Abstract 1065: MicroRNA-1296 and genistein regulate DNA licensing proteins (MCM-genes) in prostate cancer

Abstract

Abstract Introduction and Objective: The minichromosome maintenance (MCM) gene-family is essential for DNA replication and is found to be frequently up-regulated in a variety of cancers. In this study we examined the role of MCM2 in prostate cancer and the effect of microRNA-1296, genistein and trichostatin A on the MCM-complex. Methods: Prostate cancer cell lines (LNCaP and PC3) were treated with different doses (0-50umol/L) of genistein (72 hours) and 100 ng/ml trichostatin-A (24 hours). Quantitative real-time PCR, immunohistochemical staining and Western analysis were performed to check mRNA and protein expression in cells and tissues. The small interfering RNA (siRNA) approach was employed to understand the functional significance of MCM2 gene. Fluorescence activated cell sorting (FACS) analysis was performed to monitor cell cycle distribution and apoptosis. PC3 cells were transfected with miR-1296 and miR-1296 inhibitor to investigate its effect on the MCM2 gene. Results: Profiling results showed that expression of MCM-genes was higher in prostate tumor sample and cell lines. Genistein and trichostatin A (TSA) significantly down-regulated expression of all MCM-genes. A specific siRNA was used to knock-down MCM2 mRNA and protein expression. FACS results showed that genistein, TSA and siRNA caused a significant decrease in the S-phase of the cell cycle. There was also down-regulation of CDT1, CDC7 and CDK2 genes which govern the loading of MCM-complex on chromatin. We found that microRNA-1296 targets the MCM2 mRNA sequence. MicroRNA-1296 expression is significantly down-regulated in prostate cancer compared to normal tissues. To investigate the relationship between MCM2 and miR-1296, a miR-1296 inhibitor was transfected into PC3 cells. Inhibition of miR-1296 up-regulated both MCM2 mRNA and protein, whereas over-expression caused a significant decrease in MCM2 mRNA, protein and the S phase of cell cycle. Conclusion: MCM-genes are excellent anti-cancer drug targets because they are essential DNA replication factors that are highly expressed in cancer cells. This is the first report showing an anti-MCM effect by miR-1296, genistein and TSA. TSA is undergoing clinical trials as a prostate cancer treatment but has high toxicity. Genistein, a natural non-toxic dietary isoflavone, may be an advantageous therapeutic agent for treating prostate cancer. The use of RNAi is currently being implemented as a gene-specific approach for molecular medicine. By the same principle, the specific down-regulation of oncogenes by micro-RNA may contribute to novel therapeutic approaches in the treatment of prostate cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1065.