Abstract 1421: Evaluation of DDR-targeting strategy using ATR inhibitor in biliary tract cancer

Abstract

Abstract Background: The DNA damage response (DDR) is a multicomplex network of signaling pathways involved in DNA damage repair, cell cycle checkpoints and apoptosis. The ataxia telangiectasia and Rad3-related (ATR) protein kinase is a key enzyme in the DDR that activates checkpoint kinase 1 (Chk1), resulting in cell cycle arrest. Tumor types with loss of ATM function and/or high replication stress are expected to be more susceptible to DDR targeting. In biliary tract cancer (BTC), DNA repair pathway, which includes BAP1, MSH6, BRCA1, ATM, MLH1, MSH2, is altered in about 20 % of cases. TP53 module is observed in 33.9% of BTC cases (Nat Genet 2015). The purpose of this study is to test DDR targeting strategy using ATR inhibitor in biliary tract cancer. Methods: Using 9 kinds of BTC cells, MTT assay and colony formation assay were done for determining growth inhibitory effect of AZD6738, an ATR inhibitor. Cell cycle analysis was done by FACS Calibur flow cytometer and the methods described by Chou and Talalay were used to determine whether a synergistic effect existed between AZD6738 and cytotoxic chemotherapeutic agents (cisplatin, 5-FU, gemcitabine). The alkaline comet assay was done to measure of DNA damage in individual cells. Tumor xenografts model was used for in vivo test of AZD6738. Results: Among 9 BTC cells, SNU478 and SNU869 were most sensitive to AZD6738, which showed low expression of both ATM and p53. AZD6738 blocked ATR-mediated Chk1 phosphorylation and increased rH2AX, a marker of DNA damage, in sensitive cells. AZD6738 significantly increased apoptosis (cleavage of PARP and caspase-7) and G2/M arrest, increased level of p21, and decreased cdc2. In addition, combination of AZD6738 and cytotoxic chemotherapeutic agents demonstrated synergistic effects in colony formation assay, cell cycle analysis and comet assay. In xenograft model of SNU478, AZD6738 monotherapy decreased tumor growth. The combination of AZD6738 and cisplatin showed more potent growth inhibitory effects, decreased Ki67, increased Tunel than monotherapy of each drug. Conclusion: In BTC, DDR targeting strategy using ATR inhibitor demonstrated promising antitumor activity alone or in combination with cytotoxic chemotherapeutic agents. This supports further clinical development of DDR targeting strategy in BTC. Citation Format: Ah Rong Nam, Ji Eun Park, Ju Hee Bang, Mei Hua Jin, Do Youn Oh, Yung Jue Bang. Evaluation of DDR-targeting strategy using ATR inhibitor in biliary tract cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1421. doi:10.1158/1538-7445.AM2017-1421