DNA Methylation of Biliary Tract Cancer Identifies Subtypes with Different Immune Characteristics and Clinical Outcomes

DNA methylation-associated studies on biliary tract cancer (BTC), including cholangiocarcinoma (CCA) and gallbladder cancer (GBC), may improve the BTC classification scheme. In our cohort including 57 CCAs and 48 GBCs, lower level of methylation changes were associated with fewer copy number variations, fewer mutations of cell cycle pathway, CCND/E1, FGF/FGFR family, and NOTCH pathway, and remarkably longer overall survival (OS, HR=0.07, 95% CI 0.01-0.65, P =0.017). Additionally, we constructed a 12-feature prognostic prediction model that effectively predicted OS (HR=0.21, 95% CI 0.10-0.43, P <0.001). Moreover, the BTCs with minimal methylation changes exhibited higher immune-related signatures, infiltration of CD8+ lymphocytes, and PD-L1 expression, indicating an inflamed tumor immune microenvironment (TIME) with PD-L1 expression elicited by immune attack, potentially suggesting better immunotherapy efficacy. In BTCs, DNA methylation is a powerful tool for molecular classification, serving as a robust indicator of genomic aberrations, survival outcomes, and tumor immune microenvironment. Our integrative analysis provides insights into prognostication after curative surgery and patient selection for immunotherapy. Funding Statement: This work was supported by the Innovation Group Project of Shanghai Municipal Health Commission (2019CXJQ03) and the Shanghai Municipal Health Commission (202040045). Declaration of Interests: Y. Xu, G. Wang, C. Li, Y. Zhang, J. Zhao, C. Wang, X. Wen, Z. Zhang, B. Li, H. Zhang, Z. Zhang, and S. Cai are employees of Burning Rock Biotech. Other authors declare no potential conflicts of interest. Ethics Approval Statement: All collection and usage of human samples and clinical data were in accordance with the principles of the Declaration of Helsinki and approved by the Ethics Committee of Eastern Hepatobiliary Surgery Hospital (EHBHKY2018-02-014). The written consents were received from all the participated patients.