Developing novel lead compounds for kynurenine-3-monooxygenase inhibition

Kynurenine Monooxygenase (KMO) is a potential drug target for neurodegenerative diseases such as Alzheimer’s disease^1-3. In this study, we tested kynurenine analogs and sulfonylureas predicted by a pharmacophore model⁴ for competitive inhibition of KMO. Given the therapeutic relevance of KMO inhibition, and that obtaining a pure recombinant human KMO is still a challenge, our lab seeks to crystallize Cytophaga hutchinsonii (ch) KMO and hopefully provide a better surrogate for human KMO. Our kinetic data using UV-vis spectrophotometry, HPLC, and fluorometry shows that kynurenine analogs with substituted aromatic ring and three spacers atoms between benzene ring and carboxylate with or without the amino group on α-carbon are competitive inhibitors of KMO with Ki and Kd values in the low micromolar range. We also observed that compounds with a deactivating group on the para position on the benzene ring consume NADPH without hydroxylation, as expected for non-substrate effectors of KMO; this phenomenon leads to accumulation of hydrogen peroxide, a potential life-threatening side effect⁵. We have docked all our lead compound using Saccharomyces cerevisiae KMO from structure PDB ID 4J36⁵; the docking results shows that indeed the carboxylate moiety or its bioisosteres are needed for binding in the active site, which ultimately validates our pharmacophore. The ongoing study focuses on looking at how these ligands bind using x-ray crystallography. We have already identified potential conditions in which chKMO crystallizes, and we are currently optimizing those conditions in order to obtain bigger and well-ordered crystals that will diffract.

References

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5. Amaral M, Levy C, Heyes DJ, et al. Nature. 2013; 496:382.