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Palatine tonsils are secondary lymphoid organs (SLOs) representing the first line of immunological defense against inhaled or ingested pathogens. We generated an atlas of the human tonsil composed of >556,000 cells profiled across five different data modalities, including single-cell transcriptome, epigenome, proteome, and immune repertoire sequencing, as well as spatial transcriptomics. This census identified 121 cell types and states, defined developmental trajectories, and enabled an understanding of the functional units of the tonsil. Exemplarily, we stratified myeloid slan-like subtypes, established a BCL6 enhancer as locally active in follicle-associated T and B cells, and identified SIX5 as putative transcriptional regulator of plasma cell maturation. Analyses of a validation cohort confirmed the presence, annotation, and markers of tonsillar cell types and provided evidence of age-related compositional shifts. We demonstrate the value of this resource by annotating cells from B cell-derived mantle cell lymphomas, linking transcriptional heterogeneity to normal B cell differentiation states of the human tonsil.
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Linfócitos B , Tonsila Palatina , Humanos , Adulto , Linfócitos B/metabolismoRESUMO
ABSTRACT: Plasma cells (PCs) are highly specialized cells representing the end stage of B-cell differentiation. We have shown that PC differentiation can be reproduced in vitro using elaborate culture systems. The molecular changes occurring during PC differentiation are recapitulated in this in vitro differentiation model. However, a major challenge exists to decipher the spatiotemporal epigenetic and transcriptional programs that drive the early stages of PC differentiation. We combined single cell (sc) RNA sequencing (RNA-seq) and assay for transposase-accessible chromatin with high throughput sequencing (scATAC-seq) to decipher the trajectories involved in PC differentiation. ScRNA-seq experiments revealed a strong heterogeneity of the preplasmablastic and plasmablastic stages. Among genes that were commonly identified using scATAC-seq and scRNA-seq, we identified several transcription factors with significant stage specific potential importance in PC differentiation. Interestingly, differentially accessible peaks characterizing the preplasmablastic stage were enriched in motifs of BATF3, FOS and BATF, belonging to activating protein 1 (AP-1) transcription factor family that may represent key transcriptional nodes involved in PC differentiation. Integration of transcriptomic and epigenetic data at the single cell level revealed that a population of preplasmablasts had already undergone epigenetic remodeling related to PC profile together with unfolded protein response activation and are committed to differentiate in PC. These results and the supporting data generated with our in vitro PC differentiation model provide a unique resource for the identification of molecular circuits that are crucial for early and mature PC maturation and biological functions. These data thus provide critical insights into epigenetic- and transcription-mediated reprogramming events that sustain PC differentiation.
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Diferenciação Celular , Cromatina , Perfilação da Expressão Gênica , Plasmócitos , Análise de Célula Única , Humanos , Diferenciação Celular/genética , Plasmócitos/metabolismo , Plasmócitos/citologia , Análise de Célula Única/métodos , Cromatina/metabolismo , Cromatina/genética , Transcriptoma , Epigênese Genética , Células CultivadasRESUMO
The histone H3K27 demethylase KDM6A is a tumor suppressor in multiple cancers, including multiple myeloma (MM). We created isogenic MM cells disrupted for KDM6A and tagged the endogenous protein to facilitate genome wide studies. KDM6A binds genes associated with immune recognition and cytokine signaling. Most importantly, KDM6A binds and activates NLRC5 and CIITA encoding regulators of Major Histocompatibility Complex (MHC) genes. Patient data indicate that NLRC5 and CIITA, are downregulated in MM with low KDM6A expression. Chromatin analysis shows that KDM6A binds poised and active enhancers and KDM6A loss led to decreased H3K27ac at enhancers, increased H3K27me3 levels in body of genes bound by KDM6A and decreased gene expression. Reestablishing histone acetylation with an HDAC3 inhibitor leads to upregulation of MHC expression, offering a strategy to restore immunogenicity of KDM6A deficient tumors. Loss of Kdm6a in murine RAS-transformed fibroblasts led to increased growth in vivo associated with decreased T cell infiltration.
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ABSTRACT: Sterile alpha motif and histidine-aspartate (HD) domain-containing protein 1 (SAMHD1) is a deoxynucleoside triphosphate triphosphohydrolase with ara-CTPase activity that confers cytarabine (ara-C) resistance in several hematological malignancies. Targeting SAMHD1's ara-CTPase activity has recently been demonstrated to enhance ara-C efficacy in acute myeloid leukemia. Here, we identify the transcription factor SRY-related HMG-box containing protein 11 (SOX11) as a novel direct binding partner and first known endogenous inhibitor of SAMHD1. SOX11 is aberrantly expressed not only in mantle cell lymphoma (MCL), but also in some Burkitt lymphomas. Coimmunoprecipitation of SOX11 followed by mass spectrometry in MCL cell lines identified SAMHD1 as the top SOX11 interaction partner, which was validated by proximity ligation assay. In vitro, SAMHD1 bound to the HMG box of SOX11 with low-micromolar affinity. In situ crosslinking studies further indicated that SOX11-SAMHD1 binding resulted in a reduced tetramerization of SAMHD1. Functionally, expression of SOX11 inhibited SAMHD1 ara-CTPase activity in a dose-dependent manner resulting in ara-C sensitization in cell lines and in a SOX11-inducible mouse model of MCL. In SOX11-negative MCL, SOX11-mediated ara-CTPase inhibition could be mimicked by adding the recently identified SAMHD1 inhibitor hydroxyurea. Taken together, our results identify SOX11 as a novel SAMHD1 interaction partner and its first known endogenous inhibitor with potentially important implications for clinical therapy stratification.
Assuntos
Linfoma de Célula do Manto , Proteína 1 com Domínio SAM e Domínio HD , Fatores de Transcrição SOXC , Linfoma de Célula do Manto/metabolismo , Linfoma de Célula do Manto/patologia , Linfoma de Célula do Manto/tratamento farmacológico , Linfoma de Célula do Manto/genética , Humanos , Proteína 1 com Domínio SAM e Domínio HD/metabolismo , Proteína 1 com Domínio SAM e Domínio HD/genética , Animais , Camundongos , Fatores de Transcrição SOXC/metabolismo , Fatores de Transcrição SOXC/genética , Ligação Proteica , Linhagem Celular Tumoral , Citarabina/farmacologiaRESUMO
The chromatin activation landscape of chronic lymphocytic leukemia (CLL) with stereotyped B-cell receptor immunoglobulin is currently unknown. In this study, we report the results of a whole-genome chromatin profiling of histone 3 lysine 27 acetylation of 22 CLLs from major subsets, which were compared against nonstereotyped CLLs and normal B-cell subpopulations. Although subsets 1, 2, and 4 did not differ much from their nonstereotyped CLL counterparts, subset 8 displayed a remarkably distinct chromatin activation profile. In particular, we identified 209 de novo active regulatory elements in this subset, which showed similar patterns with U-CLLs undergoing Richter transformation. These regions were enriched for binding sites of 9 overexpressed transcription factors. In 78 of 209 regions, we identified 113 candidate overexpressed target genes, 11 regions being associated with more than 2 adjacent genes. These included blocks of up to 7 genes, suggesting local coupregulation within the same genome compartment. Our findings further underscore the uniqueness of subset 8 CLL, notable for the highest risk of Richter's transformation among all CLLs and provide additional clues to decipher the molecular basis of its clinical behavior.
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Leucemia Linfocítica Crônica de Células B , Linfoma Difuso de Grandes Células B , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Cromatina/genética , Linfócitos B , Receptores de Antígenos de Linfócitos B/genéticaRESUMO
Mantle cell lymphoma (MCL) is clinically and biologically heterogeneous. While various prognostic features have been proposed, none currently impact therapy selection, particularly in older patients, for whom treatment is primarily dictated by age and comorbidities. Herein, we undertook a comprehensive comparison of clinicopathological features in a cohort of patients 60 years and older, uniformly treated with bendamustine and rituximab, with a median survival of >8 years. The strongest prognostic indicators in this cohort were a high-risk call by a simplified MCL international prognostic index (s-MIPI) (HR: 3.32, 95% CI: 1.65-6.68 compared to low risk), a high-risk call by MCL35 (HR: 10.34, 95% CI: 2.37-45.20 compared to low risk) and blastoid cytology (HR: 4.21, 95% CR: 1.92-9.22 compared to classic). Patients called high risk by both the s-MIPI and MCL35 had the most dismal prognosis (HR: 11.58, 95% CI: 4.10-32.72), while those with high risk by either had a moderate but clinically relevant prognosis (HR: 2.95, 95% CI: 1.49-5.82). A robust assay to assess proliferation, such as MCL35, along with stringent guidelines for cytological evaluation of MCL, in combination with MIPI, may be a strong path to risk-stratify older MCL patients in future clinical trials.
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Linfoma de Célula do Manto , Adulto , Humanos , Idoso , Linfoma de Célula do Manto/patologia , Rituximab/efeitos adversos , Cloridrato de Bendamustina/uso terapêutico , Biomarcadores , Prognóstico , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversosRESUMO
Chronic lymphocytic leukaemia (CLL) is not only characterised by driver genetic alterations but by extensive epigenetic changes. Over the last decade, epigenomic studies have described the DNA methylome, chromatin accessibility, histone modifications and the three-dimensional (3D) genome architecture of CLL. Beyond its regulatory role, the DNA methylome contains imprints of the cellular origin and proliferative history of CLL cells. These two aspects are strong independent prognostic factors. Integrative analyses of chromatin marks have uncovered novel regulatory elements and altered transcription factor networks as non-genetic means mediating gene deregulation in CLL. Additionally, CLL cells display a disease-specific pattern of 3D genome interactions. From the technological perspective, we are currently witnessing a transition from bulk omics to single-cell analyses. This review aims at summarising the major findings from the epigenomics field as well as providing a prospect of the present and future of single-cell analyses in CLL.
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Leucemia Linfocítica Crônica de Células B , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Epigenômica , Metilação de DNA , Epigênese Genética , Cromatina/genéticaRESUMO
BACKGROUND: Cortical spreading depolarization, the cause of migraine aura, is a short-lasting depolarization wave that moves across the brain cortex, transiently suppressing neuronal activity. Prophylactic treatments for migraine, such as topiramate or valproate, reduce the number of cortical spreading depression events in rodents. OBJECTIVE: To investigate whether cortical spreading depolarization with and without chronic treatment with topiramate or valproate affect the DNA methylation of the cortex. METHODS: Sprague-Dawley rats were intraperitoneally injected with saline, topiramate or valproate for four weeks when cortical spreading depolarization were induced and genome-wide DNA methylation was performed in the cortex of six rats per group. RESULTS: The DNA methylation profile of the cortex was significantly modified after cortical spreading depolarization, with and without topiramate or valproate. Interestingly, topiramate reduced by almost 50% the number of differentially methylated regions, whereas valproate increased them by 17%, when comparing to the non-treated group after cortical spreading depolarization induction. The majority of the differentially methylated regions lay within intragenic regions, and the analyses of functional group over-representation retrieved several enriched functions, including functions related to protein processing in the cortical spreading depolarization without treatment group; functions related to metabolic processes in the cortical spreading depolarization with topiramate group; and functions related to synapse and ErbB, MAPK or retrograde endocannabinoid signaling in the cortical spreading depolarization with valproate group. CONCLUSIONS: Our results may provide insights into the underlying physiological mechanisms of migraine with aura and emphasize the role of epigenetics in migraine susceptibility.
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Depressão Alastrante da Atividade Elétrica Cortical , Transtornos de Enxaqueca , Ratos , Animais , Ácido Valproico/farmacologia , Ácido Valproico/uso terapêutico , Topiramato/farmacologia , Topiramato/uso terapêutico , Ratos Sprague-Dawley , Metilação de DNA , Transtornos de Enxaqueca/tratamento farmacológico , Transtornos de Enxaqueca/genética , Depressão Alastrante da Atividade Elétrica Cortical/fisiologiaRESUMO
Chronic lymphocytic leukemia (CLL) has heterogeneous clinical and biological behavior. Whole-genome and -exome sequencing has contributed to the characterization of the mutational spectrum of the disease, but the underlying transcriptional profile is still poorly understood. We have performed deep RNA sequencing in different subpopulations of normal B-lymphocytes and CLL cells from a cohort of 98 patients, and characterized the CLL transcriptional landscape with unprecedented resolution. We detected thousands of transcriptional elements differentially expressed between the CLL and normal B cells, including protein-coding genes, noncoding RNAs, and pseudogenes. Transposable elements are globally derepressed in CLL cells. In addition, two thousand genes-most of which are not differentially expressed-exhibit CLL-specific splicing patterns. Genes involved in metabolic pathways showed higher expression in CLL, while genes related to spliceosome, proteasome, and ribosome were among the most down-regulated in CLL. Clustering of the CLL samples according to RNA-seq derived gene expression levels unveiled two robust molecular subgroups, C1 and C2. C1/C2 subgroups and the mutational status of the immunoglobulin heavy variable (IGHV) region were the only independent variables in predicting time to treatment in a multivariate analysis with main clinico-biological features. This subdivision was validated in an independent cohort of patients monitored through DNA microarrays. Further analysis shows that B-cell receptor (BCR) activation in the microenvironment of the lymph node may be at the origin of the C1/C2 differences.
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Linfócitos B , Regulação Neoplásica da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Leucemia Linfocítica Crônica de Células B/genética , Idoso , Sequência de Bases , Feminino , Perfilação da Expressão Gênica , Humanos , Região Variável de Imunoglobulina , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Pessoa de Meia-Idade , Mutação , Ribossomos/genética , Spliceossomos/genéticaRESUMO
Deregulation of the epigenome is an important pathogenetic mechanism in acute lymphoblastic leukemia (ALL) with lysine (K)-specific methyltransferase 2A rearrangement (KMT2Ar). We performed array-based DNA methylation profiling of KMT2Ar ALL cells from 26 children in comparison to normal B-cell precursors. Significant changes in DNA methylation in KMT2Ar ALL were identified in 2,545 CpG loci, influenced by age and the translocation partners AFF1 and MLLT1. In KMT2Ar ALL, DNA methylation loss was enriched at enhancers and for certain transcription factor binding sites such as BCL11A, EBF, and MEF2A. In summary, DNA methylation changes in KMT2Ar ALL target enhancers, genes involved in leukemogenesis and normal hematopoiesis, as well as transcription factor networks.
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Biomarcadores Tumorais/genética , Metilação de DNA , Perfilação da Expressão Gênica , Rearranjo Gênico , Histona-Lisina N-Metiltransferase/genética , Proteína de Leucina Linfoide-Mieloide/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Linfócitos B/metabolismo , Estudos de Casos e Controles , Hibridização Genômica Comparativa , Feminino , Seguimentos , Humanos , Lactente , Masculino , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia , Prognóstico , Regiões Promotoras Genéticas/genéticaRESUMO
Most of the studies characterizing DNA methylation patterns have been restricted to particular genomic loci in a limited number of human samples and pathological conditions. Herein, we present a compromise between an extremely comprehensive study of a human sample population with an intermediate level of resolution of CpGs at the genomic level. We obtained a DNA methylation fingerprint of 1628 human samples in which we interrogated 1505 CpG sites. The DNA methylation patterns revealed show this epigenetic mark to be critical in tissue-type definition and stemness, particularly around transcription start sites that are not within a CpG island. For disease, the generated DNA methylation fingerprints show that, during tumorigenesis, human cancer cells underwent a progressive gain of promoter CpG-island hypermethylation and a loss of CpG methylation in non-CpG-island promoters. Although transformed cells are those in which DNA methylation disruption is more obvious, we observed that other common human diseases, such as neurological and autoimmune disorders, had their own distinct DNA methylation profiles. Most importantly, we provide proof of principle that the DNA methylation fingerprints obtained might be useful for translational purposes by showing that we are able to identify the tumor type origin of cancers of unknown primary origin (CUPs). Thus, the DNA methylation patterns identified across the largest spectrum of samples, tissues, and diseases reported to date constitute a baseline for developing higher-resolution DNA methylation maps and provide important clues concerning the contribution of CpG methylation to tissue identity and its changes in the most prevalent human diseases.
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Metilação de DNA , Linhagem Celular , Transformação Celular Neoplásica/genética , Análise por Conglomerados , Ilhas de CpG , Epigenômica/métodos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Neoplasias/genéticaRESUMO
Mantle cell lymphoma (MCL) is one of the most aggressive lymphoid neoplasms whose pathogenesis is not fully understood. The neural transcription factor SOX11 is overexpressed in most MCL but is not detected in other mature B-cell lymphomas or normal lymphoid cells. The specific expression of SOX11 in MCL suggests that it may be an important element in the development of this tumor, but its potential function is not known. Here, we show that SOX11 promotes tumor growth in a MCL-xenotransplant mouse model. Using chromatin immunoprecipitation microarray analysis combined with gene expression profiling upon SOX11 knockdown, we identify target genes and transcriptional programs regulated by SOX11 including the block of mature B-cell differentiation, modulation of cell cycle, apoptosis, and stem cell development. PAX5 emerges as one of the major SOX11 direct targets. SOX11 silencing downregulates PAX5, induces BLIMP1 expression, and promotes the shift from a mature B cell into the initial plasmacytic differentiation phenotype in both primary tumor cells and an in vitro model. Our results suggest that SOX11 contributes to tumor development by altering the terminal B-cell differentiation program of MCL and provide perspectives that may have clinical implications in the diagnosis and design of new therapeutic strategies.
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Linfócitos B/fisiologia , Diferenciação Celular/genética , Linfoma de Célula do Manto/genética , Fator de Transcrição PAX5/genética , Fatores de Transcrição SOXC/fisiologia , Animais , Linfócitos B/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Linfoma de Célula do Manto/patologia , Linfoma de Célula do Manto/fisiopatologia , Linfopoese/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos SCID , Invasividade Neoplásica , Fator de Transcrição PAX5/metabolismo , Fatores de Transcrição SOXC/genética , Fatores de Transcrição SOXC/metabolismo , Transplante HeterólogoRESUMO
Lymphoid neoplasms represent a heterogeneous group of disease entities and subtypes with markedly different molecular and clinical features. Beyond genetic alterations, lymphoid tumors also show widespread epigenomic changes. These severely affect the levels and distribution of DNA methylation, histone modifications, chromatin accessibility, and three-dimensional genome interactions. DNA methylation stands out as a tracer of cell identity and memory, as B cell neoplasms show epigenetic imprints of their cellular origin and proliferative history, which can be quantified by an epigenetic mitotic clock. Chromatin-associated marks are informative to uncover altered regulatory regions and transcription factor networks contributing to the development of distinct lymphoid tumors. Tumor-intrinsic epigenetic and genetic aberrations cooperate and interact with microenvironmental cells to shape the transcriptome at different phases of lymphoma evolution, and intraclonal heterogeneity can now be characterized by single-cell profiling. Finally, epigenetics offers multiple clinical applications, including powerful diagnostic and prognostic biomarkers as well as therapeutic targets.
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Epigenômica , Linfoma , Humanos , Cromatina , Epigênese Genética , MutaçãoRESUMO
Chronic lymphocytic leukemia (CLL) is characterized by widespread alterations in the genetic and epigenetic landscapes which seem to underlie the variable clinical manifestations observed in patients. Over the last decade, epigenomic studies have described the whole-genome maps of DNA methylation and chromatin features of CLL and normal B cells, identifying distinct epigenetic mechanisms operating in tumoral cells. DNA methylation analyses have identified that the CLL methylome contains imprints of the cell of origin, as well as of the proliferative history of the tumor cells, with both being strong independent prognostic predictors. Moreover, single-cell analysis revealed a higher degree of DNA methylation noise in CLL cells, which associates with transcriptional plasticity and disease aggressiveness. Integrative analysis of chromatin has uncovered chromatin signatures, as well as regulatory regions specifically active in each CLL subtype or in Richter transformed samples. Unique transcription factor (TF) binding motifs are overrepresented on those regions, suggesting that altered TF networks operate from disease initiation to progression as nongenetic factors mediating the oncogenic transcriptional profiles. Multiomics analysis has identified that response to treatment is modulated by an epigenetic imprint, and that treatments affect chromatin through the activity of particular set of TFs. Additionally, the epigenome is an axis of therapeutic vulnerability in CLL, as it can be targeted by inhibitors of histone modifying enzymes, that have shown promising preclinical results. Altogether, this review aims at summarizing the major findings derived from published literature to distill how altered epigenomic mechanisms contribute to CLL origin, evolution, clinical behavior, and response to treatment.
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Metilação de DNA , Epigênese Genética , Epigenoma , Leucemia Linfocítica Crônica de Células B , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/terapia , Leucemia Linfocítica Crônica de Células B/metabolismo , Prognóstico , Metilação de DNA/genética , Epigenômica , Cromatina/genética , Cromatina/metabolismoRESUMO
Introduction: Follicular Lymphoma (FL) results from the malignant transformation of germinal center (GC) B cells. FL B cells display recurrent and diverse genetic alterations, some of them favoring their direct interaction with their cell microenvironment, including follicular helper T cells (Tfh). Although FL-Tfh key role is well-documented, the impact of their regulatory counterpart, the follicular regulatory T cell (Tfr) compartment, is still sparse. Methods: The aim of this study was to characterize FL-Tfr phenotype by cytometry, gene expression profile, FL-Tfr origin by transcriptomic analysis, and functionality by in vitro assays. Results: CD4+CXCR5+CD25hiICOS+ FL-Tfr displayed a regulatory program that is close to classical regulatory T cell (Treg) program, at the transcriptomic and methylome levels. Accordingly, Tfr imprinting stigmata were found on FL-Tfh and FL-B cells, compared to their physiological counterparts. In addition, FL-Tfr co-culture with autologous FL-Tfh or cytotoxic FL-CD8+ T cells inhibited their proliferation in vitro. Finally, although FL-Tfr shared many characteristics with Treg, TCR sequencing analyses demonstrated that part of them derived from precursors shared with FL-Tfh. Discussion: Altogether, these findings uncover the role and origin of a Tfr subset in FL niche and may be useful for lymphomagenesis knowledge and therapeutic management.
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Linfoma Folicular , Linfócitos T Reguladores , Linfoma Folicular/imunologia , Linfoma Folicular/genética , Linfoma Folicular/patologia , Humanos , Linfócitos T Reguladores/imunologia , Perfilação da Expressão Gênica , Transcriptoma , Microambiente Tumoral/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Células T Auxiliares Foliculares/imunologia , Células T Auxiliares Foliculares/metabolismo , Masculino , Feminino , Técnicas de Cocultura , Centro Germinativo/imunologiaRESUMO
The histone H3K27 demethylase KDM6A is a tumor suppressor in multiple cancers, including multiple myeloma (MM). We created isogenic MM cells disrupted for KDM6A and tagged the endogenous protein to facilitate genome wide studies. KDM6A binds genes associated with immune recognition and cytokine signaling. Most importantly, KDM6A binds and activates NLRC5 and CIITA encoding regulators of Major Histocompatibility Complex (MHC) genes. Patient data indicate that NLRC5 and CIITA, are downregulated in MM with low KDM6A expression. Chromatin analysis shows that KDM6A binds poised and active enhancers and KDM6A loss led to decreased H3K27ac at enhancers, increased H3K27me3 levels in body of genes bound by KDM6A and decreased gene expression. Reestablishing histone acetylation with an HDAC3 inhibitor leads to upregulation of MHC expression, offering a strategy to restore immunogenicity of KDM6A deficient tumors. Loss of Kdm6a in murine RAS-transformed fibroblasts led to increased growth in vivo associated with decreased T cell infiltration. Statement of significance: We show that KDM6A participates in immune recognition of myeloma tumor cells by directly regulating the expression of the master regulators of MHC-I and II, NLRC5 and CIITA. The expression of these regulators can by rescued by the HDAC3 inhibitors in KDM6A-null cell lines.
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Despite the development of novel therapies for acute myeloid leukemia, outcomes remain poor for most patients, and therapeutic improvements are an urgent unmet need. Although treatment regimens promoting differentiation have succeeded in the treatment of acute promyelocytic leukemia, their role in other acute myeloid leukemia subtypes needs to be explored. Here we identify and characterize two lysine deacetylase inhibitors, CM-444 and CM-1758, exhibiting the capacity to promote myeloid differentiation in all acute myeloid leukemia subtypes at low non-cytotoxic doses, unlike other commercial histone deacetylase inhibitors. Analyzing the acetylome after CM-444 and CM-1758 treatment reveals modulation of non-histone proteins involved in the enhancer-promoter chromatin regulatory complex, including bromodomain proteins. This acetylation is essential for enhancing the expression of key transcription factors directly involved in the differentiation therapy induced by CM-444/CM-1758 in acute myeloid leukemia. In summary, these compounds may represent effective differentiation-based therapeutic agents across acute myeloid leukemia subtypes with a potential mechanism for the treatment of acute myeloid leukemia.
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Diferenciação Celular , Epigênese Genética , Inibidores de Histona Desacetilases , Leucemia Mieloide Aguda , Humanos , Diferenciação Celular/efeitos dos fármacos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Epigênese Genética/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/uso terapêutico , Linhagem Celular Tumoral , Acetilação/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , AnimaisRESUMO
Mammary carcinomas developing in SV40 transgenic WAP-T mice arise in two distinct histological phenotypes: as differentiated low-grade and undifferentiated high-grade tumors. We integrated different types of information such as histological grading, analysis of aCGH-based gene copy number and gene expression profiling to provide a comprehensive molecular description of mammary tumors in WAP-T mice. Applying a novel procedure for the correlation of gene copy number with gene expression on a global scale, we observed in tumor samples a global coherence between genotype and transcription. This coherence can be interpreted as a matched transcriptional regulation inherited from the cells of tumor origin and determined by the activity of cancer driver genes. Despite common recurrent genomic aberrations, e.g. gain of chr. 15 in most WAP-T tumors, loss of chr. 19 frequently occurs only in low-grade tumors. These tumors show features of "basal-like" epithelial differentiation, particularly expression of keratin 14. The high-grade tumors are clearly separated from the low-grade tumors by strong expression of the Met gene and by coexpression of epithelial (e.g. keratin 18) and mesenchymal (e.g. vimentin) markers. In high-grade tumors, the expression of the nonmutated Met protein is associated with Met-locus amplification and Met activity. The role of Met as a cancer driver gene is supported by the contribution of active Met signaling to motility and growth of mammary tumor-derived cells. Finally, we discuss the independent origin of low- and high-grade tumors from distinct cells of tumor origin, possibly luminal progenitors, distinguished by Met gene expression and Met signaling.
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Neoplasias Mamárias Experimentais/patologia , Proteínas do Leite/genética , Proteínas Proto-Oncogênicas c-met/fisiologia , Animais , Linhagem Celular Tumoral , Hibridização Genômica Comparativa , Feminino , Neoplasias Mamárias Experimentais/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Gradação de Tumores , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Proto-Oncogênicas c-met/genética , Proteína Supressora de Tumor p53/fisiologiaRESUMO
Rhabdoid tumors of early infancy are highly aggressive with consequent poor prognosis. Most cases show inactivation of the SMARCB1 (also known as INI1 and hSNF5) tumor suppressor, a core member of the ATP-dependent SWI/SNF chromatin-remodeling complex. Familial cases, described as rhabdoid tumor predisposition syndrome (RTPS), have been linked to heterozygous SMARCB1 germline mutations. We identified inactivation of another member of the SWI/SNF chromatin-remodeling complex, its ATPase subunit SMARCA4 (also known as BRG1), due to a SMARCA4/BRG1 germline mutation and loss of heterozygosity by uniparental disomy in the tumor cells of two sisters with rhabdoid tumors lacking SMARCB1 mutations. SMARCA4 is thus a second member of the SWI/SNF complex involved in cancer predisposition. Its general involvement in other tumor entities remains to be established.
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Códon sem Sentido/genética , DNA Helicases/genética , Inativação Gênica , Predisposição Genética para Doença , Mutação em Linhagem Germinativa/genética , Proteínas Nucleares/genética , Tumor Rabdoide/genética , Fatores de Transcrição/genética , Sequência de Bases , DNA Helicases/química , Análise Mutacional de DNA , Evolução Fatal , Feminino , Humanos , Imuno-Histoquímica , Lactente , Imageamento por Ressonância Magnética , Masculino , Dados de Sequência Molecular , Proteínas Nucleares/química , Linhagem , Tumor Rabdoide/patologia , Síndrome , Fatores de Transcrição/químicaRESUMO
Most DNA methylation studies in classic Philadelphia-negative myeloproliferative neoplasms have been performed on a gene-by-gene basis. Therefore, a more comprehensive methylation profiling is needed to study the implications of this epigenetic marker in myeloproliferative neoplasms. Here, we have analyzed 71 chronic (24 polycythemia vera, 23 essential thrombocythemia and 24 primary myelofibrosis) and 13 transformed myeloproliferative neoplasms using genome-wide DNA methylation arrays. The three types of chronic Philadelphia-negative myeloproliferative neoplasms showed a similar aberrant DNA methylation pattern when compared to control samples. Differentially methylated regions were enriched in a gene network centered on the NF-κB pathway, indicating that they may be involved in the pathogenesis of these diseases. In the case of transformed myeloproliferative neoplasms, we detected an increased number of differentially methylated regions with respect to chronic myeloproliferative neoplasms. Interestingly, these genes were enriched in a list of differentially methylated regions in primary acute myeloid leukemia and in a gene network centered around the IFN pathway. Our results suggest that alterations in the DNA methylation landscape play an important role in the pathogenesis and leukemic transformation of myeloproliferative neoplasms. The therapeutic modulation of epigenetically-deregulated pathways may allow us to design targeted therapies for these patients.