Variants in Oxidative Stress-Related Genes Affect the Chemosensitivity Through Nrf2-Mediated Signaling Pathway in Biliary Tract Cancer

Oxidative stress (OS) and their effectors play critical roles in carcinogenesis and chemoresistance. However, the role of OS-related genes variants in biliary tract cancer (BTC) chemoresistance remains unknown. In this work, we aim to investigate OS-dependent molecular mechanisms underlying chemoresistance, and find potential biomarkers to predict chemotherapy response for BTC. Sixty-six SNPs in 21 OS-related genes were genotyped and analyzed in 367 BTC patients. Immunoblot, immunohistochemical, immunofluorescent, quantitative PCR, chromatin immunoprecipitation analysis and study of animal xenograft models were performed to discover OS-related susceptibility genes underlying chemoresistance mechanism of BTC. We found that 3 functional polymorphisms (CAT_rs769217, GPX4_rs4807542, and GSR_rs3779647), which were shown to affect their respective gene expression levels, modified the effect of chemotherapy on postoperative survival (POS). We then demonstrated that knockdown of GPX4, CAT, or GSR induced chemoresistance through elevation of ROS level and activation of Nrf2-ABCG2 pathway in BTC cell lines. The association between Nrf2 expression and BTC prognosis is only found in patients who received chemotherapy. Knockdown of Nrf2 enhanced chemosensitivity or even eliminated postoperative recurrence in BTC xenograft mouse models. Importantly, upon chemotherapy treatment patients harboring high OS-related score received higher survival benefit from adjuvant chemotherapy compared with patients with low OS-related score. The result of our study suggests, for the first time, that the variations in CAT, GPX4, and GSR can be used as novel biomarkers for prediction of chemosensitivity in BTC, and it also suggests that potential Nrf2 suppression could be used for BTC treatment. Funding Statement: This work was supported by the National Science Foundation of China (81773184, 81802424, and 81803014), Foundation of Shanghai Shen Kang Hospital Development Center (16CR2002A and 16CR3028A), and Shanghai Outstanding Academic Leaders Plan (2016, JW). Declaration of Interests: The authors declare no financial or other relationships that may lead to a conflict of interest in this study. Ethics Approval Statement: This study was granted approval by the Ethical Committee of the Renji hospital, Shanghai Jiao Tong University School of Medicine. All of the subjects in this study provided written informed consent according to the protocols approved by the Ethics Committees, and their data were analyzed anonymously.