Identification of FDA-approved IMPDH inhibitors boosting the broad-spectrum antiviral potential of ribavirin through depleting cellular GTP pool

Nucleoside analogues are among the most promising broad-spectrum antiviral agents. We developed a high-throughput synergistic screening system to identify US Food and Drug Administration-approved drugs that enhance the effectiveness of nucleoside analogues. Unlike conventional combination approaches, this strategy does not require boost agents to have direct antiviral activity; instead, they can enhance antiviral agent's effect through novel synergistic mechanisms. We identified mycophenolate mofetil (MMF), which significantly boosts ribavirin's (RBV) antiviral efficacy against influenza A virus, even at concentrations where MMF alone shows minimal activity. MMF enhances RBV by inhibiting IMPDH (inosine-5'-monophosphate dehydrogenase) and reducing cellular GTP levels, potentially increasing RBV incorporation and its antiviral potency. This mechanism was confirmed by adding exogenous GTP, which reversed the synergy. In a mouse model, MMF also enhanced RBV's effectiveness against PR8 in vivo. Moreover, MMF showed synergistic effects against other influenza subtypes and RNA viruses including ZIKV and HCoV-OC43, demonstrating the potential wide applicability of this strategy. Our study introduces a new screening system and identifies novel RBV combinations, offering a promising approach for developing effective broad-spectrum antiviral combinations against emerging viral threats.