Icariside I inhibits KSHV lytic replication via targeting cellular PPAT

Kaposi’s sarcoma-associated herpesvirus (KSHV) causes malignancies, including Kaposi’s sarcoma and primary effusion lymphoma, but effective therapies are lacking. High-throughput screening of natural compounds identified icariside I (ICSI), a flavonoid from Epimedii Folium , as a potent inhibitor of KSHV lytic replication in both epithelial and endothelial cells. Mechanistically, ICSI directly targets host amidophosphoribosyltransferase (PPAT), a key enzyme in de novo purine synthesis. Drug affinity, responsive target stability and surface plasmon resonance assays confirmed ICSI–PPAT binding dependent on residues Glu124 and Asp389. Genetic PPAT knockout or pharmacological inhibition with 6-mercaptopurine reduced lytic reactivation, phenocopying ICSI, while supplementation with the purine precursor hypoxanthine restored viral replication under ICSI treatment or PPAT loss. These results reveal that ICSI inhibits KSHV lytic replication by disrupting host purine biosynthesis via PPAT inhibition, highlighting PPAT as a host-directed antiviral target and suggesting ICSI or PPAT inhibitors as potential therapies for KSHV-associated malignancies.