Macrocyclic Azapeptide Nitriles: Structure-Based Discovery of Potent SARS-CoV-2 Main Protease Inhibitors as Antiviral Drugs.

Given the crucial role of the main protease (M ^pro ) in the replication cycle of SARS-CoV-2, this viral cysteine protease constitutes a high-profile drug target. We investigated peptidomimetic azapeptide nitriles as auspicious, irreversibly acting inhibitors of M ^pro . Our systematic approach combined an M ^pro active-site scanning by combinatorially assembled azanitriles with structure-based design. Encouraged by the bioactive conformation of open-chain inhibitors, we conceptualized the novel chemotype of macrocyclic azanitriles whose binding mode was elucidated by cocrystallization. This strategy provided a favorable entropic contribution to target binding and resulted in the development of the extraordinarily potent M ^pro inhibitor 84 with an IC ₅₀ value of 3.23 nM and a second-order rate constant of inactivation, k _inac / K _i , of 448,000 M ^-1 s ^-1 . The open-chain M ^pro inhibitor 58 , along with the macrocyclic compounds 83 and 84 , a broad-spectrum anticoronaviral agent, demonstrated the highest antiviral activity with EC ₅₀ values in the single-digit micromolar range. Our findings are expected to promote the future development of peptidomimetic M ^pro inhibitors as anti-SARS-CoV-2 agents.