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1.
Leuk Res ; 87: 106269, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31751766

RESUMO

Bromodomain and Extra-Terminal inhibitors (BETi) such as OTX015 are active in Acute Myeloid Leukaemias (AML). Their activity on Leukemic Stem Cells (LSCs) is less documented. We interrogated the anti-LSC activity of OTX015 in a niche-like long-term culture in 26 primary AML samples and validated our findings in vivo. OTX015 impaired LSCs in AMLs harbouring Core Binding Factor or KMT2A gene fusions, NPM1 or chromatin/spliceosome genes mutations, but not in those with aneuploidy/TP53 mutations. In four patients, we dissected the transcriptomic footprint of Bet inhibition on LSCs versus blasts. Our results can instruct future clinical trials of BETi in AML.

2.
Blood Adv ; 3(20): 2962-2979, 2019 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-31648317

RESUMO

Standardized variant curation is essential for clinical care recommendations for patients with inherited disorders. Clinical Genome Resource (ClinGen) variant curation expert panels are developing disease-associated gene specifications using the 2015 American College of Medical Genetics and Genomics (ACMG) and Association for Molecular Pathology (AMP) guidelines to reduce curation discrepancies. The ClinGen Myeloid Malignancy Variant Curation Expert Panel (MM-VCEP) was created collaboratively between the American Society of Hematology and ClinGen to perform gene- and disease-specific modifications for inherited myeloid malignancies. The MM-VCEP began optimizing ACMG/AMP rules for RUNX1 because many germline variants have been described in patients with familial platelet disorder with a predisposition to acute myeloid leukemia, characterized by thrombocytopenia, platelet functional/ultrastructural defects, and a predisposition to hematologic malignancies. The 28 ACMG/AMP codes were tailored for RUNX1 variants by modifying gene/disease specifications, incorporating strength adjustments of existing rules, or both. Key specifications included calculation of minor allele frequency thresholds, formulating a semi-quantitative approach to counting multiple independent variant occurrences, identifying functional domains and mutational hotspots, establishing functional assay thresholds, and characterizing phenotype-specific guidelines. Preliminary rules were tested by using a pilot set of 52 variants; among these, 50 were previously classified as benign/likely benign, pathogenic/likely pathogenic, variant of unknown significance (VUS), or conflicting interpretations (CONF) in ClinVar. The application of RUNX1-specific criteria resulted in a reduction in CONF and VUS variants by 33%, emphasizing the benefit of gene-specific criteria and sharing internal laboratory data.

3.
Nat Genet ; 51(10): 1518-1529, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31570891

RESUMO

RNA modifications are emerging as key determinants of gene expression. However, compelling genetic demonstrations of their relevance to human disease are lacking. Here, we link ribosomal RNA 2'-O-methylation (2'-O-Me) to the etiology of dyskeratosis congenita. We identify nucleophosmin (NPM1) as an essential regulator of 2'-O-Me on rRNA by directly binding C/D box small nucleolar RNAs, thereby modulating translation. We demonstrate the importance of 2'-O-Me-regulated translation for cellular growth, differentiation and hematopoietic stem cell maintenance, and show that Npm1 inactivation in adult hematopoietic stem cells results in bone marrow failure. We identify NPM1 germline mutations in patients with dyskeratosis congenita presenting with bone marrow failure and demonstrate that they are deficient in small nucleolar RNA binding. Mice harboring a dyskeratosis congenita germline Npm1 mutation recapitulate both hematological and nonhematological features of dyskeratosis congenita. Thus, our findings indicate that impaired 2'-O-Me can be etiological to human disease.

4.
Nat Med ; 25(9): 1396-1401, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31501599

RESUMO

Fanconi anemia (FA) is a DNA repair syndrome generated by mutations in any of the 22 FA genes discovered to date1,2. Mutations in FANCA account for more than 60% of FA cases worldwide3,4. Clinically, FA is associated with congenital abnormalities and cancer predisposition. However, bone marrow failure is the primary pathological feature of FA that becomes evident in 70-80% of patients with FA during the first decade of life5,6. In this clinical study (ClinicalTrials.gov, NCT03157804 ; European Clinical Trials Database, 2011-006100-12), we demonstrate that lentiviral-mediated hematopoietic gene therapy reproducibly confers engraftment and proliferation advantages of gene-corrected hematopoietic stem cells (HSCs) in non-conditioned patients with FA subtype A. Insertion-site analyses revealed the multipotent nature of corrected HSCs and showed that the repopulation advantage of these cells was not due to genotoxic integrations of the therapeutic provirus. Phenotypic correction of blood and bone marrow cells was shown by the acquired resistance of hematopoietic progenitors and T lymphocytes to DNA cross-linking agents. Additionally, an arrest of bone marrow failure progression was observed in patients with the highest levels of gene marking. The progressive engraftment of corrected HSCs in non-conditioned patients with FA supports that gene therapy should constitute an innovative low-toxicity therapeutic option for this life-threatening disorder.


Assuntos
Proteína do Grupo de Complementação A da Anemia de Fanconi/genética , Anemia de Fanconi/terapia , Terapia Genética , Transplante de Células-Tronco Hematopoéticas , Adolescente , Adulto , Células da Medula Óssea/citologia , Criança , Pré-Escolar , Anemia de Fanconi/genética , Anemia de Fanconi/fisiopatologia , Feminino , Vetores Genéticos/genética , Células-Tronco Hematopoéticas/metabolismo , Humanos , Lactente , Lentivirus/genética , Masculino , Mutação/genética , Espanha/epidemiologia , Reparo Gênico Alvo-Dirigido , Transdução Genética , Adulto Jovem
5.
Blood ; 134(17): 1441-1444, 2019 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-31484648

RESUMO

Germline DDX41 mutations are involved in familial myelodysplastic syndromes (MDSs) and acute myeloid leukemias (AMLs). We analyzed the prevalence and characteristics of DDX41-related myeloid malignancies in an unselected cohort of 1385 patients with MDS or AML. Using targeted next-generation sequencing, we identified 28 different germline DDX41 variants in 43 unrelated patients, which we classified as causal (n = 21) or unknown significance (n = 7) variants. We focused on the 33 patients having causal variants, representing 2.4% of our cohort. The median age was 69 years; most patients were men (79%). Only 9 patients (27%) had a family history of hematological malignancy, and 15 (46%) had a personal history of cytopenia years before MDS/AML diagnosis. Most patients had a normal karyotype (85%), and the most frequent somatic alteration was a second DDX41 mutation (79%). High-risk DDX41 MDS/AML patients treated with intensive chemotherapy (n = 9) or azacitidine (n = 11) had an overall response rate of 100% or 73%, respectively, with a median overall survival of 5.2 years. Our study highlights that germline DDX41 mutations are relatively common in adult MDS/AML, often without known family history, arguing for systematic screening. Salient features of DDX41-related myeloid malignancies include male preponderance, frequent preexisting cytopenia, additional somatic DDX41 mutation, and relatively good outcome.

8.
Genes Chromosomes Cancer ; 58(8): 595-601, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30779244

RESUMO

Burkitt lymphoma (BL) is characterized by a translocation of the MYC oncogene that leads to the upregulation of MYC expression, cell growth and proliferation. It is well-established that MYC translocation is not a sufficient genetic event to cause BL. Next-generation sequencing has recently provided a comprehensive analysis of the landscape of additional genetic events that contribute to BL lymphomagenesis. Refractory BL or relapsing BL are almost always incurable as a result of the selection of a highly chemoresistant clonally related cell population. Conversely, a few BL recurrence cases arising from clonally distinct tumors have been reported and were associated with a favorable outcome similar to that reported for first-line treatment. Here, we used an unusual case of recurrent but clonally distinct EBV+ BL to highlight the key genetic events that drive BL lymphomagenesis. By whole exome sequencing, we established that ID3 gene was targeted by distinct mutations in the two clonally unrelated diseases, highlighting the crucial role of this gene during lymphomagenesis. We also detected a heterozygous E1021K PIK3CD mutation, thus increasing the spectrum of somatic mutations altering the PI3K signaling pathway in BL. Interestingly, this mutation is known to be associated with activated phosphoinositide 3-kinase delta syndrome (APDS). Finally, we also identified an inherited heterozygous truncating c.5791CT FANCM mutation that may contribute to the unusual recurrence of BL.

12.
Stem Cell Reports ; 11(5): 1075-1091, 2018 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-30449320

RESUMO

Fanconi anemia (FA) causes bone marrow failure early during childhood, and recent studies indicate that a hematopoietic defect could begin in utero. We performed a unique kinetics study of hematopoiesis in Fancg-/- mouse embryos, between the early embryonic day 11.5 (E11.5) to E12.5 developmental window (when the highest level of hematopoietic stem cells [HSC] amplification takes place) and E14.5. This study reveals a deep HSC defect with exhaustion of proliferative and self-renewal capacities very early during development, together with severe FA clinical and biological manifestations, which are mitigated at E14.5 due to compensatory mechanisms that help to ensure survival of Fancg-/- embryos. It also reports that a deep HSC defect is also observed during human FA development, and that human FA fetal liver (FL) HSCs present a transcriptome profile similar to that of mouse E12.5 Fancg-/- FL HSCs. Altogether, our results highlight that early mouse FL could represent a good alternative model for studying Fanconi pathology.


Assuntos
Desenvolvimento Embrionário , Anemia de Fanconi/patologia , Células-Tronco Hematopoéticas/patologia , Animais , Apoptose , Ciclo Celular , Dano ao DNA , Embrião de Mamíferos/patologia , Eritrócitos/metabolismo , Proteína do Grupo de Complementação G da Anemia de Fanconi/deficiência , Proteína do Grupo de Complementação G da Anemia de Fanconi/metabolismo , Feminino , Ontologia Genética , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/metabolismo , Humanos , Fígado/embriologia , Fígado/metabolismo , Camundongos Endogâmicos C57BL , Fenótipo , Placenta/metabolismo , Gravidez , Transcriptoma/genética
13.
Cancer Discov ; 8(12): 1614-1631, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30266814

RESUMO

Deletion of chromosome 6q is a well-recognized abnormality found in poor-prognosis T-cell acute lymphoblastic leukemia (T-ALL). Using integrated genomic approaches, we identified two candidate haploinsufficient genes contiguous at 6q14, SYNCRIP (encoding hnRNP-Q) and SNHG5 (that hosts snoRNAs), both involved in regulating RNA maturation and translation. Combined silencing of both genes, but not of either gene alone, accelerated leukemogeneis in a Tal1/Lmo1/Notch1-driven mouse model, demonstrating the tumor-suppressive nature of the two-gene region. Proteomic and translational profiling of cells in which we engineered a short 6q deletion by CRISPR/Cas9 genome editing indicated decreased ribosome and mitochondrial activities, suggesting that the resulting metabolic changes may regulate tumor progression. Indeed, xenograft experiments showed an increased leukemia-initiating cell activity of primary human leukemic cells upon coextinction of SYNCRIP and SNHG5. Our findings not only elucidate the nature of 6q deletion but also highlight the role of ribosomes and mitochondria in T-ALL tumor progression. SIGNIFICANCE: The oncogenic role of 6q deletion in T-ALL has remained elusive since this chromosomal abnormality was first identified more than 40 years ago. We combined genomic analysis and functional models to show that the codeletion of two contiguous genes at 6q14 enhances malignancy through deregulation of a ribosome-mitochondria axis, suggesting the potential for therapeutic intervention.This article is highlighted in the In This Issue feature, p. 1494.

15.
Eur J Med Chem ; 154: 280-293, 2018 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-29807333

RESUMO

Aggregation of amyloid proteins is currently involved in more than 20 serious human diseases that are actually untreated, such as Alzheimer's disease (AD). Despite many efforts made to target the amyloid cascade in AD, finding an aggregation inhibiting compound and especially modulating early oligomerization remains a relevant and challenging strategy. We report herein the first examples of small and non-peptide mimics of acyclic beta-hairpins, showing an ability to delay the fibrillization of amyloid-ß (Aß1-42) peptide and deeply modify its early oligomerization process. Modifications providing better druggability properties such as increased hydrophilicity and reduced peptidic character were performed. We also demonstrate that an appropriate balance between flexibility and stability of the ß-hairpin must be reached to adapt to the different shape of the various aggregated forms of the amyloid peptide. This strategy can be investigated to target other challenging amyloid proteins.


Assuntos
Peptídeos beta-Amiloides/antagonistas & inibidores , Piperidinas/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Peptídeos beta-Amiloides/metabolismo , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Piperidinas/síntese química , Piperidinas/química , Agregados Proteicos/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/síntese química , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade
16.
Cancer Discov ; 8(5): 616-631, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29496663

RESUMO

Leukemia is caused by the accumulation of multiple genomic lesions in hematopoietic precursor cells. However, how these events cooperate during oncogenic transformation remains poorly understood. We studied the cooperation between activated JAK3/STAT5 signaling and HOXA9 overexpression, two events identified as significantly co-occurring in T-cell acute lymphoblastic leukemia. Expression of mutant JAK3 and HOXA9 led to a rapid development of leukemia originating from multipotent or lymphoid-committed progenitors, with a significant decrease in disease latency compared with JAK3 or HOXA9 alone. Integrated RNA sequencing, chromatin immunoprecipitation sequencing, and Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) revealed that STAT5 and HOXA9 have co-occupancy across the genome, resulting in enhanced STAT5 transcriptional activity and ectopic activation of FOS/JUN (AP1). Our data suggest that oncogenic transcription factors such as HOXA9 provide a fertile ground for specific signaling pathways to thrive, explaining why JAK/STAT pathway mutations accumulate in HOXA9-expressing cells.Significance: The mechanism of oncogene cooperation in cancer development remains poorly characterized. In this study, we model the cooperation between activated JAK/STAT signaling and ectopic HOXA9 expression during T-cell leukemia development. We identify a direct cooperation between STAT5 and HOXA9 at the transcriptional level and identify PIM1 kinase as a possible drug target in mutant JAK/STAT/HOXA9-positive leukemia cases. Cancer Discov; 8(5); 616-31. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 517.


Assuntos
Transformação Celular Neoplásica/metabolismo , Proteínas de Homeodomínio/metabolismo , Janus Quinases/metabolismo , Leucemia/etiologia , Leucemia/metabolismo , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Animais , Transplante de Medula Óssea , Montagem e Desmontagem da Cromatina , Modelos Animais de Doenças , Expressão Gênica , Células-Tronco Hematopoéticas/metabolismo , Proteínas de Homeodomínio/genética , Humanos , Janus Quinase 3/genética , Janus Quinase 3/metabolismo , Janus Quinases/genética , Masculino , Camundongos , Mutação , Leucemia-Linfoma Linfoblástico de Células T Precursoras/etiologia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Ligação Proteica , Fatores de Transcrição STAT/genética , Fator de Transcrição AP-1/metabolismo , Transdução Genética , Transgenes
18.
Oncotarget ; 9(3): 3779-3793, 2018 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-29423082

RESUMO

The Fanconi anemia (FA) pathway is implicated in the repair of DNA interstrand crosslinks (ICL). In this process, it has been shown that FA factors regulate the choice for DNA double strand break repair towards homologous recombination (HR). As this mechanism is impaired in FA deficient cells exposed to crosslinking agents, an inappropriate usage of non-homologous end joining (NHEJ) leads to the accumulation of toxic chromosomal abnormalities. We studied a family with two FANCG patients and found a genetically inherited attenuation of mitomycin C sensitivity resulting in-vitro in an attenuated phenotype for one patient or in increased resistance for two healthy relatives. A heterozygous mutation in ATM was identified in these 3 subjects but was not directly linked to the observed phenotype. However, the attenuation of ICL sensitivity was associated with a reduced recruitment of 53BP1 during the course of ICL repair, and increased HR levels. These results further demonstrate the importance of favoring HR over NHEJ for the survival of cells challenged with ICLs.

19.
Blood ; 131(7): 717-732, 2018 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-29146883

RESUMO

Bone marrow (BM) failure (BMF) in children and young adults is often suspected to be inherited, but in many cases diagnosis remains uncertain. We studied a cohort of 179 patients (from 173 families) with BMF of suspected inherited origin but unresolved diagnosis after medical evaluation and Fanconi anemia exclusion. All patients had cytopenias, and 12.0% presented ≥5% BM blast cells. Median age at genetic evaluation was 11 years; 20.7% of patients were aged ≤2 years and 36.9% were ≥18 years. We analyzed genomic DNA from skin fibroblasts using whole-exome sequencing, and were able to assign a causal or likely causal germ line mutation in 86 patients (48.0%), involving a total of 28 genes. These included genes in familial hematopoietic disorders (GATA2, RUNX1), telomeropathies (TERC, TERT, RTEL1), ribosome disorders (SBDS, DNAJC21, RPL5), and DNA repair deficiency (LIG4). Many patients had an atypical presentation, and the mutated gene was often not clinically suspected. We also found mutations in genes seldom reported in inherited BMF (IBMF), such as SAMD9 and SAMD9L (N = 16 of the 86 patients, 18.6%), MECOM/EVI1 (N = 6, 7.0%), and ERCC6L2 (N = 7, 8.1%), each of which was associated with a distinct natural history; SAMD9 and SAMD9L patients often experienced transient aplasia and monosomy 7, whereas MECOM patients presented early-onset severe aplastic anemia, and ERCC6L2 patients, mild pancytopenia with myelodysplasia. This study broadens the molecular and clinical portrait of IBMF syndromes and sheds light on newly recognized disease entities. Using a high-throughput sequencing screen to implement precision medicine at diagnosis can improve patient management and family counseling.


Assuntos
Doenças da Medula Óssea/genética , Mutação em Linhagem Germinativa , Adolescente , Doenças da Medula Óssea/epidemiologia , Criança , Pré-Escolar , Estudos de Coortes , Análise Mutacional de DNA , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lactente , Recém-Nascido , Masculino , Síndromes Mielodisplásicas/epidemiologia , Síndromes Mielodisplásicas/genética , Sequenciamento Completo do Exoma
20.
Blood ; 131(4): 421-425, 2018 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-29187379

RESUMO

The Janus kinase 3 (JAK3) tyrosine kinase is mutated in 10% to 16% of T-cell acute lymphoblastic leukemia (T-ALL) cases. JAK3 mutants induce constitutive JAK/STAT signaling and cause leukemia when expressed in the bone marrow cells of mice. Surprisingly, we observed that one third of JAK3-mutant T-ALL cases harbor 2 JAK3 mutations, some of which are monoallelic and others that are biallelic. Our data suggest that wild-type JAK3 competes with mutant JAK3 (M511I) for binding to the common γ chain and thereby suppresses its oncogenic potential. We demonstrate that JAK3 (M511I) can increase its limited oncogenic potential through the acquisition of an additional mutation in the mutant JAK3 allele. These double JAK3 mutants show increased STAT5 activation and increased potential to transform primary mouse pro-T cells to interleukin-7-independent growth and were not affected by wild-type JAK3 expression. These data extend our insight into the oncogenic properties of JAK3 mutations and provide an explanation of why progression of JAK3-mutant T-ALL cases can be associated with the accumulation of additional JAK3 mutations.


Assuntos
Janus Quinase 3/genética , Mutação Puntual , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Transdução de Sinais , Alelos , Linhagem Celular Tumoral , Humanos , Janus Quinase 3/metabolismo , Modelos Moleculares , Taxa de Mutação , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo
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