title: Substrate and Inhibitor Specificity of the Plasmodium berghei Equilibrative Nucleoside Transporter Type 1.
abstract: Malaria is a critical public health issue in the tropical world, causing extensive morbidity and mortality. Infection by unicellular, obligate intracellular Plasmodium parasites causes malaria. The emergence of resistance to current antimalarial drugs necessitates the development of novel therapeutics. A potential novel drug target is the purine import transporter. Because Plasmodium parasites are purine auxotrophic, they must import purines from their host to fulfill metabolic requirements. They import purines via equilibrative nucleoside transporter 1 (ENT1) homologs. Recently, we used a yeast-based high-throughput screen to identify inhibitors of the P. falciparum ENT1 (PfENT1) that kill P. falciparum parasites in culture. P. berghei infection of mice is an animal model for human malaria. Because P. berghei ENT1 (PbENT1) shares only 60% amino acid sequence identity with PfENT1, we sought to characterize PbENT1 and its sensitivity to our PfENT1 inhibitors. We expressed PbENT1 in purine auxotrophic yeast and used radiolabeled substrate uptake to characterize its function. We showed that PbENT1 transports both purines and pyrimidines. It preferred nucleosides compared with nucleobases. Inosine (IC50 = 3.7 µM) and guanosine (IC50 = 21.3 µM) had the highest affinities. Our recently discovered PfENT1 inhibitors were equally effective against both PbENT1- and PfENT1-mediated purine uptake. The PfENT1 inhibitors are at least 10-fold more potent against PfENT1 than human hENT1. They kill P. berghei parasites in 24-hour ex vivo culture. Thus, the P. berghei murine malaria model may be useful to evaluate the efficacy of PfENT1 inhibitors in vivo and their therapeutic potential for treatment of malaria.
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York.
authors.forename: Avish
authors.initials: A
authors.lastname: Arora
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York.
authors.forename: Roman
authors.initials: R
authors.lastname: Deniskin
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York.
authors.forename: Yvett
authors.initials: Y
authors.lastname: Sosa
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York.
authors.forename: Sita Nirupama
authors.initials: SN
authors.lastname: Nishtala
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York.
authors.forename: Philipp P
authors.initials: PP
authors.lastname: Henrich
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York.
authors.forename: T R Santha
authors.initials: TR
authors.lastname: Kumar
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York.
authors.forename: David A
authors.initials: DA
authors.lastname: Fidock
authors.affiliation: Departments of Physiology and Biophysics (A.A., R.D., Y.S., S.N.N., M.H.A.) and Neuroscience and Medicine (M.H.A.), Albert Einstein College of Medicine, Bronx, New York; and Departments Microbiology and Immunology (P.P.H., T.R.S.K., D.A.F.) and Medicine (D.A.F.), Columbia University Medical Center, New York, New York myles.akabas@einstein.yu.edu.
authors.forename: Myles H
authors.initials: MH
authors.lastname: Akabas
mesh_terms: D000241:Adenosine / Q000378:metabolism; D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D033721:Equilibrative Nucleoside Transporter 1 / Q000378:metabolism*; D005260:Female; D006801:Humans; D020128:Inhibitory Concentration 50; D051379:Mice; D010962:Plasmodium berghei / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D013379:Substrate Specificity / Q000187:drug effects; D013997:Time Factors; D014316:Tritium / Q000378:metabolism; D014529:Uridine / Q000378:metabolism
chemical_list: D000962:Antimalarials; D033721:Equilibrative Nucleoside Transporter 1; D014316:Tritium; D000241:Adenosine; D014529:Uridine