Zika virus (ZIKV) has become a public health threat due to its global transmission and link to severe congenital disorders. The host immune responses to ZIKV infection have not been fully elucidated, and effective therapeutics are not currently available. Here, we identified a molecule called 25-hydroxycholesterol (25HC) that is produced by cells in response to ZIKV infection. While the endogenous production of 25HC does not suffice to ward off ZIKV, we found that augmentation with synthetically derived 25HC can provide significant protective benefits, blocking viral entry in cultured cells , and infection of both mice and rhesus macaques exposed to ZIKV. Moreover, we found that 25HC administration can markedly reduce ZIKV-associated neural tissue damage in human cortical organoids/mini-brains and in developing micel. Collectively, our findings highlight the protective role of 25HC as a natural antiviral agent to combat ZIKV infection and prevent ZIKV-associated outcomes, such as fetal brain damage.