One of the most challenging goals of hepatitis C virus (HCV) research is to 
develop well-tolerated regimens with high cure rates across a variety of patient 
populations. Such a regimen will likely require a combination of at least two 
distinct direct-acting antivirals (DAAs). Combining two or more DAAs with 
different resistance profiles increases the number of mutations required for 
viral breakthrough. Currently, most DAAs inhibit HCV replication. We recently 
reported that the combination of two distinct classes of HCV inhibitors, entry 
inhibitors and replication inhibitors, prolonged reductions in extracellular HCV 
in persistently infected cells. We therefore sought to identify new inhibitors 
targeting aspects of the HCV replication cycle other than RNA replication. We 
report here the discovery of the first small-molecule HCV infectivity inhibitor, 
GS-563253, also called HCV infectivity inhibitor 1 (HCV II-1). HCV II-1 is a 
substituted tetrahydroquinoline that selectively inhibits genotype 1 and 2 HCVs 
with low-nanomolar 50% effective concentrations. It was identified through a 
high-throughput screen and subsequent chemical optimization. HCV II-1 only 
permits the production and release of noninfectious HCV particles from cells. 
Moreover, infectious HCV is rapidly inactivated in its presence. HCV II-1 
resistance mutations map to HCV E2. In addition, HCV-II prevents HCV endosomal 
fusion, suggesting that it either locks the viral envelope in its prefusion 
state or promotes a viral envelope conformation change incapable of fusion. 
Importantly, the discovery of HCV II-1 opens up a new class of HCV inhibitors 
that prolong viral suppression by HCV replication inhibitors in persistently 
infected cell cultures.