Potent and Selective Inhibitors of CDPK1 from T. gondii and C. parvum Based on a 5-Aminopyrazole-4-carboxamide Scaffold

Abstract

5-Aminopyrazole-4-carboxamide was used as an alternative scaffold to substitute for the pyrazolopyrimidine of a known “bumped kinase inhibitor” to create selective inhibitors of calcium-dependent protein kinase-1 from both Toxoplasma gondii and Cryptosporidium parvum. Compounds with low nanomolar inhibitory potencies against the target enzymes were obtained. The most selective inhibitors also exhibited submicromolar activities in T. gondii cell proliferation assays and were shown to be non-toxic to mammalian cells.

In Vitro and In Vivo Effects of the Bumped Kinase Inhibitor 1294 in the Related Cyst-Forming Apicomplexans Toxoplasma gondii and Neospora caninum

“…Selective and potent inhibitors have been generated that take advantage of the enlarged space in the CDPK1 ATPbinding site (37)(38)(39). Benzoyl benzimidazole-based selective inhibitors and BKIs based on a pyrazolopyrimidine scaffold can block T. gondii CDPK1 and affect host cell invasion (40)(41)(42).…”

Novel Bumped Kinase Inhibitors Are Safe and Effective Therapeutics in the Calf Clinical Model for Cryptosporidiosis

“…The propagation of C. parvum (Iowa isolate) in calves and oocyst purification [34][35][36][37], synthesis of BKIs [22,25,27,38], in vitro CpCDPK1 enzyme assay [22,25,38], in vitro determination of C. parvum BKI sensitivity [22,25,38,39], neonatal mouse [36,40] and calf models of C. parvum infection [37,[41][42][43], and pharmacologic measurement of BKI plasma and stool levels and plasma protein binding have all been previously described [27,44]. BKI-1294 and BKI-1553 were synthesized on the pyrazolo [2,3-d] pyrimidine scaffold, while BKI-1517 was synthesized on a 5-aminopyrazole-4-carboxamide scaffold [25] (Figure 1).…”

“…BKI-1294 and BKI-1553 were synthesized on the pyrazolo [2,3-d] pyrimidine scaffold, while BKI-1517 was synthesized on a 5-aminopyrazole-4-carboxamide scaffold [25] (Figure 1). Previously published details of these methods are specified in the Supplementary Materials.…”

“…Most recently, the calcium-dependent protein kinase 1 of C. parvum (CpCDPK1) has shown promising potential as a drug target in treatment of cryptosporidiosis, as well as other diseases caused by apicomplexan parasites [22][23][24][25][26]. Apicomplexan CDPK homologs have crucial roles in host cell invasion, micronemal secretion, and gliding motility [23,[27][28][29][30].…”

“…BKIs project a bulky aromatic substituent (a "bump") from the C-3 position into the hydrophobic pocket next to the gatekeeper residue [22]. Large numbers of BKIs have been synthesized, differing mainly by R1 and R2 groups displayed from a few core scaffolds, which selectively target CDPK orthologs of multiple apicomplexan parasites [22,25,26,31]. Treatment with BKI-1294 reduced C. parvum infection in vitro and eliminated infection in 6 of 7 immunodeficient mice [23].…”