ATPS-48A HIGH-THROUGHPUT SCREEN IDENTIFIES microRNA-1300 AS A POTENTIAL THERAPEUTIC microRNA CAUSING CYTOKINESIS FAILURE AND APOPTOSIS IN GLIOBLASTOMA CELLS

Abstract

INTRODUCTION: There is a pressing need for new therapeutic approaches for the treatment of glioblastoma (GBM). MicroRNAs are single-stranded 22-24nt non-coding RNAs which function by reducing translation or causing degradation of target mRNAs. MiR-1300 was identified in a high-throughput miRNA mimic library screen as a promising therapeutic candidate, inducing cell death. METHOD: To validate the cytotoxic effect of miR-1300, a panel of four established cell lines (U251,LN229,U373,KNS42)and two patient-derived stem-like cell lines were utilised for RTqPCR, imaging and flow-cytometry to assess endogenous expression levels, cellular morphology, apoptosis, and cell cycle delay respectively. Bioformatic approaches identified potential miR-1300 targets as mediators of this effect and validation of this involvement in the cytotoxic phenotype using siRNA knock-down is being investigated. Validation of the direct targeting by miR-1300 is being performed using 3' UTR Luciferase assays. RESULTS: Expression, of miR-1300, by transfection of oligonucleotide mimics, in our panel of GBM cells consistently led to G2M arrest followed by apoptosis within 72h. The associated phenotype is binucleation, indicative of cytokinesis failure.Preliminary data suggest this may be mediated through ECT2, a predicted target of miR-1300 known to be involved in formation of the cleavage furrow. CONCLUSION: We have identified a highly potent pro-apoptotic microRNA that may be exploitable for GBM therapy. Importantly, there is no detectable endogenous miR-1300 expression in any GBM samples or cell lines. On-going work will confirm the targets of miR-1300 that drive this effect as well as exploring potential approaches to deliver this as a biotherapeutic agent.