Luteolin exhibits broad-spectrum antiviral activity against swine enteric coronaviruses by targeting 3CLpro

Swine enteric coronaviruses (SECoVs), including porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), and porcine deltacoronavirus (PDCoV), pose a major threat to global swine production, with current control measures remaining suboptimal. These infections are often characterized by mixed infections, highlighting the urgent need for anti-SECoV drugs. This study elucidates the potential of luteolin as a broad-spectrum antiviral agent targeting the 3 C like protease (3CLpro). Molecular docking and dynamics simulations indicated a stable and high-affinity interaction between luteolin and PEDV 3CLpro, which was confirmed by molecular layer interferometry, showing direct binding with a high affinity (KD = 236 nM). In vitro, luteolin significantly inhibited 3CLpro enzymatic activity (IC₅₀ = 18.74 µM) and potently suppressed PEDV replication. In a piglet model, both prophylactic and therapeutic administration of luteolin achieved a 75 % protection rate (3/4), significantly alleviated diarrhea, mitigated intestinal tissue damage, and reduced the viral load in the jejunum and cecum following PEDV challenge. Furthermore, luteolin exhibited direct binding to TGEV and PDCoV 3CLpro with high affinities (KD = 342 nM and 282 nM, respectively). Consistent with this, it demonstrated broad-spectrum antiviral activity in vitro, the half-effective concentrations (EC₅₀) for TGEV and PDCoV were 13.14 μM and 4.10 μM, respectively. Collectively, our findings not only validate luteolin as an effective PEDV inhibitor but also highlight its promise as a broad-spectrum agent against SECoVs, providing a crucial foundation for developing novel anti-coronavirus therapeutics.