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3.
Blood ; 133(16): 1766-1777, 2019 04 18.
Artigo em Inglês | MEDLINE | ID: mdl-30755419

RESUMO

In acute myeloid leukemia (AML), acquired genetic aberrations carry prognostic implications and guide therapeutic decisions. Clinical algorithms have been improved by the incorporation of novel aberrations. Here, we report the presence and functional characterization of mutations in the transcription factor NFE2 in patients with AML and in a patient with myelosarcoma. We previously described NFE2 mutations in patients with myeloproliferative neoplasms and demonstrated that expression of mutant NFE2 in mice causes a myeloproliferative phenotype. Now, we show that, during follow-up, 34% of these mice transform to leukemia presenting with or without concomitant myelosarcomas, or develop isolated myelosarcomas. These myelosarcomas and leukemias acquired AML-specific alterations, including the murine equivalent of trisomy 8, loss of the AML commonly deleted region on chromosome 5q, and mutations in the tumor suppressor Trp53 Our data show that mutations in NFE2 predispose to the acquisition of secondary changes promoting the development of myelosarcoma and/or AML.


Assuntos
Transformação Celular Neoplásica/genética , Leucemia Mieloide Aguda/genética , Subunidade p45 do Fator de Transcrição NF-E2/genética , Subunidade p45 do Fator de Transcrição NF-E2/metabolismo , Sarcoma Mieloide/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Aberrações Cromossômicas , Feminino , Humanos , Leucemia Mieloide Aguda/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Mutação , Sarcoma Mieloide/etiologia , Proteína Supressora de Tumor p53/genética , Adulto Jovem
4.
Blood ; 132(14): 1526-1534, 2018 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-30049810

RESUMO

The tendency of 5-methylcytosine (5mC) to undergo spontaneous deamination has had a major role in shaping the human genome, and this methylation damage remains the primary source of somatic mutations that accumulate with age. How 5mC deamination contributes to cancer risk in different tissues remains unclear. Genomic profiling of 3 early-onset acute myeloid leukemias (AMLs) identified germ line loss of MBD4 as an initiator of 5mC-dependent hypermutation. MBD4-deficient AMLs display a 33-fold higher mutation burden than AML generally, with >95% being C>T in the context of a CG dinucleotide. This distinctive signature was also observed in sporadic cancers that acquired biallelic mutations in MBD4 and in Mbd4 knockout mice. Sequential sampling of germ line cases demonstrated repeated expansion of blood cell progenitors with pathogenic mutations in DNMT3A, a key driver gene for both clonal hematopoiesis and AML. Our findings reveal genetic and epigenetic factors that shape the mutagenic influence of 5mC. Within blood cells, this links methylation damage to the driver landscape of clonal hematopoiesis and reveals a conserved path to leukemia. Germ line MBD4 deficiency enhances cancer susceptibility and predisposes to AML.


Assuntos
DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA , Endodesoxirribonucleases/genética , Regulação Leucêmica da Expressão Gênica , Hematopoese , Leucemia Mieloide Aguda/genética , Adulto , DNA Metiltransferase 3A , Feminino , Deleção de Genes , Células Germinativas/metabolismo , Células Germinativas/patologia , Humanos , Leucemia Mieloide Aguda/patologia , Masculino , Mutação , Acúmulo de Mutações
5.
N Engl J Med ; 378(13): 1189-1199, 2018 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-29601269

RESUMO

BACKGROUND: Patients with acute myeloid leukemia (AML) often reach complete remission, but relapse rates remain high. Next-generation sequencing enables the detection of molecular minimal residual disease in virtually every patient, but its clinical value for the prediction of relapse has yet to be established. METHODS: We conducted a study involving patients 18 to 65 years of age who had newly diagnosed AML. Targeted next-generation sequencing was carried out at diagnosis and after induction therapy (during complete remission). End points were 4-year rates of relapse, relapse-free survival, and overall survival. RESULTS: At least one mutation was detected in 430 out of 482 patients (89.2%). Mutations persisted in 51.4% of those patients during complete remission and were present at various allele frequencies (range, 0.02 to 47%). The detection of persistent DTA mutations (i.e., mutations in DNMT3A, TET2, and ASXL1), which are often present in persons with age-related clonal hematopoiesis, was not correlated with an increased relapse rate. After the exclusion of persistent DTA mutations, the detection of molecular minimal residual disease was associated with a significantly higher relapse rate than no detection (55.4% vs. 31.9%; hazard ratio, 2.14; P<0.001), as well as with lower rates of relapse-free survival (36.6% vs. 58.1%; hazard ratio for relapse or death, 1.92; P<0.001) and overall survival (41.9% vs. 66.1%; hazard ratio for death, 2.06; P<0.001). Multivariate analysis confirmed that the persistence of non-DTA mutations during complete remission conferred significant independent prognostic value with respect to the rates of relapse (hazard ratio, 1.89; P<0.001), relapse-free survival (hazard ratio for relapse or death, 1.64; P=0.001), and overall survival (hazard ratio for death, 1.64; P=0.003). A comparison of sequencing with flow cytometry for the detection of residual disease showed that sequencing had significant additive prognostic value. CONCLUSIONS: Among patients with AML, the detection of molecular minimal residual disease during complete remission had significant independent prognostic value with respect to relapse and survival rates, but the detection of persistent mutations that are associated with clonal hematopoiesis did not have such prognostic value within a 4-year time frame. (Funded by the Queen Wilhelmina Fund Foundation of the Dutch Cancer Society and others.).


Assuntos
Análise Mutacional de DNA , DNA de Neoplasias/análise , Leucemia Mieloide Aguda/genética , Mutação , Neoplasia Residual/genética , Adolescente , Adulto , Idoso , Análise Mutacional de DNA/métodos , Feminino , Citometria de Fluxo , Hematopoese/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Leucemia Mieloide Aguda/mortalidade , Masculino , Pessoa de Meia-Idade , Neoplasia Residual/diagnóstico , Prognóstico , Modelos de Riscos Proporcionais , Recidiva , Indução de Remissão , Análise de Sobrevida , Adulto Jovem
7.
Blood ; 125(1): 133-9, 2015 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-25381062

RESUMO

Myeloid malignancies bearing chromosomal inv(3)/t(3;3) abnormalities are among the most therapy-resistant leukemias. Deregulated expression of EVI1 is the molecular hallmark of this disease; however, the genome-wide spectrum of cooperating mutations in this disease subset has not been systematically elucidated. Here, we show that 98% of inv(3)/t(3;3) myeloid malignancies harbor mutations in genes activating RAS/receptor tyrosine kinase (RTK) signaling pathways. In addition, hemizygous mutations in GATA2, as well as heterozygous alterations in RUNX1, SF3B1, and genes encoding epigenetic modifiers, frequently co-occur with the inv(3)/t(3;3) aberration. Notably, neither mutational patterns nor gene expression profiles differ across inv(3)/t(3;3) acute myeloid leukemia, chronic myeloid leukemia, and myelodysplastic syndrome cases, suggesting recognition of inv(3)/t(3;3) myeloid malignancies as a single disease entity irrespective of blast count. The high incidence of activating RAS/RTK signaling mutations may provide a target for a rational treatment strategy in this high-risk patient group.


Assuntos
Inversão Cromossômica , Leucemia Mieloide Aguda/genética , Síndromes Mielodisplásicas/genética , Receptores Proteína Tirosina Quinases/metabolismo , Transdução de Sinais , Translocação Genética , Proteínas ras/metabolismo , Alelos , Bandeamento Cromossômico , Cromossomos Humanos Par 3 , Análise Mutacional de DNA , Epigênese Genética , Exoma , Perfilação da Expressão Gênica , Humanos , Análise de Sequência de DNA , Análise de Sequência de RNA
8.
Haematologica ; 99(5): 848-57, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24441149

RESUMO

Acute myeloid leukemia is a neoplasm characterized by recurrent molecular aberrations traditionally demonstrated by cytogenetic analyses. We used high density genome-wide genotyping and gene expression profiling to reveal acquired cryptic abnormalities in acute myeloid leukemia. By genome-wide genotyping of 137 cases of primary acute myeloid leukemia, we disclosed a recurrent focal amplification on chromosome 14q32, which included the genes BCL11B, CCNK, C14orf177 and SETD3, in two cases. In the affected cases, the BCL11B gene showed consistently high mRNA expression, whereas the expression of the other genes was unperturbed. Fluorescence in situ hybridization on 40 cases of acute myeloid leukemia with high BCL11B mRNA expression [2.5-fold above median; 40 out of 530 cases (7.5%)] revealed 14q32 abnormalities in two additional cases. In the four BCL11B-rearranged cases the 14q32 locus was fused to different partner chromosomes. In fact, in two cases, we demonstrated that the focal 14q32 amplifications were integrated into transcriptionally active loci. The translocations involving BCL11B result in increased expression of full-length BCL11B protein. The BCL11B-rearranged acute myeloid leukemias expressed both myeloid and T-cell markers. These biphenotypic acute leukemias all carried FLT3 internal tandem duplications, a characteristic marker of acute myeloid leukemia. BCL11B mRNA expression in acute myeloid leukemia appeared to be strongly associated with expression of other T-cell-specific genes. Myeloid 32D(GCSF-R) cells ectopically expressing Bcl11b showed decreased proliferation rate and less maturation. In conclusion, by an integrated approach involving high-throughput genome-wide genotyping and gene expression profiling we identified BCL11B as a candidate oncogene in acute myeloid leukemia.


Assuntos
Aberrações Cromossômicas , Cromossomos Humanos Par 14 , Leucemia Mieloide Aguda/genética , Oncogenes , Proteínas Repressoras/genética , Proteínas Supressoras de Tumor/genética , Antígenos de Superfície/metabolismo , Estudos de Casos e Controles , Proliferação de Células , Dosagem de Genes , Expressão Gênica , Perfilação da Expressão Gênica , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Imunofenotipagem , Hibridização in Situ Fluorescente , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/metabolismo , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNA , Linfócitos T/metabolismo , Translocação Genética
9.
Blood ; 119(24): 5824-31, 2012 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-22490330

RESUMO

The prevalence, the prognostic effect, and interaction with other molecular markers of DNMT3A mutations was studied in 415 patients with acute myeloid leukemia (AML) younger than 60 years. We show mutations in DNMT3A in 96 of 415 patients with newly diagnosed AML (23.1%). Univariate Cox regression analysis showed that patients with DNMT3A(mutant) AML show significantly worse overall survival (OS; P = .022; hazard ratio [HR], 1.38; 95% confidence interval [CI], 1.04-1.81), and relapse-free survival (RFS; P = .005; HR, 1.52; 95% CI, 1.13-2.05) than DNMT3A(wild-type) AMLs. In a multivariable analysis, DNMT3A mutations express independent unfavorable prognostic value for OS (P = .003; HR, 1.82; 95% CI, 1.2-2.7) and RFS (P < .001; HR, 2.2; 95% CI, 1.4-3.3). In a composite genotypic subset of cytogenetic intermediate-risk AML without FLT3-ITD and NPM1 mutations, this association is particularly evident (OS: P = .013; HR, 2.09; 95% CI, 1.16-3.77; RFS: P = .001; HR, 2.65; 95% CI, 1.48-4.89). The effect of DNMT3A mutations in human AML remains elusive, because DNMT3A(mutant) AMLs did not express a methylation or gene expression signature that discriminates them from patients with DNMT3A(wild-type) AML. We conclude that DNMT3A mutation status is an important factor to consider for risk stratification of patients with AML.


Assuntos
DNA (Citosina-5-)-Metiltransferases/genética , Leucemia Mieloide Aguda/enzimologia , Leucemia Mieloide Aguda/genética , Proteínas Mutantes/metabolismo , Adolescente , Adulto , Biomarcadores Tumorais , Análise por Conglomerados , Metilação de DNA/genética , DNA Metiltransferase 3A , Feminino , Perfilação da Expressão Gênica , Regulação Leucêmica da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Mutação/genética , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Nucleofosmina , Prognóstico , Recidiva , Análise de Sobrevida , Resultado do Tratamento , Adulto Jovem
10.
Blood ; 116(12): 2122-6, 2010 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-20538800

RESUMO

Somatic mutations in isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) were recently demonstrated in acute myeloid leukemia (AML), but their prevalence and prognostic impact remain to be explored in large extensively characterized AML series, and also in various other hematologic malignancies. Here, we demonstrate in 893 newly diagnosed cases of AML mutations in the IDH1 (6%) and IDH2 (11%) genes. Moreover, we identified IDH mutations in 2 JAK2 V617F myeloproliferative neoplasias (n = 96), a single case of acute lymphoblastic leukemia (n = 96), and none in chronic myeloid leukemias (n = 81). In AML, IDH1 and IDH2 mutations are more common among AML with normal karyotype and NPM1(mutant) genotypes. IDH1 mutation status is an unfavorable prognostic factor as regards survival in a composite genotypic subset lacking FLT3(ITD) and NPM1(mutant). Thus, IDH1 and IDH2 mutations are common genetic aberrations in AML, and IDH1 mutations may carry prognostic value in distinct subtypes of AML.


Assuntos
Isocitrato Desidrogenase/genética , Leucemia Mieloide Aguda/genética , Mutação , Adolescente , Adulto , Idoso , Feminino , Doenças Hematológicas , Humanos , Janus Quinase 2/genética , Leucemia Mieloide Aguda/classificação , Masculino , Pessoa de Meia-Idade , Nucleofosmina , Prevalência , Prognóstico , Adulto Jovem
11.
Exp Cell Res ; 297(2): 434-43, 2004 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-15212946

RESUMO

The critical factors in the regulation of telomere length are not yet clearly defined. Telomerase is a key player in telomere elongation, although previous studies have shown that telomeres are differentially elongated after telomerase reconstitution. Moreover, a clear relation between the level of telomerase activity and telomere length was not observed. To investigate which factors are critical in telomere length regulation, we generated 24 telomerase-reconstituted primary human fibroblast clones. In these clones, in vitro telomerase activity level is clearly related to telomere length. High levels of telomerase activity are associated with longer telomeres and better telomere maintenance over time. The correlation coefficient, however, indicates that the level of telomerase activity is not the only factor in the regulation of telomere length. Clearly, factors that are not measured in an in vitro telomerase activity assay are involved in telomere length regulation in vivo. To investigate which telomerase components are critical in regulating telomerase activity levels, we studied expression levels of hTERT mRNA and hTR. Expression is highly variable between individual clones, but not related to the level of telomerase activity or telomere length. Our results indicate that expression levels of hTERT mRNA and hTR do not regulate the activity level of the telomerase complex, suggesting posttranscriptional modification of hTERT or the presence of additional proteins that modulate telomerase enzyme activity.


Assuntos
Fibroblastos/enzimologia , Regulação Enzimológica da Expressão Gênica , Telomerase/análise , Telômero/metabolismo , Células Cultivadas , Células Clonais , Proteínas de Ligação a DNA , Citometria de Fluxo , Proteínas de Fluorescência Verde , Humanos , Hibridização in Situ Fluorescente , Proteínas Luminescentes , Reação em Cadeia da Polimerase , RNA , Retroviridae/genética , Pele/citologia
12.
Mol Endocrinol ; 18(6): 1499-508, 2004 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15016840

RESUMO

In this study, molecular simulations have been combined with site-directed mutagenesis experiments to explore M398(2.43), a LH (lutropin) receptor (LHR) site in helix 2 susceptible to spontaneous activating mutations, and to develop a computational tool for predicting the functionality (i.e. active or nonactive) of LHR mutants.Site-directed mutagenesis experiments engineered 15 different substitutions for M389(2.43), which resulted in variable levels of constitutive activity, inversely correlated with the size of the replacing amino acid. This inverse correlation is suggested to be mediated by I460(3.46), M571(6.37), and Y623(7.53), the tyrosine of the NPxxY motif. In fact, size reduction at position 398(2.43), which is concurrent with constitutive receptor activity, releases the van der Waals interactions found in the wild-type LHR between M398(2.43) and these three amino acids, resulting in structural modifications in the proximity to the E/DRY/W motif. An increment, above a threshold value, in the solvent accessibility of the cytosolic ends of helices 3 and 6 is the main structural feature shared by the active mutants of the LHR. This feature has been successfully used for predicting the functionality of the engineered mutants at M398(2.43), proving that molecular simulations can be useful for in silico screening of LHR mutants.


Assuntos
Mutação , Receptores do LH/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Sítios de Ligação , Linhagem Celular , AMP Cíclico/metabolismo , Citosol/metabolismo , DNA Complementar/metabolismo , Relação Dose-Resposta a Droga , Genes Reporter , Humanos , Cinética , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese , Mutagênese Sítio-Dirigida , Oligonucleotídeos/química , Plasmídeos/metabolismo , Ligação Proteica , Conformação Proteica , Rodopsina/química , Homologia de Sequência de Aminoácidos , Software , Fatores de Tempo , Transfecção , Tirosina/química
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