The Landscape of Cell-Free HBV Integrations and Mutations in Cirrhosis and Hepatocellular Carcinoma Patients
Bo Zheng, Xiao-Long Liu, Rong Fan, Jian Bai, Hao Wen, Lutao Du, Guo‐Qing Jiang, Chunying Wang, Xiaotang Fan, Yi-Nong Ye, Yun-Song Qian, Yingchao Wang, Gao-Jing Liu, Guohong Deng, Feng Shen, Heping Hu, Hui Wang, Qingzheng Zhang, Lan-Lan Ru, Jing Zhang, Yanhang Gao, Jie Xia, Huadong Yan, Min-Feng Liang, Yan-Long Yu, Fu-Ming Sun, Yujing Gao, Jian Sun, Chun-Xiu Zhong, Yin Wang, Fei Kong, Jinming Chen, Dan Zheng, Yuan Yang, Chuanxin Wang, Lin Wu, Jinlin Hou, Jingfeng Liu, Hongyang Wang, Lei Chen
Abstract
Abstract Purpose: Intratumoral hepatitis B virus (HBV) integrations and mutations are related to hepatocellular carcinoma (HCC) progression. Circulating cell-free DNA (cfDNA) has shown itself as a powerful noninvasive biomarker for cancer. However, the HBV integration and mutation landscape on cfDNA remains unclear. Experimental Design: A cSMART (Circulating Single-Molecule Amplification and Resequencing Technology)-based method (SIM) was developed to simultaneously investigate HBV integration and mutation landscapes on cfDNA with HBV-specific primers covering the whole HBV genome. Patients with HCC (n = 481) and liver cirrhosis (LC; n = 517) were recruited in the study. Results: A total of 6,861 integration breakpoints including TERT and KMT2B were discovered in HCC cfDNA, more than in LC. The concentration of circulating tumor DNA (ctDNA) was positively correlated with the detection rate of these integration hotspots and total HBV integration events in cfDNA. To track the origin of HBV integrations in cfDNA, whole-genome sequencing (WGS) was performed on their paired tumor tissues. The paired comparison of WGS data from tumor tissues and SIM data from cfDNA confirmed most recurrent integration events in cfDNA originated from tumor tissue. The mutational landscape across the whole HBV genome was first generated for both HBV genotype C and B. A region from nt1100 to nt1500 containing multiple HCC risk mutation sites (OR > 1) was identified as a potential HCC-related mutational hot zone. Conclusions: Our study provides an in-depth delineation of HBV integration/mutation landscapes at cfDNA level and did a comparative analysis with their paired tissues. These findings shed light on the possibilities of noninvasive detection of virus insertion/mutation.