Evaluation of 6-substituted coumarins as SARS-CoV-2 main protease inhibitors: enzymatic, cellular, and computational insights

Abstract

The SARS-CoV-2 main protease (Mpro) plays a crucial role in viral replication, making it an attractive target for antiviral drug development. This study evaluates the inhibitory potential of nineteen 6-substituted coumarin derivatives (1a–p and 2a–c) against Mpro using a FRET-based enzymatic assay. Among them, the chlorinated derivative 1f (AKE-9306) exhibited the highest potency with an IC50 value of 61.77 µM. Enzyme kinetics analysis through Michaelis–Menten and Lineweaver–Burk plots revealed that compound 1f may act through a mixed-type inhibition mechanism with a preference for the free enzyme. In cellular assays, 1f showed significant antiviral efficacy (IC50 = 16.02 µM, CC50 >50 µM, selectivity index >3.12). Molecular docking and dynamics simulations confirmed its stable interaction within the Mpro active site, supported by MM-PBSA calculations. These findings identify compound 1f as a hit coumarin-based scaffold for further optimization toward improved SARS-CoV-2 Mpro inhibition and antiviral activity.