RESUMO
Chromosomal translocations are a hallmark of cancer. Unraveling the molecular mechanism of these rare genetic events requires a clear distinction between correlative and causative risk-determinants, where technical and analytical issues can be excluded. To meet this goal, we performed in-depth analyses of publicly available genome-wide datasets. In contrast to several recent reports, we demonstrate that chromosomal translocation risk is causally unrelated to promoter stalling (Spt5), transcriptional activity, or off-targeting activity of the activation-induced cytidine deaminase. Rather, an open chromatin configuration, which is not promoter-specific, explained the elevated translocation risk of promoter regions. Furthermore, the fact that gene size directly correlates with the translocation risk in mice and human cancers further demonstrated the general irrelevance of promoter-specific activities. Interestingly, a subset of translocations observed in cancer patients likely initiates from double-strand breaks induced by an access-independent process. Together, these unexpected and novel insights are fundamental in understanding the origin of chromosome translocations and, consequently, cancer.
Assuntos
Neoplasias/genética , Translocação Genética , Animais , Cromatina/genética , Genoma , Humanos , Camundongos , Neoplasias/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Regiões Promotoras Genéticas , Transcrição Gênica , Fatores de Elongação da Transcrição/genética , Fatores de Elongação da Transcrição/metabolismoRESUMO
Histone deacetylases (HDACs) are epigenetic erasers of lysine-acetyl marks. Inhibition of HDACs using small molecule inhibitors (HDACi) is a potential strategy in the treatment of various diseases and is approved for treating hematological malignancies. Harnessing the therapeutic potential of HDACi requires knowledge of HDAC-function in vivo. Here, we generated a thymocyte-specific gradient of HDAC-activity using compound conditional knockout mice for Hdac1 and Hdac2. Unexpectedly, gradual loss of HDAC-activity engendered a dosage-dependent accumulation of immature thymocytes and correlated with the incidence and latency of monoclonal lymphoblastic thymic lymphomas. Strikingly, complete ablation of Hdac1 and Hdac2 abrogated lymphomagenesis due to a block in early thymic development. Genomic, biochemical and functional analyses of pre-leukemic thymocytes and tumors revealed a critical role for Hdac1/Hdac2-governed HDAC-activity in regulating a p53-dependent barrier to constrain Myc-overexpressing thymocytes from progressing into lymphomas by regulating Myc-collaborating genes. One Myc-collaborating and p53-suppressing gene, Jdp2, was derepressed in an Hdac1/2-dependent manner and critical for the survival of Jdp2-overexpressing lymphoma cells. Although reduced HDAC-activity facilitates oncogenic transformation in normal cells, resulting tumor cells remain highly dependent on HDAC-activity, indicating that a critical level of Hdac1 and Hdac2 governed HDAC-activity is required for tumor maintenance.
Assuntos
Dosagem de Genes/fisiologia , Genes Supressores de Tumor/fisiologia , Histona Desacetilase 1/genética , Histona Desacetilase 2/genética , Proteínas Proto-Oncogênicas c-myc/fisiologia , Proteína Supressora de Tumor p53/fisiologia , Animais , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Células Cultivadas , Epistasia Genética/fisiologia , Dosagem de Genes/genética , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/fisiologia , Genes/fisiologia , Histona Desacetilase 1/metabolismo , Histona Desacetilase 1/fisiologia , Histona Desacetilase 2/metabolismo , Histona Desacetilase 2/fisiologia , Linfoma/genética , Linfoma/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Histone deacetylases (HDACs) counterbalance acetylation of lysine residues, a protein modification involved in numerous biological processes. Here, Hdac1 and Hdac2 conditional knock-out alleles were used to study the function of class I Hdac1 and Hdac2 in cell cycle progression and haematopoietic differentiation. Combined deletion of Hdac1 and Hdac2, or inactivation of their deacetylase activity in primary or oncogenic-transformed fibroblasts, results in a senescence-like G(1) cell cycle arrest, accompanied by up-regulation of the cyclin-dependent kinase inhibitor p21(Cip). Notably, concomitant genetic inactivation of p53 or p21(Cip) indicates that Hdac1 and Hdac2 regulate p53-p21(Cip)-independent pathways critical for maintaining cell cycle progression. In vivo, we show that Hdac1 and Hdac2 are not essential for liver homeostasis. In contrast, total levels of Hdac1 and Hdac2 in the haematopoietic system are critical for erythrocyte-megakaryocyte differentiation. Dual inactivation of Hdac1 and Hdac2 results in apoptosis of megakaryocytes and thrombocytopenia. Together, these data indicate that Hdac1 and Hdac2 have overlapping functions in cell cycle regulation and haematopoiesis. In addition, this work provides insights into mechanism-based toxicities observed in patients treated with HDAC inhibitors.
Assuntos
Ciclo Celular , Hematopoese , Histona Desacetilase 1/metabolismo , Histona Desacetilase 2/metabolismo , Anemia/enzimologia , Animais , Apoptose , Biocatálise , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p21/deficiência , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Histona Desacetilase 1/deficiência , Histona Desacetilase 2/deficiência , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Trombocitopenia/enzimologia , Trombocitopenia/patologia , Proteína Supressora de Tumor p53/metabolismoRESUMO
Class I histone deacetylases are critical regulators of gene transcription by erasing lysine acetylation. Targeting histone deacetylases using relative non-specific small molecule inhibitors is of major interest in the treatment of cancer, neurological disorders and acquired immune deficiency syndrome. Harnessing the therapeutic potential of histone deacetylase inhibitors requires full knowledge of individual histone deacetylases in vivo. As hematologic malignancies show increased sensitivity towards histone deacetylase inhibitors we targeted deletion of class I Hdac1 and Hdac2 to hematopoietic cell lineages. Here, we show that Hdac1 and Hdac2 together control hematopoietic stem cell homeostasis, in a cell-autonomous fashion. Simultaneous loss of Hdac1 and Hdac2 resulted in loss of hematopoietic stem cells and consequently bone marrow failure. Bone-marrow-specific deletion of Sin3a, a major Hdac1/2 co-repressor, phenocopied loss of Hdac1 and Hdac2 indicating that Sin3a-associated HDAC1/2-activity is essential for hematopoietic stem cell homeostasis. Although Hdac1 and Hdac2 show compensatory and overlapping functions in hematopoiesis, mice expressing mono-allelic Hdac1 or Hdac2 revealed that Hdac1 and Hdac2 contribute differently to the development of specific hematopoietic lineages.
Assuntos
Hematopoese/fisiologia , Células-Tronco Hematopoéticas/metabolismo , Histona Desacetilase 1/deficiência , Histona Desacetilase 2/deficiência , Homeostase/fisiologia , Proteínas Repressoras/deficiência , Animais , Células da Medula Óssea/fisiologia , Linhagem da Célula/fisiologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Complexo Correpressor Histona Desacetilase e Sin3RESUMO
B-lymphocyte development is dictated by the protein products of functionally rearranged Ig heavy (H) and light (L) chain genes. Ig rearrangement begins in pro-B cells at the IgH locus. If pro-B cells generate a productive allele, they assemble a pre-B cell receptor complex, which signals their differentiation into pre-B cells and their clonal expansion. Pre-B cell receptor signals are also thought to contribute to allelic exclusion by preventing further IgH rearrangements. Here we show in two independent mouse models that the accumulation of a stabilized µH mRNA that does not encode µH chain protein specifically impairs pro-B cell differentiation and reduces the frequency of rearranged IgH genes in a dose-dependent manner. Because noncoding IgH mRNA is usually rapidly degraded by the nonsense-mediated mRNA decay machinery, we propose that the difference in mRNA stability allows pro-B cells to distinguish between productive and nonproductive Ig gene rearrangements and that µH mRNA may thus contribute to efficient H chain allelic exclusion.
Assuntos
Linfócitos B/imunologia , Cadeias Pesadas de Imunoglobulinas/genética , Biossíntese Peptídica , Alelos , Animais , Camundongos , RNA Mensageiro/genética , RNA não Traduzido/genética , VDJ Recombinases/metabolismoRESUMO
Epidermal growth factor receptor (EGFR) is highly expressed in triple-negative breast cancer (TNBC), and elevated levels correlate with poor prognosis. In analogy with the paradigm of oncogene addiction, blocking EGFR in TNBC was expected to have clinical efficacy but this has not been the case. Reasons for these results have remained elusive. Recently, Meyer and colleagues showed interplay between EGFR and the epithelial-to-mesenchymal transition-associated AXL receptor in TNBC cells, which might provide some clues.
Assuntos
Neoplasias da Mama/metabolismo , Receptores ErbB/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Transdução de Sinais , Feminino , HumanosRESUMO
INTRODUCTION: Targeting receptor tyrosine kinases (RTKs) with kinase inhibitors is a clinically validated anti-cancer approach. However, blocking one signaling pathway is often not sufficient to cause tumor regression and the effectiveness of individual inhibitors is often short-lived. As alterations in fibroblast growth factor receptor (FGFR) activity have been implicated in breast cancer, we examined in breast cancer models with autocrine FGFR activity the impact of targeting FGFRs in vivo with a selective kinase inhibitor in combination with an inhibitor of PI3K/mTOR or with a pan-ErbB inhibitor. METHODS: Using 4T1 or 67NR models of basal-like breast cancer, tumor growth was measured in mice treated with an FGFR inhibitor (dovitinib/TKI258), a PI3K/mTOR inhibitor (NVP-BEZ235) or a pan-ErbB inhibitor (AEE788) individually or in combination. To uncover mechanisms underlying inhibitor action, signaling pathway activity was examined in tumor lysates and transcriptome analysis carried out to identify pathways upregulated by FGFR inhibition. Anti-phosphotyrosine receptor antibody arrays (P-Tyr RTK) were also used to screen 4T1 tumors. RESULTS: The combination of dovitinib + NVP-BEZ235 causes tumor stasis and strong down-regulation of the FRS2/Erk and PI3K/Akt/mTOR signaling pathways. P-Tyr RTK arrays identified high levels of P-EGFR and P-ErbB2 in 4T1 tumors. Testing AEE788 in the tumor models revealed that the combination of dovitinib + AEE788 resulted in blockade of the PI3K/Akt/mTOR pathway, prolonged tumor stasis and in the 4T1 model, a significant decrease in lung metastasis. The results show that in vivo these breast cancer models become dependent upon co-activation of FGFR and ErbB receptors for PI3K pathway activity. CONCLUSIONS: The work presented here shows that in the breast cancer models examined, the combination of dovitinib + NVP-BEZ235 or dovitinib + AEE788 results in strong inhibition of tumor growth and a block in metastatic spread. Only these combinations strongly down-regulate the FGFR/FRS2/Erk and PI3K/Akt/mTOR signaling pathways. The resultant decrease in mitosis and increase in apoptosis was consistently stronger in the dovitinib + AEE788 treatment-group, suggesting that targeting ErbB receptors has broader downstream effects compared to targeting only PI3K/mTOR. Considering that sub-classes of human breast tumors co-express ErbB receptors and FGFRs, these results have implications for targeted therapy.
Assuntos
Neoplasias da Mama/genética , Proliferação de Células/efeitos dos fármacos , Receptores ErbB/genética , Receptores de Fatores de Crescimento de Fibroblastos/genética , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Receptores ErbB/antagonistas & inibidores , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Imidazóis/administração & dosagem , Camundongos , Terapia de Alvo Molecular , Fosfatidilinositol 3-Quinases/administração & dosagem , Fosfatidilinositol 3-Quinases/genética , Inibidores de Proteínas Quinases/administração & dosagem , Proteínas Proto-Oncogênicas c-akt/genética , Purinas/administração & dosagem , Quinolinas/administração & dosagem , Receptores de Fatores de Crescimento de Fibroblastos/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The two histone deacetylases (Hdacs), Hdac1 and Hdac2, are erasers of acetylation marks on histone tails, and are important regulators of gene expression that were shown to play important roles in hematological malignancies. However, several recent studies reported opposing tumor-suppressive or tumor-promoting roles for Hdac1 and Hdac2. Here, we investigated the functional role of Hdac1 and Hdac2 using the Eµ-myc mouse model of B cell lymphoma. We demonstrate that Hdac1 and Hdac2 have a pro-oncogenic role in both Eµ-myc tumorigenesis and tumor maintenance. Hdac1 and Hdac2 promote tumorigenesis in a gene dose-dependent manner, with a predominant function of Hdac1. Our data show that Hdac1 and Hdac2 impact on Eµ-myc B cell proliferation and apoptosis and suggest that a critical level of Hdac activity may be required for Eµ-myc tumorigenesis and proper B cell development. This provides the rationale for utilization of selective Hdac1 and Hdac2 inhibitors in the treatment of hematological malignancies.
Assuntos
Genes myc , Histona Desacetilase 1/metabolismo , Linfoma de Células B/genética , Oncogenes , Animais , Histona Desacetilase 2/metabolismo , Humanos , CamundongosRESUMO
The extracellular serine protease inhibitor serpinE2 is overexpressed in breast cancer and has been shown to foster metastatic spread. Here, we investigated the hypothesis that serpinE2 creates tumor-promoting conditions in the tumor microenvironment (TME) by affecting extracellular matrix remodeling. Using two different breast cancer models, we show that blocking serpinE2, either by knock-down (KD) in tumor cells or in response to a serpinE2 binding antibody, decreases metastatic dissemination from primary tumors to the lungs. We demonstrate that in response to serpinE2 KD or antibody treatment there are dramatic changes in the TME. Multiphoton intravital imaging revealed deposition of a dense extracellular collagen I matrix encapsulating serpinE2 KD or antibody-treated tumors. This is accompanied by a reduction in the population of tumor-promoting macrophages, as well as a decrease in chemokine ligand 2, which is known to affect macrophage abundance and polarization. In addition, TIMP-1 secretion is increased, which may directly inhibit matrix metalloproteases critical for collagen degradation in the tumor. In summary, our findings suggest that serpinE2 is required in the extracellular milieu of tumors where it acts in multiple ways to regulate tumor matrix deposition, thereby controlling tumor cell dissemination.
Assuntos
Neoplasias da Mama/metabolismo , Movimento Celular , Matriz Extracelular/metabolismo , Neoplasias Pulmonares/metabolismo , Macrófagos/metabolismo , Serpina E2/metabolismo , Animais , Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Quimiocina CCL2/metabolismo , Colágeno Tipo I/metabolismo , Matriz Extracelular/patologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Camundongos Endogâmicos BALB C , Camundongos SCID , Invasividade Neoplásica , Fenótipo , Interferência de RNA , Serpina E2/antagonistas & inibidores , Serpina E2/genética , Transdução de Sinais , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Transfecção , Microambiente Tumoral , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The majority of patients with breast cancer present with an estrogen receptor-positive (ER(+)) tumor, and the endocrine agent tamoxifen is a mainstay for their treatment. Unfortunately, however, resistance remains a major problem because most patients who respond eventually have a recurrence. Thus, an enduring challenge in the breast cancer field is to identify mechanisms underlying tamoxifen resistance. Jin and colleagues describe a novel ER/HOXB7 signaling loop in tamoxifen-resistant breast cancer models. Importantly, they reveal that targeting this signaling loop has great promise as an approach to treat patients with tamoxifen-resistant breast cancer.
Assuntos
Antineoplásicos Hormonais/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Receptor alfa de Estrogênio/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Receptor ErbB-2/metabolismo , Animais , Feminino , HumanosRESUMO
Recent observations connected DNA cytosine deaminase APOBEC3B to the genetic evolution of breast cancer. We addressed whether APOBEC3B is associated with breast cancer clinical outcomes. APOBEC3B messenger RNA (mRNA) levels were related in 1,491 primary breast cancers to disease-free (DFS), metastasis-free (MFS), and overall survival (OS). For independent validation, APOBEC3B mRNA expression was associated with patient outcome data in five additional cohorts (over 3,500 breast cancer cases). In univariate Cox regression analysis, increasing APOBEC3B expression as a continuous variable was associated with worse DFS, MFS, and OS (hazard ratio [HR] = 1.20, 1.21, and 1.24, respectively; all P < .001). Also, in untreated ER-positive (ER+), but not in ER-, lymph-node-negative patients, high APOBEC3B levels were associated with a poor DFS (continuous variable: HR = 1.29, P = .001; dichotomized at the median level, HR = 1.66, P = .0002). This implies that APOBEC3B is a marker of pure prognosis in ER + disease. These findings were confirmed in the analyses of five independent patient sets. In these analyses, APOBEC3B expression dichotomized at the median level was associated with adverse outcomes (METABRIC discovery and validation, 788 and 706 ER + cases, disease-specific survival (DSS), HR = 1.77 and HR = 1.77, respectively, both P < .001; Affymetrix dataset, 754 ER + cases, DFS, HR = 1.57, P = 2.46E-04; NKI295, 181 ER + cases, DFS, HR = 1.72, P = .054; and BIG 1-98, 1,219 ER + cases, breast-cancer-free interval (BCFI), HR = 1.42, P = 0.0079). APOBEC3B is a marker of pure prognosis and poor outcomes for ER + breast cancer, which strongly suggests that genetic aberrations induced by APOBEC3B contribute to breast cancer progression.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/diagnóstico , Citidina Desaminase/metabolismo , Adulto , Idoso , Neoplasias da Mama/genética , Neoplasias da Mama/mortalidade , Estudos de Coortes , Citidina Desaminase/genética , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Humanos , Análise em Microsséries , Pessoa de Meia-Idade , Antígenos de Histocompatibilidade Menor , Estadiamento de Neoplasias , Prognóstico , RNA Mensageiro/análise , Receptores de Estrogênio/metabolismo , Análise de Sobrevida , Regulação para CimaRESUMO
The Aicda locus encodes the activation induced cytidine deaminase (AID) and is highly expressed in germinal center (GC) B cells to initiate somatic hypermutation (SHM) and class switch recombination (CSR) of immunoglobulin (Ig) genes. Besides these Ig specific activities in B cells, AID has been implicated in active DNA demethylation in non-B cell systems. We here determined a potential role of AID as an epigenetic eraser and transcriptional regulator in B cells. RNA-Seq on different B cell subsets revealed that Aicda(-/-) B cells are developmentally affected. However as shown by RNA-Seq, MethylCap-Seq, and SNP analysis these transcriptome alterations may not relate to AID, but alternatively to a CBA mouse strain derived region around the targeted Aicda locus. These unexpected confounding parameters provide alternative, AID-independent interpretations on genotype-phenotype correlations previously reported in numerous studies on AID using the Aicda(-/-) mouse strain.
Assuntos
Linfócitos B/enzimologia , Linfócitos B/metabolismo , Citidina Desaminase/deficiência , Animais , Citidina Desaminase/genética , Metilação de DNA/genética , Genótipo , Camundongos , Camundongos MutantesRESUMO
Proliferating cell nuclear antigen (PCNA) encircles DNA as a ring-shaped homotrimer and, by tethering DNA polymerases to their template, PCNA serves as a critical replication factor. In contrast to high-fidelity DNA polymerases, the activation of low-fidelity translesion synthesis (TLS) DNA polymerases seems to require damage-inducible monoubiquitylation (Ub) of PCNA at lysine residue 164 (PCNA-Ub). TLS polymerases can tolerate DNA damage, i.e. they can replicate across DNA lesions. The lack of proofreading activity, however, renders TLS highly mutagenic. The advantage is that B cells use mutagenic TLS to introduce somatic mutations in immunoglobulin (Ig) genes to generate high-affinity antibodies. Given the critical role of PCNA-Ub in activating TLS and the role of TLS in establishing somatic mutations in immunoglobulin genes, we analysed the mutation spectrum of somatically mutated immunoglobulin genes in B cells from PCNAK164R knock-in mice. A 10-fold reduction in A/T mutations is associated with a compensatory increase in G/C mutations-a phenotype similar to Poleta and mismatch repair-deficient B cells. Mismatch recognition, PCNA-Ub and Poleta probably act within one pathway to establish the majority of mutations at template A/T. Equally relevant, the G/C mutator(s) seems largely independent of PCNAK(164) modification.