121 18841055 The second type of FLT3 mutations, point mutations within the activation loop of the tyrosine kinase domain (TKD), are more frequent in inv(16) patients (6-24%), but occur in only 2-7% of t(8;21) patients. ('tyrosine kinase', 'Gene', (85, 100)) ('FLT3', 'Gene', (19, 23)) ('patients', 'Species', '9606', (196, 204)) ('patients', 'Species', '9606', (144, 152)) ('point mutations', 'Var', (35, 50)) ('tyrosine kinase', 'Gene', '7294', (85, 100)) 5123 18841055 Given their relatively high incidence in inv(16) patients, large studies analyzing prognostic impact of FLT3 mutations, which constitute potential therapeutic targets for tyrosine kinase inhibitors, are warranted. ('tyrosine kinase', 'Gene', '7294', (171, 186)) ('FLT3', 'Gene', (104, 108)) ('tyrosine kinase', 'Gene', (171, 186)) ('mutations', 'Var', (109, 118)) ('patients', 'Species', '9606', (49, 57)) 5124 18841055 The most recent addition to the list of potentially leukemogenic mutations in CBF-AML is the activating, gain-of-function JAK2V617F mutation, previously reported in most patients with chronic myeloproliferative disorders. ('patients', 'Species', '9606', (170, 178)) ('JAK2', 'Gene', '3717', (122, 126)) ('gain-of-function', 'PosReg', (105, 121)) ('myeloproliferative disorders', 'Disease', (192, 220)) ('leukemogenic', 'Disease', (52, 64)) ('CBF-AML', 'Disease', (78, 85)) ('CBF-AML', 'Disease', 'MESH:D015470', (78, 85)) ('myeloproliferative disorders', 'Phenotype', 'HP:0005547', (192, 220)) ('JAK2', 'Gene', (122, 126)) ('myeloproliferative disorders', 'Disease', 'MESH:D009196', (192, 220)) ('AML', 'Phenotype', 'HP:0004808', (82, 85)) ('mutations', 'Var', (65, 74)) 5125 18841055 JAK2 mutations are infrequent in CBF-AML, being identified in 4 of 64 (6%) t(8;21) patients and none of 99 inv(16)/t(16;16) patients in one study, and in 5 of 138 (3.6%) CBF-AML patients [2 had t(8;21) and 3 inv(16)/t(16;16)] in another. ('AML', 'Phenotype', 'HP:0004808', (174, 177)) ('CBF-AML', 'Disease', 'MESH:D015470', (33, 40)) ('CBF-AML', 'Disease', (170, 177)) ('CBF-AML', 'Disease', 'MESH:D015470', (170, 177)) ('CBF-AML', 'Disease', (33, 40)) ('JAK2', 'Gene', '3717', (0, 4)) ('patients', 'Species', '9606', (178, 186)) ('mutations', 'Var', (5, 14)) ('AML', 'Phenotype', 'HP:0004808', (37, 40)) ('JAK2', 'Gene', (0, 4)) ('patients', 'Species', '9606', (83, 91)) ('patients', 'Species', '9606', (124, 132)) 5126 18841055 found JAK2 mutations in two of three t(8;21)-positive patients with therapy-related AML, but in none of 20 t(8;21)-positive de novo AML patients, and suggested JAK2 mutations might be associated with therapy-related CBF-AML after treatment with anthracyclines and topoisomerase inhibitors. ('mutations', 'Var', (11, 20)) ('anthracyclines', 'Chemical', 'MESH:D018943', (245, 259)) ('patients', 'Species', '9606', (54, 62)) ('JAK2', 'Gene', (160, 164)) ('JAK2', 'Gene', '3717', (6, 10)) ('CBF-AML', 'Disease', 'MESH:D015470', (216, 223)) ('mutations', 'Var', (165, 174)) ('associated with', 'Reg', (184, 199)) ('JAK2', 'Gene', (6, 10)) ('CBF-AML', 'Disease', (216, 223)) ('AML', 'Disease', 'MESH:D015470', (220, 223)) ('AML', 'Phenotype', 'HP:0004808', (220, 223)) ('AML', 'Disease', 'MESH:D015470', (132, 135)) ('AML', 'Disease', (220, 223)) ('AML', 'Disease', (132, 135)) ('AML', 'Disease', 'MESH:D015470', (84, 87)) ('AML', 'Phenotype', 'HP:0004808', (132, 135)) ('AML', 'Phenotype', 'HP:0004808', (84, 87)) ('AML', 'Disease', (84, 87)) ('patients', 'Species', '9606', (136, 144)) ('men', 'Species', '9606', (235, 238)) ('JAK2', 'Gene', '3717', (160, 164)) 5129 18841055 Importantly, disease-free survival (DFS) of CBF-AML patients with JAK2 mutations was significantly worse than DFS of patients without these mutations in one study. ('disease-free survival', 'CPA', (13, 34)) ('AML', 'Phenotype', 'HP:0004808', (48, 51)) ('CBF-AML', 'Disease', 'MESH:D015470', (44, 51)) ('CBF-AML', 'Disease', (44, 51)) ('JAK2', 'Gene', '3717', (66, 70)) ('mutations', 'Var', (71, 80)) ('patients', 'Species', '9606', (52, 60)) ('patients', 'Species', '9606', (117, 125)) ('JAK2', 'Gene', (66, 70)) ('worse', 'NegReg', (99, 104)) 5130 18841055 JAK2 mutations are potential targets for small-molecule inhibitors of JAK2 kinases. ('JAK2', 'Gene', (70, 74)) ('JAK2', 'Gene', '3717', (0, 4)) ('mutations', 'Var', (5, 14)) ('JAK2', 'Gene', (0, 4)) ('JAK2', 'Gene', '3717', (70, 74)) 5132 18841055 Mutated NRAS may become a target for molecularly targeted therapy. ('Mutated', 'Var', (0, 7)) ('NRAS', 'Gene', '4893', (8, 12)) ('NRAS', 'Gene', (8, 12)) 5139 18841055 Initial microarray gene-expression profiling (GEP) studies identified signatures allowing reliable separation of patients with inv(16)/t(16;16) from those with t(8;21) and from other cytogenetic, molecular genetic AML subgroups, or both. ('AML', 'Disease', 'MESH:D015470', (214, 217)) ('patients', 'Species', '9606', (113, 121)) ('AML', 'Phenotype', 'HP:0004808', (214, 217)) ('inv', 'Var', (127, 130)) ('AML', 'Disease', (214, 217)) 5147 18841055 As aberrant expression of many microRNAs has been demonstrated in several types of cancer, including AML, the role of microRNAs in CBF-AML has only begun to be investigated. ('AML', 'Disease', 'MESH:D015470', (101, 104)) ('AML', 'Disease', 'MESH:D015470', (135, 138)) ('aberrant', 'Var', (3, 11)) ('cancer', 'Phenotype', 'HP:0002664', (83, 89)) ('AML', 'Phenotype', 'HP:0004808', (135, 138)) ('AML', 'Phenotype', 'HP:0004808', (101, 104)) ('AML', 'Disease', (101, 104)) ('demonstrated', 'Reg', (50, 62)) ('AML', 'Disease', (135, 138)) ('CBF-AML', 'Disease', (131, 138)) ('cancer', 'Disease', 'MESH:D009369', (83, 89)) ('expression', 'MPA', (12, 22)) ('cancer', 'Disease', (83, 89)) ('CBF-AML', 'Disease', 'MESH:D015470', (131, 138)) 5155 18841055 Epigenetic alterations silencing gene function without changing the DNA coding sequence are intrinsic to CBF-AML leukemogenesis. ('Epigenetic alterations', 'Var', (0, 22)) ('silencing', 'NegReg', (23, 32)) ('CBF-AML leukemogenesis', 'Disease', (105, 127)) ('AML', 'Phenotype', 'HP:0004808', (109, 112)) ('CBF-AML leukemogenesis', 'Disease', 'MESH:D015470', (105, 127)) 5156 18841055 In t(8;21)-AML, the p300-binding site is replaced by the fusion partner RUNX1T1, which recruits histone deacetylases (HDACs) and induces transcriptional repression of the target genes. ('p300-binding', 'Var', (20, 32)) ('RUNX1T1', 'Gene', (72, 79)) ('transcriptional repression', 'MPA', (137, 163)) ('AML', 'Disease', 'MESH:D015470', (11, 14)) ('AML', 'Phenotype', 'HP:0004808', (11, 14)) ('induces', 'PosReg', (129, 136)) ('recruits', 'PosReg', (87, 95)) ('AML', 'Disease', (11, 14)) 5158 18841055 In addition to recruiting HDACs, RUNX1-RUNX1T1 also recruits DNA-methyltransferase 1 (DNMT). ('DNMT', 'Gene', '1786', (86, 90)) ('DNMT', 'Gene', (86, 90)) ('DNA-methyltransferase 1', 'Gene', (61, 84)) ('DNA-methyltransferase 1', 'Gene', '1786', (61, 84)) ('RUNX1-RUNX1T1', 'Var', (33, 46)) 5160 18841055 Therapeutic targeting of epigenetic alterations in CBF-AML is now being pursued in the clinic. ('AML', 'Phenotype', 'HP:0004808', (55, 58)) ('CBF-AML', 'Disease', (51, 58)) ('epigenetic alterations', 'Var', (25, 47)) ('CBF-AML', 'Disease', 'MESH:D015470', (51, 58)) 5161 18841055 Epigenetic alterations may also represent secondary 'hits' contributing to CBF-AML leukemogenesis. ('Epigenetic alterations', 'Var', (0, 22)) ('CBF-AML leukemogenesis', 'Disease', 'MESH:D015470', (75, 97)) ('CBF-AML leukemogenesis', 'Disease', (75, 97)) ('AML', 'Phenotype', 'HP:0004808', (79, 82)) 5163 18841055 Downregulation of CEBPA in inv(16)-AML may also be achieved through posttranscriptional interaction of CEBPA mRNA with the RNA-binding protein calreticulin or by microRNA-124a, which is also regulated by promoter methylation. ('CEBPA', 'Gene', '1050', (103, 108)) ('AML', 'Disease', (35, 38)) ('calreticulin', 'Gene', (143, 155)) ('Downregulation', 'NegReg', (0, 14)) ('calreticulin', 'Gene', '811', (143, 155)) ('AML', 'Disease', 'MESH:D015470', (35, 38)) ('microRNA-124a', 'Var', (162, 175)) ('interaction', 'Interaction', (88, 99)) ('CEBPA', 'Gene', (18, 23)) ('CEBPA', 'Gene', (103, 108)) ('CEBPA', 'Gene', '1050', (18, 23)) ('AML', 'Phenotype', 'HP:0004808', (35, 38)) 5183 18841055 Recent discoveries of gene mutations, changes in gene-expression and microRNA-expression levels and epigenetic events have increased our understanding of CBF-AML leukemogenesis. ('mutations', 'Var', (27, 36)) ('increased', 'PosReg', (123, 132)) ('microRNA-expression levels', 'MPA', (69, 95)) ('CBF-AML leukemogenesis', 'Disease', 'MESH:D015470', (154, 176)) ('CBF-AML leukemogenesis', 'Disease', (154, 176)) ('changes', 'Reg', (38, 45)) ('AML', 'Phenotype', 'HP:0004808', (158, 161)) ('gene-expression', 'MPA', (49, 64)) 5188 23533770 This paper will discuss the most well studied epigenetic modifications and how these are linked to cancer, will give a brief overview of the clinical use of epigenetics as biomarkers, and will focus in more detail on epigenetic drugs and their use in solid and blood cancers. ('cancer', 'Disease', (267, 273)) ('cancer', 'Disease', 'MESH:D009369', (267, 273)) ('cancer', 'Disease', 'MESH:D009369', (99, 105)) ('cancers', 'Phenotype', 'HP:0002664', (267, 274)) ('epigenetic', 'Var', (46, 56)) ('blood cancers', 'Disease', 'MESH:D009369', (261, 274)) ('cancer', 'Disease', (99, 105)) ('blood cancers', 'Disease', (261, 274)) ('cancer', 'Phenotype', 'HP:0002664', (267, 273)) ('blood cancers', 'Phenotype', 'HP:0001909', (261, 274)) ('cancer', 'Phenotype', 'HP:0002664', (99, 105)) ('linked', 'Reg', (89, 95)) 5192 23533770 Traditionally, the development of cancer is thought to be largely due to the accumulation of genetic defects such as mutations, amplifications, deletions, and translocations affecting the cancer cell machinery and providing the cancer cell with the advantage to survive and metastasize. ('cancer', 'Phenotype', 'HP:0002664', (34, 40)) ('mutations', 'Var', (117, 126)) ('cancer', 'Phenotype', 'HP:0002664', (188, 194)) ('cancer', 'Disease', (228, 234)) ('translocations', 'Var', (159, 173)) ('deletions', 'Var', (144, 153)) ('cancer', 'Disease', 'MESH:D009369', (228, 234)) ('affecting', 'Reg', (174, 183)) ('cancer', 'Disease', 'MESH:D009369', (34, 40)) ('cancer', 'Disease', 'MESH:D009369', (188, 194)) ('cancer', 'Phenotype', 'HP:0002664', (228, 234)) ('cancer', 'Disease', (34, 40)) ('amplifications', 'Var', (128, 142)) ('cancer', 'Disease', (188, 194)) ('genetic defects', 'Disease', 'MESH:D030342', (93, 108)) ('survive', 'CPA', (262, 269)) ('genetic defects', 'Disease', (93, 108)) 5196 23533770 This paper will discuss the most well studied epigenetic modifications and how these are linked to cancer, give a brief overview of the clinical use of epigenetics as biomarkers, and focus in more detail on epigenetic drugs and their use in solid and blood cancers. ('cancer', 'Phenotype', 'HP:0002664', (257, 263)) ('blood cancers', 'Phenotype', 'HP:0001909', (251, 264)) ('cancer', 'Disease', 'MESH:D009369', (99, 105)) ('blood cancers', 'Disease', 'MESH:D009369', (251, 264)) ('blood cancers', 'Disease', (251, 264)) ('cancer', 'Disease', (99, 105)) ('cancer', 'Disease', 'MESH:D009369', (257, 263)) ('cancers', 'Phenotype', 'HP:0002664', (257, 264)) ('epigenetic modifications', 'Var', (46, 70)) ('cancer', 'Disease', (257, 263)) ('cancer', 'Phenotype', 'HP:0002664', (99, 105)) ('linked', 'Reg', (89, 95)) 5207 23533770 Histone acetylation can also promote transcription by providing binding sites to proteins that are involved in gene activation, such as the bromodomain-containing family of proteins. ('binding', 'Interaction', (64, 71)) ('promote', 'PosReg', (29, 36)) ('acetyl', 'Chemical', 'MESH:D003545', (8, 14)) ('acetylation', 'Var', (8, 19)) ('Histone acetylation', 'Var', (0, 19)) ('proteins', 'Protein', (81, 89)) ('transcription', 'MPA', (37, 50)) 5209 23533770 Lysine residues can be monomethylated, dimethylated, or trimethylated whereas arginine residues can be mono- or dimethylated. ('arginine', 'Chemical', 'MESH:D001120', (78, 86)) ('Lysine', 'Protein', (0, 6)) ('trimethylated', 'Var', (56, 69)) ('Lysine', 'Chemical', 'MESH:D008239', (0, 6)) ('dimethylated', 'Var', (39, 51)) 5213 23533770 Arginine methylation of histone proteins has recently been shown to antagonize other histone marks, further increasing the histone code complexity. ('histone code complexity', 'MPA', (123, 146)) ('Arginine', 'Chemical', 'MESH:D001120', (0, 8)) ('increasing', 'PosReg', (108, 118)) ('histone proteins', 'Protein', (24, 40)) ('antagonize', 'NegReg', (68, 78)) ('Arginine methylation', 'Var', (0, 20)) 5214 23533770 In cancer, a global process of genomic hypomethylation occurs mostly at DNA-repetitive regions which results in activation of genes with growth and tumour promoting functions and loss of genome stability and imprinting. ('tumour', 'Disease', (148, 154)) ('genome stability', 'CPA', (187, 203)) ('activation', 'PosReg', (112, 122)) ('tumour', 'Disease', 'MESH:D009369', (148, 154)) ('genes', 'Gene', (126, 131)) ('cancer', 'Disease', 'MESH:D009369', (3, 9)) ('tumour', 'Phenotype', 'HP:0002664', (148, 154)) ('cancer', 'Disease', (3, 9)) ('loss', 'NegReg', (179, 183)) ('hypomethylation', 'Var', (39, 54)) ('cancer', 'Phenotype', 'HP:0002664', (3, 9)) 5217 23533770 Histone modifiers have been shown to be targets of aberrations and/or mutations in cancer such as mutated deacetylases, and amplified histone methyltransferases and demethylases. ('acetyl', 'Chemical', 'MESH:D003545', (108, 114)) ('cancer', 'Disease', 'MESH:D009369', (83, 89)) ('cancer', 'Phenotype', 'HP:0002664', (83, 89)) ('demethylases', 'Enzyme', (165, 177)) ('amplified', 'PosReg', (124, 133)) ('histone methyltransferases', 'Enzyme', (134, 160)) ('mutated', 'Var', (98, 105)) ('cancer', 'Disease', (83, 89)) ('deacetylases', 'Enzyme', (106, 118)) 5219 23533770 Somatic DNMT3A mutations have been described in approximately 20% of acute myeloid leukemia (AML) patients, especially in those with an intermediate risk cytogenetic profile and although they did not affect the 5-methylcytosine content they were associated with poor clinical outcome. ('DNMT3A', 'Gene', (8, 14)) ('DNMT3A', 'Gene', '1788', (8, 14)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (69, 91)) ('AML', 'Phenotype', 'HP:0004808', (93, 96)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (75, 91)) ('AML', 'Disease', (93, 96)) ('described', 'Reg', (35, 44)) ('mutations', 'Var', (15, 24)) ('5-methylcytosine', 'Chemical', 'MESH:D044503', (211, 227)) ('leukemia', 'Phenotype', 'HP:0001909', (83, 91)) ('patients', 'Species', '9606', (98, 106)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (69, 91)) ('associated', 'Reg', (246, 256)) ('acute myeloid leukemia', 'Disease', (69, 91)) ('AML', 'Disease', 'MESH:D015470', (93, 96)) 5220 23533770 How the lack of effect of DNMT3A mutations on 5-methylcytosine content is linked to an otherwise poor clinical outcome is not yet fully understood. ('DNMT3A', 'Gene', (26, 32)) ('DNMT3A', 'Gene', '1788', (26, 32)) ('mutations', 'Var', (33, 42)) ('5-methylcytosine', 'Chemical', 'MESH:D044503', (46, 62)) ('5-methylcytosine content', 'MPA', (46, 70)) 5221 23533770 It has been suggested that the R882 DNMT3A mutations alter functions of DNMT3A such as its ability to bind other proteins involved in transcriptional regulation and localization to chromatin regions containing methylated DNA. ('DNMT3A', 'Gene', (36, 42)) ('functions', 'MPA', (59, 68)) ('DNMT3A', 'Gene', '1788', (36, 42)) ('localization', 'MPA', (165, 177)) ('R882', 'Var', (31, 35)) ('mutations', 'Var', (43, 52)) ('bind', 'Interaction', (102, 106)) ('DNMT3A', 'Gene', (72, 78)) ('DNMT3A', 'Gene', '1788', (72, 78)) ('ability', 'MPA', (91, 98)) 5222 23533770 Loss-of-function TET2 mutations were also identified in myeloid neoplasms in 20-30% and have been associated with both good and bad prognoses. ('Loss-of-function', 'NegReg', (0, 16)) ('myeloid neoplasms', 'Phenotype', 'HP:0012324', (56, 73)) ('TET2', 'Gene', '54790', (17, 21)) ('myeloid neoplasms', 'Disease', (56, 73)) ('neoplasms', 'Phenotype', 'HP:0002664', (64, 73)) ('mutations', 'Var', (22, 31)) ('myeloid neoplasms', 'Disease', 'MESH:D007951', (56, 73)) ('TET2', 'Gene', (17, 21)) 5223 23533770 Genome sequencing has also revealed the presence of metabolic mutations in patients with myelodysplastic syndromes (MDS) and AML related to the isocitrate dehydrogenase (IDH) 1 and IDH2 genes. ('AML', 'Disease', 'MESH:D015470', (125, 128)) ('AML', 'Disease', (125, 128)) ('patients', 'Species', '9606', (75, 83)) ('IDH2', 'Gene', (181, 185)) ('AML', 'Phenotype', 'HP:0004808', (125, 128)) ('MDS', 'Phenotype', 'HP:0002863', (116, 119)) ('myelodysplastic syndromes', 'Phenotype', 'HP:0002863', (89, 114)) ('myelodysplastic syndromes', 'Disease', (89, 114)) ('myelodysplastic syndromes', 'Disease', 'MESH:D009190', (89, 114)) ('isocitrate dehydrogenase (IDH) 1', 'Gene', '3417', (144, 176)) ('MDS', 'Disease', (116, 119)) ('MDS', 'Disease', 'MESH:D009190', (116, 119)) ('mutations', 'Var', (62, 71)) ('IDH2', 'Gene', '3418', (181, 185)) 5224 23533770 These mutations have been reported in approximately 30% of patients with normal karyotype AML and have been linked to the disruption of various processes such as bone marrow microenvironment changes and impaired differentiation suggesting a proleukemogenic effect. ('reported', 'Reg', (26, 34)) ('impaired differentiation', 'CPA', (203, 227)) ('AML', 'Phenotype', 'HP:0004808', (90, 93)) ('AML', 'Disease', (90, 93)) ('linked', 'Reg', (108, 114)) ('patients', 'Species', '9606', (59, 67)) ('AML', 'Disease', 'MESH:D015470', (90, 93)) ('mutations', 'Var', (6, 15)) 5225 23533770 In an AML cohort, IDH1 and IDH2 mutations were mutually exclusive with TET2 mutations while they shared the similar epigenetic defects with the TET2 mutants. ('mutations', 'Var', (76, 85)) ('TET2', 'Gene', '54790', (71, 75)) ('AML', 'Phenotype', 'HP:0004808', (6, 9)) ('IDH1', 'Gene', '3417', (18, 22)) ('genetic defects', 'Disease', 'MESH:D030342', (119, 134)) ('genetic defects', 'Disease', (119, 134)) ('TET2', 'Gene', '54790', (144, 148)) ('mutations', 'Var', (32, 41)) ('IDH2', 'Gene', (27, 31)) ('AML', 'Disease', 'MESH:D015470', (6, 9)) ('TET2', 'Gene', (71, 75)) ('IDH1', 'Gene', (18, 22)) ('IDH2', 'Gene', '3418', (27, 31)) ('TET2', 'Gene', (144, 148)) ('AML', 'Disease', (6, 9)) 5226 23533770 Epigenetic profiling revealed that AML patients with IDH1/2 mutations displayed global hypermethylation and a specific hypermethylation signature. ('AML', 'Disease', (35, 38)) ('patients', 'Species', '9606', (39, 47)) ('IDH1/2', 'Gene', (53, 59)) ('AML', 'Disease', 'MESH:D015470', (35, 38)) ('mutations', 'Var', (60, 69)) ('IDH1/2', 'Gene', '3417;3418', (53, 59)) ('AML', 'Phenotype', 'HP:0004808', (35, 38)) 5227 23533770 MLL is another epigenetic modifier that is commonly mutated in acute leukemias and mainly due to translocations. ('leukemias', 'Disease', (69, 78)) ('acute leukemias', 'Phenotype', 'HP:0002488', (63, 78)) ('translocations', 'Var', (97, 111)) ('MLL', 'Gene', '4297', (0, 3)) ('leukemia', 'Phenotype', 'HP:0001909', (69, 77)) ('leukemias', 'Disease', 'MESH:D007938', (69, 78)) ('MLL', 'Gene', (0, 3)) ('leukemias', 'Phenotype', 'HP:0001909', (69, 78)) 5230 23533770 As MLL is a H3 K4 methyltransferase, translocations that replace the methyltransferase domain affect its function and have been linked with leukaemic transformation. ('MLL', 'Gene', (3, 6)) ('affect', 'Reg', (94, 100)) ('MLL', 'Gene', '4297', (3, 6)) ('translocations', 'Var', (37, 51)) ('leukaemic transformation', 'Disease', (140, 164)) ('function', 'MPA', (105, 113)) ('linked with', 'Reg', (128, 139)) 5231 23533770 Mutations affecting the Polycomb repressive complex (PRC) components, such as EZH2, can also affect histone modifications and have recently been reported. ('histone modifications', 'MPA', (100, 121)) ('affect', 'Reg', (93, 99)) ('PRC', 'Gene', (53, 56)) ('Mutations', 'Var', (0, 9)) ('EZH2', 'Gene', (78, 82)) ('EZH2', 'Gene', '2146', (78, 82)) 5234 23533770 Activating mutations of EZH2 have been reported in B-cell lymphomas whereas missense, nonsense, and frameshift mutations have been reported in various myeloid malignancies. ('frameshift mutations', 'Var', (100, 120)) ('lymphomas', 'Disease', (58, 67)) ('myeloid malignancies', 'Disease', (151, 171)) ('Activating', 'PosReg', (0, 10)) ('lymphomas', 'Disease', 'MESH:D008223', (58, 67)) ('EZH2', 'Gene', '2146', (24, 28)) ('lymphomas', 'Phenotype', 'HP:0002665', (58, 67)) ('EZH2', 'Gene', (24, 28)) ('reported', 'Reg', (131, 139)) ('missense', 'Var', (76, 84)) ('B-cell lymphomas', 'Phenotype', 'HP:0012191', (51, 67)) ('reported', 'Reg', (39, 47)) ('cell lymphoma', 'Phenotype', 'HP:0012191', (53, 66)) ('myeloid malignancies', 'Disease', 'MESH:D009369', (151, 171)) ('lymphoma', 'Phenotype', 'HP:0002665', (58, 66)) ('nonsense', 'Var', (86, 94)) 5235 23533770 In AML, 3 cases so far have been described to carry EZH2 mutations. ('AML', 'Disease', 'MESH:D015470', (3, 6)) ('EZH2', 'Gene', '2146', (52, 56)) ('EZH2', 'Gene', (52, 56)) ('mutations', 'Var', (57, 66)) ('AML', 'Phenotype', 'HP:0004808', (3, 6)) ('AML', 'Disease', (3, 6)) 5236 23533770 Second, in the majority of cases methylation in CpG is acquired during malignant transformation and is therefore specific to neoplasia. ('CpG', 'Gene', (48, 51)) ('methylation', 'Var', (33, 44)) ('neoplasia', 'Disease', 'MESH:D009369', (125, 134)) ('neoplasia', 'Phenotype', 'HP:0002664', (125, 134)) ('neoplasia', 'Disease', (125, 134)) 5240 23533770 In multiple myeloma, methylation of the VHL promoter has been shown to correlate with bone disease and methylation of the bcl-2 interacting killer (BIK) promoter has been shown to predict relapsed/refractory disease, while methylated FHIT has been shown to be an independent adverse prognostic factor. ('BIK', 'Gene', '638', (148, 151)) ('bone disease', 'Disease', (86, 98)) ('correlate', 'Reg', (71, 80)) ('BIK', 'Gene', (148, 151)) ('predict', 'Reg', (180, 187)) ('VHL', 'Gene', '7428', (40, 43)) ('methylated', 'Var', (223, 233)) ('relapsed/refractory disease', 'Disease', (188, 215)) ('methylation', 'Var', (103, 114)) ('bone disease', 'Disease', 'MESH:D001847', (86, 98)) ('multiple myeloma', 'Phenotype', 'HP:0006775', (3, 19)) ('bcl-2 interacting killer', 'Gene', '638', (122, 146)) ('methylation', 'Var', (21, 32)) ('FHIT', 'Gene', (234, 238)) ('multiple myeloma', 'Disease', 'MESH:D009101', (3, 19)) ('FHIT', 'Gene', '2272', (234, 238)) ('multiple myeloma', 'Disease', (3, 19)) ('VHL', 'Gene', (40, 43)) ('bcl-2 interacting killer', 'Gene', (122, 146)) 5242 23533770 Quantitative pyrosequencing in a large cohort showed that patients with higher levels of methylation for these genes had shorter median overall and progressive-free survival (PFS) independent of age, sex, and the International Prognostic Scoring System (IPSS). ('patients', 'Species', '9606', (58, 66)) ('progressive-free survival', 'CPA', (148, 173)) ('shorter', 'NegReg', (121, 128)) ('methylation', 'Var', (89, 100)) 5245 23533770 Of note, methylation-dependent silencing of the methyl transferase MGMT in glioblastoma multiforme confers sensitivity to the alkylating agent temozolomide but as with many such candidate biomarkers, clinical application to inform patient management is not yet routine. ('temozolomide', 'Chemical', 'MESH:D000077204', (143, 155)) ('glioblastoma multiforme', 'Disease', (75, 98)) ('patient', 'Species', '9606', (231, 238)) ('MGMT', 'Gene', '4255', (67, 71)) ('glioblastoma', 'Phenotype', 'HP:0012174', (75, 87)) ('MGMT', 'Gene', (67, 71)) ('glioblastoma multiforme', 'Disease', 'MESH:D005909', (75, 98)) ('sensitivity to the alkylating agent temozolomide', 'MPA', (107, 155)) ('methylation-dependent', 'Var', (9, 30)) ('silencing', 'NegReg', (31, 40)) 5246 23533770 The list of genes reported to be methylated in haematological neoplasms is extensive, and although several have been linked to clinical parameters and have been associated with survival or response to treatment, none of these markers has been used so far in the clinic to guide diagnosis or treatment, as opposed to gene mutations such as NPM1 and FLT3 that are now widely used to risk classify AML patients. ('AML', 'Phenotype', 'HP:0004808', (395, 398)) ('haematological neoplasms', 'Disease', 'MESH:D019337', (47, 71)) ('linked', 'Reg', (117, 123)) ('FLT3', 'Gene', '2322', (348, 352)) ('associated', 'Reg', (161, 171)) ('haematological neoplasms', 'Phenotype', 'HP:0004377', (47, 71)) ('patients', 'Species', '9606', (399, 407)) ('FLT3', 'Gene', (348, 352)) ('NPM1', 'Gene', (339, 343)) ('AML', 'Disease', 'MESH:D015470', (395, 398)) ('haematological neoplasms', 'Disease', (47, 71)) ('neoplasms', 'Phenotype', 'HP:0002664', (62, 71)) ('methylated', 'Var', (33, 43)) ('AML', 'Disease', (395, 398)) ('NPM1', 'Gene', '4869', (339, 343)) 5253 23533770 Both are approved for use in the myelodysplastic syndromes and low-blast count AML and have improved the survival of patients with these diseases. ('AML', 'Disease', (79, 82)) ('myelodysplastic syndromes', 'Phenotype', 'HP:0002863', (33, 58)) ('myelodysplastic syndromes', 'Disease', 'MESH:D009190', (33, 58)) ('AML', 'Phenotype', 'HP:0004808', (79, 82)) ('low-blast count', 'Var', (63, 78)) ('myelodysplastic syndromes', 'Disease', (33, 58)) ('AML', 'Disease', 'MESH:D015470', (79, 82)) ('patients', 'Species', '9606', (117, 125)) ('improved', 'PosReg', (92, 100)) 5255 23533770 A phase 1 study of decitabine with interleukin-2 in melanoma and renal cell carcinoma showed that decitabine caused grade 4 neutropenia in most patients. ('interleukin-2', 'Gene', '3558', (35, 48)) ('melanoma', 'Phenotype', 'HP:0002861', (52, 60)) ('neutropenia', 'Disease', (124, 135)) ('carcinoma', 'Phenotype', 'HP:0030731', (76, 85)) ('interleukin-2', 'Gene', (35, 48)) ('patients', 'Species', '9606', (144, 152)) ('melanoma and renal cell carcinoma', 'Disease', 'MESH:C538614', (52, 85)) ('decitabine', 'Chemical', 'MESH:D000077209', (98, 108)) ('neutropenia', 'Phenotype', 'HP:0001875', (124, 135)) ('neutropenia', 'Disease', 'MESH:D009503', (124, 135)) ('decitabine', 'Chemical', 'MESH:D000077209', (19, 29)) ('decitabine', 'Var', (98, 108)) ('renal cell carcinoma', 'Phenotype', 'HP:0005584', (65, 85)) 5261 23533770 A second explanation for these results is that agents such as azacytidine, which cause global hypomethylation, likely reactivate expression of multiple silenced genes including oncogenes and tumour suppressors in different cell types and in different cancers. ('tumour', 'Disease', 'MESH:D009369', (191, 197)) ('azacytidine', 'Chemical', 'MESH:D001374', (62, 73)) ('tumour', 'Disease', (191, 197)) ('reactivate', 'PosReg', (118, 128)) ('expression', 'MPA', (129, 139)) ('cancer', 'Phenotype', 'HP:0002664', (251, 257)) ('cancers', 'Phenotype', 'HP:0002664', (251, 258)) ('cancers', 'Disease', (251, 258)) ('cancers', 'Disease', 'MESH:D009369', (251, 258)) ('tumour', 'Phenotype', 'HP:0002664', (191, 197)) ('azacytidine', 'Var', (62, 73)) 5262 23533770 For example, the oncogene NT5E is overexpressed in aggressive metastatic melanomas, yet transcriptionally silenced by methylation in breast cancer with more favorable prognosis. ('breast cancer', 'Disease', (133, 146)) ('melanoma', 'Phenotype', 'HP:0002861', (73, 81)) ('NT5E', 'Gene', '4907', (26, 30)) ('breast cancer', 'Phenotype', 'HP:0003002', (133, 146)) ('cancer', 'Phenotype', 'HP:0002664', (140, 146)) ('NT5E', 'Gene', (26, 30)) ('aggressive metastatic melanomas', 'Disease', (51, 82)) ('melanomas', 'Phenotype', 'HP:0002861', (73, 82)) ('aggressive metastatic melanomas', 'Disease', 'MESH:D008545', (51, 82)) ('methylation', 'Var', (118, 129)) ('breast cancer', 'Disease', 'MESH:D001943', (133, 146)) ('overexpressed', 'PosReg', (34, 47)) 5279 23533770 Givinostat has been shown to selectively target cells harboring the JAK2 V617F mutation and has been tested in combination with hydroxyurea in patients with polycythemia vera in a phase II study (NCT00928707). ('JAK2', 'Gene', '3717', (68, 72)) ('polycythemia vera', 'Disease', (157, 174)) ('JAK2', 'Gene', (68, 72)) ('V617F', 'SUBSTITUTION', 'None', (73, 78)) ('patients', 'Species', '9606', (143, 151)) ('polycythemia', 'Phenotype', 'HP:0001901', (157, 169)) ('V617F', 'Var', (73, 78)) ('polycythemia vera', 'Disease', 'MESH:D011087', (157, 174)) ('hydroxyurea', 'Chemical', 'MESH:D006918', (128, 139)) 5286 23533770 The recognition that a subset of TSGs are silenced by a combination of CpG hypermethylation and histone hypoacetylation has prompted testing of combinations of the two classes of agents and trials of these are in progress. ('acetyl', 'Chemical', 'MESH:D003545', (108, 114)) ('CpG', 'Protein', (71, 74)) ('histone', 'MPA', (96, 103)) ('TSGs', 'Disease', (33, 37)) ('hypermethylation', 'Var', (75, 91)) ('silenced', 'NegReg', (42, 50)) 5297 23533770 In total 25% of breast cancers have the estrogen receptor-alpha (ER alpha) repressed mainly due to hypermethylation of the ER promoter and do not respond to endocrine therapy, and almost all hormone-sensitive tumors turn to be refractory at some point. ('tumors', 'Disease', (209, 215)) ('tumors', 'Phenotype', 'HP:0002664', (209, 215)) ('estrogen receptor-alpha', 'Gene', (40, 63)) ('tumors', 'Disease', 'MESH:D009369', (209, 215)) ('cancers', 'Phenotype', 'HP:0002664', (23, 30)) ('estrogen receptor-alpha', 'Gene', '2099', (40, 63)) ('ER alpha', 'Gene', '2099', (65, 73)) ('breast cancers', 'Phenotype', 'HP:0003002', (16, 30)) ('hypermethylation', 'Var', (99, 115)) ('ER alpha', 'Gene', (65, 73)) ('cancer', 'Phenotype', 'HP:0002664', (23, 29)) ('breast cancers', 'Disease', 'MESH:D001943', (16, 30)) ('tumor', 'Phenotype', 'HP:0002664', (209, 214)) ('breast cancers', 'Disease', (16, 30)) ('breast cancer', 'Phenotype', 'HP:0003002', (16, 29)) 5304 23533770 Identifying the epigenetically modified genes, which are principally involved in tumor resistance, can be achieved by comparative analysis of diagnostic (pretreatment) biopsy with a second biopsy at disease relapse. ('tumor', 'Disease', 'MESH:D009369', (81, 86)) ('tumor', 'Phenotype', 'HP:0002664', (81, 86)) ('epigenetically modified genes', 'Var', (16, 45)) ('tumor', 'Disease', (81, 86)) 5306 23533770 The ability of the physician to exploit therapeutic opportunities created by epigenetic changes in the cancer cell epigenome may also offer new approaches to cancer management. ('epigenetic changes', 'Var', (77, 95)) ('cancer', 'Phenotype', 'HP:0002664', (103, 109)) ('cancer', 'Disease', 'MESH:D009369', (158, 164)) ('cancer', 'Disease', (158, 164)) ('cancer', 'Disease', (103, 109)) ('cancer', 'Disease', 'MESH:D009369', (103, 109)) ('cancer', 'Phenotype', 'HP:0002664', (158, 164)) 5307 23533770 For example, ASS1, which encodes arginine succinate synthetase, the rate-limiting enzyme in arginine biosynthesis, is silenced by methylation in some cancer types including renal cell carcinoma, hepatocellular carcinoma, malignant melanoma, glioblastoma multiforme (GBM), and platinum-resistant epithelial ovarian cancer. ('silenced', 'NegReg', (118, 126)) ('carcinoma', 'Phenotype', 'HP:0030731', (210, 219)) ('arginine', 'Chemical', 'MESH:D001120', (92, 100)) ('carcinoma', 'Phenotype', 'HP:0030731', (184, 193)) ('melanoma', 'Phenotype', 'HP:0002861', (231, 239)) ('epithelial ovarian cancer', 'Disease', (295, 320)) ('ASS1', 'Gene', (13, 17)) ('ASS1', 'Gene', '445', (13, 17)) ('hepatocellular carcinoma', 'Phenotype', 'HP:0001402', (195, 219)) ('malignant melanoma', 'Disease', (221, 239)) ('cancer', 'Disease', 'MESH:D009369', (314, 320)) ('epithelial ovarian cancer', 'Disease', 'MESH:D000077216', (295, 320)) ('cancer', 'Disease', (150, 156)) ('renal cell carcinoma', 'Disease', 'MESH:C538614', (173, 193)) ('ovarian cancer', 'Phenotype', 'HP:0100615', (306, 320)) ('cancer', 'Phenotype', 'HP:0002664', (150, 156)) ('hepatocellular carcinoma', 'Disease', 'MESH:D006528', (195, 219)) ('methylation', 'Var', (130, 141)) ('arginine', 'Chemical', 'MESH:D001120', (33, 41)) ('malignant melanoma', 'Phenotype', 'HP:0002861', (221, 239)) ('glioblastoma multiforme', 'Disease', (241, 264)) ('malignant melanoma', 'Disease', 'MESH:D008545', (221, 239)) ('glioblastoma', 'Phenotype', 'HP:0012174', (241, 253)) ('glioblastoma multiforme', 'Disease', 'MESH:D005909', (241, 264)) ('renal cell carcinoma', 'Disease', (173, 193)) ('hepatocellular carcinoma', 'Disease', (195, 219)) ('renal cell carcinoma', 'Phenotype', 'HP:0005584', (173, 193)) ('cancer', 'Disease', (314, 320)) ('cancer', 'Disease', 'MESH:D009369', (150, 156)) ('epithelial ovarian cancer', 'Phenotype', 'HP:0025318', (295, 320)) ('cancer', 'Phenotype', 'HP:0002664', (314, 320)) 5311 23533770 What seems to be needed though is a better selection of patients who will benefit from such treatments as well as identification of new druggable targets and compounds such as histone kinases or inhibitors of histone methyltransferases and sirtuins. ('patients', 'Species', '9606', (56, 64)) ('inhibitors', 'Var', (195, 205)) ('sirtuins', 'Enzyme', (240, 248)) 5315 32782381 RUNX1 mutations in blast-phase chronic myeloid leukemia associate with distinct phenotypes, transcriptional profiles, and drug responses Blast-phase chronic myeloid leukemia (BP-CML) is associated with additional chromosomal aberrations, RUNX1 mutations being one of the most common. ('leukemia', 'Phenotype', 'HP:0001909', (47, 55)) ('chronic myeloid leukemia', 'Disease', 'MESH:D015464', (149, 173)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (39, 55)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (39, 55)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (157, 173)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (157, 173)) ('RUNX1', 'Gene', (238, 243)) ('leukemia', 'Phenotype', 'HP:0001909', (165, 173)) ('chromosomal aberrations', 'Phenotype', 'HP:0040012', (213, 236)) ('chronic myeloid leukemia', 'Disease', (31, 55)) ('chronic myeloid leukemia', 'Phenotype', 'HP:0005506', (31, 55)) ('chronic myeloid leukemia', 'Disease', 'MESH:D015464', (31, 55)) ('RUNX1', 'Gene', '861', (238, 243)) ('RUNX1', 'Gene', (0, 5)) ('chronic myeloid leukemia', 'Phenotype', 'HP:0005506', (149, 173)) ('chronic myeloid leukemia', 'Disease', (149, 173)) ('RUNX1', 'Gene', '861', (0, 5)) ('mutations', 'Var', (6, 15)) 5317 32782381 Using whole-exome and RNA sequencing we demonstrate that PHF6 and BCORL1 mutations, IKZF1 deletions, and AID/RAG-mediated rearrangements are enriched in RUNX1mut BP-CML leading to typical mutational signature. ('IKZF1', 'Gene', (84, 89)) ('PHF6', 'Gene', (57, 61)) ('PHF6', 'Gene', '84295', (57, 61)) ('BCORL1', 'Gene', '63035', (66, 72)) ('AID', 'Gene', '57379', (105, 108)) ('AID', 'Gene', (105, 108)) ('IKZF1', 'Gene', '10320', (84, 89)) ('BCORL1', 'Gene', (66, 72)) ('deletions', 'Var', (90, 99)) ('mutational signature', 'MPA', (188, 208)) ('mutations', 'Var', (73, 82)) 5322 32782381 Overall, our study provides insights into the pathogenic role of RUNX1 mutations and highlights personalized targeted therapy and CAR T-cell immunotherapy as potentially promising strategies for treating RUNX1mut BP-CML patients. ('CAR', 'Gene', (130, 133)) ('RUNX1mut', 'Gene', (204, 212)) ('mutations', 'Var', (71, 80)) ('patients', 'Species', '9606', (220, 228)) ('CAR', 'Gene', '653108', (130, 133)) ('RUNX1', 'Gene', (65, 70)) 5324 32782381 Somatic and germline alterations involving RUNX1 gene are commonly encountered in a variety of hematological malignancies. ('hematological malignancies', 'Disease', (95, 121)) ('hematological malignancies', 'Disease', 'MESH:D019337', (95, 121)) ('encountered', 'Reg', (67, 78)) ('RUNX1', 'Gene', (43, 48)) ('hematological malignancies', 'Phenotype', 'HP:0004377', (95, 121)) ('alterations', 'Var', (21, 32)) 5325 32782381 RUNX1 germline mutations are associated with familial platelet disorders (FPD) with predisposition to hematological malignancies. ('predisposition to hematological malignancies', 'Phenotype', 'HP:0001871', (84, 128)) ('hematological malignancies', 'Disease', (102, 128)) ('platelet disorders', 'Phenotype', 'HP:0001872', (54, 72)) ('hematological malignancies', 'Disease', 'MESH:D019337', (102, 128)) ('familial platelet disorders', 'Disease', (45, 72)) ('FPD', 'Disease', 'MESH:C563324', (74, 77)) ('germline mutations', 'Var', (6, 24)) ('hematological malignancies', 'Phenotype', 'HP:0004377', (102, 128)) ('RUNX1', 'Gene', (0, 5)) ('associated', 'Reg', (29, 39)) ('familial platelet disorders', 'Disease', 'MESH:D001791', (45, 72)) ('FPD', 'Disease', (74, 77)) 5327 32782381 These include t(8;21) RUNX1-RUNX1T1 translocation in 15% of AML patients, t(12;21) ETV6-RUNX1 translocation in 25% of BCP-ALL patients, and t(3;21) RUNX1-MECOM in therapy-related MDS/AML patients. ('patients', 'Species', '9606', (187, 195)) ('ETV6', 'Gene', (83, 87)) ('AML', 'Disease', 'MESH:D015470', (60, 63)) ('patients', 'Species', '9606', (64, 72)) ('t(3;21', 'Var', (140, 146)) ('AML', 'Disease', (183, 186)) ('AML', 'Disease', 'MESH:D015470', (183, 186)) ('MECOM', 'Gene', (154, 159)) ('ETV6', 'Gene', '2120', (83, 87)) ('RUNX1T1', 'Gene', (28, 35)) ('AML', 'Disease', (60, 63)) ('patients', 'Species', '9606', (126, 134)) ('MECOM', 'Gene', '2122', (154, 159)) ('RUNX1T1', 'Gene', '862', (28, 35)) ('MDS', 'Disease', (179, 182)) ('MDS', 'Disease', 'MESH:D009190', (179, 182)) 5328 32782381 In the t(12;21) ETV6-RUNX1 ALL, it has been reported that the aberrant RAG recombination activity mediates off-target deletions and is the driver mutagenic mechanism. ('ETV6', 'Gene', '2120', (16, 20)) ('RAG recombination activity', 'MPA', (71, 97)) ('aberrant', 'Var', (62, 70)) ('ETV6', 'Gene', (16, 20)) ('deletions', 'Var', (118, 127)) 5330 32782381 Somatic RUNX1 mutations are frequent among hematological malignancies like AML, ALL, MDS, and MDS/MPN (CMML). ('MDS', 'Disease', 'MESH:D009190', (94, 97)) ('hematological malignancies', 'Disease', (43, 69)) ('hematological malignancies', 'Disease', 'MESH:D019337', (43, 69)) ('RUNX1', 'Gene', (8, 13)) ('MDS', 'Disease', (94, 97)) ('CMML', 'Disease', (103, 107)) ('AML', 'Disease', 'MESH:D015470', (75, 78)) ('AML', 'Disease', (75, 78)) ('hematological malignancies', 'Phenotype', 'HP:0004377', (43, 69)) ('MDS', 'Disease', (85, 88)) ('MDS', 'Disease', 'MESH:D009190', (85, 88)) ('CMML', 'Disease', 'MESH:D054429', (103, 107)) ('frequent', 'Reg', (28, 36)) ('ALL', 'Disease', (80, 83)) ('mutations', 'Var', (14, 23)) 5331 32782381 AML with mutated RUNX1 (RUNX1mut AML) is a provisional entity which accounts for 10% of the newly diagnosed patients and associates with an inferior prognosis. ('AML', 'Disease', 'MESH:D015470', (33, 36)) ('RUNX1mut AML', 'Gene', (24, 36)) ('mutated', 'Var', (9, 16)) ('AML', 'Disease', (33, 36)) ('AML', 'Disease', 'MESH:D015470', (0, 3)) ('RUNX1mut AML', 'Gene', '861', (24, 36)) ('patients', 'Species', '9606', (108, 116)) ('RUNX1', 'Gene', (17, 22)) ('AML', 'Disease', (0, 3)) 5332 32782381 RUNX1 mutations are frequently encountered in AML patients with minimal differentiation (AML-M0), where it demonstrates a unique molecular signature with upregulation of B-lymphoid genes. ('patients', 'Species', '9606', (50, 58)) ('AML', 'Disease', 'MESH:D015470', (89, 92)) ('minimal differentiation', 'Disease', (64, 87)) ('AML', 'Disease', (46, 49)) ('RUNX1', 'Gene', (0, 5)) ('AML', 'Disease', 'MESH:D015470', (46, 49)) ('AML', 'Disease', (89, 92)) ('upregulation', 'PosReg', (154, 166)) ('mutations', 'Var', (6, 15)) ('encountered', 'Reg', (31, 42)) 5333 32782381 Aberrant expression of the lymphoid marker CD19 is frequently observed in t(8;21)-AML, representing an interesting target for immunotherapy. ('Aberrant', 'Var', (0, 8)) ('AML', 'Disease', 'MESH:D015470', (82, 85)) ('CD19', 'Gene', (43, 47)) ('expression', 'MPA', (9, 19)) ('AML', 'Disease', (82, 85)) ('CD19', 'Gene', '930', (43, 47)) ('observed', 'Reg', (62, 70)) 5334 32782381 RUNX1mut AML shows associations with mutations affecting spliceosome (SRSF2 and SF3B1), epigenetic modifiers (ASXL1 and EZH2), and PHF6 and BCOR genes. ('SF3B1', 'Gene', '23451', (80, 85)) ('RUNX1mut AML', 'Gene', (0, 12)) ('EZH2', 'Gene', (120, 124)) ('ASXL1', 'Gene', (110, 115)) ('SRSF2', 'Gene', '6427', (70, 75)) ('BCOR', 'Gene', '54880', (140, 144)) ('PHF6', 'Gene', '84295', (131, 135)) ('spliceosome', 'MPA', (57, 68)) ('mutations', 'Var', (37, 46)) ('RUNX1mut AML', 'Gene', '861', (0, 12)) ('associations', 'Interaction', (19, 31)) ('SF3B1', 'Gene', (80, 85)) ('SRSF2', 'Gene', (70, 75)) ('BCOR', 'Gene', (140, 144)) ('ASXL1', 'Gene', '171023', (110, 115)) ('PHF6', 'Gene', (131, 135)) ('EZH2', 'Gene', '2146', (120, 124)) 5335 32782381 Furthermore, FLT3-ITD and MLL-PTD frequently coexist with RUNX1 mutations, while fusion genes and NPM1 mutations are mutually exclusive with RUNX1 mutations. ('NPM1', 'Gene', (98, 102)) ('FLT3-ITD and MLL-PTD', 'Disease', 'MESH:C537633', (13, 33)) ('NPM1', 'Gene', '4869', (98, 102)) ('mutations', 'Var', (64, 73)) ('RUNX1', 'Gene', (58, 63)) 5336 32782381 RUNX1 mutations have also been found in CML patients and linked to disease progression and inferior treatment responses. ('patients', 'Species', '9606', (44, 52)) ('CML', 'Disease', (40, 43)) ('linked to', 'Reg', (57, 66)) ('RUNX1', 'Gene', (0, 5)) ('found', 'Reg', (31, 36)) ('mutations', 'Var', (6, 15)) 5337 32782381 In our previous study, RUNX1 mutations were identified as recurrent events in BP-CML (3 out of 20 patients). ('BP-CML', 'Disease', (78, 84)) ('RUNX1', 'Gene', (23, 28)) ('mutations', 'Var', (29, 38)) ('patients', 'Species', '9606', (98, 106)) 5338 32782381 In concord, functional studies in mice have shown that RUNX1 mutations can contribute to blast transformation of CML. ('RUNX1', 'Gene', (55, 60)) ('blast transformation', 'CPA', (89, 109)) ('mice', 'Species', '10090', (34, 38)) ('contribute', 'Reg', (75, 85)) ('mutations', 'Var', (61, 70)) 5340 32782381 This approach allowed us to enlighten a novel mutagenesis role of RUNX1 mutations that is coupled with the activation of AID/RAG axis. ('RUNX1', 'Gene', (66, 71)) ('mutations', 'Var', (72, 81)) ('AID', 'Gene', '57379', (121, 124)) ('AID', 'Gene', (121, 124)) 5355 32782381 RUNX1 mutations included three missense mutations (p.R162K, p.R204Q, and p.R107C) and one nonsense mutation (p.K117*), that were all located in the Runt domain (Fig. ('p.R204Q', 'Mutation', 'p.R204Q', (60, 67)) ('p.R107C', 'Var', (73, 80)) ('p.R107C', 'Mutation', 'p.R107C', (73, 80)) ('p.K117*', 'Mutation', 'p.K117*', (109, 116)) ('RUNX1', 'Gene', (0, 5)) ('p.R162K', 'Mutation', 'rs1057519750', (51, 58)) ('p.R204Q', 'Var', (60, 67)) ('p.R162K', 'Var', (51, 58)) 5357 32782381 RUNX1mut BP-CML patients showed a notable population of plasmacytoid dendritic cells (pDCs) in contrast to RUNX1wt group (Supplementary Fig. ('RUNX1mut BP-CML', 'Var', (0, 15)) ('patients', 'Species', '9606', (16, 24)) ('plasmacytoid dendritic cells', 'CPA', (56, 84)) 5359 32782381 We recovered four RUNX1 mutations (p.T176fs, p.L175Q, p.D198G, c.508+2T>C splice donor) in three patients (Supplementary Table 4). ('p.L175Q', 'Var', (45, 52)) ('c.508+2T>C', 'Mutation', 'c.508+2T>C', (63, 73)) ('p.T176fs', 'Mutation', 'p.T176fsX', (35, 43)) ('p.L175Q', 'Mutation', 'p.L175Q', (45, 52)) ('c.508+2T>C splice donor', 'Var', (63, 86)) ('p.D198G', 'Var', (54, 61)) ('p.T176fs', 'Var', (35, 43)) ('RUNX1', 'Gene', (18, 23)) ('p.D198G', 'Mutation', 'p.D198G', (54, 61)) ('patients', 'Species', '9606', (97, 105)) 5360 32782381 Frequent co-occuring mutations in RUNX1mut patients included PHF6 and BCORL1 mutations (Fig. ('BCORL1', 'Gene', '63035', (70, 76)) ('patients', 'Species', '9606', (43, 51)) ('PHF6', 'Gene', (61, 65)) ('PHF6', 'Gene', '84295', (61, 65)) ('mutations', 'Var', (77, 86)) ('BCORL1', 'Gene', (70, 76)) ('RUNX1mut', 'Gene', (34, 42)) 5361 32782381 IKZF1 deletions were more common in RUNX1mut patients, but also found in lymphoid RUNX1wt BP-CML patients (Fig. ('IKZF1', 'Gene', '10320', (0, 5)) ('deletions', 'Var', (6, 15)) ('IKZF1', 'Gene', (0, 5)) ('common', 'Reg', (26, 32)) ('patients', 'Species', '9606', (45, 53)) ('patients', 'Species', '9606', (97, 105)) 5362 32782381 In mut2 patient with longitudinal samples, a RUNX1 mutation (p.R162K) was seen also in diagnosis (CP) sample (variant allele frequency, VAF = 58%), with acquisition of loss of heterozygosity and loss of the wild-type allele prior sampling at BP (VAF = 99%) (Supplementary Fig. ('loss', 'NegReg', (168, 172)) ('RUNX1', 'Gene', (45, 50)) ('heterozygosity', 'MPA', (176, 190)) ('p.R162K', 'Mutation', 'rs1057519750', (61, 68)) ('p.R162K', 'Var', (61, 68)) ('patient', 'Species', '9606', (8, 15)) 5366 32782381 Notably, several AID/RAG components, including RAG1, RAG2, AICDA, and DNTT genes, were overexpressed in RUNX1mut compared with RUNX1wt patients (Fig. ('AID', 'Gene', '57379', (17, 20)) ('RAG2', 'Gene', '5897', (53, 57)) ('patients', 'Species', '9606', (135, 143)) ('RUNX1mut', 'Var', (104, 112)) ('RAG1', 'Gene', '5896', (47, 51)) ('AICDA', 'Gene', '57379', (59, 64)) ('RAG2', 'Gene', (53, 57)) ('overexpressed', 'PosReg', (87, 100)) ('RAG1', 'Gene', (47, 51)) ('DNTT', 'Gene', '1791', (70, 74)) ('AICDA', 'Gene', (59, 64)) ('DNTT', 'Gene', (70, 74)) ('AID', 'Gene', (17, 20)) 5367 32782381 Extension of the analysis to the combined data of 20 BP-CML patients (RUNX1mut; n = 7, RUNX1wt; n = 13) showed no significant differences in the mutational load or structural variants (SV) between RUNX1mut and RUNX1wt patient samples (Supplementary Fig. ('patient', 'Species', '9606', (60, 67)) ('patient', 'Species', '9606', (218, 225)) ('mutational', 'Var', (145, 155)) ('patients', 'Species', '9606', (60, 68)) ('RUNX1mut', 'Var', (197, 205)) ('structural variants', 'Var', (164, 183)) 5374 32782381 Interestingly, we observed RAG-RSS at both ends of an intragenic IKZF1 deletion in a RUNX1mut patient. ('IKZF1', 'Gene', '10320', (65, 70)) ('IKZF1', 'Gene', (65, 70)) ('patient', 'Species', '9606', (94, 101)) ('deletion', 'Var', (71, 79)) 5375 32782381 To gain insights into the transcriptional changes induced by RUNX1 mutations, diagnostic samples from four RUNX1mut and five RUNX1wt patients, were analyzed using RNA-sequencing (Fig. ('RUNX1mut', 'Disease', (107, 115)) ('patients', 'Species', '9606', (133, 141)) ('RUNX1', 'Gene', (61, 66)) ('mutations', 'Var', (67, 76)) 5379 32782381 RUNX1 mutations were associated with dysregulation of several immune regulatory molecules, including CIITA, CD74, B7-H6 (NCR3LG1), CD69, and multiple HLA-DR and TLR molecules, in addition to cytokine receptors (IL2RA, IL21R, and IL12RB2) (Fig. ('IL12RB2', 'Gene', '3595', (229, 236)) ('CD69', 'Gene', '969', (131, 135)) ('IL12RB2', 'Gene', (229, 236)) ('CD69', 'Gene', (131, 135)) ('B7-H6', 'Gene', '374383', (114, 119)) ('NCR3LG1', 'Gene', '374383', (121, 128)) ('B7-H6', 'Gene', (114, 119)) ('CD74', 'Gene', '972', (108, 112)) ('NCR3LG1', 'Gene', (121, 128)) ('mutations', 'Var', (6, 15)) ('IL2RA', 'Gene', (211, 216)) ('associated', 'Reg', (21, 31)) ('IL21R', 'Gene', (218, 223)) ('RUNX1', 'Gene', (0, 5)) ('CIITA', 'Gene', '4261', (101, 106)) ('IL2RA', 'Gene', '3559', (211, 216)) ('CD74', 'Gene', (108, 112)) ('dysregulation', 'MPA', (37, 50)) ('IL21R', 'Gene', '50615', (218, 223)) ('CIITA', 'Gene', (101, 106)) 5386 32782381 Furthermore, GSEA demonstrated similarities of upregulated pathways between RUNX1mut lymphoid-BP and CBF-related AML in contrast to NPM1mut-AML, highlighting RUNX1mut-specific transcriptional signature (Supplementary Fig. ('NPM1', 'Gene', '4869', (132, 136)) ('upregulated', 'PosReg', (47, 58)) ('pathways', 'Pathway', (59, 67)) ('RUNX1mut', 'Var', (76, 84)) ('AML', 'Disease', 'MESH:D015470', (113, 116)) ('AML', 'Disease', 'MESH:D015470', (140, 143)) ('GSEA', 'Chemical', '-', (13, 17)) ('AML', 'Disease', (113, 116)) ('AML', 'Disease', (140, 143)) ('NPM1', 'Gene', (132, 136)) 5391 32782381 Interestingly, cells from a patient with nonsense RUNX1 mutation demonstrated enhanced sensitivity to glucocorticoids and mTOR inhibitors and reduced sensitivity to navitoclax, compared with those with RUNX1 missense mutations (two patients) (Supplementary Fig. ('patient', 'Species', '9606', (232, 239)) ('patients', 'Species', '9606', (232, 240)) ('reduced', 'NegReg', (142, 149)) ('RUNX1', 'Gene', (50, 55)) ('sensitivity to navitoclax', 'MPA', (150, 175)) ('patient', 'Species', '9606', (28, 35)) ('mTOR', 'Gene', (122, 126)) ('mTOR', 'Gene', '2475', (122, 126)) ('mutation', 'Var', (56, 64)) ('enhanced', 'PosReg', (78, 86)) 5395 32782381 Given the complex genetic background of BP-CML patients, we next validated whether the identified transcriptional and drug sensitivity characteristics are truly specific to RUNX1 mutations. ('drug sensitivity', 'Phenotype', 'HP:0020174', (118, 134)) ('RUNX1', 'Gene', (173, 178)) ('patients', 'Species', '9606', (47, 55)) ('mutations', 'Var', (179, 188)) 5396 32782381 We used a mouse Ba/f3 cell line transduced with P210-BCR-ABL1 expression vector as a model of CP-CML to simulate the impact of RUNX1 mutations on the disease phenotype. ('RUNX1', 'Gene', (127, 132)) ('mouse', 'Species', '10090', (10, 15)) ('mutations', 'Var', (133, 142)) ('P210-BCR-ABL1', 'Var', (48, 61)) 5402 32782381 Furthermore, introduction of RUNX1 p.R162K mutation in Baf3-BCR-ABL1 or K562 cell lines induced changes in the sensitivity profiles, including enhanced activity of navitoclax, AZD8055, and axitinib similar to RUNX1mut patients' profiles (Supplementary Fig. ('enhanced', 'PosReg', (143, 151)) ('activity', 'MPA', (152, 160)) ('axitinib', 'Enzyme', (189, 197)) ('RUNX1', 'Gene', (29, 34)) ('sensitivity profiles', 'MPA', (111, 131)) ('patients', 'Species', '9606', (218, 226)) ('AZD8055', 'Chemical', 'MESH:C546624', (176, 183)) ('changes', 'Reg', (96, 103)) ('K562', 'CellLine', 'CVCL:0004', (72, 76)) ('axitinib', 'Chemical', 'MESH:D000077784', (189, 197)) ('p.R162K', 'Var', (35, 42)) ('p.R162K', 'Mutation', 'rs1057519750', (35, 42)) ('navitoclax', 'MPA', (164, 174)) 5403 32782381 Interestingly, differential drug activity associations with somatic mutation types (e.g., enhanced mTOR activity with nonsense mutations and navitoclax activity with missense mutations) were also notable in the cell line models (Fig. ('nonsense mutations', 'Var', (118, 136)) ('enhanced', 'PosReg', (90, 98)) ('missense mutations', 'Var', (166, 184)) ('mTOR', 'Gene', '2475', (99, 103)) ('mTOR', 'Gene', (99, 103)) 5404 32782381 Given the RUNX1mut-associated distinct phenotype, namely the aberrant expression of CD19 lymphoid marker in myeloid blast cells, we investigated the potential use of CD19-CAR T-cell immunotherapy in RUNX1mut BP-CML patients. ('CD19', 'Gene', (166, 170)) ('aberrant', 'Var', (61, 69)) ('patients', 'Species', '9606', (215, 223)) ('CD19', 'Gene', (84, 88)) ('CAR', 'Gene', (171, 174)) ('CD19', 'Gene', '930', (166, 170)) ('CD19', 'Gene', '930', (84, 88)) ('RUNX1mut-associated', 'Disease', (10, 29)) ('CAR', 'Gene', '653108', (171, 174)) 5409 32782381 At an E-T ratio of 2:1, CD19-CAR T cells-induced killing was superior to killing by imatinib (100 nM), not only in mut1 patient who carried ABL1-T315I resistance mutation, but also in mut2 with no TKI-resistance mutation. ('patient', 'Species', '9606', (120, 127)) ('ABL1-T315I', 'Var', (140, 150)) ('CAR', 'Gene', '653108', (29, 32)) ('imatinib', 'Chemical', 'MESH:D000068877', (84, 92)) ('CD19', 'Gene', (24, 28)) ('T315I', 'Mutation', 'rs121913459', (145, 150)) ('CD19', 'Gene', '930', (24, 28)) ('CAR', 'Gene', (29, 32)) 5417 32782381 In this work, we systematically studied the genomic, transcriptional, and drug sensitivity profiles of BP-CML primary patient samples with and without RUNX1 mutations. ('mutations', 'Var', (157, 166)) ('patient', 'Species', '9606', (118, 125)) ('RUNX1', 'Gene', (151, 156)) ('drug sensitivity', 'Phenotype', 'HP:0020174', (74, 90)) 5418 32782381 Our study coupled RUNX1 mutations in BP-CML with recombination events caused by off-target activity of AID/RAG complex. ('RUNX1', 'Gene', (18, 23)) ('mutations', 'Var', (24, 33)) ('AID', 'Gene', '57379', (103, 106)) ('AID', 'Gene', (103, 106)) 5422 32782381 The incidence of RUNX1 mutations in BP-CML patients ranges between 12.9 and 33.3%, varying with the cohort size, disease phenotype (myeloid or lymphoid), and the sequencing method used. ('patients', 'Species', '9606', (43, 51)) ('mutations', 'Var', (23, 32)) ('RUNX1', 'Gene', (17, 22)) 5423 32782381 In our own discovery cohort, we identified four mutations of RUNX1, that were located within the Runt domain, in line with reports of BP-CML and AML. ('AML', 'Disease', 'MESH:D015470', (145, 148)) ('AML', 'Disease', (145, 148)) ('mutations', 'Var', (48, 57)) ('RUNX1', 'Gene', (61, 66)) 5424 32782381 The identified variants have been reported in AML, displaying variable effects on RUNX1 protein functions, including CBFB dimerization and DNA binding, in addition to leukemia transformation. ('CBFB', 'Gene', '865', (117, 121)) ('DNA', 'MPA', (139, 142)) ('AML', 'Disease', 'MESH:D015470', (46, 49)) ('RUNX1', 'Gene', (82, 87)) ('AML', 'Disease', (46, 49)) ('CBFB', 'Gene', (117, 121)) ('effects', 'Reg', (71, 78)) ('leukemia transformation', 'Disease', 'MESH:D002472', (167, 190)) ('dimerization', 'MPA', (122, 134)) ('variants', 'Var', (15, 23)) ('leukemia transformation', 'Disease', (167, 190)) ('leukemia', 'Phenotype', 'HP:0001909', (167, 175)) 5425 32782381 In a myeloid-BP patient, the RUNX1 mutation was the sole leukemia-associated mutation identified both in CP (SNV) and progression (SNV and LOH) samples. ('leukemia', 'Disease', (57, 65)) ('RUNX1', 'Gene', (29, 34)) ('mutation', 'Var', (35, 43)) ('leukemia', 'Phenotype', 'HP:0001909', (57, 65)) ('patient', 'Species', '9606', (16, 23)) ('leukemia', 'Disease', 'MESH:D007938', (57, 65)) 5426 32782381 Giustacchini et al.. similarly reported a RUNX1 mutation in both CP stem cells (SCs) and BP-SCs of a lymphoid-BP patient. ('patient', 'Species', '9606', (113, 120)) ('mutation', 'Var', (48, 56)) ('RUNX1', 'Gene', (42, 47)) 5427 32782381 RUNX1 aberrations contribute to mutagenesis and leukemic predisposition, and associate with downregulation of DNA repair genes in AML. ('aberrations', 'Var', (6, 17)) ('downregulation', 'NegReg', (92, 106)) ('AML', 'Disease', (130, 133)) ('AML', 'Disease', 'MESH:D015470', (130, 133)) ('DNA repair genes', 'Gene', (110, 126)) ('RUNX1', 'Gene', (0, 5)) ('leukemic', 'Disease', (48, 56)) ('mutagenesis', 'CPA', (32, 43)) ('contribute', 'Reg', (18, 28)) 5429 32782381 Aberrant AID/RAG activity is implicated in lymphoid malignancies, namely in ETV6-RUNX1 ALL. ('Aberrant', 'Var', (0, 8)) ('lymphoid malignancies', 'Disease', 'MESH:D008223', (43, 64)) ('implicated', 'Reg', (29, 39)) ('lymphoid malignancies', 'Disease', (43, 64)) ('AID', 'Gene', (9, 12)) ('ETV6', 'Gene', (76, 80)) ('AID', 'Gene', '57379', (9, 12)) ('lymphoid malignancies', 'Phenotype', 'HP:0002665', (43, 64)) ('ETV6', 'Gene', '2120', (76, 80)) 5432 32782381 We demonstrated presence of RAG off-target activity in an IKZF1 deletion in RUNX1mut BP-CML, like previously reported in Philadelphia-positive ALL (Ph-ALL). ('IKZF1', 'Gene', (58, 63)) ('RUNX1mut', 'Gene', (76, 84)) ('deletion', 'Var', (64, 72)) ('IKZF1', 'Gene', '10320', (58, 63)) 5435 32782381 Interestingly, they reported a RUNX1mut myeloid-BP patient with exceptionally high RAG expression and aberrant lymphoid markers phenotype supporting the role of RUNX1mut-induced RAG activity. ('RAG expression', 'MPA', (83, 97)) ('high', 'PosReg', (78, 82)) ('RUNX1mut myeloid-BP', 'Var', (31, 50)) ('aberrant lymphoid markers', 'Phenotype', 'HP:0004332', (102, 127)) ('patient', 'Species', '9606', (51, 58)) 5437 32782381 RUNX1mut BP-CML exhibited other mutations in BCORL1 and PHF6 genes, as well as IKZF1 deletions, comparable to the mutational landscape of RUNX1mut AML. ('PHF6', 'Gene', '84295', (56, 60)) ('BCORL1', 'Gene', '63035', (45, 51)) ('deletions', 'Var', (85, 94)) ('IKZF1', 'Gene', '10320', (79, 84)) ('RUNX1mut AML', 'Gene', (138, 150)) ('mutations', 'Var', (32, 41)) ('BCORL1', 'Gene', (45, 51)) ('PHF6', 'Gene', (56, 60)) ('IKZF1', 'Gene', (79, 84)) ('RUNX1mut', 'Gene', (0, 8)) ('RUNX1mut AML', 'Gene', '861', (138, 150)) 5438 32782381 In addition, upregulation of early HSC signature, lymphoid markers, and various AML prognostic markers in RUNX1mut BP-CML was another similarity with RUNX1mut AML. ('RUNX1mut AML', 'Gene', (150, 162)) ('RUNX1mut BP-CML', 'Var', (106, 121)) ('AML', 'Disease', (159, 162)) ('AML', 'Disease', (80, 83)) ('RUNX1mut AML', 'Gene', '861', (150, 162)) ('upregulation', 'PosReg', (13, 25)) ('lymphoid markers', 'MPA', (50, 66)) ('early HSC signature', 'MPA', (29, 48)) ('AML', 'Disease', 'MESH:D015470', (159, 162)) ('AML', 'Disease', 'MESH:D015470', (80, 83)) 5439 32782381 Downregulation of the coagulation pathway and megakaryocytic markers is consistent with the role of RUNX1 mutations in FPD/AML. ('FPD', 'Disease', (119, 122)) ('RUNX1', 'Gene', (100, 105)) ('coagulation pathway', 'Pathway', (22, 41)) ('AML', 'Disease', 'MESH:D015470', (123, 126)) ('Downregulation', 'NegReg', (0, 14)) ('mutations', 'Var', (106, 115)) ('FPD', 'Disease', 'MESH:C563324', (119, 122)) ('AML', 'Disease', (123, 126)) ('megakaryocytic markers', 'CPA', (46, 68)) 5441 32782381 Aberrant expression of CD19 has been described in t(8;21)-AML to relate with PAX5 overexpression. ('PAX5', 'Gene', (77, 81)) ('Aberrant', 'Var', (0, 8)) ('PAX5', 'Gene', '5079', (77, 81)) ('AML', 'Disease', 'MESH:D015470', (58, 61)) ('CD19', 'Gene', (23, 27)) ('CD19', 'Gene', '930', (23, 27)) ('AML', 'Disease', (58, 61)) 5442 32782381 We demonstrated overexpression of PAX5 in RUNX1mut BP-CML patients, in concord with data from RUNX1mut AML. ('RUNX1mut AML', 'Gene', '861', (94, 106)) ('overexpression', 'PosReg', (16, 30)) ('PAX5', 'Gene', (34, 38)) ('PAX5', 'Gene', '5079', (34, 38)) ('RUNX1mut AML', 'Gene', (94, 106)) ('patients', 'Species', '9606', (58, 66)) ('RUNX1mut BP-CML', 'Var', (42, 57)) 5443 32782381 RUNX1mut BP-CML patients showed upregulation of many pDC markers. ('pDC', 'Disease', (53, 56)) ('RUNX1mut BP-CML', 'Var', (0, 15)) ('patients', 'Species', '9606', (16, 24)) ('upregulation', 'PosReg', (32, 44)) 5445 32782381 A recently described AML entity, "AML with pDC differentiation", demonstrated frequent RUNX1 mutations and expression of lymphoid antigens, comparable to RUNX1mut BP-CML. ('mutations', 'Var', (93, 102)) ('expression', 'MPA', (107, 117)) ('AML', 'Disease', 'MESH:D015470', (21, 24)) ('AML', 'Disease', 'MESH:D015470', (34, 37)) ('AML', 'Disease', (21, 24)) ('RUNX1', 'Gene', (87, 92)) ('AML', 'Disease', (34, 37)) 5446 32782381 Noteworthy, a study including 47 RUNX1mut-AML patients showed that RUNX1mut blasts shared a common gene expression signature in contrast with RUNX1wt blasts, and transcriptional differences between missense and nonsense RUNX1 mutations were demonstrated in some RUNX1 target genes. ('RUNX1mut', 'Var', (67, 75)) ('patients', 'Species', '9606', (46, 54)) ('nonsense', 'Var', (211, 219)) ('AML', 'Disease', (42, 45)) ('AML', 'Disease', 'MESH:D015470', (42, 45)) ('mutations', 'Var', (226, 235)) ('missense', 'Var', (198, 206)) ('RUNX1', 'Gene', (220, 225)) 5447 32782381 We identified potentially useful targeted drugs for RUNX1mut BP-CML patients, including mTOR inhibitors, glucocorticoids, VEGFR inhibitors, and BCL2 inhibitors. ('VEGFR', 'Gene', '3791', (122, 127)) ('RUNX1mut', 'Var', (52, 60)) ('BCL2', 'Gene', (144, 148)) ('mTOR', 'Gene', (88, 92)) ('patients', 'Species', '9606', (68, 76)) ('VEGFR', 'Gene', (122, 127)) ('mTOR', 'Gene', '2475', (88, 92)) ('BCL2', 'Gene', '596', (144, 148)) 5450 32782381 In RUNX1mutAML, glucocorticoid sensitivity is associated with RUNX1 mutations and wild-type RUNX1 activity, which potentially explains variances in glucocorticoid activity in our samples also. ('glucocorticoid sensitivity', 'MPA', (16, 42)) ('glucocorticoid sensitivity', 'Phenotype', 'HP:0008163', (16, 42)) ('AML', 'Disease', 'MESH:D015470', (11, 14)) ('associated', 'Reg', (46, 56)) ('mutations', 'Var', (68, 77)) ('RUNX1', 'Gene', (62, 67)) ('AML', 'Disease', (11, 14)) 5455 32782381 RUNX1 mutations are associated with upregulation of several molecular targets for immunotherapy, including CD19 and CD133. ('CD19', 'Gene', (107, 111)) ('CD19', 'Gene', '930', (107, 111)) ('upregulation', 'PosReg', (36, 48)) ('RUNX1', 'Gene', (0, 5)) ('CD133', 'Gene', (116, 121)) ('CD133', 'Gene', '8842', (116, 121)) ('mutations', 'Var', (6, 15)) 5463 32782381 In conclusion, this study provides insights into the role of RUNX1 mutations in CML progression by induced transcriptional reprogramming and aberrant mutagenic AID/RAG activity. ('CML', 'Disease', (80, 83)) ('AID', 'Gene', (160, 163)) ('RUNX1', 'Gene', (61, 66)) ('mutations', 'Var', (67, 76)) ('AID', 'Gene', '57379', (160, 163)) ('transcriptional reprogramming', 'CPA', (107, 136)) 5466 32782381 SB is a member of Member of the advisory board of Qiagen, Novartis, and Cepheid and has received research funding from Novartis, honoraria from Novartis, Qiagen, Cepheid, and Bristol-Myers Squibb and support from the National Health and Medical Research Council of Australia APP1117718 and APP1104425, the Ray and Shirl Norman Cancer Research Trust, and the Royal Adelaide Hospital Research Foundation. ('Cancer', 'Disease', (327, 333)) ('Cancer', 'Phenotype', 'HP:0002664', (327, 333)) ('Cancer', 'Disease', 'MESH:D009369', (327, 333)) ('APP1104425', 'Var', (290, 300)) 5471 33821592 This review summarizes the recent advances in epigenetic therapies for AML and discusses future research directions. ('epigenetic', 'Var', (46, 56)) ('AML', 'Phenotype', 'HP:0004808', (71, 74)) ('AML', 'Disease', 'MESH:D015470', (71, 74)) ('AML', 'Disease', (71, 74)) 5474 33821592 Treatment strategies could be modified by risk groups according to cytogenetic profiles including promyelocytic leukemia/retinoic acid receptor alpha (PML/RARA), core-binding factor subunit beta/myosin heavy chain 11 (CBFb/MYH11), runt-related transcription factor1/runt-related transcription factor1 translocation partner 1 (RUNX1/RUNX1T1) translocation that were known before the next-generation sequencing (NGS) era, however, only to decide whether to give several cycles of consolidation chemotherapy or to undergo HSCT. ('promyelocytic leukemia', 'Disease', (98, 120)) ('leukemia', 'Phenotype', 'HP:0001909', (112, 120)) ('CBFb', 'Gene', (218, 222)) ('runt-related transcription factor1', 'Gene', '861', (266, 300)) ('RARA', 'Gene', (155, 159)) ('retinoic acid', 'Chemical', 'MESH:D014212', (121, 134)) ('myosin heavy chain 11', 'Gene', '4629', (195, 216)) ('MYH11', 'Gene', '4629', (223, 228)) ('runt-related transcription factor1', 'Gene', '861', (231, 265)) ('core-binding factor subunit beta', 'Gene', '865', (162, 194)) ('RUNX1T1', 'Gene', (332, 339)) ('CBFb', 'Gene', '865', (218, 222)) ('MYH11', 'Gene', (223, 228)) ('PML', 'Gene', (151, 154)) ('promyelocytic leukemia', 'Disease', 'MESH:D015473', (98, 120)) ('runt-related transcription factor1', 'Gene', (266, 300)) ('myosin heavy chain 11', 'Gene', (195, 216)) ('RUNX1', 'Gene', (326, 331)) ('RUNX1', 'Gene', '861', (326, 331)) ('runt-related transcription factor1', 'Gene', (231, 265)) ('core-binding factor subunit beta', 'Gene', (162, 194)) ('PML', 'Gene', '5371', (151, 154)) ('RARA', 'Gene', '5914', (155, 159)) ('promyelocytic leukemia', 'Phenotype', 'HP:0004836', (98, 120)) ('translocation', 'Var', (341, 354)) ('RUNX1', 'Gene', (332, 337)) ('RUNX1T1', 'Gene', '862', (332, 339)) ('RUNX1', 'Gene', '861', (332, 337)) 5480 33821592 Based on this genomic information, novel therapeutic targets and drugs have been developed, including an FLT3 inhibitor that acts on a tyrosine kinase receptor and an IDH1/2 inhibitor that interrupts epigenetic changes. ('tyrosine kinase receptor', 'MPA', (135, 159)) ('IDH1/2', 'Gene', (167, 173)) ('FLT3', 'Gene', '2322', (105, 109)) ('inhibitor', 'Var', (110, 119)) ('FLT3', 'Gene', (105, 109)) ('interrupts', 'NegReg', (189, 199)) ('epigenetic changes', 'MPA', (200, 218)) ('IDH1/2', 'Gene', '3417;3418', (167, 173)) 5484 33821592 Of these, mutations in epigenetic regulators are the most frequent, implying that epigenetic dysregulation is critical in AML pathogenesis. ('AML', 'Disease', 'MESH:D015470', (122, 125)) ('mutations', 'Var', (10, 19)) ('AML', 'Phenotype', 'HP:0004808', (122, 125)) ('frequent', 'Reg', (58, 66)) ('AML', 'Disease', (122, 125)) 5485 33821592 It is well known that altered gene expression caused by epigenetic dysregulations is an important carcinogenic mechanism. ('carcinogenic', 'Disease', 'MESH:D063646', (98, 110)) ('altered', 'Reg', (22, 29)) ('epigenetic dysregulations', 'Var', (56, 81)) ('carcinogenic', 'Disease', (98, 110)) 5486 33821592 A recent NGS study reported that more than 70% of AML patients showed mutations at genes related in DNA methylation or histone acetylation. ('mutations', 'Var', (70, 79)) ('AML', 'Disease', 'MESH:D015470', (50, 53)) ('patients', 'Species', '9606', (54, 62)) ('AML', 'Phenotype', 'HP:0004808', (50, 53)) ('AML', 'Disease', (50, 53)) 5487 33821592 For this reason, most new drugs for AML are mainly focused on the epigenetic modifications, except FLT3 inhibitors and CAR-T therapy. ('AML', 'Disease', 'MESH:D015470', (36, 39)) ('CAR-T', 'Chemical', '-', (119, 124)) ('AML', 'Disease', (36, 39)) ('FLT3', 'Gene', '2322', (99, 103)) ('AML', 'Phenotype', 'HP:0004808', (36, 39)) ('epigenetic modifications', 'Var', (66, 90)) ('FLT3', 'Gene', (99, 103)) 5493 33821592 Hence, inhibition of DNMT through hypomethylation can restore normal molecular function. ('DNMT', 'Gene', '1786', (21, 25)) ('molecular function', 'MPA', (69, 87)) ('DNMT', 'Gene', (21, 25)) ('inhibition', 'NegReg', (7, 17)) ('restore', 'PosReg', (54, 61)) ('hypomethylation', 'Var', (34, 49)) 5497 33821592 Decitabine and azacitidine can also cause direct DNA damage and promote the expression of tumor-associated antigens, thus eliciting an anti-leukemic immune reaction. ('promote', 'PosReg', (64, 71)) ('cause', 'Reg', (36, 41)) ('direct DNA damage', 'MPA', (42, 59)) ('anti-leukemic immune reaction', 'CPA', (135, 164)) ('tumor', 'Disease', (90, 95)) ('Decitabine', 'Chemical', 'MESH:D000077209', (0, 10)) ('tumor', 'Phenotype', 'HP:0002664', (90, 95)) ('azacitidine', 'Var', (15, 26)) ('expression', 'MPA', (76, 86)) ('azacitidine', 'Chemical', 'MESH:D001374', (15, 26)) ('tumor', 'Disease', 'MESH:D009369', (90, 95)) ('eliciting', 'Reg', (122, 131)) 5498 33821592 Furthermore, azacitidine incorporates into RNA and interrupts protein synthesis and induces apoptosis. ('apoptosis', 'CPA', (92, 101)) ('interrupts', 'NegReg', (51, 61)) ('azacitidine', 'Var', (13, 24)) ('azacitidine', 'Chemical', 'MESH:D001374', (13, 24)) ('protein synthesis', 'MPA', (62, 79)) ('induces', 'Reg', (84, 91)) 5507 33821592 The difference between the 2 agents is based on the result of a retrospective study that compared azacitidine and decitabine in older patients and reported that azacitidine had a longer OS. ('azacitidine', 'Var', (161, 172)) ('azacitidine', 'Chemical', 'MESH:D001374', (161, 172)) ('longer', 'MPA', (179, 185)) ('decitabine', 'Chemical', 'MESH:D000077209', (114, 124)) ('patients', 'Species', '9606', (134, 142)) ('azacitidine', 'Chemical', 'MESH:D001374', (98, 109)) 5521 33821592 If mutations occur in IDH1 and IDH2, alpha-ketoglutarate is converted to 2-hydroxyglutarate (2-HG) in a nicotinamide adenine dinucleotide phosphate hydrogen-dependent manner. ('IDH2', 'Gene', '3418', (31, 35)) ('2-hydroxyglutarate', 'Chemical', 'MESH:C019417', (73, 91)) ('alpha-ketoglutarate', 'Chemical', 'MESH:D007656', (37, 56)) ('IDH1', 'Gene', (22, 26)) ('2-HG', 'Chemical', 'MESH:C019417', (93, 97)) ('hydrogen', 'Chemical', 'MESH:D006859', (148, 156)) ('nicotinamide adenine dinucleotide phosphate', 'Chemical', 'MESH:D009249', (104, 147)) ('IDH1', 'Gene', '3417', (22, 26)) ('IDH2', 'Gene', (31, 35)) ('mutations', 'Var', (3, 12)) 5522 33821592 Increased 2-HG leads to gene hypermethylation, resulting in enhanced cellular proliferation, aberrant gene expression, and the inhibition of myeloid differentiation. ('cellular proliferation', 'CPA', (69, 91)) ('myeloid differentiation', 'CPA', (141, 164)) ('2-HG', 'Chemical', 'MESH:C019417', (10, 14)) ('aberrant', 'Var', (93, 101)) ('hypermethylation', 'Var', (29, 45)) ('inhibition', 'NegReg', (127, 137)) ('enhanced', 'PosReg', (60, 68)) ('gene', 'MPA', (24, 28)) 5523 33821592 Frequent co-occurrence of IDH and DNMT3A mutations is reported. ('mutations', 'Var', (41, 50)) ('DNMT3A', 'Gene', (34, 40)) ('DNMT3A', 'Gene', '1788', (34, 40)) ('IDH', 'Gene', (26, 29)) ('IDH', 'Gene', '3417', (26, 29)) 5525 33821592 TET2 loss-of-function mutation is frequently found in AML. ('TET2', 'Gene', (0, 4)) ('AML', 'Disease', 'MESH:D015470', (54, 57)) ('mutation', 'Var', (22, 30)) ('TET2', 'Gene', '54790', (0, 4)) ('AML', 'Phenotype', 'HP:0004808', (54, 57)) ('AML', 'Disease', (54, 57)) ('loss-of-function', 'NegReg', (5, 21)) 5526 33821592 Therefore, co-occurrence of IDH and TET2 mutations leads to global DNA hypermethylation, thus contributing to leukemogenesis. ('mutations', 'Var', (41, 50)) ('IDH', 'Gene', (28, 31)) ('TET2', 'Gene', '54790', (36, 40)) ('leads to', 'Reg', (51, 59)) ('IDH', 'Gene', '3417', (28, 31)) ('TET2', 'Gene', (36, 40)) ('global DNA hypermethylation', 'MPA', (60, 87)) ('leukemogenesis', 'Disease', (110, 124)) ('contributing to', 'Reg', (94, 109)) 5528 33821592 Both are oral small molecules that inactivate mutated IDH proteins via allosteric inhibition and reduction of the oncometabolite 2-HG. ('allosteric', 'MPA', (71, 81)) ('reduction', 'NegReg', (97, 106)) ('mutated', 'Var', (46, 53)) ('oncometabolite 2-HG', 'MPA', (114, 133)) ('IDH', 'Gene', (54, 57)) ('inactivate', 'NegReg', (35, 45)) ('IDH', 'Gene', '3417', (54, 57)) ('2-HG', 'Chemical', 'MESH:C019417', (129, 133)) 5538 33821592 Acetylation of the lysine residues in the N-terminal region leads to gene expression by enabling DNA binding to transcription factors. ('Acetylation', 'Var', (0, 11)) ('DNA binding', 'Interaction', (97, 108)) ('lysine', 'Chemical', 'MESH:D008239', (19, 25)) ('gene expression', 'MPA', (69, 84)) ('enabling', 'PosReg', (88, 96)) ('leads to', 'Reg', (60, 68)) 5542 33821592 Deacetylation of P53, a well-known TSG, induces the suppression of P53-dependent growth arrest and apoptosis, and increases leukemogenesis risk. ('P53', 'Gene', '7157', (67, 70)) ('growth arrest', 'Disease', (81, 94)) ('suppression', 'NegReg', (52, 63)) ('P53', 'Gene', (17, 20)) ('growth arrest', 'Disease', 'MESH:D006323', (81, 94)) ('apoptosis', 'CPA', (99, 108)) ('leukemogenesis', 'Disease', (124, 138)) ('Deacetylation', 'Var', (0, 13)) ('increases', 'PosReg', (114, 123)) ('P53', 'Gene', '7157', (17, 20)) ('growth arrest', 'Phenotype', 'HP:0001510', (81, 94)) ('P53', 'Gene', (67, 70)) 5544 33821592 However, other genes that affect HDACs, including enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2) and additional sex combs like 1, transcriptional regulator 1 (ASXL1), are commonly mutated in AML. ('EZH2', 'Gene', '2146', (109, 113)) ('ASXL1', 'Gene', '171023', (177, 182)) ('HDAC', 'Gene', (33, 37)) ('ASXL1', 'Gene', (177, 182)) ('AML', 'Disease', 'MESH:D015470', (209, 212)) ('HDAC', 'Gene', '9734', (33, 37)) ('mutated', 'Var', (198, 205)) ('enhancer', 'PosReg', (50, 58)) ('AML', 'Phenotype', 'HP:0004808', (209, 212)) ('AML', 'Disease', (209, 212)) ('additional sex combs like 1, transcriptional regulator 1', 'Gene', '171023', (119, 175)) ('EZH2', 'Gene', (109, 113)) 5545 33821592 HDAC inhibitors increase histone acetylation to reactivated TSGs, and most are still under development for AML treatment; however, several agents have been FDA approved for other hematologic malignancies. ('HDAC', 'Gene', (0, 4)) ('HDAC', 'Gene', '9734', (0, 4)) ('AML', 'Phenotype', 'HP:0004808', (107, 110)) ('AML', 'Disease', (107, 110)) ('inhibitors', 'Var', (5, 15)) ('increase', 'PosReg', (16, 24)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (179, 203)) ('histone acetylation', 'MPA', (25, 44)) ('AML', 'Disease', 'MESH:D015470', (107, 110)) ('hematologic malignancies', 'Disease', (179, 203)) 5553 33821592 Besides the many epigenetic agents mentioned above, many novel target molecules are in clinical development, including the DOT1 like histone lysine methyltransferase (DOT1L) inhibitors (EPZ004777, EPZ-5676), bromodomain and extra-terminal domain (BET) bromodomain inhibitor, lysine demethylase 1 (LSD1) inhibitor, EZH inhibitor, and B-cell chronic lymphocytic leukemia/lymphoma 2 (BCL-2; venetoclax) inhibitors. ('lysine', 'Chemical', 'MESH:D008239', (141, 147)) ('lymphoma', 'Phenotype', 'HP:0002665', (369, 377)) ('lysine demethylase 1', 'Gene', (275, 295)) ('lysine demethylase 1', 'Gene', '23028', (275, 295)) ('EPZ004777', 'Var', (186, 195)) ('lysine', 'Chemical', 'MESH:D008239', (275, 281)) ('DOT1', 'Gene', '84444', (123, 127)) ('DOT1', 'Gene', (167, 171)) ('BCL-2', 'Gene', '596', (381, 386)) ('DOT1L', 'Gene', (167, 172)) ('B-cell chronic lymphocytic leukemia/lymphoma 2', 'Gene', '596', (333, 379)) ('venetoclax', 'Chemical', 'MESH:C579720', (388, 398)) ('LSD1', 'Gene', (297, 301)) ('BCL-2', 'Gene', (381, 386)) ('LSD1', 'Gene', '23028', (297, 301)) ('chronic lymphocytic leukemia', 'Phenotype', 'HP:0005550', (340, 368)) ('leukemia', 'Phenotype', 'HP:0001909', (360, 368)) ('B-cell chronic lymphocytic leukemia/lymphoma 2', 'Gene', (333, 379)) ('DOT1', 'Gene', '84444', (167, 171)) ('DOT1L', 'Gene', '84444', (167, 172)) ('DOT1', 'Gene', (123, 127)) 5554 33821592 The suppression of Hox expression by DOT1L inhibitors results from the rearrangement of the myeloid/lymphoid or mixed-lineage leukemia (MLL) gene, and was responsible for its observed antileukemic activities in early preclinical and early phase clinical research; however, this efficacy was minimal in later phase trials. ('DOT1L', 'Gene', (37, 42)) ('antileukemic activities', 'CPA', (184, 207)) ('suppression', 'NegReg', (4, 15)) ('MLL', 'Gene', (136, 139)) ('leukemia', 'Disease', (126, 134)) ('MLL', 'Gene', '4297', (136, 139)) ('leukemia', 'Disease', 'MESH:D007938', (126, 134)) ('leukemia', 'Phenotype', 'HP:0001909', (126, 134)) ('Hox', 'Gene', (19, 22)) ('DOT1L', 'Gene', '84444', (37, 42)) ('rearrangement', 'Var', (71, 84)) 5570 33821592 As increased 2-HG by IDH mutations leads to DNA hypermethylation and frequent co-occurrence of IDH and DNMT3A mutations is reported, combination of IDH inhibitors and DNMT inhibitors is logical therapeutic strategy. ('increased', 'PosReg', (3, 12)) ('IDH', 'Gene', (21, 24)) ('DNMT', 'Gene', (103, 107)) ('DNA hypermethylation', 'MPA', (44, 64)) ('mutations', 'Var', (25, 34)) ('IDH', 'Gene', '3417', (21, 24)) ('IDH', 'Gene', (148, 151)) ('DNMT', 'Gene', '1786', (167, 171)) ('IDH', 'Gene', '3417', (148, 151)) ('DNMT3A', 'Gene', (103, 109)) ('DNMT', 'Gene', (167, 171)) ('2-HG', 'MPA', (13, 17)) ('IDH', 'Gene', (95, 98)) ('DNMT3A', 'Gene', '1788', (103, 109)) ('IDH', 'Gene', '3417', (95, 98)) ('DNMT', 'Gene', '1786', (103, 107)) ('2-HG', 'Chemical', 'MESH:C019417', (13, 17)) 5581 33821592 Clinical trials of more specific FLT inhibitors including quizartinib or gilteritinib combination with HMAs are currently being conducted (NCT03661307, NCT01892371, and NCT02752035). ('NCT02752035', 'Var', (169, 180)) ('FLT', 'Gene', '2321', (33, 36)) ('NCT01892371', 'Var', (152, 163)) ('quizartinib', 'Chemical', 'MESH:C544967', (58, 69)) ('HMAs', 'Chemical', '-', (103, 107)) ('gilteritinib', 'Chemical', 'MESH:C000609080', (73, 85)) ('FLT', 'Gene', (33, 36)) ('NCT03661307', 'Var', (139, 150)) 5594 33821592 FLT3 mutation, one of the most common mutation in AML presents in only around 20% of patients. ('AML', 'Disease', 'MESH:D015470', (50, 53)) ('AML', 'Phenotype', 'HP:0004808', (50, 53)) ('patients', 'Species', '9606', (85, 93)) ('AML', 'Disease', (50, 53)) ('FLT3', 'Gene', '2322', (0, 4)) ('FLT3', 'Gene', (0, 4)) ('mutation', 'Var', (5, 13)) 5602 33821592 It is reported in previous researches that epigenetic silencing of immune-related genes is important to immune modulation involving T-cells in cancer. ('cancer', 'Phenotype', 'HP:0002664', (143, 149)) ('cancer', 'Disease', (143, 149)) ('cancer', 'Disease', 'MESH:D009369', (143, 149)) ('epigenetic silencing', 'Var', (43, 63)) 5615 33821592 IDH mutation sensitizes leukemic cells to BCL-2 inhibition by 2-HG accumulation. ('IDH', 'Gene', (0, 3)) ('BCL-2', 'Gene', '596', (42, 47)) ('IDH', 'Gene', '3417', (0, 3)) ('mutation', 'Var', (4, 12)) ('BCL-2', 'Gene', (42, 47)) ('2-HG', 'Chemical', 'MESH:C019417', (62, 66)) ('sensitizes', 'Reg', (13, 23)) 5617 33821592 As IDH mutated relapsed/refractory AML had a superior ORR (33% vs. 10%) to wild-type in a venetoclax monotherapy trial, combination studies of IDH inhibitors and venetoclax have been conducted. ('AML', 'Disease', (35, 38)) ('IDH', 'Gene', (3, 6)) ('IDH', 'Gene', '3417', (3, 6)) ('IDH', 'Gene', '3417', (143, 146)) ('AML', 'Phenotype', 'HP:0004808', (35, 38)) ('AML', 'Disease', 'MESH:D015470', (35, 38)) ('mutated', 'Var', (7, 14)) ('venetoclax', 'Chemical', 'MESH:C579720', (162, 172)) ('venetoclax', 'Chemical', 'MESH:C579720', (90, 100)) ('IDH', 'Gene', (143, 146)) 5632 33821592 Various types of structural chromosomal abnormalities and intra-genetic aberrations occur in AML, as well as epigenetic changes. ('AML', 'Phenotype', 'HP:0004808', (93, 96)) ('AML', 'Disease', (93, 96)) ('epigenetic changes', 'Var', (109, 127)) ('chromosomal abnormalities', 'Disease', (28, 53)) ('chromosomal abnormalities', 'Disease', 'MESH:D002869', (28, 53)) ('occur', 'Reg', (84, 89)) ('structural', 'Var', (17, 27)) ('AML', 'Disease', 'MESH:D015470', (93, 96)) 5635 33821592 In a recent trial with panobinostat, panobinostat-induced increase of histone acetylation was regarded as a predictor of response, however, more general factors to apply all patients and more earlier predictors to forecast before treatment initiation are needed. ('increase', 'PosReg', (58, 66)) ('panobinostat-induced', 'Var', (37, 57)) ('patients', 'Species', '9606', (174, 182)) ('panobinostat', 'Chemical', 'MESH:D000077767', (37, 49)) ('panobinostat', 'Chemical', 'MESH:D000077767', (23, 35)) ('histone acetylation', 'MPA', (70, 89)) 5637 33821592 Despite being incompletely elucidated, epigenetic therapies for AML might be one of the most important future treatment options, with present rapid development that correspond to clinical requirements. ('AML', 'Disease', (64, 67)) ('AML', 'Phenotype', 'HP:0004808', (64, 67)) ('AML', 'Disease', 'MESH:D015470', (64, 67)) ('epigenetic therapies', 'Var', (39, 59)) 5638 31086779 JAK2-mutated acute myeloid leukemia: comparison of next-generation sequencing (NGS) and single nucleotide polymorphism array (SNPa) findings between two cases JAK2 mutations are rare in de novo acute myeloid leukemia (AML), and JAK2-mutated acute myeloid leukemia (AML) patients usually have a previous history of myeloproliferative neoplasms (MPNs). ('neoplasm', 'Phenotype', 'HP:0002664', (333, 341)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (241, 263)) ('AML', 'Phenotype', 'HP:0004808', (265, 268)) ('AML', 'Disease', 'MESH:D015470', (218, 221)) ('JAK2', 'Gene', '3717', (0, 4)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (241, 263)) ('AML', 'Disease', (218, 221)) ('myeloproliferative neoplasms', 'Phenotype', 'HP:0005547', (314, 342)) ('JAK2', 'Gene', '3717', (159, 163)) ('AML', 'Phenotype', 'HP:0004808', (218, 221)) ('myeloproliferative neoplasms', 'Disease', (314, 342)) ('acute myeloid leukemia', 'Disease', (13, 35)) ('JAK2', 'Gene', (0, 4)) ('JAK2', 'Gene', '3717', (228, 232)) ('acute myeloid leukemia', 'Disease', (194, 216)) ('JAK2', 'Gene', (159, 163)) ('myeloproliferative neoplasms', 'Disease', 'MESH:D009196', (314, 342)) ('leukemia', 'Phenotype', 'HP:0001909', (27, 35)) ('acute myeloid leukemia', 'Disease', (241, 263)) ('mutations', 'Var', (164, 173)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (13, 35)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (19, 35)) ('myeloproliferative neoplasm', 'Phenotype', 'HP:0005547', (314, 341)) ('leukemia', 'Phenotype', 'HP:0001909', (255, 263)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (13, 35)) ('JAK2', 'Gene', (228, 232)) ('patients', 'Species', '9606', (270, 278)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (194, 216)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (247, 263)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (194, 216)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (200, 216)) ('AML', 'Disease', 'MESH:D015470', (265, 268)) ('neoplasms', 'Phenotype', 'HP:0002664', (333, 342)) ('leukemia', 'Phenotype', 'HP:0001909', (208, 216)) ('MPNs', 'Phenotype', 'HP:0005547', (344, 348)) ('AML', 'Disease', (265, 268)) 5639 31086779 Current advances in laboratory techniques, such as single nucleotide polymorphism array (SNPa) and next-generation sequencing (NGS), have facilitated new insight into the molecular basis of hematologic diseases. ('single nucleotide polymorphism', 'Var', (51, 81)) ('hematologic diseases', 'Phenotype', 'HP:0001871', (190, 210)) ('hematologic diseases', 'Disease', 'MESH:D006402', (190, 210)) ('hematologic diseases', 'Disease', (190, 210)) 5640 31086779 Herein, we present two cases of JAK2-mutated AML in which both SNPa and NGS methods added valuable information. ('AML', 'Phenotype', 'HP:0004808', (45, 48)) ('JAK2-mutated', 'Var', (32, 44)) ('AML', 'Disease', 'MESH:D015470', (45, 48)) ('AML', 'Disease', (45, 48)) 5641 31086779 Both cases had leukemogenic collaboration, namely, copy-neutral loss of heterozygosity (CN-LOH), detected on chromosome 9. ('leukemogenic collaboration', 'Disease', 'MESH:C536035', (15, 41)) ('leukemogenic collaboration', 'Disease', (15, 41)) ('copy-neutral', 'Var', (51, 63)) 5642 31086779 One of the cases exhibited both JAK2 and IDH2 mutations, most likely having originated as an MPN with leukemic transformation, while the other case was classified as a de novo AML with JAK2, CEBPA, and FLT3 mutations. ('leukemic transformation', 'Disease', (102, 125)) ('JAK2', 'Gene', (32, 36)) ('mutations', 'Var', (46, 55)) ('CEBPA', 'Gene', (191, 196)) ('AML', 'Phenotype', 'HP:0004808', (176, 179)) ('leukemic transformation', 'Disease', 'MESH:D007938', (102, 125)) ('AML', 'Disease', (176, 179)) ('CEBPA', 'Gene', '1050', (191, 196)) ('IDH2', 'Gene', (41, 45)) ('FLT3', 'Gene', '2322', (202, 206)) ('FLT3', 'Gene', (202, 206)) ('AML', 'Disease', 'MESH:D015470', (176, 179)) ('exhibited', 'Reg', (17, 26)) 5649 31086779 Mutations in JAK2 have been identified in the majority of patients with PV, ET, and PM, underscoring the importance of constitutive activation of JAK2 signaling caused by mutations. ('identified', 'Reg', (28, 38)) ('JAK2', 'Gene', (13, 17)) ('Mutations', 'Var', (0, 9)) ('patients', 'Species', '9606', (58, 66)) 5651 31086779 Patients who exhibit the JAK2 mutation in MPNs progress to AML, suggesting that this mutation might be associated with the acquisition of additional genetic alterations. ('AML', 'Phenotype', 'HP:0004808', (59, 62)) ('progress', 'PosReg', (47, 55)) ('MPNs', 'Phenotype', 'HP:0005547', (42, 46)) ('MPNs', 'Gene', (42, 46)) ('Patients', 'Species', '9606', (0, 8)) ('JAK2', 'Var', (25, 29)) ('AML', 'Disease', 'MESH:D015470', (59, 62)) ('AML', 'Disease', (59, 62)) 5653 31086779 It is important to highlight that, despite the cooccurrence of JAK2 mutations with other genetic alterations, JAK2 mutations have rarely been reported in de novo AML (<5%). ('AML', 'Phenotype', 'HP:0004808', (162, 165)) ('AML', 'Disease', (162, 165)) ('JAK2', 'Gene', (63, 67)) ('mutations', 'Var', (68, 77)) ('AML', 'Disease', 'MESH:D015470', (162, 165)) ('JAK2', 'Gene', (110, 114)) 5654 31086779 Advances in laboratory techniques, such as single nucleotide polymorphism array (SNPa), a molecular cytogenetic method, and high-throughput sequencing, have facilitated new insight into the molecular basis of hematologic diseases. ('hematologic diseases', 'Disease', (209, 229)) ('hematologic diseases', 'Disease', 'MESH:D006402', (209, 229)) ('single', 'Var', (43, 49)) ('hematologic diseases', 'Phenotype', 'HP:0001871', (209, 229)) 5655 31086779 Next-generation sequencing (NGS) is capable of detecting single nucleotide variants (SNVs) or small insertions and deletions that have recently been shown as important molecular phenomena in AML. ('AML', 'Disease', 'MESH:D015470', (191, 194)) ('AML', 'Phenotype', 'HP:0004808', (191, 194)) ('AML', 'Disease', (191, 194)) ('insertions', 'Var', (100, 110)) ('single nucleotide variants', 'Var', (57, 83)) ('deletions', 'Var', (115, 124)) 5656 31086779 To substantiate this idea, we describe two JAK2-mutated AML cases for which both SNPa and NGS added valuable information. ('AML', 'Phenotype', 'HP:0004808', (56, 59)) ('JAK2-mutated', 'Var', (43, 55)) ('AML', 'Disease', 'MESH:D015470', (56, 59)) ('AML', 'Disease', (56, 59)) 5665 31086779 Bone marrow aspirate smear revealed 20% of blasts, and multiparametric flow cytometry immunophenotyping was compatible with AML (Positive: CD45 moderate, CD34, HLA-DR, CD117, CD33 heterogeneous, CD13 heterogeneous, CD11b partial, CD64 partial heterogeneous, CD36 partial, CD7 partial. ('CD117', 'Gene', '3815', (168, 173)) ('CD7', 'Gene', (272, 275)) ('CD45', 'Gene', (139, 143)) ('CD34', 'Gene', (154, 158)) ('CD64', 'Gene', (230, 234)) ('CD45', 'Gene', '5788', (139, 143)) ('CD33', 'Gene', '945', (175, 179)) ('CD33', 'Gene', (175, 179)) ('CD36', 'MPA', (258, 262)) ('CD117', 'Gene', (168, 173)) ('CD13', 'Gene', (195, 199)) ('CD11b', 'Var', (215, 220)) ('CD64', 'Gene', '2209', (230, 234)) ('CD36', 'Species', '42374', (258, 262)) ('CD34', 'Gene', '947', (154, 158)) ('AML', 'Disease', 'MESH:D015470', (124, 127)) ('AML', 'Phenotype', 'HP:0004808', (124, 127)) ('AML', 'Disease', (124, 127)) ('CD13', 'Gene', '290', (195, 199)) ('CD7', 'Gene', '924', (272, 275)) 5666 31086779 Negative: CD14, CD65, CD41, CD2, CD4, CD56, CD19, CD10). ('CD56', 'Gene', '4684', (38, 42)) ('CD41', 'Gene', '3674', (22, 26)) ('CD4', 'Gene', '920', (22, 25)) ('CD19', 'Gene', (44, 48)) ('CD2', 'Gene', '914', (28, 31)) ('CD2', 'Gene', (28, 31)) ('CD10', 'Gene', '4311', (50, 54)) ('CD56', 'Gene', (38, 42)) ('CD14', 'Gene', (10, 14)) ('CD10', 'Gene', (50, 54)) ('CD19', 'Gene', '930', (44, 48)) ('CD4', 'Gene', (22, 25)) ('CD14', 'Gene', '929', (10, 14)) ('CD4', 'Gene', (33, 36)) ('CD41', 'Gene', (22, 26)) ('CD4', 'Gene', '920', (33, 36)) ('CD65', 'Var', (16, 20)) 5668 31086779 NGS analysis revealed IDH2 R140Q and JAK2 V617F mutations. ('R140Q', 'Var', (27, 32)) ('V617F', 'Var', (42, 47)) ('IDH2', 'Gene', (22, 26)) ('R140Q', 'Mutation', 'rs121913502', (27, 32)) ('V617F', 'SUBSTITUTION', 'None', (42, 47)) 5672 31086779 Negative: CD11b, CD10, CD19, CD15, CD36, CD14, CD65, CD2, CD56). ('CD14', 'Gene', (41, 45)) ('CD11b', 'Var', (10, 15)) ('CD56', 'Gene', '4684', (58, 62)) ('CD19', 'Gene', (23, 27)) ('CD14', 'Gene', '929', (41, 45)) ('CD56', 'Gene', (58, 62)) ('CD36', 'Species', '42374', (35, 39)) ('CD19', 'Gene', '930', (23, 27)) ('CD10', 'Gene', '4311', (17, 21)) ('CD10', 'Gene', (17, 21)) ('CD15', 'Var', (29, 33)) ('CD36', 'Var', (35, 39)) ('CD65', 'Var', (47, 51)) ('CD2', 'Gene', '914', (53, 56)) ('CD2', 'Gene', (53, 56)) 5673 31086779 NGS results revealed mutations in FLT3-ITD, CEBPA E309_T310insE and JAK2 V617F. ('T310insE', 'Mutation', 'c.310insT,E', (55, 63)) ('V617F', 'SUBSTITUTION', 'None', (73, 78)) ('CEBPA', 'Gene', (44, 49)) ('FLT3', 'Gene', '2322', (34, 38)) ('CEBPA', 'Gene', '1050', (44, 49)) ('V617F', 'Var', (73, 78)) ('E309_T310insE', 'Var', (50, 63)) ('FLT3', 'Gene', (34, 38)) 5682 31086779 JAK2 and FLT3 mutations share a common mechanism that confers augmented reactive oxygen species, which induce DNA damage, resulting in homologous recombination events that initiate CN-LOH. ('FLT3', 'Gene', '2322', (9, 13)) ('augmented', 'PosReg', (62, 71)) ('reactive oxygen species', 'MPA', (72, 95)) ('FLT3', 'Gene', (9, 13)) ('homologous recombination events', 'MPA', (135, 166)) ('JAK2', 'Gene', (0, 4)) ('mutations', 'Var', (14, 23)) ('reactive oxygen species', 'Chemical', 'MESH:D017382', (72, 95)) 5683 31086779 The JAK2 V617F mutation was identified in both cases. ('V617F', 'SUBSTITUTION', 'None', (9, 14)) ('V617F', 'Var', (9, 14)) ('JAK2', 'Gene', (4, 8)) 5684 31086779 In MPNs, the high JAK2 V617F allele burden in PV represents a risk factor for progression to myelofibrosis. ('myelofibrosis', 'Disease', 'MESH:D055728', (93, 106)) ('MPNs', 'Phenotype', 'HP:0005547', (3, 7)) ('V617F', 'SUBSTITUTION', 'None', (23, 28)) ('myelofibrosis', 'Phenotype', 'HP:0011974', (93, 106)) ('V617F', 'Var', (23, 28)) ('JAK2', 'Gene', (18, 22)) ('myelofibrosis', 'Disease', (93, 106)) 5685 31086779 In contrast, high JAK2 V617F allele burden is not significantly related to leukemic transformation. ('V617F', 'SUBSTITUTION', 'None', (23, 28)) ('V617F', 'Var', (23, 28)) ('leukemic transformation', 'Disease', (75, 98)) ('leukemic transformation', 'Disease', 'MESH:D007938', (75, 98)) 5686 31086779 However, some groups have identified the IDH2 mutation in a subset of patients with MPN who underwent leukemic transformation. ('leukemic transformation', 'Disease', (102, 125)) ('patients', 'Species', '9606', (70, 78)) ('leukemic transformation', 'Disease', 'MESH:D007938', (102, 125)) ('IDH2', 'Gene', (41, 45)) ('mutation', 'Var', (46, 54)) ('MPN', 'Disease', (84, 87)) 5687 31086779 In addition, combined expression of JAK2 V617F and IDH2 R140Q induces MPN progression, alters stem/progenitor cell function and impairs differentiation in mice. ('V617F', 'SUBSTITUTION', 'None', (41, 46)) ('MPN progression', 'CPA', (70, 85)) ('R140Q', 'Mutation', 'rs121913502', (56, 61)) ('stem/progenitor cell function', 'CPA', (94, 123)) ('induces', 'Reg', (62, 69)) ('V617F', 'Var', (41, 46)) ('mice', 'Species', '10090', (155, 159)) ('IDH2', 'Gene', (51, 55)) ('impairs', 'NegReg', (128, 135)) ('differentiation', 'CPA', (136, 151)) ('alters', 'Reg', (87, 93)) 5689 31086779 Previous studies suggest that monoallelic mutations in CEBPA are prognostically neutral, whereas biallelic mutations in trans are associated with a favorable prognosis in cytogenetically normal AML. ('biallelic mutations', 'Var', (97, 116)) ('CEBPA', 'Gene', '1050', (55, 60)) ('AML', 'Disease', 'MESH:D015470', (194, 197)) ('AML', 'Disease', (194, 197)) ('AML', 'Phenotype', 'HP:0004808', (194, 197)) ('monoallelic mutations', 'Var', (30, 51)) ('CEBPA', 'Gene', (55, 60)) 5691 31086779 JAK2 mutations are rare in de novo AML, and AML patients with JAK2 mutations usually have a history of an antecedent MPN. ('AML', 'Disease', (35, 38)) ('AML', 'Disease', 'MESH:D015470', (44, 47)) ('mutations', 'Var', (67, 76)) ('AML', 'Disease', (44, 47)) ('patients', 'Species', '9606', (48, 56)) ('AML', 'Phenotype', 'HP:0004808', (44, 47)) ('AML', 'Disease', 'MESH:D015470', (35, 38)) ('JAK2', 'Gene', (62, 66)) ('AML', 'Phenotype', 'HP:0004808', (35, 38)) 5693 31086779 Despite the infrequent incidence of JAK2 mutation in AML, mutated JAK2 could be therapeutically targeted in some cases with JAK2 mutations. ('JAK2', 'Gene', (36, 40)) ('AML', 'Disease', 'MESH:D015470', (53, 56)) ('mutation', 'Var', (41, 49)) ('AML', 'Disease', (53, 56)) ('AML', 'Phenotype', 'HP:0004808', (53, 56)) ('JAK2', 'Gene', (66, 70)) 5694 31086779 SNPa and NGS added valuable information to these analyses, as both cases exhibited leukemogenic collaboration of the detected mutations: one had JAK2 and IDH2 mutations, being most likely an MPN with leukemic transformation, while the other could be a de novo AML with JAK2, CEBPA, and FLT3 mutations. ('JAK2', 'Gene', (145, 149)) ('FLT3', 'Gene', '2322', (286, 290)) ('exhibited', 'Reg', (73, 82)) ('CEBPA', 'Gene', '1050', (275, 280)) ('FLT3', 'Gene', (286, 290)) ('mutations', 'Var', (159, 168)) ('AML', 'Disease', 'MESH:D015470', (260, 263)) ('leukemogenic collaboration', 'Disease', (83, 109)) ('leukemic transformation', 'Disease', (200, 223)) ('IDH2', 'Gene', (154, 158)) ('AML', 'Phenotype', 'HP:0004808', (260, 263)) ('AML', 'Disease', (260, 263)) ('mutations', 'Var', (126, 135)) ('leukemic transformation', 'Disease', 'MESH:D007938', (200, 223)) ('CEBPA', 'Gene', (275, 280)) ('leukemogenic collaboration', 'Disease', 'MESH:C536035', (83, 109)) 5697 30623626 FLT3 Internal Tandem Duplication in Patients With Acute Myeloid Leukemia Is Readily Detectable in a Single Next-Generation Sequencing Assay Using the Pindel Algorithm Dear Editor, Next-generation sequencing (NGS) is rapidly being adapted in clinical practice, and numerous clinical laboratories are using this technology to assess patients with acute myeloid leukemia (AML). ('AML', 'Disease', 'MESH:D015470', (369, 372)) ('AML', 'Disease', (369, 372)) ('AML', 'Phenotype', 'HP:0004808', (369, 372)) ('patients', 'Species', '9606', (331, 339)) ('Acute Myeloid Leukemia', 'Disease', 'MESH:D015470', (50, 72)) ('Leukemia', 'Phenotype', 'HP:0001909', (64, 72)) ('Acute Myeloid Leukemia', 'Phenotype', 'HP:0004808', (50, 72)) ('acute myeloid leukemia', 'Disease', (345, 367)) ('Myeloid Leukemia', 'Phenotype', 'HP:0012324', (56, 72)) ('Acute Myeloid Leukemia', 'Disease', (50, 72)) ('Pindel', 'Chemical', '-', (150, 156)) ('Internal Tandem Duplication', 'Var', (5, 32)) ('FLT3', 'Gene', (0, 4)) ('Patients', 'Species', '9606', (36, 44)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (345, 367)) ('leukemia', 'Phenotype', 'HP:0001909', (359, 367)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (345, 367)) ('FLT3', 'Gene', '2322', (0, 4)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (351, 367)) 5698 30623626 Among recurrent somatic mutations in AML, the FLT3 internal tandem duplication (ITD) has major clinical implications and is associated with adverse outcomes. ('FLT3', 'Gene', '2322', (46, 50)) ('AML', 'Disease', 'MESH:D015470', (37, 40)) ('internal tandem duplication', 'Var', (51, 78)) ('FLT3', 'Gene', (46, 50)) ('associated with', 'Reg', (124, 139)) ('AML', 'Phenotype', 'HP:0004808', (37, 40)) ('AML', 'Disease', (37, 40)) 5699 30623626 Furthermore, as activated kinase targets such as FLT3 and JAK are under active clinical investigation, it has become increasingly important to identify activating FLT3 mutations. ('FLT3', 'Gene', '2322', (163, 167)) ('FLT3', 'Gene', (49, 53)) ('FLT3', 'Gene', (163, 167)) ('FLT3', 'Gene', '2322', (49, 53)) ('mutations', 'Var', (168, 177)) 5703 30623626 To determine whether FLT3 ITD mutations could be accurately identified from targeted multigene NGS data, we analyzed the NGS data with different algorithms and compared the results with those of PCR and fragment analysis, a current standard test. ('FLT3', 'Gene', (21, 25)) ('FLT3', 'Gene', '2322', (21, 25)) ('mutations', 'Var', (30, 39)) 5705 30623626 GATK MuTect2 (3.8-0), and Pindel (0.2.0) algorithms were used to detect FLT3 ITD mutations, which were implemented in the DxSeq algorithm (Dxome, Seoul, Korea). ('FLT3', 'Gene', (72, 76)) ('FLT3', 'Gene', '2322', (72, 76)) ('mutations', 'Var', (81, 90)) 5707 30623626 GATK MuTect2 detected FLT3 ITD mutations in 15 of 30 cases, most of which had a relatively short duplicated segment. ('FLT3', 'Gene', '2322', (22, 26)) ('mutations', 'Var', (31, 40)) ('FLT3', 'Gene', (22, 26)) 5708 30623626 The Pindel algorithm detected FLT3 ITD mutations in all cases. ('FLT3', 'Gene', (30, 34)) ('FLT3', 'Gene', '2322', (30, 34)) ('mutations', 'Var', (39, 48)) 5709 30623626 Inclusion of these reads is critical for identifying medium-size insertions, such as those in FLT3 ITD mutations, because this size range is too large to be detected on the basis of indels within single reads and is often too small for detection by analysis of discordant insert sizes. ('mutations', 'Var', (103, 112)) ('FLT3', 'Gene', '2322', (94, 98)) ('FLT3', 'Gene', (94, 98)) 5712 30623626 Although the true mutant allele burden could be deduced from the variant allele frequency of each tool, the existence of a larger portion of leukemic cells harboring the mutation warrants consideration. ('leukemic', 'Disease', 'MESH:D007938', (141, 149)) ('leukemic', 'Disease', (141, 149)) ('mutation', 'Var', (170, 178)) 5713 30623626 In conclusion, the Pindel algorithm was highly effective in detecting FLT3 ITD mutations in the NGS assessment of AML patients and was superior to GATK MuTect2. ('AML', 'Disease', (114, 117)) ('AML', 'Phenotype', 'HP:0004808', (114, 117)) ('FLT3', 'Gene', (70, 74)) ('FLT3', 'Gene', '2322', (70, 74)) ('AML', 'Disease', 'MESH:D015470', (114, 117)) ('patients', 'Species', '9606', (118, 126)) ('mutations', 'Var', (79, 88)) 5714 30623626 Although the mutant allele fraction calculated tended to be lower than that obtained by PCR fragment analysis, Pindel has clinical utility for identifying clinically significant FLT3 ITD mutations in a single NGS analysis. ('mutations', 'Var', (187, 196)) ('FLT3', 'Gene', (178, 182)) ('FLT3', 'Gene', '2322', (178, 182)) 5748 29560000 Real-time PCR was performed using an ABI Step One Plus System (Applied Biosystems, Foster City, CA) with standard reagents (TaqMan Gene Expression Master Mix, Applied Biosystems, UK), which showed positive results for CMV (5000 copy/ml). ('CMV (5000 copy/ml', 'Var', (218, 235)) ('Mix', 'Gene', (154, 157)) ('Mix', 'Gene', '83881', (154, 157)) 5763 29560000 Although available data have been confirmed direct association between high CMV viral load and development of CMV retinitis in patients with CMV viremia after hematopoietic stem cell transplantation (HSCT), there is lack of enough evidence to support this relationship in leukemia. ('CMV retinitis', 'Disease', 'MESH:D012173', (110, 123)) ('high', 'Var', (71, 75)) ('leukemia', 'Phenotype', 'HP:0001909', (272, 280)) ('retinitis', 'Phenotype', 'HP:0032118', (114, 123)) ('leukemia', 'Disease', 'MESH:D007938', (272, 280)) ('viremia', 'Phenotype', 'HP:0020071', (145, 152)) ('CMV viremia', 'Phenotype', 'HP:0032247', (141, 152)) ('patients', 'Species', '9606', (127, 135)) ('CMV retinitis', 'Disease', (110, 123)) ('leukemia', 'Disease', (272, 280)) ('CMV viremia', 'Disease', 'MESH:D014766', (141, 152)) ('CMV viremia', 'Disease', (141, 152)) 5772 29320732 Here, we report that stabilization of NF-kappaB-inducing kinase (NIK) suppresses AML. ('NF-kappaB-inducing kinase', 'Gene', '53859', (38, 63)) ('AML', 'Disease', 'MESH:D015470', (81, 84)) ('AML', 'Phenotype', 'HP:0004808', (81, 84)) ('AML', 'Disease', (81, 84)) ('stabilization', 'Var', (21, 34)) ('NF-kappaB-inducing kinase', 'Gene', (38, 63)) ('suppresses', 'NegReg', (70, 80)) 5773 29320732 Mechanistically, stabilization of NIK activates NF-kappaB non-canonical signaling and represses NF-kappaB canonical signaling. ('NF-kappaB', 'Gene', '18033', (48, 57)) ('NF-kappaB', 'Gene', '18033', (96, 105)) ('represses', 'NegReg', (86, 95)) ('stabilization', 'Var', (17, 30)) ('NIK', 'Gene', (34, 37)) ('activates', 'PosReg', (38, 47)) ('NF-kappaB', 'Gene', (48, 57)) ('NF-kappaB', 'Gene', (96, 105)) 5776 29320732 Our data provide a scientific rationale for developing small molecules to stabilize NIK specifically in myeloid leukemias as an attractive therapeutic option. ('leukemia', 'Phenotype', 'HP:0001909', (112, 120)) ('myeloid leukemias', 'Disease', (104, 121)) ('myeloid leukemias', 'Phenotype', 'HP:0012324', (104, 121)) ('NIK', 'Protein', (84, 87)) ('leukemias', 'Phenotype', 'HP:0001909', (112, 121)) ('stabilize', 'Var', (74, 83)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (104, 120)) ('myeloid leukemias', 'Disease', 'MESH:D007951', (104, 121)) 5782 29320732 Suppression of canonical signaling by ectopic expression of the so-called super-repressor form of IkappaBalpha attenuates AML development, validating targeting the pathway as potential therapy in AML in combination with current treatment. ('AML', 'Phenotype', 'HP:0004808', (122, 125)) ('attenuates', 'NegReg', (111, 121)) ('AML', 'Phenotype', 'HP:0004808', (196, 199)) ('AML', 'Disease', 'MESH:D015470', (196, 199)) ('ectopic expression', 'Var', (38, 56)) ('canonical signaling', 'Pathway', (15, 34)) ('AML', 'Disease', 'MESH:D015470', (122, 125)) ('Suppression', 'NegReg', (0, 11)) ('AML', 'Disease', (196, 199)) ('IkappaBalpha', 'Gene', (98, 110)) ('AML', 'Disease', (122, 125)) ('IkappaBalpha', 'Gene', '18035', (98, 110)) 5785 29320732 In contrast, using a biphenotypic B-myelomonocytic leukemia cell line with lymphoblastic morphology, expression of RelB was shown to repress a tumor suppressor, death-associated protein kinase 1. ('tumor', 'Disease', (143, 148)) ('tumor', 'Phenotype', 'HP:0002664', (143, 148)) ('leukemia', 'Phenotype', 'HP:0001909', (51, 59)) ('expression', 'Var', (101, 111)) ('death-associated protein kinase 1', 'Gene', (161, 194)) ('leukemia', 'Disease', 'MESH:D007938', (51, 59)) ('death-associated protein kinase 1', 'Gene', '69635', (161, 194)) ('B-myelomonocytic leukemia', 'Phenotype', 'HP:0004812', (34, 59)) ('leukemia', 'Disease', (51, 59)) ('RelB', 'Gene', '19698', (115, 119)) ('RelB', 'Gene', (115, 119)) ('tumor', 'Disease', 'MESH:D009369', (143, 148)) ('repress', 'NegReg', (133, 140)) 5787 29320732 NIK activates NF-kappaB non-canonical signaling by directly phosphorylating IKKalpha, which, in turn, phosphorylates p100 and induces its processing to p52, facilitating the formation and nuclear translocation of RelB/p52 complexes. ('processing', 'MPA', (138, 148)) ('RelB', 'Gene', (213, 217)) ('IKKalpha', 'Gene', '12675', (76, 84)) ('NF-kappaB', 'Gene', '18033', (14, 23)) ('p52', 'Gene', '18034', (152, 155)) ('induces', 'Reg', (126, 133)) ('IKKalpha', 'Gene', (76, 84)) ('formation', 'MPA', (174, 183)) ('RelB', 'Gene', '19698', (213, 217)) ('p52', 'Gene', '18034', (218, 221)) ('facilitating', 'PosReg', (157, 169)) ('p52', 'Gene', (152, 155)) ('NF-kappaB', 'Gene', (14, 23)) ('p100', 'Var', (117, 121)) ('nuclear translocation', 'MPA', (188, 209)) ('activates', 'PosReg', (4, 13)) ('p52', 'Gene', (218, 221)) 5789 29320732 To constitutively or conditionally stabilize NIK in the hematopoietic system, mice carrying a NIKDeltaT3flSTOP allele were crossed with either Vav-Cre mice (hereinafter termed caNIK) or Rosa-CreERT2 mice (hereinafter termed NIKERT2). ('NIKDeltaT3flSTOP', 'Var', (94, 110)) ('ERT2', 'Gene', '26417', (194, 198)) ('ERT2', 'Gene', (227, 231)) ('mice', 'Species', '10090', (199, 203)) ('mice', 'Species', '10090', (151, 155)) ('Vav', 'Gene', (143, 146)) ('Vav', 'Gene', '22324', (143, 146)) ('ERT2', 'Gene', (194, 198)) ('ERT2', 'Gene', '26417', (227, 231)) ('mice', 'Species', '10090', (78, 82)) 5790 29320732 We used these mutant mice in combination with the well-characterized MLL-AF9-induced AML mouse model to investigate the role of NIK-induced non-canonical signaling in AML. ('mice', 'Species', '10090', (21, 25)) ('mutant', 'Var', (14, 20)) ('AML', 'Disease', 'MESH:D015470', (85, 88)) ('AML', 'Disease', 'MESH:D015470', (167, 170)) ('AML', 'Phenotype', 'HP:0004808', (85, 88)) ('AML', 'Disease', (85, 88)) ('AML', 'Disease', (167, 170)) ('AML', 'Phenotype', 'HP:0004808', (167, 170)) ('mouse', 'Species', '10090', (89, 94)) 5791 29320732 Unexpectedly, we found that stabilization of NIK suppressed MLL-AF9-induced AML, which is different from the tumor-promoting role of NIK in B cell neoplasms. ('neoplasms', 'Disease', 'MESH:D009369', (147, 156)) ('neoplasms', 'Disease', (147, 156)) ('AML', 'Disease', (76, 79)) ('AML', 'Phenotype', 'HP:0004808', (76, 79)) ('tumor', 'Disease', 'MESH:D009369', (109, 114)) ('stabilization', 'Var', (28, 41)) ('suppressed', 'NegReg', (49, 59)) ('NIK', 'Gene', (45, 48)) ('neoplasms', 'Phenotype', 'HP:0002664', (147, 156)) ('tumor', 'Phenotype', 'HP:0002664', (109, 114)) ('B cell neoplasms', 'Phenotype', 'HP:0012191', (140, 156)) ('AML', 'Disease', 'MESH:D015470', (76, 79)) ('tumor', 'Disease', (109, 114)) 5802 29320732 These results suggest that NIK impairs the initiation of MLL-AF9-induced AML, which encouraged us to further investigate whether NIK has a similar effect on fully developed AML. ('NIK', 'Var', (27, 30)) ('AML', 'Disease', (73, 76)) ('AML', 'Phenotype', 'HP:0004808', (173, 176)) ('AML', 'Phenotype', 'HP:0004808', (73, 76)) ('AML', 'Disease', (173, 176)) ('initiation', 'MPA', (43, 53)) ('impairs', 'NegReg', (31, 38)) ('AML', 'Disease', 'MESH:D015470', (173, 176)) ('AML', 'Disease', 'MESH:D015470', (73, 76)) 5810 29320732 Most importantly, stabilization of NIK has a broad anti-human myeloid leukemia role (Figure S2). ('NIK', 'Protein', (35, 38)) ('myeloid leukemia', 'Disease', (62, 78)) ('human', 'Species', '9606', (56, 61)) ('leukemia', 'Phenotype', 'HP:0001909', (70, 78)) ('stabilization', 'Var', (18, 31)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (62, 78)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (62, 78)) 5821 29320732 In addition, stabilization of NIK upregulated Notch signaling, which has been show to inhibit MLL-AF9-induced AML (Figures S3B and S3C). ('AML', 'Phenotype', 'HP:0004808', (110, 113)) ('Notch signaling', 'MPA', (46, 61)) ('NIK', 'Gene', (30, 33)) ('AML', 'Disease', (110, 113)) ('upregulated', 'PosReg', (34, 45)) ('stabilization', 'Var', (13, 26)) ('inhibit', 'NegReg', (86, 93)) ('AML', 'Disease', 'MESH:D015470', (110, 113)) 5823 29320732 NIK protein was ~2-fold increased in NIK-stabilized AML cells and was associated with increased IKKalpha phosphorylation, upregulation of RelB, and activation of p100 processing to p52 (Figure 3A). ('phosphorylation', 'MPA', (105, 120)) ('p100', 'Protein', (162, 166)) ('increased', 'PosReg', (86, 95)) ('AML', 'Phenotype', 'HP:0004808', (52, 55)) ('p52', 'Gene', (181, 184)) ('activation', 'PosReg', (148, 158)) ('AML', 'Disease', (52, 55)) ('RelB', 'Gene', (138, 142)) ('NIK-stabilized', 'Var', (37, 51)) ('RelB', 'Gene', '19698', (138, 142)) ('IKKalpha', 'Gene', '12675', (96, 104)) ('upregulation', 'PosReg', (122, 134)) ('p52', 'Gene', '18034', (181, 184)) ('IKKalpha', 'Gene', (96, 104)) ('increased', 'PosReg', (24, 33)) ('AML', 'Disease', 'MESH:D015470', (52, 55)) 5827 29320732 It has been suggested that inhibition of canonical NF-kappaB signaling leads to robust anti-leukemia effects, phenocopying loss of functional MLL oncoprotein. ('leukemia', 'Disease', (92, 100)) ('leukemia', 'Phenotype', 'HP:0001909', (92, 100)) ('leukemia', 'Disease', 'MESH:D007938', (92, 100)) ('NF-kappaB', 'Gene', '18033', (51, 60)) ('inhibition', 'Var', (27, 37)) ('NF-kappaB', 'Gene', (51, 60)) ('MLL', 'Protein', (142, 145)) 5828 29320732 However, overexpression of RelA, confirmed by real-time PCR and western blot, failed to restore the reduced colony formation and the delay in leukemogenesis caused by NIK stabilization (Figures S4A-S4D). ('delay', 'NegReg', (133, 138)) ('NIK stabilization', 'Var', (167, 184)) ('RelA', 'Gene', '19697', (27, 31)) ('reduced', 'NegReg', (100, 107)) ('RelA', 'Gene', (27, 31)) ('leukemogenesis', 'Disease', (142, 156)) ('colony formation', 'CPA', (108, 124)) 5831 29320732 The latter is consistent with RelA overexpression, which accelerates AML with MLL mutations. ('AML', 'Phenotype', 'HP:0004808', (69, 72)) ('MLL', 'Gene', (78, 81)) ('AML', 'Disease', (69, 72)) ('accelerates', 'PosReg', (57, 68)) ('RelA', 'Gene', '19697', (30, 34)) ('RelA', 'Gene', (30, 34)) ('mutations', 'Var', (82, 91)) ('AML', 'Disease', 'MESH:D015470', (69, 72)) 5837 29320732 The inactivating DNMT3A mutation enhances HSPC selfrenewal and is a driver mutation in about 30% of AML. ('AML', 'Phenotype', 'HP:0004808', (100, 103)) ('enhances', 'PosReg', (33, 41)) ('HSPC', 'Gene', '5688', (42, 46)) ('HSPC', 'Gene', (42, 46)) ('DNMT3A', 'Gene', (17, 23)) ('DNMT3A', 'Gene', '13435', (17, 23)) ('AML', 'Disease', 'MESH:D015470', (100, 103)) ('mutation', 'Var', (24, 32)) ('inactivating', 'Var', (4, 16)) ('AML', 'Disease', (100, 103)) 5839 29320732 In addition, Mef2c is a known MLL-AF9 downstream target, and deletion of Mef2c impairs MLL-AF9-induced leukemogenesis. ('impairs MLL-AF9-induced leukemogenesis', 'Disease', 'MESH:D009422', (79, 117)) ('deletion', 'Var', (61, 69)) ('Mef2c', 'Gene', '17260', (73, 78)) ('Mef2c', 'Gene', (13, 18)) ('impairs MLL-AF9-induced leukemogenesis', 'Disease', (79, 117)) ('Mef2c', 'Gene', (73, 78)) ('Mef2c', 'Gene', '17260', (13, 18)) 5841 29320732 We found that RelB directly binds to Mef2c and one of the Dnmt3a promoters in front of exon 7, which functionally regulates the expression of an active DNMT3a variant, but not Tifab promoter (Figure 3E). ('regulates', 'Reg', (114, 123)) ('Mef2c', 'Gene', (37, 42)) ('RelB', 'Gene', (14, 18)) ('Dnmt3a', 'Gene', '13435', (58, 64)) ('Dnmt3a', 'Gene', (58, 64)) ('RelB', 'Gene', '19698', (14, 18)) ('active', 'MPA', (145, 151)) ('DNMT3a', 'Gene', '13435', (152, 158)) ('Mef2c', 'Gene', '17260', (37, 42)) ('expression', 'MPA', (128, 138)) ('DNMT3a', 'Gene', (152, 158)) ('variant', 'Var', (159, 166)) 5844 29320732 In contrast, knockdown of Relb partially restored NIK's anti-leukemic effect, suggesting that upregulation of DNMT3a and downregulation of MEF2C, along with other RelB-regulated molecules, contribute to AML suppression (Figure 3H). ('Relb', 'Gene', '19698', (26, 30)) ('AML', 'Phenotype', 'HP:0004808', (203, 206)) ('leukemic', 'Disease', 'MESH:D007938', (61, 69)) ('upregulation', 'PosReg', (94, 106)) ('leukemic', 'Disease', (61, 69)) ('DNMT3a', 'Gene', '13435', (110, 116)) ('RelB', 'Gene', (163, 167)) ('MEF2C', 'Gene', '17260', (139, 144)) ('DNMT3a', 'Gene', (110, 116)) ('MEF2C', 'Gene', (139, 144)) ('AML', 'Disease', 'MESH:D015470', (203, 206)) ('downregulation', 'NegReg', (121, 135)) ('RelB', 'Gene', '19698', (163, 167)) ('knockdown', 'Var', (13, 22)) ('AML', 'Disease', (203, 206)) ('Relb', 'Gene', (26, 30)) 5847 29320732 We noted that NIK impaired the function of HSPCs and LSCs through different sets of genes. ('HSPC', 'Gene', '5688', (43, 47)) ('function', 'MPA', (31, 39)) ('LSCs', 'Disease', (53, 57)) ('HSPC', 'Gene', (43, 47)) ('NIK', 'Var', (14, 17)) ('impaired', 'NegReg', (18, 26)) 5856 29320732 Importantly, the inhibitory effect of verteporfin was largely attenuated by knockdown of RelB and has no additive effects on NIK-stabilized AML cells (Figure 4I). ('RelB', 'Gene', (89, 93)) ('knockdown', 'Var', (76, 85)) ('attenuated', 'NegReg', (62, 72)) ('RelB', 'Gene', '19698', (89, 93)) ('AML', 'Disease', 'MESH:D015470', (140, 143)) ('verteporfin', 'Chemical', 'MESH:D000077362', (38, 49)) ('AML', 'Disease', (140, 143)) ('inhibitory effect', 'MPA', (17, 34)) ('AML', 'Phenotype', 'HP:0004808', (140, 143)) 5859 29320732 In this study, we have shown that stabilization of NIK suppresses myeloid leukemia through the activation of NF-kappaB non-canonical signaling and simultaneous repression of NF-kappaB canonical signaling. ('repression', 'NegReg', (160, 170)) ('NF-kappaB', 'Gene', (109, 118)) ('stabilization', 'Var', (34, 47)) ('NF-kappaB', 'Gene', (174, 183)) ('activation', 'PosReg', (95, 105)) ('myeloid leukemia', 'Disease', (66, 82)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (66, 82)) ('NF-kappaB', 'Gene', '18033', (109, 118)) ('NIK', 'Gene', (51, 54)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (66, 82)) ('suppresses', 'NegReg', (55, 65)) ('leukemia', 'Phenotype', 'HP:0001909', (74, 82)) ('NF-kappaB', 'Gene', '18033', (174, 183)) 5864 29320732 Future studies are required to determine the genes differentially regulated by RelA and RelB in AML and whether deletion of RelB will enhance AML development and block stabilization of NIK-induced impaired leukemogenesis using genetic RelB-deficient or NIK-stabilized/RelB-deficient compound mutant mice. ('RelA', 'Gene', '19697', (79, 83)) ('mice', 'Species', '10090', (299, 303)) ('deletion', 'Var', (112, 120)) ('RelB', 'Gene', (88, 92)) ('AML', 'Disease', 'MESH:D015470', (96, 99)) ('RelB', 'Gene', '19698', (88, 92)) ('AML', 'Phenotype', 'HP:0004808', (96, 99)) ('AML', 'Disease', (96, 99)) ('impaired leukemogenesis', 'Disease', 'MESH:D009422', (197, 220)) ('impaired leukemogenesis', 'Disease', (197, 220)) ('AML', 'Disease', 'MESH:D015470', (142, 145)) ('AML', 'Disease', (142, 145)) ('RelB', 'Gene', (124, 128)) ('AML', 'Phenotype', 'HP:0004808', (142, 145)) ('RelB', 'Gene', '19698', (268, 272)) ('RelB', 'Gene', (268, 272)) ('RelB', 'Gene', '19698', (124, 128)) ('RelB', 'Gene', (235, 239)) ('RelA', 'Gene', (79, 83)) ('RelB', 'Gene', '19698', (235, 239)) ('enhance', 'PosReg', (134, 141)) 5865 29320732 Nevertheless, the key finding of the present study is that AML is significantly suppressed by stabilization of NIK protein. ('AML', 'Phenotype', 'HP:0004808', (59, 62)) ('AML', 'Disease', 'MESH:D015470', (59, 62)) ('NIK protein', 'Protein', (111, 122)) ('suppressed', 'NegReg', (80, 90)) ('stabilization', 'Var', (94, 107)) ('AML', 'Disease', (59, 62)) 5866 29320732 Although activating mutations in components of both canonical and non-canonical NF-kappaB signaling are frequently detected in B cell neoplasms, no gain- or loss-of-function mutations in canonical and non-canonical signaling have been reported in AML. ('loss-of-function', 'NegReg', (157, 173)) ('NF-kappaB', 'Gene', (80, 89)) ('neoplasms', 'Phenotype', 'HP:0002664', (134, 143)) ('NF-kappaB', 'Gene', '18033', (80, 89)) ('AML', 'Phenotype', 'HP:0004808', (247, 250)) ('B cell', 'Disease', (127, 133)) ('gain-', 'PosReg', (148, 153)) ('activating', 'PosReg', (9, 19)) ('AML', 'Disease', 'MESH:D015470', (247, 250)) ('B cell neoplasms', 'Phenotype', 'HP:0012191', (127, 143)) ('neoplasms', 'Disease', 'MESH:D009369', (134, 143)) ('neoplasms', 'Disease', (134, 143)) ('AML', 'Disease', (247, 250)) ('mutations', 'Var', (20, 29)) 5869 29320732 We have shown that either blocking or activating non-canonical signaling impairs HSPC self-renewal, and other research has shown that deletion of NIK also impairs HSPC function through non-canonical signaling. ('HSPC', 'Gene', '5688', (81, 85)) ('activating', 'MPA', (38, 48)) ('impairs', 'NegReg', (155, 162)) ('non-canonical', 'Protein', (49, 62)) ('deletion', 'Var', (134, 142)) ('impairs', 'NegReg', (73, 80)) ('HSPC', 'Gene', (163, 167)) ('HSPC', 'Gene', '5688', (163, 167)) ('non-canonical', 'MPA', (185, 198)) ('NIK', 'Gene', (146, 149)) ('HSPC', 'Gene', (81, 85)) 5871 29320732 However, stem cells whose ability to self-renew is impaired are not "good targets" for leukemic transformation, and we speculate that this is why no loss-of-function mutations in non-canonical signaling components have been detected. ('leukemic transformation', 'Disease', (87, 110)) ('leukemic transformation', 'Disease', 'MESH:D007938', (87, 110)) ('mutations', 'Var', (166, 175)) 5874 29320732 Furthermore, we want to emphasize that NIK protein is constantly degraded, its expression in normal BM cells and AML blasts was very low to undetectable, and it even cannot be detected after WT NIK is overexpressed, generating a "null, naturally mutated condition" even without mutations (Figures 3A, S2D, and S2E). ('NIK', 'Var', (39, 42)) ('expression', 'MPA', (79, 89)) ('low', 'NegReg', (133, 136)) ('degraded', 'NegReg', (65, 73)) ('AML', 'Disease', 'MESH:D015470', (113, 116)) ('AML', 'Phenotype', 'HP:0004808', (113, 116)) ('AML', 'Disease', (113, 116)) ('protein', 'Protein', (43, 50)) 5891 29080039 FLT3ITD-harboring AML cell lines and primary cells treated with the FLT3 inhibitor PKC412 displayed a rapid decline in the levels of hTERT mRNA and telomerase activity. ('AML', 'Disease', (18, 21)) ('AML', 'Phenotype', 'HP:0004808', (18, 21)) ('decline', 'NegReg', (108, 115)) ('telomerase activity', 'MPA', (148, 167)) ('hTERT', 'Gene', '7015', (133, 138)) ('AML', 'Disease', 'MESH:D015470', (18, 21)) ('FLT3', 'Gene', (68, 72)) ('hTERT', 'Gene', (133, 138)) ('PKC412', 'Var', (83, 89)) 5892 29080039 Moreover, PKC412 inhibited hTERT gene transcription in a c-MYC-dependent manner. ('hTERT', 'Gene', '7015', (27, 32)) ('hTERT', 'Gene', (27, 32)) ('inhibited', 'NegReg', (17, 26)) ('PKC412', 'Var', (10, 16)) 5894 29080039 Mechanistically, hTERT enhanced the activity of FLT3 downstream effectors or alternative RTK signaling, thereby enhancing AKT phosphorylation, in AML cells treated with PKC412. ('enhanced', 'PosReg', (23, 31)) ('enhancing', 'PosReg', (112, 121)) ('FLT3', 'Gene', (48, 52)) ('PKC412', 'Var', (169, 175)) ('hTERT', 'Gene', (17, 22)) ('AML', 'Disease', (146, 149)) ('activity', 'MPA', (36, 44)) ('AML', 'Phenotype', 'HP:0004808', (146, 149)) ('AKT', 'Gene', '207', (122, 125)) ('AML', 'Disease', 'MESH:D015470', (146, 149)) ('hTERT', 'Gene', '7015', (17, 22)) ('AKT', 'Gene', (122, 125)) 5898 29080039 FMS-like tyrosine kinase 3 (FLT3), a type III receptor tyrosine kinase (RTK), is one of the most frequently mutated genes and occurs in approximately one third of AML, with the majority of mutations exhibiting the internal tandem duplication (ITD) in its justamembrance domain. ('FLT3', 'Gene', (28, 32)) ('AML', 'Phenotype', 'HP:0004808', (163, 166)) ('FMS-like tyrosine kinase 3', 'Gene', (0, 26)) ('AML', 'Disease', (163, 166)) ('FMS-like tyrosine kinase 3', 'Gene', '2322', (0, 26)) ('mutations', 'Var', (189, 198)) ('AML', 'Disease', 'MESH:D015470', (163, 166)) 5900 29080039 In addition, many AML patients who receive FLT3TKIs eventually acquire drug resistance. ('AML', 'Disease', (18, 21)) ('drug resistance', 'MPA', (71, 86)) ('AML', 'Phenotype', 'HP:0004808', (18, 21)) ('patients', 'Species', '9606', (22, 30)) ('acquire', 'Reg', (63, 70)) ('AML', 'Disease', 'MESH:D015470', (18, 21)) ('drug resistance', 'Phenotype', 'HP:0020174', (71, 86)) ('FLT3TKIs', 'Var', (43, 51)) 5901 29080039 It has been well-characterized that the occurrence of new mutations in the FLT3 genome is an important mechanism contributing to patients' resistance to FLT3TKIs. ('patients', 'Species', '9606', (129, 137)) ('mutations', 'Var', (58, 67)) ('FLT3', 'Gene', (75, 79)) 5918 29080039 Patient 2: 79 years old, diagnosed as AML with del(20) and FLT3ITD, WBCC = 161.8 x 109/l with blasts 91.5%. ('FLT3ITD', 'Gene', (59, 66)) ('AML', 'Disease', (38, 41)) ('del(20', 'Var', (47, 53)) ('AML', 'Disease', 'MESH:D015470', (38, 41)) ('Patient', 'Species', '9606', (0, 7)) ('AML', 'Phenotype', 'HP:0004808', (38, 41)) 5930 29080039 MV4,11 and MOLM-13 cells cultured in 24-well plates at 0.5 x 106 were transfected with p181wt and p181MYC- plasmids using Lipofectamine2000 (Life Technology) according to the manufacturer's protocol, followed by the treatment with PKC412. ('Lipofectamine2000', 'Chemical', 'MESH:C086724', (122, 139)) ('MOLM', 'Chemical', '-', (11, 15)) ('MYC', 'Gene', '4609', (102, 105)) ('p181wt', 'Var', (87, 93)) ('MYC', 'Gene', (102, 105)) 5938 29080039 PKC412 is a TKI specifically targeting FLT3ITD, and as expected, the PKC412 treatment of MV4,11 and MOLM-13 cells inhibited FLT3 phosphorylation and activity (Fig. ('FLT3', 'Protein', (124, 128)) ('MOLM', 'Chemical', '-', (100, 104)) ('activity', 'MPA', (149, 157)) ('PKC412', 'Var', (69, 75)) ('inhibited', 'NegReg', (114, 123)) 5939 29080039 To determine the effect of PKC412 on hTERT expression, we incubated FLT3ITD+ cells with PKC412. ('hTERT', 'Gene', (37, 42)) ('PKC412', 'Var', (88, 94)) ('hTERT', 'Gene', '7015', (37, 42)) 5940 29080039 The PCR analysis showed that MV4,11 cells displayed a time-dependent downregulation of hTERT mRNA expression in the presence of PKC412 at 0.1 muM (Fig. ('PKC412 at', 'Var', (128, 137)) ('muM', 'Gene', '56925', (142, 145)) ('hTERT', 'Gene', (87, 92)) ('muM', 'Gene', (142, 145)) ('downregulation', 'NegReg', (69, 83)) ('hTERT', 'Gene', '7015', (87, 92)) 5941 29080039 This inhibitory effect was also dose-dependent, and hTERT mRNA became undetectable in MV4,11 cells treated with PKC412 at 0.1 muM for 24 h (Fig. ('muM', 'Gene', '56925', (126, 129)) ('hTERT', 'Gene', '7015', (52, 57)) ('muM', 'Gene', (126, 129)) ('PKC412', 'Var', (112, 118)) ('hTERT', 'Gene', (52, 57)) ('undetectable', 'NegReg', (70, 82)) 5943 29080039 In addition, we treated wild-type FLT3-carrying HL60 cells and HeLa cells with PKC412 and did not observe detectable changes in hTERT mRNA levels in these cells (data not shown), which suggests that PKC412 inhibited hTERT expression via FLT3ITD. ('PKC412', 'Var', (79, 85)) ('hTERT', 'Gene', (216, 221)) ('HL60', 'CellLine', 'CVCL:0002', (48, 52)) ('hTERT', 'Gene', '7015', (128, 133)) ('hTERT', 'Gene', '7015', (216, 221)) ('PKC412', 'Var', (199, 205)) ('hTERT', 'Gene', (128, 133)) ('HeLa', 'CellLine', 'CVCL:0030', (63, 67)) ('inhibited', 'NegReg', (206, 215)) 5944 29080039 Consistent with diminished hTERT mRNA expression, telomerase activity was significantly repressed in MV4,11 and MOLM-13 cells treated with PKC412 (Fig. ('hTERT', 'Gene', '7015', (27, 32)) ('PKC412', 'Var', (139, 145)) ('hTERT', 'Gene', (27, 32)) ('telomerase', 'Enzyme', (50, 60)) ('repressed', 'NegReg', (88, 97)) ('MOLM', 'Chemical', '-', (112, 116)) 5947 29080039 1g, PKC412 treatment of these AML cells similarly led to the substantial decline in hTERT mRNA levels. ('AML', 'Disease', 'MESH:D015470', (30, 33)) ('decline', 'NegReg', (73, 80)) ('hTERT', 'Gene', '7015', (84, 89)) ('AML', 'Disease', (30, 33)) ('AML', 'Phenotype', 'HP:0004808', (30, 33)) ('hTERT', 'Gene', (84, 89)) ('PKC412', 'Var', (4, 10)) 5948 29080039 We next determined the mechanism responsible for PKC412-mediated downregulation of hTERT expression in the FLT3ITD-carrying cells. ('PKC412-mediated', 'Var', (49, 64)) ('expression', 'MPA', (89, 99)) ('hTERT', 'Gene', (83, 88)) ('downregulation', 'NegReg', (65, 79)) ('hTERT', 'Gene', '7015', (83, 88)) 5950 29080039 2a, b, there were about 50% of GFP-positive cells in the DMSO-containing culture while the presence of PKC412 led to diminished GFP+ cells. ('DMSO', 'Chemical', 'MESH:D004121', (57, 61)) ('presence', 'Var', (91, 99)) ('GFP+ cells', 'CPA', (128, 138)) ('diminished', 'NegReg', (117, 127)) ('PKC412', 'Var', (103, 109)) 5952 29080039 The hTERT promoter activity, reflected as the level of luciferase activity, was significantly inhibited in the cells exposed to PKC412 compared to the DMSO-treated ones (Fig. ('hTERT', 'Gene', (4, 9)) ('luciferase', 'Enzyme', (55, 65)) ('PKC412', 'Var', (128, 134)) ('activity', 'MPA', (66, 74)) ('DMSO', 'Chemical', 'MESH:D004121', (151, 155)) ('hTERT', 'Gene', '7015', (4, 9)) ('inhibited', 'NegReg', (94, 103)) 5953 29080039 Thus, PKC412 inhibited the hTERT transcription and clearly, the hTERT proximal promoter was sufficient for its inhibitory effect. ('hTERT', 'Gene', '7015', (27, 32)) ('hTERT', 'Gene', (64, 69)) ('hTERT', 'Gene', (27, 32)) ('inhibited', 'NegReg', (13, 22)) ('hTERT', 'Gene', '7015', (64, 69)) ('PKC412', 'Var', (6, 12)) 5955 29080039 The treatment of FLT3ITD-carrying cells with PKC412 led to a fast and robust inhibition of c-MYC mRNA and protein expression in time- and dose-dependent manners, which preceded a decline in hTERT expression (Fig. ('hTERT', 'Gene', '7015', (190, 195)) ('inhibition', 'NegReg', (77, 87)) ('hTERT', 'Gene', (190, 195)) ('PKC412', 'Var', (45, 51)) ('decline', 'NegReg', (179, 186)) 5958 29080039 2c), wt hTERT promoter activity declined significantly in PKC412-treated cells compared to that in control cells (DMSO-treated). ('DMSO', 'Chemical', 'MESH:D004121', (114, 118)) ('declined', 'NegReg', (32, 40)) ('hTERT', 'Gene', '7015', (8, 13)) ('PKC412-treated', 'Var', (58, 72)) ('hTERT', 'Gene', (8, 13)) 5959 29080039 However, PKC412 did not affect the hTERT promoter activity any longer once two MYC binding sites on the promoter were disrupted (Fig. ('disrupted', 'NegReg', (118, 127)) ('PKC412', 'Var', (9, 15)) ('hTERT', 'Gene', '7015', (35, 40)) ('MYC', 'Gene', (79, 82)) ('hTERT', 'Gene', (35, 40)) ('MYC', 'Gene', '4609', (79, 82)) 5961 29080039 Having demonstrated the downregulation of hTERT expression by PKC412 treatment in FLT3ITD-harboring AML cells, we further asked whether this PKC412 effect was associated with its AML cell killing. ('AML', 'Disease', (179, 182)) ('AML', 'Phenotype', 'HP:0004808', (100, 103)) ('hTERT', 'Gene', (42, 47)) ('AML', 'Phenotype', 'HP:0004808', (179, 182)) ('downregulation', 'NegReg', (24, 38)) ('AML', 'Disease', 'MESH:D015470', (100, 103)) ('AML', 'Disease', 'MESH:D015470', (179, 182)) ('PKC412', 'Var', (62, 68)) ('AML', 'Disease', (100, 103)) ('hTERT', 'Gene', '7015', (42, 47)) 5964 29080039 4b, IC50 was 17.2 and 34.1 muM for MOLM-13-pBMN (left panel) and MOLM-13-hTERT (right panel), respectively. ('muM', 'Gene', '56925', (27, 30)) ('MOLM-13-pBMN', 'Chemical', '-', (35, 47)) ('MOLM-13-hTERT', 'CellLine', 'CVCL:2119', (65, 78)) ('muM', 'Gene', (27, 30)) ('MOLM-13-hTERT', 'Var', (65, 78)) ('MOLM-13-pBMN', 'Var', (35, 47)) 5967 29080039 Consistent with cell counting results, the FACS analysis revealed that PKC412 induced apoptosis in 35% of these cells, and the ectopic hTERT expression significantly attenuated apoptosis of MOLM-13 cells mediated by PKC412 (18%) (MOLM-13-hTERT vs MOLM-13-pBMN cells, P = 0.02) (Fig. ('apoptosis', 'CPA', (177, 186)) ('MOLM-13-hTERT', 'CellLine', 'CVCL:2119', (230, 243)) ('MOLM', 'Chemical', '-', (230, 234)) ('hTERT', 'Gene', '7015', (135, 140)) ('PKC412', 'Var', (71, 77)) ('MOLM', 'Chemical', '-', (247, 251)) ('hTERT', 'Gene', '7015', (238, 243)) ('MOLM', 'Chemical', '-', (190, 194)) ('hTERT', 'Gene', (135, 140)) ('attenuated', 'NegReg', (166, 176)) ('apoptosis', 'CPA', (86, 95)) ('hTERT', 'Gene', (238, 243)) ('PKC412', 'Var', (216, 222)) ('MOLM-13-pBMN', 'Chemical', '-', (247, 259)) 5968 29080039 In addition, PKC412 significantly decreased cells at S and G2/M of both MOLM-13-hTERT and MOLM-13-pBMN cells (Fig. ('cells at S', 'CPA', (44, 54)) ('decreased', 'NegReg', (34, 43)) ('MOLM-13-hTERT', 'CellLine', 'CVCL:2119', (72, 85)) ('MOLM-13-pBMN', 'Chemical', '-', (90, 102)) ('PKC412', 'Var', (13, 19)) 5970 29080039 Intriguingly, we identified that the ectopic hTERT expression significantly affected FLT3 and other RTK signaling pathways (Fig. ('hTERT', 'Gene', '7015', (45, 50)) ('ectopic', 'Var', (37, 44)) ('hTERT', 'Gene', (45, 50)) ('RTK signaling pathways', 'Pathway', (100, 122)) ('FLT3', 'Gene', (85, 89)) ('affected', 'Reg', (76, 84)) 5973 29080039 Second, DOC3, an endogenous inhibitor of the RSA-MAPK signaling, was downregulated in MOLM-13-hTERT cells and the PKC412 treatment led to further dramatic decline in DOC3 levels (Fig. ('DOC3', 'Gene', (8, 12)) ('decline', 'NegReg', (155, 162)) ('PKC412', 'Var', (114, 120)) ('MOLM-13-hTERT', 'CellLine', 'CVCL:2119', (86, 99)) ('downregulated', 'NegReg', (69, 82)) ('DOC3 levels', 'MPA', (166, 177)) 5974 29080039 Finally, SULF2 which activates the PDGF signaling pathway exhibited enhanced expression in MOLM-13-hTERT cells and its robust increase was observed following PKC412 treatment of these cells, whereas there was no detectable alteration in its expression in MOLM-13-pBMN cells with and without PKC412 (Fig. ('expression', 'MPA', (77, 87)) ('enhanced', 'PosReg', (68, 76)) ('MOLM-13-pBMN', 'Chemical', '-', (255, 267)) ('SULF2', 'Gene', (9, 14)) ('PDGF signaling pathway', 'Pathway', (35, 57)) ('PKC412', 'Var', (158, 164)) ('increase', 'PosReg', (126, 134)) ('SULF2', 'Gene', '55959', (9, 14)) ('MOLM-13-hTERT', 'CellLine', 'CVCL:2119', (91, 104)) ('activates', 'PosReg', (21, 30)) 5976 29080039 Given all the above observations, we determined the AKT Ser473 phosphorylation between MOLM-13-pBMN and MOLM-13-hTERT cells with and without PKC412. ('Ser473', 'Chemical', '-', (56, 62)) ('MOLM-13-pBMN', 'Chemical', '-', (87, 99)) ('AKT', 'Gene', (52, 55)) ('MOLM-13-hTERT', 'CellLine', 'CVCL:2119', (104, 117)) ('phosphorylation', 'MPA', (63, 78)) ('Ser473', 'Var', (56, 62)) ('AKT', 'Gene', '207', (52, 55)) 5979 29080039 Inhibiting FLT3ITD by a specific TKI PKC412 led to diminished hTERT expression and telomerase activity. ('FLT3ITD', 'Gene', (11, 18)) ('telomerase', 'CPA', (83, 93)) ('Inhibiting', 'NegReg', (0, 10)) ('PKC412', 'Var', (37, 43)) ('hTERT', 'Gene', (62, 67)) ('diminished', 'NegReg', (51, 61)) ('hTERT', 'Gene', '7015', (62, 67)) 5980 29080039 PKC412-mediated downregulation of hTERT is likely important for apoptotic cell death induced by FLT3ITD inhibition, because ectopic hTERT expression significantly increased cell survival of PKC412-treated MOLM-13 cells. ('hTERT', 'Gene', (34, 39)) ('increased', 'PosReg', (163, 172)) ('MOLM', 'Chemical', '-', (205, 209)) ('hTERT', 'Gene', '7015', (132, 137)) ('hTERT', 'Gene', (132, 137)) ('cell survival', 'CPA', (173, 186)) ('ectopic', 'Var', (124, 131)) ('hTERT', 'Gene', '7015', (34, 39)) ('FLT3ITD', 'Gene', (96, 103)) 5981 29080039 Collectively, our findings reveal a functional link between the mutant FLT3 and hTERT or telomerase, which may be implicated in AML pathogenesis and therapy. ('AML', 'Disease', (128, 131)) ('AML', 'Phenotype', 'HP:0004808', (128, 131)) ('hTERT', 'Gene', '7015', (80, 85)) ('hTERT', 'Gene', (80, 85)) ('FLT3', 'Gene', (71, 75)) ('telomerase', 'Enzyme', (89, 99)) ('AML', 'Disease', 'MESH:D015470', (128, 131)) ('mutant', 'Var', (64, 70)) ('implicated', 'Reg', (114, 124)) 5987 29080039 Of note, the PKC412 effect occurred only in FLT3ITD-carrying MOLM-13 and MV4-11 cell lines but not in HL60 or HeLa cells bearing wt FLT3. ('FLT3ITD-carrying', 'Var', (44, 60)) ('PKC412', 'Var', (13, 19)) ('MOLM', 'Chemical', '-', (61, 65)) ('HeLa', 'CellLine', 'CVCL:0030', (110, 114)) ('HL60', 'CellLine', 'CVCL:0002', (102, 106)) 5988 29080039 Although MOLM-13 and MV4-11 cell lines also harbor MLL mutations, there is no evidence that PKC412 interfere with the MLL pathway. ('mutations', 'Var', (55, 64)) ('interfere', 'NegReg', (99, 108)) ('MLL', 'Gene', '4297', (51, 54)) ('MOLM', 'Chemical', '-', (9, 13)) ('MLL', 'Gene', (51, 54)) ('MLL', 'Gene', (118, 121)) ('MLL', 'Gene', '4297', (118, 121)) 5993 29080039 However, it is unclear whether this is the case in targeted cancer therapy against FLT3 mutation. ('FLT3', 'Gene', (83, 87)) ('mutation', 'Var', (88, 96)) ('cancer', 'Disease', 'MESH:D009369', (60, 66)) ('cancer', 'Disease', (60, 66)) ('cancer', 'Phenotype', 'HP:0002664', (60, 66)) 5994 29080039 To address this issue, we utilized the targeted therapeutic reagent PKC412, specifically to FLT3ITD, to treat FLT3ITD-carrying AML MOLM-13 cells. ('MOLM', 'Chemical', '-', (131, 135)) ('FLT3ITD-carrying', 'Var', (110, 126)) ('AML', 'Disease', 'MESH:D015470', (127, 130)) ('AML', 'Phenotype', 'HP:0004808', (127, 130)) ('AML', 'Disease', (127, 130)) 5995 29080039 We found that the ectopic expression of hTERT significantly prevented apoptosis of MOLM-13 cells induced by PKC412, which suggests that hTERT does have a protective effect against targeted cancer therapy. ('hTERT', 'Gene', (40, 45)) ('prevented', 'NegReg', (60, 69)) ('hTERT', 'Gene', (136, 141)) ('PKC412', 'Var', (108, 114)) ('MOLM', 'Chemical', '-', (83, 87)) ('cancer', 'Phenotype', 'HP:0002664', (189, 195)) ('hTERT', 'Gene', '7015', (40, 45)) ('hTERT', 'Gene', '7015', (136, 141)) ('apoptosis', 'CPA', (70, 79)) ('cancer', 'Disease', 'MESH:D009369', (189, 195)) ('cancer', 'Disease', (189, 195)) 5996 29080039 On the other hand, PKC412 decreased S and G2/M cells independently of hTERT expression, and thus, hTERT does not affect PKC412-mediated cell cycle arrest. ('hTERT', 'Gene', '7015', (70, 75)) ('PKC412', 'Var', (19, 25)) ('hTERT', 'Gene', (70, 75)) ('hTERT', 'Gene', '7015', (98, 103)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (136, 153)) ('hTERT', 'Gene', (98, 103)) ('decreased', 'NegReg', (26, 35)) 5999 29080039 The ectopic hTERT expression substantially enhanced the expression of c-KIT, another TK receptor structurally and functionally similar to FLT3, and c-KIT level was even much higher in the presence of PKC412. ('enhanced', 'PosReg', (43, 51)) ('c-KIT', 'Gene', (70, 75)) ('hTERT', 'Gene', '7015', (12, 17)) ('higher', 'PosReg', (174, 180)) ('c-KIT', 'Gene', (148, 153)) ('expression', 'MPA', (56, 66)) ('c-KIT', 'Gene', '3815', (70, 75)) ('c-KIT', 'Gene', '3815', (148, 153)) ('hTERT', 'Gene', (12, 17)) ('PKC412', 'Var', (200, 206)) 6000 29080039 The increased c-KIT expression likely compensates for diminished AKT and MAPK activities due to FLT3ITD inhibition by PKC412. ('inhibition', 'NegReg', (104, 114)) ('PKC412', 'Var', (118, 124)) ('increased', 'PosReg', (4, 13)) ('diminished', 'NegReg', (54, 64)) ('expression', 'MPA', (20, 30)) ('c-KIT', 'Gene', (14, 19)) ('AKT', 'Gene', (65, 68)) ('c-KIT', 'Gene', '3815', (14, 19)) ('MAPK activities', 'CPA', (73, 88)) ('AKT', 'Gene', '207', (65, 68)) ('FLT3ITD', 'Gene', (96, 103)) 6001 29080039 In addition, the ectopic hTERT expression in the presence of PKC412 upregulated the expression of SULF2, an activator in the PDGF signaling pathway. ('hTERT', 'Gene', '7015', (25, 30)) ('upregulated', 'PosReg', (68, 79)) ('SULF2', 'Gene', '55959', (98, 103)) ('hTERT', 'Gene', (25, 30)) ('PKC412', 'Var', (61, 67)) ('expression', 'MPA', (84, 94)) ('SULF2', 'Gene', (98, 103)) 6002 29080039 Intriguingly, DOC3, a negative regulator of the RAS signaling pathway, was highly repressed by hTERT over-expression, especially when cells were exposed to PKC412. ('DOC3', 'Gene', (14, 18)) ('over-expression', 'Var', (101, 116)) ('hTERT', 'Gene', '7015', (95, 100)) ('hTERT', 'Gene', (95, 100)) 6003 29080039 Indeed, we observed the increased accumulation of phosphorylated AKT in MOLM-13-hTERT cells treated with PKC412. ('AKT', 'Gene', (65, 68)) ('PKC412', 'Var', (105, 111)) ('MOLM-13-hTERT', 'CellLine', 'CVCL:2119', (72, 85)) ('accumulation', 'PosReg', (34, 46)) ('phosphorylated', 'MPA', (50, 64)) ('AKT', 'Gene', '207', (65, 68)) 6005 29080039 However, it remains unclear why the upregulation of SULF2, c-KIT expression, and AKT phosphorylation occur up on PKC412 exposure in hTERT-over-expressed cells, which calls for further investigations. ('c-KIT', 'Gene', (59, 64)) ('AKT', 'Gene', '207', (81, 84)) ('SULF2', 'Gene', (52, 57)) ('upregulation', 'PosReg', (36, 48)) ('hTERT', 'Gene', '7015', (132, 137)) ('c-KIT', 'Gene', '3815', (59, 64)) ('PKC412', 'Var', (113, 119)) ('expression', 'MPA', (65, 75)) ('hTERT', 'Gene', (132, 137)) ('AKT', 'Gene', (81, 84)) ('SULF2', 'Gene', '55959', (52, 57)) 6006 29080039 In summary, we identified that PKC412, a TKI specifically targeting FLT3ITD mutation, repressed hTERT transcription and telomerase activity in FLT3ITD-carrying AML cells in a MYC-dependent manner. ('MYC', 'Gene', (175, 178)) ('hTERT', 'Gene', '7015', (96, 101)) ('AML', 'Disease', 'MESH:D015470', (160, 163)) ('activity', 'MPA', (131, 139)) ('AML', 'Disease', (160, 163)) ('hTERT', 'Gene', (96, 101)) ('MYC', 'Gene', '4609', (175, 178)) ('AML', 'Phenotype', 'HP:0004808', (160, 163)) ('telomerase', 'Enzyme', (120, 130)) ('FLT3ITD', 'Gene', (68, 75)) ('mutation', 'Var', (76, 84)) 6007 29080039 This effect of PKC412 is likely associated with its therapeutic efficacy on AML. ('AML', 'Phenotype', 'HP:0004808', (76, 79)) ('AML', 'Disease', 'MESH:D015470', (76, 79)) ('AML', 'Disease', (76, 79)) ('PKC412', 'Var', (15, 21)) 6008 29080039 Importantly, we demonstrate that hTERT significantly attenuates the apoptotic cell death mediated by PKC412, which strongly indicates that hTERT is capable of mediating resistance to cancer-targeted therapy. ('cancer', 'Disease', (183, 189)) ('PKC412', 'Var', (101, 107)) ('hTERT', 'Gene', '7015', (33, 38)) ('hTERT', 'Gene', '7015', (139, 144)) ('apoptotic cell death', 'CPA', (68, 88)) ('cancer', 'Phenotype', 'HP:0002664', (183, 189)) ('attenuates', 'NegReg', (53, 63)) ('hTERT', 'Gene', (33, 38)) ('hTERT', 'Gene', (139, 144)) ('cancer', 'Disease', 'MESH:D009369', (183, 189)) 6015 26205403 Distinct mechanisms such as transcriptional, epigenetic or interference with miRNA processing machinery have been involved. ('interference', 'NegReg', (59, 71)) ('miR', 'Gene', (77, 80)) ('miR', 'Gene', '220972', (77, 80)) ('epigenetic', 'Var', (45, 55)) 6022 26205403 The genome of HTLV-1 encodes common retrovirus structural and enzymatic proteins, Gag, Pro, Pol, and Env, and additional accessory and regulatory proteins such as Tax, Rex, P30, p12, p13, and HTLV-1 basic leucine zipper factor protein (HBZ). ('Env', 'Gene', '1491939', (101, 104)) ('p12', 'Var', (178, 181)) ('Gag', 'Gene', '1491934', (82, 85)) ('Gag', 'Gene', (82, 85)) ('p13', 'Var', (183, 186)) ('P30', 'Var', (173, 176)) ('HTLV-1', 'Gene', (192, 198)) ('HBZ', 'Gene', '3050', (236, 239)) ('Rex', 'Gene', '1491937', (168, 171)) ('HTLV-1', 'Species', '11908', (192, 198)) ('Rex', 'Gene', (168, 171)) ('HBZ', 'Gene', (236, 239)) ('HTLV-1', 'Species', '11908', (14, 20)) ('HTLV-1', 'Gene', (14, 20)) ('Env', 'Gene', (101, 104)) 6099 26205403 Bai and colleagues proposed a model where the modulation of miR-28-3p expression affected HTLV-1 virus spreading. ('miR-28', 'Gene', '407020', (60, 66)) ('miR-28', 'Gene', (60, 66)) ('affected', 'Reg', (81, 89)) ('HTLV-1', 'Gene', (90, 96)) ('modulation', 'Var', (46, 56)) ('spreading', 'CPA', (103, 112)) ('HTLV-1', 'Species', '11908', (90, 96)) 6195 26205403 It has been reported that loss of TP53INP1 correlates with the development of cancers and its induction promotes G1 cell cycle arrest and apoptosis. ('TP53INP1', 'Gene', (34, 42)) ('cancer', 'Phenotype', 'HP:0002664', (78, 84)) ('arrest', 'Disease', (127, 133)) ('cancers', 'Disease', 'MESH:D009369', (78, 85)) ('TP53INP1', 'Gene', '94241', (34, 42)) ('cancers', 'Phenotype', 'HP:0002664', (78, 85)) ('promotes', 'PosReg', (104, 112)) ('cancers', 'Disease', (78, 85)) ('apoptosis', 'CPA', (138, 147)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (116, 133)) ('loss', 'Var', (26, 30)) ('arrest', 'Disease', 'MESH:D006323', (127, 133)) 6217 26205403 The authors have shown that ectopic expression of HBZ does not decrease cellular growth in DNA-damaged cells. ('HBZ', 'Gene', (50, 53)) ('cellular growth', 'CPA', (72, 87)) ('ectopic expression', 'Var', (28, 46)) ('HBZ', 'Gene', '3050', (50, 53)) 6219 26205403 This phenotype can be reversed by ectopic expression of OBFC2A, which leads to a decrease of proliferation rates and restores the DNA damage response. ('decrease', 'NegReg', (81, 89)) ('DNA damage response', 'MPA', (130, 149)) ('proliferation rates', 'CPA', (93, 112)) ('OBFC2A', 'Gene', (56, 62)) ('ectopic expression', 'Var', (34, 52)) ('restores', 'PosReg', (117, 125)) ('OBFC2A', 'Gene', '64859', (56, 62)) 6238 25973391 Nevertheless, no clear association between constitutional 47,XXY and acute leukemias has yet been established. ('leukemias', 'Disease', 'MESH:D007938', (75, 84)) ('acute leukemias', 'Phenotype', 'HP:0002488', (69, 84)) ('acute leukemia', 'Phenotype', 'HP:0002488', (69, 83)) ('leukemias', 'Phenotype', 'HP:0001909', (75, 84)) ('XXY', 'Disease', (61, 64)) ('constitutional 47', 'Var', (43, 60)) ('leukemias', 'Disease', (75, 84)) ('leukemia', 'Phenotype', 'HP:0001909', (75, 83)) 6249 25973391 Subsequent molecular genetic analysis revealed an NPM1 mutation at exon 12, but no FLT3 or CEBPA mutations. ('NPM1', 'Gene', (50, 54)) ('FLT3', 'Gene', (83, 87)) ('NPM1', 'Gene', '4869', (50, 54)) ('CEBPA', 'Gene', (91, 96)) ('CEBPA', 'Gene', '1050', (91, 96)) ('revealed', 'Reg', (38, 46)) ('FLT3', 'Gene', '2322', (83, 87)) ('mutation', 'Var', (55, 63)) 6250 25973391 The final diagnosis was acute myeloid leukemia with mutated NPM1. ('NPM1', 'Gene', (60, 64)) ('mutated', 'Var', (52, 59)) ('leukemia', 'Phenotype', 'HP:0001909', (38, 46)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (24, 46)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (30, 46)) ('NPM1', 'Gene', '4869', (60, 64)) ('acute myeloid leukemia', 'Disease', (24, 46)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (24, 46)) 6255 25973391 Given the relatively favorable prognostic risk associated with an isolated NPM1 mutation, the patient has gone on to receive cycles of consolidation chemotherapy with high dose cytarabine. ('cytarabine', 'Chemical', 'MESH:D003561', (177, 187)) ('NPM1', 'Gene', (75, 79)) ('isolated', 'Var', (66, 74)) ('NPM1', 'Gene', '4869', (75, 79)) ('patient', 'Species', '9606', (94, 101)) ('mutation', 'Var', (80, 88)) 6258 25973391 For example, children with trisomy 21 (Down syndrome) have a 10-20 fold higher risk of developing acute leukemia than other children. ('children', 'Species', '9606', (124, 132)) ('leukemia', 'Disease', (104, 112)) ('leukemia', 'Phenotype', 'HP:0001909', (104, 112)) ('leukemia', 'Disease', 'MESH:D007938', (104, 112)) ('acute leukemia', 'Phenotype', 'HP:0002488', (98, 112)) ('children', 'Species', '9606', (13, 21)) ('trisomy 21', 'Var', (27, 37)) 6306 33841567 F-PDOs were established in our previous study, and three lung F-PDOs (RLUN001, RLUN021, and RLUN023), each established from a different patient, were used in this study. ('patient', 'Species', '9606', (136, 143)) ('RLUN023', 'Var', (92, 99)) ('RLUN021', 'Var', (79, 86)) ('lung F-PDOs', 'Disease', 'OMIM:102510', (57, 68)) ('lung F-PDOs', 'Disease', (57, 68)) 6352 33841567 We examined the use of F-PDO (RLUN001 and RLUN023) established from lung tumors, which is difficult to assay using reported methods. ('lung tumor', 'Phenotype', 'HP:0100526', (68, 78)) ('lung tumors', 'Phenotype', 'HP:0100526', (68, 79)) ('lung tumors', 'Disease', (68, 79)) ('tumor', 'Phenotype', 'HP:0002664', (73, 78)) ('tumors', 'Phenotype', 'HP:0002664', (73, 79)) ('RLUN023', 'Var', (42, 49)) ('lung tumors', 'Disease', 'MESH:D008175', (68, 79)) 6354 33841567 RLUN001 and RLUN023, which possess a sensitivity mutation (p.E746-A750del) to EGFR inhibitors, may indicate a high sensitivity. ('EGFR', 'Gene', (78, 82)) ('p.E746-A750del', 'Var', (59, 73)) ('EGFR', 'Gene', '1956', (78, 82)) ('p.E746-A750del', 'Mutation', 'p.746,750delA', (59, 73)) 6357 33841567 We have previously reported that the IC50 values of erlotinib and gefitinib for RLUN021 without EGFR mutations were very high at >20 and 6 microM, respectively. ('mutations', 'Var', (101, 110)) ('gefitinib', 'Chemical', 'MESH:D000077156', (66, 75)) ('EGFR', 'Gene', (96, 100)) ('IC50', 'MPA', (37, 41)) ('erlotinib', 'Chemical', 'MESH:D000069347', (52, 61)) ('EGFR', 'Gene', '1956', (96, 100)) 6358 33841567 These results indicate that an F-PDO with susceptibility mutations is highly sensitive to EGFR inhibitors, consistent with general clinical results. ('mutations', 'Var', (57, 66)) ('sensitive', 'MPA', (77, 86)) ('EGFR', 'Gene', (90, 94)) ('EGFR', 'Gene', '1956', (90, 94)) 6403 33841567 In the EGFR inhibitor sensitivity test of lung tumor-derived F-PDOs, those with mutant EGFRs that are clinically sensitive to EGFR inhibitors were more sensitive than the wild type. ('lung tumor', 'Phenotype', 'HP:0100526', (42, 52)) ('EGFR', 'Gene', '1956', (87, 91)) ('EGFR', 'Gene', (126, 130)) ('EGFR', 'Gene', (7, 11)) ('mutant', 'Var', (80, 86)) ('EGFR', 'Gene', (87, 91)) ('lung tumor', 'Disease', 'MESH:D008175', (42, 52)) ('tumor', 'Phenotype', 'HP:0002664', (47, 52)) ('lung tumor', 'Disease', (42, 52)) ('EGFR', 'Gene', '1956', (7, 11)) ('sensitive', 'MPA', (152, 161)) ('EGFR', 'Gene', '1956', (126, 130)) 6448 32269970 In addition, these cells express a set of cell surface markers such as CD133, CD44, CD33, CD29, CD24, aldehyde dehydrogenase1 (ALDH1), and others, which can be enriched by other cells within the tumor. ('CD29', 'Gene', (90, 94)) ('tumor', 'Disease', (195, 200)) ('ALDH1', 'Gene', '216', (127, 132)) ('aldehyde dehydrogenase1', 'Gene', '216', (102, 125)) ('CD44', 'Gene', (78, 82)) ('CD133', 'Gene', '8842', (71, 76)) ('CD29', 'Gene', '3688', (90, 94)) ('CD133', 'Gene', (71, 76)) ('CD24', 'Gene', (96, 100)) ('CD33', 'Var', (84, 88)) ('aldehyde dehydrogenase1', 'Gene', (102, 125)) ('tumor', 'Disease', 'MESH:D009369', (195, 200)) ('ALDH1', 'Gene', (127, 132)) ('CD24', 'Gene', '100133941', (96, 100)) ('tumor', 'Phenotype', 'HP:0002664', (195, 200)) ('CD44', 'Gene', '960', (78, 82)) 6467 32269970 Also, a new finding has shown the feasibility of engrafting CD33+-derived cord blood cells with multi-lineage hematopoiesis. ('CD33+-derived', 'Var', (60, 73)) ('hematopoiesis', 'Disease', (110, 123)) ('hematopoiesis', 'Disease', 'MESH:C536227', (110, 123)) 6469 32269970 In scenario 1, both the preliminary and subsequent mutations occur at the level of pluripotent CD33- progenitors; these are referred to as "immature" leukemias. ('leukemias', 'Disease', 'MESH:D007938', (150, 159)) ('mutations', 'Var', (51, 60)) ('leukemia', 'Phenotype', 'HP:0001909', (150, 158)) ('leukemias', 'Phenotype', 'HP:0001909', (150, 159)) ('leukemias', 'Disease', (150, 159)) 6472 32269970 The preliminary mutation specifically occurs in pluripotent HSCs in the second scenario; however, the cooperating mutations result in the full transformation to AML, and then clonal expansion only happens at a later stage, perhaps at the committed CD33-myeloid progenitor level. ('mutations', 'Var', (114, 123)) ('AML', 'Phenotype', 'HP:0004808', (161, 164)) ('AML', 'Disease', 'MESH:D015470', (161, 164)) ('pluripotent HSCs', 'Disease', 'None', (48, 64)) ('transformation', 'MPA', (143, 157)) ('pluripotent HSCs', 'Disease', (48, 64)) ('AML', 'Disease', (161, 164)) ('result in', 'Reg', (124, 133)) 6482 32269970 The practicality of CD33-targeted therapeutics is supported by the observation that, in some leukemias with clonal influence limited to monocytes/granulocytes, ablation of CD33+ cells could result in the resumption of normal hematopoiesis. ('hematopoiesis', 'Disease', 'MESH:C536227', (225, 238)) ('leukemias', 'Phenotype', 'HP:0001909', (93, 102)) ('result in', 'Reg', (190, 199)) ('leukemias', 'Disease', (93, 102)) ('ablation', 'Var', (160, 168)) ('hematopoiesis', 'Disease', (225, 238)) ('leukemia', 'Phenotype', 'HP:0001909', (93, 101)) ('leukemias', 'Disease', 'MESH:D007938', (93, 102)) ('CD33+', 'Gene', (172, 177)) 6483 32269970 In other words, unconjugated anti-CD33 antibodies were greatly useless in patients with distinct CD33+ non-APL AML. ('APL', 'Phenotype', 'HP:0004836', (107, 110)) ('patients', 'Species', '9606', (74, 82)) ('APL AML', 'Disease', (107, 114)) ('AML', 'Phenotype', 'HP:0004808', (111, 114)) ('APL AML', 'Disease', 'MESH:D015473', (107, 114)) ('CD33+', 'Var', (97, 102)) 6504 32269970 Up to now, one report of a patient with r/r AML, who was treated with anti-CD33 CAR-T (CART-33) cell, has been published. ('AML', 'Disease', 'MESH:D015470', (44, 47)) ('CART-33', 'Gene', (87, 94)) ('CAR-T', 'Gene', '9607', (80, 85)) ('CART-33', 'Gene', '9607', (87, 94)) ('patient', 'Species', '9606', (27, 34)) ('AML', 'Disease', (44, 47)) ('AML', 'Phenotype', 'HP:0004808', (44, 47)) ('CAR-T', 'Gene', (80, 85)) ('anti-CD33', 'Var', (70, 79)) 6510 32269970 This is probably because of the heterogeneous and extremely polyclonal character of AML, and approaches that target peculiar mutations in AML cells may result in the eradication just single subclasses, and thus they are insufficient to rid a majority of patients of the disease. ('mutations', 'Var', (125, 134)) ('AML', 'Phenotype', 'HP:0004808', (138, 141)) ('AML', 'Disease', (138, 141)) ('AML', 'Disease', 'MESH:D015470', (84, 87)) ('AML', 'Phenotype', 'HP:0004808', (84, 87)) ('AML', 'Disease', (84, 87)) ('AML', 'Disease', 'MESH:D015470', (138, 141)) ('patients', 'Species', '9606', (254, 262)) ('eradication', 'MPA', (166, 177)) 6520 30625392 The presence of U2AF1 (p=0.007) or DNMT3A (p=0.034) mutations was associated with worse OS. ('mutations', 'Var', (52, 61)) ('associated', 'Reg', (66, 76)) ('U2AF1', 'Gene', (16, 21)) ('U2AF1', 'Gene', '7307', (16, 21)) ('DNMT3A', 'Gene', (35, 41)) ('DNMT3A', 'Gene', '1788', (35, 41)) ('worse OS', 'Disease', (82, 90)) 6529 30625392 Although activation of the JAK-STAT signaling pathway remains the hallmark of MPN pathogenesis, it has become clear that the presence of additional genomic events, such as mutations in TET2, EZH2 and TP53, alters the biology of disease in preclinical models. ('mutations', 'Var', (172, 181)) ('MPN', 'Gene', '83886', (78, 81)) ('TET2', 'Gene', (185, 189)) ('biology of', 'CPA', (217, 227)) ('MPN', 'Gene', (78, 81)) ('JAK-STAT signaling pathway', 'Pathway', (27, 53)) ('TP53', 'Gene', '7157', (200, 204)) ('TP53', 'Gene', (200, 204)) ('EZH2', 'Gene', (191, 195)) ('alters', 'Reg', (206, 212)) ('EZH2', 'Gene', '2146', (191, 195)) ('TET2', 'Gene', '54790', (185, 189)) 6531 30625392 For example, the presence of ASXL1, EZH2, SRSF2, and IDH1/2 mutations, as well as a lack of canonical JAK2, MPL or CALR mutations (so-called triple negative status) are associated with increased risk of leukemic transformation and poor survival and the presence of any of these mutations is considered high molecular risk disease (HMR). ('ASXL1', 'Gene', '171023', (29, 34)) ('mutations', 'Var', (120, 129)) ('IDH1/2', 'Gene', '3417;3418', (53, 59)) ('CALR', 'Gene', (115, 119)) ('JAK2', 'Gene', '3717', (102, 106)) ('IDH1/2', 'Gene', (53, 59)) ('CALR', 'Gene', '811', (115, 119)) ('ASXL1', 'Gene', (29, 34)) ('leukemic transformation', 'Disease', 'MESH:D002472', (203, 226)) ('SRSF2', 'Gene', '6427', (42, 47)) ('leukemic transformation', 'Disease', (203, 226)) ('MPL', 'Gene', '4352', (108, 111)) ('SRSF2', 'Gene', (42, 47)) ('JAK2', 'Gene', (102, 106)) ('EZH2', 'Gene', '2146', (36, 40)) ('EZH2', 'Gene', (36, 40)) ('mutations', 'Var', (60, 69)) ('MPL', 'Gene', (108, 111)) ('poor survival', 'CPA', (231, 244)) 6532 30625392 Moreover, the presence of certain genotypes appears to predict less-durable response to JAK inhibitor therapy in MF patients . ('patients', 'Species', '9606', (116, 124)) ('presence', 'Var', (14, 22)) ('less-durable', 'NegReg', (63, 75)) ('MF', 'Phenotype', 'HP:0011974', (113, 115)) 6548 30625392 The majority of patients had activating JAK2 mutations (56.4%). ('mutations', 'Var', (45, 54)) ('JAK2', 'Gene', (40, 44)) ('patients', 'Species', '9606', (16, 24)) ('JAK2', 'Gene', '3717', (40, 44)) ('activating', 'PosReg', (29, 39)) 6549 30625392 Mutations in chromatin modifiers (ASXL1 18% and EZH2 4%) as well splicing factors (SRSF2 12%, U2AF1 10% and SF3B1 4%) were the most frequently observed class of non-JAK-STAT mutations in this cohort. ('SF3B1', 'Gene', '23451', (108, 113)) ('SRSF2', 'Gene', '6427', (83, 88)) ('U2AF1', 'Gene', (94, 99)) ('U2AF1', 'Gene', '7307', (94, 99)) ('EZH2', 'Gene', '2146', (48, 52)) ('Mutations', 'Var', (0, 9)) ('EZH2', 'Gene', (48, 52)) ('SRSF2', 'Gene', (83, 88)) ('observed', 'Reg', (143, 151)) ('SF3B1', 'Gene', (108, 113)) ('ASXL1', 'Gene', '171023', (34, 39)) ('ASXL1', 'Gene', (34, 39)) 6551 30625392 Notably, we identified recurrent mutations in KMT2C in 11% of patients. ('KMT2C', 'Gene', (46, 51)) ('patients', 'Species', '9606', (62, 70)) ('mutations', 'Var', (33, 42)) ('KMT2C', 'Gene', '58508', (46, 51)) 6552 30625392 As mentioned above, the presence of mutations in ASXL1, SRSF2, IDH1/2, and EZH2 have been previously associated with increased risk of leukemic transformation, and TP53 mutations are enriched in post-MPN AML. ('EZH2', 'Gene', (75, 79)) ('ASXL1', 'Gene', (49, 54)) ('IDH1/2', 'Gene', '3417;3418', (63, 69)) ('leukemic transformation', 'Disease', (135, 158)) ('IDH1/2', 'Gene', (63, 69)) ('presence', 'Var', (24, 32)) ('TP53', 'Gene', (164, 168)) ('MPN', 'Gene', (200, 203)) ('associated', 'Reg', (101, 111)) ('ASXL1', 'Gene', '171023', (49, 54)) ('TP53', 'Gene', '7157', (164, 168)) ('AML', 'Disease', 'MESH:D015470', (204, 207)) ('SRSF2', 'Gene', '6427', (56, 61)) ('AML', 'Disease', (204, 207)) ('mutations', 'Var', (36, 45)) ('SRSF2', 'Gene', (56, 61)) ('MPN', 'Gene', '83886', (200, 203)) ('leukemic transformation', 'Disease', 'MESH:D002472', (135, 158)) ('EZH2', 'Gene', '2146', (75, 79)) 6557 30625392 Further, prior data has indicated that increasing numbers of mutations per patient are associated with increased risk of leukemic transformation and impaired survival . ('leukemic transformation', 'Disease', (121, 144)) ('impaired', 'NegReg', (149, 157)) ('survival', 'CPA', (158, 166)) ('mutations', 'Var', (61, 70)) ('leukemic transformation', 'Disease', 'MESH:D002472', (121, 144)) ('patient', 'Species', '9606', (75, 82)) 6576 30625392 Furthermore, the presence of HMR mutations did not impact survival of patients in this cohort (HR for mortality with HMR mutation compared to none was 1.42, 95% CI 0.77-2.61, p=0.2603, Figure 3B). ('mortality', 'Disease', (102, 111)) ('patients', 'Species', '9606', (70, 78)) ('mortality', 'Disease', 'MESH:D003643', (102, 111)) ('mutation', 'Var', (121, 129)) 6577 30625392 Analysis of the impact of individual mutations revealed that the presence of U2AF1 or DNMT3A was associated with worse OS (U2AF1: hazard ratio for death 2.76; 95% confidence interval, 1.28 to 5.99, p=0.007, DNM3TA: HR 2.91; 95% CI, 1.03 - 8.24, p=0.034, Figure 3C and 3D respectively). ('U2AF1', 'Gene', '7307', (123, 128)) ('death', 'Disease', 'MESH:D003643', (147, 152)) ('death', 'Disease', (147, 152)) ('U2AF1', 'Gene', (77, 82)) ('worse OS', 'Disease', (113, 121)) ('DNMT3A', 'Gene', '1788', (86, 92)) ('U2AF1', 'Gene', '7307', (77, 82)) ('DNMT3A', 'Gene', (86, 92)) ('U2AF1', 'Gene', (123, 128)) ('presence', 'Var', (65, 73)) 6578 30625392 Notably, three out of four cases of mortality due to graft failure occurred in patients with U2AF1 mutations. ('U2AF1', 'Gene', '7307', (93, 98)) ('mortality', 'Disease', 'MESH:D003643', (36, 45)) ('U2AF1', 'Gene', (93, 98)) ('occurred', 'Reg', (67, 75)) ('graft failure', 'Disease', (53, 66)) ('graft failure', 'Disease', 'MESH:D006333', (53, 66)) ('mortality', 'Disease', (36, 45)) ('mutations', 'Var', (99, 108)) ('patients', 'Species', '9606', (79, 87)) 6579 30625392 As well, the presences of U2AF1, DNMT3A, or IDH2 mutations were associated with increased risk of NRM (Supplemental Table 3). ('DNMT3A', 'Gene', (33, 39)) ('DNMT3A', 'Gene', '1788', (33, 39)) ('IDH2', 'Gene', (44, 48)) ('mutations', 'Var', (49, 58)) ('IDH2', 'Gene', '3418', (44, 48)) ('presences', 'Var', (13, 22)) ('U2AF1', 'Gene', '7307', (26, 31)) ('NRM', 'Disease', (98, 101)) ('U2AF1', 'Gene', (26, 31)) 6580 30625392 Recent data has indicated that the variant allele fraction (VAF) of mutant genes such as TP53 in myelodysplastic syndrome may impact the clinical outcomes of patients. ('TP53', 'Gene', '7157', (89, 93)) ('TP53', 'Gene', (89, 93)) ('myelodysplastic syndrome', 'Disease', 'MESH:D009190', (98, 122)) ('myelodysplastic syndrome', 'Phenotype', 'HP:0002863', (98, 122)) ('impact', 'Reg', (127, 133)) ('clinical outcomes', 'CPA', (138, 155)) ('variant', 'Var', (35, 42)) ('patients', 'Species', '9606', (159, 167)) ('myelodysplastic syndrome', 'Disease', (98, 122)) ('mutant', 'Var', (68, 74)) 6584 30625392 In multivariate analysis, both RIC (HR 5.38, 95% CI 1.29-22.39, p=0.02) and the presence of U2AF1 mutations (HR 2.83, 95% CI 1.29-6.19, p=0.009) remained negatively associated with OS. ('mutations', 'Var', (98, 107)) ('U2AF1', 'Gene', (92, 97)) ('U2AF1', 'Gene', '7307', (92, 97)) ('RIC', 'Disease', (31, 34)) ('negatively', 'NegReg', (154, 164)) 6586 30625392 Univariate analysis demonstrated that RIC was associated with worse RFS compared to MAC (HR 2.96, 95% CI 1.06-8.26, p=0.03) and presence of U2AF1 mutations (HR 2.37, 95% CI 1.10-5.08, p=0.026) and DNMT3A mutations (HR 4.02 95% CI 1.56-10.35, p=0.0018) were associated with worse RFS (Supplemental Table 4, Supplemental Figure 1). ('U2AF1', 'Gene', '7307', (140, 145)) ('mutations', 'Var', (146, 155)) ('DNMT3A', 'Gene', (197, 203)) ('DNMT3A', 'Gene', '1788', (197, 203)) ('mutations', 'Var', (204, 213)) ('RIC', 'Disease', (38, 41)) ('RFS', 'MPA', (68, 71)) ('U2AF1', 'Gene', (140, 145)) 6593 30625392 In multivariate analysis, mutations previously associated with worse outcome in MF patients, such as ASXL1, EZH2, SRSF2, IDH1/2, and TP53 mutations were not found to affect OS or RFS in MF patients undergoing transplant. ('MF', 'Phenotype', 'HP:0011974', (186, 188)) ('ASXL1', 'Gene', (101, 106)) ('SRSF2', 'Gene', '6427', (114, 119)) ('IDH1/2', 'Gene', '3417;3418', (121, 127)) ('RFS', 'Disease', (179, 182)) ('ASXL1', 'Gene', '171023', (101, 106)) ('EZH2', 'Gene', (108, 112)) ('affect', 'Reg', (166, 172)) ('EZH2', 'Gene', '2146', (108, 112)) ('MF', 'Phenotype', 'HP:0011974', (80, 82)) ('IDH1/2', 'Gene', (121, 127)) ('patients', 'Species', '9606', (83, 91)) ('patients', 'Species', '9606', (189, 197)) ('TP53', 'Gene', '7157', (133, 137)) ('SRSF2', 'Gene', (114, 119)) ('TP53', 'Gene', (133, 137)) ('mutations', 'Var', (138, 147)) 6596 30625392 We identified U2AF1 mutations as a risk factor for decreased OS, and U2AF1 and DNMT3A mutations were both associated with impaired RFS. ('mutations', 'Var', (86, 95)) ('U2AF1', 'Gene', (69, 74)) ('U2AF1', 'Gene', '7307', (69, 74)) ('U2AF1', 'Gene', '7307', (14, 19)) ('associated', 'Reg', (106, 116)) ('decreased OS', 'Disease', (51, 63)) ('impaired RFS', 'Disease', (122, 134)) ('DNMT3A', 'Gene', (79, 85)) ('DNMT3A', 'Gene', '1788', (79, 85)) ('decreased OS', 'Disease', 'MESH:C567932', (51, 63)) ('impaired RFS', 'Disease', 'MESH:D005198', (122, 134)) ('mutations', 'Var', (20, 29)) ('U2AF1', 'Gene', (14, 19)) 6597 30625392 Mutations in U2AF1 have been reported in about 10-15% of patients with MF and have been shown to strongly correlate with the degree of anemia and also with worse OS compared to patients with unmutated U2AF1. ('correlate with', 'Reg', (106, 120)) ('U2AF1', 'Gene', (202, 207)) ('U2AF1', 'Gene', '7307', (202, 207)) ('anemia', 'Disease', 'MESH:D000740', (135, 141)) ('anemia', 'Disease', (135, 141)) ('patients', 'Species', '9606', (57, 65)) ('patients', 'Species', '9606', (178, 186)) ('anemia', 'Phenotype', 'HP:0001903', (135, 141)) ('Mutations', 'Var', (0, 9)) ('reported', 'Reg', (29, 37)) ('U2AF1', 'Gene', '7307', (13, 18)) ('U2AF1', 'Gene', (13, 18)) ('MF', 'Phenotype', 'HP:0011974', (71, 73)) 6598 30625392 Interestingly, in our cohort, 4 cases of mortality were secondary to graft failure, 3 of which had mutated U2AF1. ('U2AF1', 'Gene', '7307', (107, 112)) ('U2AF1', 'Gene', (107, 112)) ('mortality', 'Disease', 'MESH:D003643', (41, 50)) ('mutated', 'Var', (99, 106)) ('graft failure', 'Disease', (69, 82)) ('graft failure', 'Disease', 'MESH:D006333', (69, 82)) ('mortality', 'Disease', (41, 50)) 6602 30625392 DNMT3A mutations appear to mediate anthracycline-based chemotherapy resistance in AML and DNMT3A R882 in particular predicts for minimal residual disease in AML. ('anthracycline', 'Chemical', 'MESH:D018943', (35, 48)) ('AML', 'Disease', 'MESH:D015470', (82, 85)) ('anthracycline-based chemotherapy resistance', 'MPA', (35, 78)) ('minimal residual disease', 'Disease', (129, 153)) ('AML', 'Disease', 'MESH:D015470', (157, 160)) ('mediate', 'Reg', (27, 34)) ('DNMT3A', 'Gene', (0, 6)) ('AML', 'Disease', (82, 85)) ('DNMT3A', 'Gene', (90, 96)) ('DNMT3A', 'Gene', '1788', (0, 6)) ('DNMT3A', 'Gene', '1788', (90, 96)) ('AML', 'Disease', (157, 160)) ('mutations', 'Var', (7, 16)) 6603 30625392 Thus, it is possible that the presence of DNMT3A mutations renders MPN hematopoietic stem cells relatively resistant to effects of conditioning. ('mutations', 'Var', (49, 58)) ('DNMT3A', 'Gene', (42, 48)) ('DNMT3A', 'Gene', '1788', (42, 48)) ('MPN', 'Gene', '83886', (67, 70)) ('MPN', 'Gene', (67, 70)) 6604 30625392 The biological impact of U2AF1 and DNMT3a mutations may thus alter the likelihood of transplant success. ('U2AF1', 'Gene', (25, 30)) ('U2AF1', 'Gene', '7307', (25, 30)) ('alter', 'Reg', (61, 66)) ('DNMT3a', 'Gene', (35, 41)) ('DNMT3a', 'Gene', '1788', (35, 41)) ('mutations', 'Var', (42, 51)) 6606 30625392 However, we did identify mutations in KMT2C. ('mutations', 'Var', (25, 34)) ('KMT2C', 'Gene', '58508', (38, 43)) ('KMT2C', 'Gene', (38, 43)) 6607 30625392 KMT2C mutations have been described in a variety of solid tumors and were recently described by Durham et al. ('solid tumors', 'Disease', 'MESH:D009369', (52, 64)) ('tumor', 'Phenotype', 'HP:0002664', (58, 63)) ('tumors', 'Phenotype', 'HP:0002664', (58, 64)) ('KMT2C', 'Gene', '58508', (0, 5)) ('KMT2C', 'Gene', (0, 5)) ('solid tumors', 'Disease', (52, 64)) ('described', 'Reg', (26, 35)) ('mutations', 'Var', (6, 15)) 6609 30625392 Chang et al also recently reported KMT2C mutations in a group of patients with TN MPN. ('reported', 'Reg', (26, 34)) ('KMT2C', 'Gene', (35, 40)) ('KMT2C', 'Gene', '58508', (35, 40)) ('mutations', 'Var', (41, 50)) ('patients', 'Species', '9606', (65, 73)) ('TN MPN', 'Disease', (79, 85)) ('TN MPN', 'Disease', 'MESH:C562719', (79, 85)) 6610 30625392 The biological contribution of KMT2C mutations to MPN pathogenesis remains to be determined. ('MPN', 'Gene', (50, 53)) ('mutations', 'Var', (37, 46)) ('MPN', 'Gene', '83886', (50, 53)) ('KMT2C', 'Gene', '58508', (31, 36)) ('KMT2C', 'Gene', (31, 36)) 6611 30625392 Most cases of mortality in this cohort were not related to relapse, and indeed the incidence of relapse was surprisingly low despite the fact that 55% of the patients in this cohort had advanced disease (Intermediate-II and high-risk disease), and many patients had HMR mutations. ('mortality', 'Disease', 'MESH:D003643', (14, 23)) ('patients', 'Species', '9606', (253, 261)) ('mutations', 'Var', (270, 279)) ('HMR', 'Gene', (266, 269)) ('patients', 'Species', '9606', (158, 166)) ('mortality', 'Disease', (14, 23)) 6612 30625392 By contrast, data from MDS and AML literature indicates that certain mutations predict for very poor prognosis post allo-HCT, mostly due to disease relapse . ('mutations', 'Var', (69, 78)) ('MDS', 'Disease', (23, 26)) ('MDS', 'Disease', 'MESH:D009190', (23, 26)) ('AML', 'Disease', 'MESH:D015470', (31, 34)) ('disease', 'Disease', (140, 147)) ('AML', 'Disease', (31, 34)) 6613 30625392 As well, our findings are in contrast to data recently published by Kroger et al that demonstrated ASXL1 mutations are associated with higher relapse risk. ('ASXL1', 'Gene', (99, 104)) ('mutations', 'Var', (105, 114)) ('ASXL1', 'Gene', '171023', (99, 104)) ('relapse', 'Disease', (142, 149)) 6614 30625392 In our cohort, among 19 patients who had mutated ASXL1, nine patients died without relapse at a median of 4 months post-transplant. ('mutated', 'Var', (41, 48)) ('ASXL1', 'Gene', (49, 54)) ('died', 'Disease', (70, 74)) ('died', 'Disease', 'MESH:D003643', (70, 74)) ('patients', 'Species', '9606', (61, 69)) ('patients', 'Species', '9606', (24, 32)) ('ASXL1', 'Gene', '171023', (49, 54)) 6619 30625392 In two patients, loss of clones containing ASXL1 and KMT2C mutations were noted, suggesting some degree of selective pressure by allo-HCT on different subclones. ('ASXL1', 'Gene', '171023', (43, 48)) ('KMT2C', 'Gene', '58508', (53, 58)) ('KMT2C', 'Gene', (53, 58)) ('ASXL1', 'Gene', (43, 48)) ('mutations', 'Var', (59, 68)) ('patients', 'Species', '9606', (7, 15)) 6627 30625392 Our data establishes that genomic alterations have predictive value with regard to allo-HCT, and are likely useful in guiding transplant treatment decision-making in MF patients. ('allo-HCT', 'Disease', (83, 91)) ('genomic alterations', 'Var', (26, 45)) ('patients', 'Species', '9606', (169, 177)) ('MF', 'Phenotype', 'HP:0011974', (166, 168)) 6628 30625392 It also suggests that mutations that are associated with poor prognosis and progression to AML do not predict for post-transplant outcomes. ('AML', 'Disease', 'MESH:D015470', (91, 94)) ('mutations', 'Var', (22, 31)) ('AML', 'Disease', (91, 94)) 6629 30625392 Moreover, these observations raise new questions about how genomic alterations may impact transplant outcomes in MF and whether interventions to eliminate the mutated clone, particularly in patient with mutated U2AF1, will impact transplant outcomes (notably, clinical trials of inhibitors targeting splicing factors are currently underway; NCT02841540). ('transplant outcomes', 'CPA', (230, 249)) ('mutated', 'Var', (203, 210)) ('mutated', 'Var', (159, 166)) ('MF', 'Phenotype', 'HP:0011974', (113, 115)) ('transplant outcomes', 'CPA', (90, 109)) ('U2AF1', 'Gene', '7307', (211, 216)) ('impact', 'Reg', (223, 229)) ('U2AF1', 'Gene', (211, 216)) ('impact', 'Reg', (83, 89)) ('patient', 'Species', '9606', (190, 197)) 6632 30625392 The presence of U2AF1 mutations was associated with worsened overall survival and relapse-free survival in patients undergoing allo-HCT for MF. ('MF', 'Phenotype', 'HP:0011974', (140, 142)) ('relapse-free survival', 'CPA', (82, 103)) ('U2AF1', 'Gene', (16, 21)) ('U2AF1', 'Gene', '7307', (16, 21)) ('mutations', 'Var', (22, 31)) ('overall survival', 'CPA', (61, 77)) ('patients', 'Species', '9606', (107, 115)) ('worsened', 'NegReg', (52, 60)) 6633 31801627 Functional characterization of the selective pan-allele anti-SIRPalpha antibody ADU-1805 that blocks the SIRPalpha-CD47 innate immune checkpoint Accumulating preclinical data indicate that targeting the SIRPalpha/CD47 axis alone or in combination with existing targeted therapies or immune checkpoint inhibitors enhances tumor rejection. ('CD47', 'Gene', '961', (213, 217)) ('SIRPalpha', 'Gene', '140885', (105, 114)) ('CD47', 'Gene', (213, 217)) ('enhances', 'PosReg', (312, 320)) ('SIRPalpha', 'Gene', '140885', (203, 212)) ('tumor', 'Disease', 'MESH:D009369', (321, 326)) ('targeting', 'Var', (189, 198)) ('CD47', 'Gene', '961', (115, 119)) ('tumor', 'Phenotype', 'HP:0002664', (321, 326)) ('SIRPalpha', 'Gene', (105, 114)) ('SIRPalpha', 'Gene', (61, 70)) ('CD47', 'Gene', (115, 119)) ('blocks', 'NegReg', (94, 100)) ('tumor', 'Disease', (321, 326)) ('SIRPalpha', 'Gene', (203, 212)) ('SIRPalpha', 'Gene', '140885', (61, 70)) 6640 31801627 In vitro characterization demonstrated that ADU-1805 promotes macrophage phagocytosis, with similar potency to anti-CD47 antibodies, and enhances neutrophil trogocytosis. ('promotes', 'PosReg', (53, 61)) ('macrophage phagocytosis', 'CPA', (62, 85)) ('enhances', 'PosReg', (137, 145)) ('neutrophil trogocytosis', 'CPA', (146, 169)) ('ADU-1805', 'Var', (44, 52)) ('ADU-1805', 'Chemical', '-', (44, 52)) 6641 31801627 Unlike CD47-targeting agents, ADU-1805 does not interfere with T-cell activation and is not expected to require frequent and extensive dosing due to the restricted expression of SIRPalpha to cells of the myeloid lineage. ('ADU-1805', 'Var', (30, 38)) ('ADU-1805', 'Chemical', '-', (30, 38)) ('SIRPalpha', 'Gene', (178, 187)) ('T-cell', 'CPA', (63, 69)) 6642 31801627 ADU-1805 is cross-reactive to cynomolgus monkey SIRPalpha and upon single-dose intravenous administration in these non-human primates (NHPs) did not show any signs of anemia, thrombocytopenia or other toxicities. ('ADU-1805', 'Chemical', '-', (0, 8)) ('anemia', 'Disease', (167, 173)) ('cynomolgus monkey', 'Species', '9541', (30, 47)) ('thrombocytopenia', 'Phenotype', 'HP:0001873', (175, 191)) ('anemia', 'Disease', 'MESH:D000740', (167, 173)) ('toxicities', 'Disease', (201, 211)) ('thrombocytopenia', 'Disease', (175, 191)) ('anemia', 'Phenotype', 'HP:0001903', (167, 173)) ('thrombocytopenia', 'Disease', 'MESH:D013921', (175, 191)) ('ADU-1805', 'Var', (0, 8)) ('toxicities', 'Disease', 'MESH:D064420', (201, 211)) ('human', 'Species', '9606', (119, 124)) 6652 31801627 Of these, Hu5F9-G4, TTI-621 and ALX148 are furthest in development and have shown encouraging clinical data either alone or in combination with other agents. ('ALX148', 'Gene', (32, 38)) ('Hu5F9-G4', 'Var', (10, 18)) ('ALX148', 'Chemical', '-', (32, 38)) ('TTI-621', 'Gene', (20, 27)) 6659 31801627 In the present study, we report the development of ADU-1805, a potentially best-in-class pan-allele SIRPalpha mAb that blocks the interaction of SIRPalpha with CD47 and lacks binding to SIRPbeta1. ('ADU-1805', 'Var', (51, 59)) ('binding', 'Interaction', (175, 182)) ('ADU-1805', 'Chemical', '-', (51, 59)) ('blocks', 'NegReg', (119, 125)) ('SIRPbeta1', 'Gene', (186, 195)) ('CD47', 'Protein', (160, 164)) ('SIRPalpha', 'Protein', (145, 154)) ('interaction', 'Interaction', (130, 141)) ('SIRPbeta1', 'Gene', '10326', (186, 195)) 6717 31801627 To assess the pharmacokinetic properties of ADU-1805 in cynomolgus monkey serum, blood was drawn on 0, 1, 8, and 24 h, and 3, 8, 11, 15, 22, 29, 36, 43, 59 days post single-dose ADU-1805. ('ADU-1805', 'Gene', (44, 52)) ('ADU-1805', 'Chemical', '-', (44, 52)) ('cynomolgus monkey', 'Species', '9541', (56, 73)) ('ADU-1805', 'Var', (178, 186)) ('ADU-1805', 'Chemical', '-', (178, 186)) 6727 31801627 In this assay human peripheral blood-derived macrophages that endogenously express SIRPalpha are co-incubated with Burkitt's lymphoma Raji cells (expressing both CD20 and CD47 (Additional file 3: Figure S1A, B)). ('SIRPalpha', 'Gene', (83, 92)) ('Raji cells', 'CellLine', 'CVCL:0511', (134, 144)) ('human', 'Species', '9606', (14, 19)) ("Burkitt's lymphoma", 'Phenotype', 'HP:0030080', (115, 133)) ('CD20', 'Gene', '54474', (162, 166)) ('CD20', 'Gene', (162, 166)) ('CD47', 'Var', (171, 175)) ("Burkitt's lymphoma", 'Disease', 'MESH:D002051', (115, 133)) ('lymphoma', 'Phenotype', 'HP:0002665', (125, 133)) ("Burkitt's lymphoma", 'Disease', (115, 133)) 6729 31801627 The relevance of the unique binding profile of hSIRPalpha.40A was illustrated by the anti-hSIRPalphaV1 allele-specific mAb that only enhanced tumor cell phagocytosis by SIRPalphaV1/SIRPalphaV1 homozygous-derived macrophages while showing moderate or no phagocytosis by macrophages obtained from SIRPalphaV1/SIRPalphaV2 or SIRPalphaV2/SIRPalphaV2 individuals, respectively. ('tumor', 'Phenotype', 'HP:0002664', (142, 147)) ('hSIRPalpha', 'Gene', (90, 100)) ('SIRPalphaV1/SIRPalphaV1', 'Var', (169, 192)) ('tumor', 'Disease', (142, 147)) ('hSIRPalpha', 'Gene', (47, 57)) ('hSIRPalpha', 'Gene', '140885', (90, 100)) ('tumor', 'Disease', 'MESH:D009369', (142, 147)) ('enhanced', 'PosReg', (133, 141)) ('hSIRPalpha', 'Gene', '140885', (47, 57)) 6742 31801627 Indeed, human IgG1 and IgG4 variants of chimeric hSIRPalpha.40A bound to FcgammaRI while the human IgG2 variant did not (data not shown). ('FcgammaRI', 'Gene', '2209', (73, 82)) ('IgG1', 'Gene', (14, 18)) ('variants', 'Var', (28, 36)) ('human', 'Species', '9606', (93, 98)) ('hSIRPalpha', 'Gene', '140885', (49, 59)) ('IgG4', 'Gene', (23, 27)) ('IgG1', 'Gene', '16017', (14, 18)) ('FcgammaRI', 'Gene', (73, 82)) ('human', 'Species', '9606', (8, 13)) ('hSIRPalpha', 'Gene', (49, 59)) ('bound', 'Interaction', (64, 69)) 6743 31801627 In addition, human IgG1 and IgG4 Fc variants that minimize antibody Fc-FcgammaR interactions restored the enhancement of rituximab-mediated phagocytosis as compared to their wild-type counterparts (Additional file 7: Figure S4A), while similar mutations of the human IgG2 Fc did not further alter macrophage-dependent phagocytosis. ('IgG4 Fc', 'Gene', (28, 35)) ('rituximab-mediated', 'MPA', (121, 139)) ('human', 'Species', '9606', (13, 18)) ('IgG1', 'Gene', '16017', (19, 23)) ('human', 'Species', '9606', (261, 266)) ('rituximab', 'Chemical', 'MESH:D000069283', (121, 130)) ('variants', 'Var', (36, 44)) ('interactions', 'Interaction', (80, 92)) ('enhancement', 'PosReg', (106, 117)) ('IgG1', 'Gene', (19, 23)) 6748 31801627 Next, ADU-1805 was shown to enhance rituximab-induced phagocytosis, in a concentration-dependent manner, by human macrophages obtained from different human individuals (Fig. ('human', 'Species', '9606', (150, 155)) ('ADU-1805', 'Chemical', '-', (6, 14)) ('rituximab-induced', 'MPA', (36, 53)) ('enhance', 'PosReg', (28, 35)) ('human', 'Species', '9606', (108, 113)) ('ADU-1805', 'Var', (6, 14)) ('rituximab', 'Chemical', 'MESH:D000069283', (36, 45)) 6749 31801627 Also, ADU-1805 was shown to enhance rituximab-mediated cell killing by neutrophils in a concentration-dependent manner, through a process called trogocytosis (Fig. ('ADU-1805', 'Chemical', '-', (6, 14)) ('rituximab-mediated cell killing', 'CPA', (36, 67)) ('trogocytosis', 'Disease', (145, 157)) ('enhance', 'PosReg', (28, 35)) ('ADU-1805', 'Var', (6, 14)) ('rituximab', 'Chemical', 'MESH:D000069283', (36, 45)) 6750 31801627 ADU-1805 lacked binding to human RBCs and platelets, and did not trigger hemagglutination, which is in line with its binding characteristics (Fig. ('ADU-1805', 'Var', (0, 8)) ('ADU-1805', 'Chemical', '-', (0, 8)) ('RBC', 'Gene', (33, 36)) ('RBC', 'Gene', '106480993', (33, 36)) ('human', 'Species', '9606', (27, 32)) ('lacked', 'NegReg', (9, 15)) ('binding', 'Interaction', (16, 23)) ('trigger', 'Reg', (65, 72)) ('not', 'NegReg', (61, 64)) ('hemagglutination', 'MPA', (73, 89)) 6752 31801627 The restricted expression of SIRPalpha was further demonstrated by comparing the reactivity of ADU-1805 and anti-CD47 towards human PBMCs. ('anti-CD47', 'Var', (108, 117)) ('human', 'Species', '9606', (126, 131)) ('ADU-1805', 'Gene', (95, 103)) ('ADU-1805', 'Chemical', '-', (95, 103)) 6754 31801627 Altogether, based on the presented in vitro data, this confirms the hypothesis that ADU-1805 will show a biological activity profile differentiated from CD47-targeting agents by its more restrictive binding pattern (i.e. ('binding', 'Interaction', (199, 206)) ('ADU-1805', 'Var', (84, 92)) ('ADU-1805', 'Chemical', '-', (84, 92)) 6759 31801627 ADU-1805 did not alter the T-cell secretion of IFNgamma triggered by the allogeneic MLR, whereas anti-CD47 mAb treatment inhibited IFNgamma secretion (Fig. ('ADU-1805', 'Chemical', '-', (0, 8)) ('IFNgamma', 'Gene', '3458', (131, 139)) ('IFNgamma', 'Gene', (131, 139)) ('T-cell secretion of', 'MPA', (27, 46)) ('IFNgamma', 'Gene', '3458', (47, 55)) ('IFNgamma', 'Gene', (47, 55)) ('anti-CD47', 'Var', (97, 106)) ('inhibited', 'NegReg', (121, 130)) 6763 31801627 To complement the nonclinical antibody development, we demonstrated that ADU-1805 did not engage FcgammaRIIA, nor did it induce ADCC via FcgammaR-bearing NK cells (Additional file 11: Figure S8A, B). ('ADU-1805', 'Var', (73, 81)) ('FcgammaRI', 'Gene', '2209', (97, 106)) ('ADCC', 'Disease', (128, 132)) ('induce', 'Reg', (121, 127)) ('ADU-1805', 'Chemical', '-', (73, 81)) ('FcgammaRI', 'Gene', (97, 106)) 6764 31801627 In addition, ADU-1805 did not induce CDC of the SIRPalpha-expressing U-937 AML cell line, consistent with the observation that human IgG2 is a poor C1q binder (Additional file 11: Figure S8C). ('C1q', 'Gene', '712', (148, 151)) ('human', 'Species', '9606', (127, 132)) ('AML', 'Disease', 'MESH:D015470', (75, 78)) ('C1q', 'Gene', (148, 151)) ('CD', 'Chemical', 'MESH:D002104', (37, 39)) ('U-937', 'CellLine', 'CVCL:0007', (69, 74)) ('AML', 'Phenotype', 'HP:0004808', (75, 78)) ('AML', 'Disease', (75, 78)) ('ADU-1805', 'Var', (13, 21)) ('ADU-1805', 'Chemical', '-', (13, 21)) 6765 31801627 Furthermore, ADU-1805 did not induce cytokine secretion in human whole blood, similar to the FDA-approved human IgG2 antibody panitumumab targeting epidermal growth factor receptor (EGFR) (Additional file 12: Figure S9). ('human', 'Species', '9606', (106, 111)) ('human', 'Species', '9606', (59, 64)) ('ADU-1805', 'Var', (13, 21)) ('EGFR', 'Gene', '1956', (182, 186)) ('EGFR', 'Gene', (182, 186)) ('ADU-1805', 'Chemical', '-', (13, 21)) ('epidermal growth factor receptor', 'Gene', (148, 180)) ('panitumumab', 'Chemical', 'MESH:D000077544', (126, 137)) ('epidermal growth factor receptor', 'Gene', '1956', (148, 180)) 6767 31801627 First, two putative variants, SIRPalphaV1 (NM_001284750.1) and SIRPalphaV2 (XP_015313155.1) were identified in cynomolgus monkey, that share 99.2% sequence identity. ('XP_015313155.1', 'Var', (76, 90)) ('NM_001284750.1', 'Var', (43, 57)) ('cynomolgus monkey', 'Species', '9541', (111, 128)) ('SIRPalphaV1', 'Gene', (30, 41)) 6768 31801627 These variants share a sequence identity of > 91% with human SIRPalphaV1 and SIRPalphaV2 and ADU-1805 bound to both cynomolgus variants with an EC50 <= 1 nM, similar to its binding affinity for human SIRPalpha (Additional file 13: Figure S10A). ('ADU-1805', 'Gene', (93, 101)) ('variants', 'Var', (6, 14)) ('SIRPalphaV2', 'Gene', (77, 88)) ('ADU-1805', 'Chemical', '-', (93, 101)) ('human', 'Species', '9606', (55, 60)) ('S10A', 'SUBSTITUTION', 'None', (238, 242)) ('cynomolgus', 'Species', '9541', (116, 126)) ('human', 'Species', '9606', (194, 199)) ('S10A', 'Var', (238, 242)) ('bound', 'Interaction', (102, 107)) 6769 31801627 Furthermore, the ADU-1805 binding profile was comparable for human and cynomolgus monkey leukocytes (Additional file 13: Figure S10B). ('cynomolgus monkey', 'Species', '9541', (71, 88)) ('S10B', 'SUBSTITUTION', 'None', (128, 132)) ('binding', 'Interaction', (26, 33)) ('human', 'Species', '9606', (61, 66)) ('ADU-1805', 'Protein', (17, 25)) ('S10B', 'Var', (128, 132)) ('ADU-1805', 'Chemical', '-', (17, 25)) 6773 31801627 This finding supports that targeting SIRPalpha via ADU-1805 may have a favorable safety profile compared to CD47-targeting agents (Fig. ('SIRPalpha', 'Gene', (37, 46)) ('ADU-1805', 'Var', (51, 59)) ('ADU-1805', 'Chemical', '-', (51, 59)) 6779 31801627 Hu5F9-G4, TTI-621) induce acute anemia and thrombocytopenia in patients which may also further depend on the Fc format. ('TTI-621', 'Gene', (10, 17)) ('thrombocytopenia', 'Disease', (43, 59)) ('acute anemia', 'Disease', (26, 38)) ('Hu5F9-G4', 'Var', (0, 8)) ('anemia', 'Phenotype', 'HP:0001903', (32, 38)) ('thrombocytopenia', 'Disease', 'MESH:D013921', (43, 59)) ('acute anemia', 'Disease', 'MESH:D000740', (26, 38)) ('patients', 'Species', '9606', (63, 71)) ('thrombocytopenia', 'Phenotype', 'HP:0001873', (43, 59)) 6781 31801627 In contrast, the acute toxicity initially observed with Hu5F9-G4 was ultimately managed by adopting a dosing strategy that involved a priming (1 mg/kg priming on day 1) and maintenance phase (30 mg/kg weekly for 3 doses and 30 mg/kg every other week thereafter). ('toxicity', 'Disease', (23, 31)) ('Hu5F9-G4', 'Var', (56, 64)) ('toxicity', 'Disease', 'MESH:D064420', (23, 31)) 6784 31801627 Hence, ALX148 may induce reduced toxicity at the expense of single agent activity, similar to that seen with high-affinity SIRPalpha variants that lack an Fc chain. ('ALX148', 'Var', (7, 13)) ('ALX148', 'Chemical', '-', (7, 13)) ('reduced', 'NegReg', (25, 32)) ('toxicity', 'Disease', 'MESH:D064420', (33, 41)) ('Fc chain', 'Protein', (155, 163)) ('variants', 'Var', (133, 141)) ('SIRPalpha', 'Gene', (123, 132)) ('toxicity', 'Disease', (33, 41)) 6798 31801627 In addition, ADU-1805 also binds to SIRPgamma expressed on T-cells, albeit with a 2.9-fold reduced EC50 compared to SIRPalpha, and thus may block the SIRPgamma-CD47 interaction. ('interaction', 'Interaction', (165, 176)) ('binds', 'Interaction', (27, 32)) ('EC50', 'MPA', (99, 103)) ('reduced', 'NegReg', (91, 98)) ('SIRPgamma', 'Gene', '55423', (150, 159)) ('SIRPgamma', 'Gene', '55423', (36, 45)) ('ADU-1805', 'Var', (13, 21)) ('SIRPgamma', 'Gene', (150, 159)) ('SIRPgamma', 'Gene', (36, 45)) ('ADU-1805', 'Chemical', '-', (13, 21)) ('block', 'NegReg', (140, 145)) 6799 31801627 Targeting of this interaction using anti-CD47 mAbs or an anti-SIRPgamma mAb, was previously shown to inhibit T-cell proliferation in an allogeneic MLR. ('SIRPgamma', 'Gene', '55423', (62, 71)) ('inhibit', 'NegReg', (101, 108)) ('anti-CD47', 'Var', (36, 45)) ('SIRPgamma', 'Gene', (62, 71)) ('T-cell proliferation', 'CPA', (109, 129)) 6800 31801627 We therefore assessed whether ADU-1805 alters T-cell activation using a similar assay. ('ADU-1805', 'Var', (30, 38)) ('ADU-1805', 'Chemical', '-', (30, 38)) ('T-cell activation', 'CPA', (46, 63)) 6801 31801627 Despite minimal binding to T-cells, ADU-1805 did not affect T-cell activation in an allogeneic MLR. ('binding', 'Interaction', (16, 23)) ('T-cell', 'CPA', (60, 66)) ('ADU-1805', 'Var', (36, 44)) ('ADU-1805', 'Chemical', '-', (36, 44)) 6805 31801627 tumor shrinkage by means of macrophage phagocytosis) with anti-CD47 have been observed in patients, longer follow-up is needed to address T-cell activation, since this may be required for durability of clinical responses. ('tumor', 'Disease', 'MESH:D009369', (0, 5)) ('anti-CD47', 'Var', (58, 67)) ('patients', 'Species', '9606', (90, 98)) ('tumor', 'Phenotype', 'HP:0002664', (0, 5)) ('tumor', 'Disease', (0, 5)) 6807 31801627 Similar observations were made for antibodies directed against colony-stimulating factor 1 receptor (CSF1R), where for H27K15, a non-ligand competitive anti-CSF1R mAb, it was shown that the Fc region participates in its mode of action, suggestive of a similar scorpion effect. ('CSF1R', 'Gene', '1436', (157, 162)) ('colony-stimulating factor 1 receptor', 'Gene', (63, 99)) ('CSF1R', 'Gene', (101, 106)) ('colony-stimulating factor 1 receptor', 'Gene', '1436', (63, 99)) ('CSF1R', 'Gene', (157, 162)) ('CSF1R', 'Gene', '1436', (101, 106)) ('H27K15', 'Var', (119, 125)) 6808 31801627 Our data suggest that SIRPalpha targeting by ADU-1805 can activate myeloid cell types such as neutrophils and macrophages. ('ADU-1805', 'Chemical', '-', (45, 53)) ('targeting', 'Var', (32, 41)) ('activate', 'PosReg', (58, 66)) ('neutrophils', 'CPA', (94, 105)) ('myeloid cell types', 'CPA', (67, 85)) ('ADU-1805', 'Gene', (45, 53)) ('SIRPalpha', 'Protein', (22, 31)) 6809 31801627 The role of neutrophils and macrophages upon SIRPalpha blockade is further confirmed in xenograft mouse models that are deficient in T-cells, B-cells and NK cells. ('mouse', 'Species', '10090', (98, 103)) ('SIRPalpha', 'Gene', (45, 54)) ('blockade', 'Var', (55, 63)) 6812 31801627 Repolarization of TAMs into M1 type macrophages may skew the microenvironment towards becoming more pro-inflammatory thereby promoting the anti-tumor immune response. ('tumor', 'Disease', (144, 149)) ('promoting', 'PosReg', (125, 134)) ('pro-inflammatory', 'MPA', (100, 116)) ('tumor', 'Disease', 'MESH:D009369', (144, 149)) ('Repolarization', 'Var', (0, 14)) ('tumor', 'Phenotype', 'HP:0002664', (144, 149)) ('TAMs', 'Chemical', 'MESH:D013629', (18, 22)) 6813 31801627 Moreover, anti-SIRPalpha treatment led to a marked increase in the number of tumor-infiltrating NK cells and CD8+ T-cells, and antibody-mediated depletion of these cells decreased the inhibitory effect of SIRPalpha blockade on tumor formation. ('increase', 'PosReg', (51, 59)) ('CD8', 'Gene', (109, 112)) ('CD8', 'Gene', '925', (109, 112)) ('tumor', 'Disease', (227, 232)) ('SIRPalpha blockade on tumor', 'Disease', (205, 232)) ('tumor', 'Disease', 'MESH:D009369', (77, 82)) ('SIRPalpha blockade on tumor', 'Disease', 'MESH:D009369', (205, 232)) ('anti-SIRPalpha', 'Var', (10, 24)) ('decreased', 'NegReg', (170, 179)) ('tumor', 'Phenotype', 'HP:0002664', (77, 82)) ('tumor', 'Disease', 'MESH:D009369', (227, 232)) ('tumor', 'Disease', (77, 82)) ('tumor', 'Phenotype', 'HP:0002664', (227, 232)) 6814 31801627 Finally, anti-SIRPalpha mAbs have the ability to enhance the activity of immune checkpoint inhibitors such as anti-PD-1 and this has been confirmed in vivo with the CD47-blocking molecule ALX148 in combination with anti-PD-1 or anti-PD-L1 therapy. ('PD-1', 'Gene', '5133', (220, 224)) ('enhance', 'PosReg', (49, 56)) ('PD-1', 'Gene', (220, 224)) ('PD-L1', 'Gene', (233, 238)) ('PD-1', 'Gene', (115, 119)) ('activity', 'MPA', (61, 69)) ('PD-1', 'Gene', '5133', (115, 119)) ('anti-SIRPalpha', 'Var', (9, 23)) ('ALX148', 'Chemical', '-', (188, 194)) ('PD-L1', 'Gene', '29126', (233, 238)) 6820 31801627 ADU-1805 is a potentially best-in-class antagonistic SIRPalpha-targeting antibody with a unique epitope that encompasses pan-allele SIRPalpha binding. ('ADU-1805', 'Var', (0, 8)) ('binding', 'Interaction', (142, 149)) ('ADU-1805', 'Chemical', '-', (0, 8)) 6821 31801627 Unlike anti-CD47 mAbs, ADU-1805 does not trigger depletion of RBCs and platelets when tested at increasing dose levels in NHPs, supporting its favorable safety profile. ('RBC', 'Gene', '106480993', (62, 65)) ('RBC', 'Gene', (62, 65)) ('depletion', 'MPA', (49, 58)) ('ADU-1805', 'Var', (23, 31)) ('ADU-1805', 'Chemical', '-', (23, 31)) 6843 30645099 Interestingly, two other minor groove DNA binders recently showed interesting anti-leukemic activities by targeting transcription factors associated with acute myeloid leukemia (AML): the alkylating agent-conjugated pyrrole-imidazole polyamide Chb-M against RUNX1 and DB2313 against PU.1. ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (154, 176)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (160, 176)) ('leukemic', 'Disease', 'MESH:D007938', (83, 91)) ('leukemia', 'Phenotype', 'HP:0001909', (168, 176)) ('DB2313', 'Var', (268, 274)) ('targeting', 'Reg', (106, 115)) ('AML', 'Disease', 'MESH:D015470', (178, 181)) ('acute myeloid leukemia', 'Disease', (154, 176)) ('RUNX1', 'Gene', '12394', (258, 263)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (154, 176)) ('AML', 'Disease', (178, 181)) ('leukemic', 'Disease', (83, 91)) ('RUNX1', 'Gene', (258, 263)) ('PU.1', 'Gene', '20375', (283, 287)) ('pyrrole-imidazole polyamide', 'Chemical', 'MESH:C009916', (216, 243)) ('PU.1', 'Gene', (283, 287)) 6845 30645099 Besides its implication in some solid tumors, HOXA9 is clearly associated with different hematologic malignancies. ('tumor', 'Phenotype', 'HP:0002664', (38, 43)) ('solid tumors', 'Disease', 'MESH:D009369', (32, 44)) ('tumors', 'Phenotype', 'HP:0002664', (38, 44)) ('associated', 'Reg', (63, 73)) ('HOXA9', 'Var', (46, 51)) ('hematologic malignancies', 'Disease', 'MESH:D019337', (89, 113)) ('solid tumors', 'Disease', (32, 44)) ('hematologic malignancies', 'Disease', (89, 113)) 6849 30645099 Indeed, deletion of its homeodomain abolishes the propensity of HOXA9 to induce leukemia in mice. ('abolishes', 'NegReg', (36, 45)) ('leukemia', 'Disease', (80, 88)) ('leukemia', 'Phenotype', 'HP:0001909', (80, 88)) ('propensity', 'MPA', (50, 60)) ('leukemia', 'Disease', 'MESH:D007938', (80, 88)) ('mice', 'Species', '10090', (92, 96)) ('deletion', 'Var', (8, 16)) 6856 30645099 We were specifically interested in heterocyclic diamidine compounds that have been previously reported to be potent in vitro transcription factor DNA binding inhibitors: DB293 against Pit-1 and Brn-3 and DB1255 against the ERG transcription factor. ('Pit-1', 'Gene', '18736', (184, 189)) ('DB293', 'CellLine', 'CVCL:8468', (170, 175)) ('DB1255', 'Chemical', 'MESH:C579039', (204, 210)) ('ERG', 'Gene', (223, 226)) ('Brn-3', 'Gene', '18996', (194, 199)) ('Brn-3', 'Gene', (194, 199)) ('DB1255', 'Var', (204, 210)) ('Pit-1', 'Gene', (184, 189)) ('diamidine', 'Chemical', 'MESH:D010419', (48, 57)) ('ERG', 'Gene', '13876', (223, 226)) ('DB293', 'Var', (170, 175)) 6860 30645099 Modifications of the R1 ring and the amidine extremities (R4 and R5) offer a large diversity of molecules to expand structure/function relationships (Table 1). ('structure/function', 'MPA', (116, 134)) ('amidine', 'Chemical', 'MESH:D000578', (37, 44)) ('mole', 'Phenotype', 'HP:0003764', (96, 100)) ('Modifications', 'Var', (0, 13)) 6865 30645099 In the case of DB928, the hydrogenolysis in addition to cleaving the N-OAc bond also cleaved the benzyl group. ('benzyl group', 'MPA', (97, 109)) ('cleaved', 'Reg', (85, 92)) ('N-OAc bond', 'MPA', (69, 79)) ('benzyl', 'Chemical', 'MESH:C503494', (97, 103)) ('DB928', 'Var', (15, 20)) ('N-OAc', 'Chemical', 'MESH:C065699', (69, 74)) 6870 30645099 In Scheme 3 the synthesis approach to five (11a-e; DB1478, DB1481, DB1879, DB2005, DB2529) heteroaryl indole diamidines is presented. ('DB1478', 'Var', (51, 57)) ('indole diamidines', 'Chemical', 'MESH:D010419', (102, 119)) ('DB2529', 'Var', (83, 89)) ('DB1481', 'Var', (59, 65)) ('DB1879', 'Var', (67, 73)) ('DB2005', 'Var', (75, 81)) 6891 30645099 By contrast, changing the thiophene ring of DB818, DB1879 or DB2529 to a furan (DB293, DB1878 or DB915, respectively) or a thiazole ring (DB2651) in the benzimidazole series strongly reduces HOXA9 inhibition efficacy whereas a pyrazole ring modification still confers some weaker inhibitory activity to DB2223. ('DB293', 'CellLine', 'CVCL:8468', (80, 85)) ('benzimidazole', 'Chemical', 'MESH:C031000', (153, 166)) ('thiazole', 'Chemical', 'MESH:D013844', (123, 131)) ('DB2529', 'Var', (61, 67)) ('thiophene', 'Chemical', 'MESH:D013876', (26, 35)) ('pyrazole', 'Chemical', 'MESH:D011720', (227, 235)) ('HOXA9', 'Enzyme', (191, 196)) ('reduces', 'NegReg', (183, 190)) ('DB1879', 'Var', (51, 57)) ('DB818', 'Var', (44, 49)) ('furan', 'Chemical', 'MESH:D005663', (73, 78)) ('inhibitory activity', 'MPA', (280, 299)) ('changing', 'Var', (13, 21)) 6892 30645099 Surprisingly, such modification of the thiophene ring to a thiazole ring in the di-phenyl series changes an inactive compound (DB351) to an effective (but low) inhibitor of HOXA9/DNA binding (DB1620). ('modification', 'Var', (19, 31)) ('di-phenyl', 'Chemical', 'MESH:C076868', (80, 89)) ('thiophene', 'Chemical', 'MESH:D013876', (39, 48)) ('thiazole', 'Chemical', 'MESH:D013844', (59, 67)) ('changes', 'Reg', (97, 104)) 6893 30645099 The addition of a methyl group on the benzimidazole ring (DB818) at the R3 position or changing the DB818 benzimidazole ring to a quinoline ring or to a N-isopropyl-benzimidazole-phenyl moiety strongly alters its propensity to inhibit the HOXA9/DNA interaction (see DB2429, DB1504 or DB2708) relative to DB818. ('inhibit', 'NegReg', (227, 234)) ('methyl', 'Chemical', 'MESH:C031105', (18, 24)) ('propensity', 'MPA', (213, 223)) ('benzimidazole', 'Chemical', 'MESH:C031000', (106, 119)) ('benzimidazole', 'Chemical', 'MESH:C031000', (165, 178)) ('alters', 'Reg', (202, 208)) ('DB818', 'Gene', (100, 105)) ('phenyl', 'Chemical', 'MESH:C064635', (179, 185)) ('benzimidazole', 'Chemical', 'MESH:C031000', (38, 51)) ('HOXA9/DNA interaction', 'MPA', (239, 260)) ('quinoline', 'Chemical', 'MESH:D011804', (130, 139)) ('changing', 'Var', (87, 95)) 6894 30645099 A methyl group on the phenyl ring in R1 also alters the inhibitory effect of DB1878 (unmethylated) as shown using DB2005 (methylated) (Figure 1A). ('inhibitory effect', 'MPA', (56, 73)) ('phenyl', 'Chemical', 'MESH:C064635', (22, 28)) ('methyl', 'Var', (2, 8)) ('DB1878', 'Gene', (77, 83)) ('methyl', 'Chemical', 'MESH:C031105', (87, 93)) ('alters', 'Reg', (45, 51)) ('methyl', 'Chemical', 'MESH:C031105', (122, 128)) ('methyl', 'Chemical', 'MESH:C031105', (2, 8)) 6895 30645099 Moreover, the addition of a methyl group to the benzimidazole ring in the R3 position alters the HOXA9 inhibition efficiency as observed with the highly active unmethylated compounds DB818 and DB2529, relative to their inactive methylated counterparts DB2429 and DB2430 (of note, this latter also differs by the presence of a furan 5-membered ring in R2 position instead of a thiophene). ('methyl', 'Chemical', 'MESH:C031105', (28, 34)) ('HOXA9', 'Enzyme', (97, 102)) ('DB818', 'Var', (183, 188)) ('methyl', 'Chemical', 'MESH:C031105', (228, 234)) ('methyl', 'Chemical', 'MESH:C031105', (162, 168)) ('benzimidazole', 'Chemical', 'MESH:C031000', (48, 61)) ('alters', 'Reg', (86, 92)) ('thiophene', 'Chemical', 'MESH:D013876', (376, 385)) ('furan', 'Chemical', 'MESH:D005663', (326, 331)) ('DB2529', 'Var', (193, 199)) ('inhibition', 'NegReg', (103, 113)) 6896 30645099 Finally, the two pyrrole derivatives bearing an additional ring (phenyl for DB884 or pyridine for DB883) at both the R1/R5 positions are identified as high (DB884) and medium (DB883) inhibitors. ('DB883', 'Var', (98, 103)) ('DB884', 'Var', (76, 81)) ('pyridine', 'Chemical', 'MESH:C023666', (85, 93)) ('phenyl', 'Chemical', 'MESH:C064635', (65, 71)) ('pyrrole', 'Chemical', 'MESH:D011758', (17, 24)) 6901 30645099 This observation indicates that the diamidine compounds bind within the groove of the HBS DNA sequence, more likely within the minor groove as strongly evidenced in the crystal and/or NMR structure for DB818, DB1055, DB884 or DB293 to 5'-AATT-containing DNA (PDB: 1VZK, 2I5A, 2I2I, 2GYX). ('PDB', 'Disease', (259, 262)) ('DB884', 'Var', (217, 222)) ('HBS DNA', 'Gene', (86, 93)) ('diamidine', 'Chemical', 'MESH:D010419', (36, 45)) ('DB293 to', 'Var', (226, 234)) ('DB1055', 'Var', (209, 215)) ('PDB', 'Disease', 'MESH:D010001', (259, 262)) ('DB293', 'CellLine', 'CVCL:8468', (226, 231)) ('DB818', 'Var', (202, 207)) 6902 30645099 To define the specific drug/DNA binding sequence, DNaseI footprinting experiments were used for selected compounds from the "High" (DB818, DB1055, DB1879, DB2529), "Medium" (DB1177, DB883), "low" (DB1878) and "inactive" (DB828) subgroups for HOXA9/DNA binding inhibition. ('DB1879', 'Var', (147, 153)) ('DB883', 'Var', (182, 187)) ('DB2529', 'Var', (155, 161)) ('DB1055', 'Var', (139, 145)) ('low" (DB1878', 'Var', (191, 203)) ('DNaseI', 'Gene', '13419', (50, 56)) ('DNaseI', 'Gene', (50, 56)) 6905 30645099 We have mutated the sequence at the 5'-ATGA (HBS-ATGAm) or the AT-rich (HBS-ATm) or containing both mutations (HBS-2mut) (oligonucleotide sequences are presented in Supplementary Table S1). ('ATm', 'Gene', '11920', (76, 79)) ('mutated', 'Var', (8, 15)) ('mutations', 'Var', (100, 109)) ('ATm', 'Gene', (76, 79)) 6906 30645099 For the HBS-ATGAm sequence, the DeltaTm values obtained with DB818 and DB1055 were similar to that obtained with wild-type HBS oligonucleotide, which suggests that the diamidine compounds do not bind at the 5'-ATG binding site of HBS (Figure 2C). ('DB1055', 'Var', (71, 77)) ('bind', 'Interaction', (195, 199)) ('diamidine', 'Chemical', 'MESH:D010419', (168, 177)) ('DB818', 'Var', (61, 66)) 6907 30645099 By contrast, when the 5'-ATTTA site was mutated alone for HBS-ATm or together with the ATGA portion for HBS-2mut, DeltaTm values decreased by 4 C, which is similar to the observed value for the DB828. ('ATm', 'Gene', (62, 65)) ('ATm', 'Gene', '11920', (62, 65)) ('mutated', 'Var', (40, 47)) ('DeltaTm values', 'MPA', (114, 128)) ('ether', 'Chemical', 'MESH:D004987', (72, 77)) ('decreased', 'NegReg', (129, 138)) 6908 30645099 This result is in agreement with the DNase I footprinting experiments which suggest that, among other, DB818 and DB1055 interact at the 5'-ATTTA-sequence of the HOXA9 binding site (Figure 2) but did not bind to an HBS-mutated DNA as shown in Supplementary Figure S2. ('DNase I', 'Gene', (37, 44)) ('DB1055', 'Var', (113, 119)) ('DB818', 'Gene', (103, 108)) ('interact', 'Interaction', (120, 128)) ('DNase I', 'Gene', '13419', (37, 44)) 6911 30645099 Two amidine groups, -N-Hs form H-bond either with T=O of dT1' base or A-N3 of dA5 base (Figure 4C-D) that are an average of 2.3 A in length. ('H-bond', 'MPA', (31, 37)) ('dT1', 'Gene', '5657400', (57, 60)) ('A-N3', 'Var', (70, 74)) ('amidine', 'Chemical', 'MESH:D000578', (4, 11)) ('dT1', 'Gene', (57, 60)) 6923 30645099 In this cell line, both DB818, DB1055 and DB828 compounds have no effect on cell survival and clonogenic propensities (Supplementary Figure S5B, Table 4, top lanes). ('lanes', 'Chemical', 'MESH:C480782', (158, 163)) ('clonogenic propensities', 'CPA', (94, 117)) ('cell survival', 'CPA', (76, 89)) ('DB1055', 'Var', (31, 37)) ('DB818', 'Var', (24, 29)) ('DB828', 'Gene', (42, 47)) 6927 30645099 GSEA analyses on MSigDB C2-Chemical and Genetic Perturbation gene sets (Figure 6B) evidenced that the significant genes up-regulated in DB818-treated MigA9 cells at the different time correspond to genes that were down-regulated in human CD34+ cells transduced with NUP98-HOXA9 fusion (TAKEDA TARGETS OF NUP98 HOXA9 FUSION 8D DN), Figure 6B. ('CD34', 'Gene', '947', (238, 242)) ('CD34', 'Gene', (238, 242)) ('human', 'Species', '9606', (232, 237)) ('up-regulated', 'PosReg', (120, 132)) ('DB818-treated', 'Var', (136, 149)) 6939 30645099 DB818 and DB1055 would abolish such negative control of Akap13 expression by HOXA9, leading to an overexpression of Akap13 as evidenced by qRT-PCR (Figure 6D) for this gene which over-expression is associated with granulocyte and monocyte differentiation (Supplementary Figure S7). ('Akap13', 'Gene', (116, 122)) ('over-expression', 'PosReg', (179, 194)) ('Akap13', 'Gene', '75547', (56, 62)) ('overexpression', 'PosReg', (98, 112)) ('Akap13', 'Gene', (56, 62)) ('Akap13', 'Gene', '75547', (116, 122)) ('DB818', 'Var', (0, 5)) ('DB1055', 'Var', (10, 16)) ('abolish', 'NegReg', (23, 30)) 6944 30645099 Interestingly, the five ring containing compounds DB883 and DB884 were the most active in terms of cell death, suggesting that cell death is the main cellular process that is responsible for the decrease of cell survival by DB883 and DB884. ('five ring', 'Species', '270470', (19, 28)) ('DB884', 'Var', (234, 239)) ('decrease', 'NegReg', (195, 203)) ('cell survival', 'CPA', (207, 220)) ('cell death', 'CPA', (127, 137)) ('DB883', 'Var', (224, 229)) 6945 30645099 This reduction of cycling cells is associated with a strong increase in the G0/G1 phase (DB1055, DB1878, DB2529), in the G2/M phase (DB818) or in both phases (DB1879, DB1620) at the tested concentrations and at 48H treatment. ('G2/M phase', 'CPA', (121, 131)) ('DB1878', 'Var', (97, 103)) ('DB2529', 'Var', (105, 111)) ('G0/G1 phase', 'CPA', (76, 87)) ('reduction', 'NegReg', (5, 14)) ('DB1620', 'Var', (167, 173)) ('DB1879', 'Var', (159, 165)) ('DB1055', 'Var', (89, 95)) ('increase', 'PosReg', (60, 68)) ('8H', 'Chemical', 'MESH:D006859', (212, 214)) ('cycling cells', 'CPA', (18, 31)) 6946 30645099 This experiment first highlights the impact of both DB818 and DB1055 on global clonogenic propensity of the HOXA9-positive MigA9 cell line, but not on the HOXA9-negative K562 cell line (Table 4 and Supplementary Figure S6C-D). ('K562', 'CellLine', 'CVCL:0004', (170, 174)) ('DB818', 'Var', (52, 57)) ('DB1055', 'Var', (62, 68)) 6947 30645099 Surprisingly, the intrinsic fluorescence of DB818 and DB828 were still visible even after 10 days of culture in Methocult using fluorescence microscopy with DAPI-filters, which indicates very high stability of the diamidine compounds within the cells (Supplementary Figure S6D). ('intrinsic fluorescence', 'MPA', (18, 40)) ('diamidine', 'Chemical', 'MESH:D010419', (214, 223)) ('DB818', 'Var', (44, 49)) ('DB828', 'Var', (54, 59)) 6948 30645099 Results for clonogenic assays clearly show a reduction in the number of colony sub-types obtained in the presence of DB818 or DB1055, but not DB828, for the untreated wells (CTR) (Table 4). ('CTR', 'Gene', (174, 177)) ('DB1055', 'Var', (126, 132)) ('CTR', 'Gene', '12311', (174, 177)) ('reduction', 'NegReg', (45, 54)) ('DB818', 'Var', (117, 122)) 6950 30645099 These results suggest that both DB818 and DB1055 induced a decrease in the number of the most immature hematopoietic progenitors cells (CFU-GM) and favored the development of colonies that are more engaged in granulocytic or monocytic differentiation (CFU-G, CFU-M). ('favored', 'PosReg', (148, 155)) ('DB818', 'Var', (32, 37)) ('granulocytic or monocytic differentiation', 'CPA', (209, 250)) ('DB1055', 'Var', (42, 48)) ('decrease', 'NegReg', (59, 67)) ('CFU-GM', 'Chemical', 'MESH:C029624', (136, 142)) 6951 30645099 This result is in agreement with gene expression analyses with DB818 (GSEA, IPA analyses), confirmed by qRT-PCR for both DB818 and DB1055 (but not for DB828) on Itgam (CD11b) as well as on CD14 expression as two markers of monocytic to macrophagic differentiation. ('Itgam', 'Gene', (161, 166)) ('CD11b', 'Gene', (168, 173)) ('CD11b', 'Gene', '16409', (168, 173)) ('CD14', 'Gene', '12475', (189, 193)) ('Itgam', 'Gene', '16409', (161, 166)) ('CD14', 'Gene', (189, 193)) ('DB1055', 'Var', (131, 137)) ('IPA', 'Chemical', 'MESH:C507406', (76, 79)) ('DB818', 'Gene', (121, 126)) 6953 30645099 In the present work, we selected a series of heterocyclic diamidines and derivatives as sequence-selective DNA ligands that directly inhibit HOXA9/DNA binding (Figure 1, Figure 2A) through groove binding (Figure 2B) on the HOXA9 cognate DNA sequence (Figure 2). ('heterocyclic diamidines', 'Chemical', 'MESH:D010419', (45, 68)) ('HOXA9/DNA', 'Protein', (141, 150)) ('inhibit', 'NegReg', (133, 140)) ('groove', 'Var', (189, 195)) 6955 30645099 DNase I footprinting experiments also evidenced that DB818 and DB1055 appeared to be very active and selective compounds on binding to the HOXA9-binding site (Figure 2) but not to the mutated HBS sequence (Supplementary Figure S2). ('DB1055', 'Gene', (63, 69)) ('DNase I', 'Gene', '13419', (0, 7)) ('DB818', 'Var', (53, 58)) ('DNase I', 'Gene', (0, 7)) ('binding', 'Interaction', (124, 131)) 6959 30645099 Such DNA distortion towards the minor groove results from interaction of amino acids of the amino-terminal arm (Pro-7 and Tyr-8) with helix-1 of the homeodomain of HOXA9 (Thr-13). ('amino', 'Chemical', 'MESH:D000596', (73, 78)) ('Tyr', 'Chemical', 'MESH:C042696', (122, 125)) ('interaction', 'Interaction', (58, 69)) ('results from', 'Reg', (45, 57)) ('amino', 'Chemical', 'MESH:D000596', (92, 97)) ('Tyr-8', 'Var', (122, 127)) ('Thr', 'Chemical', 'MESH:C055175', (171, 174)) 6960 30645099 At the cellular level, both DB818 and DB1055 control HOXA9-directed transcription based on luciferase gene expression assays (Figure 5B) and induce, among other derivatives, cell death, cell cycle arrest and cell differentiation in a model of Hoxa9-transformed murine MigA9 leukemia cell line (Tables 2-4, Figure 6 and Supplementary Figure S6). ('Hoxa9', 'Gene', (243, 248)) ('cell cycle arrest', 'Phenotype', 'HP:0011018', (186, 203)) ('induce', 'Reg', (141, 147)) ('cell death', 'CPA', (174, 184)) ('DB1055', 'Var', (38, 44)) ('Hoxa9', 'Gene', '15405', (243, 248)) ('cell differentiation', 'CPA', (208, 228)) ('HOXA9-directed transcription', 'Gene', (53, 81)) ('cell cycle arrest', 'CPA', (186, 203)) ('DB818', 'Var', (28, 33)) ('murine', 'Species', '10090', (261, 267)) ('control', 'Reg', (45, 52)) ('leukemia', 'Phenotype', 'HP:0001909', (274, 282)) ('leukemia', 'Disease', (274, 282)) ('leukemia', 'Disease', 'MESH:D007938', (274, 282)) 6961 30645099 DB828 also failed to alter cell survival (MTS, Table 2), to induce cell death (PI-positive or sub-G1 populations analysis, Table 2) and MigA9 cell differentiation (anti-clonogenic activities and CD11b/CD14 differentiation markers expression, Table 4 and Figure 6). ('CD11b', 'Gene', (195, 200)) ('CD11b', 'Gene', '16409', (195, 200)) ('DB828', 'Var', (0, 5)) ('CD14', 'Gene', '12475', (201, 205)) ('CD14', 'Gene', (201, 205)) ('MigA9 cell differentiation', 'CPA', (136, 162)) ('cell death', 'CPA', (67, 77)) 6965 30645099 Some other heterocyclic diamidines were previously identified as transcription factors inhibitors, DB293 against Pit-1 and Brn-3, DB1255 against ERG and DB2113 against PU.1 but none were previously evaluated against the HOXA9 transcription factor. ('Pit-1', 'Gene', (113, 118)) ('Brn-3', 'Gene', '18996', (123, 128)) ('Brn-3', 'Gene', (123, 128)) ('PU.1', 'Gene', '20375', (168, 172)) ('heterocyclic diamidines', 'Chemical', 'MESH:D010419', (11, 34)) ('DB293', 'Var', (99, 104)) ('ERG', 'Gene', (145, 148)) ('DB2113', 'Var', (153, 159)) ('PU.1', 'Gene', (168, 172)) ('Pit-1', 'Gene', '18736', (113, 118)) ('DB1255', 'Chemical', 'MESH:C579039', (130, 136)) ('DB293', 'CellLine', 'CVCL:8468', (99, 104)) ('DB1255', 'Var', (130, 136)) ('ERG', 'Gene', '13876', (145, 148)) 6966 30645099 HOXA9 is one of the master regulators of the differentiation blockade in acute myeloid leukemia, being at the crossroads of many cell signaling perturbations resulting from well-characterized mutations or translocations defining AML subgroups such as for instance MLL translocations, MYST3-CREBBP or NUP98-NSD1 fusion proteins and NPM1 or EZH2 mutations. ('acute myeloid leukemia', 'Disease', (73, 95)) ('CREBBP', 'Gene', (290, 296)) ('mutations', 'Var', (344, 353)) ('leukemia', 'Phenotype', 'HP:0001909', (87, 95)) ('MLL', 'Gene', (264, 267)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (79, 95)) ('mutations', 'Var', (192, 201)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (73, 95)) ('NSD1', 'Gene', (306, 310)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (73, 95)) ('NPM1', 'Gene', (331, 335)) ('AML', 'Disease', 'MESH:D015470', (229, 232)) ('perturbations', 'Disease', (144, 157)) ('EZH2', 'Gene', '14056', (339, 343)) ('perturbations', 'Disease', 'MESH:C536875', (144, 157)) ('NSD1', 'Gene', '18193', (306, 310)) ('AML', 'Disease', (229, 232)) ('MYST3', 'Gene', (284, 289)) ('MLL', 'Gene', '214162', (264, 267)) ('MYST3', 'Gene', '244349', (284, 289)) ('CREBBP', 'Gene', '12914', (290, 296)) ('EZH2', 'Gene', (339, 343)) ('NPM1', 'Gene', '18148', (331, 335)) 6967 30645099 Therefore, treatment of such AML subtypes with DB818/DB1055 needs to be further evaluated in the future to offer new potential differentiation therapies. ('AML', 'Disease', (29, 32)) ('AML', 'Disease', 'MESH:D015470', (29, 32)) ('DB818/DB1055', 'Var', (47, 59)) 6968 30645099 HOXA9 is also associated with different solid tumor models such as glioblastoma, gastric and prostate cancers against which DB818, DB1055 and/or derivatives might also be evaluated to propose future therapeutic strategies. ('glioblastoma', 'Disease', (67, 79)) ('cancer', 'Phenotype', 'HP:0002664', (102, 108)) ('glioblastoma', 'Disease', 'MESH:D005909', (67, 79)) ('DB818', 'Var', (124, 129)) ('tumor', 'Phenotype', 'HP:0002664', (46, 51)) ('glioblastoma', 'Phenotype', 'HP:0012174', (67, 79)) ('tumor', 'Disease', (46, 51)) ('associated', 'Reg', (14, 24)) ('gastric and prostate cancers', 'Disease', 'MESH:D011471', (81, 109)) ('cancers', 'Phenotype', 'HP:0002664', (102, 109)) ('prostate cancers', 'Phenotype', 'HP:0012125', (93, 109)) ('tumor', 'Disease', 'MESH:D009369', (46, 51)) 7082 30645099 It was synthesized from 15 (0.33 g, 1.6 mmol), NBS (0.3 g, 1.8 mmol) in dry DMF according to a general procedure to give 0.47 g of 3 in 93% yield, mp 216-218 C. 1H NMR (DMSO-d6) delta 8.57 (s, 1H), 8.50 (d, J = 9 Hz, 1H), 8.20 (d, J = 9 Hz, 1H), 8.03 (m, 2H), 7.90 (d, J = 3.9 Hz, 1H), 7.36 (d, J = 3.9 Hz, 1H). ('2H', 'Chemical', 'MESH:D006859', (256, 258)) ('NBS', 'Disease', 'MESH:D049932', (47, 50)) ('DMF', 'Chemical', 'MESH:C503868', (76, 79)) ('NBS', 'Disease', (47, 50)) ('DMSO-d6', 'Chemical', 'MESH:D004121', (170, 177)) ('mp 216-218 C.', 'Var', (147, 161)) 7086 30645099 It was synthesized from 4 (95 mg, 0.27 mmol), LiN(TMS)2 (1M, 1.6 mL, 1.6 mmol) in dry THF according to the procedure described above to give after work up 79 mg (79%) of DB1504, mp > 300 C. 1H NMR (DMSO-d6) delta 9.46 (s, 2 H, exchangeable with D2O) 9.50 (s, 2 H, exchangeable with D2O), 9.42 (s, 2 H, exchangeable with D2O), 9.29 (s, 2 H, exchangeable with D2O), 8.58 (m, 2H), 8.34 (d, J = 8.4 Hz, 1H), 8.20 (d, J = 3.6 Hz, 1H), 8.10 (m, 4H), 7.94 (m, 3H). ('2H', 'Chemical', 'MESH:D006859', (374, 376)) ('DMSO-d6', 'Chemical', 'MESH:D004121', (199, 206)) ('THF', 'Gene', (86, 89)) ('D2O', 'Chemical', 'MESH:D013481', (359, 362)) ('D2O', 'Chemical', 'MESH:D013481', (246, 249)) ('LiN(TMS)2', 'Disease', 'MESH:C565664', (46, 55)) ('D2O', 'Chemical', 'MESH:D013481', (321, 324)) ('mp >', 'Var', (178, 182)) ('D2O', 'Chemical', 'MESH:D013481', (283, 286)) ('4H', 'Chemical', 'MESH:D006859', (440, 442)) ('THF', 'Gene', '21831', (86, 89)) 7092 30645099 for C28H25NS m/z 505.1923 (M++1), found m/z 505.1876; Anal. ('m/z', 'Var', (40, 43)) ('8H', 'Chemical', 'MESH:D006859', (6, 8)) ('C28H25NS', 'Var', (4, 12)) 7098 30645099 The HOXA9/DNA complex was revealed by addition of 85ng of a mouse anti-TY1-tag antibody (C1520054, Diagenode, Belgium) in TBS-T for 1H at room temperature followed by three washing steps and addition of a 1/2000 dilution of a secondary sheep HRP-conjugated anti-mouse antibody (NXA931, GE Healthcare, France). ('C1520054', 'Var', (89, 97)) ('C1520054', 'Chemical', 'MESH:D002244', (89, 97)) ('TBS-T', 'Disease', 'MESH:C536974', (122, 127)) ('mouse', 'Species', '10090', (262, 267)) ('sheep', 'Species', '9940', (236, 241)) ('TBS-T', 'Disease', (122, 127)) ('mouse', 'Species', '10090', (60, 65)) 7114 30645099 One-site binding models have been applied for DB818 with HP-HBS and HP-HBS-ATm DNA and also for DB1055 with HP-HBS-ATm DNA. ('ATm', 'Gene', '11920', (75, 78)) ('HP-HBS', 'Chemical', 'MESH:C528383', (108, 114)) ('ATm', 'Gene', (115, 118)) ('ATm', 'Gene', (75, 78)) ('HP-HBS', 'Chemical', 'MESH:C528383', (57, 63)) ('DB1055', 'Var', (96, 102)) ('HP-HBS', 'Chemical', 'MESH:C528383', (68, 74)) ('ATm', 'Gene', '11920', (115, 118)) ('DB818', 'Var', (46, 51)) 7133 30645099 MigA9 cells were treated in presence and absence of 5 and 10 muM of DB818, DB1055 and DB828 at n=6 per condition for 48H treatments prior to the purification of total RNA. ('DB818', 'Var', (68, 73)) ('DB1055', 'Var', (75, 81)) ('8H', 'Chemical', 'MESH:D006859', (118, 120)) ('DB828', 'Var', (86, 91)) 7135 30645099 For the MTS assay, MigA9 cells were seeded into 96-well plates at a density of 1.5x104/well in 100muL of complete medium and treated with DB818, DB1055 or DB828 compounds at 0.1 to 50muM or the corresponding control vehicle for 72H before analysis using CellTiter 96 AQueous One Solution Cell Proliferation Assay kit following the manufacturer's instruction. ('DB1055', 'Var', (145, 151)) ('2H', 'Chemical', 'MESH:D006859', (229, 231)) ('DB828', 'Var', (155, 160)) ('DB818', 'Var', (138, 143)) 7175 30057994 Apart from the known genetic and epigenetic mechanisms involved in ineffective hematopoiesis and clonal evolution to AML, immune mechanisms with bone marrow microenvironment inflammatory dysregulation and aberrant signaling of innate immune pathways, might be involved in MDS. ('MDS', 'Disease', (272, 275)) ('MDS', 'Disease', 'MESH:D009190', (272, 275)) ('MDS', 'Phenotype', 'HP:0002863', (272, 275)) ('AML', 'Disease', (117, 120)) ('hematopoiesis', 'Disease', (79, 92)) ('AML', 'Phenotype', 'HP:0004808', (117, 120)) ('ineffective hematopoiesis', 'Phenotype', 'HP:0010972', (67, 92)) ('hematopoiesis', 'Disease', 'MESH:C536227', (79, 92)) ('aberrant', 'Var', (205, 213)) ('AML', 'Disease', 'MESH:D015470', (117, 120)) ('signaling', 'MPA', (214, 223)) ('innate immune pathways', 'Pathway', (227, 249)) 7193 28902850 We investigated the association between copy number variation (CNV) in glutathione S-transferases (GST) and cytochromes (CYP) and the response rate to TKI. ('GST', 'Gene', '373156', (99, 102)) ('copy number variation', 'Var', (40, 61)) ('CYP', 'Gene', '9360', (121, 124)) ('glutathione S-transferases', 'Gene', (71, 97)) ('GST', 'Gene', (99, 102)) ('glutathione S-transferases', 'Gene', '373156', (71, 97)) ('CYP', 'Gene', (121, 124)) 7196 28902850 Patients with optimal response and with failure of TKI therapy showed different frequencies of wild type and mutated CYPs and GST (p<0.0013). ('GST', 'Gene', '373156', (126, 129)) ('mutated', 'Var', (109, 116)) ('Patients', 'Species', '9606', (0, 8)) ('CYP', 'Gene', '9360', (117, 120)) ('CYP', 'Gene', (117, 120)) ('GST', 'Gene', (126, 129)) 7204 28902850 Studies of metabolizing enzymes reveal some valuable associations between polymorphisms in glutathione S-transferases (GST) and cytochromes (CYP) and the response rate to TKI therapy. ('polymorphisms', 'Var', (74, 87)) ('GST', 'Gene', '373156', (119, 122)) ('glutathione S-transferases', 'Gene', (91, 117)) ('CYP', 'Gene', '9360', (141, 144)) ('CYP', 'Gene', (141, 144)) ('GST', 'Gene', (119, 122)) ('glutathione S-transferases', 'Gene', '373156', (91, 117)) 7207 28902850 CNV analysis of these genes may become a valuable prediction marker of TKI efficiency in CML patients. ('CML', 'Disease', (89, 92)) ('CML', 'Disease', 'MESH:D015464', (89, 92)) ('patients', 'Species', '9606', (93, 101)) ('CNV', 'Var', (0, 3)) 7223 28902850 CNV detection was performed using SALSA MLPA P128-C1 Cytochrome P450 probe mix (#P128-100R, MRC-Holland, Amsterdam, Netherlands) following the manufacturer's protocol. ('MRC', 'CellLine', 'CVCL:0440', (92, 95)) ('Cytochrome P450', 'Gene', (53, 68)) ('Cytochrome P450', 'Gene', '4051', (53, 68)) ('#P128-100R', 'Var', (80, 90)) 7227 28902850 We mentioned earlier that CNV in GSTM1 are frequent in all groups and may be non-specific for prognosing therapy outcome. ('GSTM1', 'Gene', '2944', (33, 38)) ('GSTM1', 'Gene', (33, 38)) ('CNV', 'Var', (26, 29)) 7240 28902850 However, we found CNV mutations in other CYP among optimal responders: CYP2A6, CYP2D6 and CYP2E1. ('CYP', 'Gene', '9360', (71, 74)) ('CYP', 'Gene', (71, 74)) ('CYP2A6', 'Gene', '1548', (71, 77)) ('CYP2A6', 'Gene', (71, 77)) ('CYP2E1', 'Gene', (90, 96)) ('CYP', 'Gene', '9360', (41, 44)) ('CYP', 'Gene', (41, 44)) ('CYP2D6', 'Gene', '1565', (79, 85)) ('CYP', 'Gene', '9360', (79, 82)) ('CYP', 'Gene', (79, 82)) ('CYP2D6', 'Gene', (79, 85)) ('mutations', 'Var', (22, 31)) ('CYP', 'Gene', '9360', (90, 93)) ('CYP', 'Gene', (90, 93)) ('CYP2E1', 'Gene', '1571', (90, 96)) 7245 28902850 B. Mitchell and M. Deininger pointed out SNPs in CYP3A5 among other molecular markers as possible predictors of primary TKI resistance. ('CYP3A5', 'Gene', (49, 55)) ('CYP3A5', 'Gene', '1577', (49, 55)) ('SNPs', 'Var', (41, 45)) ('primary TKI resistance', 'Disease', (112, 134)) 7248 28902850 The main difference between our groups of patients is contributed by the CNVs in GSTT1 (S3 Table). ('GSTT1', 'Gene', (81, 86)) ('CNVs', 'Var', (73, 77)) ('patients', 'Species', '9606', (42, 50)) ('GSTT1', 'Gene', '2952', (81, 86)) 7249 28902850 It was previously shown that polymorphisms of the GSTs correlate with a risk of acute myeloid leukemia (AML) in accordance with the meta-analysis of case-control studies published between 1998 and 2009. ('acute myeloid leukemia', 'Disease', (80, 102)) ('AML', 'Disease', (104, 107)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (80, 102)) ('GSTs', 'Gene', (50, 54)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (86, 102)) ('AML', 'Disease', 'MESH:D015470', (104, 107)) ('leukemia', 'Phenotype', 'HP:0001909', (94, 102)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (80, 102)) ('polymorphisms', 'Var', (29, 42)) ('GSTs', 'Gene', '373156', (50, 54)) 7250 28902850 Specifically, deletions of GSTM1 and GSTT1 were associated with poor prognosis of AML and worse chemotherapy efficacy. ('GSTT1', 'Gene', '2952', (37, 42)) ('GSTT1', 'Gene', (37, 42)) ('associated', 'Reg', (48, 58)) ('AML', 'Disease', 'MESH:D015470', (82, 85)) ('deletions', 'Var', (14, 23)) ('AML', 'Disease', (82, 85)) ('GSTM1', 'Gene', '2944', (27, 32)) ('GSTM1', 'Gene', (27, 32)) 7253 28902850 The pooled OR of acute leukemia risks associated with GSTM1 null genotype, GSTP1 Val105 allele and GSTT1 null genotype, were 1.22 (95% CI 1.07-1.38), 1.07 (95% CI 1.00-1.13) and 1.19 (95% CI 1.00-1.41), respectively. ('acute leukemia', 'Disease', 'MESH:D015470', (17, 31)) ('acute leukemia', 'Disease', (17, 31)) ('acute leukemia', 'Phenotype', 'HP:0002488', (17, 31)) ('null genotype', 'Var', (60, 73)) ('GSTT1', 'Gene', (99, 104)) ('GSTM1', 'Gene', '2944', (54, 59)) ('GSTP1', 'Gene', '2950', (75, 80)) ('GSTM1', 'Gene', (54, 59)) ('GSTT1', 'Gene', '2952', (99, 104)) ('null', 'Var', (105, 109)) ('Val105 allele', 'Var', (81, 94)) ('leukemia', 'Phenotype', 'HP:0001909', (23, 31)) ('GSTP1', 'Gene', (75, 80)) 7254 28902850 A significant rise in acute lymphoblastic leukemia risk was observed in patients with GSTM1 and GSTT1 null genotypes. ('rise', 'PosReg', (14, 18)) ('acute lymphoblastic leukemia', 'Disease', 'MESH:D054198', (22, 50)) ('null genotypes', 'Var', (102, 116)) ('acute lymphoblastic leukemia', 'Phenotype', 'HP:0006721', (22, 50)) ('patients', 'Species', '9606', (72, 80)) ('leukemia', 'Phenotype', 'HP:0001909', (42, 50)) ('GSTT1', 'Gene', '2952', (96, 101)) ('GSTT1', 'Gene', (96, 101)) ('lymphoblastic leukemia', 'Phenotype', 'HP:0005526', (28, 50)) ('GSTM1', 'Gene', '2944', (86, 91)) ('acute lymphoblastic leukemia', 'Disease', (22, 50)) ('GSTM1', 'Gene', (86, 91)) 7257 28902850 In addition, the presence of double-null genotypes increased the risk of AML in both Caucasians and East Asians. ('double-null', 'Var', (29, 40)) ('AML', 'Disease', 'MESH:D015470', (73, 76)) ('AML', 'Disease', (73, 76)) 7260 28902850 Later it was observed that deletions of GSTT1 associate with imatinib failure, while CNV in GSTM1 or GSTP1 have no influence on risk of failure. ('GSTP1', 'Gene', '2950', (101, 106)) ('GSTT1', 'Gene', '2952', (40, 45)) ('GSTT1', 'Gene', (40, 45)) ('associate', 'Reg', (46, 55)) ('GSTM1', 'Gene', '2944', (92, 97)) ('GSTM1', 'Gene', (92, 97)) ('imatinib', 'Chemical', 'MESH:D000068877', (61, 69)) ('GSTP1', 'Gene', (101, 106)) ('deletions', 'Var', (27, 36)) ('imatinib failure', 'MPA', (61, 77)) 7261 28902850 The authors of that study estimated failure of treatment during at least 36 months and moreover analyzed patients with secondary failure including confirmed mutations of BCR/ABL. ('secondary failure', 'Disease', (119, 136)) ('patients', 'Species', '9606', (105, 113)) ('mutations', 'Var', (157, 166)) ('BCR/ABL', 'Gene', '25;613', (170, 177)) ('BCR/ABL', 'Gene', (170, 177)) ('secondary failure', 'Disease', 'MESH:D060085', (119, 136)) 7281 28039479 Our studies show that such treatments are most effective in M4/5 subtypes of AML. ('M4/5', 'Var', (60, 64)) ('AML', 'Disease', 'MESH:D015470', (77, 80)) ('AML', 'Phenotype', 'HP:0004808', (77, 80)) ('AML', 'Disease', (77, 80)) 7292 28039479 Also, inactivation of NF-kappaB signaling selectively eradicates LSCs in vitro especially when combined with chemotherapeutic drugs without significant influence on the survival and growth of normal HSPCs. ('inactivation', 'Var', (6, 18)) ('SP', 'Chemical', 'MESH:C000604007', (200, 202)) ('eradicates', 'NegReg', (54, 64)) ('NF-kappaB', 'Protein', (22, 31)) ('LSCs', 'Disease', (65, 69)) 7295 28039479 We determined that inhibition of TNF-JNK signaling provided improved treatment for TNF-expressing AML when combined with NF-kappaB inhibitors. ('TNF-JNK', 'MPA', (33, 40)) ('inhibition', 'Var', (19, 29)) ('AML', 'Disease', 'MESH:D015470', (98, 101)) ('AML', 'Phenotype', 'HP:0004808', (98, 101)) ('improved', 'PosReg', (60, 68)) ('AML', 'Disease', (98, 101)) ('TNF-expressing', 'Gene', (83, 97)) ('treatment', 'MPA', (69, 78)) 7299 28039479 Our study suggests that inhibition of both TNF and IL1beta signaling could represent an improved treatment for inflammatory cytokine-secreting AML when combined with an NF-kappaB inhibitor. ('inflammatory cytokine-secreting', 'Disease', (111, 142)) ('AML', 'Disease', 'MESH:D015470', (143, 146)) ('inhibition', 'Var', (24, 34)) ('AML', 'Disease', (143, 146)) ('AML', 'Phenotype', 'HP:0004808', (143, 146)) 7301 28039479 Consistent with this observation, we found that combined treatment with both the NF-kappaB inhibitor BAY11-7802 (BAY hereafter) and the JNK inhibitor SP600125 (SP hereafter) profoundly reduced the tumor burden and prolonged the survival of leukemic mice developed from MA9 transduction (Figure 1A-1C). ('BAY11-7802', 'Chemical', '-', (101, 111)) ('survival', 'CPA', (228, 236)) ('prolonged', 'PosReg', (214, 223)) ('leukemic', 'Disease', 'MESH:D007938', (240, 248)) ('tumor', 'Disease', 'MESH:D009369', (197, 202)) ('MA9', 'Gene', '4297', (269, 272)) ('MA9', 'Gene', (269, 272)) ('SP', 'Chemical', 'MESH:C000604007', (160, 162)) ('leukemic', 'Disease', (240, 248)) ('NF-kappaB', 'Protein', (81, 90)) ('SP600125', 'Var', (150, 158)) ('SP600125', 'Chemical', 'MESH:C432165', (150, 158)) ('SP', 'Chemical', 'MESH:C000604007', (150, 152)) ('mice', 'Species', '10090', (249, 253)) ('tumor', 'Phenotype', 'HP:0002664', (197, 202)) ('tumor', 'Disease', (197, 202)) ('BAY11-7802', 'Var', (101, 111)) ('reduced', 'NegReg', (185, 192)) 7328 28039479 Although IL1beta also promotes the clone-forming capacity of HSPCs (Figure 4D), we found that it compromises the hematopoietic reconstitutive ability of hematopoietic stem cells (HSCs) as demonstrated by ex vivo treatment and transplantation studies (Figure 4E). ('compromises', 'NegReg', (97, 108)) ('promotes', 'PosReg', (22, 30)) ('clone-forming capacity of HSPCs', 'CPA', (35, 66)) ('IL1beta', 'Var', (9, 16)) ('hematopoietic reconstitutive ability of', 'CPA', (113, 152)) ('SP', 'Chemical', 'MESH:C000604007', (62, 64)) 7332 28039479 We found that genetic inhibition of IL1R also sensitized LCs to anti-TNF treatment (Figure 4I). ('IL1R', 'Gene', (36, 40)) ('LCs', 'Disease', (57, 60)) ('genetic inhibition', 'Var', (14, 32)) ('IL1R', 'Gene', '3554', (36, 40)) ('sensitized', 'Reg', (46, 56)) 7333 28039479 LCs with IL1R knockdown generated far fewer CFUs in the presence of anti-TNF when compared to Scr-shRNA-transduced LCs. ('IL1R', 'Gene', (9, 13)) ('CFUs', 'CPA', (44, 48)) ('IL1R', 'Gene', '3554', (9, 13)) ('knockdown', 'Var', (14, 23)) ('fewer', 'NegReg', (38, 43)) 7344 28039479 As a consequence, we found that disease onset in mice that had received either IL1RA, anti-TNF, or BAY single-treated LCs was significantly delayed compared to mice receiving vehicle-treated LCs, and was even further delayed in mice receiving LCs treated with any of these two chemical combinations. ('IL1RA', 'Gene', '16181', (79, 84)) ('disease', 'CPA', (32, 39)) ('delayed', 'NegReg', (140, 147)) ('mice', 'Species', '10090', (49, 53)) ('mice', 'Species', '10090', (160, 164)) ('mice', 'Species', '10090', (228, 232)) ('IL1RA', 'Gene', (79, 84)) ('anti-TNF', 'Var', (86, 94)) 7355 28039479 Taken together, our data suggest that inactivation of both IL1beta and TNF signaling is more effective in facilitating NF-kappaB inhibitor-induced repression of leukemia development. ('IL1beta', 'Gene', (59, 66)) ('leukemia', 'Disease', (161, 169)) ('leukemia', 'Phenotype', 'HP:0001909', (161, 169)) ('leukemia', 'Disease', 'MESH:D007938', (161, 169)) ('inactivation', 'Var', (38, 50)) ('repression', 'NegReg', (147, 157)) ('TNF', 'Gene', (71, 74)) ('NF-kappaB', 'Protein', (119, 128)) 7358 28039479 However, in LCs, the expression of MKP5 turns off JNK signaling shortly after its activation. ('MKP5', 'Gene', '63953', (35, 39)) ('turns', 'Reg', (40, 45)) ('JNK signaling', 'MPA', (50, 63)) ('expression', 'Var', (21, 31)) ('MKP5', 'Gene', (35, 39)) 7364 28039479 Using CFU assay, we demonstrated that inactivation of c-Jun and JunB but not JunD repressed the clonogenic capacity of LCs (Figure 7D). ('JunD', 'Gene', (77, 81)) ('c-Jun', 'Gene', '16476', (54, 59)) ('JunB', 'Gene', '16477', (64, 68)) ('inactivation', 'Var', (38, 50)) ('JunB', 'Gene', (64, 68)) ('clonogenic capacity', 'CPA', (96, 115)) ('c-Jun', 'Gene', (54, 59)) ('JunD', 'Gene', '16478', (77, 81)) 7365 28039479 LCs with either c-Jun or JunB knockdown showed increased sensitivity to NB-kappaB inhibitor treatment compared to Scr-shRNA-transduced control (Figure 7E). ('c-Jun', 'Gene', '16476', (16, 21)) ('c-Jun', 'Gene', (16, 21)) ('increased', 'PosReg', (47, 56)) ('knockdown', 'Var', (30, 39)) ('sensitivity', 'MPA', (57, 68)) ('JunB', 'Gene', '16477', (25, 29)) ('JunB', 'Gene', (25, 29)) 7370 28039479 IL1beta induces TNF-independent activation of JNK signaling through the RIP1-MYD88-IRAK pathway. ('RIP1', 'Gene', (72, 76)) ('activation', 'PosReg', (32, 42)) ('JNK signaling', 'MPA', (46, 59)) ('MYD88', 'Gene', '17874', (77, 82)) ('IRAK', 'Gene', (83, 87)) ('MYD88', 'Gene', (77, 82)) ('IRAK', 'Gene', '16179', (83, 87)) ('RIP1', 'Gene', '19766', (72, 76)) ('IL1beta', 'Var', (0, 7)) 7374 28039479 To determine whether inhibition of both TNF and IL1 can also promote the anti-leukemic ability of NF-kappaB inhibition in primary human AML, we treated primary human LCs freshly isolated from peripheral blood of AML patients with BAY and TNF/IL-1 antagonists (Enbrel and Kinaret , respectively) individually or in combination in methylcellulose for CFU assay. ('leukemic', 'Disease', (78, 86)) ('promote', 'PosReg', (61, 68)) ('methylcellulose', 'Chemical', 'MESH:D008747', (330, 345)) ('AML', 'Disease', 'MESH:D015470', (212, 215)) ('patients', 'Species', '9606', (216, 224)) ('inhibition', 'Var', (21, 31)) ('human', 'Species', '9606', (130, 135)) ('AML', 'Disease', 'MESH:D015470', (136, 139)) ('leukemic', 'Disease', 'MESH:D007938', (78, 86)) ('AML', 'Phenotype', 'HP:0004808', (136, 139)) ('AML', 'Phenotype', 'HP:0004808', (212, 215)) ('AML', 'Disease', (136, 139)) ('human', 'Species', '9606', (160, 165)) ('AML', 'Disease', (212, 215)) 7381 28039479 Thus inactivation of one cytokine-stimulated signal in most cases is insufficient to repress tumor growth due to the compensation of signaling stimulated by other cytokines. ('insufficient to repress tumor', 'Disease', 'MESH:D000309', (69, 98)) ('inactivation', 'Var', (5, 17)) ('tumor', 'Phenotype', 'HP:0002664', (93, 98)) ('insufficient to repress tumor', 'Disease', (69, 98)) 7391 28039479 However, despite of the effective repressive role of IL1 antagonists (such as sIL-R or IL1RA) on the growth of LCs in in vitro culture, inhibition of IL1 signaling might not be able to successfully repress AML in vivo due to compensation by TNF-stimulated signaling. ('AML', 'Phenotype', 'HP:0004808', (206, 209)) ('IL1RA', 'Gene', (87, 92)) ('inhibition', 'Var', (136, 146)) ('IL1RA', 'Gene', '16181', (87, 92)) ('AML', 'Disease', 'MESH:D015470', (206, 209)) ('repress', 'NegReg', (198, 205)) ('AML', 'Disease', (206, 209)) 7396 28039479 MLL rearrangements are commonly detected in such AML samples. ('AML', 'Disease', (49, 52)) ('rearrangements', 'Var', (4, 18)) ('MLL', 'Gene', (0, 3)) ('AML', 'Disease', 'MESH:D015470', (49, 52)) ('MLL', 'Gene', '214162', (0, 3)) ('AML', 'Phenotype', 'HP:0004808', (49, 52)) 7402 28039479 We found that in these TNF/IL1-expressing AML cells, as is the case with inactivation of JNK signaling, inhibition of both IL1 and TNF represses growth and significantly sensitizes clonogenic LPs and leukemogenic LSCs to NF-kappaB inhibitor treatment. ('LPs', 'Chemical', '-', (192, 195)) ('AML', 'Phenotype', 'HP:0004808', (42, 45)) ('AML', 'Disease', (42, 45)) ('IL1', 'Gene', (123, 126)) ('sensitizes', 'Reg', (170, 180)) ('growth', 'CPA', (145, 151)) ('TNF', 'Gene', (131, 134)) ('leukemogenic', 'Disease', (200, 212)) ('AML', 'Disease', 'MESH:D015470', (42, 45)) ('inhibition', 'Var', (104, 114)) ('clonogenic LPs', 'CPA', (181, 195)) 7423 28039479 Ten thousand LCs (CD45.2+) were plated in each well in a suspension culture and treated with indicated doses of BAY11-7085, SP600125, anti-TNF antibody (Amgen), and IL-1Ra (Anakinra, Amgen) in indicated combinations for 12 hours. ('SP600125', 'Var', (124, 132)) ('CD45', 'Gene', (18, 22)) ('IL-1Ra', 'Gene', (165, 171)) ('BAY11-7085', 'Chemical', 'MESH:C416282', (112, 122)) ('BAY11-7085', 'Var', (112, 122)) ('anti-TNF antibody', 'Protein', (134, 151)) ('IL-1Ra', 'Gene', '16181', (165, 171)) ('SP600125', 'Chemical', 'MESH:C432165', (124, 132)) ('CD45', 'Gene', '19264', (18, 22)) 7429 28039479 Twenty days after transplantation, mice were treated with 10mg/kg InVivoMAb anti m-TNFalpha (BioXcell), 10 mg/kg BAY11-7085, 30 mg/kg SP600125, or IL-1RA (Anakinra, Amgen) individually or in combinations every day for 10 days. ('TNFalpha', 'Gene', (83, 91)) ('IL-1RA', 'Gene', (147, 153)) ('BAY11-7085', 'Chemical', 'MESH:C416282', (113, 123)) ('BAY11-7085', 'Var', (113, 123)) ('SP600125', 'Chemical', 'MESH:C432165', (134, 142)) ('IL-1RA', 'Gene', '16181', (147, 153)) ('mice', 'Species', '10090', (35, 39)) ('SP600125', 'Var', (134, 142)) ('TNFalpha', 'Gene', '21926', (83, 91)) 7432 28039479 LCs were transduced with retrovirus-expressing shRNAs (Origene) specifically targeted to c-Jun (TG501139), JunB (TG516091), JunD (TG501140), and IL-1R (TG501076). ('JunB', 'Gene', (107, 111)) ('c-Jun', 'Gene', '16476', (89, 94)) ('IL-1R', 'Gene', '3554', (145, 150)) ('TG501140', 'Var', (130, 138)) ('JunD', 'Gene', '16478', (124, 128)) ('IL-1R', 'Gene', (145, 150)) ('TG501076', 'Var', (152, 160)) ('TG516091', 'Var', (113, 121)) ('JunD', 'Gene', (124, 128)) ('TG501139', 'Var', (96, 104)) ('c-Jun', 'Gene', (89, 94)) ('targeted', 'Reg', (77, 85)) ('JunB', 'Gene', '16477', (107, 111)) 7437 28039479 A portion of MNC was used for RNA extraction and TNF expression analysis; another fraction of MNC was plated in StemSpan serum-free medium (StemCell) supplemented with recombinant human SCF (100 ng/mL), Flt-3L (100 ng/mL), TPO (20 ng/mL), IL-6 (20 ng/mL), and IL-3 (20 ng/mL). ('IL-3', 'Gene', '3562', (260, 264)) ('TPO', 'Gene', '7173', (223, 226)) ('Flt-3L', 'Gene', (203, 209)) ('human', 'Species', '9606', (180, 185)) ('StemSpan', 'Chemical', '-', (112, 120)) ('TPO', 'Gene', (223, 226)) ('Flt-3L', 'Gene', '2323', (203, 209)) ('100 ng/mL', 'Var', (211, 220)) ('IL-3', 'Gene', (260, 264)) 7438 28039479 Following overnight culturing, 3x105 cells from each sample were harvested and treated with the indicated doses of BAY11-7085, anti-TNF (Etanercept ), or anti-IL1 (Kinaret ), and plated into methylcellulose (StemCell) for CFU assay. ('BAY11-7085', 'Gene', (115, 125)) ('methylcellulose', 'Chemical', 'MESH:D008747', (191, 206)) ('anti-TNF', 'Var', (127, 135)) ('anti-IL1', 'Var', (154, 162)) ('BAY11-7085', 'Chemical', 'MESH:C416282', (115, 125)) 7464 27771292 Modifications on the C-1 of the ceramide via glyosidic or phosphodiester linkage results in different subclasses of complex sphingolipids: sphingomyelin (phosphocholine), and the glycosphingolipids (sugar). ('sphingolipids', 'Chemical', 'MESH:D013107', (124, 137)) ('sugar', 'Chemical', 'MESH:D000073893', (199, 204)) ('results in', 'Reg', (81, 91)) ('subclasses of complex sphingolipids: sphingomyelin', 'MPA', (102, 152)) ('glycosphingolipids', 'Chemical', 'MESH:D006028', (179, 197)) ('ceramide', 'Chemical', 'MESH:D002518', (32, 40)) ('glyosidic', 'Disease', 'None', (45, 54)) ('Modifications', 'Var', (0, 13)) ('glyosidic', 'Disease', (45, 54)) ('phosphocholine', 'Chemical', 'MESH:D010767', (154, 168)) ('sphingolipids', 'Chemical', 'MESH:D013107', (184, 197)) ('sphingomyelin', 'Chemical', 'MESH:D013109', (139, 152)) ('glycosphingolipids', 'MPA', (179, 197)) 7471 27771292 At that point modifications on the C-1 can generate the three subclasses mentioned earlier as well as phosphorylation that generates ceramide-1-phosphate. ('modifications', 'Var', (14, 27)) ('phosphorylation', 'MPA', (102, 117)) ('ceramide-1-phosphate', 'Chemical', 'MESH:C065576', (133, 153)) ('rat', 'Species', '10116', (47, 50)) ('rat', 'Species', '10116', (127, 130)) ('C-1', 'Gene', (35, 38)) 7480 27771292 In the salvage pathway, sphingolipids are initially hydrolyzed mostly to sphingosine in the lysosome, which can then reenter sphingolipid metabolism via either phosphorylation by the sphingosine kinases and/or acylation by one of the six ceramide synthases. ('phosphorylation', 'MPA', (160, 175)) ('sphingosine', 'Enzyme', (183, 194)) ('acylation', 'Var', (210, 219)) ('ceramide', 'Chemical', 'MESH:D002518', (238, 246)) ('sphingolipid metabolism', 'Disease', (125, 148)) ('sphingosine', 'Chemical', 'MESH:D013110', (73, 84)) ('reenter', 'Reg', (117, 124)) ('sphingolipids', 'Chemical', 'MESH:D013107', (24, 37)) ('sphingosine', 'Chemical', 'MESH:D013110', (183, 194)) ('sphingolipid metabolism', 'Disease', 'MESH:D013106', (125, 148)) ('ceramide synthases', 'Enzyme', (238, 256)) 7496 27771292 Mutations of AC in humans are associated with Farber's disease (OMIM #228000). ('associated', 'Reg', (30, 40)) ("Farber's disease", 'Disease', 'MESH:D055577', (46, 62)) ('humans', 'Species', '9606', (19, 25)) ('Mutations', 'Var', (0, 9)) ('AC', 'Gene', '427', (13, 15)) ("Farber's disease", 'Disease', (46, 62)) 7502 27771292 They also observed the accumulation of radiolabeled ceramide in lysosomes and therefore they proposed that Farber's disease is a lysosomal storage disorder (LSD) associated with a biallelic mutation of AC. ("Farber's disease", 'Disease', 'MESH:D055577', (107, 123)) ('lysosomal storage disorder', 'Disease', (129, 155)) ('associated', 'Reg', (162, 172)) ('ceramide', 'Chemical', 'MESH:D002518', (52, 60)) ("Farber's disease", 'Disease', (107, 123)) ('AC', 'Gene', '427', (202, 204)) ('lysosomal storage disorder', 'Disease', 'MESH:D016464', (129, 155)) ('biallelic mutation', 'Var', (180, 198)) ('accumulation', 'MPA', (23, 35)) 7535 27771292 They confirmed this result by introducing arginine residues in this pocket at positions Gly124, Ala211, and Gly465 and determined that those mutations inhibit NC activity. ('Gly465', 'Var', (108, 114)) ('Ala211', 'Chemical', '-', (96, 102)) ('Gly465', 'Chemical', '-', (108, 114)) ('arginine', 'Chemical', 'MESH:D001120', (42, 50)) ('Gly124', 'Var', (88, 94)) ('Gly124', 'Chemical', '-', (88, 94)) ('inhibit', 'NegReg', (151, 158)) ('introducing', 'Reg', (30, 41)) 7542 27771292 Using a model of glycolysis inhibition via 2-deoxyglucose (2DG) and mitochondrial electron transport via antimycin A, Sundaram et al showed that loss of NC protected cells from nutrient deprivation-induced necroptosis via autophagy and clearance of damaged mitochondria. ('2-deoxyglucose', 'Chemical', 'MESH:D003847', (43, 57)) ('antimycin A', 'Chemical', 'MESH:D000968', (105, 116)) ('necroptosis via autophagy', 'Disease', 'MESH:C564093', (206, 231)) ('necroptosis via autophagy', 'Disease', (206, 231)) ('loss', 'Var', (145, 149)) ('clearance', 'CPA', (236, 245)) ('2DG', 'Chemical', 'MESH:D003847', (59, 62)) 7545 27771292 The authors found that silencing the expression of ASAH2 significantly decreased cell growth as well as neuronal differentiation phenotypes. ('ASAH2', 'Gene', '56624', (51, 56)) ('cell growth', 'CPA', (81, 92)) ('silencing', 'Var', (23, 32)) ('ASAH2', 'Gene', (51, 56)) ('decreased', 'NegReg', (71, 80)) ('neuronal differentiation phenotypes', 'CPA', (104, 139)) 7546 27771292 demonstrated, using a DSS inflammation model, that knock out of NC resulted in paradoxical elevation of sphingosine, possibly due to induction of other CDases. ('elevation', 'PosReg', (91, 100)) ('sphingosine', 'MPA', (104, 115)) ('sphingosine', 'Chemical', 'MESH:D013110', (104, 115)) ('inflammation', 'Disease', 'MESH:D007249', (26, 38)) ('rat', 'Species', '10116', (7, 10)) ('inflammation', 'Disease', (26, 38)) ('knock out', 'Var', (51, 60)) 7552 27771292 The role of sphingolipids in inflammation has been also studied by Huang et al., they demonstrated that TGF-beta increased the expression of SphK1 and S1P lyase in human lung fibroblasts and that knockdown of SphK1 or its inhibition attenuated TGF-beta mediated signal transduction and the pathophysiology of lung fibrosis. ('attenuated', 'NegReg', (233, 243)) ('TGF-beta', 'Gene', (244, 252)) ('sphingolipids', 'Chemical', 'MESH:D013107', (12, 25)) ('SphK1', 'Gene', '8877', (209, 214)) ('inflammation', 'Disease', 'MESH:D007249', (29, 41)) ('knockdown', 'Var', (196, 205)) ('lung fibrosis', 'Disease', (309, 322)) ('S1P', 'Gene', '13609', (151, 154)) ('SphK1', 'Gene', (209, 214)) ('lung fibrosis', 'Phenotype', 'HP:0002206', (309, 322)) ('TGF-beta', 'Gene', '7039', (104, 112)) ('inflammation', 'Disease', (29, 41)) ('rat', 'Species', '10116', (93, 96)) ('TGF-beta', 'Gene', (104, 112)) ('expression', 'MPA', (127, 137)) ('SphK1', 'Gene', '8877', (141, 146)) ('increased', 'PosReg', (113, 122)) ('S1P', 'Gene', (151, 154)) ('human', 'Species', '9606', (164, 169)) ('TGF-beta', 'Gene', '7039', (244, 252)) ('SphK1', 'Gene', (141, 146)) ('lung fibrosis', 'Disease', 'MESH:D005355', (309, 322)) 7556 27771292 This study also demonstrated that inhibition of NC in a xenograft model delayed tumor growth and increased ceramide. ('tumor', 'Disease', 'MESH:D009369', (80, 85)) ('increased', 'PosReg', (97, 106)) ('tumor', 'Phenotype', 'HP:0002664', (80, 85)) ('inhibition', 'Var', (34, 44)) ('rat', 'Species', '10116', (23, 26)) ('delayed', 'NegReg', (72, 79)) ('ceramide', 'Chemical', 'MESH:D002518', (107, 115)) ('tumor', 'Disease', (80, 85)) ('ceramide', 'MPA', (107, 115)) 7589 27771292 They also reported that ACER3 knockdown inhibited cell proliferation and upregulated the cyclin-dependent kinase inhibitor p21(CIP1/WAF1). ('CIP1', 'Gene', '1026', (127, 131)) ('ACER3', 'Gene', (24, 29)) ('inhibited', 'NegReg', (40, 49)) ('WAF1', 'Gene', '1026', (132, 136)) ('p21', 'Gene', '1026', (123, 126)) ('knockdown', 'Var', (30, 39)) ('upregulated', 'PosReg', (73, 84)) ('rat', 'Species', '10116', (62, 65)) ('WAF1', 'Gene', (132, 136)) ('CIP1', 'Gene', (127, 131)) ('p21', 'Gene', (123, 126)) ('cell proliferation', 'CPA', (50, 68)) 7594 27771292 ACER3 deficient mice demonstrated impairment in motor coordination and balance capabilities. ('rat', 'Species', '10116', (28, 31)) ('deficient', 'Var', (6, 15)) ('balance capabilities', 'CPA', (71, 91)) ('motor coordination', 'CPA', (48, 66)) ('ACER3', 'Gene', (0, 5)) ('mice', 'Species', '10090', (16, 20)) ('impairment', 'NegReg', (34, 44)) 7596 27771292 suggested that homozygous mutation in ACER3 in patients was responsible for early childhood leukodystrophy. ('ACER3', 'Gene', (38, 43)) ('patients', 'Species', '9606', (47, 55)) ('leukodystrophy', 'Phenotype', 'HP:0002415', (92, 106)) ('responsible', 'Reg', (60, 71)) ('homozygous mutation', 'Var', (15, 34)) ('leukodystrophy', 'Disease', 'MESH:D007966', (92, 106)) ('leukodystrophy', 'Disease', (92, 106)) 7598 27771292 demonstrated that loss of the mouse Acer3 aggravates colitis and colitis-associated colorectal cancer in a murine model likely by disrupting the homeostasis of ceramides and other sphingolipids in the digestive system. ('colitis', 'Phenotype', 'HP:0002583', (53, 60)) ('colitis-associated colorectal cancer', 'Disease', (65, 101)) ('colitis', 'Disease', (65, 72)) ('rat', 'Species', '10116', (7, 10)) ('colorectal cancer', 'Phenotype', 'HP:0003003', (84, 101)) ('Acer3', 'Gene', (36, 41)) ('colitis', 'Disease', 'MESH:D003092', (65, 72)) ('ceramides', 'Chemical', 'MESH:D002518', (160, 169)) ('colitis-associated colorectal cancer', 'Disease', 'MESH:D015179', (65, 101)) ('colitis', 'Disease', (53, 60)) ('cancer', 'Phenotype', 'HP:0002664', (95, 101)) ('murine', 'Species', '10090', (107, 113)) ('mouse', 'Species', '10090', (30, 35)) ('Acer3', 'Gene', '66190', (36, 41)) ('colitis', 'Phenotype', 'HP:0002583', (65, 72)) ('colitis', 'Disease', 'MESH:D003092', (53, 60)) ('homeostasis of ceramides', 'MPA', (145, 169)) ('aggravates', 'PosReg', (42, 52)) ('sphingolipids', 'Chemical', 'MESH:D013107', (180, 193)) ('loss', 'Var', (18, 22)) ('disrupting', 'NegReg', (130, 140)) 7601 27771292 As such, it is not surprising that mutations in certain ceramidase genes or their dysregulation through other mechanisms can lead to different diseases, such as inflammatory diseases, neurodegenerative diseases, or tumorigenesis. ('inflammatory diseases', 'Disease', 'MESH:D007249', (161, 182)) ('neurodegenerative diseases', 'Disease', (184, 210)) ('tumor', 'Disease', 'MESH:D009369', (215, 220)) ('neurodegenerative diseases', 'Phenotype', 'HP:0002180', (184, 210)) ('inflammatory diseases', 'Disease', (161, 182)) ('tumor', 'Phenotype', 'HP:0002664', (215, 220)) ('mutations', 'Var', (35, 44)) ('tumor', 'Disease', (215, 220)) ('lead to', 'Reg', (125, 132)) ('neurodegenerative diseases', 'Disease', 'MESH:D019636', (184, 210)) ('ceramidase genes', 'Gene', (56, 72)) 7624 27613707 If myeloid cells are also particularly susceptible to these factors, this could explain the association between low birthweight and AML. ('low birthweight', 'Var', (112, 127)) ('AML', 'Disease', (132, 135)) ('low birthweight', 'Phenotype', 'HP:0001518', (112, 127)) ('AML', 'Disease', 'MESH:D015470', (132, 135)) ('association', 'Interaction', (92, 103)) 7627 27613707 Whereas pre-pregnancy underweight and insufficient gestational weight gain, which have been linked to restricted fetal growth, will result in an increased risk of childhood cancers that have been associated with lower birthweight. ('restricted fetal growth', 'Phenotype', 'HP:0001558', (102, 125)) ('-pregnancy underweight and insufficient gestational weight', 'Phenotype', 'HP:0001622', (11, 69)) ('underweight', 'Var', (22, 33)) ('insufficient gestational weight gain', 'Disease', (38, 74)) ('cancers', 'Phenotype', 'HP:0002664', (173, 180)) ('childhood cancers', 'Disease', 'MESH:C536928', (163, 180)) ('cancer', 'Phenotype', 'HP:0002664', (173, 179)) ('childhood cancers', 'Disease', (163, 180)) ('insufficient gestational weight', 'Phenotype', 'HP:0001518', (38, 69)) ('lower birthweight', 'Phenotype', 'HP:0001518', (212, 229)) ('weight gain', 'Phenotype', 'HP:0004324', (63, 74)) ('insufficient gestational weight gain', 'Disease', 'MESH:D000078064', (38, 74)) 7636 27613707 We also excluded children that were missing sex (n=3), births that were likely not viable (gestational age <20 weeks and/or birthweight <500g) (n=169), and children diagnosed with Down syndrome (n=151). ('children', 'Species', '9606', (156, 164)) ('Down syndrome', 'Disease', (180, 193)) ('birthweight <500g', 'Phenotype', 'HP:0001518', (124, 141)) ('missing sex', 'Phenotype', 'HP:0008187', (36, 47)) ('<500g', 'Var', (136, 141)) ('children', 'Species', '9606', (17, 25)) 7648 27613707 Pre-pregnancy BMI was categorized according to the World Health Organization criteria: <18.5 kg/m2 (underweight), 18.5-24.9 kg/m2 (normal), 25-29.9 kg/m2 (overweight), >=30 kg/m2 (obese). ('obese', 'Disease', (180, 185)) ('BMI', 'Disease', (14, 17)) ('<18.5 kg/m2', 'Var', (87, 98)) ('25-29.9 kg/m2', 'Var', (140, 153)) ('obese', 'Disease', 'MESH:D009765', (180, 185)) ('overweight', 'Phenotype', 'HP:0025502', (155, 165)) 7698 27613707 In contrast, underweight appeared to increase the risk of germ cell tumors. ('germ cell tumor', 'Phenotype', 'HP:0100728', (58, 73)) ('tumors', 'Phenotype', 'HP:0002664', (68, 74)) ('cell tumors', 'Disease', (63, 74)) ('germ cell tumors', 'Phenotype', 'HP:0100728', (58, 74)) ('germ', 'Disease', (58, 62)) ('underweight', 'Var', (13, 24)) ('cell tumors', 'Disease', 'MESH:D005935', (63, 74)) ('tumor', 'Phenotype', 'HP:0002664', (68, 73)) 7719 27613707 Errors in pre-pregnancy weight seem to be the main source of misclassification of pre-pregnancy BMI and gestational weight gain, which is plausible since pre-pregnancy weight recorded on the birth certificate is typically ascertained by maternal recall at delivery. ('gestational weight gain', 'Disease', (104, 127)) ('weight gain', 'Phenotype', 'HP:0004324', (116, 127)) ('Errors', 'Var', (0, 6)) ('gestational weight gain', 'Disease', 'MESH:D000078064', (104, 127)) ('BMI', 'Disease', (96, 99)) 7784 26798643 Nevertheless, some genetic or chromosomal alterations are found in specific subtypes of leukaemia and have a role in conditioning the severity of the clinical course. ('leukaemia', 'Disease', (88, 97)) ('leukaemia', 'Disease', 'MESH:D007938', (88, 97)) ('chromosomal alterations', 'Var', (30, 53)) 7785 26798643 Rearrangement of the mixed lineage leukaemia (MLL) gene, which occurs in the 11q23 translocation, leads to aggressive acute leukaemia and may be present in both AML and ALL; it occurs in 34% of this subgroup. ('Rearrangement', 'Var', (0, 13)) ('leukaemia', 'Disease', (35, 44)) ('aggressive acute leukaemia', 'Disease', 'MESH:D007938', (107, 133)) ('leukaemia', 'Disease', 'MESH:D007938', (124, 133)) ('leukaemia', 'Disease', (124, 133)) ('AML', 'Phenotype', 'HP:0004808', (161, 164)) ('AML', 'Disease', 'MESH:D015470', (161, 164)) ('aggressive acute leukaemia', 'Disease', (107, 133)) ('ALL', 'Phenotype', 'HP:0006721', (169, 172)) ('leukaemia', 'Disease', 'MESH:D007938', (35, 44)) ('MLL', 'Gene', (46, 49)) ('MLL', 'Gene', '4297', (46, 49)) ('leads to', 'Reg', (98, 106)) ('AML', 'Disease', (161, 164)) ('acute leukaemia', 'Phenotype', 'HP:0002488', (118, 133)) 7786 26798643 On the other hand, spontaneous remission can occur in a subset of neonates affected by AML M4 with t(8;16)(p11;p13). ('AML', 'Phenotype', 'HP:0004808', (87, 90)) ('t(8;16)(p11;p13', 'Var', (99, 114)) ('AML', 'Disease', (87, 90)) ('affected', 'Reg', (75, 83)) ('t(8;16)(p11;p13)', 'STRUCTURAL_ABNORMALITY', 'None', (99, 115)) ('AML', 'Disease', 'MESH:D015470', (87, 90)) 7806 26798643 Because TMD appears to be restricted to patients with DS or trisomy 21 (T21) mosaicism, the origin of AML must be related to a cytogenetic abnormality in chromosome 21. ('TMD', 'Disease', 'MESH:D049310', (8, 11)) ('patients', 'Species', '9606', (40, 48)) ('T21', 'Gene', (72, 75)) ('AML', 'Disease', 'MESH:D015470', (102, 105)) ('trisomy 21', 'Var', (60, 70)) ('AML', 'Disease', (102, 105)) ('mosaicism', 'Var', (77, 86)) ('AML', 'Phenotype', 'HP:0004808', (102, 105)) ('TMD', 'Phenotype', 'HP:0005534', (8, 11)) ('TMD', 'Disease', (8, 11)) 7807 26798643 TMD is caused by cooperation between T21 and acquired somatic N-terminal truncating mutations in the key haematopoietic transcription factor GATA1. ('TMD', 'Disease', 'MESH:D049310', (0, 3)) ('GATA1', 'Gene', (141, 146)) ('TMD', 'Phenotype', 'HP:0005534', (0, 3)) ('N-terminal truncating mutations', 'Var', (62, 93)) ('TMD', 'Disease', (0, 3)) ('GATA1', 'Gene', '2623', (141, 146)) 7808 26798643 Analysis of primary human fetal liver haematopoietic cells and of human embryonic stem cells demonstrates that T21 itself substantially alters human fetal haematopoietic development. ('human', 'Species', '9606', (20, 25)) ('human', 'Species', '9606', (143, 148)) ('alters', 'Reg', (136, 142)) ('T21', 'Var', (111, 114)) ('human', 'Species', '9606', (66, 71)) ('human fetal haematopoietic development', 'CPA', (143, 181)) 7824 26798643 BRAFV600E has been detected in more than half of the cases of LCH in adult age. ('BRAFV600E', 'Mutation', 'rs113488022', (0, 9)) ('LCH', 'Disease', (62, 65)) ('BRAFV600E', 'Var', (0, 9)) 7826 26798643 Mutations in MAP2K1, which encodes the dual-specificity kinase MEK1 protein in the MAPK pathway, have been identified in 27.5% of cases with LCH, thus explaining MAPK pathway activation in the absence of the BRAF mutation. ('BRAF', 'Gene', '673', (208, 212)) ('LCH', 'Disease', (141, 144)) ('MAPK', 'Gene', '5594', (83, 87)) ('MAPK', 'Gene', '5594', (162, 166)) ('identified', 'Reg', (107, 117)) ('BRAF', 'Gene', (208, 212)) ('activation', 'PosReg', (175, 185)) ('MAPK', 'Gene', (83, 87)) ('MAP2K1', 'Gene', '5604', (13, 19)) ('MAPK', 'Gene', (162, 166)) ('MEK1', 'Gene', '5604', (63, 67)) ('Mutations', 'Var', (0, 9)) ('MAP2K1', 'Gene', (13, 19)) ('MEK1', 'Gene', (63, 67)) 7827 26798643 MAP2K1 and BRAF mutations were found to be mutually exclusive, as would be expected since MEK1 is directly downstream BRAF within the MAPK pathway. ('BRAF', 'Gene', '673', (118, 122)) ('MAP2K1', 'Gene', (0, 6)) ('MAPK', 'Gene', (134, 138)) ('BRAF', 'Gene', (118, 122)) ('mutations', 'Var', (16, 25)) ('BRAF', 'Gene', (11, 15)) ('BRAF', 'Gene', '673', (11, 15)) ('MAP2K1', 'Gene', '5604', (0, 6)) ('MEK1', 'Gene', '5604', (90, 94)) ('MAPK', 'Gene', '5594', (134, 138)) ('MEK1', 'Gene', (90, 94)) 7828 26798643 Additional cases of other mutations in the MAPK genes have also been reported, including ARAF and ERBB3. ('ARAF', 'Gene', (89, 93)) ('ERBB3', 'Gene', '2065', (98, 103)) ('mutations', 'Var', (26, 35)) ('ARAF', 'Gene', '369', (89, 93)) ('MAPK', 'Gene', '5594', (43, 47)) ('ERBB3', 'Gene', (98, 103)) ('MAPK', 'Gene', (43, 47)) 7841 26798643 The amplification of MYCN and the subsequent overexpression of the protein directly contribute to tumorigenesis. ('contribute', 'Reg', (84, 94)) ('MYCN', 'Gene', (21, 25)) ('MYCN', 'Gene', '4613', (21, 25)) ('tumor', 'Disease', 'MESH:D009369', (98, 103)) ('tumor', 'Phenotype', 'HP:0002664', (98, 103)) ('amplification', 'Var', (4, 17)) ('tumor', 'Disease', (98, 103)) ('overexpression', 'PosReg', (45, 59)) 7843 26798643 Missense mutations of Paired Homeobox 2b (PHOX2B) on chromosome 4p, frequently associated with other neural crest disorders or malignancies (Ondine's and Hirschsprung's disease), were the first germline mutations to be identified in NB predisposition. ("Hirschsprung's disease", 'Phenotype', 'HP:0002251', (154, 176)) ("Hirschsprung's disease", 'Disease', 'MESH:D006627', (154, 176)) ('PHOX2B', 'Gene', (42, 48)) ('associated', 'Reg', (79, 89)) ('malignancies', 'Disease', 'MESH:D009369', (127, 139)) ('NB', 'Phenotype', 'HP:0003006', (233, 235)) ('malignancies', 'Disease', (127, 139)) ('neural crest disorders', 'Disease', (101, 123)) ('PHOX2B', 'Gene', '8929', (42, 48)) ('neural crest disorders', 'Disease', 'MESH:C536408', (101, 123)) ("Hirschsprung's disease", 'Disease', (154, 176)) ('Missense mutations', 'Var', (0, 18)) 7844 26798643 More recently, whole exome and whole genome sequencing analyses have identified loss-of-function mutations/deletions in chromatin modifiers including ATRX, ARID1A, and ARID1B. ('ARID1B', 'Gene', (168, 174)) ('mutations/deletions', 'Var', (97, 116)) ('ATRX', 'Gene', (150, 154)) ('ARID1B', 'Gene', '57492', (168, 174)) ('ATRX', 'Gene', '546', (150, 154)) ('loss-of-function', 'NegReg', (80, 96)) ('ARID1A', 'Gene', '8289', (156, 162)) ('ARID1A', 'Gene', (156, 162)) 7845 26798643 However, ATRX mutations were more common in patients over 5 years of age and no mutations/deletions were identified in the youngest age group (<18 months). ('ATRX', 'Gene', '546', (9, 13)) ('patients', 'Species', '9606', (44, 52)) ('common', 'Reg', (34, 40)) ('ATRX', 'Gene', (9, 13)) ('mutations', 'Var', (14, 23)) 7865 26798643 In RT, genetic mutations are characterized by somatically acquired biallelic inactivating truncating mutations within the tumor cells, associated or not with a predisposing germline mutation. ('biallelic inactivating truncating mutations', 'Var', (67, 110)) ('tumor', 'Disease', (122, 127)) ('tumor', 'Disease', 'MESH:D009369', (122, 127)) ('tumor', 'Phenotype', 'HP:0002664', (122, 127)) 7873 26798643 Moreover, the presence of a mutation of the hsNF5/INI1 gene located in chromosome 22q11 is helpful in establishing the diagnosis. ('INI1', 'Gene', (50, 54)) ('hsNF5', 'Gene', '6598', (44, 49)) ('hsNF5', 'Gene', (44, 49)) ('mutation', 'Var', (28, 36)) ('INI1', 'Gene', '6598', (50, 54)) 7882 26798643 Alveolar RMS is associated with specific translocation, t(2;13)(q37;q14) or its variant t(1;13)(p36;q14). ('t(1;13)(p36;q14', 'Var', (88, 103)) ('t(1;13)(p36;q14)', 'STRUCTURAL_ABNORMALITY', 'None', (88, 104)) ('RMS', 'Phenotype', 'HP:0002859', (9, 12)) ('t(2;13)(q37;q14', 'Var', (56, 71)) ('associated', 'Reg', (16, 26)) ('Alveolar RMS', 'Disease', (0, 12)) ('t(2;13)(q37;q14)', 'STRUCTURAL_ABNORMALITY', 'None', (56, 72)) 7919 23565126 Children with blasts after the second induction (HAM) or still in aplasia 4 weeks after second induction were qualified for allo-HSCT from an human leukocyte antigen (HLA)-identical unrelated donor. ('Children', 'Species', '9606', (0, 8)) ('blasts', 'Var', (14, 20)) ('human', 'Species', '9606', (142, 147)) ('aplasia', 'Disease', 'MESH:C566720', (66, 73)) ('donor', 'Species', '9606', (192, 197)) ('aplasia', 'Disease', (66, 73)) 7972 23359317 The role of tumor suppressor p15Ink4b in the regulation of hematopoietic progenitor cell fate Epigenetic silencing of the tumor suppressor gene p15Ink4b (CDKN2B) is a frequent event in blood disorders like acute myeloid leukemia and myelodysplastic syndromes. ('p15Ink4b', 'Gene', (29, 37)) ('blood disorders', 'Disease', 'MESH:D006402', (185, 200)) ('tumor', 'Disease', (12, 17)) ('acute myeloid leukemia', 'Disease', (206, 228)) ('tumor', 'Disease', (122, 127)) ('leukemia', 'Phenotype', 'HP:0001909', (220, 228)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (212, 228)) ('tumor', 'Disease', 'MESH:D009369', (12, 17)) ('tumor', 'Disease', 'MESH:D009369', (122, 127)) ('p15Ink4b', 'Gene', '12579', (144, 152)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (206, 228)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (206, 228)) ('CDKN2B', 'Gene', (154, 160)) ('tumor', 'Phenotype', 'HP:0002664', (12, 17)) ('p15Ink4b', 'Gene', (144, 152)) ('myelodysplastic syndromes', 'Phenotype', 'HP:0002863', (233, 258)) ('myelodysplastic syndromes', 'Disease', (233, 258)) ('tumor', 'Phenotype', 'HP:0002664', (122, 127)) ('Epigenetic silencing', 'Var', (94, 114)) ('p15Ink4b', 'Gene', '12579', (29, 37)) ('myelodysplastic syndromes', 'Disease', 'MESH:D009190', (233, 258)) ('blood disorders', 'Disease', (185, 200)) 7973 23359317 Our previous study demonstrated that loss of p15Ink4b in mice results in skewing of the differentiation pattern of the common myeloid progenitor towards the myeloid lineage. ('mice', 'Species', '10090', (57, 61)) ('skewing', 'Reg', (73, 80)) ('rat', 'Species', '10116', (26, 29)) ('loss', 'Var', (37, 41)) ('p15Ink4b', 'Var', (45, 53)) 7975 23359317 More importantly, mice lacking p15Ink4b have lower numbers of primitive red cell progenitors and a severely impaired response to 5-fluorouracil- and phenylhydrazine-induced hematopoietic stress. ('phenylhydrazine', 'Chemical', 'MESH:C030299', (149, 164)) ('mice', 'Species', '10090', (18, 22)) ('5-fluorouracil', 'Chemical', 'MESH:D005472', (129, 143)) ('lower', 'NegReg', (45, 50)) ('p15Ink4b', 'Var', (31, 39)) ('impaired', 'NegReg', (108, 116)) 7976 23359317 Introduction of p15Ink4b into multipotential progenitors produced changes at the molecular level, including activation of mitogen-activated protein kinase\extracellular signal-regulated kinase (MEK/ERK) signaling, increase GATA-1, erythropoietin receptor (EpoR) and decrease Pu1, GATA-2 expression. ('ERK', 'Gene', '26413', (198, 201)) ('erythropoietin receptor', 'Gene', '13857', (231, 254)) ('GATA-1', 'Gene', '14460', (223, 229)) ('Pu1', 'Gene', (275, 278)) ('Pu1', 'Gene', '20375', (275, 278)) ('EpoR', 'Gene', (256, 260)) ('p15Ink4b', 'Var', (16, 24)) ('MEK', 'Gene', '17242', (194, 197)) ('increase', 'PosReg', (214, 222)) ('GATA-2', 'Gene', (280, 286)) ('ERK', 'Gene', (198, 201)) ('EpoR', 'Gene', '13857', (256, 260)) ('erythropoietin receptor', 'Gene', (231, 254)) ('MEK', 'Gene', (194, 197)) ('GATA-1', 'Gene', (223, 229)) ('decrease', 'NegReg', (266, 274)) ('expression', 'MPA', (287, 297)) ('activation', 'PosReg', (108, 118)) 7978 23359317 Our results indicate that p15Ink4b functions in hematopoiesis, by maintaining proper lineage commitment of progenitors and assisting in rapid red blood cells replenishment following stress. ('hematopoiesis', 'Disease', (48, 61)) ('assisting', 'PosReg', (123, 132)) ('p15Ink4b', 'Var', (26, 34)) ('hematopoiesis', 'Disease', 'MESH:C536227', (48, 61)) ('rapid red blood cells replenishment', 'MPA', (136, 171)) ('maintaining', 'PosReg', (66, 77)) 7983 23359317 Lack of p15INK4B expression in MDS patients is positively associated with leukemic transformation and poor prognosis. ('MDS', 'Disease', (31, 34)) ('MDS', 'Disease', 'MESH:D009190', (31, 34)) ('associated', 'Reg', (58, 68)) ('Lack', 'NegReg', (0, 4)) ('expression', 'MPA', (17, 27)) ('leukemic transformation', 'Disease', 'MESH:D007938', (74, 97)) ('patients', 'Species', '9606', (35, 43)) ('leukemic transformation', 'Disease', (74, 97)) ('p15INK4B', 'Var', (8, 16)) ('MDS', 'Phenotype', 'HP:0002863', (31, 34)) 7984 23359317 Despite the compelling evidence supporting the important role of p15INK4B in the development and progression of MDS and leukemia, its function in normal blood cell formation is just beginning to emerge. ('leukemia', 'Disease', (120, 128)) ('leukemia', 'Phenotype', 'HP:0001909', (120, 128)) ('leukemia', 'Disease', 'MESH:D007938', (120, 128)) ('MDS', 'Phenotype', 'HP:0002863', (112, 115)) ('MDS', 'Disease', (112, 115)) ('MDS', 'Disease', 'MESH:D009190', (112, 115)) ('p15INK4B', 'Var', (65, 73)) 7986 23359317 Interestingly, loss of p15Ink4b does not alter cell proliferation, self-renewal or apoptosis in blood progenitors, implicating a specific role in cellular differentiation. ('self-renewal', 'CPA', (67, 79)) ('p15Ink4b', 'Gene', (23, 31)) ('loss', 'Var', (15, 19)) ('apoptosis', 'CPA', (83, 92)) ('rat', 'Species', '10116', (59, 62)) ('cell proliferation', 'CPA', (47, 65)) 7987 23359317 Based upon these studies and its implied role in human blood diseases, we hypothesized that p15Ink4b has a role in commitment of blood progenitors to the erythroid lineage. ('human', 'Species', '9606', (49, 54)) ('commitment', 'CPA', (115, 125)) ('p15Ink4b', 'Var', (92, 100)) ('blood diseases', 'Disease', 'MESH:D006402', (55, 69)) ('blood diseases', 'Disease', (55, 69)) 7998 23359317 A mouse myelomonocytic leukemia cell line, M1, was cultured in RPMI medium supplemented with 10% heat-inactivated horse serum, P/S and used as a positive control for p15Ink4b mRNA and protein detection. ('myelomonocytic leukemia', 'Disease', 'MESH:D054429', (8, 31)) ('P', 'Chemical', 'MESH:D010758', (64, 65)) ('p15Ink4b', 'Var', (166, 174)) ('RPMI medium', 'Chemical', '-', (63, 74)) ('leukemia', 'Phenotype', 'HP:0001909', (23, 31)) ('P', 'Chemical', 'MESH:D010758', (127, 128)) ('horse', 'Species', '9796', (114, 119)) ('mouse', 'Species', '10090', (2, 7)) ('S', 'Chemical', 'MESH:D013455', (129, 130)) ('myelomonocytic leukemia', 'Disease', (8, 31)) ('M1', 'CellLine', 'CVCL:J159', (43, 45)) 7999 23359317 The HEK 293T-derived Lenti-X293T cell line (Clontech, Mountain View, CA, USA) was maintained in Dulbecco's modified Eagle's medium with high glucose medium supplemented with 10% heat-inactivated, Tet system-approved, fetal bovine serum (FBS), 4 mML-glutamine, 3.7 g/l sodium bicarbonate and 1 mM sodium pyruvate. ('Tet', 'Chemical', 'MESH:C010349', (196, 199)) ('glucose', 'Chemical', 'MESH:D005947', (141, 148)) ('HEK 293T', 'CellLine', 'CVCL:0063', (4, 12)) ('sodium pyruvate', 'Chemical', '-', (296, 311)) ('sodium bicarbonate', 'Chemical', 'MESH:D017693', (268, 286)) ('bovine', 'Species', '9913', (223, 229)) ('glutamine', 'Chemical', 'MESH:D005973', (249, 258)) ('S', 'Chemical', 'MESH:D013455', (74, 75)) ('X293T', 'SUBSTITUTION', 'None', (27, 32)) ('sodium bicarbonate', 'MPA', (268, 286)) ('X293T', 'Var', (27, 32)) ('S', 'Chemical', 'MESH:D013455', (239, 240)) ('high glucose', 'Phenotype', 'HP:0003074', (136, 148)) 8007 23359317 The following number of cells was plated per 35 mm pretested culture dish (StemCell Technologies): for M3436, 3000 EML cells or 37 500 whole bone marrow cells, or 10 000 lineage-negative (Lin-) cells per dish, or in vitro-differentiated progenitors; and for M3534, 3750 whole bone marrow cells or 1000 Lin- cells per dish, or in vitro-differentiated progenitors. ('M3436', 'Var', (103, 108)) ('M3534', 'Var', (258, 263)) ('S', 'Chemical', 'MESH:D013455', (75, 76)) 8015 23359317 Removal of SH by washing the cells resulted in a quick reduction of p15Ink4b protein to background levels. ('p15Ink4b', 'Var', (68, 76)) ('S', 'Chemical', 'MESH:D013455', (11, 12)) ('reduction', 'NegReg', (55, 64)) 8037 23359317 Expression of p15Ink4b was induced by the addition of SH (5-500 nM) into the culture medium. ('Expression', 'MPA', (0, 10)) ('S', 'Chemical', 'MESH:D013455', (54, 55)) ('induced', 'Reg', (27, 34)) ('p15Ink4b', 'Var', (14, 22)) 8058 23359317 To determine how the loss of p15Ink4b expression might affect the formation of committed erythroid progenitor cells, we employed a methylcellulose-based colony forming assay (MethoCult) that allows the detection of early RBC precursor cells termed BFU-E. ('loss', 'Var', (21, 25)) ('methylcellulose', 'Chemical', 'MESH:D008747', (131, 146)) ('affect', 'Reg', (55, 61)) ('p15Ink4b', 'Var', (29, 37)) 8059 23359317 Bone marrow of mice lacking p15Ink4b gave rise to significantly lower numbers of early BFU-E colonies compared with mice carrying a normal p15Ink4b locus (wild type) (Figure 1d and e). ('mice', 'Species', '10090', (116, 120)) ('lower', 'NegReg', (64, 69)) ('lacking', 'NegReg', (20, 27)) ('early BFU-E colonies', 'CPA', (81, 101)) ('mice', 'Species', '10090', (15, 19)) ('p15Ink4b', 'Var', (28, 36)) 8063 23359317 Quantitative real-time polymerase chain reaction analysis of cDNA derived from these cells determined that MEPs expressed twofold higher levels of p15Ink4b mRNA compared with CMPs, and fourfold higher levels than GMPs (Figure 1g). ('P', 'Chemical', 'MESH:D010758', (109, 110)) ('P', 'Chemical', 'MESH:D010758', (215, 216)) ('GMP', 'Gene', (213, 216)) ('higher', 'PosReg', (130, 136)) ('GMP', 'Gene', '76952', (213, 216)) ('p15Ink4b', 'Var', (147, 155)) ('MEP', 'Gene', (107, 110)) ('P', 'Chemical', 'MESH:D010758', (177, 178)) ('MEP', 'Gene', '13848', (107, 110)) 8066 23359317 Although these two genes function cooperatively in many tissues to inhibit the cell cycle through the binding of cyclin-dependent kinases (Cdks), our findings suggest a novel role for p15Ink4b in MEPs. ('cell cycle', 'CPA', (79, 89)) ('Cdks', 'Gene', (139, 143)) ('inhibit', 'NegReg', (67, 74)) ('binding', 'Interaction', (102, 109)) ('p15Ink4b', 'Var', (184, 192)) ('rat', 'Species', '10116', (39, 42)) ('MEP', 'Gene', (196, 199)) ('Cdks', 'Gene', '12567;12571', (139, 143)) ('MEP', 'Gene', '13848', (196, 199)) 8067 23359317 The association of p15Ink4b expression with erythroid commitment was further supported by the identification of mRNA encoding p15Ink4b in several erythroleukemia cell lines that are blocked at early stages of RBC development (Figure 1h). ('association', 'Interaction', (4, 15)) ('erythroleukemia', 'Disease', 'MESH:D004915', (146, 161)) ('erythroleukemia', 'Disease', (146, 161)) ('p15Ink4b', 'Gene', (19, 27)) ('erythroid', 'CPA', (44, 53)) ('p15Ink4b', 'Var', (126, 134)) ('leukemia', 'Phenotype', 'HP:0001909', (153, 161)) 8071 23359317 For these experiments, knockout and wild-type mice were treated with two different stimuli, both inducing anemia, but acting through different cellular mechanisms: 5-FU and PHZ. ('anemia', 'Phenotype', 'HP:0001903', (106, 112)) ('PHZ', 'Var', (173, 176)) ('5-FU', 'Chemical', 'MESH:D005472', (164, 168)) ('inducing', 'Reg', (97, 105)) ('mice', 'Species', '10090', (46, 50)) ('anemia', 'Disease', (106, 112)) ('anemia', 'Disease', 'MESH:D000740', (106, 112)) ('PHZ', 'Chemical', 'MESH:C030299', (173, 176)) 8078 23359317 These data suggest that p15Ink4b facilitates RBC formation under conditions of severe anemic stress. ('anemic stress', 'Disease', (86, 99)) ('p15Ink4b', 'Var', (24, 32)) ('RBC formation', 'CPA', (45, 58)) ('facilitates', 'PosReg', (33, 44)) ('anemic stress', 'Disease', 'MESH:D004194', (86, 99)) 8084 23359317 We observed that the animals lacking p15Ink4b showed no increase in MEP (Figure 3d) and BFU-E (Figure 3e) when treated with PHZ, whereas wild-type mice showed a continual increase in these cells over a 40-h period post treatment. ('mice', 'Species', '10090', (147, 151)) ('p15Ink4b', 'Var', (37, 45)) ('MEP', 'Gene', (68, 71)) ('BFU-E', 'CPA', (88, 93)) ('MEP', 'Gene', '13848', (68, 71)) ('PHZ', 'Chemical', 'MESH:C030299', (124, 127)) 8086 23359317 In all, loss of p15Ink4b in mice impairs the balance of erythroid and myeloid progenitor cell formation, preventing sufficient erythropoiesis to allow recovery from anemia. ('impairs', 'NegReg', (33, 40)) ('anemia', 'Phenotype', 'HP:0001903', (165, 171)) ('mice', 'Species', '10090', (28, 32)) ('p15Ink4b', 'Var', (16, 24)) ('preventing', 'NegReg', (105, 115)) ('erythropoiesis', 'MPA', (127, 141)) ('recovery', 'MPA', (151, 159)) ('loss', 'Var', (8, 12)) ('balance', 'MPA', (45, 52)) ('anemia', 'Disease', (165, 171)) ('anemia', 'Disease', 'MESH:D000740', (165, 171)) ('sufficient erythropoiesis', 'Phenotype', 'HP:0010972', (116, 141)) 8088 23359317 Using this system, we were able to efficiently induce expression of low levels of p15Ink4b by simply adding of an appropriate concentration of the inducer named SH. ('expression', 'MPA', (54, 64)) ('S', 'Chemical', 'MESH:D013455', (161, 162)) ('p15Ink4b', 'Var', (82, 90)) ('rat', 'Species', '10116', (133, 136)) 8090 23359317 Expression of p15Ink4b in bone marrow derived from knockout mice restored the BFU-E colony morphology (Figure 4b) to resemble wild-type mice. ('mice', 'Species', '10090', (136, 140)) ('restored', 'PosReg', (65, 73)) ('mice', 'Species', '10090', (60, 64)) ('BFU-E colony morphology', 'CPA', (78, 101)) ('p15Ink4b', 'Var', (14, 22)) 8092 23359317 Overall, this work demonstrates that p15Ink4b has a direct role in regulating the formation of early erythroid progenitor cells in normal bone marrow. ('p15Ink4b', 'Var', (37, 45)) ('rat', 'Species', '10116', (26, 29)) ('formation of early erythroid progenitor cells', 'CPA', (82, 127)) 8093 23359317 To gain molecular insight into the mechanisms through which p15Ink4b protein affects erythropoiesis, we explored the potential utility of the mouse multipotent blood progenitor cell line, EML. ('affects', 'Reg', (77, 84)) ('mouse', 'Species', '10090', (142, 147)) ('protein', 'Protein', (69, 76)) ('p15Ink4b', 'Var', (60, 68)) ('erythropoiesis', 'MPA', (85, 99)) 8101 23359317 Interestingly, even extremely low levels of p15Ink4b that were constitutively produced in the absence of SH, due to leakage in the overexpression system, were able to induce an increase in numbers of colonies (Figure 5f) and alter morphology (Figure 5g), supporting the idea that low p15Ink4b protein levels are sufficient to produce a developmental change. ('morphology', 'CPA', (231, 241)) ('increase', 'PosReg', (177, 185)) ('p15Ink4b', 'Var', (44, 52)) ('S', 'Chemical', 'MESH:D013455', (105, 106)) ('alter', 'Reg', (225, 230)) 8106 23359317 As shown in Figure 6c and d, restoration of p15Ink4b expression in bone marrow progenitors returned the balance of myeloid and erythroid lineage commitment even in the absence of pRb. ('returned', 'PosReg', (91, 99)) ('pRb', 'Gene', '18667', (179, 182)) ('p15Ink4b', 'Var', (44, 52)) ('rat', 'Species', '10116', (34, 37)) ('balance', 'MPA', (104, 111)) ('pRb', 'Gene', (179, 182)) 8110 23359317 Collectively, our data suggest that p15Ink4b when expressed at low levels does not affect Cdk4/6 and might have an additional cell-cycle-independent function (Figure 6e). ('cell-cycle-independent function', 'CPA', (126, 157)) ('p15Ink4b', 'Var', (36, 44)) ('Cdk4/6', 'Gene', '12567;12571', (90, 96)) ('Cdk4/6', 'Gene', (90, 96)) 8112 23359317 The observed dynamic changes following p15Ink4b expression were concomitant with transcriptional upregulation of the EpoR (Figure 7d), a target of GATA-1. ('EpoR', 'Gene', '13857', (117, 121)) ('upregulation', 'PosReg', (97, 109)) ('expression', 'MPA', (48, 58)) ('p15Ink4b', 'Var', (39, 47)) ('EpoR', 'Gene', (117, 121)) ('GATA-1', 'Gene', (147, 153)) ('GATA-1', 'Gene', '14460', (147, 153)) 8114 23359317 As shown in Figure 7e, the expression of p15Ink4b in EML cells specifically results in the phosphorylation of mitogen-activated protein kinase\extracellular signal-regulated kinase (MEK/ERK1/2), a signaling cascade shown previously to be essential for erythropoiesis. ('phosphorylation', 'MPA', (91, 106)) ('MEK', 'Gene', (182, 185)) ('ERK1/2', 'Gene', (186, 192)) ('p15Ink4b', 'Var', (41, 49)) ('MEK', 'Gene', '17242', (182, 185)) ('ERK1/2', 'Gene', '26417;26413', (186, 192)) ('results in', 'Reg', (76, 86)) 8115 23359317 Treatment of p15Ink4b-expressing EML cells with inhibitors of ERK1/2 phosphorylation (U0126 and ERKII) or proteasome inhibitor (MG132) prevented the loss of GATA-2 (Figure 7e). ('ERK1/2', 'Gene', (62, 68)) ('ERK', 'Gene', '26413', (62, 65)) ('ERK', 'Gene', '26413', (96, 99)) ('U0126', 'Chemical', 'MESH:C113580', (86, 91)) ('ERK1/2', 'Gene', '26417;26413', (62, 68)) ('p15Ink4b-expressing', 'Var', (13, 32)) ('MG132', 'Chemical', 'MESH:C072553', (128, 133)) ('U0126', 'Var', (86, 91)) ('ERK', 'Gene', (62, 65)) ('ERK', 'Gene', (96, 99)) ('GATA-2', 'Gene', (157, 163)) 8117 23359317 Our data suggest that p15Ink4b expression, through mechanisms that involve ERK1/2 phosphorylation, might regulate proteasome-mediated degradation of GATA-2, leading to the increase in GATA-1 and EpoR mRNA. ('regulate', 'Reg', (105, 113)) ('GATA-1', 'Gene', (184, 190)) ('increase', 'PosReg', (172, 180)) ('GATA-1', 'Gene', '14460', (184, 190)) ('proteasome-mediated degradation', 'MPA', (114, 145)) ('ERK1/2', 'Gene', '26417;26413', (75, 81)) ('p15Ink4b', 'Var', (22, 30)) ('EpoR', 'Gene', (195, 199)) ('ERK1/2', 'Gene', (75, 81)) ('EpoR', 'Gene', '13857', (195, 199)) 8119 23359317 The AML tumor suppressor p15Ink4b is demonstrated here to have a novel biological function in erythropoiesis. ('AML tumor', 'Disease', (4, 13)) ('tumor', 'Phenotype', 'HP:0002664', (8, 13)) ('rat', 'Species', '10116', (44, 47)) ('AML tumor', 'Disease', 'MESH:D015470', (4, 13)) ('p15Ink4b', 'Var', (25, 33)) ('erythropoiesis', 'Disease', (94, 108)) ('AML', 'Phenotype', 'HP:0004808', (4, 7)) 8120 23359317 Its function in regulating production of erythroid cells may provide an explanation for the anemia observed in MDS and AML patients, 80% of which show a methylation-mediated repression of p15INK4B expression. ('anemia', 'Disease', (92, 98)) ('MDS', 'Disease', (111, 114)) ('MDS', 'Disease', 'MESH:D009190', (111, 114)) ('MDS', 'Phenotype', 'HP:0002863', (111, 114)) ('anemia', 'Disease', 'MESH:D000740', (92, 98)) ('AML', 'Disease', 'MESH:D015470', (119, 122)) ('patients', 'Species', '9606', (123, 131)) ('repression', 'NegReg', (174, 184)) ('p15INK4B', 'Gene', (188, 196)) ('AML', 'Phenotype', 'HP:0004808', (119, 122)) ('AML', 'Disease', (119, 122)) ('anemia', 'Phenotype', 'HP:0001903', (92, 98)) ('expression', 'MPA', (197, 207)) ('methylation-mediated', 'Var', (153, 173)) 8121 23359317 Based on our study, our view of the normal role of p15Ink4b in organisms is to assist the blood system in regulating the lineage fate of progenitor cells by promoting erythroid commitment while suppressing myeloid cell formation, a role that becomes exaggerated under anemic stress. ('suppressing', 'NegReg', (194, 205)) ('erythroid commitment', 'CPA', (167, 187)) ('rat', 'Species', '10116', (256, 259)) ('anemic stress', 'Disease', (268, 281)) ('myeloid cell formation', 'CPA', (206, 228)) ('lineage fate of progenitor cells', 'CPA', (121, 153)) ('assist', 'PosReg', (79, 85)) ('anemic stress', 'Disease', 'MESH:D004194', (268, 281)) ('promoting', 'PosReg', (157, 166)) ('p15Ink4b', 'Var', (51, 59)) 8123 23359317 Our functional demonstration of a role for p15Ink4b in erythropoiesis and blood progenitor homeostasis provides a missing link in the regulation of such networks. ('rat', 'Species', '10116', (22, 25)) ('p15Ink4b', 'Var', (43, 51)) ('erythropoiesis', 'CPA', (55, 69)) 8124 23359317 This knowledge will not only promote further investigation of p15Ink4b in cellular differentiation and regulation of signal-transduction pathways but will also advance our understanding of p15Ink4b in the etiology of the diseases like anemia and cancer. ('cancer', 'Disease', 'MESH:D009369', (246, 252)) ('cancer', 'Disease', (246, 252)) ('anemia', 'Disease', (235, 241)) ('anemia', 'Disease', 'MESH:D000740', (235, 241)) ('cancer', 'Phenotype', 'HP:0002664', (246, 252)) ('anemia', 'Phenotype', 'HP:0001903', (235, 241)) ('p15Ink4b', 'Var', (189, 197)) ('p15Ink4b', 'Var', (62, 70)) ('advance', 'PosReg', (160, 167)) 8125 23359317 One observation made here was that expression of p15Ink4b results in phosphorylation of MEK and ERK1/2, a signaling cascade shown previously to be essential for erythropoiesis. ('ERK1/2', 'Gene', (96, 102)) ('MEK', 'Gene', '17242', (88, 91)) ('results in', 'Reg', (58, 68)) ('phosphorylation', 'MPA', (69, 84)) ('MEK', 'Gene', (88, 91)) ('p15Ink4b', 'Var', (49, 57)) ('ERK1/2', 'Gene', '26417;26413', (96, 102)) 8126 23359317 Interestingly, ERK1/2 was also observed to be an important effector downstream of p15Ink4b in the development of dendritic cells. ('development of dendritic cells', 'CPA', (98, 128)) ('p15Ink4b', 'Var', (82, 90)) ('ERK1/2', 'Gene', (15, 21)) ('ERK1/2', 'Gene', '26417;26413', (15, 21)) 8128 23359317 p15Ink4b-induced signaling may impact a replacement of GATA-2 with GATA-1 at some promoters, a process known as the 'GATA switch'. ('GATA', 'Gene', (117, 121)) ('GATA', 'Gene', '76563', (117, 121)) ('replacement', 'MPA', (40, 51)) ('p15Ink4b-induced', 'Var', (0, 16)) ('GATA-1', 'Gene', '14460', (67, 73)) ('GATA', 'Gene', (67, 71)) ('GATA-1', 'Gene', (67, 73)) ('GATA', 'Gene', '76563', (67, 71)) ('impact', 'Reg', (31, 37)) ('GATA', 'Gene', (55, 59)) ('GATA', 'Gene', '76563', (55, 59)) 8131 23359317 Indeed, altered expression of either p15INK4B or GATA-2 has been linked to poor prognosis in a high number of AML patients. ('p15INK4B', 'Var', (37, 45)) ('linked', 'Reg', (65, 71)) ('GATA-2', 'Gene', (49, 55)) ('AML', 'Disease', (110, 113)) ('AML', 'Phenotype', 'HP:0004808', (110, 113)) ('altered', 'Reg', (8, 15)) ('AML', 'Disease', 'MESH:D015470', (110, 113)) ('patients', 'Species', '9606', (114, 122)) ('expression', 'MPA', (16, 26)) 8132 23359317 Moreover, it has been reported recently that heritable GATA-2 mutations are associated with familial MDS and AML. ('familial MDS', 'Disease', 'MESH:D009190', (92, 104)) ('GATA-2', 'Gene', (55, 61)) ('familial MDS', 'Disease', (92, 104)) ('AML', 'Disease', 'MESH:D015470', (109, 112)) ('MDS', 'Phenotype', 'HP:0002863', (101, 104)) ('associated', 'Reg', (76, 86)) ('AML', 'Disease', (109, 112)) ('AML', 'Phenotype', 'HP:0004808', (109, 112)) ('mutations', 'Var', (62, 71)) 8133 23359317 It has been demonstrated previously, using a large cohort of patients representing various types of hematological malignancies, that loss of p15INK4B but not p16INK4A is characteristic of adult and pediatric AML and pediatric B-ALL. ('loss', 'Var', (133, 137)) ('p15INK4B', 'Var', (141, 149)) ('hematological malignancies', 'Phenotype', 'HP:0004377', (100, 126)) ('AML', 'Disease', 'MESH:D015470', (208, 211)) ('patients', 'Species', '9606', (61, 69)) ('AML', 'Phenotype', 'HP:0004808', (208, 211)) ('AML', 'Disease', (208, 211)) ('rat', 'Species', '10116', (19, 22)) ('hematological malignancies', 'Disease', (100, 126)) ('hematological malignancies', 'Disease', 'MESH:D019337', (100, 126)) 8134 23359317 Inactivation of both genes was rather uncommon and occurred only in pediatric T-ALL and Burkitt's lymphoma. ("Burkitt's lymphoma", 'Disease', (88, 106)) ('lymphoma', 'Phenotype', 'HP:0002665', (98, 106)) ('T-ALL', 'Disease', (78, 83)) ('rat', 'Species', '10116', (31, 34)) ("Burkitt's lymphoma", 'Phenotype', 'HP:0030080', (88, 106)) ('Inactivation', 'Var', (0, 12)) ("Burkitt's lymphoma", 'Disease', 'MESH:D002051', (88, 106)) 8135 23359317 The tumor suppressor function of p16INK4A has been predominantly linked to cancers of epithelial origin and it is associated to its ability to bind CDK4/6 and inhibit cell cycle. ('tumor', 'Disease', (4, 9)) ('CDK4/6', 'Gene', (148, 154)) ('linked', 'Reg', (65, 71)) ('cancers', 'Disease', 'MESH:D009369', (75, 82)) ('cancers', 'Phenotype', 'HP:0002664', (75, 82)) ('cancers', 'Disease', (75, 82)) ('CDK4/6', 'Gene', '12567;12571', (148, 154)) ('cancer', 'Phenotype', 'HP:0002664', (75, 81)) ('tumor', 'Disease', 'MESH:D009369', (4, 9)) ('tumor', 'Phenotype', 'HP:0002664', (4, 9)) ('bind', 'Interaction', (143, 147)) ('p16INK4A', 'Var', (33, 41)) ('inhibit', 'NegReg', (159, 166)) ('cell cycle', 'CPA', (167, 177)) 8138 23359317 There was an increase in the expression of p16Ink4a in the absence of p15Ink4b that could represent a compensation mechanism. ('p16Ink4a', 'Gene', '12578', (43, 51)) ('p15Ink4b', 'Var', (70, 78)) ('p16Ink4a', 'Gene', (43, 51)) ('increase', 'PosReg', (13, 21)) ('expression', 'MPA', (29, 39)) 8143 23359317 Loss of p15Ink4b in mice impairs the balance between erythroid and myeloid progenitor cell formation, preventing sufficient erythropoiesis to allow recovery from anemia. ('p15Ink4b', 'Var', (8, 16)) ('anemia', 'Disease', 'MESH:D000740', (162, 168)) ('anemia', 'Disease', (162, 168)) ('balance', 'MPA', (37, 44)) ('mice', 'Species', '10090', (20, 24)) ('sufficient erythropoiesis', 'Phenotype', 'HP:0010972', (113, 138)) ('anemia', 'Phenotype', 'HP:0001903', (162, 168)) ('impairs', 'NegReg', (25, 32)) ('preventing', 'NegReg', (102, 112)) ('Loss', 'Var', (0, 4)) ('erythropoiesis', 'MPA', (124, 138)) ('recovery', 'MPA', (148, 156)) 8145 23359317 Indeed, we have previously demonstrated that loss of p15Ink4b in mice results in monocytosis and predisposition to myeloid leukemia. ('monocytosis', 'Phenotype', 'HP:0012311', (81, 92)) ('leukemia', 'Disease', (123, 131)) ('leukemia', 'Phenotype', 'HP:0001909', (123, 131)) ('leukemia', 'Disease', 'MESH:D007938', (123, 131)) ('results', 'Reg', (70, 77)) ('mice', 'Species', '10090', (65, 69)) ('p15Ink4b', 'Var', (53, 61)) ('monocytosis', 'MPA', (81, 92)) ('rat', 'Species', '10116', (34, 37)) ('predisposition', 'Reg', (97, 111)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (115, 131)) ('loss', 'Var', (45, 49)) 8236 19344392 MTIs may cause hyperlipidemia, mild myelosuppression, hypertension, and mucositis. ('hypertension', 'Disease', (54, 66)) ('MTIs', 'Var', (0, 4)) ('myelosuppression', 'Disease', 'MESH:D001855', (36, 52)) ('hypertension', 'Phenotype', 'HP:0000822', (54, 66)) ('myelosuppression', 'Disease', (36, 52)) ('mucositis', 'Disease', 'MESH:D052016', (72, 81)) ('hyperlipidemia', 'Disease', 'MESH:D006949', (15, 29)) ('hyperlipidemia', 'Phenotype', 'HP:0003077', (15, 29)) ('hypertension', 'Disease', 'MESH:D006973', (54, 66)) ('hyperlipidemia', 'Disease', (15, 29)) ('cause', 'Reg', (9, 14)) ('MTIs', 'Chemical', '-', (0, 4)) ('mucositis', 'Disease', (72, 81)) 8257 19344392 IGFR, HER/EGFR), point mutations in the PIK3CA (p110alpha PI3K) gene, loss of tumor suppressor genes (e.g. ('IGFR', 'Gene', (0, 4)) ('EGFR', 'Gene', '1956', (10, 14)) ('PIK3CA', 'Gene', (40, 46)) ('loss', 'NegReg', (70, 74)) ('EGFR', 'Gene', (10, 14)) ('tumor', 'Disease', 'MESH:D009369', (78, 83)) ('PIK3CA', 'Gene', '5290', (40, 46)) ('IGFR', 'Gene', '3480', (0, 4)) ('tumor', 'Phenotype', 'HP:0002664', (78, 83)) ('point mutations', 'Var', (17, 32)) ('tumor', 'Disease', (78, 83)) ('p110alpha', 'Gene', (48, 57)) ('p110alpha', 'Gene', '5290', (48, 57)) 8258 19344392 PTEN or TSC1:TSC2 complex), and gain-of-function mutations in mTOR or mTOR-linked pathways (e.g. ('TSC2', 'Gene', (13, 17)) ('mTOR', 'Gene', '2475', (62, 66)) ('TSC1', 'Gene', '7248', (8, 12)) ('gain-of-function', 'PosReg', (32, 48)) ('mTOR', 'Gene', (70, 74)) ('mTOR', 'Gene', (62, 66)) ('mutations', 'Var', (49, 58)) ('TSC1', 'Gene', (8, 12)) ('mTOR', 'Gene', '2475', (70, 74)) ('TSC2', 'Gene', '7249', (13, 17)) ('PTEN', 'Gene', (0, 4)) ('PTEN', 'Gene', '5728', (0, 4)) 8259 19344392 formation of the aberrant protein BCR-ABL in Ph+ leukemia cells or stimulation of PI3K by aberrant ras/raf/MAPK pathway intermediates). ('raf', 'Gene', '22882', (103, 106)) ('leukemia', 'Disease', (49, 57)) ('leukemia', 'Phenotype', 'HP:0001909', (49, 57)) ('BCR-ABL', 'Gene', '25', (34, 41)) ('raf', 'Gene', (103, 106)) ('leukemia', 'Disease', 'MESH:D007938', (49, 57)) ('aberrant', 'Var', (90, 98)) ('BCR-ABL', 'Gene', (34, 41)) 8267 19344392 As mTOR inhibitors have activity against lymphocytes and abnormal signaling can lead to neoplastic transformation, our group hypothesized that ALL cells may be dependent on mTOR signaling and studied the effects of MTIs on ALL blasts. ('mTOR', 'Gene', '2475', (3, 7)) ('lead to', 'Reg', (80, 87)) ('al', 'Chemical', 'MESH:D000535', (182, 184)) ('al', 'Chemical', 'MESH:D000535', (63, 65)) ('mTOR', 'Gene', '2475', (173, 177)) ('al', 'Chemical', 'MESH:D000535', (70, 72)) ('neoplastic transformation', 'CPA', (88, 113)) ('mTOR', 'Gene', (173, 177)) ('MTIs', 'Chemical', '-', (215, 219)) ('abnormal', 'Var', (57, 65)) ('mTOR', 'Gene', (3, 7)) 8322 19344392 As other inhibitors of the PI3K/AKT/mTOR pathway are developed and tested in clinical trials, these agents may prove superior to mTOR inhibitors, since targeting PI3K with LY294002, AKT with perifosine, and both mTOR and PI3K with the dual inhibitor PI103 have shown promise in preclinical studies. ('perifosine', 'Chemical', 'MESH:C105905', (191, 201)) ('LY294002', 'Var', (172, 180)) ('mTOR', 'Gene', '2475', (36, 40)) ('PI3K', 'Var', (162, 166)) ('mTOR', 'Gene', (129, 133)) ('al', 'Chemical', 'MESH:D000535', (237, 239)) ('AKT', 'Gene', '207', (32, 35)) ('mTOR', 'Gene', '2475', (129, 133)) ('AKT', 'Gene', '207', (182, 185)) ('PI3K/AKT', 'Gene', (27, 35)) ('LY294002', 'Chemical', 'MESH:C085911', (172, 180)) ('PI3K/AKT', 'Gene', '5290;207', (27, 35)) ('mTOR', 'Gene', (212, 216)) ('mTOR', 'Gene', (36, 40)) ('al', 'Chemical', 'MESH:D000535', (83, 85)) ('AKT', 'Gene', (32, 35)) ('al', 'Chemical', 'MESH:D000535', (287, 289)) ('al', 'Chemical', 'MESH:D000535', (89, 91)) ('mTOR', 'Gene', '2475', (212, 216)) ('AKT', 'Gene', (182, 185)) 8326 19344392 MDS are a heterogenous group of disorders characterized by cytopenias from defects in haematopoietic stem cell differentiation which frequently transform into acute myelogenous leukemia. ('acute myelogenous leukemia', 'Phenotype', 'HP:0004808', (159, 185)) ('leukemia', 'Phenotype', 'HP:0001909', (177, 185)) ('defects', 'Var', (75, 82)) ('acute myelogenous leukemia', 'Disease', 'MESH:D015470', (159, 185)) ('transform', 'Reg', (144, 153)) ('acute myelogenous leukemia', 'Disease', (159, 185)) ('cytopenias', 'Disease', 'MESH:D006402', (59, 69)) ('cytopenias', 'Disease', (59, 69)) ('myelogenous leukemia', 'Phenotype', 'HP:0012324', (165, 185)) ('MDS', 'Disease', (0, 3)) ('MDS', 'Disease', 'MESH:D009190', (0, 3)) 8329 19344392 hypothesized that aberrant activation of these survival signals may lead to transformation of MDS into AML. ('MDS', 'Disease', 'MESH:D009190', (94, 97)) ('AML', 'Disease', (103, 106)) ('lead to', 'Reg', (68, 75)) ('MDS', 'Disease', (94, 97)) ('transformation', 'CPA', (76, 90)) ('aberrant', 'Var', (18, 26)) ('al', 'Chemical', 'MESH:D000535', (53, 55)) ('activation', 'PosReg', (27, 37)) ('AML', 'Disease', 'MESH:D015470', (103, 106)) ('al', 'Chemical', 'MESH:D000535', (60, 62)) 8353 19344392 Imatinib was the first TKI to be used in CML; unfortunately, approximately 25% of patients will either have innate resistance or more commonly acquire resistance to imatinib, because of Bcr-Abl mutations. ('Imatinib', 'Chemical', 'MESH:D000068877', (0, 8)) ('patients', 'Species', '9606', (82, 90)) ('acquire', 'Reg', (143, 150)) ('mutations', 'Var', (194, 203)) ('Bcr-Abl', 'Gene', (186, 193)) ('Bcr-Abl', 'Gene', '25', (186, 193)) ('imatinib', 'Chemical', 'MESH:D000068877', (165, 173)) ('innate resistance', 'MPA', (108, 125)) ('resistance to imatinib', 'MPA', (151, 173)) 8354 19344392 The majority of these patients will respond to second line TKIs, but a subset have a particular mutation (T3151) that is not treatable with current Bcr-Abl targeting TKIs. ('abl', 'Gene', '25', (130, 133)) ('abl', 'Gene', (130, 133)) ('patients', 'Species', '9606', (22, 30)) ('T3151', 'Var', (106, 111)) ('Bcr-Abl', 'Gene', (148, 155)) ('Bcr-Abl', 'Gene', '25', (148, 155)) 8359 19344392 Sirolimus has also been shown to be effective in vitro against resistant CML, including cells with T3151 mutations. ('T3151 mutations', 'Var', (99, 114)) ('al', 'Chemical', 'MESH:D000535', (14, 16)) ('CML', 'Disease', (73, 76)) ('Sirolimus', 'Chemical', 'MESH:D020123', (0, 9)) 8369 19344392 hypothesized that targeting mTOR would be an effective treatment in mantle cell lymphoma by down-regulating cyclin D1 expression. ('mantle cell lymphoma', 'Disease', (68, 88)) ('mTOR', 'Gene', '2475', (28, 32)) ('cyclin D1', 'Gene', (108, 117)) ('mantle cell lymphoma', 'Disease', 'MESH:D020522', (68, 88)) ('mTOR', 'Gene', (28, 32)) ('lymphoma', 'Phenotype', 'HP:0002665', (80, 88)) ('down-regulating', 'NegReg', (92, 107)) ('targeting', 'Var', (18, 27)) ('cell lymphoma', 'Phenotype', 'HP:0012191', (75, 88)) ('cyclin D1', 'Gene', '595', (108, 117)) 8372 19344392 Other preclinical work has demonstrated that mTOR inhibitors synergize in vitro with a number of agents used to treat mantle cell, including rituximab, vincristine, doxorubicin, and bortezomib. ('inhibitors', 'Var', (50, 60)) ('bortezomib', 'Chemical', 'MESH:D000069286', (182, 192)) ('rituximab', 'Chemical', 'MESH:D000069283', (141, 150)) ('doxorubicin', 'Chemical', 'MESH:D004317', (165, 176)) ('mTOR', 'Gene', '2475', (45, 49)) ('al', 'Chemical', 'MESH:D000535', (15, 17)) ('vincristine', 'Chemical', 'MESH:D014750', (152, 163)) ('mTOR', 'Gene', (45, 49)) 8376 19344392 AKT and the down-stream intermediates, 4E-BP1 and S6K1, were shown to be activated in Hodgkin's lymphoma cell lines; however, targeting either PI3K with LY294002 or mTOR with sirolimus only showed a modest effect in vitro. ('AKT', 'Gene', (0, 3)) ('LY294002', 'Chemical', 'MESH:C085911', (153, 161)) ("Hodgkin's lymphoma", 'Phenotype', 'HP:0012189', (86, 104)) ('mTOR', 'Gene', (165, 169)) ('4E-BP1', 'Gene', (39, 45)) ('LY294002', 'Var', (153, 161)) ('AKT', 'Gene', '207', (0, 3)) ('4E-BP1', 'Gene', '1978', (39, 45)) ("Hodgkin's lymphoma", 'Disease', 'MESH:D006689', (86, 104)) ('mTOR', 'Gene', '2475', (165, 169)) ('lymphoma', 'Phenotype', 'HP:0002665', (96, 104)) ('S6K1', 'Gene', (50, 54)) ('activated', 'PosReg', (73, 82)) ("Hodgkin's lymphoma", 'Disease', (86, 104)) ('sirolimus', 'Chemical', 'MESH:D020123', (175, 184)) ('S6K1', 'Gene', '6198', (50, 54)) 8395 19344392 In other studies, sirolimus was found to sensitize multiple myeloma cells to dexamethasone, and mTOR inhibitors were found to synergize with other targeted agents, including sunitinib, a HSP90 inhibitor, and lenalidomide. ('sensitize', 'Reg', (41, 50)) ('mTOR', 'Gene', (96, 100)) ('inhibitors', 'Var', (101, 111)) ('sunitinib', 'Chemical', 'MESH:D000077210', (174, 183)) ('multiple myeloma', 'Phenotype', 'HP:0006775', (51, 67)) ('dexamethasone', 'Chemical', 'MESH:D003907', (77, 90)) ('sirolimus', 'Chemical', 'MESH:D020123', (18, 27)) ('multiple myeloma', 'Disease', (51, 67)) ('lenalidomide', 'Chemical', 'MESH:D000077269', (208, 220)) ('synergize', 'Reg', (126, 135)) ('HSP90', 'Gene', (187, 192)) ('HSP90', 'Gene', '3320', (187, 192)) ('mTOR', 'Gene', '2475', (96, 100)) ('multiple myeloma', 'Disease', 'MESH:D009101', (51, 67)) 8397 19344392 In addition to the recent interest of MTIs in haematologic malignancies, targeting mTOR signaling has been studied in non-malignant haematologic disorders, particularly autoimmune disorders, for two reasons: (1) MTIs can cause apoptosis in abnormal lymphocytes, whereas many immunosuppressive agents only inhibit growth; and (2) mTOR inhibitors increase peripheral blood regulatory T cells (Tregs). ('MTIs', 'Chemical', '-', (38, 42)) ('mTOR', 'Gene', (83, 87)) ('haematologic disorders', 'Disease', 'MESH:D006402', (132, 154)) ('al', 'Chemical', 'MESH:D000535', (246, 248)) ('cause', 'Reg', (221, 226)) ('mTOR', 'Gene', (329, 333)) ('MTIs', 'Var', (212, 216)) ('haematologic malignancies', 'Disease', (46, 71)) ('mTOR', 'Gene', '2475', (83, 87)) ('haematologic disorders', 'Phenotype', 'HP:0001871', (132, 154)) ('peripheral blood regulatory T cells', 'CPA', (354, 389)) ('abnormal lymphocytes', 'Phenotype', 'HP:0004332', (240, 260)) ('haematologic disorders', 'Disease', (132, 154)) ('apoptosis', 'CPA', (227, 236)) ('mTOR', 'Gene', '2475', (329, 333)) ('increase', 'PosReg', (345, 353)) ('autoimmune disorders', 'Disease', (169, 189)) ('al', 'Chemical', 'MESH:D000535', (362, 364)) ('haematologic disorder', 'Phenotype', 'HP:0001871', (132, 153)) ('haematologic malignancies', 'Disease', 'MESH:D019337', (46, 71)) ('al', 'Chemical', 'MESH:D000535', (92, 94)) ('al', 'Chemical', 'MESH:D000535', (60, 62)) ('al', 'Chemical', 'MESH:D000535', (123, 125)) ('autoimmune disorders', 'Phenotype', 'HP:0002960', (169, 189)) ('MTIs', 'Chemical', '-', (212, 216)) ('autoimmune disorders', 'Disease', 'MESH:D001327', (169, 189)) 8423 18087601 Some epidemiologic studies have suggested that pesticides might increase the risk of childhood hematopoietic malignancies. ('hematopoietic malignancies', 'Disease', 'MESH:D019337', (95, 121)) ('hematopoietic malignancies', 'Disease', (95, 121)) ('pesticides', 'Var', (47, 57)) ('child', 'Species', '9606', (85, 90)) 8527 33182397 Novel DNMT3A Germline Variant in a Patient with Multiple Paragangliomas and Papillary Thyroid Carcinoma The use of next generation technologies has helped to unravel the genetics of rare inherited diseases, facilitating the discovery of new susceptibility genes. ('inherited diseases', 'Disease', 'MESH:D030342', (187, 205)) ('Patient', 'Species', '9606', (35, 42)) ('Paragangliomas and Papillary Thyroid Carcinoma', 'Disease', 'MESH:D000077273', (57, 103)) ('Thyroid Carcinoma', 'Phenotype', 'HP:0002890', (86, 103)) ('Papillary Thyroid Carcinoma', 'Phenotype', 'HP:0002895', (76, 103)) ('Carcinoma', 'Phenotype', 'HP:0030731', (94, 103)) ('Paragangliomas', 'Phenotype', 'HP:0002668', (57, 71)) ('inherited diseases', 'Disease', (187, 205)) ('Variant', 'Var', (22, 29)) ('DNMT3A', 'Gene', (6, 12)) ('DNMT3A', 'Gene', '1788', (6, 12)) 8529 33182397 Here we describe a novel DNMT3A germline variant identified by whole-exome sequencing in a patient with multiple paragangliomas and papillary thyroid carcinoma. ('thyroid carcinoma', 'Phenotype', 'HP:0002890', (142, 159)) ('paragangliomas', 'Phenotype', 'HP:0002668', (113, 127)) ('paraganglioma', 'Phenotype', 'HP:0002668', (113, 126)) ('carcinoma', 'Phenotype', 'HP:0030731', (150, 159)) ('DNMT3A', 'Gene', (25, 31)) ('multiple paragangliomas and papillary thyroid carcinoma', 'Disease', 'MESH:D000077273', (104, 159)) ('papillary thyroid carcinoma', 'Phenotype', 'HP:0002895', (132, 159)) ('variant', 'Var', (41, 48)) ('patient', 'Species', '9606', (91, 98)) 8530 33182397 The increased methylation of DNMT3A target genes observed in the proband's sample points towards a gain-of-function effect of the variant, contrasting with the inactivation caused by loss-of-function alterations commonly seen in other neoplasia and in patients with Tatton-Brown-Rahman syndrome. ('gain-of-function', 'PosReg', (99, 115)) ('Tatton-Brown-Rahman syndrome', 'Disease', (266, 294)) ('neoplasia', 'Disease', (235, 244)) ('methylation', 'MPA', (14, 25)) ('Tatton-Brown-Rahman syndrome', 'Disease', 'OMIM:615879', (266, 294)) ('patients', 'Species', '9606', (252, 260)) ('variant', 'Var', (130, 137)) ('neoplasia', 'Disease', 'MESH:D009369', (235, 244)) ('neoplasia', 'Phenotype', 'HP:0002664', (235, 244)) ('increased', 'PosReg', (4, 13)) 8532 33182397 We recently found germline DNMT3A gain-of-function variants in two patients with head and neck paragangliomas causing a characteristic hypermethylated DNA profile. ('paraganglioma', 'Phenotype', 'HP:0002668', (95, 108)) ('patients', 'Species', '9606', (67, 75)) ('head and neck paragangliomas', 'Phenotype', 'HP:0002864', (81, 109)) ('neck paragangliomas', 'Disease', 'MESH:D010235', (90, 109)) ('variants', 'Var', (51, 59)) ('gain-of-function', 'PosReg', (34, 50)) ('hypermethylated DNA profile', 'MPA', (135, 162)) ('DNMT3A', 'Gene', (27, 33)) ('neck paragangliomas', 'Disease', (90, 109)) ('paragangliomas', 'Phenotype', 'HP:0002668', (95, 109)) 8533 33182397 Here, whole-exome sequencing identifies a novel germline DNMT3A variant (p.Gly332Arg) in a patient with bilateral carotid paragangliomas, papillary thyroid carcinoma and idiopathic intellectual disability. ('papillary thyroid carcinoma', 'Disease', (138, 165)) ('papillary thyroid carcinoma', 'Disease', 'MESH:D000077273', (138, 165)) ('DNMT3A', 'Gene', (57, 63)) ('paragangliomas', 'Phenotype', 'HP:0002668', (122, 136)) ('carcinoma', 'Phenotype', 'HP:0030731', (156, 165)) ('paraganglioma', 'Phenotype', 'HP:0002668', (122, 135)) ('thyroid carcinoma', 'Phenotype', 'HP:0002890', (148, 165)) ('bilateral carotid paragangliomas', 'Disease', (104, 136)) ('papillary thyroid carcinoma', 'Phenotype', 'HP:0002895', (138, 165)) ('carotid paragangliomas', 'Phenotype', 'HP:0100635', (114, 136)) ('patient', 'Species', '9606', (91, 98)) ('p.Gly332Arg', 'Var', (73, 84)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (73, 84)) ('bilateral carotid paragangliomas', 'Disease', 'MESH:D002345', (104, 136)) ('intellectual disability', 'Phenotype', 'HP:0001249', (181, 204)) 8534 33182397 The variant, located in the Pro-Trp-Trp-Pro (PWWP) domain of the protein involved in chromatin targeting, affects a residue mutated in papillary thyroid tumors and located between the two residues found mutated in microcephalic dwarfism patients. ('Trp', 'Chemical', 'MESH:D014364', (36, 39)) ('papillary thyroid tumors', 'Phenotype', 'HP:0002895', (135, 159)) ('Pro', 'Chemical', 'MESH:D011392', (40, 43)) ('mutated', 'Var', (124, 131)) ('papillary thyroid tumors', 'Disease', 'MESH:D000077273', (135, 159)) ('tumor', 'Phenotype', 'HP:0002664', (153, 158)) ('microcephalic dwarfism', 'Phenotype', 'HP:0008873', (214, 236)) ('affects', 'Reg', (106, 113)) ('microcephalic dwarfism', 'Disease', (214, 236)) ('microcephalic dwarfism', 'Disease', 'MESH:C537533', (214, 236)) ('tumors', 'Phenotype', 'HP:0002664', (153, 159)) ('Pro', 'Chemical', 'MESH:D011392', (28, 31)) ('patients', 'Species', '9606', (237, 245)) ('papillary thyroid tumors', 'Disease', (135, 159)) ('Trp', 'Chemical', 'MESH:D014364', (32, 35)) ('dwarfism', 'Phenotype', 'HP:0003510', (228, 236)) ('Pro-Trp', 'Chemical', '-', (28, 35)) 8535 33182397 An increased methylation of DNMT3A target genes, compatible with a gain-of-function effect of the alteration, was observed in saliva DNA from the proband and in one independent acute myeloid leukemia sample carrying the same p.Gly332Arg variant. ('increased', 'PosReg', (3, 12)) ('methylation', 'MPA', (13, 24)) ('acute myeloid leukemia', 'Disease', (177, 199)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (225, 236)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (177, 199)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (177, 199)) ('leukemia', 'Phenotype', 'HP:0001909', (191, 199)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (183, 199)) ('p.Gly332Arg', 'Var', (225, 236)) 8536 33182397 Although further studies are needed to support a causal role of DNMT3A variants in paraganglioma, the description of a new DNMT3A alteration in a patient with multiple clinical features suggests a heterogeneous phenotypic spectrum related to DNMT3A germline variants. ('variants', 'Var', (71, 79)) ('alteration', 'Var', (130, 140)) ('paraganglioma', 'Phenotype', 'HP:0002668', (83, 96)) ('DNMT3A', 'Gene', (123, 129)) ('paraganglioma', 'Disease', (83, 96)) ('patient', 'Species', '9606', (146, 153)) ('paraganglioma', 'Disease', 'MESH:D010235', (83, 96)) 8537 33182397 The drip of new susceptibility genes involved in the development of pheochromocytoma (PCC) and paraganglioma (PGL), altogether PPGLs, has not ceased since the discovery of pathogenic variants in the succinate dehydrogenase (SDH) genes in the early 2000s. ('SDH', 'Gene', '6390', (224, 227)) ('succinate dehydrogenase', 'Gene', (199, 222)) ('paraganglioma', 'Disease', (95, 108)) ('succinate dehydrogenase', 'Gene', '6390', (199, 222)) ('paraganglioma', 'Phenotype', 'HP:0002668', (95, 108)) ('pheochromocytoma', 'Disease', (68, 84)) ('variants', 'Var', (183, 191)) ('PPGL', 'Chemical', '-', (127, 131)) ('pheochromocytoma', 'Phenotype', 'HP:0002666', (68, 84)) ('SDH', 'Gene', (224, 227)) ('PGL', 'Phenotype', 'HP:0002668', (110, 113)) ('pheochromocytoma', 'Disease', 'MESH:D010673', (68, 84)) ('paraganglioma', 'Disease', 'MESH:D010235', (95, 108)) ('PGL', 'Phenotype', 'HP:0002668', (128, 131)) ('PCC', 'Phenotype', 'HP:0002666', (86, 89)) 8538 33182397 Thus, up to 35-40% of PPGL patients carry a germline variant in one of the almost twenty high:or low:penetrant susceptibility genes related to the disease described so far. ('PGL', 'Phenotype', 'HP:0002668', (23, 26)) ('PPGL', 'Disease', (22, 26)) ('germline variant', 'Var', (44, 60)) ('PPGL', 'Chemical', '-', (22, 26)) ('patients', 'Species', '9606', (27, 35)) 8543 33182397 On the one hand, germline de novo loss-of-function variants in DNMT3A cause Tatton-Brown-Rahman syndrome (TBRS), an autosomal dominant condition characterized by overgrowth, intellectual disability and distinctive facial appearance. ('distinctive facial appearance', 'Phenotype', 'HP:0001999', (202, 231)) ('Tatton-Brown-Rahman syndrome', 'Disease', (76, 104)) ('Tatton-Brown-Rahman syndrome', 'Disease', 'OMIM:615879', (76, 104)) ('variants', 'Var', (51, 59)) ('loss-of-function', 'NegReg', (34, 50)) ('overgrowth', 'Phenotype', 'HP:0001548', (162, 172)) ('DNMT3A', 'Gene', (63, 69)) ('intellectual disability', 'Phenotype', 'HP:0001249', (174, 197)) 8544 33182397 On the other hand and despite not being identified until 2010, somatic variants in DNMT3A are one of the most recurrent events across blood cancers of the myeloid lineage. ('variants', 'Var', (71, 79)) ('blood cancers', 'Disease', (134, 147)) ('blood cancers', 'Phenotype', 'HP:0001909', (134, 147)) ('age', 'Gene', (167, 170)) ('cancers', 'Phenotype', 'HP:0002664', (140, 147)) ('blood cancers', 'Disease', 'MESH:D007022', (134, 147)) ('age', 'Gene', '5973', (167, 170)) ('DNMT3A', 'Gene', (83, 89)) ('cancer', 'Phenotype', 'HP:0002664', (140, 146)) 8545 33182397 Up to 22% of acute myeloid leukemia (AML) patients carry a somatic variant in DNMT3A and these alterations have been associated with poor prognosis and adverse survival outcomes for AML patients. ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (13, 35)) ('leukemia', 'Phenotype', 'HP:0001909', (27, 35)) ('associated', 'Reg', (117, 127)) ('DNMT3A', 'Gene', (78, 84)) ('variant', 'Var', (67, 74)) ('AML', 'Disease', 'MESH:D015470', (37, 40)) ('AML', 'Disease', 'MESH:D015470', (182, 185)) ('AML', 'Phenotype', 'HP:0004808', (37, 40)) ('acute myeloid leukemia', 'Disease', (13, 35)) ('AML', 'Disease', (37, 40)) ('AML', 'Phenotype', 'HP:0004808', (182, 185)) ('patients', 'Species', '9606', (42, 50)) ('AML', 'Disease', (182, 185)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (13, 35)) ('patients', 'Species', '9606', (186, 194)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (19, 35)) 8546 33182397 Interestingly, there is overlap of the spectrum of DNMT3A variants in TBRS and hematological malignancies, with some examples of both diseases occurring in the same patient. ('variants', 'Var', (58, 66)) ('hematological malignancies', 'Disease', (79, 105)) ('patient', 'Species', '9606', (165, 172)) ('hematological malignancies', 'Disease', 'MESH:D019337', (79, 105)) ('TBRS', 'Disease', (70, 74)) ('DNMT3A', 'Gene', (51, 57)) ('hematological malignancies', 'Phenotype', 'HP:0004377', (79, 105)) 8547 33182397 Moreover, a germline DNMT3A loss-of-function variant has been found causing familial AML, in this case in the absence of TBRS symptoms. ('loss-of-function', 'NegReg', (28, 44)) ('DNMT3A', 'Gene', (21, 27)) ('familial AML', 'Disease', 'MESH:D015470', (76, 88)) ('familial AML', 'Disease', (76, 88)) ('AML', 'Phenotype', 'HP:0004808', (85, 88)) ('variant', 'Var', (45, 52)) 8548 33182397 We recently identified missense germline variants in DNMT3A in two patients with multiple head and neck PGLs. ('DNMT3A', 'Gene', (53, 59)) ('patients', 'Species', '9606', (67, 75)) ('missense germline variants', 'Var', (23, 49)) ('PGL', 'Phenotype', 'HP:0002668', (104, 107)) 8549 33182397 Unlike what happens in patients with TBRS and in AML and other neoplasia, in which the gene is usually inactivated by loss-of-function alterations, we demonstrated that DNMT3A-mutated PGLs exhibited a significant overall methylation, indicating an activating role of the variants. ('PGL', 'Phenotype', 'HP:0002668', (184, 187)) ('AML', 'Disease', (49, 52)) ('neoplasia', 'Phenotype', 'HP:0002664', (63, 72)) ('PGLs', 'Gene', (184, 188)) ('DNMT3A-mutated', 'Var', (169, 183)) ('neoplasia', 'Disease', 'MESH:D009369', (63, 72)) ('patients', 'Species', '9606', (23, 31)) ('methylation', 'MPA', (221, 232)) ('activating', 'PosReg', (248, 258)) ('neoplasia', 'Disease', (63, 72)) ('AML', 'Disease', 'MESH:D015470', (49, 52)) ('AML', 'Phenotype', 'HP:0004808', (49, 52)) 8551 33182397 This duality has been also reported for the histone methyltransferase EZH2, in which somatic gain-of-function variants are frequently found in lymphoma, while inactivating germline alterations cause the Weaver overgrowth syndrome. ('lymphoma', 'Disease', (143, 151)) ('lymphoma', 'Disease', 'MESH:D008223', (143, 151)) ('overgrowth syndrome', 'Disease', 'MESH:D001765', (210, 229)) ('variants', 'Var', (110, 118)) ('histone methyltransferase', 'Gene', '56979', (44, 69)) ('overgrowth', 'Phenotype', 'HP:0001548', (210, 220)) ('overgrowth syndrome', 'Disease', (210, 229)) ('lymphoma', 'Phenotype', 'HP:0002665', (143, 151)) ('histone methyltransferase', 'Gene', (44, 69)) ('EZH2', 'Gene', '2146', (70, 74)) ('EZH2', 'Gene', (70, 74)) ('gain-of-function', 'PosReg', (93, 109)) 8553 33182397 The filtering process resulted in 541 variants, six of which were found in the public archive ClinVar (Table S1) and only one was found in a gene previously associated to PPGLs, DNMT3A (c.994G > A, p.Gly332Arg; VAF = 0.48). ('AF', 'Disease', 'MESH:D001281', (213, 215)) ('resulted in', 'Reg', (22, 33)) ('variants', 'Var', (38, 46)) ('p.Gly332Arg', 'Var', (199, 210)) ('c.994G > A', 'Var', (187, 197)) ('c.994G > A', 'Mutation', 'rs760854242', (187, 197)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (199, 210)) ('PGL', 'Phenotype', 'HP:0002668', (173, 176)) ('PPGL', 'Chemical', '-', (172, 176)) 8555 33182397 This frequency is below the frequency found for pathogenic variants in SDHB, a gene whose alterations have the lowest penetrance amongst those found in the known PPGL susceptibility genes. ('SDHB', 'Gene', '6390', (71, 75)) ('variants', 'Var', (59, 67)) ('SDHB', 'Gene', (71, 75)) ('PGL', 'Phenotype', 'HP:0002668', (163, 166)) ('PPGL', 'Chemical', '-', (162, 166)) 8556 33182397 The Gly332 residue is located within the Pro-Trp-Trp-Pro (PWWP) domain of DNMT3A, described to function as a chromatin methylation reader by recognizing both DNA and histone methylated lysines. ('lysines', 'Chemical', 'MESH:D008239', (185, 192)) ('Gly332', 'Var', (4, 10)) ('DNA', 'MPA', (158, 161)) ('Pro', 'Chemical', 'MESH:D011392', (53, 56)) ('DNMT3A', 'Gene', (74, 80)) ('Pro', 'Chemical', 'MESH:D011392', (41, 44)) ('Trp', 'Chemical', 'MESH:D014364', (45, 48)) ('Pro-Trp', 'Chemical', '-', (41, 48)) ('Trp', 'Chemical', 'MESH:D014364', (49, 52)) ('Gly332', 'Chemical', '-', (4, 10)) ('histone', 'Protein', (166, 173)) ('recognizing', 'Reg', (141, 152)) 8557 33182397 There are 31 missense variants in gnomAD, including p.Gly332Arg, located within the PWWP domain of DNMT3A, all of them with frequencies bellow 6.4 x 10-5 (Table S2). ('gnomAD', 'Gene', (34, 40)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (52, 63)) ('p.Gly332Arg', 'Var', (52, 63)) 8559 33182397 Three different missense somatic variants have been reported in five independent samples in COSMIC or cBioPortal databases affecting the Gly332 residue (Figure 1a): p.Gly332Glu (n = 1; found in one clear cell renal cell carcinoma), p.Gly332Arg (n = 3; found in PTC, AML and essential thrombocythemia) and p.Gly332Val (n = 1; found in one PTC); although the causality of these variants has not been assessed. ('clear cell renal cell carcinoma', 'Disease', (200, 231)) ('renal cell carcinoma', 'Phenotype', 'HP:0005584', (211, 231)) ('p.Gly332Glu', 'Var', (167, 178)) ('p.Gly332Arg', 'Var', (234, 245)) ('affecting', 'Reg', (125, 134)) ('essential thrombocythemia', 'Disease', (276, 301)) ('Gly332', 'Chemical', '-', (309, 315)) ('p.Gly332Val', 'Var', (307, 318)) ('Gly332', 'Chemical', '-', (236, 242)) ('essential thrombocythemia', 'Disease', 'MESH:D013920', (276, 301)) ('clear cell renal cell carcinoma', 'Phenotype', 'HP:0006770', (200, 231)) ('carcinoma', 'Phenotype', 'HP:0030731', (222, 231)) ('PTC', 'Phenotype', 'HP:0002895', (263, 266)) ('Gly332', 'Chemical', '-', (139, 145)) ('p.Gly332Val', 'Mutation', 'p.G332V', (307, 318)) ('thrombocythemia', 'Phenotype', 'HP:0001894', (286, 301)) ('p.Gly332Glu', 'Mutation', 'rs751360082', (167, 178)) ('PTC', 'Phenotype', 'HP:0002895', (340, 343)) ('AML', 'Disease', 'MESH:D015470', (268, 271)) ('clear cell renal cell carcinoma', 'Disease', 'MESH:C538614', (200, 231)) ('AML', 'Disease', (268, 271)) ('Gly332', 'Chemical', '-', (169, 175)) ('AML', 'Phenotype', 'HP:0004808', (268, 271)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (234, 245)) 8562 33182397 In the predicted structure of the PWWP domain of the DNMT3A Gly332Arg mutant, the position of the Arg lies opposite to the Phe303 residue and could form an additional side to the conserved aromatic cage formed by the amino acids Phe303, Trp306, Trp330 and Asp333. ('DNMT3A', 'Gene', (53, 59)) ('Trp306', 'Var', (237, 243)) ('Phe303', 'Chemical', '-', (229, 235)) ('age', 'Gene', (199, 202)) ('Gly332Arg', 'SUBSTITUTION', 'None', (60, 69)) ('Phe303', 'Var', (229, 235)) ('Gly332Arg', 'Var', (60, 69)) ('Asp333', 'Chemical', '-', (256, 262)) ('Trp330', 'Chemical', '-', (245, 251)) ('Phe303', 'Chemical', '-', (123, 129)) ('Arg', 'Chemical', 'MESH:D001120', (98, 101)) ('Trp306', 'Chemical', '-', (237, 243)) ('age', 'Gene', '5973', (199, 202)) ('Arg', 'Chemical', 'MESH:D001120', (66, 69)) 8563 33182397 Thus, it is possible that an Arg residue in the 332 position could promote the stabilization of the binding to the trimethyl lysine present in histone H3. ('trimethyl lysine', 'Chemical', 'MESH:C003712', (115, 131)) ('binding', 'Interaction', (100, 107)) ('Arg residue', 'Var', (29, 40)) ('Arg', 'Chemical', 'MESH:D001120', (29, 32)) ('stabilization', 'MPA', (79, 92)) ('trimethyl lysine', 'MPA', (115, 131)) ('promote', 'PosReg', (67, 74)) 8565 33182397 Hierarchical clustering grouped all the controls together and separated from the proband's sample that showed a recognizable methylated profile (Figure 3a), suggesting that the p.Gly332Arg DNMT3A variant produces a similar alteration in germline global methylation than the observed for the previously reported PPGL patients. ('p.Gly332Arg', 'Mutation', 'rs760854242', (177, 188)) ('PPGL', 'Disease', (311, 315)) ('PGL', 'Phenotype', 'HP:0002668', (312, 315)) ('PPGL', 'Chemical', '-', (311, 315)) ('germline global methylation', 'MPA', (237, 264)) ('DNMT3A', 'Gene', (189, 195)) ('patients', 'Species', '9606', (316, 324)) ('p.Gly332Arg', 'Var', (177, 188)) ('alteration', 'Reg', (223, 233)) 8566 33182397 We also speculated that the methylated profile observed upon alteration of DNMT3A in PPGL patients could be similar to the one described for patients with microcephalic dwarfism harboring germline gain-of-function DNMT3A variants. ('microcephalic dwarfism', 'Phenotype', 'HP:0008873', (155, 177)) ('alteration', 'Var', (61, 71)) ('DNMT3A', 'Gene', (75, 81)) ('microcephalic dwarfism', 'Disease', (155, 177)) ('microcephalic dwarfism', 'Disease', 'MESH:C537533', (155, 177)) ('methylated profile', 'MPA', (28, 46)) ('PPGL', 'Gene', (85, 89)) ('variants', 'Var', (221, 229)) ('PGL', 'Phenotype', 'HP:0002668', (86, 89)) ('PPGL', 'Chemical', '-', (85, 89)) ('dwarfism', 'Phenotype', 'HP:0003510', (169, 177)) ('patients', 'Species', '9606', (90, 98)) ('patients', 'Species', '9606', (141, 149)) ('gain-of-function', 'PosReg', (197, 213)) 8567 33182397 To explore this, we investigated the methylation status of the 307 differentially methylated probes in lymphocyte DNAs from the previously reported PGL DNMT3A variant carriers and in blood samples from patients with either overgrowth or dwarfism due to DNMT3A germline variants. ('investigated', 'Reg', (20, 32)) ('PGL DNMT3A', 'Gene', (148, 158)) ('overgrowth', 'CPA', (223, 233)) ('overgrowth', 'Phenotype', 'HP:0001548', (223, 233)) ('dwarfism', 'Phenotype', 'HP:0003510', (237, 245)) ('patients', 'Species', '9606', (202, 210)) ('PGL', 'Phenotype', 'HP:0002668', (148, 151)) ('dwarfism', 'CPA', (237, 245)) ('variant', 'Var', (159, 166)) 8571 33182397 In addition, we were able to collect an AML sample carrying the p.Gly332Arg DNMT3A variant and investigated whether this somatic substitution could also be causing a gain-of-function in AML. ('AML', 'Disease', 'MESH:D015470', (40, 43)) ('p.Gly332Arg', 'Var', (64, 75)) ('AML', 'Disease', 'MESH:D015470', (186, 189)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (64, 75)) ('AML', 'Phenotype', 'HP:0004808', (186, 189)) ('AML', 'Disease', (186, 189)) ('AML', 'Phenotype', 'HP:0004808', (40, 43)) ('AML', 'Disease', (40, 43)) ('DNMT3A', 'Gene', (76, 82)) ('gain-of-function', 'PosReg', (166, 182)) 8572 33182397 To explore this, we profiled one AML sample carrying the p.Gly332Arg DNMT3A variant (this sample also carries another DNMT3A variant and two frameshift variants in TET2), with AML samples extracted from TCGA and carriers of DNMT3A somatic variants known to cause global methylation alterations (i.e., the bona fide p.Arg882 DNMT3A loss-of-function alteration). ('Arg882', 'Chemical', '-', (317, 323)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (57, 68)) ('AML', 'Disease', 'MESH:D015470', (33, 36)) ('AML', 'Phenotype', 'HP:0004808', (176, 179)) ('AML', 'Disease', (176, 179)) ('AML', 'Disease', (33, 36)) ('p.Arg882', 'Var', (315, 323)) ('TET2', 'Gene', '54790', (164, 168)) ('AML', 'Phenotype', 'HP:0004808', (33, 36)) ('loss-of-function', 'NegReg', (331, 347)) ('DNMT3A', 'Gene', (69, 75)) ('AML', 'Disease', 'MESH:D015470', (176, 179)) ('p.Gly332Arg', 'Var', (57, 68)) ('methylation alterations', 'MPA', (270, 293)) ('TET2', 'Gene', (164, 168)) 8573 33182397 We also included three samples carrying additional frameshift TET2 variants. ('TET2', 'Gene', (62, 66)) ('frameshift', 'Var', (51, 61)) ('TET2', 'Gene', '54790', (62, 66)) 8574 33182397 The unsupervised clustering, using the aforementioned methylation signature, revealed that the p.Gly332Arg mutated AML showed a completely different profile to the one observed for the samples carrying the recurrent p.Arg882 DNMT3A inactivating variant (Figure S1). ('AML', 'Disease', 'MESH:D015470', (115, 118)) ('p.Gly332Arg', 'Var', (95, 106)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (95, 106)) ('AML', 'Phenotype', 'HP:0004808', (115, 118)) ('AML', 'Disease', (115, 118)) ('Arg882', 'Chemical', '-', (218, 224)) 8576 33182397 Overall, these results suggest that the p.Gly332Arg DNMT3A variant led, in germline and also in AML, to a characteristic methylation profile, distinct to the profile caused by well-known DNMT3A inactivating variants. ('AML', 'Disease', (96, 99)) ('p.Gly332Arg', 'Var', (40, 51)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (40, 51)) ('methylation profile', 'MPA', (121, 140)) ('DNMT3A', 'Gene', (52, 58)) ('AML', 'Disease', 'MESH:D015470', (96, 99)) ('AML', 'Phenotype', 'HP:0004808', (96, 99)) 8577 33182397 Although the sample carrying the p.Gly332Arg variant harbors an additional variant in the MTase domain of DNMT3A (p.Arg720Cys), its low allele frequency together with the fact that variants in this domain likely disrupt the catalytic activity of the enzyme, supports that the observed methylated profile is caused by the PWWP variant. ('disrupt', 'NegReg', (212, 219)) ('p.Gly332Arg', 'Var', (33, 44)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (33, 44)) ('p.Arg720Cys', 'Mutation', 'rs1197133406', (114, 125)) ('catalytic activity', 'MPA', (224, 242)) 8580 33182397 Later it was demonstrated that this was a consequence of oncometabolite accumulation, caused by pathogenic variants in Krebs cycle genes. ('Krebs', 'Chemical', '-', (119, 124)) ('Krebs', 'Gene', (119, 124)) ('variants', 'Var', (107, 115)) ('oncometabolite accumulation', 'MPA', (57, 84)) 8581 33182397 More recently, disrupting variants in ATRX and in other chromatin-remodeling genes, including post-zygotic H3F3A gain-of-function alterations, have also been found recurrently in PPGLs. ('gain-of-function', 'PosReg', (113, 129)) ('ATRX', 'Gene', (38, 42)) ('H3F3A', 'Gene', '3020', (107, 112)) ('alterations', 'Var', (130, 141)) ('PPGL', 'Chemical', '-', (179, 183)) ('PGL', 'Phenotype', 'HP:0002668', (180, 183)) ('ATRX', 'Gene', '546', (38, 42)) ('H3F3A', 'Gene', (107, 112)) ('PPGLs', 'Disease', (179, 184)) ('variants', 'Var', (26, 34)) 8582 33182397 The identification of DNMT3A activating variants leading to a specific CIMP further supported the association between epigenetic modifications and PPGL development. ('association', 'Interaction', (98, 109)) ('DNMT3A', 'Gene', (22, 28)) ('CIMP', 'Disease', (71, 75)) ('PPGL', 'Chemical', '-', (147, 151)) ('variants', 'Var', (40, 48)) ('PGL', 'Phenotype', 'HP:0002668', (148, 151)) ('leading to', 'Reg', (49, 59)) 8583 33182397 In addition to the variants reported in patients with PGLs, germline gain-of-function DNMT3A variants located in the PWWP domain which cause widespread DNA hypermethylation at polycomb-regulated regions (with the H3K27me3 mark) have been found in patients with microcephalic dwarfism, an extreme growth disorder. ('growth disorder', 'Disease', (296, 311)) ('PGL', 'Phenotype', 'HP:0002668', (54, 57)) ('microcephalic dwarfism', 'Phenotype', 'HP:0008873', (261, 283)) ('microcephalic dwarfism', 'Disease', 'MESH:C537533', (261, 283)) ('patients', 'Species', '9606', (40, 48)) ('dwarfism', 'Phenotype', 'HP:0003510', (275, 283)) ('microcephalic dwarfism', 'Disease', (261, 283)) ('variants', 'Var', (93, 101)) ('patients', 'Species', '9606', (247, 255)) ('growth disorder', 'Phenotype', 'HP:0001507', (296, 311)) ('gain-of-function', 'PosReg', (69, 85)) ('DNMT3A', 'Gene', (86, 92)) ('growth disorder', 'Disease', 'MESH:D006130', (296, 311)) 8584 33182397 A similar growth deficiency has been observed in mice carrying a DNMT3A PWWP germline variant in the Asp329 residue (corresponding to the human Asp333). ('variant in', 'Var', (86, 96)) ('human', 'Species', '9606', (138, 143)) ('growth deficiency', 'Phenotype', 'HP:0001510', (10, 27)) ('mice', 'Species', '10090', (49, 53)) ('growth deficiency', 'Disease', 'MESH:D006130', (10, 27)) ('Asp333', 'Chemical', '-', (144, 150)) ('Asp329', 'Chemical', '-', (101, 107)) ('growth deficiency', 'Disease', (10, 27)) 8585 33182397 This results in DNA hypermethylation and de-repression of developmental regulatory genes that manifests phenotypically as dominant postnatal growth retardation. ('growth retardation', 'Phenotype', 'HP:0001510', (141, 159)) ('postnatal growth retardation', 'Disease', (131, 159)) ('hypermethylation', 'Var', (20, 36)) ('de-repression', 'NegReg', (41, 54)) ('postnatal growth retardation', 'Phenotype', 'HP:0008897', (131, 159)) ('postnatal growth retardation', 'Disease', 'MESH:D006130', (131, 159)) ('developmental regulatory genes', 'Gene', (58, 88)) ('results in DNA', 'Reg', (5, 19)) 8586 33182397 In the same way, the murine equivalent to the human Lys299Ile found in patients with PGL, disrupts both DNA and H3K36me2/3 binding by altering the aromatic cage conformation of the PWWP domain of DNMT3A, finally leading to disruption of the sub-nuclear localization of DNMT3A. ('binding', 'Interaction', (123, 130)) ('altering', 'Reg', (134, 142)) ('H3K36me2/3', 'Protein', (112, 122)) ('PGL', 'Phenotype', 'HP:0002668', (85, 88)) ('Lys299Ile', 'Var', (52, 61)) ('patients', 'Species', '9606', (71, 79)) ('DNA', 'Protein', (104, 107)) ('age', 'Gene', (157, 160)) ('DNMT3A', 'Gene', (196, 202)) ('human', 'Species', '9606', (46, 51)) ('age', 'Gene', '5973', (157, 160)) ('Lys299Ile', 'SUBSTITUTION', 'None', (52, 61)) ('disrupts', 'NegReg', (90, 98)) ('sub-nuclear localization', 'MPA', (241, 265)) ('DNMT3A', 'Gene', (269, 275)) ('murine', 'Species', '10090', (21, 27)) ('disruption', 'Reg', (223, 233)) 8587 33182397 In PGL patients, we demonstrated that germline DNMT3A variants in residues within the PWWP domain caused significant hypermethylation of homeobox-containing genes involved in early embryonic development. ('hypermethylation', 'MPA', (117, 133)) ('DNMT3A', 'Gene', (47, 53)) ('homeobox-containing', 'Gene', (137, 156)) ('PGL', 'Phenotype', 'HP:0002668', (3, 6)) ('variants', 'Var', (54, 62)) ('patients', 'Species', '9606', (7, 15)) 8588 33182397 This contrasts with the widespread hypomethylation observed in TBRS patients carrying germline DNMT3A variants, especially enriched at genes involved in morphogenesis, development and malignancy predisposition pathways. ('variants', 'Var', (102, 110)) ('DNMT3A', 'Gene', (95, 101)) ('malignancy', 'Disease', 'MESH:D009369', (184, 194)) ('malignancy', 'Disease', (184, 194)) ('patients', 'Species', '9606', (68, 76)) 8590 33182397 The conservation of Gly332 within the PWWP domain and its location between Trp330 and Asp333, the two residues altered in patients with microcephalic dwarfism, highly suggest a disruptive role of the variant. ('Asp333', 'Chemical', '-', (86, 92)) ('dwarfism', 'Phenotype', 'HP:0003510', (150, 158)) ('patients', 'Species', '9606', (122, 130)) ('Trp330', 'Chemical', '-', (75, 81)) ('Gly332', 'Chemical', '-', (20, 26)) ('microcephalic dwarfism', 'Phenotype', 'HP:0008873', (136, 158)) ('microcephalic dwarfism', 'Disease', (136, 158)) ('microcephalic dwarfism', 'Disease', 'MESH:C537533', (136, 158)) ('Gly332', 'Var', (20, 26)) 8591 33182397 The substitution of the wild-type Gly332 by an Arg could stabilize the link with trimethylated histone H3 causing, as occurred with the previously reported variants found in PGL patients, a gain of alternative activities by DNMT3A. ('trimethylated histone', 'Chemical', '-', (81, 102)) ('stabilize', 'Reg', (57, 66)) ('PGL', 'Phenotype', 'HP:0002668', (174, 177)) ('link', 'Interaction', (71, 75)) ('alternative activities', 'MPA', (198, 220)) ('patients', 'Species', '9606', (178, 186)) ('DNMT3A', 'Enzyme', (224, 230)) ('Gly332 by an Arg', 'Mutation', 'rs760854242', (34, 50)) ('gain', 'PosReg', (190, 194)) ('PGL', 'Disease', (174, 177)) ('Gly332', 'Var', (34, 40)) 8593 33182397 Moreover, overall alterations of DNA methylation have been also observed in blood cells from TET2 (hypermethylation) or EZH2 (hypomethylation) germline pathogenic loss-of-function variant carriers. ('alterations', 'Reg', (18, 29)) ('EZH2', 'Gene', (120, 124)) ('variant', 'Var', (180, 187)) ('TET2', 'Gene', (93, 97)) ('loss-of-function', 'NegReg', (163, 179)) ('DNA', 'Gene', (33, 36)) ('TET2', 'Gene', '54790', (93, 97)) ('EZH2', 'Gene', '2146', (120, 124)) 8596 33182397 Interestingly, an AML pedigree carrying a germline DNMT3A loss-of-function mutation has no symptoms of TBRS and only three patients with TBRS have been reported to develop AML. ('AML', 'Disease', (18, 21)) ('AML', 'Phenotype', 'HP:0004808', (172, 175)) ('AML', 'Phenotype', 'HP:0004808', (18, 21)) ('mutation', 'Var', (75, 83)) ('patients', 'Species', '9606', (123, 131)) ('loss-of-function', 'NegReg', (58, 74)) ('AML', 'Disease', 'MESH:D015470', (172, 175)) ('DNMT3A', 'Gene', (51, 57)) ('AML', 'Disease', 'MESH:D015470', (18, 21)) ('AML', 'Disease', (172, 175)) 8597 33182397 Somatic alterations in the epigenetic regulators TET2 and DNMT3A represent a frequent cause of clonal hematopoiesis, an aging-related mechanism in which somatic variants in early blood cell progenitors confer an advantage to mutated clones. ('variants', 'Var', (161, 169)) ('age', 'Gene', (218, 221)) ('TET2', 'Gene', (49, 53)) ('alterations', 'Var', (8, 19)) ('hematopoiesis', 'Disease', 'MESH:C536227', (102, 115)) ('age', 'Gene', '5973', (218, 221)) ('DNMT3A', 'Gene', (58, 64)) ('cause', 'Reg', (86, 91)) ('TET2', 'Gene', '54790', (49, 53)) ('hematopoiesis', 'Disease', (102, 115)) 8599 33182397 Interestingly, several variants affecting the Gly332 residue (including p.Gly332Arg and p.Gly332Glu) have been found in healthy individuals with clonal hematopoietic mutations. ('found', 'Reg', (111, 116)) ('Gly332', 'Chemical', '-', (90, 96)) ('p.Gly332Glu', 'Var', (88, 99)) ('Gly332', 'Chemical', '-', (46, 52)) ('Gly332', 'Chemical', '-', (74, 80)) ('p.Gly332Arg', 'Var', (72, 83)) ('p.Gly332Glu', 'Mutation', 'rs751360082', (88, 99)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (72, 83)) 8600 33182397 In fact, the Gly332Arg variant is included in a list of leukemogenic driver mutations affecting genes known to promote clonal expansion of hematopoietic stem cells (Table S2). ('mutations', 'Var', (76, 85)) ('leukemogenic', 'Disease', (56, 68)) ('Gly332Arg', 'SUBSTITUTION', 'None', (13, 22)) ('Gly332Arg', 'Var', (13, 22)) ('promote', 'PosReg', (111, 118)) 8601 33182397 It has been suggested that the presence of some pathogenic TBRS variants amongst the general population is due to age-related clonal hematopoiesis, questioning the utility of databases such as gnomAD in DNMT3A variant pathogenicity stratification. ('age', 'Gene', '5973', (114, 117)) ('hematopoiesis', 'Disease', 'MESH:C536227', (133, 146)) ('TBRS', 'Gene', (59, 63)) ('age', 'Gene', (114, 117)) ('hematopoiesis', 'Disease', (133, 146)) ('pathogenic', 'Reg', (48, 58)) ('variants', 'Var', (64, 72)) 8602 33182397 The methylated profile observed in the AML sample carrying the p.Gly332Arg variant further suggests that alteration of specific residues within the PWWP domain of DNMT3A leading to activation of its methylation capacities, may have oncogenic potential to both endocrine malignancies (PPGL) and AML. ('methylation capacities', 'MPA', (199, 221)) ('alteration', 'Var', (105, 115)) ('AML', 'Disease', 'MESH:D015470', (39, 42)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (63, 74)) ('endocrine malignancies', 'Disease', (260, 282)) ('PPGL', 'Chemical', '-', (284, 288)) ('AML', 'Disease', 'MESH:D015470', (294, 297)) ('endocrine malignancies', 'Disease', 'MESH:D004701', (260, 282)) ('endocrine malignancies', 'Phenotype', 'HP:0100568', (260, 282)) ('AML', 'Phenotype', 'HP:0004808', (294, 297)) ('AML', 'Disease', (39, 42)) ('activation', 'PosReg', (181, 191)) ('AML', 'Disease', (294, 297)) ('p.Gly332Arg', 'Var', (63, 74)) ('DNMT3A', 'Gene', (163, 169)) ('PGL', 'Phenotype', 'HP:0002668', (285, 288)) ('AML', 'Phenotype', 'HP:0004808', (39, 42)) ('oncogenic potential', 'CPA', (232, 251)) 8606 33182397 Moreover, COSMIC reports only one DNMT3A variant affecting the microcephalic dwarfism-mutated residue Asp333 and it affects an anaplastic thyroid carcinoma, further supporting the relevance of the PWWP domain in thyroid cancer. ('dwarfism', 'Phenotype', 'HP:0003510', (77, 85)) ('anaplastic thyroid carcinoma', 'Phenotype', 'HP:0011779', (127, 155)) ('thyroid carcinoma', 'Disease', 'MESH:D013964', (138, 155)) ('variant', 'Var', (41, 48)) ('microcephalic dwarfism', 'Disease', 'MESH:C537533', (63, 85)) ('thyroid carcinoma', 'Disease', (138, 155)) ('thyroid cancer', 'Disease', 'MESH:D013964', (212, 226)) ('affects', 'Reg', (116, 123)) ('DNMT3A', 'Gene', (34, 40)) ('thyroid cancer', 'Phenotype', 'HP:0002890', (212, 226)) ('thyroid carcinoma', 'Phenotype', 'HP:0002890', (138, 155)) ('microcephalic dwarfism', 'Phenotype', 'HP:0008873', (63, 85)) ('Asp333', 'MPA', (102, 108)) ('carcinoma', 'Phenotype', 'HP:0030731', (146, 155)) ('cancer', 'Phenotype', 'HP:0002664', (220, 226)) ('microcephalic dwarfism', 'Disease', (63, 85)) ('Asp333', 'Chemical', '-', (102, 108)) ('affecting', 'Reg', (49, 58)) ('thyroid cancer', 'Disease', (212, 226)) 8608 33182397 In summary, we describe and perform methylome analysis for a novel DNMT3A activating variant in a patient with multiple clinical features. ('activating', 'PosReg', (74, 84)) ('DNMT3A', 'Gene', (67, 73)) ('variant', 'Var', (85, 92)) ('patient', 'Species', '9606', (98, 105)) 8609 33182397 Although the causal role of DNMT3A variants in PPGL has not been demonstrated, our results stress the heterogeneous phenotypic spectrum related to DNMT3A germline variants and support a dual nature of this gene in disease. ('variants', 'Var', (35, 43)) ('PPGL', 'Chemical', '-', (47, 51)) ('PPGL', 'Gene', (47, 51)) ('DNMT3A', 'Gene', (147, 153)) ('PGL', 'Phenotype', 'HP:0002668', (48, 51)) 8615 33182397 Tumor DNA from a patient with AML and carrying two somatic variants in DNMT3A (p.Gly332Arg and p.Arg720Cys) and two in TET2 (p.Arg1216Ter and p.Arg544Ter) was used for methylation profiling. ('Tumor', 'Phenotype', 'HP:0002664', (0, 5)) ('DNMT3A', 'Gene', (71, 77)) ('p.Arg720Cys', 'Var', (95, 106)) ('AML', 'Disease', 'MESH:D015470', (30, 33)) ('p.Arg544Ter', 'Mutation', 'rs1440692352', (142, 153)) ('TET2', 'Gene', '54790', (119, 123)) ('patient', 'Species', '9606', (17, 24)) ('p.Arg544Ter', 'Var', (142, 153)) ('AML', 'Disease', (30, 33)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (79, 90)) ('AML', 'Phenotype', 'HP:0004808', (30, 33)) ('p.Arg1216Ter', 'Var', (125, 137)) ('p.Gly332Arg', 'Var', (79, 90)) ('p.Arg720Cys', 'Mutation', 'rs1197133406', (95, 106)) ('TET2', 'Gene', (119, 123)) ('p.Arg1216Ter', 'Mutation', 'rs1009194427', (125, 137)) 8618 33182397 Filtering was applied to exclude variants present in gnomAD with AF > 0.00005, variants present in internal exomes or in the CIBERER Spanish Variant Server , noncoding substitutions and variants in genomic regions with low mappability, with low depth readings or quality or with the alternative allele present in < 20% of reads. ('substitutions', 'Var', (168, 181)) ('mappability', 'MPA', (223, 234)) ('variants', 'Var', (79, 87)) ('AF', 'Disease', 'MESH:D001281', (65, 67)) ('variants', 'Var', (186, 194)) 8619 33182397 The PredictSNP1 consensus classifier was used to predict the effect of the only nucleotide substitution affecting a gene previously associated with PPGL and that passed all filtering steps. ('PredictSNP1', 'Gene', '6625', (4, 15)) ('nucleotide substitution', 'Var', (80, 103)) ('PredictSNP1', 'Gene', (4, 15)) ('PGL', 'Phenotype', 'HP:0002668', (149, 152)) ('PPGL', 'Chemical', '-', (148, 152)) 8623 33182397 The H3K36me3 peptide was modelled into the DNMT3A PWWP Gly332Arg mutant structure obtained using the DNMT3B PWWP domain structure bound to H3K36me3 peptide (PDB code 5CIU) as a reference to ensure no clashes with the methylated lysine. ('Gly332Arg', 'SUBSTITUTION', 'None', (55, 64)) ('Gly332Arg', 'Var', (55, 64)) ('DNMT3B', 'Gene', '1789', (101, 107)) ('DNMT3B', 'Gene', (101, 107)) ('lysine', 'Chemical', 'MESH:D008239', (228, 234)) 8627 33182397 In a subsequent analysis, we used the same list of 307 probes to profile methylation data from blood of patients with microcephalic dwarfism (n = 1) or overgrowth syndrome (n = 2) (GSE120428) and compare them to controls (n = 2) and to PGLs carrying DNMT3A variants (n = 2). ('variants', 'Var', (257, 265)) ('methylation', 'MPA', (73, 84)) ('microcephalic dwarfism', 'Phenotype', 'HP:0008873', (118, 140)) ('microcephalic dwarfism', 'Disease', (118, 140)) ('microcephalic dwarfism', 'Disease', 'MESH:C537533', (118, 140)) ('overgrowth syndrome', 'Disease', 'MESH:D001765', (152, 171)) ('dwarfism', 'Phenotype', 'HP:0003510', (132, 140)) ('overgrowth syndrome', 'Disease', (152, 171)) ('overgrowth', 'Phenotype', 'HP:0001548', (152, 162)) ('patients', 'Species', '9606', (104, 112)) ('PGL', 'Phenotype', 'HP:0002668', (236, 239)) 8628 33182397 Finally, we extracted methylation data from 12 AML samples available from The Cancer Genome Atlas (TCGA), corresponding to 213 out of the 307 probes aforementioned for hierarchical clustering with one additional AML sample carrying the p.Gly332Arg variant, another DNMT3A variant (p.Arg720Cys) and two frameshift TET2 mutations. ('p.Gly332Arg', 'Mutation', 'rs760854242', (236, 247)) ('TET2', 'Gene', (313, 317)) ('Cancer', 'Disease', (78, 84)) ('p.Arg720Cys', 'Mutation', 'rs1197133406', (281, 292)) ('Cancer', 'Disease', 'MESH:D009369', (78, 84)) ('Cancer', 'Phenotype', 'HP:0002664', (78, 84)) ('AML', 'Disease', 'MESH:D015470', (47, 50)) ('AML', 'Disease', 'MESH:D015470', (212, 215)) ('AML', 'Disease', (47, 50)) ('TET2', 'Gene', '54790', (313, 317)) ('AML', 'Phenotype', 'HP:0004808', (47, 50)) ('AML', 'Phenotype', 'HP:0004808', (212, 215)) ('p.Gly332Arg', 'Var', (236, 247)) ('AML', 'Disease', (212, 215)) 8629 33182397 The TCGA samples were all carriers of mutations in DNMT3A known to cause global methylation alterations (i.e., p.Arg882Cys/His) and three of them also harbored TET2 frameshift mutations. ('cause', 'Reg', (67, 72)) ('TET2', 'Gene', '54790', (160, 164)) ('His', 'Chemical', 'MESH:D006639', (123, 126)) ('TET2', 'Gene', (160, 164)) ('p.Arg882Cys', 'Var', (111, 122)) ('mutations', 'Var', (38, 47)) ('DNMT3A', 'Gene', (51, 57)) ('global methylation alterations', 'MPA', (73, 103)) ('p.Arg882Cys', 'SUBSTITUTION', 'None', (111, 122)) 8630 33182397 We describe a novel germline DNMT3A variant (p.Gly332Arg) identified by whole-exome sequencing in a patient with multiple clinical features: bilateral carotid PGLs, PTC and idiopathic intellectual disability. ('intellectual disability', 'Phenotype', 'HP:0001249', (184, 207)) ('p.Gly332Arg', 'Var', (45, 56)) ('patient', 'Species', '9606', (100, 107)) ('PGL', 'Phenotype', 'HP:0002668', (159, 162)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (45, 56)) ('PTC', 'Phenotype', 'HP:0002895', (165, 168)) ('DNMT3A', 'Gene', (29, 35)) ('PTC', 'Disease', (165, 168)) 8631 33182397 The variant affects a residue found mutated in papillary thyroid tumors and the structural modelling of the PWWP domain predicts the substitution to alter the interaction of DNMT3A with H3K36me3. ('alter', 'Reg', (149, 154)) ('papillary thyroid tumors', 'Disease', (47, 71)) ('substitution', 'Var', (133, 145)) ('papillary thyroid tumors', 'Phenotype', 'HP:0002895', (47, 71)) ('interaction', 'Interaction', (159, 170)) ('tumors', 'Phenotype', 'HP:0002664', (65, 71)) ('DNMT3A', 'Protein', (174, 180)) ('variant', 'Var', (4, 11)) ('papillary thyroid tumors', 'Disease', 'MESH:D000077273', (47, 71)) ('H3K36me3', 'Protein', (186, 194)) ('tumor', 'Phenotype', 'HP:0002664', (65, 70)) ('affects', 'Reg', (12, 19)) 8632 33182397 Moreover, we observed an increased methylation of DNMT3A target genes in saliva DNA from the proband and in one independent AML sample carrying the same p.Gly332Arg variant, compatible with a gain-of-function effect of the alteration. ('DNMT3A', 'Gene', (50, 56)) ('AML', 'Phenotype', 'HP:0004808', (124, 127)) ('increased', 'PosReg', (25, 34)) ('AML', 'Disease', (124, 127)) ('methylation', 'MPA', (35, 46)) ('AML', 'Disease', 'MESH:D015470', (124, 127)) ('p.Gly332Arg', 'Var', (153, 164)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (153, 164)) 8633 33182397 Although the causal role of DNMT3A variants in PPGL has not been demonstrated, our finding stresses the heterogeneous phenotypic spectrum related to DNMT3A germline variants. ('PGL', 'Phenotype', 'HP:0002668', (48, 51)) ('PPGL', 'Chemical', '-', (47, 51)) ('PPGL', 'Gene', (47, 51)) ('variants', 'Var', (35, 43)) 8634 33182397 The following are available online at , Figure S1: Hierarchical clustering performed with one AML sample carrying the p.Gly332Arg DNMT3A mutation (black circle) and 12 AML samples from TCGA carrying mutations known to cause global methylation alterations: loss-of-function p.Arg882Cys/His DNMT3A mutations (blue boxes) and TET2 frameshift mutations (purple boxes). ('AML', 'Disease', (94, 97)) ('TET2', 'Gene', (323, 327)) ('AML', 'Phenotype', 'HP:0004808', (94, 97)) ('p.Arg882Cys', 'SUBSTITUTION', 'None', (273, 284)) ('p.Arg882Cys', 'Var', (273, 284)) ('His', 'Chemical', 'MESH:D006639', (285, 288)) ('AML', 'Disease', 'MESH:D015470', (168, 171)) ('AML', 'Phenotype', 'HP:0004808', (168, 171)) ('loss-of-function', 'NegReg', (256, 272)) ('DNMT3A', 'Gene', (289, 295)) ('AML', 'Disease', (168, 171)) ('AML', 'Disease', 'MESH:D015470', (94, 97)) ('p.Gly332Arg', 'Var', (118, 129)) ('TET2', 'Gene', '54790', (323, 327)) ('p.Gly332Arg', 'Mutation', 'rs760854242', (118, 129)) 8635 33182397 Profiling was based on methylation data from the 213 probes, out of the 307 significantly differentially methylated between DNMT3A-mutated and non-mutated tissues in patients with PPGL, contained in the Infinium HumanMethylation450 BeadChip, Figure S2: Schematic representation of the methylation analysis pipeline, Table S1: Filtered exome variants found in ClinVar, Table S2: Variants affecting the PWWP domain found in gnomAD, Table S3: List of differentially methylated CpGs between DNMT3A-mutated and non-mutated samples. ('PPGL', 'Chemical', '-', (180, 184)) ('Variants', 'Var', (378, 386)) ('PGL', 'Phenotype', 'HP:0002668', (181, 184)) ('variants', 'Var', (341, 349)) ('Pro', 'Chemical', 'MESH:D011392', (0, 3)) ('patients', 'Species', '9606', (166, 174)) 8643 32099035 Epigenetic regulation of CD141Hi cDC differentiation offers an intriguing opportunity for intervention and a potential adjunct to immunotherapy for patients with MDS. ('patients', 'Species', '9606', (148, 156)) ('MDS', 'Disease', (162, 165)) ('MDS', 'Disease', 'MESH:D009190', (162, 165)) ('CD141Hi cDC', 'Gene', (25, 36)) ('Epigenetic regulation', 'Var', (0, 21)) ('CD141Hi cDC', 'Gene', '7056', (25, 36)) 8667 32099035 Finally, we rescued differentiation of CD141Hi cDCs from MDS progenitors using pharmacologic inhibition of Lysine-Specific Demethylase 1A (LSD1). ('CD141Hi cDC', 'Gene', (39, 50)) ('Lysine-Specific Demethylase 1A', 'Gene', (107, 137)) ('CD141Hi cDC', 'Gene', '7056', (39, 50)) ('Lysine-Specific Demethylase 1A', 'Gene', '23028', (107, 137)) ('MDS', 'Disease', (57, 60)) ('MDS', 'Disease', 'MESH:D009190', (57, 60)) ('inhibition', 'Var', (93, 103)) 8696 32099035 Our data mirror outcome studies in patients with solid tumors and suggest that deficiency of CD141Hi cDCs may contribute to inferior outcome in MDS patients. ('MDS', 'Disease', (144, 147)) ('patients', 'Species', '9606', (148, 156)) ('MDS', 'Disease', 'MESH:D009190', (144, 147)) ('tumor', 'Phenotype', 'HP:0002664', (55, 60)) ('deficiency', 'Var', (79, 89)) ('solid tumors', 'Disease', (49, 61)) ('tumors', 'Phenotype', 'HP:0002664', (55, 61)) ('patients', 'Species', '9606', (35, 43)) ('solid tumors', 'Disease', 'MESH:D009369', (49, 61)) ('CD141Hi cDC', 'Gene', (93, 104)) ('CD141Hi cDC', 'Gene', '7056', (93, 104)) 8718 32099035 MDS patients with IRF8Lo MDPs produced fewer CDPs and CD141Hi cDCs compared to those with MDPs expressing higher levels of IRF8 (IRF8Hi; Figure 2G). ('IRF8Lo', 'Var', (18, 24)) ('CD141Hi cDC', 'Gene', (54, 65)) ('MDPs', 'Chemical', '-', (90, 94)) ('IRF8Hi', 'Gene', (129, 135)) ('CDPs', 'CPA', (45, 49)) ('fewer', 'NegReg', (39, 44)) ('CD141Hi cDC', 'Gene', '7056', (54, 65)) ('MDPs', 'Chemical', '-', (25, 29)) ('patients', 'Species', '9606', (4, 12)) ('IRF8Hi', 'Gene', '3394', (129, 135)) ('MDS', 'Disease', (0, 3)) ('MDS', 'Disease', 'MESH:D009190', (0, 3)) ('CDPs', 'Chemical', 'MESH:D003565', (45, 49)) 8721 32099035 These results suggest that induction of IRF8 expression might enhance DC differentiation of MDS progenitors offering a strategy to increase the number of CD141Hi cDCs in patients with MDS. ('patients', 'Species', '9606', (170, 178)) ('CD141Hi cDC', 'Gene', (154, 165)) ('enhance', 'PosReg', (62, 69)) ('IRF8', 'Gene', (40, 44)) ('induction', 'Var', (27, 36)) ('CD141Hi cDC', 'Gene', '7056', (154, 165)) ('MDS', 'Disease', (184, 187)) ('MDS', 'Disease', 'MESH:D009190', (184, 187)) ('DC', 'Gene', '13179', (70, 72)) ('DC', 'Gene', '13179', (163, 165)) ('MDS', 'Disease', (92, 95)) ('MDS', 'Disease', 'MESH:D009190', (92, 95)) 8722 32099035 Inhibition of LSD1 induces differentiation of myeloid leukemia cells and several clinical trials are currently underway to test the efficacy of LSD1 inhibitors in myeloid malignancy. ('myeloid malignancy', 'Disease', (163, 181)) ('LSD1', 'Gene', (14, 18)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (46, 62)) ('myeloid leukemia', 'Disease', (46, 62)) ('Inhibition', 'Var', (0, 10)) ('myeloid malignancy', 'Disease', 'MESH:D009369', (163, 181)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (46, 62)) ('leukemia', 'Phenotype', 'HP:0001909', (54, 62)) ('induces', 'Reg', (19, 26)) ('differentiation', 'CPA', (27, 42)) 8724 32099035 We hypothesized that inhibition of LSD1 in HD and MDS CD34+ progenitors would promote CD141Hi cDC differentiation. ('CD34', 'Gene', (54, 58)) ('promote', 'PosReg', (78, 85)) ('CD34', 'Gene', '947', (54, 58)) ('HD', 'Disease', 'MESH:D006816', (43, 45)) ('LSD1', 'Gene', (35, 39)) ('MDS', 'Disease', (50, 53)) ('MDS', 'Disease', 'MESH:D009190', (50, 53)) ('inhibition', 'Var', (21, 31)) ('CD141Hi cDC', 'Gene', (86, 97)) ('CD141Hi cDC', 'Gene', '7056', (86, 97)) 8725 32099035 We tested this hypothesis using an in vitro model of DC differentiation and used two previously described compounds, GSK2879552 (GSK) and ORY-1001 (ORY), as pharmacologic tools to inhibit LSD1 activity. ('DC', 'Gene', '13179', (53, 55)) ('GSK2879552', 'Chemical', 'MESH:C000602008', (117, 127)) ('ORY-1001', 'Var', (138, 146)) ('GSK2879552', 'Var', (117, 127)) ('inhibit', 'NegReg', (180, 187)) ('GSK', 'Chemical', 'MESH:C000602008', (117, 120)) ('activity', 'MPA', (193, 201)) ('tested', 'Reg', (3, 9)) ('ORY-1001', 'Chemical', '-', (138, 146)) ('GSK', 'Chemical', 'MESH:C000602008', (129, 132)) ('LSD1', 'Enzyme', (188, 192)) 8737 32099035 Together, these data suggest that inhibition of LSD1 can promote the differentiation of mature CD141Hi DCs from progenitors derived from both HD and patients with MDS. ('LSD1', 'Gene', (48, 52)) ('patients', 'Species', '9606', (149, 157)) ('differentiation', 'CPA', (69, 84)) ('MDS', 'Disease', (163, 166)) ('promote', 'PosReg', (57, 64)) ('inhibition', 'Var', (34, 44)) ('HD', 'Disease', 'MESH:D006816', (142, 144)) ('MDS', 'Disease', 'MESH:D009190', (163, 166)) ('DC', 'Gene', '13179', (103, 105)) ('CD141Hi', 'Gene', (95, 102)) 8748 32099035 Together, these data suggest that LSD1 inhibition can increase expression of IRF8 in primary MDS specimens, but in some cases, this may not be sufficient to induce CD141Hi cDC differentiation. ('inhibition', 'Var', (39, 49)) ('induce', 'Reg', (157, 163)) ('CD141Hi cDC', 'Gene', (164, 175)) ('MDS', 'Disease', (93, 96)) ('MDS', 'Disease', 'MESH:D009190', (93, 96)) ('IRF8', 'Gene', (77, 81)) ('increase', 'PosReg', (54, 62)) ('expression', 'MPA', (63, 73)) ('CD141Hi cDC', 'Gene', '7056', (164, 175)) ('LSD1', 'Gene', (34, 38)) 8750 32099035 The role of IRF8 in cDC differentiation is conserved between human and mouse and IRF8 expression is necessary for differentiation of CD24+ cDCs, the mouse homolog of CD141Hi cDCs. ('DC', 'Gene', '13179', (140, 142)) ('CD141Hi cDC', 'Gene', '7056', (166, 177)) ('DC', 'Gene', '13179', (21, 23)) ('IRF8', 'Gene', (81, 85)) ('DC', 'Gene', '13179', (175, 177)) ('mouse', 'Species', '10090', (149, 154)) ('CD24+', 'Var', (133, 138)) ('human', 'Species', '9606', (61, 66)) ('mouse', 'Species', '10090', (71, 76)) ('CD141Hi cDC', 'Gene', (166, 177)) 8754 32099035 As expected, treatment of WT c-kit+ cells with ORY resulted in an increased number of CD24+ cDCs following our in vitro differentiation experiment (Figures 5A and 5B). ('DC', 'Gene', '13179', (93, 95)) ('increased', 'PosReg', (66, 75)) ('ORY', 'Var', (47, 50)) 8763 32099035 We know less about how defects in immune activating populations impact MDS progression and development. ('development', 'CPA', (91, 102)) ('MDS', 'Disease', (71, 74)) ('MDS', 'Disease', 'MESH:D009190', (71, 74)) ('impact', 'Reg', (64, 70)) ('defects', 'Var', (23, 30)) 8769 32099035 In addition, we have found that rationally designed epigenetic therapy can improve differentiation of CD141Hi cDCs from MDS progenitors. ('differentiation', 'CPA', (83, 98)) ('CD141Hi cDC', 'Gene', (102, 113)) ('CD141Hi cDC', 'Gene', '7056', (102, 113)) ('improve', 'PosReg', (75, 82)) ('epigenetic therapy', 'Var', (52, 70)) ('MDS', 'Disease', (120, 123)) ('MDS', 'Disease', 'MESH:D009190', (120, 123)) 8782 32099035 These data support prior work demonstrating that LSD1 inhibition drives differentiation programs in myeloid leukemia cells. ('inhibition', 'Var', (54, 64)) ('myeloid leukemia', 'Disease', (100, 116)) ('differentiation programs', 'CPA', (72, 96)) ('leukemia', 'Phenotype', 'HP:0001909', (108, 116)) ('myeloid leukemia', 'Disease', 'MESH:D007951', (100, 116)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (100, 116)) ('LSD1', 'Gene', (49, 53)) 8783 32099035 We confirm prior reports that LSD1 inhibition enhances IRF8 expression by modulating local chromatin structure but it is possible that LSD1 inhibition may also interact with IRF8 at other loci to regulate the chromatin structure during DC lineage differentiation. ('enhances', 'PosReg', (46, 54)) ('expression', 'MPA', (60, 70)) ('local chromatin structure', 'MPA', (85, 110)) ('chromatin structure', 'MPA', (209, 228)) ('interact', 'Reg', (160, 168)) ('DC', 'Gene', '13179', (236, 238)) ('regulate', 'Reg', (196, 204)) ('inhibition', 'Var', (35, 45)) ('IRF8', 'Gene', (55, 59)) ('modulating', 'Reg', (74, 84)) 8788 32099035 These investigators also showed that TET2 mutations were associated with the greatest response to the combination of LSD1 inhibition and 5-azacitidine, suggesting a hypothesis that the response to LSD1 inhibition is partly regulated by the MDS mutational landscape. ('5-azacitidine', 'Chemical', 'MESH:D001374', (137, 150)) ('MDS', 'Disease', (240, 243)) ('MDS', 'Disease', 'MESH:D009190', (240, 243)) ('TET2', 'Gene', '54790', (37, 41)) ('TET2', 'Gene', (37, 41)) ('mutations', 'Var', (42, 51)) 8793 32099035 Pre-clinically, LSD1 inhibition has already been demonstrated to improve the efficacy of anti-PD-1 therapy. ('PD-1', 'Gene', '6622', (94, 98)) ('efficacy', 'MPA', (77, 85)) ('improve', 'PosReg', (65, 72)) ('LSD1', 'Gene', (16, 20)) ('inhibition', 'Var', (21, 31)) ('PD-1', 'Gene', (94, 98)) 8799 31203423 Acute Myeloid Leukemia developed in 36.7% of NHD13 mice with a Bok gene knockout between the age of 8 and 13 months and presented a similar overall survival to the NHD13 mice. ('knockout', 'Var', (72, 80)) ('Bok', 'Gene', (63, 66)) ('Acute Myeloid Leukemia', 'Disease', (0, 22)) ('mice', 'Species', '10090', (51, 55)) ('Bok', 'Gene', '51800', (63, 66)) ('Myeloid Leukemia', 'Phenotype', 'HP:0012324', (6, 22)) ('Acute Myeloid Leukemia', 'Disease', 'MESH:D015470', (0, 22)) ('Acute Myeloid Leukemia', 'Phenotype', 'HP:0004808', (0, 22)) ('mice', 'Species', '10090', (170, 174)) ('Leukemia', 'Phenotype', 'HP:0001909', (14, 22)) 8800 31203423 The loss of BOK exacerbated anemia in NHD13 mice, and NHD13/BOK-deficient mice exhibited significantly lower hemoglobin, lower mean cell hemoglobin concentration, and higher mean cell volume than NHD13 mice. ('higher', 'PosReg', (167, 173)) ('anemia', 'Phenotype', 'HP:0001903', (28, 34)) ('mice', 'Species', '10090', (202, 206)) ('BOK', 'Gene', (12, 15)) ('exacerbated', 'PosReg', (16, 27)) ('anemia', 'Disease', 'MESH:D000740', (28, 34)) ('lower', 'NegReg', (103, 108)) ('lower', 'NegReg', (121, 126)) ('lower hemoglobin', 'Phenotype', 'HP:0001903', (103, 119)) ('anemia', 'Disease', (28, 34)) ('hemoglobin', 'MPA', (109, 119)) ('mean cell volume', 'MPA', (174, 190)) ('mean cell hemoglobin concentration', 'MPA', (127, 161)) ('mice', 'Species', '10090', (74, 78)) ('mice', 'Species', '10090', (44, 48)) ('lower mean cell hemoglobin concentration', 'Phenotype', 'HP:0025547', (121, 161)) ('loss', 'Var', (4, 8)) 8805 31203423 BCL-2, the BCL-2 family member PUMA, and the PUMA transcriptional regulator p53, which is mutated in 814% of MDS, all contribute to the antecedent apoptosis and subsequent development of AML. ('contribute', 'Reg', (118, 128)) ('p53', 'Gene', (76, 79)) ('BCL-2', 'Gene', '12043', (11, 16)) ('BCL-2', 'Gene', (11, 16)) ('BCL-2', 'Gene', '12043', (0, 5)) ('BCL-2', 'Gene', (0, 5)) ('MDS', 'Phenotype', 'HP:0002863', (109, 112)) ('PUMA', 'Gene', '170770', (45, 49)) ('mutated', 'Var', (90, 97)) ('PUMA', 'Gene', '170770', (31, 35)) ('p53', 'Gene', '22059', (76, 79)) ('MDS', 'Disease', 'MESH:D009190', (109, 112)) ('apoptosis', 'CPA', (147, 156)) ('AML', 'Disease', 'MESH:D015470', (187, 190)) ('PUMA', 'Gene', (45, 49)) ('PUMA', 'Gene', (31, 35)) ('AML', 'Phenotype', 'HP:0004808', (187, 190)) ('AML', 'Disease', (187, 190)) ('MDS', 'Disease', (109, 112)) 8810 31203423 In agreement, the induction of CHOP during diethylnitrosamine-induced hepatocarcinogenesis was also greatly attenuated by the loss of BOK. ('hepatocarcinogenesis', 'Disease', 'MESH:D063646', (70, 90)) ('attenuated', 'NegReg', (108, 118)) ('hepatocarcinogenesis', 'Disease', (70, 90)) ('CHOP', 'Gene', '13198', (31, 35)) ('BOK', 'Gene', (134, 137)) ('diethylnitrosamine-induced', 'Disease', (43, 69)) ('diethylnitrosamine', 'Chemical', 'MESH:D004052', (43, 61)) ('CHOP', 'Gene', (31, 35)) ('loss', 'Var', (126, 130)) 8817 31203423 Approximately onethird of NHD13 mice progress to AML within 10-14 months with high frequency acquisition of NRas, Kras, or Cbl mutations. ('AML', 'Disease', (49, 52)) ('Kras', 'Gene', '16653', (114, 118)) ('AML', 'Disease', 'MESH:D015470', (49, 52)) ('NRas', 'Gene', '18176', (108, 112)) ('mice', 'Species', '10090', (32, 36)) ('NHD13', 'Gene', (26, 31)) ('Cbl', 'Gene', '12402', (123, 126)) ('Cbl', 'Gene', (123, 126)) ('mutations', 'Var', (127, 136)) ('NRas', 'Gene', (108, 112)) ('Kras', 'Gene', (114, 118)) ('AML', 'Phenotype', 'HP:0004808', (49, 52)) 8818 31203423 In this model, both overexpression of anti-apoptotic BCL-2 and deletion of pro-apoptotic PUMA rescue cytopenias, but surprisingly delay progression to AML. ('cytopenias', 'Disease', 'MESH:D006402', (101, 111)) ('PUMA', 'Gene', '170770', (89, 93)) ('BCL-2', 'Gene', (53, 58)) ('cytopenias', 'Disease', (101, 111)) ('delay', 'NegReg', (130, 135)) ('rescue', 'PosReg', (94, 100)) ('overexpression', 'PosReg', (20, 34)) ('AML', 'Disease', 'MESH:D015470', (151, 154)) ('deletion', 'Var', (63, 71)) ('AML', 'Disease', (151, 154)) ('BCL-2', 'Gene', '12043', (53, 58)) ('PUMA', 'Gene', (89, 93)) ('AML', 'Phenotype', 'HP:0004808', (151, 154)) 8819 31203423 We hypothesized that loss of BOK may delay progression to AML in NHD13 mice similar to the aforementioned overexpression of BCL-2 and deletion of PUMA. ('loss', 'Var', (21, 25)) ('mice', 'Species', '10090', (71, 75)) ('PUMA', 'Gene', (146, 150)) ('AML', 'Disease', (58, 61)) ('PUMA', 'Gene', '170770', (146, 150)) ('progression', 'MPA', (43, 54)) ('AML', 'Disease', 'MESH:D015470', (58, 61)) ('BCL-2', 'Gene', '12043', (124, 129)) ('BOK', 'Gene', (29, 32)) ('BCL-2', 'Gene', (124, 129)) ('AML', 'Phenotype', 'HP:0004808', (58, 61)) ('delay', 'NegReg', (37, 42)) 8820 31203423 Notwithstanding, we found that loss of BOK does not appear to affect the development of AML in the NHD13 mice. ('AML', 'Disease', 'MESH:D015470', (88, 91)) ('loss', 'Var', (31, 35)) ('AML', 'Disease', (88, 91)) ('BOK', 'Gene', (39, 42)) ('AML', 'Phenotype', 'HP:0004808', (88, 91)) ('mice', 'Species', '10090', (105, 109)) 8826 31203423 For genotyping, loss of Bok was routinely confirmed by PCR as previously described and NHD13 transgene was also routinely confirmed with PCR procedure following the Jackson Laboratory's instruction. ('Bok', 'Gene', (24, 27)) ('Bok', 'Gene', '51800', (24, 27)) ('loss', 'Var', (16, 20)) ('NHD13', 'Gene', (87, 92)) 8852 31203423 Median survival of NHD13/Bok-/- and NHD13 mice were 355 and 340 days, respectively, which was not statistically significant by the Log-rank or Gehan-Breslow-Wilcoxon tests. ('Bok', 'Gene', '51800', (25, 28)) ('mice', 'Species', '10090', (42, 46)) ('NHD13', 'Var', (36, 41)) ('Bok', 'Gene', (25, 28)) 8853 31203423 Overall survival of NHD13/Bok-/- mice was similar to that of NHD13 mice suggesting that the loss of BOK does not affect the leukemogenesis in NHD13 mice (Fig. ('leukemogenesis', 'Disease', (124, 138)) ('mice', 'Species', '10090', (148, 152)) ('Bok', 'Gene', (26, 29)) ('mice', 'Species', '10090', (33, 37)) ('loss', 'Var', (92, 96)) ('Bok', 'Gene', '51800', (26, 29)) ('BOK', 'Gene', (100, 103)) ('mice', 'Species', '10090', (67, 71)) 8855 31203423 Comparison of hemoglobin (Hb) at 3 months showed that the Hb of NHD13 mice is significantly lower than that of wild type (WT) mice (Fig. ('mice', 'Species', '10090', (70, 74)) ('NHD13', 'Var', (64, 69)) ('lower', 'NegReg', (92, 97)) ('mice', 'Species', '10090', (126, 130)) 8860 31203423 In other words, the loss of BOK exacerbated the anemia of the NHD13 mice, which raised a potential connection between BOK and the regulation of erythropoiesis in cells experiencing stress from NHD13 translocation. ('mice', 'Species', '10090', (68, 72)) ('BOK', 'Gene', (28, 31)) ('loss', 'Var', (20, 24)) ('anemia', 'Phenotype', 'HP:0001903', (48, 54)) ('exacerbated', 'PosReg', (32, 43)) ('anemia', 'Disease', (48, 54)) ('anemia', 'Disease', 'MESH:D000740', (48, 54)) 8865 31203423 Isolation of various stages of erythroid progenitors in the bone marrow by Fluorescence Activated Cell Sorting (FACS), as described previously, revealed that both NHD13 and NHD13/Bok-/- mice have an increase in proerythroblasts relative to more mature red blood cells. ('NHD13', 'Var', (163, 168)) ('Bok', 'Gene', '51800', (179, 182)) ('mice', 'Species', '10090', (186, 190)) ('proerythroblasts', 'CPA', (211, 227)) ('increase', 'PosReg', (199, 207)) ('Bok', 'Gene', (179, 182)) 8872 31203423 We found that expression levels of ATF4 in NHD13, NHD13 /Bok-/- and Bok-/- are significantly lower than those of WT in the RBC population, as well (Fig. ('ATF4', 'Gene', (35, 39)) ('expression levels', 'MPA', (14, 31)) ('Bok', 'Gene', '51800', (68, 71)) ('NHD13', 'Var', (43, 48)) ('ATF4', 'Gene', '11911', (35, 39)) ('Bok', 'Gene', (57, 60)) ('lower', 'NegReg', (93, 98)) ('Bok', 'Gene', (68, 71)) ('Bok', 'Gene', '51800', (57, 60)) 8873 31203423 Finally, we found that the expression levels of CHOP in NHD13, NHD13 /Bok-/- and Bok-/- are significantly lower than those of WT in RBC population, (Fig. ('lower', 'NegReg', (106, 111)) ('expression levels', 'MPA', (27, 44)) ('Bok', 'Gene', (70, 73)) ('CHOP', 'Gene', '13198', (48, 52)) ('Bok', 'Gene', '51800', (70, 73)) ('Bok', 'Gene', (81, 84)) ('Bok', 'Gene', '51800', (81, 84)) ('NHD13', 'Var', (56, 61)) ('CHOP', 'Gene', (48, 52)) 8874 31203423 These findings imply that mechanism of decreased erythropoiesis in NHD13 and NHD13/Bok-/-mice may involve the Activating Transcription Factor 4 (ATF4) pathway. ('ATF4', 'Gene', (145, 149)) ('decreased erythropoiesis', 'Phenotype', 'HP:0010972', (39, 63)) ('Bok', 'Gene', (83, 86)) ('mice', 'Species', '10090', (89, 93)) ('NHD13', 'Var', (67, 72)) ('Bok', 'Gene', '51800', (83, 86)) ('ATF4', 'Gene', '11911', (145, 149)) ('Activating Transcription Factor 4', 'Gene', (110, 143)) ('erythropoiesis', 'CPA', (49, 63)) ('decreased', 'NegReg', (39, 48)) ('Activating Transcription Factor 4', 'Gene', '11911', (110, 143)) 8877 31203423 Similarly, the loss of PUMA reduced apoptosis and significantly delayed the development of AML. ('delayed', 'NegReg', (64, 71)) ('reduced', 'NegReg', (28, 35)) ('AML', 'Disease', 'MESH:D015470', (91, 94)) ('apoptosis', 'CPA', (36, 45)) ('AML', 'Disease', (91, 94)) ('PUMA', 'Gene', (23, 27)) ('loss', 'Var', (15, 19)) ('AML', 'Phenotype', 'HP:0004808', (91, 94)) ('PUMA', 'Gene', '170770', (23, 27)) 8878 31203423 The loss of pro-apoptotic BOK was expected to delay the progression to AML in NHD13 mice, but overall survival of NHD13/Bok-/- mice was not significantly different from that of NHD13 mice. ('mice', 'Species', '10090', (127, 131)) ('Bok', 'Gene', '51800', (120, 123)) ('AML', 'Disease', 'MESH:D015470', (71, 74)) ('mice', 'Species', '10090', (84, 88)) ('AML', 'Disease', (71, 74)) ('mice', 'Species', '10090', (183, 187)) ('BOK', 'Gene', (26, 29)) ('AML', 'Phenotype', 'HP:0004808', (71, 74)) ('delay', 'NegReg', (46, 51)) ('progression', 'MPA', (56, 67)) ('Bok', 'Gene', (120, 123)) ('loss', 'Var', (4, 8)) 8879 31203423 The percentages of AML development in NHD13/Bok-/- and NHD13 mice were 36.7% and 11.1%, respectively, which were not significantly different. ('mice', 'Species', '10090', (61, 65)) ('AML', 'Disease', 'MESH:D015470', (19, 22)) ('AML', 'Disease', (19, 22)) ('AML', 'Phenotype', 'HP:0004808', (19, 22)) ('Bok', 'Gene', (44, 47)) ('Bok', 'Gene', '51800', (44, 47)) ('NHD13', 'Var', (55, 60)) 8880 31203423 This suggests that loss of BOK does not appear to affect the development of AML. ('AML', 'Disease', (76, 79)) ('AML', 'Phenotype', 'HP:0004808', (76, 79)) ('loss', 'Var', (19, 23)) ('BOK', 'Gene', (27, 30)) ('AML', 'Disease', 'MESH:D015470', (76, 79)) 8888 31203423 Previously, knock-out of ATF4 in mice resulted in severe fetal anemia caused by proliferation defect of BFU-E and erythrocyte colony-forming unit (CFU-E). ('ATF4', 'Gene', '11911', (25, 29)) ('anemia', 'Disease', 'MESH:D000740', (63, 69)) ('erythrocyte colony-forming unit', 'CPA', (114, 145)) ('knock-out', 'Var', (12, 21)) ('anemia', 'Phenotype', 'HP:0001903', (63, 69)) ('resulted in', 'Reg', (38, 49)) ('proliferation', 'CPA', (80, 93)) ('ATF4', 'Gene', (25, 29)) ('defect', 'NegReg', (94, 100)) ('mice', 'Species', '10090', (33, 37)) ('anemia', 'Disease', (63, 69)) 8909 30636933 Cas9 generates DSBs at a target genetic locus like ZFNs and TALENs. ('DSBs', 'Chemical', '-', (15, 19)) ('Cas9', 'Chemical', '-', (0, 4)) ('DSBs', 'MPA', (15, 19)) ('TALENs', 'Disease', (60, 66)) ('Cas9', 'Var', (0, 4)) 8910 30636933 The advantage of Cas9 editing is that its nuclease function is guided either by a natural dual-RNA complex or a chimeric single-guide RNA (sgRNA) that recognizes target sequences via Watson-Crick base pairing. ('Cas9', 'Chemical', '-', (17, 21)) ('editing', 'Var', (22, 29)) ('Watson-Crick base pairing', 'Var', (183, 208)) 8913 30636933 Repair by NHEJ introduces insertion/deletion (Indel) mutations and larger deletions in the genome. ('Indel', 'Chemical', '-', (46, 51)) ('insertion/deletion', 'Var', (26, 44)) ('NHEJ', 'Gene', (10, 14)) 8918 30636933 The type I systems use Cas3 enzyme, type II systems use Cas9, type III systems use Cas10 enzyme, type IV systems use Csf1 protein, type V systems use Cpf1, C2c1 or C2c3, and type VI use protein Cas13a. ('Csf1', 'Gene', '1435', (117, 121)) ('Cas', 'Chemical', 'MESH:D002118', (23, 26)) ('C2c1', 'Var', (156, 160)) ('Cas3', 'Gene', (23, 27)) ('Cas3', 'Gene', '10278', (23, 27)) ('C2c3', 'Var', (164, 168)) ('Cas', 'Chemical', 'MESH:D002118', (83, 86)) ('Cas', 'Chemical', 'MESH:D002118', (56, 59)) ('Cas9', 'Chemical', '-', (56, 60)) ('Cas', 'Chemical', 'MESH:D002118', (194, 197)) ('Csf1', 'Gene', (117, 121)) 8924 30636933 Also, Cpf1 recognizes T-rich PAM sequences adjacent to the target DNA unlike Cas9, which recognizes G-rich PAM. ('Cas9', 'Chemical', '-', (77, 81)) ('Cpf1', 'Gene', (6, 10)) ('T-rich', 'Var', (22, 28)) 8930 30636933 The associated proteins Csx27 represses, whereas Csx28 enhances the Cas513b mediated RNA interference. ('Csx28', 'Var', (49, 54)) ('enhances', 'PosReg', (55, 63)) ('Cas', 'Chemical', 'MESH:D002118', (68, 71)) ('Cas513b', 'Var', (68, 75)) 8931 30636933 Mutating both nuclease domains of Cas9 renders the protein catalytically dead (dCas9). ('Mutating', 'Var', (0, 8)) ('protein', 'Protein', (51, 58)) ('catalytically', 'MPA', (59, 72)) ('Cas9', 'Chemical', '-', (34, 38)) ('Cas9', 'Gene', (34, 38)) ('Cas9', 'Chemical', '-', (80, 84)) 8934 30636933 dCas9 can also be used for gene activation, this approach is known as CRISPR activator (CRISPRa); dCas9 can also mediate reversible CRISPR interference (CRISPRi). ('CRISPR', 'Disease', (132, 138)) ('dCas9', 'Var', (98, 103)) ('Cas9', 'Chemical', '-', (1, 5)) ('Cas9', 'Chemical', '-', (99, 103)) 8960 30636933 The Degron-KI system has also helped identify the link between putative oncogene SF3B1 hotspot mutations and splicing alterations. ('3B1', 'Species', '1413326', (83, 86)) ('splicing', 'MPA', (109, 117)) ('SF3B1', 'Gene', (81, 86)) ('mutations', 'Var', (95, 104)) 8966 30636933 Isogenic organoids with mutations in tumor suppressor genes APC, SMAD4, TP53 and oncogenes KRAS and/or PIK3Ca are selected and implanted in mice where they form tumors and successfully mimic colon cancer in vitro. ('PIK3Ca', 'Gene', (103, 109)) ('tumor', 'Disease', 'MESH:D009369', (37, 42)) ('tumors', 'Disease', (161, 167)) ('TP53', 'Gene', '22059', (72, 76)) ('APC', 'Gene', '11789', (60, 63)) ('mutations', 'Var', (24, 33)) ('tumor', 'Disease', (161, 166)) ('colon cancer', 'Phenotype', 'HP:0003003', (191, 203)) ('SMAD4', 'Gene', '17128', (65, 70)) ('mice', 'Species', '10090', (140, 144)) ('tumors', 'Disease', 'MESH:D009369', (161, 167)) ('tumor', 'Disease', 'MESH:D009369', (161, 166)) ('tumor', 'Phenotype', 'HP:0002664', (37, 42)) ('PIK3Ca', 'Gene', '18706', (103, 109)) ('KRAS', 'Gene', (91, 95)) ('colon cancer', 'Disease', 'MESH:D015179', (191, 203)) ('tumor', 'Phenotype', 'HP:0002664', (161, 166)) ('SMAD4', 'Gene', (65, 70)) ('cancer', 'Phenotype', 'HP:0002664', (197, 203)) ('APC', 'Gene', (60, 63)) ('tumors', 'Phenotype', 'HP:0002664', (161, 167)) ('colon cancer', 'Disease', (191, 203)) ('mimic', 'Reg', (185, 190)) ('TP53', 'Gene', (72, 76)) ('tumor', 'Disease', (37, 42)) 8967 30636933 CRISPR/Cas9 mediated deletion of DNA repair genes in colon organoids has been successfully modeled mismatch repair deficient colorectal cancer. ('colorectal cancer', 'Phenotype', 'HP:0003003', (125, 142)) ('deletion', 'Var', (21, 29)) ('Cas9', 'Chemical', '-', (7, 11)) ('deficient colorectal cancer', 'Disease', 'MESH:D015179', (115, 142)) ('deficient colorectal cancer', 'Disease', (115, 142)) ('DNA repair genes', 'Gene', (33, 49)) ('cancer', 'Phenotype', 'HP:0002664', (136, 142)) 8969 30636933 CRISPR/Cas9 driven mutagenesis has been used to mutate 5 genes (Tet1, 2, 3, Sry, Uty-8 alleles) in mouse ES cells in a single step, which reduces the complexity of generating multi-gene cancer models. ('mouse', 'Species', '10090', (99, 104)) ('multi-gene cancer', 'Disease', (175, 192)) ('Tet1, 2, 3', 'Gene', '52463;214133;194388', (64, 74)) ('cancer', 'Phenotype', 'HP:0002664', (186, 192)) ('Cas9', 'Chemical', '-', (7, 11)) ('mutate', 'Var', (48, 54)) ('Sry', 'Gene', (76, 79)) ('multi-gene cancer', 'Disease', 'MESH:D009369', (175, 192)) ('Sry', 'Gene', '21674', (76, 79)) 8970 30636933 Tet1 and Tet2 gene mutations were generated with 80% efficiency by coinjecting Cas9 mRNA and sgRNAs directly into the mice zygote. ('Tet2', 'Gene', (9, 13)) ('mutations', 'Var', (19, 28)) ('Tet1', 'Gene', '52463', (0, 4)) ('Cas9', 'Chemical', '-', (79, 83)) ('Tet2', 'Gene', '214133', (9, 13)) ('Tet1', 'Gene', (0, 4)) ('mice', 'Species', '10090', (118, 122)) 8975 30636933 Cre dependent Cas9 knock in mouse was generated and using these mice, KRAS, p53 and LKB1 gene mutations were modeled for lung adenocarcinoma. ('lung adenocarcinoma', 'Disease', (121, 140)) ('carcinoma', 'Phenotype', 'HP:0030731', (131, 140)) ('mouse', 'Species', '10090', (28, 33)) ('mutations', 'Var', (94, 103)) ('p53', 'Gene', (76, 79)) ('p53', 'Gene', '22059', (76, 79)) ('lung adenocarcinoma', 'Disease', 'MESH:D000077192', (121, 140)) ('Cas9', 'Chemical', '-', (14, 18)) ('LKB1', 'Gene', (84, 88)) ('lung adenocarcinoma', 'Phenotype', 'HP:0030078', (121, 140)) ('LKB1', 'Gene', '20869', (84, 88)) ('mice', 'Species', '10090', (64, 68)) 8990 30636933 Ongoing clinical trials include PD-1 knockout autologous T-cells in prostate cancer (NCT02867345), bladder cancer (NCT02863913) and renal cell carcinoma (NCT02867332). ('bladder cancer', 'Phenotype', 'HP:0009725', (99, 113)) ('renal cell carcinoma', 'Disease', (132, 152)) ('renal cell carcinoma', 'Phenotype', 'HP:0005584', (132, 152)) ('NCT02867345', 'Var', (85, 96)) ('cancer', 'Phenotype', 'HP:0002664', (107, 113)) ('renal cell carcinoma', 'Disease', 'MESH:C538614', (132, 152)) ('prostate cancer', 'Disease', (68, 83)) ('NCT02867332', 'Var', (154, 165)) ('bladder cancer', 'Disease', 'MESH:D001749', (99, 113)) ('NCT02863913', 'Var', (115, 126)) ('bladder cancer', 'Disease', (99, 113)) ('cancer', 'Phenotype', 'HP:0002664', (77, 83)) ('PD-1', 'Gene', (32, 36)) ('carcinoma', 'Phenotype', 'HP:0030731', (143, 152)) ('prostate cancer', 'Disease', 'MESH:D011471', (68, 83)) ('prostate cancer', 'Phenotype', 'HP:0012125', (68, 83)) 8991 30636933 CRISPR/Cas-9 mediated simultaneous knockout of 4 loci of PD-1 and CTLA-4 have been successful in generating allogeneic universal T-donor cells. ('CTLA-4', 'Gene', (66, 72)) ('donor', 'Species', '9606', (131, 136)) ('Cas', 'Chemical', 'MESH:D002118', (7, 10)) ('CTLA-4', 'Gene', '1493', (66, 72)) ('knockout', 'Var', (35, 43)) ('PD-1', 'Gene', (57, 61)) 8992 30636933 CRISPR/Cas9 mediated generation of lymphocyte activating gene-3 (LAG-3) knockout CAR-T cells shows better specificity and anti-tumor potential in xenograft mouse models. ('better', 'PosReg', (99, 105)) ('knockout', 'Var', (72, 80)) ('tumor', 'Phenotype', 'HP:0002664', (127, 132)) ('tumor', 'Disease', (127, 132)) ('specificity', 'MPA', (106, 117)) ('Cas9', 'Chemical', '-', (7, 11)) ('mouse', 'Species', '10090', (156, 161)) ('tumor', 'Disease', 'MESH:D009369', (127, 132)) ('lymphocyte activating gene-3 (LAG-3', 'Gene', '16768', (35, 70)) 8995 30636933 The study showed enhanced efficacy of immunotherapy by deletion of PTPN2 via interferon gamma mediated pathway. ('enhanced', 'PosReg', (17, 25)) ('PTPN2', 'Gene', (67, 72)) ('immunotherapy', 'CPA', (38, 51)) ('PTPN2', 'Gene', '5771', (67, 72)) ('deletion', 'Var', (55, 63)) ('efficacy', 'CPA', (26, 34)) 8997 30636933 In this trial, the immune cells from recipient blood was removed followed by ex vivo CRISPR/Cas9 editing thereby disabling PD-1 protein. ('disabling', 'NegReg', (113, 122)) ('Cas9', 'Chemical', '-', (92, 96)) ('editing', 'Var', (97, 104)) ('PD-1 protein', 'Protein', (123, 135)) 8999 30636933 A new clinical trail (NCT03057912) proposes to use a combination of TALENs and CRISPR/Cas9 in the treatment of HPV-related cervical neoplasma by targeting HPV16 and HPV18 E6/E7 DNA; this approach promises to reduce off-target effects. ('HPV-related cervical neoplasma', 'Disease', (111, 141)) ('E6/E7 DNA', 'Var', (171, 180)) ('HPV18', 'Gene', (165, 170)) ('Cas9', 'Chemical', '-', (86, 90)) ('HPV16', 'Species', '333760', (155, 160)) ('HPV16', 'Gene', (155, 160)) ('cervical neoplasma', 'Phenotype', 'HP:0032241', (123, 141)) 9014 30636933 Some strategies to avoid or at least minimize off-target effects of CRISPR/Cas9 editing include (i) choosing unique target sites that lack homology to any other region of the genome, (ii) modifying Cas9 by replacing wild type Cas9 with dCas9, (iii) fusing dCas9 with FokI nuclease (fCas9) which has higher sequence targeting specificity, (iv) modifying sgRNA by truncating the 3'end of sgRNA, and (v) reducing the concentration of Cas9-sgRNA delivered to cells. ('Cas9', 'Chemical', '-', (198, 202)) ('Cas9', 'Chemical', '-', (283, 287)) ('Cas9', 'Chemical', '-', (431, 435)) ('reducing', 'NegReg', (401, 409)) ('Cas9', 'Chemical', '-', (75, 79)) ('Cas9', 'Chemical', '-', (226, 230)) ('modifying', 'Var', (188, 197)) ('Cas9', 'Chemical', '-', (257, 261)) ('concentration', 'MPA', (414, 427)) ('sgRNA', 'Protein', (386, 391)) ('modifying', 'Reg', (343, 352)) ('sgRNA', 'Gene', (353, 358)) ('Cas9', 'Chemical', '-', (237, 241)) ('fusing', 'Var', (249, 255)) ('truncating', 'NegReg', (362, 372)) ("3'end of", 'MPA', (377, 385)) 9017 30636933 Also, Cas9 from S. aureus and S. pyogenes has been shown to cause infectious diseases in humans. ('cause', 'Reg', (60, 65)) ('infectious diseases', 'Disease', 'MESH:D003141', (66, 85)) ('S. pyogenes', 'Var', (30, 41)) ('Cas9', 'Chemical', '-', (6, 10)) ('humans', 'Species', '9606', (89, 95)) ('S. aureus', 'Species', '1280', (16, 25)) ('Cas9', 'Var', (6, 10)) ('S. pyogenes', 'Species', '1314', (30, 41)) ('infectious diseases', 'Disease', (66, 85)) 9037 28933777 A substantial proportion of primary AML patient samples, in particular less aggressive clinical subtypes such as those bearing mutations in core binding factor and those classified as acute promyelocytic leukemia (APL), failed to engraft in NOD/SCID/IL2R-gamma null (NSG) mice or did so at low levels that do not mimic clinical human disease. ('patient', 'Species', '9606', (40, 47)) ('AML', 'Disease', 'MESH:D015470', (36, 39)) ('SCID', 'Disease', 'MESH:D053632', (245, 249)) ('APL', 'Disease', 'MESH:D015473', (214, 217)) ('acute promyelocytic leukemia', 'Disease', (184, 212)) ('SCID', 'Disease', (245, 249)) ('AML', 'Disease', (36, 39)) ('mice', 'Species', '10090', (272, 276)) ('acute promyelocytic leukemia', 'Phenotype', 'HP:0004836', (184, 212)) ('acute promyelocytic leukemia', 'Disease', 'MESH:D015473', (184, 212)) ('human', 'Species', '9606', (328, 333)) ('failed', 'NegReg', (220, 226)) ('APL', 'Disease', (214, 217)) ('mutations', 'Var', (127, 136)) ('leukemia', 'Phenotype', 'HP:0001909', (204, 212)) 9123 28933777 63323-540-57) L-Glutamax 100x (200 mM, Fisher, cat. ('L-Glutamax 100x', 'Protein', (14, 29)) ('200 mM', 'Var', (31, 37)) ('L-Glutamax 100x', 'Chemical', '-', (14, 29)) 9199 28933777 Antibodies for CD34, CD38, CD45RA, and CD90 are added individually with a final concentration ranging from 1:25 to 1:50, but optimal concentration should be determined empirically by each investigator. ('CD45RA', 'Var', (27, 33)) ('CD34', 'Gene', (15, 19)) ('CD34', 'Gene', '947', (15, 19)) ('CD38', 'Gene', '952', (21, 25)) ('CD90', 'Gene', '7070', (39, 43)) ('CD90', 'Gene', (39, 43)) ('CD38', 'Gene', (21, 25)) 9256 28933777 Define human HSPCs based on expression of CD34, CD38, CD90, CD45RA and CD123 (Figure 5a) on FACSAria II or other three laser machines with similar laser setup. ('CD123', 'Gene', '3563', (71, 76)) ('CD38', 'Gene', (48, 52)) ('human', 'Species', '9606', (7, 12)) ('CD90', 'Gene', '7070', (54, 58)) ('CD45RA', 'Var', (60, 66)) ('CD38', 'Gene', '952', (48, 52)) ('CD34', 'Gene', (42, 46)) ('CD34', 'Gene', '947', (42, 46)) ('CD90', 'Gene', (54, 58)) ('CD123', 'Gene', (71, 76)) 9361 28280079 In Fanconi anemia, loss of activity of the ALDH2 isoform in concert with FANC-D deficiency predisposes to the development of acute myeloid leukemia (AML) and bone marrow failure. ('FANC-D', 'Gene', (73, 79)) ('ALDH2', 'Gene', (43, 48)) ('Fanconi anemia', 'Phenotype', 'HP:0001994', (3, 17)) ('acute myeloid leukemia', 'Disease', (125, 147)) ('bone marrow failure', 'Disease', 'MESH:D000080983', (158, 177)) ('loss', 'NegReg', (19, 23)) ('FANC-D', 'Gene', '675', (73, 79)) ('AML', 'Disease', 'MESH:D015470', (149, 152)) ('bone marrow failure', 'Disease', (158, 177)) ('ALDH2', 'Gene', '217', (43, 48)) ('AML', 'Disease', (149, 152)) ('acute myeloid leukemia', 'Disease', 'MESH:D015470', (125, 147)) ('AML', 'Phenotype', 'HP:0004808', (149, 152)) ('acute myeloid leukemia', 'Phenotype', 'HP:0004808', (125, 147)) ('bone marrow failure', 'Phenotype', 'HP:0005528', (158, 177)) ('leukemia', 'Phenotype', 'HP:0001909', (139, 147)) ('Fanconi anemia', 'Disease', (3, 17)) ('myeloid leukemia', 'Phenotype', 'HP:0012324', (131, 147)) ('Fanconi anemia', 'Disease', 'MESH:D005199', (3, 17)) ('anemia', 'Phenotype', 'HP:0001903', (11, 17)) ('deficiency', 'Var', (80, 90)) 9400 28280079 In addition, low-level expression of the ALDH1A1 isoform was correlated with improved event-free survival (EFS) and overall survival (OS) in the entire cohort of AML patients (Figure 2D). ('ALDH1A1', 'Gene', (41, 48)) ('AML', 'Phenotype', 'HP:0004808', (162, 165)) ('AML', 'Disease', (162, 165)) ('event-free survival', 'CPA', (86, 105)) ('low-level expression', 'Var', (13, 33)) ('AML', 'Disease', 'MESH:D015470', (162, 165)) ('overall survival', 'CPA', (116, 132)) ('patients', 'Species', '9606', (166, 174)) ('improved', 'PosReg', (77, 85)) 9403 28280079 In addition, an in