Bratton, S.M. and Mosher, C.M. and Khallouki, F. and Finel, M. and Court, M.H. and Moran, J.H. and Radominska-Pandya, A. (2012) Analysis of R- and S-hydroxywarfarin glucuronidation catalyzed by human liver microsomes and recombinant udp-glucuronosyltransferases. Journal of Pharmacology and Experimental Therapeutics, 340 (1). pp. 46-55.

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Abstract

Coumadin (R-, S-warfarin) is a challenging drug to accurately dose, both initially and for maintenance, because of its narrow therapeutic range and wide interpatient variability and is typically administered as a racemic (Rac) mixture, which complicates the biotransformation pathways. The goal of the current work was to identify the human UDP-glucuronosyltransferases (UGTs) involved in the glucuronidation of the separated R- and S-enantiomers of 6-, 7-, and 8-hydroxywarfarin and the possible interactions between these enantiomers. The kinetic and inhibition constants for human recombinant 1A family UGTs toward these separated enantiomers have been assessed using high-performance liquid chromatography (HPLC)-UV-visible analysis, and product confirmations have been made using HPLC-mass spectrometry/mass spectrometry. We found that separated R- and S-enantiomers of 6-, 7-, and 8-hydroxywarfarin demonstrate significantly different glucuronidation kinetics and can be mutually inhibitory. In some cases significant substrate inhibition was observed, as shown by K m, V max, and K i, comparisons. In particular, UGT1A1 and extrahepatic UGT1A10 have significantly higher capacities than other isoforms for S-7-hydroxywarfarin and R-7-hydroxywarfarin glucuronidation, respectively. Activity data generated using a set of well characterized human liver microsomes supported the recombinant enzyme data, suggesting an important (although not exclusive) role for UGT1A1 in glucuronidation of the main warfarin metabolites, including Rac-6- and 7-hydroxywarfarin and their R- and S-enantiomers in the liver. This is the first demonstration that the R- and S-enantiomers of hydroxywarfarins are glucuronidated, with significantly different enzymatic affinity and capacity, and supports the importance of UGT1A1 as the major hepatic isoform involved. Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics.

Item Type: Article
Uncontrolled Keywords: 6 hydroxywarfarin; 7 hydroxywarfarin; 8 hydroxywarfarin; drug metabolite; glucuronosyltransferase; glucuronosyltransferase 1A1; glucuronosyltransferase 1A10; isoenzyme; recombinant enzyme; unclassified drug; warfarin, article; catalysis; controlled study; drug glucuronidation; drug structure; enantiomer; enzyme activity; enzyme inhibition; enzyme kinetics; enzyme substrate; high performance liquid chromatography; human; human cell; liver microsome; mass spectrometry; priority journal; ultraviolet spectroscopy, Chromatography, High Pressure Liquid; Glucuronides; Glucuronosyltransferase; Humans; Kinetics; Microsomes, Liver; Recombinant Proteins; Spectrophotometry, Ultraviolet; Stereoisomerism; Tandem Mass Spectrometry; Tissue Banks; Warfarin
Subjects: Biochemistry, Genetics and Molecular Biology
Divisions: SCIENTIFIC PRODUCTION > Biochemistry, Genetics and Molecular Biology
Depositing User: Administrateur Eprints Administrateur Eprints
Last Modified: 31 Jan 2020 15:44
URI: http://eprints.umi.ac.ma/id/eprint/1576

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