Structure-Based Design of Submicromolar, Biologically Active Inhibitors of Trypanosomatid Glyceraldehyde-3-Phosphate Dehydrogenase

• Published in: Proceedings of the National Academy of Sciences - PNAS 1999-04, Vol.96 (8), p.4273-4278
• Main Authors: Aronov, Alex M., Suresh, Stephen, Buckner, Frederick S., Van Voorhis, Wesley C., Christophe L. M. J. Verlinde, Opperdoes, Fred R., Wim G. J. Hol, Gelb, Michael H.
• Format: Article
• Language: English
• Subjects: 3T3 Cells; Adenosine - analogs & derivatives; Adenosine - chemistry; Adenosine - pharmacology; Amastigotes; Animals; Binding Sites; Biochemistry; Biological Sciences; Dehydrogenases; Drug Design; Enzyme Inhibitors - chemical synthesis; Enzyme Inhibitors - chemistry; Enzyme Inhibitors - pharmacology; Enzymes; Fibroblasts; Glyceraldehyde-3-Phosphate Dehydrogenases - antagonists & inhibitors; Glyceraldehyde-3-Phosphate Dehydrogenases - chemistry; Glycolysis; Glycolysis - drug effects; Humans; Inhibitory concentration 50; Lead compounds; Leishmania mexicana - drug effects; Leishmania mexicana - enzymology; Leishmania mexicana - growth & development; Mice; Models, Molecular; Molecular Conformation; Molecular Sequence Data; Parasite hosts; Parasites; Protein Conformation; Trypanocidal Agents - chemical synthesis; Trypanocidal Agents - chemistry; Trypanocidal Agents - pharmacology; Trypanosoma; Trypanosoma brucei brucei - drug effects; Trypanosoma brucei brucei - enzymology; Trypanosoma brucei brucei - growth & development
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.96.8.4273

The bloodstream stage of Trypanosoma brucei and probably the intracellular (amastigote) stage of Trypanosoma cruzi derive all of their energy from glycolysis. Inhibiting glycolytic enzymes may be a novel approach for the development of antitrypanosomatid drugs provided that sufficient parasite versus host selectivity can be obtained. Guided by the crystal structures of human, T. brucei, and Leishmania mexicana glyceraldehyde-3-phosphate dehydrogenase, we designed adenosine analogs as tight binding inhibitors that occupy the pocket on the enzyme that accommodates the adenosyl moiety of the NAD+cosubstrate. Although adenosine is a very poor inhibitor, IC50≈ 50 mM, addition of substituents to the 2′position of ribose and the N6-position of adenosine led to disubstituted nucleosides with micromolar to submicromolar potency in glyceraldehyde-3-phosphate dehydrogenase assays, an improvement of 5 orders of magnitude over the lead. The designed compounds do not inhibit the human glycolytic enzyme when tested up to their solubility limit (≈ 40 μ M). When tested against cultured bloodstream T. brucei and intracellular T. cruzi,$N^{6}\text{-(1-naphthalenemethyl)-2}^{\prime}\text{-(3-chlorobenzam ido)adenosine}$inhibited growth in the low micromolar range. Within minutes after adding this compound to bloodstream T. brucei, production of glucose-derived pyruvate ceased, parasite motility was lost, and a mixture of grossly deformed and lysed parasites was observed. These studies underscore the feasibility of using structure-based drug design to transform a mediocre lead compound into a potent enzyme inhibitor. They also suggest that energy production can be blocked in trypanosomatids with a tight binding competitive inhibitor of an enzyme in the glycolytic pathway.