Yeast-based high-throughput screen identifies plasmodium falciparum equilibrative nucleoside transporter 1 inhibitors that kill malaria parasites

I. J. Frame, Roman Deniskin, Alison Rinderspacher, Francine Katz, Shi Xian Deng, Robyn D. Moir, Sophie H. Adjalley, Olivia Coburn-Flynn, David A. Fidock, Ian M. Willis, Donald W. Landry, Myles Akabas

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Equilibrative transporters are potential drug targets; however, most functional assays involve radioactive substrate uptake that is unsuitable for high-throughput screens (HTS). We developed a robust yeast-based growth assay that is potentially applicable to many equilibrative transporters. As proof of principle, we applied our approach to Equilibrative Nucleoside Transporter 1 of the malarial parasite Plasmodium falciparum (PfENT1). PfENT1 inhibitors might serve as novel antimalarial drugs since PfENT1-mediated purine import is essential for parasite proliferation. To identify PfENT1 inhibitors, we screened 64560 compounds and identified 171 by their ability to rescue the growth of PfENT1-expressing fui1Δ yeast in the presence of a cytotoxic PfENT1 substrate, 5-fluorouridine (5-FUrd). In secondary assays, nine of the highest activity compounds inhibited PfENT1-dependent growth of a purine auxotrophic yeast strain with adenosine as the sole purine source (IC50 0.2-2 μM). These nine compounds completely blocked [3H]adenosine uptake into PfENT1-expressing yeast and erythrocyte-free trophozoite-stage parasites (IC50 5-50 nM), and inhibited chloroquine-sensitive and -resistant parasite proliferation (IC50 5-50 μM). Wild-type (WT) parasite IC50 values were up to 4-fold lower compared to PfENT1-knockout (pfent1Δ) parasites. pfent1Δ parasite killing showed a delayed-death phenotype not observed with WT. We infer that, in parasites, the compounds inhibit both PfENT1 and a secondary target with similar efficacy. The secondary target identity is unknown, but its existence may reduce the likelihood of parasites developing resistance to PfENT1 inhibitors. Our data support the hypothesis that blocking purine transport through PfENT1 may be a novel and compelling approach for antimalarial drug development.

Original languageEnglish (US)
Pages (from-to)775-783
Number of pages9
JournalACS Chemical Biology
Volume10
Issue number3
DOIs
StatePublished - Mar 20 2015

Fingerprint

Equilibrative Nucleoside Transporter 1
Plasmodium falciparum
Yeast
Malaria
Parasites
Yeasts
Throughput
Inhibitory Concentration 50
Assays
Antimalarials
Adenosine
Growth
Trophozoites
Chloroquine
Substrates
Erythrocytes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine

Cite this

Yeast-based high-throughput screen identifies plasmodium falciparum equilibrative nucleoside transporter 1 inhibitors that kill malaria parasites. / Frame, I. J.; Deniskin, Roman; Rinderspacher, Alison; Katz, Francine; Deng, Shi Xian; Moir, Robyn D.; Adjalley, Sophie H.; Coburn-Flynn, Olivia; Fidock, David A.; Willis, Ian M.; Landry, Donald W.; Akabas, Myles.

In: ACS Chemical Biology, Vol. 10, No. 3, 20.03.2015, p. 775-783.

Research output: Contribution to journalArticle

Frame, IJ, Deniskin, R, Rinderspacher, A, Katz, F, Deng, SX, Moir, RD, Adjalley, SH, Coburn-Flynn, O, Fidock, DA, Willis, IM, Landry, DW & Akabas, M 2015, 'Yeast-based high-throughput screen identifies plasmodium falciparum equilibrative nucleoside transporter 1 inhibitors that kill malaria parasites', ACS Chemical Biology, vol. 10, no. 3, pp. 775-783. https://doi.org/10.1021/cb500981y
Frame, I. J. ; Deniskin, Roman ; Rinderspacher, Alison ; Katz, Francine ; Deng, Shi Xian ; Moir, Robyn D. ; Adjalley, Sophie H. ; Coburn-Flynn, Olivia ; Fidock, David A. ; Willis, Ian M. ; Landry, Donald W. ; Akabas, Myles. / Yeast-based high-throughput screen identifies plasmodium falciparum equilibrative nucleoside transporter 1 inhibitors that kill malaria parasites. In: ACS Chemical Biology. 2015 ; Vol. 10, No. 3. pp. 775-783.
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AU - Katz, Francine

AU - Deng, Shi Xian

AU - Moir, Robyn D.

AU - Adjalley, Sophie H.

AU - Coburn-Flynn, Olivia

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AU - Willis, Ian M.

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AU - Akabas, Myles

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