HBV Integration into Host Superenhancers Might Not Be a Risk Factor for Hepatocellular Carcinoma

Abstract

Hepatitis B virus (HBV) DNA integration into the host genome can be found in all phases of chronic HBV infection and plays a role in hepatocarcinogenesis. Integration into CpG islands has been reported to be a risk factor for hepatocellular carcinoma (HCC), but the relationship between HBV integration into other regulatory elements and HCC remains unclear. Superenhancers (SEs) contribute to the cancer cell state by regulating oncogenes. This study aimed to analyze whether integration into host SEs is a risk factor for HCC. We systematically annotated 21,520 HBV integration sites in the human genome obtained from the ViMIC database to determine their distribution in regular elements, including SEs, CpG islands, CCCTC-binding factors (CTCFs)-binding sites, transcription factor-binding sites (TFBSs), and transcription start sites (TSSs). Then, we constructed a logistic regression model to evaluate the relationship between the integration sites and HCC. Integration into CpG islands and TFBS were risk factors for HCC (P = 0.000, OR = 2.65 and P = 0.041, OR = 1.06, respectively. OR = odds ratio), and integration into SEs and CTCF-binding sites were significantly associated with nontumor tissues (P = 0.000, OR = 0.58 and P = 0.000, OR = 0.48, respectively). To further investigate the underlying mechanism, we analyzed CpG methylation and histone modifications at flanking integration sites. We found that regions flanking HBV integration sites in SEs were more likely to be hypermethylated (P = 0.000). Moreover, the hypermethylated SE regions flanking HBV integration sites showed lower levels of epigenomic markers. Our results suggested that integration into SEs might not be a risk factor for HCC. The protective effect observed for integration into host SEs might be associated with hypermethylation.

Received: 24 December 2024 | Revised: 28 May 2025 | Accepted: 12 June 2025 

Conflicts of Interest

The authors declare that they have no conflicts of interest to this work.

Data Availability Statement

The data that support the findings of this study are available in public repositories. HBV integration sites in ViMIC: http://bmtongji. cn/ViMIC/downloaddata/integration/HBV_integration.csv. Human genome regulatory elements in hg38: www.genome.ucsc.edu. SEdb: https://bio.liclab.net/sedb/index.php. CpG methylation data of HepG2: https://www.encodeproject.org/experiments/ENCSR786DCL/. Methylation panel data for fetal livers: https://www.ncbi.nlm.nih.gov/ geo/query/acc.cgi?acc=GSM1014211. Chip-seq data from the ENCODE project: https://www.encodeproject.org/.

Author Contribution Statement

Mengna Zhang: Methodology, Software, Formal analysis, Investigation, Writing – original draft, Visualization. Ying Ming: Conceptualization, Writing – review & editing, Supervision. Yunling Du: Validation. Ziwen Guo: Formal analysis. Ziyuan Xin: Data curation, Investigation. Yanjun Li: Data curation. Ge Yang: Resources. Zhaoyang Jiang: Visualization.