A target-based high throughput screen yields Trypanosoma brucei hexokinase small molecule inhibitors with antiparasitic activity.
BACKGROUND
The parasitic protozoan Trypanosoma brucei utilizes glycolysis exclusively for ATP production during infection of the mammalian host. The first step in this metabolic pathway is mediated by hexokinase (TbHK), an enzyme essential to the parasite that transfers the gamma-phospho of ATP to a hexose. Here we describe the identification and confirmation of novel small molecule inhibitors of bacterially expressed TbHK1, one of two TbHKs expressed by T. brucei, using a high throughput screening assay.


RESULTS
Exploiting optimized high throughput screening assay procedures, we interrogated 220,233 unique compounds and identified 239 active compounds from which ten small molecules were further characterized. Computation chemical cluster analyses indicated that six compounds were structurally related while the remaining four compounds were classified as unrelated or singletons. All ten compounds were approximately 20-17,000-fold more potent than lonidamine, a previously identified TbHK1 inhibitor. Seven compounds inhibited T. brucei blood stage form parasite growth (0.03<or=EC(50)<3 microM) with parasite specificity of the compounds being demonstrated using insect stage T. brucei parasites, Leishmania promastigotes, and mammalian cell lines. Analysis of two structurally related compounds, ebselen and SID 17387000, revealed that both were mixed inhibitors of TbHK1 with respect to ATP. Additionally, both compounds inhibited parasite lysate-derived HK activity. None of the compounds displayed structural similarity to known hexokinase inhibitors or human African trypanosomiasis therapeutics.


CONCLUSIONS
The novel chemotypes identified here could represent leads for future therapeutic development against the African trypanosome.
Sharlow
Elizabeth R
ER
University of Pittsburgh Drug Discovery Institute and Pittsburgh Molecular Libraries Screening Center, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
Lyda
Todd A
TA
Dodson
Heidi C
HC
Mustata
Gabriela
G
Morris
Meredith T
MT
Leimgruber
Stephanie S
SS
Lee
Kuo-Hsiung
KH
Kashiwada
Yoshiki
Y
Close
David
D
Lazo
John S
JS
Morris
James C
JC
D000977:Antiparasitic Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D006593:Hexokinase / Q000037:antagonists & inhibitors*; D006801:Humans; D015800:Protozoan Proteins / Q000037:antagonists & inhibitors*; D014346:Trypanosoma brucei brucei / Q000187:drug effects* / Q000201:enzymology*
D000977:Antiparasitic Agents; D004791:Enzyme Inhibitors; D015800:Protozoan Proteins; D006593:Hexokinase
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