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1.
Cell ; 168(5): 801-816.e13, 2017 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-28215704

RESUMO

DNMT3A mutations occur in ∼25% of acute myeloid leukemia (AML) patients. The most common mutation, DNMT3AR882H, has dominant negative activity that reduces DNA methylation activity by ∼80% in vitro. To understand the contribution of DNMT3A-dependent methylation to leukemogenesis, we performed whole-genome bisulfite sequencing of primary leukemic and non-leukemic cells in patients with or without DNMT3AR882 mutations. Non-leukemic hematopoietic cells with DNMT3AR882H displayed focal methylation loss, suggesting that hypomethylation antedates AML. Although virtually all AMLs with wild-type DNMT3A displayed CpG island hypermethylation, this change was not associated with gene silencing and was essentially absent in AMLs with DNMT3AR882 mutations. Primary hematopoietic stem cells expanded with cytokines were hypermethylated in a DNMT3A-dependent manner, suggesting that hypermethylation may be a response to, rather than a cause of, cellular proliferation. Our findings suggest that hypomethylation is an initiating phenotype in AMLs with DNMT3AR882, while DNMT3A-dependent CpG island hypermethylation is a consequence of AML progression.


Assuntos
Ilhas de CpG , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA , Leucemia Mieloide Aguda/genética , Células da Medula Óssea/patologia , DNA (Citosina-5-)-Metiltransferases/genética , DNA Metiltransferase 3A , Epigênese Genética , Humanos , Leucemia Mieloide Aguda/patologia , Mutação , Análise de Sequência de DNA
2.
Cell ; 165(5): 1134-1146, 2016 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-27156452

RESUMO

Innate lymphoid cells (ILCs) serve as sentinels in mucosal tissues, sensing release of soluble inflammatory mediators, rapidly communicating danger via cytokine secretion, and functioning as guardians of tissue homeostasis. Although ILCs have been extensively studied in model organisms, little is known about these "first responders" in humans, especially their lineage and functional kinships to cytokine-secreting T helper (Th) cell counterparts. Here, we report gene regulatory circuitries for four human ILC-Th counterparts derived from mucosal environments, revealing that each ILC subset diverges as a distinct lineage from Th and circulating natural killer cells but shares circuitry devoted to functional polarization with their Th counterparts. Super-enhancers demarcate cohorts of cell-identity genes in each lineage, uncovering new modes of regulation for signature cytokines, new molecules that likely impart important functions to ILCs, and potential mechanisms for autoimmune disease SNP associations within ILC-Th subsets.


Assuntos
Linfócitos/citologia , Linfócitos/imunologia , Imunidade Adaptativa , Animais , Citocinas/imunologia , Citocinas/metabolismo , Elementos Facilitadores Genéticos , Humanos , Imunidade Inata , Imunidade nas Mucosas , Células Matadoras Naturais , Linfócitos/metabolismo , Camundongos , Tonsila Palatina/citologia , Linfócitos T Auxiliares-Indutores/citologia , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Fatores de Transcrição/metabolismo , Transcriptoma
3.
Cell ; 150(2): 264-78, 2012 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-22817890

RESUMO

Most mutations in cancer genomes are thought to be acquired after the initiating event, which may cause genomic instability and drive clonal evolution. However, for acute myeloid leukemia (AML), normal karyotypes are common, and genomic instability is unusual. To better understand clonal evolution in AML, we sequenced the genomes of M3-AML samples with a known initiating event (PML-RARA) versus the genomes of normal karyotype M1-AML samples and the exomes of hematopoietic stem/progenitor cells (HSPCs) from healthy people. Collectively, the data suggest that most of the mutations found in AML genomes are actually random events that occurred in HSPCs before they acquired the initiating mutation; the mutational history of that cell is "captured" as the clone expands. In many cases, only one or two additional, cooperating mutations are needed to generate the malignant founding clone. Cells from the founding clone can acquire additional cooperating mutations, yielding subclones that can contribute to disease progression and/or relapse.


Assuntos
Evolução Clonal , Leucemia Mieloide Aguda/genética , Mutação , Adulto , Idoso , Análise Mutacional de DNA , Progressão da Doença , Feminino , Estudo de Associação Genômica Ampla , Células-Tronco Hematopoéticas/metabolismo , Humanos , Leucemia Mieloide Aguda/fisiopatologia , Masculino , Pessoa de Meia-Idade , Proteínas de Fusão Oncogênica/genética , Recidiva , Pele/metabolismo , Adulto Jovem
4.
N Engl J Med ; 384(10): 924-935, 2021 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-33704937

RESUMO

BACKGROUND: Genomic analysis is essential for risk stratification in patients with acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS). Whole-genome sequencing is a potential replacement for conventional cytogenetic and sequencing approaches, but its accuracy, feasibility, and clinical utility have not been demonstrated. METHODS: We used a streamlined whole-genome sequencing approach to obtain genomic profiles for 263 patients with myeloid cancers, including 235 patients who had undergone successful cytogenetic analysis. We adapted sample preparation, sequencing, and analysis to detect mutations for risk stratification using existing European Leukemia Network (ELN) guidelines and to minimize turnaround time. We analyzed the performance of whole-genome sequencing by comparing our results with findings from cytogenetic analysis and targeted sequencing. RESULTS: Whole-genome sequencing detected all 40 recurrent translocations and 91 copy-number alterations that had been identified by cytogenetic analysis. In addition, we identified new clinically reportable genomic events in 40 of 235 patients (17.0%). Prospective sequencing of samples obtained from 117 consecutive patients was performed in a median of 5 days and provided new genetic information in 29 patients (24.8%), which changed the risk category for 19 patients (16.2%). Standard AML risk groups, as defined by sequencing results instead of cytogenetic analysis, correlated with clinical outcomes. Whole-genome sequencing was also used to stratify patients who had inconclusive results by cytogenetic analysis into risk groups in which clinical outcomes were measurably different. CONCLUSIONS: In our study, we found that whole-genome sequencing provided rapid and accurate genomic profiling in patients with AML or MDS. Such sequencing also provided a greater diagnostic yield than conventional cytogenetic analysis and more efficient risk stratification on the basis of standard risk categories. (Funded by the Siteman Cancer Research Fund and others.).


Assuntos
Análise Citogenética , Leucemia Mieloide Aguda/genética , Síndromes Mielodisplásicas/genética , Sequenciamento Completo do Genoma , Estudos de Viabilidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Modelos de Riscos Proporcionais , Análise de Sobrevida , Sequenciamento Completo do Genoma/métodos
5.
Blood ; 140(13): 1533-1548, 2022 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-35895896

RESUMO

We have developed a deep-scale proteome and phosphoproteome database from 44 representative acute myeloid leukemia (AML) patients from the LAML TCGA dataset and 6 healthy bone marrow-derived controls. After confirming data quality, we orthogonally validated several previously undescribed features of AML revealed by the proteomic data. We identified examples of posttranscriptionally regulated proteins both globally (ie, in all AML samples) and also in patients with recurrent AML driver mutations. For example, samples with IDH1/2 mutations displayed elevated levels of the 2-oxoglutarate-dependent histone demethylases KDM4A/B/C, despite no changes in messenger RNA levels for these genes; we confirmed this finding in vitro. In samples with NPMc mutations, we identified several nuclear importins with posttranscriptionally increased protein abundance and showed that they interact with NPMc but not wild-type NPM1. We identified 2 cell surface proteins (CD180 and MRC1/CD206) expressed on AML blasts of many patients (but not healthy CD34+ stem/progenitor cells) that could represent novel targets for immunologic therapies and confirmed these targets via flow cytometry. Finally, we detected nearly 30 000 phosphosites in these samples; globally, AML samples were associated with the abnormal phosphorylation of specific residues in PTPN11, STAT3, AKT1, and PRKCD. FLT3-TKD samples were associated with increased phosphorylation of activating tyrosines on the cytoplasmic Src-family tyrosine kinases FGR and HCK and related signaling proteins. PML-RARA-initiated AML samples displayed a unique phosphorylation signature, and TP53-mutant samples showed abundant phosphorylation of serine-183 on TP53 itself. This publicly available database will serve as a foundation for further investigations of protein dysregulation in AML pathogenesis.


Assuntos
Leucemia Mieloide Aguda , Proteínas Nucleares , Histona Desmetilases/metabolismo , Humanos , Histona Desmetilases com o Domínio Jumonji , Carioferinas/genética , Ácidos Cetoglutáricos , Leucemia Mieloide Aguda/patologia , Proteínas de Membrana/genética , Mutação , Proteínas Nucleares/genética , Nucleofosmina , Proteoma/metabolismo , Proteômica , RNA Mensageiro , Serina/genética , Tirosina Quinase 3 Semelhante a fms/genética , Quinases da Família src/metabolismo
6.
Immunity ; 42(1): 186-98, 2015 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-25607463

RESUMO

Most B-cell lymphomas arise in the germinal center (GC), where humoral immune responses evolve from potentially oncogenic cycles of mutation, proliferation, and clonal selection. Although lymphoma gene expression diverges significantly from GC B cells, underlying mechanisms that alter the activities of corresponding regulatory elements (REs) remain elusive. Here we define the complete pathogenic circuitry of human follicular lymphoma (FL), which activates or decommissions REs from normal GC B cells and commandeers enhancers from other lineages. Moreover, independent sets of transcription factors, whose expression was deregulated in FL, targeted commandeered versus decommissioned REs. Our approach revealed two distinct subtypes of low-grade FL, whose pathogenic circuitries resembled GC B or activated B cells. FL-altered enhancers also were enriched for sequence variants, including somatic mutations, which disrupt transcription-factor binding and expression of circuit-linked genes. Thus, the pathogenic regulatory circuitry of FL reveals distinct genetic and epigenetic etiologies for GC B-cell transformation.


Assuntos
Linfócitos B/fisiologia , Redes Reguladoras de Genes , Centro Germinativo/patologia , Linfoma de Células B/genética , Elementos Reguladores de Transcrição/imunologia , Adulto , Idoso , Transformação Celular Neoplásica , Epigênese Genética , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Ativação Linfocitária/genética , Masculino , Pessoa de Meia-Idade , Mutação/genética , Elementos Reguladores de Transcrição/genética , Fatores de Transcrição/metabolismo
7.
Proc Natl Acad Sci U S A ; 118(49)2021 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-34845035

RESUMO

Acute myeloid leukemia (AML) patients rarely have long first remissions (LFRs; >5 y) after standard-of-care chemotherapy, unless classified as favorable risk at presentation. Identification of the mechanisms responsible for long vs. more typical, standard remissions may help to define prognostic determinants for chemotherapy responses. Using exome sequencing, RNA-sequencing, and functional immunologic studies, we characterized 28 normal karyotype (NK)-AML patients with >5 y first remissions after chemotherapy (LFRs) and compared them to a well-matched group of 31 NK-AML patients who relapsed within 2 y (standard first remissions [SFRs]). Our combined analyses indicated that genetic-risk profiling at presentation (as defined by European LeukemiaNet [ELN] 2017 criteria) was not sufficient to explain the outcomes of many SFR cases. Single-cell RNA-sequencing studies of 15 AML samples showed that SFR AML cells differentially expressed many genes associated with immune suppression. The bone marrow of SFR cases had significantly fewer CD4+ Th1 cells; these T cells expressed an exhaustion signature and were resistant to activation by T cell receptor stimulation in the presence of autologous AML cells. T cell activation could be restored by removing the AML cells or blocking the inhibitory major histocompatibility complex class II receptor, LAG3. Most LFR cases did not display these features, suggesting that their AML cells were not as immunosuppressive. These findings were confirmed and extended in an independent set of 50 AML cases representing all ELN 2017 risk groups. AML cell-mediated suppression of CD4+ T cell activation at presentation is strongly associated with unfavorable outcomes in AML patients treated with standard chemotherapy.


Assuntos
Tolerância Imunológica/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/imunologia , Adulto , Linfócitos T CD4-Positivos/imunologia , Feminino , Humanos , Tolerância Imunológica/imunologia , Cariótipo , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Prognóstico , Recidiva , Indução de Remissão , Fatores de Risco , Análise de Sequência de RNA/métodos , Células Th1/imunologia , Transcriptoma/genética , Resultado do Tratamento
8.
PLoS Pathog ; 17(5): e1009577, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-34019588

RESUMO

Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus that causes an aggressive T-cell malignancy and a variety of inflammatory conditions. The integrated provirus includes a single binding site for the epigenomic insulator, CCCTC-binding protein (CTCF), but its function remains unclear. In the current study, a mutant virus was examined that eliminates the CTCF-binding site. The mutation did not disrupt the kinetics and levels of virus gene expression, or establishment of or reactivation from latency. However, the mutation disrupted the epigenetic barrier function, resulting in enhanced DNA CpG methylation downstream of the CTCF binding site on both strands of the integrated provirus and H3K4Me3, H3K36Me3, and H3K27Me3 chromatin modifications both up- and downstream of the site. A majority of clonal cell lines infected with wild type HTLV-1 exhibited increased plus strand gene expression with CTCF knockdown, while expression in mutant HTLV-1 clonal lines was unaffected. These findings indicate that CTCF binding regulates HTLV-1 gene expression, DNA and histone methylation in an integration site dependent fashion.


Assuntos
Epigênese Genética , Genoma Viral/genética , Infecções por HTLV-I/virologia , Vírus Linfotrópico T Tipo 1 Humano/genética , Leucemia de Células T/virologia , Sítios de Ligação , Fator de Ligação a CCCTC/genética , Fator de Ligação a CCCTC/metabolismo , Linhagem Celular , Cromatina/genética , Metilação de DNA , Epigenômica , Vírus Linfotrópico T Tipo 1 Humano/fisiologia , Humanos , Mutação , Integração Viral , Latência Viral/genética
9.
Nucleic Acids Res ; 49(16): e93, 2021 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-34157105

RESUMO

Epigenetic changes, such as aberrant DNA methylation, contribute to cancer clonal expansion and disease progression. However, identifying subpopulation-level changes in a heterogeneous sample remains challenging. Thus, we have developed a computational approach, DXM, to deconvolve the methylation profiles of major allelic subpopulations from the bisulfite sequencing data of a heterogeneous sample. DXM does not require prior knowledge of the number of subpopulations or types of cells to expect. We benchmark DXM's performance and demonstrate improvement over existing methods. We further experimentally validate DXM predicted allelic subpopulation-methylation profiles in four Diffuse Large B-Cell Lymphomas (DLBCLs). Lastly, as proof-of-concept, we apply DXM to a cohort of 31 DLBCLs and relate allelic subpopulation methylation profiles to relapse. We thus demonstrate that DXM can robustly find allelic subpopulation methylation profiles that may contribute to disease progression using bisulfite sequencing data of any heterogeneous sample.


Assuntos
Algoritmos , Metilação de DNA , Linfoma Difuso de Grandes Células B/genética , Análise de Sequência de DNA/métodos , Linhagem Celular Tumoral , Epigenômica/métodos , Epigenômica/normas , Heterogeneidade Genética , Humanos , Análise de Sequência de DNA/normas
10.
J Immunol ; 205(3): 595-607, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32571842

RESUMO

Of the thousands of long noncoding RNAs (lncRNA) identified in lymphocytes, very few have defined functions. In this study, we report the discovery and functional elucidation of a human B cell-specific lncRNA with high levels of expression in three types of B cell cancer and normal B cells. The AC099524.1 gene is upstream of the gene encoding the B cell-specific phospholipase C γ 2 (PLCG2), a B cell-specific enzyme that stimulates intracellular Ca2+ signaling in response to BCR activation. AC099524.1 (B cell-associated lncRNA modulator of BCR-mediated Ca+ signaling [BCALM]) transcripts are localized in the cytoplasm and, as expected, CRISPR/Cas9 knockout of AC099524.1 did not affect PLCG2 mRNA or protein expression. lncRNA interactome, RNA immunoprecipitation, and coimmunoprecipitation studies identified BCALM-interacting proteins in B cells, including phospholipase D 1 (PLD1), and kinase adaptor proteins AKAP9 (AKAP450) and AKAP13 (AKAP-Lbc). These two AKAP proteins form signaling complexes containing protein kinases A and C, which phosphorylate and activate PLD1 to produce phosphatidic acid (PA). BCR stimulation of BCALM-deficient B cells resulted in decreased PLD1 phosphorylation and increased intracellular Ca+ flux relative to wild-type cells. These results suggest that BCALM promotes negative feedback that downmodulates BCR-mediated Ca+ signaling by promoting phosphorylation of PLD1 by AKAP-associated kinases, enhancing production of PA. PA activates SHP-1, which negatively regulates BCR signaling. We propose the name BCALM for B-Cell Associated LncRNA Modulator of BCR-mediated Ca+ signaling. Our findings suggest a new, to our knowledge, paradigm for lncRNA-mediated modulation of lymphocyte activation and signaling, with implications for B cell immune response and BCR-dependent cancers.


Assuntos
Linfócitos B/imunologia , Sinalização do Cálcio/imunologia , RNA Longo não Codificante/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Proteínas de Ancoragem à Quinase A/genética , Proteínas de Ancoragem à Quinase A/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B/citologia , Sinalização do Cálcio/genética , Linhagem Celular , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/imunologia , Fosfolipase C gama/genética , Fosfolipase C gama/imunologia , Fosfolipase D/genética , Fosfolipase D/imunologia , Proteína Tirosina Fosfatase não Receptora Tipo 6/genética , Proteína Tirosina Fosfatase não Receptora Tipo 6/imunologia , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/imunologia , RNA Longo não Codificante/genética , Receptores de Antígenos de Linfócitos B/genética
11.
Mol Cell ; 56(6): 808-18, 2014 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-25435138

RESUMO

The structure of broken DNA ends is a critical determinant of the pathway used for DNA double-strand break (DSB) repair. Here, we develop an approach involving the hairpin capture of DNA end structures (HCoDES), which elucidates chromosomal DNA end structures at single-nucleotide resolution. HCoDES defines structures of physiologic DSBs generated by the RAG endonuclease, as well as those generated by nucleases widely used for genome editing. Analysis of G1 phase cells deficient in H2AX or 53BP1 reveals DNA ends that are frequently resected to form long single-stranded overhangs that can be repaired by mutagenic pathways. In addition to 3' overhangs, many of these DNA ends unexpectedly form long 5' single-stranded overhangs. The divergence in DNA end structures resolved by HCoDES suggests that H2AX and 53BP1 may have distinct activities in end protection. Thus, the high-resolution end structures obtained by HCoDES identify features of DNA end processing during DSB repair.


Assuntos
Cromossomos Humanos/genética , Análise de Sequência de DNA/métodos , Sequência de Bases , Células Cultivadas , Quebras de DNA de Cadeia Dupla , Humanos
12.
N Engl J Med ; 379(24): 2330-2341, 2018 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-30380364

RESUMO

BACKGROUND: As consolidation therapy for acute myeloid leukemia (AML), allogeneic hematopoietic stem-cell transplantation provides a benefit in part by means of an immune-mediated graft-versus-leukemia effect. We hypothesized that the immune-mediated selective pressure imposed by allogeneic transplantation may cause distinct patterns of tumor evolution in relapsed disease. METHODS: We performed enhanced exome sequencing on paired samples obtained at initial presentation with AML and at relapse from 15 patients who had a relapse after hematopoietic stem-cell transplantation (with transplants from an HLA-matched sibling, HLA-matched unrelated donor, or HLA-mismatched unrelated donor) and from 20 patients who had a relapse after chemotherapy. We performed RNA sequencing and flow cytometry on a subgroup of these samples and on additional samples for validation. RESULTS: On exome sequencing, the spectrum of gained and lost mutations observed with relapse after transplantation was similar to the spectrum observed with relapse after chemotherapy. Specifically, relapse after transplantation was not associated with the acquisition of previously unknown AML-specific mutations or structural variations in immune-related genes. In contrast, RNA sequencing of samples obtained at relapse after transplantation revealed dysregulation of pathways involved in adaptive and innate immunity, including down-regulation of major histocompatibility complex (MHC) class II genes ( HLA-DPA1, HLA-DPB1, HLA-DQB1, and HLA-DRB1) to levels that were 3 to 12 times lower than the levels seen in paired samples obtained at presentation. Flow cytometry and immunohistochemical analysis confirmed decreased expression of MHC class II at relapse in 17 of 34 patients who had a relapse after transplantation. Evidence suggested that interferon-γ treatment could rapidly reverse this phenotype in AML blasts in vitro. CONCLUSIONS: AML relapse after transplantation was not associated with the acquisition of relapse-specific mutations in immune-related genes. However, it was associated with dysregulation of pathways that may influence immune function, including down-regulation of MHC class II genes, which are involved in antigen presentation. These epigenetic changes may be reversible with appropriate therapy. (Funded by the National Cancer Institute and others.).


Assuntos
Genes MHC da Classe II/fisiologia , Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/imunologia , Mutação , Adolescente , Adulto , Idoso , Regulação para Baixo , Epigênese Genética , Feminino , Citometria de Fluxo , Humanos , Imunidade/genética , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Masculino , Pessoa de Meia-Idade , RNA Neoplásico/análise , Recidiva , Análise de Sequência de RNA , Linfócitos T/imunologia , Transplante Homólogo , Sequenciamento do Exoma
13.
BMC Genomics ; 20(1): 137, 2019 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-30767760

RESUMO

BACKGROUND: Long non-coding RNAs (lncRNAs) exhibit remarkable cell-type specificity and disease association. LncRNA's functional versatility includes epigenetic modification, nuclear domain organization, transcriptional control, regulation of RNA splicing and translation, and modulation of protein activity. However, most lncRNAs remain uncharacterized due to a shortage of predictive tools available to guide functional experiments. RESULTS: To address this gap for lymphoma-associated lncRNAs identified in our studies, we developed a new computational method, Predicting LncRNA Activity through Integrative Data-driven 'Omics and Heuristics (PLAIDOH), which has several unique features not found in other methods. PLAIDOH integrates transcriptome, subcellular localization, enhancer landscape, genome architecture, chromatin interaction, and RNA-binding (eCLIP) data and generates statistically defined output scores. PLAIDOH's approach identifies and ranks functional connections between individual lncRNA, coding gene, and protein pairs using enhancer, transcript cis-regulatory, and RNA-binding protein interactome scores that predict the relative likelihood of these different lncRNA functions. When applied to 'omics datasets that we collected from lymphoma patients, or to publicly available cancer (TCGA) or ENCODE datasets, PLAIDOH identified and prioritized well-known lncRNA-target gene regulatory pairs (e.g., HOTAIR and HOX genes, PVT1 and MYC), validated hits in multiple lncRNA-targeted CRISPR screens, and lncRNA-protein binding partners (e.g., NEAT1 and NONO). Importantly, PLAIDOH also identified novel putative functional interactions, including one lymphoma-associated lncRNA based on analysis of data from our human lymphoma study. We validated PLAIDOH's predictions for this lncRNA using knock-down and knock-out experiments in lymphoma cell models. CONCLUSIONS: Our study demonstrates that we have developed a new method for the prediction and ranking of functional connections between individual lncRNA, coding gene, and protein pairs, which were validated by genetic experiments and comparison to published CRISPR screens. PLAIDOH expedites validation and follow-on mechanistic studies of lncRNAs in any biological system. It is available at https://github.com/sarahpyfrom/PLAIDOH .


Assuntos
Biologia Computacional/métodos , Linfoma não Hodgkin/genética , RNA Longo não Codificante/genética , RNA Neoplásico/genética , Algoritmos , Processamento Alternativo , Linfócitos B/metabolismo , Linhagem Celular Tumoral , Bases de Dados Genéticas , Éxons , Código das Histonas/genética , Humanos , Proteínas de Ligação a RNA/análise , Transcrição Gênica , Transcriptoma
14.
Trends Genet ; 31(12): 720-731, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26604030

RESUMO

B cell lymphomas (BCLs) are characterized by widespread deregulation of gene expression compared with their normal B cell counterparts. Recent epigenomic studies defined cis-regulatory elements (REs) whose activities are altered in BCL to drive some of these pathogenic expression changes. During transformation, multiple mechanisms are employed to alter RE activities, including perturbations in the function of chromatin modifiers, which can lead to revision of the B cell epigenome. Inherited and somatic variants also alter RE function via disruption of transcription factor (TF) binding. Aberrant expression of noncoding RNAs (ncRNAs) deregulates genes involved in B cell differentiation via direct repression and post-transcriptional targeting. These discoveries have established epigenetic etiologies for B cell transformation that are being exploited in novel therapeutic approaches.


Assuntos
Redes Reguladoras de Genes , Linfoma de Células B/genética , Animais , Epigênese Genética , Humanos , Camundongos
15.
J Immunol ; 196(11): 4805-13, 2016 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-27183590

RESUMO

It is now clear that recognition of nascent tumors by the immune system is critical for survival of the host against cancer. During cancer immunoediting, the ability of the tumor to escape immune recognition is important for tumor development. The immune system recognizes tumors via the presence of classical Ags and also by conserved innate mechanisms. One of these mechanisms is the NKG2D receptor that recognizes ligands whose expression is induced by cell transformation. In this study, we show that in NKG2D receptor-deficient mice, increasing numbers of B cells begin to express NKG2D ligands as they age. Their absence in wild-type mice suggests that these cells are normally cleared by NKG2D-expressing cells. NKG2D-deficient mice and mice constitutively expressing NKG2D ligands had increased incidence of B cell tumors, confirming that the inability to clear NKG2D ligand-expressing cells was important in tumor suppression and that NKG2D ligand expression is a marker of nascent tumors. Supporting a role for NKG2D ligand expression in controlling the progression of early-stage B cell lymphomas in humans, we found higher expression of a microRNA that inhibits human NKG2D ligand expression in tumor cells from high-grade compared with low-grade follicular lymphoma patients.


Assuntos
Linfoma de Células B/metabolismo , Linfoma de Células B/patologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Animais , Progressão da Doença , Humanos , Ligantes , Linfoma de Células B/genética , Linfoma de Células B/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , MicroRNAs/genética , Subfamília K de Receptores Semelhantes a Lectina de Células NK/antagonistas & inibidores , Subfamília K de Receptores Semelhantes a Lectina de Células NK/deficiência , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética
16.
Nature ; 481(7382): 506-10, 2012 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-22237025

RESUMO

Most patients with acute myeloid leukaemia (AML) die from progressive disease after relapse, which is associated with clonal evolution at the cytogenetic level. To determine the mutational spectrum associated with relapse, we sequenced the primary tumour and relapse genomes from eight AML patients, and validated hundreds of somatic mutations using deep sequencing; this allowed us to define clonality and clonal evolution patterns precisely at relapse. In addition to discovering novel, recurrently mutated genes (for example, WAC, SMC3, DIS3, DDX41 and DAXX) in AML, we also found two major clonal evolution patterns during AML relapse: (1) the founding clone in the primary tumour gained mutations and evolved into the relapse clone, or (2) a subclone of the founding clone survived initial therapy, gained additional mutations and expanded at relapse. In all cases, chemotherapy failed to eradicate the founding clone. The comparison of relapse-specific versus primary tumour mutations in all eight cases revealed an increase in transversions, probably due to DNA damage caused by cytotoxic chemotherapy. These data demonstrate that AML relapse is associated with the addition of new mutations and clonal evolution, which is shaped, in part, by the chemotherapy that the patients receive to establish and maintain remissions.


Assuntos
Evolução Clonal/genética , Genoma Humano/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Antineoplásicos/efeitos adversos , Antineoplásicos/uso terapêutico , Células Clonais/efeitos dos fármacos , Células Clonais/metabolismo , Células Clonais/patologia , Dano ao DNA/efeitos dos fármacos , Análise Mutacional de DNA , Genes Neoplásicos/genética , Genoma Humano/efeitos dos fármacos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Mutagênese/efeitos dos fármacos , Mutagênese/genética , Recidiva , Reprodutibilidade dos Testes
17.
Hematol Oncol ; 35(4): 397-407, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28378425

RESUMO

The 13th International Conference on Malignant Lymphoma held in Lugano in June 2015 was preceded by a closed workshop (organized in collaboration with the American Association for Cancer Research and the European School of Oncology) with the aim of developing an up-to-date understanding of the biology of follicular lymphoma and the clinical implications of new findings in the field. Discussed topics included the mutational spectrum at diagnosis, the clinical correlates of genetic and epigenetic alterations, the mechanisms of clonal evolution and histological transformation, the cross talk between tumor cells and microenvironment, and the development of novel treatments. This report represents a summary of the workshop.


Assuntos
Linfoma Folicular , Feminino , História do Século XXI , Humanos , Masculino , Suíça
18.
N Engl J Med ; 368(22): 2059-74, 2013 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-23634996

RESUMO

BACKGROUND: Many mutations that contribute to the pathogenesis of acute myeloid leukemia (AML) are undefined. The relationships between patterns of mutations and epigenetic phenotypes are not yet clear. METHODS: We analyzed the genomes of 200 clinically annotated adult cases of de novo AML, using either whole-genome sequencing (50 cases) or whole-exome sequencing (150 cases), along with RNA and microRNA sequencing and DNA-methylation analysis. RESULTS: AML genomes have fewer mutations than most other adult cancers, with an average of only 13 mutations found in genes. Of these, an average of 5 are in genes that are recurrently mutated in AML. A total of 23 genes were significantly mutated, and another 237 were mutated in two or more samples. Nearly all samples had at least 1 nonsynonymous mutation in one of nine categories of genes that are almost certainly relevant for pathogenesis, including transcription-factor fusions (18% of cases), the gene encoding nucleophosmin (NPM1) (27%), tumor-suppressor genes (16%), DNA-methylation-related genes (44%), signaling genes (59%), chromatin-modifying genes (30%), myeloid transcription-factor genes (22%), cohesin-complex genes (13%), and spliceosome-complex genes (14%). Patterns of cooperation and mutual exclusivity suggested strong biologic relationships among several of the genes and categories. CONCLUSIONS: We identified at least one potential driver mutation in nearly all AML samples and found that a complex interplay of genetic events contributes to AML pathogenesis in individual patients. The databases from this study are widely available to serve as a foundation for further investigations of AML pathogenesis, classification, and risk stratification. (Funded by the National Institutes of Health.).


Assuntos
Leucemia Mieloide Aguda/genética , Mutação , Adulto , Ilhas de CpG , Metilação de DNA , Epigenômica , Feminino , Expressão Gênica , Fusão Gênica , Genoma Humano , Humanos , Leucemia Mieloide Aguda/classificação , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Nucleofosmina , Análise de Sequência de DNA/métodos
19.
JAMA ; 314(8): 811-22, 2015 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-26305651

RESUMO

IMPORTANCE: Tests that predict outcomes for patients with acute myeloid leukemia (AML) are imprecise, especially for those with intermediate risk AML. OBJECTIVES: To determine whether genomic approaches can provide novel prognostic information for adult patients with de novo AML. DESIGN, SETTING, AND PARTICIPANTS: Whole-genome or exome sequencing was performed on samples obtained at disease presentation from 71 patients with AML (mean age, 50.8 years) treated with standard induction chemotherapy at a single site starting in March 2002, with follow-up through January 2015. In addition, deep digital sequencing was performed on paired diagnosis and remission samples from 50 patients (including 32 with intermediate-risk AML), approximately 30 days after successful induction therapy. Twenty-five of the 50 were from the cohort of 71 patients, and 25 were new, additional cases. EXPOSURES: Whole-genome or exome sequencing and targeted deep sequencing. Risk of identification based on genetic data. MAIN OUTCOMES AND MEASURES: Mutation patterns (including clearance of leukemia-associated variants after chemotherapy) and their association with event-free survival and overall survival. RESULTS: Analysis of comprehensive genomic data from the 71 patients did not improve outcome assessment over current standard-of-care metrics. In an analysis of 50 patients with both presentation and documented remission samples, 24 (48%) had persistent leukemia-associated mutations in at least 5% of bone marrow cells at remission. The 24 with persistent mutations had significantly reduced event-free and overall survival vs the 26 who cleared all mutations. Patients with intermediate cytogenetic risk profiles had similar findings. [table: see text]. CONCLUSIONS AND RELEVANCE: The detection of persistent leukemia-associated mutations in at least 5% of bone marrow cells in day 30 remission samples was associated with a significantly increased risk of relapse, and reduced overall survival. These data suggest that this genomic approach may improve risk stratification for patients with AML.


Assuntos
Quimioterapia de Indução , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Mutação , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Medula Óssea , Citarabina/administração & dosagem , Daunorrubicina/administração & dosagem , Intervalo Livre de Doença , Feminino , Genoma Humano , Humanos , Idarubicina/administração & dosagem , Leucemia Mieloide Aguda/mortalidade , Masculino , MicroRNAs/análise , Pessoa de Meia-Idade , Avaliação de Resultados em Cuidados de Saúde , Polimorfismo Genético , Prognóstico , RNA Mensageiro/análise , Recidiva , Análise de Sequência de RNA/métodos
20.
Nature ; 456(7218): 66-72, 2008 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-18987736

RESUMO

Acute myeloid leukaemia is a highly malignant haematopoietic tumour that affects about 13,000 adults in the United States each year. The treatment of this disease has changed little in the past two decades, because most of the genetic events that initiate the disease remain undiscovered. Whole-genome sequencing is now possible at a reasonable cost and timeframe to use this approach for the unbiased discovery of tumour-specific somatic mutations that alter the protein-coding genes. Here we present the results obtained from sequencing a typical acute myeloid leukaemia genome, and its matched normal counterpart obtained from the same patient's skin. We discovered ten genes with acquired mutations; two were previously described mutations that are thought to contribute to tumour progression, and eight were new mutations present in virtually all tumour cells at presentation and relapse, the function of which is not yet known. Our study establishes whole-genome sequencing as an unbiased method for discovering cancer-initiating mutations in previously unidentified genes that may respond to targeted therapies.


Assuntos
Regulação Neoplásica da Expressão Gênica/genética , Genoma Humano/genética , Leucemia Mieloide Aguda/genética , Estudos de Casos e Controles , Progressão da Doença , Perfilação da Expressão Gênica , Genômica , Humanos , Mutagênese Insercional , Mutação , Polimorfismo de Nucleotídeo Único , Recidiva , Análise de Sequência de DNA , Deleção de Sequência , Pele/metabolismo
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