Discovery of a novel antiviral agent targeting the nonstructural protein 4 (nsP4) of chikungunya virus

Chikungunya fever (CHIKF) is caused by chikungunya virus (CHIKV) infection which is a re-emerging mosquito-borne zoonosis. At present, there are no approved therapeutics for CHIKF. Herein, we have investigated candidate compounds which can inhibit CHIKV infection. Screening of chemical compound libraries were performed and one candidate, a benzimidazole-related compound designated Compound-A was found to inhibit infection by several CHIKV strains and a Sindbis virus strain at nanomolar concentrations. To investigate the inhibitory mechanism of action, a Compound-A resistant CHIKV (res-CHIKV) was isolated and a key mutation associated with resistance was identified by reverse-genetic recombinant CHIKVs containing amino acid substitutions present in res-CHIKV. These results demonstrated that the target site of Compound-A was the M2295 residue in the nonstructural protein 4 (nsP4), which is located in one of the functional domains of RNA-dependent RNA-polymerase (RdRp). We also confirmed that Compound-A inhibits RdRp function of CHIKV by using CHIKV replicons.