RESUMO
Drugs that block the activity of the methyltransferase EZH2 are in clinical development for the treatment of non-Hodgkin lymphomas harboring EZH2 gain-of-function mutations that enhance its polycomb repressive function. We have previously reported that EZH2 can act as a transcriptional activator in castration-resistant prostate cancer (CRPC). Now we show that EZH2 inhibitors can also block the transactivation activity of EZH2 and inhibit the growth of CRPC cells. Gene expression and epigenomics profiling of cells treated with EZH2 inhibitors demonstrated that in addition to derepressing gene expression, these compounds also robustly down-regulate a set of DNA damage repair (DDR) genes, especially those involved in the base excision repair (BER) pathway. Methylation of the pioneer factor FOXA1 by EZH2 contributes to the activation of these genes, and interaction with the transcriptional coactivator P300 via the transactivation domain on EZH2 directly turns on the transcription. In addition, CRISPR-Cas9-mediated knockout screens in the presence of EZH2 inhibitors identified these BER genes as the determinants that underlie the growth-inhibitory effect of EZH2 inhibitors. Interrogation of public data from diverse types of solid tumors expressing wild-type EZH2 demonstrated that expression of DDR genes is significantly correlated with EZH2 dependency and cellular sensitivity to EZH2 inhibitors. Consistent with these findings, treatment of CRPC cells with EZH2 inhibitors dramatically enhances their sensitivity to genotoxic stress. These studies reveal a previously unappreciated mechanism of action of EZH2 inhibitors and provide a mechanistic basis for potential combination cancer therapies.
Assuntos
Dano ao DNA/genética , Dano ao DNA/fisiologia , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Ativação Transcricional , Sistemas CRISPR-Cas , Linhagem Celular Tumoral , Reparo do DNA/genética , Reparo do DNA/fisiologia , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Técnicas de Inativação de Genes , Fator 3-alfa Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Humanos , Masculino , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/metabolismoRESUMO
BACKGROUND: Cyclin-dependent kinase 9 (CDK9) stimulates oncogenic transcriptional pathways in cancer and CDK9 inhibitors have emerged as promising therapeutic candidates. METHODS: The activity of an orally bioavailable CDK9 inhibitor, CDKI-73, was evaluated in prostate cancer cell lines, a xenograft mouse model, and patient-derived tumor explants and organoids. Expression of CDK9 was evaluated in clinical specimens by mining public datasets and immunohistochemistry. Effects of CDKI-73 on prostate cancer cells were determined by cell-based assays, molecular profiling and transcriptomic/epigenomic approaches. RESULTS: CDKI-73 inhibited proliferation and enhanced cell death in diverse in vitro and in vivo models of androgen receptor (AR)-driven and AR-independent models. Mechanistically, CDKI-73-mediated inhibition of RNA polymerase II serine 2 phosphorylation resulted in reduced expression of BCL-2 anti-apoptotic factors and transcriptional defects. Transcriptomic and epigenomic approaches revealed that CDKI-73 suppressed signaling pathways regulated by AR, MYC, and BRD4, key drivers of dysregulated transcription in prostate cancer, and reprogrammed cancer-associated super-enhancers. These latter findings prompted the evaluation of CDKI-73 with the BRD4 inhibitor AZD5153, a combination that was synergistic in patient-derived organoids and in vivo. CONCLUSION: Our work demonstrates that CDK9 inhibition disrupts multiple oncogenic pathways and positions CDKI-73 as a promising therapeutic agent for prostate cancer, particularly aggressive, therapy-resistant subtypes.
RESUMO
The evolution of prostate cancer from an androgen-dependent state to one that is androgen-independent marks its lethal progression. The androgen receptor (AR) is essential in both, though its function in androgen-independent cancers is poorly understood. We have defined the direct AR-dependent target genes in both androgen-dependent and -independent cancer cells by generating AR-dependent gene expression profiles and AR cistromes. In contrast to what is found in androgen-dependent cells, AR selectively upregulates M-phase cell-cycle genes in androgen-independent cells, including UBE2C, a gene that inactivates the M-phase checkpoint. We find that epigenetic marks at the UBE2C enhancer, notably histone H3K4 methylation and FoxA1 transcription factor binding, are present in androgen-independent cells and direct AR-enhancer binding and UBE2C activation. Thus, the role of AR in androgen-independent cancer cells is not to direct the androgen-dependent gene expression program without androgen, but rather to execute a distinct program resulting in androgen-independent growth.
Assuntos
Regulação Neoplásica da Expressão Gênica , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Androgênios/metabolismo , Divisão Celular , Linhagem Celular Tumoral , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Histonas/metabolismo , Humanos , Masculino , Neoplasias da Próstata/genética , Ativação Transcricional , Enzimas de Conjugação de Ubiquitina/metabolismoRESUMO
Adverse neighborhood social and natural (green space) environments may contribute to the etiology of prostate cancer (CaP), but mechanisms are unclear. We examined associations between neighborhood environment and prostate intratumoral inflammation in 967 men diagnosed with CaP with available tissue samples from 1986-2009 in the Health Professionals Follow-up Study. Exposures were linked to work or residential addresses in 1988. We estimated indices of neighborhood socioeconomic status (nSES) and segregation (Index of Concentration at the Extremes (ICE)) using US Census tract-level data. Surrounding greenness was estimated using seasonal averaged Normalized Difference Vegetation Index (NDVI) data. Surgical tissue underwent pathological review for acute and chronic inflammation, corpora amylacea, and focal atrophic lesions. Adjusted odds ratios (aORs) for inflammation (ordinal) and focal atrophy (binary) were estimated using logistic regression. No associations were observed for acute or chronic inflammation. Each interquartile-range increase in NDVI within 1,230 m of the participant's work or home address (aOR = 0.74, 95% confidence interval (CI): 0.59, 0.93), in ICE-income (aOR = 0.79, 95% CI: 0.61, 1.04), and in ICE-race/income (aOR = 0.79, 95% CI: 0.63, 0.99) was associated with lower odds of postatrophic hyperplasia. Interquartile-range increases in nSES (aOR = 0.76, 95% CI: 0.57, 1.02) and ICE-race/income (aOR = 0.73, 95% CI: 0.54, 0.99) were associated with lower odds of tumor corpora amylacea. Histopathological inflammatory features of prostate tumors may be influenced by neighborhood.
Assuntos
Meio Ambiente , Neoplasias da Próstata , Humanos , Masculino , Seguimentos , Inflamação , Neoplasias da Próstata/epidemiologia , Características de Residência , Classe Social , Fatores SocioeconômicosRESUMO
The androgen receptor (AR) plays a pivotal role in driving prostate cancer (PCa) development. However, when stimulated by high levels of androgens, AR can also function as a tumor suppressor in PCa cells. While the high-dose testosterone (high-T) treatment is currently being tested in clinical trials of castration-resistant prostate cancer (CRPC), there is still a pressing need to fully understand the underlying mechanism and thus develop treatment strategies to exploit this tumor-suppressive activity of AR. In this study, we demonstrate that retinoblastoma (Rb) family proteins play a central role in maintaining the global chromatin binding and transcriptional repression program of AR and that Rb inactivation desensitizes CRPC to the high-dose testosterone treatment in vitro and in vivo. Using a series of patient-derived xenograft (PDX) CRPC models, we further show that the efficacy of high-T treatment can be fully exploited by a CDK4/6 inhibitor, which strengthens the chromatin binding of the Rb-E2F repressor complex by blocking the hyperphosphorylation of Rb proteins. Overall, our study provides strong mechanistic and preclinical evidence on further developing clinical trials to combine high-T with CDK4/6 inhibitors in treating CRPC.
Assuntos
Neoplasias de Próstata Resistentes à Castração , Receptores Androgênicos , Linhagem Celular Tumoral , Cromatina , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 4 Dependente de Ciclina/uso terapêutico , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/uso terapêutico , Genes Supressores de Tumor , Humanos , Masculino , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Proteína do Retinoblastoma/genética , Testosterona/uso terapêuticoRESUMO
BACKGROUND: Men of African ancestry (AA) with prostate cancer suffer from worse outcomes. However, a recent analysis of patients treated with the dendritic cell vaccine sipuleucel-T for prostate cancer suggested that AA patients could have improved outcomes relative to whites. METHODS: We conducted a focused literature review of Medline-indexed articles and clinical trials listed on clinicaltrials.gov. RESULTS: We identify several studies pointing to enrichment of inflammatory cellular infiltrates and cytokine signaling among AA patients with prostate cancer. We outline potential genomic and transcriptomic alterations that may contribute to immunogenicity. Last, we investigate differences in host immunity and vaccine responsiveness that may be enhanced in AA patients. CONCLUSIONS: AA patients with prostate cancer may be enriched for an immunogenic phenotype. Dedicated studies are needed to better understand the immune mechanisms that contribute to existing cancer disparities and test immune-based therapies in this population.
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Negro ou Afro-Americano , Neoplasias da Próstata , Negro ou Afro-Americano/genética , População Negra/genética , Humanos , Masculino , Neoplasias da Próstata/terapia , Transcriptoma , População BrancaRESUMO
BACKGROUND: B7 homolog 3 (B7-H3) is an immunomodulatory molecule that is highly expressed in prostate cancer (PCa) and belongs to the B7 superfamily, which includes PD-L1. Immunotherapies (antibodies, antibody-drug conjugates, and chimeric antigen receptor T cells) targeting B7-H3 are currently in clinical trials; therefore, elucidating the molecular and immune microenvironment correlates of B7-H3 expression may help to guide trial design and interpretation. The authors tested the interconnected hypotheses that B7-H3 expression is associated with genetic racial ancestry, immune cell composition, and androgen receptor signaling in PCa. METHODS: An automated, clinical-grade immunohistochemistry assay was developed by to digitally quantify B7-H3 protein expression across 2 racially diverse cohorts of primary PCa (1 with previously reported transcriptomic data) and pretreatment and posttreatment PCa tissues from a trial of intensive neoadjuvant hormonal therapy. RESULTS: B7-H3 protein expression was significantly lower in self-identified Black patients and was inversely correlated with the percentage African ancestry. This association with race was independent of the significant association of B7-H3 protein expression with ERG/ETS and PTEN status. B7-H3 messenger RNA expression, but not B7-H3 protein expression, was significantly correlated with regulatory (FOXP3-positive) T-cell density. Finally, androgen receptor activity scores were significantly correlated with B7-H3 messenger RNA expression, and neoadjuvant intensive hormonal therapy was associated with a significant decrease in B7-H3 protein expression. CONCLUSIONS: The current data underscore the importance of studying racially and molecularly diverse PCa cohorts in the immunotherapy era. This study is among the first to use genetic ancestry markers to add to the emerging evidence that PCa in men of African ancestry may have a distinct biology associated with B7-H3 expression. LAY SUMMARY: B7-H3 is an immunomodulatory molecule that is highly expressed in prostate cancer and is under investigation in clinical trials. The authors determined that B7-H3 protein expression is inversely correlated with an individual's proportion of African ancestry. The results demonstrate that B7-H3 messenger RNA expression is correlated with the density of tumor T-regulatory cells. Finally, in the first paired analysis of B7-H3 protein expression before and after neoadjuvant intensive hormone therapy, the authors determined that hormone therapy is associated with a decrease in B7-H3 protein levels, suggesting that androgen signaling may positively regulate B7-H3 expression. These results may help to guide the design of future clinical trials and to develop biomarkers of response in such trials.
Assuntos
Neoplasias da Próstata , Receptores Androgênicos , Androgênios , Antígenos B7/genética , Antígenos B7/metabolismo , Antígeno B7-H1/genética , Contagem de Células , Humanos , Masculino , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , RNA Mensageiro , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Microambiente TumoralRESUMO
Invariant natural killer T cells (iNKT cells) express a semi-invariant T cell receptor that recognizes certain glycolipids (including α-galactosylceramide, αGC) bound to CD1d, and can induce potent antitumor responses. Here, we assessed whether αGC could enhance the efficacy of a GM-CSF-producing tumor cell vaccine in the transgenic SV40 T antigen-driven TRAMP prostate cancer model. In healthy mice, we initially found that optimal T cell responses were obtained with αGC-pulsed TRAMP-C2 cells secreting GM-CSF and milk fat globule epidermal growth factor protein-8 (MFG-E8) with an RGD to RGE mutation (GM-CSF/RGE TRAMP-C2), combined with systemic low dose IL-12. In a therapeutic model, transgenic TRAMP mice were then castrated at ~ 20 weeks, followed by treatment with the combination vaccine. Untreated mice succumbed to tumor by ~ 40 weeks, but survival was markedly prolonged by vaccine treatment, with most mice surviving past 80 weeks. Prostates in the treated mice were heavily infiltrated with T cells and iNKT cells, which both secreted IFNγ in response to tumor cells. The vaccine was not effective if the αGC, IL-12, or GM-CSF secretion was eliminated. Finally, immunized mice were fully resistant to challenge with TRAMP-C2 cells. Together these findings support further development of therapeutic vaccines that exploit iNKT cell activation.
Assuntos
Vacinas Anticâncer , Células T Matadoras Naturais , Neoplasias da Próstata , Masculino , Camundongos , Animais , Humanos , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Ativação Linfocitária , Galactosilceramidas , Interleucina-12/farmacologia , Neoplasias da Próstata/terapia , Neoplasias da Próstata/metabolismo , Vacinas Combinadas/farmacologia , Antígenos Virais de Tumores , Família de Proteínas EGF/metabolismo , Família de Proteínas EGF/farmacologia , Oligopeptídeos/farmacologia , Camundongos Endogâmicos C57BLRESUMO
Abiraterone blocks androgen synthesis and prolongs survival in patients with castration-resistant prostate cancer, which is otherwise driven by intratumoral androgen synthesis. Abiraterone is metabolized in patients to Δ(4)-abiraterone (D4A), which has even greater anti-tumour activity and is structurally similar to endogenous steroidal 5α-reductase substrates, such as testosterone. Here, we show that D4A is converted to at least three 5α-reduced and three 5ß-reduced metabolites in human serum. The initial 5α-reduced metabolite, 3-keto-5α-abiraterone, is present at higher concentrations than D4A in patients with prostate cancer taking abiraterone, and is an androgen receptor agonist, which promotes prostate cancer progression. In a clinical trial of abiraterone alone, followed by abiraterone plus dutasteride (a 5α-reductase inhibitor), 3-keto-5α-abiraterone and downstream metabolites were depleted by the addition of dutasteride, while D4A concentrations rose, showing that dutasteride effectively blocks production of a tumour-promoting metabolite and permits D4A accumulation. Furthermore, dutasteride did not deplete the three 5ß-reduced metabolites, which were also clinically detectable, demonstrating the specific biochemical effects of pharmacological 5α-reductase inhibition on abiraterone metabolism. Our findings suggest a previously unappreciated and biochemically specific method of clinically fine-tuning abiraterone metabolism to optimize therapy.
Assuntos
Inibidores de 5-alfa Redutase/farmacologia , Androgênios/biossíntese , Androstenos/metabolismo , Dutasterida/farmacologia , Dutasterida/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo , 3-Oxo-5-alfa-Esteroide 4-Desidrogenase/metabolismo , Inibidores de 5-alfa Redutase/uso terapêutico , Acetato de Abiraterona/administração & dosagem , Acetato de Abiraterona/sangue , Acetato de Abiraterona/metabolismo , Acetato de Abiraterona/uso terapêutico , Administração Oral , Antagonistas de Androgênios/farmacologia , Antagonistas de Androgênios/uso terapêutico , Androstenos/administração & dosagem , Androstenos/sangue , Androstenos/farmacologia , Animais , Linhagem Celular Tumoral , Progressão da Doença , Humanos , Masculino , Camundongos , Oxirredução/efeitos dos fármacos , Neoplasias da Próstata/sangue , Neoplasias de Próstata Resistentes à Castração/sangue , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores Androgênicos/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Invariant natural killer T (iNKT) cells are evolutionarily conserved innate T cells that influence inflammatory responses. We have shown that iNKT cells, previously thought to be rare in humans, were highly enriched in human and murine adipose tissue, and that as adipose tissue expanded in obesity, iNKT cells were depleted, correlating with proinflammatory macrophage infiltration. iNKT cell numbers were restored in mice and humans after weight loss. Mice lacking iNKT cells had enhanced weight gain, larger adipocytes, fatty livers, and insulin resistance on a high-fat diet. Adoptive transfer of iNKT cells into obese mice or in vivo activation of iNKT cells via their lipid ligand, alpha-galactocylceramide, decreased body fat, triglyceride levels, leptin, and fatty liver and improved insulin sensitivity through anti-inflammatory cytokine production by adipose-derived iNKT cells. This finding highlights the potential of iNKT cell-targeted therapies, previously proven to be safe in humans, in the management of obesity and its consequences.
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Tecido Adiposo/imunologia , Citocinas/imunologia , Doenças Metabólicas/imunologia , Células T Matadoras Naturais/imunologia , Obesidade/imunologia , Tecido Adiposo/metabolismo , Transferência Adotiva , Adulto , Animais , Antígenos CD1d/genética , Antígenos CD1d/imunologia , Antígenos CD1d/metabolismo , Antígeno CD11c/imunologia , Antígeno CD11c/metabolismo , Citocinas/metabolismo , Dieta Hiperlipídica/efeitos adversos , Feminino , Citometria de Fluxo , Humanos , Fígado/imunologia , Fígado/metabolismo , Contagem de Linfócitos , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Doenças Metabólicas/etiologia , Doenças Metabólicas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Obesos , Pessoa de Meia-Idade , Células T Matadoras Naturais/metabolismo , Células T Matadoras Naturais/transplante , Obesidade/etiologia , Obesidade/metabolismo , Baço/imunologia , Baço/metabolismo , Adulto JovemRESUMO
Major problems in cancer chemotherapy are toxicity, resistance, and cancer heterogeneity. A new theranostic paradigm has been proposed by the authors. Many million small molecules (SM) are bound to the proteins extracted from a patient's cancer. SM that also bind proteins extracted from normal human tissues are subtracted from the cancer protein bound SM leaving a large array of SM targeting many sites on each of the cancer biomarkers. Targeting many more than the conventional 1 - 4 cancer biomarkers will reduce development of tumor resistance. After several cycles of selection and counter selection, DNA codes appended to the SM will be PCR amplified to provide templates for restricted libraries of the SM to improve selectivity and sensitivity. The large array of selected and counter selected SM assures that many of the compounds in the array will penetrate the cell membrane and bind to intracellular targets, low tumor resistance, low background for imaging, low therapeutic toxicity, and targeting of the diverse biomarkers present in the heterogeneous mixture of cells in primary and metastatic cancer. Theranostic use of radiolabeled SM binding many sites on many, not necessarily critical, biomarkers provides high cancer cell killing. Experiments to provide proof of principle of this novel concept suggested by the authors.
Assuntos
DNA , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias/terapia , Bibliotecas de Moléculas Pequenas/farmacologia , Nanomedicina Teranóstica/métodos , Biomarcadores Tumorais/metabolismo , Corantes Fluorescentes/química , Corantes Fluorescentes/metabolismo , Humanos , Terapia de Alvo Molecular/métodos , Neoplasias/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/metabolismoRESUMO
PURPOSE OF REVIEW: Prostate cancer (PCa) is diagnosed in one out of every nine men and is the second leading cause of cancer death among men. Although therapies targeting the androgen receptor (AR) are highly effective, development of resistance is universal and remains a major therapeutic challenge. Nonetheless, signaling via AR is frequently maintained despite standard androgen-signaling inhibition. We review the current understanding of mechanisms of resistance as well as therapeutic approaches to improving treatment of PCa via targeting of the AR. RECENT FINDINGS: Resistance to AR-targeting therapies may be mediated by several mechanisms, including amplification, mutation, and alternative splicing of AR; intratumoral androgen synthesis; activation of alternative signaling pathways; and in a minority of cases, emergence of AR-independent phenotypes. Recent trials demonstrate that intensification of androgen blockade in metastatic castration-sensitive PCa can significantly improve survival. Similar strategies are being explored in earlier disease states. In addition, several other cellular signaling pathways have been identified as mechanisms of resistance, offering opportunities for cotargeted therapy. Finally, immune-based approaches are in development to complement AR-targeted therapies. SUMMARY: Targeting the AR remains a critical focus in the treatment of PCa.
Assuntos
Antagonistas de Receptores de Andrógenos/uso terapêutico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/metabolismo , Antagonistas de Receptores de Andrógenos/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Humanos , Masculino , Terapia de Alvo Molecular , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologiaRESUMO
Myeloproliferative neoplasms (MPNs) driver mutations are usually found in JAK2, MPL, and CALR genes; however, 10%-15% of cases are triple negative (TN). A previous study showed lower rate of JAK2 V617F in primary myelofibrosis patients exposed to low doses of ionizing radiation (IR) from Chernobyl accident. To examine distinct driver mutations, we enrolled 281 Ukrainian IR-exposed and unexposed MPN patients. Genomic DNA was obtained from peripheral blood leukocytes. JAK2 V617F, MPL W515, types 1- and 2-like CALR mutations were identified by Sanger Sequencing and real time polymerase chain reaction. Chromosomal alterations were assessed by oligo-SNP microarray platform. Additional genetic variants were identified by whole exome and targeted sequencing. Statistical significance was evaluated by Fisher's exact test and Wilcoxon's rank sum test (R, version 3.4.2). IR-exposed MPN patients exhibited a different genetic profile vs unexposed: lower rate of JAK2 V617F (58.4% vs 75.4%, P = .0077), higher rate of type 1-like CALR mutation (12.2% vs 3.1%, P = .0056), higher rate of TN cases (27.8% vs 16.2%, P = .0366), higher rate of potentially pathogenic sequence variants (mean numbers: 4.8 vs 3.1, P = .0242). Furthermore, we identified several potential drivers specific to IR-exposed TN MPN patients: ATM p.S1691R with copy-neutral loss of heterozygosity at 11q; EZH2 p.D659G at 7q and SUZ12 p.V71 M at 17q with copy number loss. Thus, IR-exposed MPN patients represent a group with distinct genomic characteristics worthy of further study.
Assuntos
Acidente Nuclear de Chernobyl , Transtornos Mieloproliferativos/etiologia , Neoplasias Induzidas por Radiação/etiologia , Poluentes Radioativos/efeitos adversos , Adulto , Idoso , Calreticulina/genética , Aberrações Cromossômicas , DNA/genética , Feminino , Dosagem de Genes , Humanos , Janus Quinase 2/genética , Perda de Heterozigosidade , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Transtornos Mieloproliferativos/epidemiologia , Transtornos Mieloproliferativos/genética , Neoplasias Induzidas por Radiação/epidemiologia , Neoplasias Induzidas por Radiação/genética , Receptores de Trombopoetina/genética , Ucrânia/epidemiologia , Sequenciamento do Exoma , Adulto JovemRESUMO
P-TEFb (CDK9/cyclin T) plays a central role in androgen receptor (AR)-mediated transactivation by phosphorylating both RNA polymerase 2 complex proteins and AR at S81. CDK9 dephosphorylation mobilizes P-TEFb from an inhibitory 7SK ribonucleoprotein complex, but mechanisms targeting phosphatases to P-TEFb are unclear. We show that AR recruits protein phosphatase 1α (PP1α), resulting in P-TEFb mobilization and CDK9-mediated AR S81 phosphorylation. This increased pS81 enhances p300 recruitment, histone acetylation, BRD4 binding and subsequent further recruitment of P-TEFb, generating a positive feedback loop that sustains transcription. AR S81 is also phosphorylated by CDK1, and blocking basal CDK1-mediated S81 phosphorylation markedly suppresses AR activity and initiation of this positive feedback loop. Finally, androgen-independent AR activity in castration-resistant prostate cancer (CRPC) cells is driven by increased CDK1-mediated S81 phosphorylation. Collectively these findings reveal a mechanism involving PP1α, CDK9 and CDK1 that is used by AR to initiate and sustain P-TEFb activity, which may be exploited to drive AR in CRPC.
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Regulação Neoplásica da Expressão Gênica , Fator B de Elongação Transcricional Positiva/metabolismo , Neoplasias da Próstata/genética , Proteína Fosfatase 1/metabolismo , Receptores Androgênicos/metabolismo , Antagonistas de Receptores de Andrógenos/farmacologia , Proteína Quinase CDC2/metabolismo , Linhagem Celular Tumoral , Cromatina/metabolismo , Quinase 9 Dependente de Ciclina/metabolismo , Retroalimentação Fisiológica , Humanos , Masculino , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/metabolismo , Ativação TranscricionalRESUMO
Next-generation sequencing promotes identification of mutations in non-BRCA1/2 genes in hereditary cancer families. The contribution of mutations in moderate penetrance genes to hereditary cancer risk is not well established. Here, we report a family with early onset breast and fallopian tube cancer that was identified as carrying germline mutations in BARD1 and ATM genes. Loss of heterozygosity studies suggest a causative role of the BARD1 mutation in the development of primary peritoneal cancer, but fail to confirm an association between germline ATM mutations and breast cancer development in this family. Complexities in interpreting implications of mutations in moderate-risk cancer susceptibility genes are discussed.
Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/genética , Neoplasias da Mama/genética , Predisposição Genética para Doença/genética , Neoplasias Ovarianas/genética , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Adulto , Idoso , Diagnóstico Diferencial , Feminino , Humanos , Mutação , LinhagemRESUMO
The androgen receptor (AR) is a key factor that regulates the behavior and fate of prostate cancer cells. The AR-regulated network is activated when AR binds enhancer elements and modulates specific enhancer-promoter looping. Kallikrein-related peptidase 3 (KLK3), which codes for prostate-specific antigen (PSA), is a well-known AR-regulated gene and its upstream enhancers produce bidirectional enhancer RNAs (eRNAs), termed KLK3e. Here, we demonstrate that KLK3e facilitates the spatial interaction of the KLK3 enhancer and the KLK2 promoter and enhances long-distance KLK2 transcriptional activation. KLK3e carries the core enhancer element derived from the androgen response element III (ARE III), which is required for the interaction of AR and Mediator 1 (Med1). Furthermore, we show that KLK3e processes RNA-dependent enhancer activity depending on the integrity of core enhancer elements. The transcription of KLK3e was detectable and its expression is significantly correlated with KLK3 (R(2) = 0.6213, P < 5 × 10(-11)) and KLK2 (R(2) = 0.5893, P < 5 × 10(-10)) in human prostate tissues. Interestingly, RNAi silencing of KLK3e resulted in a modest negative effect on prostate cancer cell proliferation. Accordingly, we report that an androgen-induced eRNA scaffolds the AR-associated protein complex that modulates chromosomal architecture and selectively enhances AR-dependent gene expression.
Assuntos
Elementos Facilitadores Genéticos , Regulação Neoplásica da Expressão Gênica , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Animais , Células COS , Linhagem Celular Tumoral , Chlorocebus aethiops , Inativação Gênica , Humanos , Calicreínas/metabolismo , Masculino , Subunidade 1 do Complexo Mediador/metabolismo , Regiões Promotoras Genéticas , Próstata/metabolismo , Antígeno Prostático Específico/metabolismo , Interferência de RNA , Sequências Reguladoras de Ácido Nucleico , Calicreínas Teciduais/metabolismo , Transcrição Gênica , Ativação TranscricionalRESUMO
INTRODUCTION: High Grade Prostatic Intraepithelial Neoplasia (HGPIN) is the putative precursor lesion to prostatic adenocarcinoma (PCa), but the precise relationship between HGPIN and PCa remains unclear. METHODS: We performed a molecular case study in which we studied mutation profiles of six tumor-associated HGPIN lesions in a single case of TMPRSS2:ERG fusion positive Gleason score 7 PCa that we had previously mapped for somatic mutations in adjacent Gleason patterns 3 and 4 foci, using microdissection and targeted deep-sequencing. RESULTS: A total of 32 tumor-specific mutated sites were successfully amplified and sequenced, including 25 truncal mutations and 7 mutations that were specific to either the Gleason pattern 3 or pattern 4 foci. All six HGPIN foci shared the same tumor-specific TMPRSS2:ERG fusion breakpoint, establishing that they were all clonally related to the adjacent invasive tumor. Among the 32 gene targets mutated in the invasive tumor, only mutation of the OR2AP1 gene, a truncal mutation, was found in a single focus of HGPIN. The remaining gene targets that were successfully sequenced were wild-type in all other HGPIN foci. DISCUSSION: This study demonstrates the feasibility of targeted mutation profiling of HGPIN lesions, which will be important to understand PCa tumorigenesis. The results in this case, showing a remarkable absence of truncal mutations in HGPIN lesions bearing the tumor-specific ERG fusion, indicate HGPIN lesions may be relatively stable genetically and argue against a stepwise clonal evolution model of HGPIN to PCa. Prostate 76:1227-1236, 2016. © 2016 Wiley Periodicals, Inc.
Assuntos
Perfilação da Expressão Gênica/métodos , Mutação/genética , Neoplasia Prostática Intraepitelial/diagnóstico , Neoplasia Prostática Intraepitelial/genética , Neoplasias da Próstata/diagnóstico , Neoplasias da Próstata/genética , Adenocarcinoma/diagnóstico , Idoso , Humanos , Masculino , Invasividade Neoplásica/diagnóstico , Invasividade Neoplásica/patologiaRESUMO
PURPOSE: Spliced variant forms of androgen receptor were recently identified in castration resistant prostate cancer cell lines and clinical samples. We identified the cistrome and gene signature of androgen receptor splice variants in castration resistant prostate cancer cell lines and determined the clinical significance of androgen receptor splice variant regulated genes. MATERIALS AND METHODS: The castration resistant prostate cancer cell line 22Rv1, which expresses full-length androgen receptor and androgen receptor splice variants endogenously, was used as the research model. We established 22Rv1-ARFL(-)/ARV(+) and 22Rv1-ARFL(-)/ARV(-) through RNA interference. Chromatin immunoprecipitation coupled with next generation sequencing and microarray techniques were used to identify the cistrome and gene expression profiles of androgen receptor splice variants in the absence of androgen. RESULTS: Androgen receptor splice variant binding sites were identified in 22Rv1-ARFL(-)/ARV(+). A gene set was regulated uniquely by androgen receptor splice variants but not by full-length androgen receptor in the absence of androgen. Integrated analysis revealed that some genes were directly modulated by androgen receptor splice variants. Unsupervised clustering analysis showed that the androgen receptor splice variant gene signature differentiated benign from malignant prostate tissue as well as localized prostate cancer from metastatic castration resistant prostate cancer specimens. Some genes that were modulated uniquely by androgen receptor splice variants also correlated with histological grade and biochemical failure. CONCLUSIONS: Androgen receptor splice variants can bind to DNA independent of full-length androgen receptor in the absence of androgen and modulate a unique set of genes that is not regulated by full-length androgen receptor. The androgen receptor splice variant gene signature correlates with disease progression. It distinguishes primary cancer from castration resistant prostate cancer specimens and benign from malignant prostate specimens.
Assuntos
Regulação Neoplásica da Expressão Gênica , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/genética , Transcriptoma , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Isoformas de Proteínas , Células Tumorais CultivadasRESUMO
Gene expression profiling has identified breast cancer (BCa) subtypes, including an aggressive basal-like (BL) subtype. The molecular signals underlying the behavior observed in BL-BCa group are largely unknown, although recent results indicate a prevalent increase in Wnt/ß-catenin activity. Our immunohistochemistry study confirmed that SOX9, one of the BL-BCa signature genes, was expressed by most BL-BCa, and its expression correlated with indicators of poor prognosis. Importantly, BCa gene expression profiling strongly associated SOX9 with the expression of Wnt/ß-catenin pathway components, LRP6 and TCF4. In cancer cell lines, SOX9 silencing reduced cell proliferation and invasion, LRP6 and TCF4 transcription, and decreased Wnt/ß-catenin activation. SOX9 expression was also increased by Wnt, indicating that SOX9 is at the center of a positive feedback loop that enhances Wnt/ß-catenin signaling. Consistently, SOX9 overexpression in BCa cell lines and transgenic SOX9 expression in breast epithelium caused increased LRP6 and TCF4 expression and Wnt/ß-catenin activation. These results identify SOX9-mediated Wnt/ß-catenin activation as one of the molecular mechanisms underlying aberrant Wnt/ß-catenin activity in BCa, especially in the BL-BCa subgroup.
Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/biossíntese , Neoplasias da Mama/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/biossíntese , Neoplasias Mamárias Animais/metabolismo , Proteínas de Neoplasias/metabolismo , Fatores de Transcrição SOX9/metabolismo , Fatores de Transcrição/biossíntese , Via de Sinalização Wnt , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Humanos , Imuno-Histoquímica , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/patologia , Glândulas Mamárias Humanas/metabolismo , Glândulas Mamárias Humanas/patologia , Neoplasias Mamárias Animais/genética , Neoplasias Mamárias Animais/patologia , Camundongos , Camundongos Transgênicos , Proteínas de Neoplasias/genética , Fatores de Transcrição SOX9/genética , Fator de Transcrição 4 , Fatores de Transcrição/genética , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , beta Catenina/genética , beta Catenina/metabolismoRESUMO
Degradation of unliganded androgen receptor (AR) in prostate cancer cells can be prevented by proteasome inhibition, but this is associated with only modest increases in polyubiquitylated AR. An inhibitor (VLX1570) of the deubiquitylases associated with the proteasome did not increase ubiquitylation of unliganded AR, indicating that AR is not targeted by these deubiquitylases. We then identified a series of AR ubiquitylation sites, including a not previously identified site at K911, as well as methylation sites and previously identified phosphorylation sites. Mutagenesis of K911 increases AR stability, chromatin binding, and transcriptional activity. We further found that K313, a previously reported ubiquitylation site, could also be methylated and acetylated. Mutagenesis of K313, in combination with K318, increases AR transcriptional activity, indicating that distinct posttranslational modifications at K313 differentially regulate AR activity. Together these studies expand the spectrum of AR posttranslational modifications, and indicate that the K911 site may regulate AR turnover on chromatin.