An Oral Antiviral Combination Against Severe COVID-19 Enabled by the Synergistic Mechanisms of Action of Auranofin and Remdesivir

Auranofin, an FDA-approved oral anti-rheumatic drug, was previously shown by our group to restrict Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) replication and pathology in preclinical models. Here, we systematically characterized its mechanism of action (MOA) through biochemical drug-protein interactions, cell-based mechanistic assays, and preclinical efficacy studies. Auranofin directly binds and inhibits SARS-CoV-2 MPro while simultaneously targeting multiple stages of the viral life cycle, disrupting endosomal trafficking, suppressing Spike S1/S2 proteolytic processing, impairing syncytia formation, and attenuating NF-κB-driven proinflammatory signaling. In cell culture, Auranofin demonstrated robust synergy with Remdesivir but competed with the MPro inhibitor Nirmatrelvir, informing rational combination design. In mice infected with the SARS-CoV-2 Delta variant, the oral combination of Auranofin and the Remdesivir prodrug GS-621763 outperformed monotherapy, markedly reducing viral replication, inflammatory cytokine responses, and lung pathology, and improving clinical outcomes. These findings establish a multi-target antiviral MOA of Auranofin and provide a mechanistic rationale for an all-oral antiviral combination regimen against COVID-19.