ATPS-01INHIBITION OF T-TYPE CALCIUM CHANNELS SENSITIZES GLIOBLASTOMA STEM CELLS TO CHEMOTHERAPY

Glioblastoma stem cells (GSC) have been implicated in tumor resistance to radio- and chemotherapy. T type calcium channels (Cav3.2) regulate cell cycle progression by mediating the necessary influx of calcium for transit past the G1/S cell cycle checkpoint. We hypothesized that treating GSCs and GSC-derived xenografts with the FDA-approved Cav3.2 channel blocker mibefradil would synchronize GSCs to enter the S phase, and consequently sensitize them to cytotoxic therapies. We demonstrated that Cav3.2 is highly expressed in the majority of human GBM specimens and all GCSs, compared to normal brain tissue or glioma cell lines, respectively. Mibefradil treatment inhibited GSC proliferation and induced cell death. Furthermore, mibefradil sensitized GSCs to temozolomide treatment (TMZ) and increased TMZ-induced cell death by 25-63% (p < 0.05) in vitro. To determine the effect of mibefradil on glioblastoma xenograft growth, we implanted GSCs in the brains of immunodeficient mice and treated the mice with mibefradil and/or TMZ and monitored tumor growth by MRI. We found that mibefradil increased TMZ-induced tumor growth inhibition by 60% (p < 0.05). Mibefradil also significantly improved the survival of TMZ-treated mice bearing GSC-derived xenografts. To further investigate the mechanism of action of mibefradil, we performed reverse phase protein arrays on GSCs treated with mibefradil. We found that mibefradil strongly regulated GSC apoptosis by regulating BCL2, PUMA and BAX expressions/activations as well as caspase cleavage. Mibefradil also altered proteins involved in autophagy and invasion including LC3, FAK and other. We are currently performing RNA-seq to determine the transcriptome wide changes that are induced by Cav3.2 inhibition. Altogether, the data provide mechanistic and functional rationales for the use of Cav3.2 inhibitors such as mibefradil as a new adjuvant therapy that enhances the efficacy of cytotoxic therapies in glioblastoma by targeting glioblastoma stem cells.