Abstract 520: The role of the calpain pathway in early response and acquired resistance to radiation in glioblastoma

Abstract

Abstract Glioblastoma (GBM) is the most aggressive, malignant brain tumor that is highly heterogeneous and often develops resistance to conventional chemotherapy and radiation treatments. The median survival time of GBM patients is 12-15 months despite aggressive treatment modalities, and there is an urgent need for the development of novel treatment strategies. The goal of this study is to identify novel mechanisms of radioresistance in GBM through mass spectrometry-based phosphoproteomic profiling, with the objective that more effective therapies can be rationally developed in the future. We and others have previously reported that GBMs are characterized by deregulated signaling pathways that control critical cellular processes including survival, migration, and proliferation. Very little is known about acquired resistance mechanisms to radiation, specifically how radiation alters protein phosphorylation patterns in tumors on a global level, or how radiation-induced changes in protein phosphorylation contribute to treatment resistance. In order to identify radiation-induced changes in protein phosphorylation, we performed phosphoproteomic profiling of 6 GBM cell lines and 1 normal human astrocyte cell line both before and after radiation treatment in 3 independent experiments. Specifically, cells were collected before treatment and at 2 time-points following a 10 Gy radiation treatment. Following protein isolation and phosphopeptide enrichment, global phosphoproteomic analysis was performed by mass spectrometry. Relative quantitation and statistical analysis were performed to determine fold changes in phosphorylation at both post-radiation time-points. An average of 3,971 phosphopeptides mapping to 1,649 phosphoproteins were quantified for each independent experiment, and a total of 621 phosphoproteins were quantified across all 3 datasets. We identified a major component of the calpain signaling pathway that consistently shows an increase in phosphorylation following radiation treatment in GBM cell lines. The post-radiation increase in phosphorylation was confirmed in 2 independent experiments and observed across all 6 GBM cell lines. This is the first study linking the calpain signaling pathway to radiation response in GBM. Mechanistic studies using phosphomimetic and nonphosphorylatable mutants of the protein of interest will determine the effect of phosphorylation on calpain activity and radiation resistance. Results of this work will contribute to our understanding of how GBM tumors become resistant to radiation therapy, and may identify a novel target to overcome radiation resistance in GBM. Citation Format: Emily A. Bassett, Arnab Chakravarti. The role of the calpain pathway in early response and acquired resistance to radiation in glioblastoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 520.