Abstract
Amphihevir, a benzofuran derivative, is the first reported NS4B inhibitor that has advanced to clinical trials (currently in Phase Ib). Here, we report the results of a preclinical study of its potency, toxicity, selectivity, DMPK, and safety profiles. Amphihevir displayed good antiviral activities against genotype 1a (EC50=0.34 nM) and genotype 1b (EC50=1.97 nM) replicons and evident cytotoxicity in twelve strains of cell lines derived from animals and humans. Amphihevir was found to be inactive against other viruses, human kinases, and GPCRs, which implies its good selectivity. A 9-day long-term treatment of genotype 1b replicon with Amphihevir resulted in a 3.8 Log10 decline of the hepatitis C viral RNA at a concentration of 25×EC90 Drug resistance screening showed that mutations occurred at H94, F98, and V105 of NS4B, which mediated the resistance to Amphihevir. This result suggests that NS4B is the main target of Amphihevir. There was no cross-resistances between Amphihevir and NS5A, NS3/4A, and NS5B inhibitors, suggesting that Amphihevir on combination of other anti- hepatitis C virus drugs could treat hepatitis C, as proven by studies of Amphihevir and other hepatitis C virus inhibitors. Pharmacokinetic studies demonstrated that Amphihevir has good oral bioavailability and appropriate T1/2 in rats and dogs, thereby supporting its use once per day. Finally, Amphihevir showed good safety profiles in rats and dogs. The results shed light on the use of Amphihevir as a potential treatment option for chronic hepatitis C patients.