Abstract B54: miR-125b and miR-17-3p: Newly identified tumor suppressors for prostate cancer

Abstract

Abstract MicroRNAs (miRs) are small molecules that post-transcriptionally regulate expression of target mRNAs and are thought to play a major role in prostate cancer (CaP) tumorigenesis. We compared miR expression in a progressive prostate cancer cell model consisting of the immortalized p69 human prostate cell line compared to M12, a highly tumorigenic, metastatic subline, and F6, a weakly tumorigenic variant using Exiqon's miRCURY LNA™ miR human panels. Most notable was a decrease in expression of miR-125b and miR-17-3p, which was confirmed by single miR analysis via RT-qPCR. Previously, miR array screens have been done on RNA extracted from a variety of whole tumors. However, due to the heterogeneous nature of prostate tumors, it is impossible to determine which cell-type is contributing to differential miR expression. To address this concern, we have developed laser capture microdissection (LCM) to validate miR expression in RNA extracted from the individual cell-types (normal stroma, normal epithelial, PIN, and tumor cell of increasing Gleason score) that comprise human prostate tumors. This analysis validated that the expression of miR-17-3p and miR-125b is reduced in the prostate tumor cell compared to normal epithelium. Previously, we have shown that miR-17-3p targets vimentin and IGF1R. Restoration of miR-17-3p expression in the M12 cell line resulted in decreased cell motility and invasion in transwell (Boyden) chamber assays as well as decreased growth of subcutaneous and orthotopic tumors in male, athymic nude mice confirming miR-17-3p is behaving as a tumor suppressor. Previously, data from miRNA array screens coupled to gene arrays have been used to predict mRNA targets. This approach assumes that all miRNAs regulate gene expression by degrading their respective targets, which is not always the case. Since the end result of miRNA targeting is reduced protein synthesis, we propose that proteomics using RPMA, Reverse Phase Protein MicroArrays, is a better approach for deducing targets of miR regulation. Proteomic analysis of these same cell lines confirmed a correlative increase in pErB2/ErB3 and vimentin protein levels in cell lines where miR-125b and miR-17-3p expression are low. MiR-125b is proposed to target ErbB2/ErbB3. Interestingly, a network analysis of oncogenic miRNA targets in the prostate revealed that the EGFR family of which ErB2/ErB3 is thought to be a major driver of CaP was one of the most highly connected proteins. Currently, we are using a variety of in vitro and in vivo experiments to prove that miR-125b can function as a tumor suppressor as we have shown for miR-17-3p. Altogether this multi-factorial approach of miRNA array analysis coupled to proteomics, validated by LCM analysis of human tumor samples and additional experiments has identified two new tumor suppressors, miR-17-3p and miR-125b, which could be relevant biomarkers for CaP. Restoration of miR-17-3p and/or miR-125 expression could provide new targets/treatments to block prostate cancer progression. Citation Format: William T. Budd, Danielle E. Weaver, Valerie Calvert, Emanuel F. Petricoin, III, Joy L. Ware, Zendra E. Zehner. miR-125b and miR-17-3p: Newly identified tumor suppressors for prostate cancer [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr B54.