Ethyl caffeate as a novel targeted inhibitor of 3CLpro with antiviral activity against porcine epidemic diarrhea virus.

Porcine epidemic diarrhea virus (PEDV) can cause severe diarrhea death in newborn piglets, resulting in significant economic losses for the pig industry. Therefore, the advancement of safe and effective anti-PEDV drugs for the treatment of PEDV is of paramount importance. In this study, molecular docking was used to screen natural drugs that can target PEDV 3C like protease (3CLpro). As well, the anti-PEDV effects of the screened drugs were evaluated in vitro and in vivo. Molecular docking and molecular dynamics (MD) simulation results showed that ethyl caffeate (EC) could efficiently bind to the active cavity of PEDV 3CLpro. Biolayer interferometry (BLI) and fluorescence resonance energy transfer (FRET) analyses demonstrated that EC directly interacts with PEDV 3CLpro (K = 1650 μM) and inhibits the activity of 3CLpro (IC = 33.87 μM). EC has been shown to significantly inhibit the replication of PEDV in Vero E6 cells. The half maximal inhibitory concentration (CC) and half-effective concentration (EC) were determined to be 283.1 μM and 8.641 μM, respectively, yielding a selectivity index as high as 32.7. Furthermore, EC was evaluated using a piglet infection model for PEDV. It demonstrated the ability to inhibit PEDV infection in vivo and improve the survival rate of piglets (3/5, 60%). Compared to the control group, oral administration of EC significantly reduced intestinal pathological damage and viral load. Our study indicated that EC, targeting PEDV 3CLpro, is a safe and effective anti-PEDV drug with promising clinical application prospects.