RESUMEN
We previously reported that miR-1 is among the most consistently down-regulated miRs in primary human prostate tumors. In this follow-up study, we further corroborated this finding in an independent data set and made the novel observation that miR-1 expression is further reduced in distant metastasis and is a candidate predictor of disease recurrence. Moreover, we performed in vitro experiments to explore the tumor suppressor function of miR-1. Cell-based assays showed that miR-1 is epigenetically silenced in human prostate cancer. Overexpression of miR-1 in these cells led to growth inhibition and down-regulation of genes in pathways regulating cell cycle progression, mitosis, DNA replication/repair and actin dynamics. This observation was further corroborated with protein expression analysis and 3'-UTR-based reporter assays, indicating that genes in these pathways are either direct or indirect targets of miR-1. A gene set enrichment analysis revealed that the miR-1-mediated tumor suppressor effects are globally similar to those of histone deacetylase inhibitors. Lastly, we obtained preliminary evidence that miR-1 alters the cellular organization of F-actin and inhibits tumor cell invasion and filipodia formation. In conclusion, our findings indicate that miR-1 acts as a tumor suppressor in prostate cancer by influencing multiple cancer-related processes and by inhibiting cell proliferation and motility.
Asunto(s)
Biomarcadores de Tumor/metabolismo , MicroARNs/metabolismo , Neoplasias de la Próstata/genética , Biomarcadores de Tumor/genética , Ciclo Celular/genética , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Reparación del ADN/genética , Epigénesis Genética , Genes Supresores de Tumor , Inhibidores de Histona Desacetilasas/farmacología , Humanos , Masculino , MicroARNs/genética , Mitosis , Metástasis de la Neoplasia , Recurrencia Local de Neoplasia/diagnóstico , Pronóstico , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Ultraconserved regions (UCR) are genomic segments of more than 200 base pairs that are evolutionarily conserved among mammalian species. They are thought to have functions as transcriptional enhancers and regulators of alternative splicing. Recently, it was shown that numerous RNAs are transcribed from these regions. These UCR-encoded transcripts (ucRNAs) were found to be expressed in a tissue- and disease-specific manner and may interfere with the function of other RNAs through RNA: RNA interactions. We hypothesized that ucRNAs have unidentified roles in the pathogenesis of human prostate cancer. In a pilot study, we examined ucRNA expression profiles in human prostate tumors. METHODS: Using a custom microarray with 962 probesets representing sense and antisense sequences for the 481 human UCRs, we examined ucRNA expression in resected, fresh-frozen human prostate tissues (57 tumors, 7 non-cancerous prostate tissues) and in cultured prostate cancer cells treated with either epigenetic drugs (the hypomethylating agent, 5-Aza 2'deoxycytidine, and the histone deacetylase inhibitor, trichostatin A) or a synthetic androgen, R1881. Expression of selected ucRNAs was also assessed by qRT-PCR and NanoString®-based assays. Because ucRNAs may function as RNAs that target protein-coding genes through direct and inhibitory RNA: RNA interactions, computational analyses were applied to identify candidate ucRNA:mRNA binding pairs. RESULTS: We observed altered ucRNA expression in prostate cancer (e.g., uc.106+, uc.477+, uc.363 + A, uc.454 + A) and found that these ucRNAs were associated with cancer development, Gleason score, and extraprostatic extension after controlling for false discovery (false discovery rate < 5% for many of the transcripts). We also identified several ucRNAs that were responsive to treatment with either epigenetic drugs or androgen (R1881). For example, experiments with LNCaP human prostate cancer cells showed that uc.287+ is induced by R1881 (P < 0.05) whereas uc.283 + A was up-regulated following treatment with combined 5-Aza 2'deoxycytidine and trichostatin A (P < 0.05). Additional computational analyses predicted RNA loop-loop interactions of 302 different sense and antisense ucRNAs with 1058 different mRNAs, inferring possible functions of ucRNAs via direct interactions with mRNAs. CONCLUSIONS: This first study of ucRNA expression in human prostate cancer indicates an altered transcript expression in the disease.
Asunto(s)
Adenocarcinoma/genética , Neoplasias de la Próstata/genética , ARN Neoplásico/genética , Transcriptoma , Adenocarcinoma/metabolismo , Anciano , Azacitidina/análogos & derivados , Azacitidina/farmacología , Estudios de Casos y Controles , Línea Celular Tumoral , Secuencia Conservada , Decitabina , Epigénesis Genética/efectos de los fármacos , Expresión Génica , Regulación Neoplásica de la Expresión Génica , Genoma Humano , Inhibidores de Histona Desacetilasas/farmacología , Humanos , Ácidos Hidroxámicos/farmacología , Masculino , Metribolona/farmacología , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Próstata/metabolismo , Neoplasias de la Próstata/metabolismo , ARN Mensajero/genética , ARN Neoplásico/metabolismo , ARN no Traducido/genética , ARN no Traducido/metabolismo , Congéneres de la Testosterona/farmacologíaRESUMEN
PURPOSE: The causes of disproportionate incidence and mortality of prostate cancer among African Americans (AA) remain elusive. The purpose of this study was to investigate the mechanistic role and assess clinical utility of the splicing factor heterogeneous nuclear ribonucleoprotein H1 (hnRNP H1) in prostate cancer progression among AA men. EXPERIMENTAL DESIGN: We employed an unbiased functional genomics approach coupled with suppressive subtractive hybridization (SSH) and custom cDNA microarrays to identify differentially expressed genes in microdissected tumors procured from age- and tumor grade-matched AA and Caucasian American (CA) men. Validation analysis was performed in independent cohorts and tissue microarrays. The underlying mechanisms of hnRNPH1 regulation and its impact on androgen receptor (AR) expression and tumor progression were explored. RESULTS: Aberrant coexpression of AR and hnRNPH1 and downregulation of miR-212 were detected in prostate tumors and correlate with disease progression in AA men compared with CA men. Ectopic expression of miR-212 mimics downregulated hnRNPH1 transcripts, which in turn reduced expression of AR and its splice variant AR-V7 (or AR3) in prostate cancer cells. hnRNPH1 physically interacts with AR and steroid receptor coactivator-3 (SRC-3) and primes activation of androgen-regulated genes in a ligand-dependent and independent manner. siRNA silencing of hnRNPH1 sensitized prostate cancer cells to bicalutamide and inhibited prostate tumorigenesis in vivo CONCLUSIONS: Our findings define novel roles for hnRNPH1 as a putative oncogene, splicing factor, and an auxiliary AR coregulator. Targeted disruption of the hnRNPH1-AR axis may have therapeutic implications to improve clinical outcomes in patients with advanced prostate cancer, especially among AA men.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Ribonucleoproteína Heterogénea-Nuclear Grupo F-H/genética , MicroARNs/genética , Neoplasias de la Próstata Resistentes a la Castración/genética , Receptores Androgénicos/genética , Empalme Alternativo , Andrógenos/metabolismo , Anilidas/farmacología , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular , Análisis por Conglomerados , Progresión de la Enfermedad , Resistencia a Antineoplásicos/genética , Perfilación de la Expresión Génica , Silenciador del Gen , Ribonucleoproteína Heterogénea-Nuclear Grupo F-H/metabolismo , Humanos , Masculino , Persona de Mediana Edad , Nitrilos/farmacología , Coactivador 3 de Receptor Nuclear/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/diagnóstico , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Unión Proteica , Interferencia de ARN , Receptores Androgénicos/metabolismo , Elementos de Respuesta , Compuestos de Tosilo/farmacologíaRESUMEN
Smokers develop metastatic prostate cancer more frequently than nonsmokers, suggesting that a tobacco-derived factor is driving metastatic progression. To identify smoking-induced alterations in human prostate cancer, we analyzed gene and protein expression patterns in tumors collected from current, past, and never smokers. By this route, we elucidated a distinct pattern of molecular alterations characterized by an immune and inflammation signature in tumors from current smokers that were either attenuated or absent in past and never smokers. Specifically, this signature included elevated immunoglobulin expression by tumor-infiltrating B cells, NF-κB activation, and increased chemokine expression. In an alternate approach to characterize smoking-induced oncogenic alterations, we also explored the effects of nicotine in human prostate cancer cells and prostate cancer-prone TRAMP mice. These investigations showed that nicotine increased glutamine consumption and invasiveness of cancer cells in vitro and accelerated metastatic progression in tumor-bearing TRAMP mice. Overall, our findings suggest that nicotine is sufficient to induce a phenotype resembling the epidemiology of smoking-associated prostate cancer progression, illuminating a novel candidate driver underlying metastatic prostate cancer in current smokers.
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Inflamación/metabolismo , Neoplasias de la Próstata/inmunología , Fumar/efectos adversos , Transcriptoma , Animales , Línea Celular Tumoral , Núcleo Celular/metabolismo , Humanos , Inmunoglobulinas/genética , Interleucina-8/sangre , Masculino , Ratones , FN-kappa B/metabolismo , Invasividad Neoplásica , Metástasis de la Neoplasia , Nicotina/farmacología , Neoplasias de la Próstata/etiología , Neoplasias de la Próstata/patología , Proteínas Proto-Oncogénicas c-akt/metabolismoRESUMEN
Inducible nitric oxide synthase (NOS2) is involved in wound healing, angiogenesis, and carcinogenesis. NOS2 upregulation and increased nitric oxide (NO) production affect the redox state of cells and can induce protein, lipid, and DNA modifications. To investigate whether NOS2 levels influence survival of breast cancer patients, we examined NOS2 expression and its association with tumor markers and survival in 248 breast tumors. In multivariable survival analysis, increased NOS2 predicted inferior survival in women with estrogen receptor α-negative (ER-negative) tumors. Microdissected tumor epithelium from ER-negative tumors with high NOS2 had increased IL-8 and a gene expression signature characteristic of basal-like breast cancer with poor prognosis. In cell culture, NO only induced selected signature genes in ER-negative breast cancer cells. ER transgene expression in ER-negative cells inhibited NO-induced upregulation of the stem cell marker CD44 and other proteins encoded by signature genes, but not of IL-8. Exposure to NO also enhanced cell motility and invasion of ER-negative cells. Last, pathway analysis linked the tumor NOS2 gene signature to c-Myc activation. Thus, NOS2 is associated with a basal-like transcription pattern and poor survival of ER-negative patients.
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Neoplasias de la Mama/enzimología , Óxido Nítrico Sintasa de Tipo II/metabolismo , Receptores de Estrógenos/metabolismo , Animales , Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/diagnóstico , Neoplasias de la Mama/patología , Línea Celular Tumoral , Movimiento Celular , Femenino , Perfilación de la Expresión Génica , Humanos , Interleucina-8/genética , Interleucina-8/metabolismo , Datos de Secuencia Molecular , Óxido Nítrico/metabolismo , Pronóstico , Análisis de Supervivencia , Tasa de SupervivenciaRESUMEN
BACKGROUND: Perineural invasion (PNI) is the dominant pathway for local invasion in prostate cancer. To date, only few studies have investigated the molecular differences between prostate tumors with PNI and those without it. METHODS: To evaluate the involvement of both microRNAs and protein-coding genes in PNI, we determined their genome-wide expression with a custom microRNA microarray and Affymetrix GeneChips in 50 prostate adenocarcinomas with PNI and 7 without it. In situ hybridization (ISH) and immunohistochemistry was used to validate candidate genes. RESULTS: Unsupervised classification of the 57 adenocarcinomas revealed two clusters of tumors with distinct global microRNA expression. One cluster contained all non-PNI tumors and a subgroup of PNI tumors. Significance analysis of microarray data yielded a list of microRNAs associated with PNI. At a false discovery rate (FDR)<10%, 19 microRNAs were higher expressed in PNI tumors than in non-PNI tumors. The most differently expressed microRNA was miR-224. ISH showed that this microRNA is expressed by perineural cancer cells. The analysis of protein-coding genes identified 34 transcripts that were differently expressed by PNI status (FDR<10%). These transcripts were down-regulated in PNI tumors. Many of those encoded metallothioneins and proteins with mitochondrial localization and involvement in cell metabolism. Consistent with the microarray data, perineural cancer cells tended to have lower metallothionein expression by immunohistochemistry than nonperineural cancer cells. CONCLUSIONS: Although preliminary, our findings suggest that alterations in microRNA expression, mitochondrial function, and cell metabolism occur at the transition from a noninvasive prostate tumor to a tumor with PNI.
Asunto(s)
Adenocarcinoma/genética , Adenocarcinoma/patología , Regulación Neoplásica de la Expresión Génica , MicroARNs/fisiología , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Diferenciación Celular/fisiología , Linaje de la Célula/fisiología , Proteína de la Membrana Similar al Receptor de Coxsackie y Adenovirus , Humanos , Masculino , Metalotioneína/genética , Mitocondrias/fisiología , Invasividad Neoplásica , Proteínas de Neoplasias/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Nervios Periféricos/patología , Receptores Virales/genéticaRESUMEN
MicroRNAs are small noncoding RNAs that regulate the expression of protein-coding genes. To evaluate the involvement of microRNAs in prostate cancer, we determined genome-wide expression of microRNAs and mRNAs in 60 primary prostate tumors and 16 nontumor prostate tissues. The mRNA analysis revealed that key components of microRNA processing and several microRNA host genes, e.g., MCM7 and C9orf5, were significantly up-regulated in prostate tumors. Consistent with these findings, tumors expressed the miR-106b-25 cluster, which maps to intron 13 of MCM7, and miR-32, which maps to intron 14 of C9orf5, at significantly higher levels than nontumor prostate. The expression levels of other microRNAs, including a number of miR-106b-25 cluster homologues, were also altered in prostate tumors. Additional differences in microRNA abundance were found between organ-confined tumors and those with extraprostatic disease extension. Lastly, we found evidence that some microRNAs are androgen-regulated and that tumor microRNAs influence transcript abundance of protein-coding target genes in the cancerous prostate. In cell culture, E2F1 and p21/WAF1 were identified as targets of miR-106b, Bim of miR-32, and exportin-6 and protein tyrosine kinase 9 of miR-1. In summary, microRNA expression becomes altered with the development and progression of prostate cancer. Some of these microRNAs regulate the expression of cancer-related genes in prostate cancer cells.