HBV integration into KMT2B increases malignant phenotype of hepatocellular carcinoma by epigenetic reprogramming.

Lysine methyltransferase 2B (KMT2B, also named MLL4) is a frequently reported target of hepatitis B virus (HBV) genomic integration in hepatocellular carcinoma (HCC). However, the mechanism by which HBV integration at KMT2B (HBV-KMT2B) promotes HCC evolution remains largely unknown. We performed HBV capture sequencing and transcriptome sequencing on paired tumoral and adjacent hepatic tissues from 50 HBV-positive HCC patients, enabling systematic characterization of HBV-KMT2B and its aberrant transcripts. We used methylome sequencing, assay for transposase-accessible chromatin using sequencing, and cleavage under targets and tagmentation to assess the impact of HBV-KMT2B on epigenetic modifications and to identify key downstream molecules. It was found that the HBV integration loads accumulated with HCC evolution. HBV-KMT2B was the second most frequently detected HBV integration in HCC. HBV-KMT2B was mostly exclusive with other high-frequent integrations and the common oncogenic driver mutations. Integrated HBV fragments, predominantly from HBx, clustered within KMT2B exons 3-4, resulting in decreased transcription of full-length KMT2B and the generation of N-terminally truncated KMT2B (KMT2B-Ct). Both knockdown of full-length KMT2B and ectopic expression of KMT2B-Ct significantly promoted cell proliferation, migration, and invasion and inhibited apoptosis in HCC. KMT2B knockdown upregulated the stemness marker CD133 and suppressed the forward embryonic developmental pathways by regulating the promoter methylation. KMT2B-Ct suppressed vanin-1 (VNN1) by decreasing H3K4me3 enrichment at the VNN1 locus and reducing chromatin accessibility of VNN1. Overexpression of VNN1 suppressed the malignant phenotype of HCC cells. Thus, HBV-KMT2B drives HCC evolution by epigenetic modifications to upregulate CD133 and suppress VNN1, thereby inducing dedifferentiation in hepatocarcinogenesis.