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Combination therapy with ribavirin, interferon, and viral protease inhibitors could be expected to elicit a high level of sustained virologic response in patients infected with hepatitis C virus (HCV). However, several severe side effects of this combination therapy have been encountered in clinical trials. In order to develop more effective and safer anti-HCV compounds, we employed the replicon systems derived from several strains of HCV to screen 84 extracts from 54 organisms that were gathered from the sea surrounding Okinawa Prefecture, Japan. The ethyl acetate-soluble extract that was prepared from marine sponge  Amphimedon  sp. showed the highest inhibitory effect on viral replication, with EC 50  values of 1.5 and 24.9 µg/ml in sub-genomic replicon cell lines derived from genotypes 1b and 2a, respectively. But the extract had no effect on interferon-inducing signaling or cytotoxicity. Treatment with the extract inhibited virus production by 30% relative to the control in the JFH1-Huh7 cell culture system. The  in vitro  enzymological assays revealed that treatment with the extract suppressed both helicase and protease activities of NS3 with IC 50  values of 18.9 and 10.9 µg/ml, respectively. Treatment with the extract of  Amphimedon  sp. inhibited RNA-binding ability but not ATPase activity. These results suggest that the novel compound(s) included in  Amphimedon  sp. can target the protease and helicase activities of HCV NS3.

Inhibition of Both Protease and Helicase Activities of Hepatitis C Virus NS3 by an Ethyl Acetate Extract of Marine Sponge Amphimedon sp