Reduction of HBV cccDNA level by stably expressed HBV C protein C-terminal siRNA in vitro

Objective To study the effect of hepatitis B virus (HBV) C protein C-terminal small interference RNA (siRNA) on reduction of HBV covalently closed circular DNA (cccDNA) level. Methods A pair of oligos with sites of restriction endonuclease on their terminal was synthesized and annealed to form double-stranded oligo. Then the siRNA oligos were inserted into recombined adeno-associated viral vector (rAAV) for construction of recombinant (rAAV-siRNA), rAAV-siRNA could stably express short hairpin RNA which could be digested by dicer to become 22-mer siRNA complementary to the C-terminal site of HBV C protein (2389nt-2410nt). When 22.1.5 cells were treated with rAAV-siRNA 72 hours later, HBV cccDNA and HBV DNA levels in cultural supernatant and cells were determined by real-time PCR. Results The levels of HBV cccDNA and HBV DNA were significantly decreased in rAAV-siRNA group compared with blank control group (P 0.01 ). No inhibitory effects on the levels of HBV cccDNA and HBV DNA in mock sequence group. Conclusion HBV C-terminal siRNA can decrease HBV cccDNA level and HBV replication in vitro.