The aim of this study was to investigate the mechanism of inhibition of hepatitis B virus replication by penciclovir-triphosphate, the active metabolite of famciclovir. A recently developed in vitro translation assay for the expression of an enzymatically active duck hepatitis B virus (DHBV) reverse transcriptase was used to assess the inhibitory activity of penciclovir-triphosphate (PCV-TP) in comparison with other guanosine analogue triphosphates. Acyclovir-triphosphate (ACV-TP), the chiral triphosphates of penciclovir (PCV), (R)-PCV-TP and (S)-PCV-TP, and carbocyclic 2′-deoxyguanosine-TP (CDG-TP) did inhibit reproducibly minus strand DNA synthesis to different extents. CDG-TP was the most potent inhibitor of dGTP incorporation. The inhibitory effect of these compounds against the incorporation of the first nucleotide of minus strand DNA, dGMP, was similar to that observed with DNA chain elongation. 2′,3′-dideoxyguanosine-TP (ddG-TP), ACV-TP and both (R) and (S)-PCV-TP inhibited the incorporation of the next nucleotides in the short DNA primer, whereas CDG-TP did not. These results demonstrate that PCV-TP inhibits hepadnavirus reverse transcription by inhibiting the synthesis of the short DNA primer. The data obtained with the inhibition of the enzymatic activity of the DHBV polymerase provides a new insight into the mechanism of action of penciclovir-triphosphate on HBV replication.
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