Abstract
BACKGROUND
Glioblastoma is the most aggressive brain tumor with poor prognosis despite the best available treatment. MicroRNAs (miRNAs) are emerging as promising, novel prognostic biomarkers and therapeutic targets in glioblastoma. In a previous study, we demonstrated that miR-4516 predicts poor prognosis and functions as an oncogene in glioblastoma. Aim of the current study is to examine the role miR-4516 in radiation resistance and identify downstream targets contributing to this phenotype
METHODS
Radiosensitization was evaluated by cell viability and clonogenic assays. Cell apoptosis was evaluated using flow cytometry and immunoblotting. Potential targets of miR-4516 were identified using bioinformatic analysis (Targetscan and miRDB) and confirmed by luciferase reporter assays. Results were validated using immunoblotting. miR-4516 expression in glioblastoma cell lines after radiation treatment was quantified by qRT-PCR.
RESULTS
Expression of miR-4516 was increased up to 15 fold following radiation treatment, peaking at around 15min-60 min in primary and established glioblastoma cell lines including GBM 08-387, GBM 30 and U87-MG. Furthermore, inhibition of miR-4516 sensitized GBM 08-387, GBM30 and U87-MG cells to radiation in comparison to control groups as determined by cell viability and clonogenic assays. Further, miR-4516 inhibition induced apoptosis in these cell lines following radiation treatment. While conducting mechanistic studies, we found that the tumor-promoting function of miR-4516 was, in part, mediated by inhibition of p21 and PTPN14, two direct targets of miR-4516
CONCLUSION
Our data suggest that radiation induces the expression of miR-4516 in glioblastoma cell lines. This miRNA plays a critical role in radio-resistance and promotes aggressive phenotypes in glioblastoma and therefore, functional analyses of its target pathways may uncover novel therapeutically vulnerable target(s) in glioblastoma. FUNDING: R01CA108633, R01CA169368, RC2CA148190, U10CA180850-01(NCI), Brain Tumor Funders Collaborative Grant, and OSU-CCC (all to AC). The Ton and Patricia Bohnenn Fund for Neuro_Oncology Research (to PR).
Neutralization of Adrenomedullin Inhibits the Growth of Human Glioblastoma Cell Lines in Vitro and Suppresses Tumor Xenograft Growth in Vivo
