Structure-Activity Relationship Study of Anti-Cryptosporidium Benzoxaboroles Yields Enhanced Potency and Curative Efficacy.

Cryptosporidiosis is a major cause of life-threatening diarrhea in children and chronic diarrhea in immunocompromised individuals. Our previous work identified pyrazolo[3,4-d]pyrimidine benzoxaborole (2) as an orally efficacious inhibitor of Cryptosporidium with a downside of relapse after 7 days post drug treatment. Here, we report a structure-activity relationship study around the benzoxaborole ring of compound 2, resulting in rac- 11 with improved in vitro potency (EC50 = 0.025 μM). Remarkably, rac- 11 is noncytotoxic and displayed curative in vivo efficacy in Cryptosporidium-infected immunocompromised NSG mice. Furthermore, resolution of rac-11 into its enantiomers demonstrated that one enantiomer is 2-fold more potent (ent2-11; EC50 = 0.011 μM), whereas the other is 400-fold less potent, highlighting the importance of the methyl group at the C3 position of benzoxaborole. The ent2-11 also had reduced adverse binding to hPDE5 compared to 2. Collectively, these findings advance new and effective candidates for the treatment of cryptosporidiosis.