MICRO-RNA MEDIATED DIFFERENTIAL EXPRESSION OF ANNEXIN A2 IN PROSTATE CANCER

Purpose: Prostate cancer (PCa)develops through many defined stages: prostatic intraepithelial neoplasia (PIN), prostate cancer in situ, hormone-dependent and independent metastatic cancer. Annexin A2 (AnxA2), has been implicated in many cancer associated functions like plasminogen activation, actin-cytoskeletal rearrangement, cellular migration, adhesion and proliferation. Interestingly, AnxA2 is lost during PIN and early prostate cancer and reappears in metastatic prostate cancer, leading to worse clinical outcome. However the molecular mechanisms behind this differential regulation of AnxA2 are not yet understood. Our objective for this study is to identify the mechanism behind this differential regulation of AnxA2 in prostate cancer. Methods: As a proof of concept, miRNA maturation enzymes Drosha and Dicer were down regulated in hormone dependent PCa cells (LNCaP) by using siRNA, and AnxA2 mRNA expression was analyzed by PCR. Identification of the specific miRNAs that regulates AnxA2 expression in PCa was carried out by comparing the miRNA array profiling of normal and prostate cancer cell lines as well as in silico analysis. Identification of specific miRNA's was further confirmed by western blot analyses of the LNCaP and PC3 (metastatic prostate cancer), transfected with specific inhibitor of the miRNA and its mimic respectively. Migration and invasion assays were performed to study the effect of the specific miRNA on the prostate cancer cells. Results: Although LNCaP cells are null for AnxA2 expression, massive induction of AnxA2 mRNA expression by Drosha and Dicer knockdown confirmed the miRNA mediated regulation of AnxA2. The microarray study revealed three specific miRNAs. Out of these three miRNAs, miR-XYZ was first analyzed because of the high complimentary of its seed sequence to the coding region of the AnxA2 mRNA. Overexpression of AnxA2 in LNCaP and downregulation of AnxA2 PC3 cell lines upon treatment with miR-XYZ inhibitor and its mimic respectively were confirmed the Western blot. Transfection of miR-XYZ in PC3 cells resulted in a marked decrease in the migration and invasion of these cells as compared to control. Conclusions: These studies revealed that differential expression of AnxA2 is regulated by specific miRNA(s). Our results demonstrate that invasion and migration of PCa cells is significantly altered by modulating the expression of miR-XYZ. Thus miR-XYZ can be used as a potential diagnostic marker and as an adjuvant therapy in metastatic prostate cancer in the future.