Twenty-five 2,4-diketolactam derivatives t1~t25 were synthesized using β-alanine ethyl ester hydrochloride and monoethyl malonyl chloride as initial materials. The inhibitory activities of these compounds against the main protease (Mpro) were tested. The experimental results showed that t5, t9, and t18 exhibited relatively high Mpro inhibitory rates. Further gradient concentration assays of three compounds were conducted to evaluate the Mpro inhibitory activities. Their half-maximal inhibitory concentrations (IC₅₀) against Mpro were 0.445, 0.425, and 0.088 μmol/L, respectively, whereas the IC₅₀ value of nirmatrelvir was 0.426 μmol/L. The inhibitory activity showed that the IC₅₀ values of t5 and t9 were comparable to those of nirmatrelvir, while the IC₅₀ value of t18 was lower. Molecular docking revealed that t5 interacted with the key residue His41, while t9 and t18 interacted with both Cys145 and His41 in the active site of Mpro. This study demonstrates that the 2,4-diketolactam moiety can serve as a privileged building block for targeting the Mpro. The above three compounds exhibit potent inhibitory activity against coronavirus Mpro and represent promising candidates for the development of anti-coro- navirus agents, providing a material basis for further pharmacodynamic evaluations.

Key words: coronavirus, 2,4-diketolactam derivatives, main protease (Mpro), enzyme inhibitors