MicroRNA-21 is overexpressed in most cancers and has been implicated in tumorigenesis. Accumulating evidence supports a central role for the miR-21 guide strand (miR-21-5p) in ovarian cancer initiation, progression, and chemoresistance. However, there is limited information regarding the biological role of the miR-21 passenger strand (miR-21-3p) in ovarian cancer cells. The aim of this study was to investigate the role of miR-21-3p and its target genes in cisplatin-resistant ovarian cancer cells. Expression profiling of miR-21-5p and miR-21-3p was performed in a panel of cancer cells by qPCR. Colony formation and invasion assays were carried out on ovarian and prostate cancer cells transfected with miR-21-5p and miR-21-3p inhibitors. Dual luciferase reporter assays were used to identify the miR-21-3p target genes in ovarian cancer cells. Our results show that miR-21-5p had higher expression levels compared to miR-21-3p on a panel of cancer cells. Moreover, inhibition of miR-21-5p or miR-21-3p resulted in a significant decrease in ovarian and prostate cancer cell proliferation and invasion. Luciferase reporter assays identify RNA Binding Protein with Multiple Splicing (RBPMS), Regulator of Chromosome Condensation and POZ Domain Containing Protein 1 (RCBTB1), and Zinc Finger protein 608 (ZNF608) as miR-21-3p target genes. SiRNA-induced RBPMS silencing reduced the sensitivity of ovarian cancer cells to cisplatin treatment. Immunohistochemical analyses of serous ovarian cancer patient samples suggest a significant decrease of RBMPS levels when compared to normal ovarian epithelium. Taken together, the data generated in this study suggests a functional role for miR-21-3p in ovarian cancer and other solid tumors.

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Alternative splicing is an essential process for the generation of protein diversity. The physiological role, cellular localization, and abundance of splice variant products compared to the wild-type protein may be completely different. This is illustrated by the five splice variants of the antiapoptotic protein survivin that are more abundant in cancerous cells compared with normal tissues. Interestingly, some survivin splice variants have been associated with drug resistance. Herein, we describe a SYBR green I-based real-time PCR method to assess the messenger RNA levels of the human survivin splice variants in taxane-sensitive versus taxane-resistant ovarian cancer cells and in human ovarian cancer samples. Furthermore, in this chapter, we describe the quantification of survivin splice variants by real-time quantitative PCR (qPCR) after in vitro and in vivo small interference RNA (siRNA)-mediated silencing of survivin splice variants.

MicroRNAs (miRNAs) are 21-25 nucleotides regulatory involved in silencing mRNAs in a sequence specific manner. Accumulated data has established the central role of various microRNAs in tumor initiation and progression. Furthermore, recent studies revealed that microRNAs participate in the resistance of cancer cells to different chemotherapeutic agents. In ovarian carcinoma, the fifth cause of cancer deaths in US, it has been shown that several miRNAs contribute to drug resistance. However, the role of these miRNAs in the cisplatin resistance of ovarian cancer cells is unclear. In a microarray study, we found that miR-21, miR27b and miR-622 were upregulated in the A2780CP20 cisplatin resistant ovarian cancer cells compared with the A2780PAR cells, which are sensitive to cisplatin treatment. Real-time PCR studies confirmed these findings. Opposite, the c-Jun transcription factor was upregulated in this microarray analysis. Then, we used TargetScan and miRBase tools to identify miR-21, miR-27b and miR-622- target genes. Interestingly, several of these miRNA-target genes were identified as downregulated in our microarray study. These results were corroborated by Western blot analysis. Moreover, cell survival experiments in the presence of a miR-21 mimic, made A2780PAR cells more resistant to cisplatin treatment. These results indicate that dysregulation of various miRNAs and their target genes contribute to the cisplatin resistance of ovarian cancer cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1105. doi:1538-7445.AM2012-1105