Inhibitors of Purine Import into Plasmodium falciparum Kill Malaria Parasites

? DESCRIPTION (provided by applicant): Infection with unicellular eukaryotic Plasmodium species parasites causes malaria. P. falciparum causes the most virulent form of malaria. Currently, artemisinin combination therapy (ACT) is the treatment of choice for infected individuals. The rise of artemisinin resistant P. falciparum in Southeast Asia makes it imperative to develop new antimalarial drugs. Malaria parasites are purine auxotrophs. They transport purine precursors from the host erythrocyte into the parasite via the P. falciparum Equilibrative Nucleoside Transporter 1 (PfENT1). In the parasite, purine salvage pathway enzymes modify the purine precursors to form the nucleotides needed for RNA and DNA synthesis and other cellular processes. At purine concentrations found in human blood ( 12, and the Z' score > 0.80, indicating a highly robust assay. We screened a 64,500 compound library and identified 171 hits. We tested nine of the top hits in a series of secondary assays. All nine inhibited [3H]adenosine uptake into both PfENT1-expressing yeast and into erythrocyte-free trophozoite stage P. falciparum with IC50 values in the 2 - 40 nM range. The nine compounds, five distinct chemical scaffolds, do not kill yeast but do kill P. falciparum parasites in culture with IC50 values in the 5 - 55 µM range. The goals of this application are 1) to improve the potency and selectivity of the PfENT1 inhibitors through medicinal chemistry; 2) to define the mechanism of action of the inhibitors and their impact on parasite biology and growth at various life cycle stages; 3) to test the efficacy of the inhibitorsin a mouse malaria model; and 4) to identify the inhibitor binding site and the conformation of PfENT1 to which the inhibitors bind. Successful completion of this project will determine the utility of targeting PfENT1 for antimalarial drug development and may identify compounds suitable for further development.