Abstract 3553: Targeting miR-361-3p suppresses the growth of human oral squamous cell carcinoma cells in vitro & in vivo

Abstract

Abstract MicroRNAs (miRNAs) are small non-coding RNAs with size of 20-25 nucleotides, and inhibit protein translation by binding the 3’-untranslated region (3’-UTR) of target mRNA. Each miRNA can regulate multiple mRNAs and each mRNA can be targeted by a number of miRNAs. In cancer, miRNAs can act as not only tumor suppressor genes but also oncogenes. Addiction to oncogenic miRNAs (OncomiRs) may provide therapeutic opportunities in human cancers such as oncogene addiction. In this study, we have attempted to identify OncomiRs in human oral squamous cell carcinoma (OSCC) cells and considered whether targeting miRNAs can be possible for cancer therapy. Functional screening for OncomiRs in human OSCC cells was performed with the use of miRCURY LNATM microRNA Knockdown Library-Human v12.0 (EXIQON). We transfected 918 locked nucleic acid (LNA)/DNA antisense oligonucleotides for specific human mature miRNAs into human OSCC cells (GFP-SAS). After 80 hours, each cell growth was evaluated by WST-8 assay. We identified LNA/DNA antisense oligonucleotide against miR-361-3p (LNA-miR-361-3p) which showed remarkable growth inhibition in GFP-SAS cells. Subsequently, we confirmed the target specificity of LNA-miR-361-3p by quantitative RT-PCR (qRT-PCR). LNA-miR-361-3p significantly reduced the expression of miR-361-3p. Co-transfection of synthetic mature miR-361-3p abrogated the growth inhibitory effect of LNA-miR-361-3p in GFP-SAS cells. Furthermore, transfection of synthetic mature miR-361-3p resulted in 20% increase of cell growth in GFP-SAS cells. Next, we identified odd-skipped related 2 (OSR2) as a target gene of miR-361-3p by the use of microarray, GeneSpring GX, and Ingenuity Pathway Analysis (IPA). The expression of OSR2 after transfection with LNA-miR-361-3p was significantly induced, and co-transfection of the OSR2 3’-UTR luciferase reporter plasmid and LNA-miR-361-3p into GFP-SAS cells produced higher luciferase activity than cells in co-transfected with LNA-non target (LNA-NT). Finally, we assessed the effect of LNA-miR-361-3p on the in vivo growth of GFP-SAS cells. We administered LNA-miR-361-3p into the xenograft tumors every 3 days. We found that LNA-miR-361-3p significantly reduced the size of subcutaneously xenografted GFP-SAS tumors, compared to the control group treated with LNA-NT. The expression of miR-361-3p in excised tumors was examined by qRT-PCR. LNA-miR-361-3p suppressed the expression levels of miR-361-3p compared with the control tumors. These results suggest that miR-361-3p support the growth of human OSCC cells and that targeting miR-361-3p may be a useful therapeutic approach for patients with OSCC. Citation Format: Norihiko Tokuzen, Koh-ichi Nakashiro, Himiko Ogawa, Hiroyuki Goda. Targeting miR-361-3p suppresses the growth of human oral squamous cell carcinoma cells in vitro & in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3553.