The Identification of Nanomolar 1,4-benzoquinone Inhibitors of Shikimate Kinase.

K. REYNOLDS1, G.S. COCKERILL1, D. MADGE2, V. DOWDELL1, C. VISINTIN2, S. CHAPMAN1, A. MORRISON2, D. RUPASSARA1, J. STABLES1, K. POWELL1, I. CHARLES1, A. HAWKINS3

Author address: 

1 Arrow Therapeutics, London, United Kingdom, 2 University College London, London, United Kingdom, 3 University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom

Abstract: 

Background: The shikimate pathway is the seven-step biosynthetic route which generates chorismic acid from phosphoenolpyruvate and erythrose 4-phosphate and is crucial for aromatic amino acid synthesis. The pathway is essential for plants and micro-organisms, but is absent from animals making the enzymes important targets for the development of potentially non-toxic antimicrobial agents and herbicides. Shikimate kinase, the fifth enzyme of the pathway, catalyses the specific phosphorylation of the 3-hydroxyl group of shikimic acid using ATP as a co-substrate. Methods: A three-enzyme linked assay employing shikimate kinase (AroL, Aspergillis), pyruvate kinase and lactate dehydrogenase has been used to identify potent inhibitors. Results: Screening of in-house compound collections identified A-00000764 to be a potent inhibitor of shikimate kinase (AroL, 500nM). Chemical exploration of the template identified substitution of 2-phenyl ring to provide compounds with improved activity vs SK (A-00026158, 40nM). The toxicity (TD50 in Vero cells) could be modified by ortho-substitution of this ring system whilst maintaining activity for SK (A-00026161; IC50 200nM, TD50(Vero) 100µM). Compounds from the class also displayed activity vs Staphylococcus aureus (A-00026161, MIC 12µM). Experiments indicated members of the series (A-00000762, IC50 100nM) to be competitive inhibitors with respect to ATP. Conclusions: The data demonstrates that potent, ATP competitive inhibitors can be identified using Arrow's assay and HTS methodology. The assay is being used for the identification of further inhibitor classes of shikimate kinase.
2002

abstract No: 

F-745

Full conference title: 

42nd ICAAC Abstracts, American Society for Microbiology, September 27 - 30, 2002, San Diego, CA
    • ICAAC 42nd