Impact of AAV2 and Hepatitis B Virus Integration Into Genome on Development of Hepatocellular Carcinoma in Patients with Prior Hepatitis B Virus Infection

Purpose: Hepatitis B viral (HBV) DNA is frequently integrated into the genomes of hepatocellular carcinoma (HCC) in patients with chronic HBV infection (chronic HBV, hereafter), whereas the frequency of HBV integration in patients after the disappearance of HBV (prior HBV, hereafter) has yet to be determined. This study aimed to detect integration of HBV and adeno-associated virus type 2 (AAV2) into the human genome as a possible oncogenic event. Experimental Design: Virome capture sequencing was performed, using HCC and liver samples obtained from 243 patients, including 73 with prior HBV without hepatitis C viral (HCV) infection and 81 with chronic HBV. Results: Clonal HBV integration events were identified in 11 (15.0%) cases of prior HBV without HCV and 61 (75.3%) cases of chronic HBV (P < 0.001). Several driver genes were commonly targeted by HBV, leading to transcriptional activation of these genes; TERT [four (5.4%) vs. 15 (18.5%)], KMT2B [two (2.7%) vs. five (6.1%)], CCNE1 [zero vs. one (1.2%)], CCNA2 [zero vs. one (1.2%)]. Conversely, CCNE1 and CCNA2 were, respectively, targeted by AAV2 only in prior HBV. In liver samples, HBV genome recurrently integrated into fibrosis-related genes FN1, HS6ST3, KNG1, and ROCK1 in chronic HBV. There was not history of alcohol abuse and 3 patients with a history of nucleoside analogue treatment for HBV in 8 prior HBV with driver gene integration. Conclusions: Despite the seroclearance of hepatitis B surface antigen, HBV or AAV2 integration in prior HBV was not rare; therefore, such patients are at risk of developing HCC.