Long Chain Fatty Acyl-Coenzyme A Synthetase as Novel Drug Targets in Cryptosporidium parvum

Abstract

Cryptosporidium parvum infects both humans and animals, and continues to be a significant opportunistic pathogen among AIDS patients and one of the leading diarrheal pathogens in children. Despite decades of research on cryptosporidiosis including screening hundreds of compounds in vitro and in vivo, fully effective therapeutic agents are still unavailable. The major goal of this study is to explore three long-chain fatty acyl-CoA synthetase (CpACSs) of the parasite as novel drug targets. The molecular and biological data showed that the three CpACSs genes are differently expressed in the parasite different life cycle stages and their proteins display different subcellular distribution patterns in the parasite, suggesting that the those genes may play different biological roles in the parasite. Using recombinant proteins, we have determined detailed enzyme kinetics for CpACS1 and CpACS2, and observed that the inhibitor triacsin C could inhibit their enzyme activities with Ki in the nanomolar range. Triacsin C also effectively inhibited the growth of C. parvum parasites in vitro (IC_(50) = 136 nM). Most importantly, triacsin C effectively reduced parasite oocyst production up to 88.1% with no apparent toxicity when administered to Cryptosporidium-infected IL-12 knock-out mice at 15 mg/kg/d for one week. These observations not only validate CpACSs as a pharmacological target, but also indicate that triacsin C and analogs may be explored as potential new therapeutics against cryptosporidiosis.