Mechanism of entecavir-induced reduction of HBV DNA integration

Nucleos(t)ide analogues (NUC) treatment can reduce the extent of HBV DNA integration in chronic hepatitis B (CHB) patients. However, the mechanism by which NUC reduces HBV integration is unclear. This study investigated the effects of entecavir (ETV), one of the commonly used NUC, on cells with HBV integration. Full genome-length HBV DNA was inserted into HepG2 cell genome using the sleeping beauty transposon system. The resulting cells, named HepG2/SB/HBV, was subjected to ETV treatment. In ETV-treated HepG2/SB/HBV, intracellular HBV DNA was reduced by 2-fold. When treated with ETV, HepG2/SB/HBV had an impaired cell survival (25% reduction in cell proliferation rate when compared with untreated cells; p = 0.043). The median integration frequency in untreated HepG2/SB/HBV was 16 integration sites per 10⁵ cells, which was reduced to 14.8 integration sites per 10⁵ cells when treated with ETV. Analysis of the expression of apoptosis and mitosis markers showed that ETV-treated HepG2/SB/HBV had a reduced expression of mitosis markers phospho-cell division control-2 (p-cdc-2) and phospho-histone H3 (p-histone H3), but that of the apoptotic markers [Poly-ADP-ribose-polymerase] (PARP) and caspase-3 were not affected. In conclusion, ETV suppressed cell proliferation of hepatoma cells with HBV integration via interference of mitosis and reduced expression of p-histone H3, thereby reducing the number of HBV-integrated hepatocytes. This may be the mechanism by which HBV integration is reduced in CHB patients who received ETV therapy.