Abstract 2598: AZD1390 radio-sensitizes p53-mutant GBM via disrupting homology directed DNA repair
Authors: Gupta Shiv K. , Chen Jiajia , Laverty Daniel J. , Talele Surabhi , Carlson Brett L. , Tuma Ann C. Mladek , Burgenske Danielle , Kitange Gaspar J. , Hamerlik Petra , Nagel Zachary D. , Durant Stephen T. , Elmquist William F. , Sarkaria Jann N. 
Published: June 2022

Abstract
The ATM inhibitor AZD1390 disrupts cellular responses to ionizing radiation (IR) and is a potent radiosensitizer being tested in clinical trials. Here, we evaluated, the effects of AZD1390 on radiation sensitivity and DNA damage repair pathways in glioblastoma (GBM) cells and patient derived xenografts (PDXs). AZD1390 (30 nM and higher) suppressed IR (5 Gy)-induced phosphorylation of ATM-Serine1981 and downstream phosphorylation sites on Kap1, Chk2 and H2AX in U251 cells and multiple PDXs. Consistent with enhanced DNA damage, AZD1390 increased IR induced G2/M arrest in U251 (80.6% with AZD1390/IR vs. 64.6% with IR, p= 0.01), GBM43 (61.9% vs. 25.7%, p= 0.01) and GBM39 (40.9% vs. 25.4%, p= 0.01). Moreover, in a clonogenic survival assay, AZD1390 sensitized U251 cells to 5 Gy IR (0.24% survival with AZD1390/IR vs. 2.3% with IR alone, p=0.01). In a reporter-based analysis of DNA repair capacity, ATM inhibition resulted in a 40 to 60% reduction in homologous recombination (HR) and modest but significant decrease in micro-homology mediated end joining (MMEJ) and gap fill-in synthesis in U251 cells but had no effect on non-homologous end joining, translesion synthesis, nucleotide or base excision repair pathways. Comparing effects of AZD1390 on repair in GBM14 (TP53-wt) and GBM43 (TP53-mutant), similar results were observed except that decreased MMEJ was seen only in GBM43 (0.03± 0.01% vs. 0.07± 0.01% in control, p=0.002). Intriguingly, RAD51 knockdown (to disrupt HR) sensitized U251 and GBM43 but not GBM14. The efficacy of AZD1390 ± IR was studied in vivo in 10 PDXs. IR was delivered to orthotopic tumors using opposed lateral 225 kVp beams. AZD1390 (20 mg/kg PO) was given just prior to each radiation dose (2 Gy x 5 fractions). AZD1390 monotherapy was mostly ineffective, IR alone was reasonably efficacious with an average 1.8 ± 0.1-fold-increase in survival relative to sham radiation (survival ratio) across all 10 models. IR/AZD1390 treatment resulted in significant survival extension relative to IR alone in 6 of 10 models. Analysis of the survival benefit of combination therapy compared to IR alone across the entire cohort of PDXs was statistically marginal (average survival ratio 2.3± 0.3 vs. 1.8 ± 0.1 with IR, p=0.08). However, when stratified by TP53 status, combination therapy was significantly more effective than IR (mean survival ratio 2.3 ± 0.1 vs. 1.6 ± 0.2 with IR alone, p=0.02) in TP53-mutant PDXs, where all 5 models benefited. In contrast, TP53-wt group had no benefit (mean survival ratio 2.2 ± 0.5 vs. 2.0 ± 0.2, p=0.61), GBM39 was only TP53-wt PDX that benefited from the combination. In conclusion, AZD1390 is an effective radio-sensitizer that causes disruption in HR and potentially other DNA repair pathways. Interestingly, in vivo radiosensitizing effects are mostly restricted to TP53-mutant GBM PDXs. Understandingmechanism of resistance in the context of different TP53 backgrounds remains an important future direction.
Citation Format: Shiv K. Gupta, Jiajia Chen, Daniel J. Laverty, Surabhi Talele, Brett L. Carlson, Ann C. Mladek Tuma, Danielle Burgenske, Gaspar J. Kitange, Petra Hamerlik, Zachary D. Nagel, Stephen T. Durant, William F. Elmquist, Jann N. Sarkaria. AZD1390 radio-sensitizes p53-mutant GBM via disrupting homology directed DNA repair [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2598.