High-Throughput Screening of HCV RNA Replication Inhibitors by Means of a Reporter Replicon System

Efforts to find effective treatment for hepatitis C virus (HCV) have been hampered by the lack of a robust in vitro infectious tissue-culture system for this virus. A subgenomic replicon system was first developed in 1999 and has since been extensively optimized to accommodate the need for conveniently measuring HCV replication in vitro and widely adopted in HCV drug-discovery efforts. Here we describe the adaptation of a modified replicon system for a high-throughput screening (HTS) in anti-HCV drug discovery. In this system, the antiviral activity and cytotoxicity of any experimental compound are measured from a single well. This duplex measurement greatly increases the efficiency of the HTS while lowering the cost. The usefulness of this approach has been supported by the recent discovery of many new lead compounds from our HTS efforts in the past two years.