Epigenetic changes have been suggested to drive prostate cancer (PCa) development and progression. Therefore, in this study, we aimed to identify novel epigenetics-related genes in PCa tissues, and to examine their expression in metastatic PCa cell lines. We analyzed the expression of epigenetics-related genes via a clustering analysis based on gene function in moderately and poorly differentiated PCa glands compared to normal glands of the peripheral zone (prostate proper) from PCa patients using Whole Human Genome Oligo Microarrays. Our analysis identified 12 epigenetics-related genes with a more than 2-fold increase or decrease in expression and a p-value <0.01. In moderately differentiated tumors compared to normal glands of the peripheral zone, we found the genes, TDRD1, IGF2, DICER1, ADARB1, HILS1, GLMN and TRIM27, to be upregulated, whereas TNRC6A and DGCR8 were found to be downregulated. In poorly differentiated tumors, we found TDRD1, ADARB and RBM3 to be upregulated, whereas DGCR8, PIWIL2 and BC069781 were downregulated. Our analysis of the expression level for each gene in the metastatic androgen-sensitive VCaP and LNCaP, and -insensitive PC3 and DU-145 PCa cell lines revealed differences in expression among the cell lines which may reflect the different biological properties of each cell line, and the potential role of each gene at different metastatic sites. The novel epigenetics-related genes that we identified in primary PCa tissues may provide further insight into the role that epigenetic changes play in PCa. Moreover, some of the genes that we identified may play important roles in primary PCa and metastasis, in primary PCa only, or in metastasis only. Follow-up studies are required to investigate the functional role and the role that the expression of these genes play in the outcome and progression of PCa using tissue microarrays.

Abstract. Prostate cancer (PCA) is a clinically heterogeneous and often multifocal disease with a clinical outcome difficult to predict. A deeper knowledge of the molecular basis of the disease may lead to a better prediction of prognosis. Therefore, in this study we investigated the molecular basis of PCA by identifying potential tumor markers in laser-microdisected PCA tissues. Among a group of PCA patients, quantitative RT-PCR analysis was performed to compare the expression of 70 genes. These genes were selected from the results of two microarrays which investigated the gene expression profile differences between moderately or poorly differentiated prostate carcinoma glands and the corresponding normal glands. Among the genes examined, CDKN2A, GATA3, CREBBP, ITGA2, NBL1 and TGM4 were down-regulated in the prostate carcinoma glands compared to the corresponding normal glands, whereas TFF3, TMPRSS2 and ERG were up-regulated. Our findings indicate that these genes may play roles as tumor suppressor genes or oncogenes in PCA, and may serve as potential tumor markers and novel therapeutic targets.

Abstract. Prostate cancer (PCa) is one of the most prevalent forms of cancer affecting males worldwide, and knowledge of the immune defenses involved in PCa remains incomplete. Since the identification of immunity-related genes may have enormous implications for the understanding of PCa immunology, we recently reported the identification of immunity-related genes in PCa tissues and found potential binding sites for the ETS family prototype, ETS-1, in the majority of genes identified. Therefore, as a continuation of our previous study, we investigated whether ETS-1 regulates these genes in an in vitro PCa cell line model, PC3 cells. We specifically blocked ETS-1 in PC3 cells by transfection with an ETS-1 inverse antisense expression vector or a mock control vector. We then assessed the effect of the blockade on the expression of the recently identified PCa immunity-related genes using a comprehensive oligo gene expression microarray analysis. The results showed that ETS-1 is involved in the activation or repression of the recently identified immunity-related genes in PCa. These findings provide insights into the regulation of immunity-related genes in PCa, and emphasize the importance of ETS-1 in prostate cancer immunology.

Abstract. The role of the immune response in tumor progression, and disease outcome is still debated, and a lack of knowledge of the immune defenses in prostate cancer still exists. In addition, the ETS family of transcription factors which is involved in translocations frequently found in prostate cancer is reported to be essential for the regulation of immunity-related genes. In order to identify immunity-related genes in prostate cancer, we performed two microarrays using RNA extracted from laser microdissected glands of the normal prostate proper (or the peripheral zone) and moderately and poorly differentiated prostate carcinomas from patients who had undergone radical prostatectomy. Many differentially expressed genes were found, however, only immunity-related genes (B cell, innate, and T cell immunity) with an expression of more than 10-fold increase or decrease and a P<0.01 between the moderately differentiated tumors and the normal glands, and the poorly differentiated tumors and the normal glands were considered significant. Based on these two microarrays, we identified a set of 37 genes that were up-or down-regulated in tumors (moderately and poorly differentiated) compared to the normal glands. Analysis of these genes revealed, strikingly, that 31/37 of these genes have potential binding sites within their promoter regions for members of the ETS family of transcription factors, and some are reported to be targets of ETS members. These findings identified immunity-related genes in prostate cancer, and provided insights into their potential regulation, which may lead to a better early detection, immunotherapy, and therapeutic drug treatment of this disease. Unraveling the dynamics of the ETS-immunity-related genes will provide an invaluable insight into understanding prostate cancer immunology.

Abstract. The ETS family of transcription factors plays important roles in both normal and neoplastic cells for different biological processes such as proliferation, differentiation, development, transformation, apoptosis, migration, invasion and angiogenesis. The 27 ETS factors are probably a part of complex regulatory networks including interactions among family members. In human prostate cancer, rearrangements have been found in several genes of the ETS family resulting in chimeric oncoproteins. In a previous study we found that the ETS family prototype, Ets-1 affects biological properties of PC3 prostate cancer cells. In a first effort to understand the cooperative interactions between different ETS factors in prostate cancer, in the present study we examined the expression pattern of all 27 ETS members using quantitative RT-PcR (qRT-PcR) in the androgen-sensitive VcaP and LNcaP, and the androgen-insensitive Pc3 and dU-145 prostate cancer cell lines as well as in human prostate cancer tissue samples. We further investigated whether the ETS family prototype, Ets-1, regulates other ETS family members by examining the effect of Ets-1 blockade in Pc3 cells on their expression. We found an expression specificity of various ETS family members in the prostate cancer cell lines which might reflect their different biological properties. In human prostate samples only 3 among the 27 ETS family members (Ehf, Elk-4 and Ets-2) showed significant expression differences between normal and cancerous prostate glands. We finally demonstrate that the family prototype, Ets-1, regulates the family members Elf-1, Elf-2, Elk-1, Etv-5 and Spi-1 in Pc3 prostate cancer cells. chimeric oncoproteins containing ETS family members arising due to frequent translocations in prostate cancer are probably part of a regulatory network involving other ETS family members as well.