Chronic infection with hepatitis C virus (HCV) remains a global health concern. Using both in vitro and cell-based assays, a series of small molecule agents specific for the viral RNA-dependent RNA polymerase have been shown to interfere with viral RNA replication. Although no agents targeting this viral enzyme have demonstrated sustained efficacy in infected patients as measured by reduction in viral load at 72 weeks post-treatment, proof-of-concept has been achieved in the clinic. A comprehensive account of the structure-activity relationship for nucleoside and non-nucleoside inhibitors of HCV polymerase, as well as consideration of early discovery biologic approaches targeting NS5B are reviewed.