Inhibition of hepatitis C virus translation and subgenomic replication by siRNAs directed against highly conserved HCV sequence and cellular HCV cofactors

Background/Aims: Small interfering RNAs (siRNAs) are an efficient tool to specifically inhibit gene expression by RNA interference. Since hepatitis C virus (HCV) replicates in the cytoplasm of liver cells without integration into the host genome, RNA-directed antiviral strategies are likely to successfully block the HCV replication cycle. Additional benefit might arise from inhibition of cellular cofactors of HCV replication, such as proteasome a-subunit 7 (PSMA7) or Hu antigen R (HuR). Methods: In this study, we investigated direct and cofactor-radiated inhibition of HCV by a panel of DNA-based retroviral vectors expressing siRNAs against highly conserved HCV sequences or the putative HCV cofactors PSMA7 and HuR. Effects were determined in HCV IRES-mediated translation assays and subgenomic HCV replicon cells. Results: PSMA7- and HuR-directed siRNAs successfully inhibited expression of the endogenous genes, and PSMA7 and HuR silencing significantly diminished HCV replicon RN X and NS5B protein levels. HCV-directed siRNAs substantially inhibited HCV IRES-mediated translation and subgenomic HCV replication. Combinations of PSMA7 and HuR-directed siRNAs with HCV-directed. siRNAs revealed additive HCV RNA inhibitory effects in monocistronic replicon cells. Conclusions: A dual approach of direct- and cofactor-mediated inhibition of HCV replication might avoid selection of mutants and thereby become a powerful strategy against ACV. (c) 2005 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.