Human norovirus is recognised as one of the world’s leading causes of acute gastroenteritis, responsible for approximately 677 million cases, 219,000 deaths, and USD $4.2 billion in global health expenditure each year. Yet despite this heavy social and economic burden, there are currently no specific antivirals available to treat norovirus infections, or for prophylaxis in outbreak settings.
In this study, a structure-based virtual screening of commercially available, drug-like compounds (~300,000) was performed on the suramin binding-site of the norovirus RNA-dependent RNA polymerase (RdRp). Following virtual screening, 62 compounds were selected and examined for inhibition of norovirus RdRp activity using a quantitative, fluorescent RdRp assay. Eight antiviral candidates demonstrated RdRp inhibition (>25% inhibition at 10 µM), which was verified using a secondary gel-shift RdRp assay. Based on these results, further structure-activity relationship enhancements of the initial hits resulted in the synthesis of 31 novel compounds, which were examined for inhibitory activity using polymerase assays, and also cytotoxicity using a cell viability assay. Of these 31 compounds, five demonstrated norovirus RdRp inhibition (>50% at 10 µM), with the best candidate, AB-1512, demonstrating a half maximal inhibitory concentration of 5.6 µM and a half maximal cytotoxic concentration of 68.6 µM.
Combinational treatment of AB-1512 with the norovirus RdRp inhibitor PPNDS, revealed antagonism with a combination index of 1.7, indicating that both compounds bind in the RdRp Site-B pocket. Additional studies were carried out on two mutant RdRps (Q414A and R419A), where the mutated residues were previously shown to be critical for the binding of specific Site-B compounds. However, no effect was observed on AB-1512 inhibitory activity, suggesting that while AB-1512 binds in Site-B, the interactions are with likely to be with distinct residues.
This study has revealed a novel antiviral scaffold, AB-1512, for further structural modifications for treating and preventing human norovirus infections.