Abstract
PfCDPK1 is a Plasmodium falciparum calcium-dependent protein kinase, which has been identified as a potential target for novel antimalarial chemotherapeutics. In order to further investigate the role of PfCDPK1, we established a high-throughput in vitro biochemical assay and used it to screen a library of over 35,000 small molecules. Five chemical series of inhibitors were initially identified from the screen, from which series 1 and 2 were selected for chemical optimization. Indicative of their mechanism of action, enzyme inhibition by these compounds was found to be sensitive to both the ATP concentration and substitution of the amino acid residue present at the “gatekeeper” position at the ATP-binding site of the enzyme. Medicinal chemistry efforts led to a series of PfCDPK1 inhibitors with 50% inhibitory concentrations (IC50s) below 10 nM against PfCDPK1 in a biochemical assay and 50% effective concentrations (EC50s) less than 100 nM for inhibition of parasite growth in vitro. Potent inhibition was combined with acceptable absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties and equipotent inhibition of Plasmodium vivax CDPK1. However, we were unable to correlate biochemical inhibition with parasite growth inhibition for this series overall. Inhibition of Plasmodium berghei CDPK1 correlated well with PfCDPK1 inhibition, enabling progression of a set of compounds to in vivo evaluation in the P. berghei rodent model for malaria. These chemical series have potential for further development as inhibitors of CDPK1.
Highlights
PfCDPK1 is a Plasmodium falciparum calcium-dependent protein kinase, which has been identified as a potential target for novel antimalarial chemotherapeutics
PfCDPK1 has been identified as a potential therapeutic target for intervention in malaria, and we set out to find novel inhibitors of PfCDPK1 using a high-throughput biochemical assay to screen a library of more than 35,000 small molecules
The physiological relevance of this activity for PfCDPK1 is unknown; ATP hydrolysis by isolated kinases is not uncommon and is considered to be a function of the ATP-binding site, which can be exploited for identification of inhibitors [27]
Summary
PfCDPK1 is a Plasmodium falciparum calcium-dependent protein kinase, which has been identified as a potential target for novel antimalarial chemotherapeutics. Inhibition of Plasmodium berghei CDPK1 correlated well with PfCDPK1 inhibition, enabling progression of a set of compounds to in vivo evaluation in the P. berghei rodent model for malaria These chemical series have potential for further development as inhibitors of CDPK1. The P. falciparum genome contains five genes encoding canonical CDPKs, and they have been implicated in a range of biological processes at different stages of the parasite life cycle [9] The fact that these enzymes are absent from the vertebrate hosts of these parasites suggests that they may represent useful targets for the development of antimicrobial agents. P. falciparum calcium-dependent protein kinase 1 (PfCDPK1) has been shown to phosphorylate MTIP and GAP45 in vitro, it remains to be established that these proteins are its principal substrates in vivo [13]
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