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BACKGROUND: Benign prostatic hyperplasia (BPH) is the most common urologic disease in aging males, affecting 50% of men over 50 and up to 80% of men over 80 years old. Its negative impact on health-related quality of life implores further investigation into its risk factors and strategies for effective management. Although the exact molecular mechanisms underlying pathophysiological onset of BPH are poorly defined, the current hypothesized contributors to BPH and lower urinary tract symptoms (LUTS) include aging, inflammation, metabolic syndrome, and hormonal changes. These processes are indirectly influenced by circadian rhythm disruption. In this article, we review the recent evidence on the potential association of light changes/circadian rhythm disruption and the onset of BPH and impact on treatment. METHODS: A narrative literature review was conducted using PubMed and Google Scholar to identify supporting evidence. The articles referenced ranged from 1975 to 2023. RESULTS: A clear relationship between BPH/LUTS and circadian rhythm disruption is yet to be established. However, common mediators influence both diseases, including proinflammatory states, metabolic syndrome, and hormonal regulation that can be asserted to circadian disruption. Some studies have identified a possible relationship between general LUTS and sleep disturbance, but little research has been done on the medical management of these diseases and how circadian rhythm disruption further affects treatment outcomes. CONCLUSIONS: There is evidence to implicate a relationship between BPH/LUTS and circadian rhythm disruptions. However, there is scarce literature on potential specific link in medical management of the disease and treatment outcomes with circadian rhythm disruption. Further study is warranted to provide BPH patients with insights into circadian rhythm directed appropriate interventions.
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Sintomas do Trato Urinário Inferior , Síndrome Metabólica , Hiperplasia Prostática , Masculino , Humanos , Idoso de 80 Anos ou mais , Qualidade de Vida , Síndrome Metabólica/complicações , Sintomas do Trato Urinário Inferior/etiologia , Fatores de RiscoRESUMO
Enhancer regions and transcription start sites of estrogen-target regulated genes are connected by means of Estrogen Receptor long-range chromatin interactions. Yet, the complete molecular mechanisms controlling the transcriptional output of engaged enhancers and subsequent activation of coding genes remain elusive. Here, we report that CTCF binding to enhancer RNAs is enriched when breast cancer cells are stimulated with estrogen. CTCF binding to enhancer regions results in modulation of estrogen-induced gene transcription by preventing Estrogen Receptor chromatin binding and by hindering the formation of additional enhancer-promoter ER looping. Furthermore, the depletion of CTCF facilitates the expression of target genes associated with cell division and increases the rate of breast cancer cell proliferation. We have also uncovered a genomic network connecting loci enriched in cell cycle regulator genes to nuclear lamina that mediates the CTCF function. The nuclear lamina and chromatin interactions are regulated by estrogen-ER. We have observed that the chromatin loops formed when cells are treated with estrogen establish contacts with the nuclear lamina. Once there, the portion of CTCF associated with the nuclear lamina interacts with enhancer regions, limiting the formation of ER loops and the induction of genes present in the loop. Collectively, our results reveal an important, unanticipated interplay between CTCF and nuclear lamina to control the transcription of ER target genes, which has great implications in the rate of growth of breast cancer cells.
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Núcleo Celular/metabolismo , Cromatina/metabolismo , Receptores de Estrogênio/metabolismo , Proteínas Repressoras/fisiologia , Sítios de Ligação , Fator de Ligação a CCCTC , Elementos Facilitadores Genéticos , Estrogênios/fisiologia , Humanos , Células MCF-7 , Ligação Proteica , Ativação TranscricionalRESUMO
BACKGROUND: Epigenetic information can be used to identify clinically relevant genomic variants single nucleotide polymorphisms (SNPs) of functional importance in cancer development. Super-enhancers are cell-specific DNA elements, acting to determine tissue or cell identity and driving tumor progression. Although previous approaches have been tried to explain risk associated with SNPs in regulatory DNA elements, so far epigenetic readers such as bromodomain containing protein 4 (BRD4) and super-enhancers have not been used to annotate SNPs. In prostate cancer (PC), androgen receptor (AR) binding sites to chromatin have been used to inform functional annotations of SNPs. RESULTS: Here we establish criteria for enhancer mapping which are applicable to other diseases and traits to achieve the optimal tissue-specific enrichment of PC risk SNPs. We used stratified Q-Q plots and Fisher test to assess the differential enrichment of SNPs mapping to specific categories of enhancers. We find that BRD4 is the key discriminant of tissue-specific enhancers, showing that it is more powerful than AR binding information to capture PC specific risk loci, and can be used with similar effect in breast cancer (BC) and applied to other diseases such as schizophrenia. CONCLUSIONS: This is the first study to evaluate the enrichment of epigenetic readers in genome-wide associations studies for SNPs within enhancers, and provides a powerful tool for enriching and prioritizing PC and BC genetic risk loci. Our study represents a proof of principle applicable to other diseases and traits that can be used to redefine molecular mechanisms of human phenotypic variation.
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Neoplasias da Mama/genética , Elementos Facilitadores Genéticos , Regulação Neoplásica da Expressão Gênica , Predisposição Genética para Doença , Proteínas Nucleares/metabolismo , Neoplasias da Próstata/genética , Locos de Características Quantitativas , Fatores de Transcrição/metabolismo , Sítios de Ligação , Neoplasias da Mama/metabolismo , Proteínas de Ciclo Celular , Mapeamento Cromossômico , Biologia Computacional/métodos , Epigênese Genética , Feminino , Estudo de Associação Genômica Ampla , Histonas/metabolismo , Humanos , Masculino , Especificidade de Órgãos/genética , Polimorfismo de Nucleotídeo Único , Neoplasias da Próstata/metabolismo , Ligação Proteica , Receptores Androgênicos/metabolismoRESUMO
BACKGROUND: Although endocrine therapy has been used for decades, its influence on the expression of microRNAs (miRNAs) in clinical tissue specimens has not been analyzed. Moreover, the effects of the TMPRSS2:ERG fusion on the expression of miRNAs in hormone naïve and endocrine-treated prostate cancers are poorly understood. METHODS: We used clinical material from a neoadjuvant trial consisting of 28 men treated with goserelin (n = 8), bicalutamide (n = 9), or no treatment (n = 11) for 3 months prior to radical prostatectomy. Freshly frozen specimens were used for microarray analysis of 723 human miRNAs. Specific miRNA expression in cancer, benign epithelium and stromal tissue compartments was predicted with an in silico Bayesian modeling tool. RESULTS: The expression of 52, 44, and 34 miRNAs was affected >1.4-fold by the endocrine treatment in the cancer, non-malignant epithelium, and stromal compartments, respectively. Of the 52 miRNAs, only 10 were equally affected by the two treatment modalities in the cancer compartment. Twenty-six of the 52 genes (50%) showed AR binding sites in their proximity in either VCaP or LNCaP cell lines. Forty-seven miRNAs were differentially expressed in TMPRSS2:ERG fusion positive compared with fusion negative cases. Endocrine treatment reduced the differences between fusion positive and negative cases. CONCLUSIONS: Goserelin treatment and bicalutamide treatment mostly affected the expression of different miRNAs. The effect clearly varied in different tissue compartments. TMPRSS2:ERG fusion positive and negative cases showed differential expression of miRNAs, and the difference was diminished by androgen ablation.
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Adenocarcinoma/tratamento farmacológico , Anilidas/uso terapêutico , Antineoplásicos/uso terapêutico , Gosserrelina/uso terapêutico , MicroRNAs/genética , Nitrilas/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Compostos de Tosil/uso terapêutico , Adenocarcinoma/metabolismo , Simulação por Computador , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Fusão Gênica/efeitos dos fármacos , Humanos , Hibridização in Situ Fluorescente , Masculino , MicroRNAs/metabolismo , Terapia Neoadjuvante , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas de Fusão Oncogênica/genética , Prostatectomia , Neoplasias da Próstata/metabolismo , Células Tumorais CultivadasRESUMO
Endocrine therapy by castration or anti-androgens is the gold standard treatment for advanced prostate cancer. Although it has been used for decades, the molecular consequences of androgen deprivation are incompletely known and biomarkers of its resistance are lacking. In this study, we studied the molecular mechanisms of hormonal therapy by comparing the effect of bicalutamide (anti-androgen), goserelin (GnRH agonist) and no therapy, followed by radical prostatectomy. For this purpose, 28 men were randomly assigned to treatment groups. Freshly frozen specimens were used for gene expression profiling for all known protein-coding genes. An in silico Bayesian modelling tool was used to assess cancer-specific gene expression from heterogeneous tissue specimens. The expression of 128 genes was > two-fold reduced by the treatments. Only 16% of the altered genes were common in both treatment groups. Of the 128 genes, only 24 were directly androgen-regulated genes, according to re-analysis of previous data on gene expression, androgen receptor-binding sites and histone modifications in prostate cancer cell line models. The tumours containing TMPRSS2-ERG fusion showed higher gene expression of genes related to proliferation compared to the fusion-negative tumours in untreated cases. Interestingly, endocrine therapy reduced the expression of one-half of these genes and thus diminished the differences between the fusion-positive and -negative samples. This study reports the significantly different effects of an anti-androgen and a GnRH agonist on gene expression in prostate cancer cells. TMPRSS2-ERG fusion seems to bring many proliferation-related genes under androgen regulation.
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Antagonistas de Androgênios/administração & dosagem , Anilidas/administração & dosagem , Antineoplásicos Hormonais/administração & dosagem , Castração/métodos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Gosserrelina/administração & dosagem , Neoplasias Hormônio-Dependentes/tratamento farmacológico , Nitrilas/administração & dosagem , Neoplasias da Próstata/tratamento farmacológico , Compostos de Tosil/administração & dosagem , Administração Oral , Análise de Variância , Teorema de Bayes , Quimioterapia Adjuvante , Distribuição de Qui-Quadrado , Finlândia , Perfilação da Expressão Gênica/métodos , Hormônio Liberador de Gonadotropina/agonistas , Hormônio Liberador de Gonadotropina/metabolismo , Humanos , Injeções Subcutâneas , Masculino , Terapia Neoadjuvante , Neoplasias Hormônio-Dependentes/genética , Neoplasias Hormônio-Dependentes/metabolismo , Neoplasias Hormônio-Dependentes/cirurgia , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas de Fusão Oncogênica/genética , Prostatectomia , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/cirurgia , Resultado do TratamentoRESUMO
Androgen Receptor (AR) signaling inhibitors, including enzalutamide, are treatment options for patients with metastatic castration-resistant prostate cancer (mCRPC), but resistance inevitably develops. Using metastatic samples from a prospective phase II clinical trial, we epigenetically profiled enhancer/promoter activities with H3K27ac chromatin immunoprecipitation followed by sequencing, before and after AR-targeted therapy. We identified a distinct subset of H3K27ac-differentially marked regions that associated with treatment responsiveness. These data were successfully validated in mCRPC patient-derived xenograft models (PDX). In silico analyses revealed HDAC3 as a critical factor that can drive resistance to hormonal interventions, which we validated in vitro . Using cell lines and mCRPC PDX tumors in vitro , we identified drug-drug synergy between enzalutamide and the pan-HDAC inhibitor vorinostat, providing therapeutic proof-of-concept. These findings demonstrate rationale for new therapeutic strategies using a combination of AR and HDAC inhibitors to improve patient outcome in advanced stages of mCRPC.
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Metastatic and high-risk localized prostate cancer respond to hormone therapy but outcomes vary. Following a pre-specified statistical plan, we used Cox models adjusted for clinical variables to test associations with survival of multi-gene expression-based classifiers from 781 patients randomized to androgen deprivation with or without abiraterone in the STAMPEDE trial. Decipher score was strongly prognostic (p<2×10-5) and identified clinically-relevant differences in absolute benefit, especially for localized cancers. In metastatic disease, classifiers of proliferation, PTEN or TP53 loss and treatment-persistent cells were prognostic. In localized disease, androgen receptor activity was protective whilst interferon signaling (that strongly associated with tumor lymphocyte infiltration) was detrimental. Post-Operative Radiation-Therapy Outcomes Score was prognostic in localized but not metastatic disease (interaction p=0.0001) suggesting the impact of tumor biology on clinical outcome is context-dependent on metastatic state. Transcriptome-wide testing has clinical utility for advanced prostate cancer and identified worse outcomes for localized cancers with tumor-promoting inflammation.
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BACKGROUND: Castration-resistant prostate cancers (CRPCs) overexpress often androgen receptor (AR). Here, we investigated the effect of AR overexpression on the dynamics of AR loading and RNA polymerase II (RNA Pol II) recruitment to chromatin. Acetylation of histone 3 (AcH3) on lysines 9 and 14 (K9 and K14) was also studied. METHODS: We used an LNCaP-based AR overexpression cell line model that includes a control line and two sublines, LNCaP-ARmo and LNCaP-ARhi, which overexpress AR twofold to threefold and fourfold to fivefold, respectively. Cells were exposed to 1 or 100 nM of dihydrotestosterone (DHT). Chromatin immunoprecipitation (ChIP) on the promoters and enhancers of prostate specific antigen (PSA) and transmembrane protease, serine 2 (TMPRSS2) genes was performed. qRT-PCR was used to measure the levels of PSA and TMPRSS2 transcripts. RESULTS: Upon stimulation with 1 nM DHT, AR and RNA Pol II were recruited onto PSA and TMPRSS2 enhancer regions to a greater extent (P < 0.05) in AR-overexpressing cells compared to control cells. The difference in AR loading between the control and AR-overexpressing cells was abolished by a higher DHT concentration. The ratio of AcH3/H3 was increased in AR-overexpressing cells. The induction of transcription of PSA and TMPRSS2 occurred earlier in the AR-overexpressing cells. CONCLUSIONS: Our findings suggest that the levels of AR potentiate the recruitment of the AR, as well as components of the basic transcription machinery, to chromatin and affect the acetylation of histones in the presence of low levels of androgens. These changes result in enhanced gene transcription of AR target genes.
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Cromatina/genética , Neoplasias da Próstata/genética , RNA Polimerase II/metabolismo , Receptores Androgênicos/metabolismo , Acetilação/efeitos dos fármacos , Androgênios/farmacologia , Linhagem Celular Tumoral , Cromatina/metabolismo , Imunoprecipitação da Cromatina , Histonas/metabolismo , Humanos , Masculino , Orquiectomia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/cirurgia , Receptores Androgênicos/genéticaRESUMO
Autophagy and autophagy-associated genes are implicated in a growing list of cellular, physiological, and pathophysiological processes and conditions. Therefore, it is ever more important to be able to reliably monitor and quantify autophagic activity. Whereas autophagic markers, such as LC3 can provide general indications about autophagy, specific and accurate detection of autophagic activity requires assessment of autophagic cargo flux. Here, we provide protocols on how to monitor bulk and selective autophagy by the use of inducible expression of exogenous probes based on the fluorescent coral protein Keima. To exemplify and demonstrate the power of this system, we provide data obtained by analyses of cytosolic and mitochondrially targeted Keima probes in human retinal epithelial cells treated with the mTOR-inhibitor Torin1 or with the iron chelator deferiprone (DFP). Our data indicate that Torin1 induces autophagic flux of cytosol and mitochondria to a similar degree, that is, compatible with induction of bulk autophagy, whereas DFP induces a highly selective form of mitophagy that efficiently excludes cytosol.
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Autofagia , Proteínas Associadas aos Microtúbulos , Autofagia/fisiologia , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias/metabolismo , MitofagiaRESUMO
Single-cell RNA sequencing studies have suggested that total mRNA content correlates with tumor phenotypes. Technical and analytical challenges, however, have so far impeded at-scale pan-cancer examination of total mRNA content. Here we present a method to quantify tumor-specific total mRNA expression (TmS) from bulk sequencing data, taking into account tumor transcript proportion, purity and ploidy, which are estimated through transcriptomic/genomic deconvolution. We estimate and validate TmS in 6,590 patient tumors across 15 cancer types, identifying significant inter-tumor variability. Across cancers, high TmS is associated with increased risk of disease progression and death. TmS is influenced by cancer-specific patterns of gene alteration and intra-tumor genetic heterogeneity as well as by pan-cancer trends in metabolic dysregulation. Taken together, our results indicate that measuring cell-type-specific total mRNA expression in tumor cells predicts tumor phenotypes and clinical outcomes.
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Neoplasias , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Heterogeneidade Genética , Genômica , RNA Mensageiro/genética , Progressão da DoençaRESUMO
BACKGROUND: Androgens play a critical role in the growth of both androgen dependent and castration-resistant prostate cancer (CRPC). Only a few micro-RNAs (miRNAs) have been suggested to be androgen regulated. We aim to identify androgen regulated miRNAs. METHODS: We utilized LNCaP derived model, we have established, and which overexpresses the androgen receptor (AR), the VCaP cell line, and 13 intact-castrated prostate cancer (PC) xenograft pairs, as well as clinical specimens of untreated (PC) and CRPC. The expression of miRNAs was analyzed by microarrays and quantitative RT-PCR (Q-RT-PCR). Transfection of pre-miR-141 and anti-miR-141 was also used. RESULTS: Seventeen miRNAs were > 1.5-fold up- or downregulated upon dihydrotestosterone (DHT) treatment in the cell lines, and 42 after castration in the AR-positive xenografts. Only four miRNAs (miR-10a, miR-141, miR-150*, and miR-1225-5p) showed similar androgen regulation in both cell lines and xenografts. Of those, miR-141 was found to be expressed more in PC and CRPC compared to benign prostate hyperplasia. Additionally, the overexpression of miR-141 enhanced growth of parental LNCaP cells while inhibition of miR-141 by anti-miR-141 suppressed the growth of the LNCaP subline overexpressing AR. CONCLUSIONS: Only a few miRNAs were found to be androgen-regulated in both cell lines and xenografts models. Of those, the expression of miR-141 was upregulated in cancer. The ectopic overexpression of miR-141 increased growth of LNCaP cell suggesting it may contribute to the progression of PC.
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Androgênios/metabolismo , Regulação Neoplásica da Expressão Gênica , MicroRNAs/metabolismo , Neoplasias Hormônio-Dependentes/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Animais , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/genética , Linhagem Celular Tumoral , Di-Hidrotestosterona/farmacologia , Perfilação da Expressão Gênica/métodos , Humanos , Masculino , Camundongos , MicroRNAs/biossíntese , MicroRNAs/genética , Neoplasias Experimentais , Neoplasias Hormônio-Dependentes/genética , Neoplasias Hormônio-Dependentes/patologia , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , RNA Neoplásico/química , RNA Neoplásico/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transfecção , Transplante HeterólogoRESUMO
The dysregulation of chromatin and epigenetics has been defined as the overarching cancer hallmark. By disrupting transcriptional regulation in normal cells and mediating tumor progression by promoting cancer cell plasticity, this process has the ability to mediate all defined hallmarks of cancer. In this review, we collect and assess evidence on the contribution of chromatin and epigenetic dysregulation in prostate cancer. We highlight important mechanisms leading to prostate carcinogenesis, the emergence of castration-resistance upon treatment with androgen deprivation therapy, and resistance to antiandrogens. We examine in particular the contribution of chromatin structure and epigenetics to cell lineage commitment, which is dysregulated during tumorigenesis, and cell plasticity, which is altered during tumor progression.
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BACKGROUND: Prostate cancer (PCa) is the second most common tumour diagnosed in men. Tumoral heterogeneity in PCa creates a significant challenge to develop robust prognostic markers and novel targets for therapy. An analysis of gene regulatory networks (GRNs) in PCa may provide insight into progressive PCa. Herein, we exploited a graph-based enrichment score to integrate data from GRNs identified in preclinical prostate orthografts and differentially expressed genes in clinical resected PCa. We identified active regulons (transcriptional regulators and their targeted genes) associated with PCa recurrence following radical prostatectomy. METHODS: The expression of known transcription factors and co-factors was analysed in a panel of prostate orthografts (n = 18). We searched for genes (as part of individual GRNs) predicted to be regulated by the highest number of transcriptional factors. Using differentially expressed gene analysis (on a per sample basis) coupled with gene graph enrichment analysis, we identified candidate genes and associated GRNs in PCa within the UTA cohort, with the most enriched regulon being JMJD6, which was further validated in two additional cohorts, namely EMC and ICGC cohorts. Cox regression analysis was performed to evaluate the association of the JMJD6 regulon activity with disease-free survival time in the three clinical cohorts as well as compared to three published prognostic gene signatures (TMCC11, BROMO-10 and HYPOXIA-28). RESULTS: 1308 regulons were correlated to transcriptomic data from the three clinical prostatectomy cohorts. The JMJD6 regulon was identified as the top enriched regulon in the UTA cohort and again validated in the EMC cohort as the top-ranking regulon. In both UTA and EMC cohorts, the JMJD6 regulon was significantly associated with cancer recurrence. Active JMJD6 regulon also correlated with disease recurrence in the ICGC cohort. Furthermore, Kaplan-Meier analysis confirmed shorter time to recurrence in patients with active JMJD6 regulon for all three clinical cohorts (UTA, EMC and ICGC), which was not the case for three published prognostic gene signatures (TMCC11, BROMO-10 and HYPOXIA-28). In multivariate analysis, the JMJD6 regulon status significantly predicted disease recurrence in the UTA and EMC, but not ICGC datasets, while none of the three published signatures significantly prognosticate for cancer recurrence. CONCLUSIONS: We have characterised gene regulatory networks from preclinical prostate orthografts and applied transcriptomic data from three clinical cohorts to evaluate the prognostic potential of the JMJD6 regulon.
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Guidelines for genetic testing have been established for multiple tumor types, frequently indicating the most confident molecularly targeted treatment options. However, considering the often-complex presentation of individual cancer patients, in addition to the combinatorial complexity and inherent uncertainties of molecular findings, deriving optimal treatment strategies frequently becomes very challenging. Here, we report a comprehensive analysis of a 68-year-old male with metastatic prostate cancer, encompassing pathology and MRI findings, transcriptomic results, and key genomics findings from whole-exome sequencing, both somatic aberrations and germline variants. We identify multiple somatic aberrations that are known to be enriched in prostate cancer, including a deletion of PTEN and a fusion transcript involving BRCA2. The gene expression patterns in the tumor biopsy were also strikingly similar to prostate tumor samples from TCGA. Furthermore, we detected multiple lines of evidence for homologous recombination repair deficiency (HRD), including a dominant contribution by mutational signature SBS3, which is specifically attributed to HRD. On the basis of the genomic and transcriptomic findings, and in light of the clinical case presentation, we discussed the personalized treatment options that exist for this patient and the various challenges that one faces in the process of translating high-throughput sequencing data towards treatment regimens.
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Long noncoding RNAs (lncRNAs) play pivotal roles in cancer development and progression, and some function in a highly cancer-specific manner. However, whether the cause of their expression is an outcome of a specific regulatory mechanism or nonspecific transcription induced by genome reorganization in cancer remains largely unknown. Here, we investigated a group of lncRNAs that we previously identified to be aberrantly expressed in prostate cancer (PC), called TPCATs. Our high-throughput real-time PCR experiments were integrated with publicly available RNA-seq and ChIP-seq data and revealed that the expression of a subset of TPCATs is driven by PC-specific transcription factors (TFs), especially androgen receptor (AR) and ETS-related gene (ERG). Our in vitro validations confirmed that AR and ERG regulated a subset of TPCATs, most notably for EPCART. Knockout of EPCART was found to reduce migration and proliferation of the PC cells in vitro. The high expression of EPCART and two other TPCATs (TPCAT-3-174133 and TPCAT-18-31849) were also associated with the biochemical recurrence of PC in prostatectomy patients and were independent prognostic markers. Our findings suggest that the expression of numerous PC-associated lncRNAs is driven by PC-specific mechanisms and not by random cellular events that occur during cancer development. Furthermore, we report three prospective prognostic markers for the early detection of advanced PC and show EPCART to be a functionally relevant lncRNA in PC.
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Regulação Neoplásica da Expressão Gênica , Neoplasias da Próstata/genética , RNA Longo não Codificante/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Fator 3-alfa Nuclear de Hepatócito/genética , Proteínas de Homeodomínio/genética , Humanos , Masculino , Estudos Prospectivos , Neoplasias da Próstata/patologia , Interferência de RNA , Receptores Androgênicos/genética , Regulador Transcricional ERG/genéticaRESUMO
Imprinted genes are highly expressed in the hypothalamus; however, whether specific imprinted genes affect hypothalamic neuromodulators and their functions is unknown. It has been suggested that Prader-Willi syndrome (PWS), a neurodevelopmental disorder caused by lack of paternal expression at chromosome 15q11-q13, is characterized by hypothalamic insufficiency. Here, we investigate the role of the paternally expressed Snord116 gene within the context of sleep and metabolic abnormalities of PWS, and we report a significant role of this imprinted gene in the function and organization of the 2 main neuromodulatory systems of the lateral hypothalamus (LH) - namely, the orexin (OX) and melanin concentrating hormone (MCH) - systems. We observed that the dynamics between neuronal discharge in the LH and the sleep-wake states of mice with paternal deletion of Snord116 (PWScrm+/p-) are compromised. This abnormal state-dependent neuronal activity is paralleled by a significant reduction in OX neurons in the LH of mutant mice. Therefore, we propose that an imbalance between OX- and MCH-expressing neurons in the LH of mutant mice reflects a series of deficits manifested in the PWS, such as dysregulation of rapid eye movement (REM) sleep, food intake, and temperature control.
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Comportamento Animal/fisiologia , Região Hipotalâmica Lateral/metabolismo , Hipotálamo/metabolismo , Orexinas/metabolismo , RNA Nucleolar Pequeno/genética , Sono/fisiologia , Animais , Modelos Animais de Doenças , Comportamento Alimentar , Região Hipotalâmica Lateral/fisiopatologia , Hormônios Hipotalâmicos/metabolismo , Melaninas/metabolismo , Camundongos , Neurônios/metabolismo , Hormônios Hipofisários/metabolismo , Síndrome de Prader-Willi/metabolismo , Síndrome de Prader-Willi/fisiopatologiaRESUMO
The MC1R/cAMP/MITF pathway is a key determinant for growth, differentiation, and survival of melanocytes and melanoma. MITF-M is the melanocyte-specific isoform of Microphthalmia-associated Transcription Factor (MITF) in human melanoma. Here we use two melanocyte cell lines to show that forced expression of hemagglutinin (HA) -tagged MITF-M through lentiviral transduction represents an oncogenic insult leading to consistent cell transformation of the immortalized melanocyte cell line Hermes 4C, being a melanocortin-1 receptor (MC1R) compound heterozygote, while not causing transformation of the MC1R wild type cell line Hermes 3C. The transformed HA-tagged MITF-M transduced Hermes 4C cells form colonies in soft agar and tumors in mice. Further, Hermes 4C cells display increased MITF chromatin binding, and transcriptional reprogramming consistent with an invasive melanoma phenotype. Mechanistically, forced expression of MITF-M drives the upregulation of the AXL tyrosine receptor kinase (AXL), with concomitant downregulation of phosphatase and tensin homolog (PTEN), leading to increased activation of the PI3K/AKT pathway. Treatment with AXL inhibitors reduces growth of the transformed cells by reverting AKT activation. In conclusion, we present a model system of melanoma development, driven by MITF-M in the context of MC1R loss of function, and independent of UV exposure. This model provides a basis for further studies of critical changes in the melanocyte transformation process.
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O-GlcNAc transferase (OGT) is a nutrient-sensitive glycosyltransferase that is overexpressed in prostate cancer, the most common cancer in males. We recently developed a specific and potent inhibitor targeting this enzyme, and here, we report a synthetic lethality screen using this compound. Our screen identified pan-cyclin-dependent kinase (CDK) inhibitor AT7519 as lethal in combination with OGT inhibition. Follow-up chemical and genetic approaches identified CDK9 as the major target for synthetic lethality with OGT inhibition in prostate cancer cells. OGT expression is regulated through retention of the fourth intron in the gene and CDK9 inhibition blunted this regulatory mechanism. CDK9 phosphorylates carboxy-terminal domain (CTD) of RNA Polymerase II to promote transcription elongation. We show that OGT inhibition augments effects of CDK9 inhibitors on CTD phosphorylation and general transcription. Finally, the combined inhibition of both OGT and CDK9 blocked growth of organoids derived from patients with metastatic prostate cancer, but had minimal effects on normal prostate spheroids. We report a novel synthetic lethal interaction between inhibitors of OGT and CDK9 that specifically kills prostate cancer cells, but not normal cells. Our study highlights the potential of combining OGT inhibitors with other treatments to exploit cancer-specific vulnerabilities. IMPLICATIONS: The primary contribution of OGT to cell proliferation is unknown, and in this study, we used a compound screen to indicate that OGT and CDK9 collaborate to sustain a cancer cell-specific pro-proliferative program. A better understanding of how OGT and CDK9 cross-talk will refine our understanding of this novel synthetic lethal interaction.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Quinase 9 Dependente de Ciclina/metabolismo , Inibidores Enzimáticos/farmacologia , N-Acetilglucosaminiltransferases/antagonistas & inibidores , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/enzimologia , Inibidores de Proteínas Quinases/farmacologia , Linhagem Celular Tumoral , Quinase 9 Dependente de Ciclina/antagonistas & inibidores , Sinergismo Farmacológico , Inibidores Enzimáticos/administração & dosagem , Humanos , Masculino , Terapia de Alvo Molecular , N-Acetilglucosaminiltransferases/metabolismo , Piperidinas/farmacologia , Neoplasias da Próstata/genética , Inibidores de Proteínas Quinases/administração & dosagem , Pirazóis/farmacologiaRESUMO
Tumor evolution is based on the ability to constantly mutate and activate different pathways under the selective pressure of targeted therapies. Epigenetic alterations including those of the chromatin structure are associated with tumor initiation, progression and drug resistance. Many cancers, including prostate cancer, present enlarged nuclei, and chromatin appears altered and irregular. These phenotypic changes are likely to result from epigenetic dysregulation. High-throughput sequencing applied to bulk samples and now to single cells has made it possible to study these processes in unprecedented detail. It is therefore timely to review the impact of chromatin relaxation and increased DNA accessibility on prostate cancer growth and drug resistance, and their effects on gene expression. In particular, we focus on the contribution of chromatin-associated proteins such as the bromodomain-containing proteins to chromatin relaxation. We discuss the consequence of this for androgen receptor transcriptional activity and briefly summarize wider gain-of-function effects on other oncogenic transcription factors and implications for more effective prostate cancer treatment.
Assuntos
Cromatina/metabolismo , Neoplasias da Próstata/metabolismo , Animais , Humanos , Masculino , Receptores Androgênicos/metabolismo , Fatores de Transcrição/metabolismoRESUMO
O-GlcNAc transferase (OGT) is overexpressed in aggressive prostate cancer. OGT modifies intra-cellular proteins via single sugar conjugation (O-GlcNAcylation) to alter their activity. We recently discovered the first fast-acting OGT inhibitor OSMI-2. Here, we probe the stability and function of the chromatin O-GlcNAc and identify transcription factors that coordinate with OGT to promote proliferation of prostate cancer cells. Methods: Chromatin immunoprecipitation (ChIP) coupled to sequencing (seq), formaldehyde-assisted isolation of regulatory elements, RNA-seq and reverse-phase protein arrays (RPPA) were used to study the importance of OGT for chromatin structure and transcription. Mass spectrometry, western blot, RT-qPCR, cell cycle analysis and viability assays were used to establish the role of OGT for MYC-related processes. Prostate cancer patient data profiled for both mRNA and protein levels were used to validate findings. Results: We show for the first time that OGT inhibition leads to a rapid loss of O-GlcNAc chromatin mark. O-GlcNAc ChIP-seq regions overlap with super-enhancers (SE) and MYC binding sites. OGT inhibition leads to down-regulation of SE-dependent genes. We establish the first O-GlcNAc chromatin consensus motif, which we use as a bait for mass spectrometry. By combining the proteomic data from oligonucleotide enrichment with O-GlcNAc and MYC ChIP-mass spectrometry, we identify host cell factor 1 (HCF-1) as an interaction partner of MYC. Inhibition of OGT disrupts this interaction and compromises MYC's ability to confer androgen-independent proliferation to prostate cancer cells. We show that OGT is required for MYC-mediated stabilization of mitotic proteins, including Cyclin B1, and/or the increased translation of their coding transcripts. This implies that increased expression of mRNA is not always required to achieve increased protein expression and confer aggressive phenotype. Indeed, high expression of Cyclin B1 protein has strong predictive value in prostate cancer patients (p=0.000014) while mRNA does not. Conclusions: OGT promotes SE-dependent gene expression. OGT activity is required for the interaction between MYC and HCF-1 and expression of MYC-regulated mitotic proteins. These features render OGT essential for the androgen-independent, MYC-driven proliferation of prostate cancer cells. Androgen-independency is the major mechanism of prostate cancer progression, and our study identifies OGT as an essential mediator in this process.