Non-Covalent 3CLpro Inhibitors with Cross-Lineage Activity Against Zoonotic Betacoronavirus

The high genetic diversity and broad host range of betacoronavirus lead to frequent zoonotic outbreaks, posing a severe threat to public health. Therefore, the development of broad-spectrum antiviral agents is critical. Our study evaluated the broad-spectrum antiviral activity of 3CL protease (3CLpro) inhibitors AKEX0730 and AKEX0757 against seven representative betacoronavirus strains from Sarbecovirus and Merbecovirus. Their efficacy was confirmed via in vitro live virus inhibition assays, and the binding mechanism and stability with conserved viral targets were elucidated using molecular docking and molecular dynamics (MD) simulations. Our experiments demonstrated that both AKEX0730 and AKEX0757 exhibit significant broad-spectrum inhibition against coronaviruses originating from diverse hosts. These findings highlight their potential as highly potent broad-spectrum antiviral agents, holding substantial promise for the prophylaxis and treatment of emerging zoonotic coronaviruses.