Cholesterol synthesis inhibitors effectively inhibit the replication of African swine fever virus in vitro.

The devastating impact of African swine fever (ASF) on the global pig industry has prompted an urgent need for the development of novel antiviral agents; however, no safe and effective vaccines are currently available. Cholesterol metabolism has emerged as a critical host pathway exploited by various viruses, but whether cholesterol synthesis inhibitors exert antiviral activity against ASFV is unknown. In this study, we evaluated the effects of two cholesterol biosynthesis inhibitors, AM580 and simvastatin, on ASFV replication in porcine alveolar macrophages (PAMs). We found that both inhibitors impaired ASFV replication in a dose-dependent manner in vitro. Neither compound affected viral entry. Instead, both significantly inhibited ASFV replication at post-entry stages of the viral life cycle, as demonstrated by time-of-addition assays. Notably, direct treatment of ASFV particles with either inhibitor did not reduce viral infectivity, indicating that the inhibitors do not act through virucidal mechanisms. Furthermore, exogenous cholesterol supplementation reversed the inhibitory effects of AM580 and simvastatin, suggesting that their antiviral activity is mediated through modulation of intracellular cholesterol levels. Taken together, our findings demonstrate that cholesterol synthesis inhibitors restrict ASFV replication in vitro by targeting host cholesterol metabolism in macrophages. Our findings highlight the critical role of macrophage cholesterol metabolism in ASFV replication and support the repurposing of cholesterol synthesis inhibitors as a promising host-directed antiviral strategy against ASFV.