Inhibition of hepatitis C virus NS3 helicase by manoalide

The hepatitis C virus (HCV) causes one of the most prevalent chronic infectious diseases in the world, hepatitis C, which ultimately develops into liver cancer through cirrhosis. The NS3 protein of HCV possesses nucleoside triphosphatase (NTPase) and RNA helicase activities. As both activities are essential for viral replication, NS3 is proposed as an ideal target for antiviral drug development. In this study, we identified manoalide (1) from marine sponge extracts as an RNA helicase inhibitor using a high-throughput screening photoinduced electron transfer (PET) system that we previously developed. Compound 1 inhibits the RNA helicase and ATPase activities of NS3 in a dose-dependent manner, with IC₅₀ values of 15 and 70 μM, respectively. Biochemical kinetic analysis demonstrated that 1 does not affect the apparent K_m value (0.31 mM) of NS3 ATPase activity, suggesting that 1 acts as a noncompetitive inhibitor. The binding of NS3 to single-stranded RNA was inhibited by 1. Manoalide (1) also has the ability to inhibit the ATPase activity of human DHX36/RHAU, a putative RNA helicase. Taken together, we conclude that 1 inhibits the ATPase, RNA binding, and helicase activities of NS3 by targeting the helicase core domain conserved in both HCV NS3 and DHX36/RHAU.