4'-Fluorouridine inhibits alphavirus replication and infection in vitro and in vivo

Chikungunya virus (CHIKV) is an enveloped, positive-sense RNA virus that has re-emerged to cause millions of human infections worldwide. In humans, acute CHIKV infection causes fever and severe muscle and joint pain. Chronic and debilitating arthritis and joint pain can persist for months to years. To date, there are no approved antivirals against CHIKV. Recently, the ribonucleoside analog 4'-fluorouridine(4'-FlU) was reported as a highly potent orally available inhibitor of SARS-CoV-2, respiratory syncytial virus, and influenzavirus replication. In this study, we assessed 4'-FlU's potency and breadth of inhibition against a panel of alphaviruses including CHIKV, and found that it broadly suppressed alphavirus production in cell culture. 4'-FlU acted on the viral RNA replication step, and the first4 hours post-infection were the critical time for its antiviral effect.In vitro replication assays identifiednsP4 as the target of inhibition. In vivo, treatment with 4'-FlU reduced disease signs, inflammatoryresponses, and viral tissue burden in mouse models of CHIKV and Mayaro virus infection. Treatment initiated at 2 hours post-infection was most effective;however, treatment initiated as late as 24-48 hours post-infection produced measurable antiviral effectsin the CHIKV mouse model. 4'-FlU showed effectiveoral delivery in our mouse model and resulted in the accumulation of both 4'-FlU and its bioactive triphosphate form in tissues relevant to arthritogenic alphavirus pathogenesis. Together, our data indicate that 4'-FlU inhibits CHIKV infection in vitro and in vivo and is a promising oral therapeutic candidate against CHIKV infection.