| Improved therapeutic approaches are needed for glioblastoma (GBM), the most common primary adult brain cancer. Despite maximal care, GBM has a dismal prognosis and the median survival is less than two years after diagnosis. Novel microRNA(miRNA)-based gene-specific targeting strategies can potentially improve the current poor median survival of GBM patients. We identified and tested the underexpressed miRNA-100 (miR-100) as a candidate `tumor suppressor' in multiple GBMs. We report that miR-100 expression reduces GBM tumorigenicity. In vitro, four GBM lines (two patient derived: 22T, and 33T; and two commercials: U87, U251,) demonstrated reduced proliferation when transiently overexpressed with the miR-100 precursor (pre-miR-00). pre-miR-100 triggered cell death on average 70% more than scrambled microRNA controls within 24hrs (p < 0.01; n = 3). miR-100-5p (Mature miRNA) targeted inhibition of the silencing mediator of retinoid or thyroid hormone receptor-2 (SMRT/NCOR2) gene expression was confirmed via reporter assays. In addition, we documented pre-miR-100 inhibitory consequence on multiple GBM tumor initiating cells (TICs) and the related pathways. The precursor of miR-100 targets SMARCA5, chromatin remodeling gene of SWI/SNF family, in TICs and diminishes STAT3 signaling. Furthermore, pre-miR-100 expression participated in declining ErbB3 (Her3) protein level and its downstream pathways (AKT and ERK). The pre-miR-100 enforces GBM TICs differentiation by lessening the amount of stem cell markers, such as Nestin and L1CAM, on average 50% compared to control miR. In vivo, Ki-67 proliferation index was decreased 40% in tumor xenografts generated from stable pre-miR-100 transfected GBM lines versus controls (p < 0.01; n = 8). Furthermore, the xenograft showed a minimum of two fold inhibitions to ErbB2 (Her2) and ErbB3 (Her3) expressions compared to the control tissue. Additionally, treatment of tumor xenografts with a single pre-mir-100 injection (60 pmol) significantly extended survival of mice bearing intracranial GBM xenografts 25% more than scrambled controls (p < 0.01; n=8). These studies establish the tumor suppressor activity of pre-miR-100 in GBMs and TICs. This suggests strong clinical potential for microRNA replacement therapy in GBM cases. |
