Structure-guided screening of FAR-1 antagonists with multi-stage anthelmintic activity.

Fatty acid and retinol binding proteins (FARs) are nematode-specific proteins that orchestrate lipid metabolism, development, and host immune response. Here, the antagonists of Nippostrongylus brasiliensis FAR-1 (NbFAR-1) were identified through integrating virtual screening, fluorescent ligand binding assay, and in vitro egg hatching assays. An in vivo mouse model was employed to evaluate anthelmintic efficacy against intestine-parasitized N. brasiliensis and brain-parasitized Angiostrongylus cantonensis. Forty-eight candidates were selected by virtual screening, six of them showed more than 40% antagonism to NbFAR-1 by fluorescent ligand-competition binding assay and suppressed N. brasiliensis egg hatching by 40%-80% at 20 μM. In mice, E002-0872 and 4340-0245 reduced intestinal N. brasiliensis burdens by 61.74% and 62.15%, respectively, and ameliorated intestinal damages. 4340-0245 reduced cerebral A. cantonensis burdens by 52.73% and alleviated meningeal bleeding and neurological signs. Moreover, treatment with 4340-0245 at 15 mg/kg shortened the body length of female worms, consistent with the higher far-1 expression in females. Alanine scanning showed I95 of NbFAR-1 as a key residue for binding fatty acid, retinol, and 4340-0245. Intraperitoneal administration of 4340-0245 at 50 mg/kg did not cause any significant toxic effects, whereas 15 mg/kg resulted in a plasma Cmax of 56,473 ng/mL at 10 min and a half-life of ~7 h. These data provide evidence that FAR-1 is a promising target for developing anthelmintic drugs.