Phenothiazines inhibit hepatitis C virus entry, likely by increasing the fluidity of cholesterol-rich membranes

Despite recent progress in the development of direct-acting antiviral agents against hepatitisCvirus (HCV), more effective therapies are still urgently needed.Weand others previously identified three phenothiazine compounds as potentHCVentry inhibitors. In this study, we show that phenothiazines inhibitHCVentry at the step of virus-host cell fusion, by intercalating into cholesterol-rich domains of the target membrane and increasing membrane fluidity. Perturbation of the alignment/packing of cholesterol in lipid membranes likely increases the energy barrier needed for virus-host fusion. A screening assay based on the ability of molecules to selectively increase the fluidity of cholesterol-rich membranes was subsequently developed. One compound that emerged from the library screen, topotecan, is able to very potently inhibit the fusion of liposomes with cell culture-derivedHCV(HCVcc). These results yield new insights intoHCVinfection and provide a platform for the identification of newHCVinhibitors.