ABSTRACT
Background: SARS-CoV-2 evolution has contributed to successive waves of infections and severely compromised the efficacy of available SARS-CoV-2 monoclonal antibodies. Decaying vaccine-induced immunity, vaccine hesitancy, and limited vaccine protection in older and immunocompromised populations further compromises vaccine efficacy at the population level. Early antiviral treatments, including intravenous remdesivir (RDV), reduce hospitalization and severe disease due to COVID-19. An orally bioavailable RDV analog could facilitate earlier widespread administration to non-hospitalized COVID-19 patients. Method(s): We synthesized monoalkyl glyceryl ether phosphodiesters of GS-441524 (RVn), lysophospholipid analogs which allow for oral bioavailability and stability in plasma. We evaluated the in vivo efficacy of our lead compound, 1-O-octadecyl-2-O-benzyl-sn-glyceryl-3-phospho-RVn (V2043), in an oral treatment model of murine SARS-CoV-2 infection. We then synthesized numerous phospholipid analogs of RVn and determined which modifications enhanced in vitro antiviral activity and selectivity. The most effective compounds against SARS-CoV-2 were then evaluated for antiviral activity against other RNA viruses. Result(s): Oral treatment of SARS-CoV-2 infected BALB/c mice with V2043 (60 mg/kg once daily for 5 days, starting 12 hrs after infection) reduced lung viral load by more than 100-fold versus vehicle at day 2 and to below the LOD at day 5. V2043 inhibited previous and contemporary SARS-CoV-2 Variants of concern to a similar degree, as measured by the half maximal effective concentration (EC50) in a human lung epithelial cell line (Calu-3). Evaluation of multiple RVn analogs with hydrophobic esters at the sn-2 of glycerol revealed that in vitro antiviral activity was improved by the introduction of a 3-fluoro-4-methoxysubstituted benzyl or a 3-or 4-cyano-substituted benzyl. These compounds showed a 2-to 6-fold improvement in antiviral activity compared to analogs having an unsubstituted benzyl, such as V2043, and were more active than RDV. These compounds also showed enhanced antiviral activity against multiple contemporary and emerging RNA viruses. Conclusion(s): Collectively, our data support the development of RVn phospholipid prodrugs as oral antiviral agents for prevention and treatment of SARS-CoV-2 infections and as preparation for future outbreaks of pandemic RNA viruses.