Background: Glioblastoma Multiforme (GBM-Grade IV) is the most aggressive form of glioma in the USA carrying only a 15-month median survival time. IDH1R132H is an early onset mutation present in 70-80% of low grade gliomas (LGG-Grade I and II) and secondary GBM patients that causes intracellular oxidative stress and impairment of post-radiation DNA damage repair (DDR) via Homologous Recombination (HR). Although IDH1R132H is associated with improved patient survival and clinical response to chemotherapy, astrocytoma (LGG) patients specifically also harbor ATRXLoss, a genetic alteration that inactivates the ATRX chromatin remodeling protein and impairs DDR via non-homologous end joining (NHEJ). Importantly, the recent discovery of glioma stem cells (GSCs), a subpopulation of chemoradioresistant, self-renewable, tumor initiating cells known to promote tumor recurrence, presents a new target for therapeutic intervention. However, the mechanisms by which IDH1R132H and ATRXLoss contribute to GSC sensitivity to proton radiotherapy (PRT), a more tumor specific and cytotoxic form of radiation therapy than conventional X-rays (XRT), is not well understood. We hypothesize that both IDH1R132H and ATRXLoss contribute to the impairment of GSC self-renewal after PRT, but not after XRT. Methods: Isogenic MGG18-IDH1WT, MGG18-IDH1R132H, TS543-ATRXWT, and TS543-ATRXLoss patient derived glioma cells were cultured as neurospheres and treated with 3-6 Gy of XRT or PRT. GSC self-renewal, based on neurosphere formation frequency, was quantified using extreme limiting dilution analysis (ELDA). Results: TS543-ATRXLoss significantly inhibited GSC self-renewal over TS543-ATRXWT when subject to PRT (Pairwise Testing Pr(>χ2)<0.01). However, TS543-ATRXWT displayed lower GSC self-renewal compared to TS543-ATRXLoss when subject to XRT (Pr(>χ2)<0.0001). Interestingly, similar results were found in the isogenic MGG18 cell lines. MGG18-IDH1R132H displayed higher radiosensitivity to PRT (Pr(>χ2)<0.05) while MGG18-IDH1WT displayed higher radiosensitivity to XRT (Pr(>χ2)<0.01), as defined by GSC self-renewal. Conclusions: We conclude that ATRXLoss and IDH1R132H both substantially contribute to PRT sensitivity in GSCs, thereby supporting the benefits of PRT for astrocytoma patients. Our future experiments will further elucidate the biological mechanisms by which ATRXLoss and IDH1R132H confer PRT sensitivity, especially via dsDNA damage and apoptosis.

Citation Format: Angel A. Garces, Lawrence Bronk, Krishna P.L. Bhat, David R. Grosshans. ATRXLoss and IDH1R132H impair self-renewal of glioma stem cells after proton radiotherapy [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-065.