Identification of potent and orally efficacious phosphodiesterase inhibitors in Cryptosporidium parvum-infected immunocompromised mice

Cryptosporidium species, mostly C. parvum and C. hominis in humans, are intestinal apicomplexan parasites that cause life-threatening diarrhea in young children and people with cell-mediated immune defects, such as due to AIDS. There is only one approved treatment for cryptosporidiosis, but it is ineffective for immunocompromised people and only modestly effective for children. In this study, screening 278 compounds from the Merck KGaA, Darmstadt, Germany collection and accelerated follow-up work enabled by prior investigation of the compounds resulted in identification of a series of pyrazolopyrimidine human phosphodiesterase (PDE)-V inhibitors with potent anticryptosporidial activity and efficacy following oral administration in C. parvum-infected immunocompromised mice. The novel PDE inhibitor leads (compounds PDEi2 and PDEi5) affect parasite egress from infected host cells. They have comparable activity against C. parvum and C. hominis, rapidly eliminate C. parvum in tissue culture, and have minimal off-target effects in a panel of safety screening assays. In comparison, the potent human PDE-V inhibitors sildenafil and the 4-aminoquinoline compound 7a have no useful activity against C. parvum. Based on homology modeling and in silico compound docking, PDEi5 interacts directly with an active-site metal ion and docks well to two C. parvum PDEs. In contrast, larger amino acid side groups (Val900/Tyr11128 and His884/Asn1112) in both C. parvum PDEs replace alanine in human PDE-V and block sildenafil binding, explaining its lack of efficacy. These results identify a promising new drug target and lead series for anticryptosporidial drug development and validates a route to target-based optimization.