Effects of Nucleos(T)Ide Analogue Therapy on Hepatocyte Clonal Expansion and Hepatitis B Virus DNA Integration in Chronic Hepatitis B Infection: Implications for Hepatitis B Treatment

Background: Integration of HBV DNA into human genome and hepatocyte clonal expansion are implicated in hepatocellular carcinoma (HCC) development in chronic hepatitis B (CHB) infection. This study investigated the effect of nucleos(t)ide analogues (NUC) treatment on HBV DNA integration and hepatocyte clonal expansion. Methods: Twenty-eight patients receiving NUCs (11 lamivudine, seven telbivudine, ten entecavir) were included. All had liver biopsies at baseline and year 1, and seven had a third biopsy at year 10. HBV DNA integration and hepatocyte clone size were assessed by inverse PCR, a semiquantitative, end-point PCR-based method which specifically detected viral-host integration junctions. Findings: All patients had detectable HBV DNA integration at baseline, with median integration frequency of 1·01×109 per liver and hepatocyte clone size of 2·21×10 5 . Neither integration frequency nor hepatocyte clone size correlated with age and HBV virologic parameters. After one year of treatment, HBV DNA integration was still detectable in all patients, with a median of 5·74×108 integration per liver (0·22 log reduction; p=0·011) and hepatocyte clone size of 1·22×105 (0·45 log reduction; p=0·026). HBV DNA integration remained detectable in the seven patients at year 10 of treatment, with a median integration frequency of 4·84×107 integration per liver (0·93 log reduction from baseline) and hepatocyte clone size of 1·89×104 (1·02 log reduction from baseline). From baseline through year 1 to year 10, there was a decreasing trend in both integration frequency and hepatocyte clone size (p=0·066 and 0·018, respectively). Interpretation: NUCs reduced both HBV DNA integration and hepatocyte clonal expansion, suggesting another alternative pathway besides direct viral suppression to reduce HCC risk. Our findings also supported an early treatment of CHB to prevent HCC. Funding: None to declare. Declaration of Interest: D Wong received speaker’s fees from Abbott Laboratories and travel support from Gilead Sciences and Chong Lap. CL Lai received speaker’s fees from Gilead Sciences, GlaxoSmithKline and AbbVie. WK Seto received speaker’s fees from AstraZeneca and Mylan, is an advisory board member of CSL Behring, is an advisory board member and received speaker’s fees from AbbVie, and is an advisory board member, received speaker’s fees and researching funding from Gilead Sciences. MF Yuen serves as advisor/consultant for AbbVie, Assembly Biosciences, Aligos Therapeutics, Arbutus Biopharma, Bristol Myer Squibb, Clear B Therapeutics, Dicerna Pharmaceuticals, Finch Therapeutics, GlaxoSmithKline, Gilead Sciences, Immunocore, Janssen, Merck Sharp and Dohme, Hoffmann-La Roche and Springbank Pharmaceuticals, Vir Biotechnology and receives grant/research support from Assembly Biosciences, Aligos Therapeutics, Arrowhead Pharmaceuticals, Bristol Myer Squibb, Fujirebio Incorporation, Gilead Sciences, Immunocore, Merck Sharp and Dohme, Hoffmann-La Roche, Springbank Pharmaceuticals and Sysmex Corporation. The remaining authors have no conflict of interests. Ethical Approval: This study was approved by the Institution Review Board of The University of Hong Kong and Hospital Authority Hong Kong West Cluster (UW 19-842).