title: Cysteine amide adduct formation from carboxylic acid drugs via UGT-mediated bioactivation in human liver microsomes.
abstract: Although chemical trapping has been widely used to evaluate cytochrome P450-mediated drug bioactivation, thus far, only a few in vitro-trapping studies have been performed on UDP-glucuronosyltransferase (UGT)-mediated drug bioactivation. In this study, we used cysteine (Cys) as trapping agent to gain new insights into the UGT-mediated bioactivation involving acyl glucuronides of carboxylic acid drugs. Diclofenac, ketoprofen and ibuprofen were incubated in human liver microsomes with UDPGA and Cys, followed by analysis using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF/MS). The N-acyl-Cys amide adduct of diclofenac was characterized by mass analysis and was detectable even in photodiode array analysis. Our data indicated that the formation of such adducts reflects the reactivity of the corresponding acyl glucuronides. In addition, it was suggested that the adduct formation requires an N-terminal Cys moiety with both a free amine and a free thiol group, from the results using various cysteine derivatives. We propose that the S-acyl-Cys thioester adduct can form via transacylation of an acyl glucuronide and can then form to an N-acyl-Cys amide adduct through intramolecular S- to N-acyl rearrangement. This series of the reactions has important implications as a possible bioactivation mechanism for covalent binding of carboxylic acid drugs to macromolecules.
authors.forename: H
authors.initials: H
authors.lastname: Harada
authors.forename: Y
authors.initials: Y
authors.lastname: Toyoda
authors.forename: T
authors.initials: T
authors.lastname: Endo
authors.forename: M
authors.initials: M
authors.lastname: Kobayashi
mesh_terms: D000577:Amides / Q000378:metabolism; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000378:metabolism; D002264:Carboxylic Acids / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003545:Cysteine / Q000378:metabolism*; D004008:Diclofenac / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000378:metabolism*; D006801:Humans; D007052:Ibuprofen / Q000378:metabolism; D066298:In Vitro Techniques; D007660:Ketoprofen / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D000577:Amides; D000894:Anti-Inflammatory Agents, Non-Steroidal; D002264:Carboxylic Acids; D004008:Diclofenac; D007660:Ketoprofen; D014453:Glucuronosyltransferase; D003545:Cysteine; D007052:Ibuprofen