Abstract 3090: miR-24 influences proliferation of prostate cancer cells <i>in vitro</i> via targeting ZNF217.
Authors: Elke Nolte, Jaroslaw Szczyrba, Martin Hart, Celina Döll, Sven Wach, Helge Taubert, Bastian Keck, Elisabeth Kremmer, Robert Stöhr, Arndt Hartmann, Bernd Wullich, Friedrich Grässer
Journal: Cancer Research (2013)

Abstract Background: Prostate cancer is one of the leading causes of tumor-related death in men. To identify the underlying mechanisms, microRNA (miRNA) expression profiles of primary prostate cancer samples and of adjacent non-tumor tissue were analyzed. Using database analysis, we sought to identify regulatory targets of aberrantly expressed miRNA miR-24. Material and Methods: We performed comparative miRNA expression profiling using deep sequencing of cDNA libraries. Potential target genes of deregulated miRNAs were identified by database analysis and subsequently validated in vitro using reporter gene constructs. miRNA and mRNA expression was determined using quantitative real-time PCR. Results: One of the miRNAs consistently deregulated in primary prostate cancer samples was miR-24 which was found downregulated in tumor samples more than 1.5-fold in the deep sequencing approach. Independent target prediction databases (including miRanda, Targetscan and PicTar) indicated that the 3’ untranslated region (3’UTR) of the zinc finger protein 217 (ZNF217) gene is a regulative target for miR-24. This prediction could be confirmed using a luciferase reporter gene construct containing the ZNF217 3’UTR. Co-expression of the reporter gene construct with miR-24 resulted in a reduction of reporter gene activity while mutation of the miRNA binding sites caused a loss of responsiveness to the respective miRNA. Furthermore we could show that transient expression of miR-24 reduced the amount of ZNF217 protein in DU145 and LNCaP prostate cancer cells. miR-24 expression was significantly reduced in tumor tissue compared to matched non-tumor tissue (p&amp;lt;0.0001, N=26, Wilcoxon-signed-rank-test) while the expression of ZNF217 mRNA was significantly higher in tumor tissue compared to matched non-tumor tissue (p=0.033, N=26, Wilcoxon-signed-rank-test). In Western blot analyses 61% (14/23) of primary tissue samples also displayed an increased amount of ZNF217 protein. A transient knock-down of ZNF217 in DU145 and LNCaP cells applying siRNA technique led to reduced cell proliferation over a time period of 72 hours. Conclusion: Our findings demonstrate that the 3’-UTR of ZNF217 is one regulative target of miR-24, which is commonly deregulated in prostate cancer. ZNF217 mRNA and protein is elevated in the majority of tumor samples analyzed. A transient knock-down of ZNF217 leads to a reduced proliferation in prostate cancer cell lines. Citation Format: Elke Nolte, Jaroslaw Szczyrba, Martin Hart, Celina Döll, Sven Wach, Helge Taubert, Bastian Keck, Elisabeth Kremmer, Robert Stöhr, Arndt Hartmann, Bernd Wullich, Friedrich Grässer. miR-24 influences proliferation of prostate cancer cells in vitro via targeting ZNF217. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3090. doi:10.1158/1538-7445.AM2013-3090