CSIG-16. INTRONIC miR-744 INHIBITS GLIOBLASTOMA INVASION THROUGH INHIBITION OF MAPK-, SMAD- AND BETA-CATENIN SIGNALING. (5th November 2018)

• Record Type: Journal Article
• Authors: Hübner, Max; Hinske, Christian; Effinger, David; Kreth, Friedrich-Wilhelm; Kreth, Simone
• Is Part Of: Neuro-oncology. Volume 20(2018)Supplement 6
• Page Start: vi46
• Page End: vi46
• Publication Date: 2018-11-05
• Subjects: Brain Neoplasms -- Periodicals; Brain -- Tumors -- Periodicals; Brain -- Cancer -- Periodicals; Nervous system -- Cancer -- Periodicals; 616.99481
• DOI: 10.1093/neuonc/noy148.182
• Languages: English
• ISSNs: 1522-8517

Abstract

Abstract: INTRODUCTION: There is accumulating evidence that intronic micro-RNAs (miRNAs) are capable of either supporting or restraining functional pathways of their host genes thereby creating intricate regulative networks. We hypothesized that such type of interaction might gain special importance in human glioblastoma (GBM): The second intron of Mitogen-Activated Protein Kinase Kinase 4 (MAP2K4), an important hub in the pro-invasive MAPK pathway, harbors miR-744. We hypothesized that miR-744 regulates GBM migration by interacting with its host′s pathways. METHODS: MiR-744 was stably overexpressed in U87 cells. U87 invasion was studied using migration and Boyden chamber assays. TGFB1, MAP2K4 and DVL2 levels were measured by quantitative Real-Time-PCR (qRT-PCR) and SDS-PAGE. Interactions of miR-744 and 3`UTRs were analysed by luciferase reporter assays, SMAD2/3, p38 and beta-Catenin activity by SDS-PAGE, and TOP/FOPflash reporter gene assays. Stereotactically obtained tumor specimens were analysed by qRT-PCR. Experiments were performed in triplicate at least 3 times. Results are presented as mean±SEM. P-Values were calculated using Student's t-test. RESULTS: MiR-744 overexpression inhibited U87 invasion in migration- and Boyden chamber assays (-46%±5, 8%, n=4; p=0.026), qRT-PCR and SDS-PAGE revealed reduced levels of TGFB1, DVL2 and MAP2K4 (mRNA:-38%±5.6%, -33.6%±4.9% and -66.2%±7.9%; protein:-35.6%±8.3%, -36.8%±5.3% and -56.2%±9.6%; n=5, p0.05). TGFB1 knockdown repressed MAP2K4 mRNA- and protein levels (mRNA:-66.2% ±7.9%, protein:-56.2% ±9.6%, n=5, p<0.05). MiR-744 levels were dramatically decreased in glioblastoma samples (-90.3%±14.7%, n=21, p<0.05). DVL2 and TGFB1 were significantly induced (DVL2:3.35-fold±0.14; n=17; p<0.001; TGFB1:2.19-fold±0.09, n=29, p=0.015). CONCLUSION: These results provide evidence that miR-744 acts as an intrinsic brake on its host: It impedes MAP2K4 functional pathways through simultaneously targeting SMAD, beta-Catenin and MAPK signaling networks, thereby strongly mitigating pro-migratory effects of MAP2K4. Reexpression of miR-744 could be a promising approach to attenuate GBM invasion.