MP61-09 PROMOTER-TARGETED DOUBLE-STRAND RNA DUPLEX ENHANCES DPYSL3 GENE EXPRESSION IN PROSTATE CANCER CELLS

Abstract

You have accessJournal of UrologyProstate Cancer: Basic Research IV1 Apr 2015MP61-09 PROMOTER-TARGETED DOUBLE-STRAND RNA DUPLEX ENHANCES DPYSL3 GENE EXPRESSION IN PROSTATE CANCER CELLS William Parker, Qingting Hu, Jiang Wencong, J. Brantley Thrasher, and Benyi Li William ParkerWilliam Parker More articles by this author , Qingting HuQingting Hu More articles by this author , Jiang WencongJiang Wencong More articles by this author , J. Brantley ThrasherJ. Brantley Thrasher More articles by this author , and Benyi LiBenyi Li More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.2190AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES CRMP4, which is derived from the gene DPYSL3, has been recognized as a tumor metastasis suppressor gene in prostate cancer. We sought to characterize DPYSL3 gene expression profiles in prostate cancer cell lines and human prostate cancer specimens and then to develop a gene therapy for DPYSL3 enhancement utilizing a double-strand small activating RNA-based approach (saRNA). METHODS Cellular gene expression profiles were assessed using SYBR Green-based PCR and correlated to protein expression of CRMP4 using Western blot in 2 benign and 6 malignant prostate cell lines as well as in thirty-one de-identified pathology specimens containing prostate cancer and individually matched benign prostate tissues from patients who underwent radical prostatectomy. Genomic sequences and promoter locations of DPYSL3 gene were extracted from UCSD Genome Browser database. Small double-strand RNA duplexes (21nt) were chemically synthesized and were transiently transfected into prostate cancer cells with these small RNA molecules using RNAiMAX reagent. RESULTS Two isoforms of DPYSL3 gene exist in the human genome. Expression profiles showed that isoform-1 is the dominant isoform expressed in DU145 and LAPC-4 cells and isoform-2 is the dominant form in LNCaP, C4-2 and PC-3 cells. Both isoforms were similarly expressed in prostate cancer 22RV1 and benign prostate epithelial BPH1 cell, and are expressed at an extremely low level in benign RWPE-1 cells. The mRNA levels correlated with protein levels as evaluated by Western blot. Analysis of patient specimens revealed that isoform-2 expression levels are 2-fold higher than isoform-1 in both cancer and matched benign tissues. However, isoform-1 expression levels within the malignant tissue are significantly lower (2-fold, p = 0.01313, paired t-Test) than that in matched benign tissue. The saRNAs targeting the isoform-1 promoter but not the isoform-2 promoter significantly enhanced DPYSL3 gene expression in the mRNA and protein levels at day 3 after transfection in 22RV1, DU145 and PC-3 cells. CONCLUSIONS Our data reveal that isoform-1 of tumor suppressor gene DPYSL3 expression is significantly reduced in malignant tissues and is responsive to up-regulation by a saRNA-based approach. Further animal based in vivo testing of the saRNAs is being undertaking by our group to evaluate their clinical significance as a novel gene therapy strategy. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e749-e750 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information William Parker More articles by this author Qingting Hu More articles by this author Jiang Wencong More articles by this author J. Brantley Thrasher More articles by this author Benyi Li More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...