Drug-S-Acyl-Glutathione Thioesters: Synthesis, Bioanalytical Properties, Chemical Reactivity, Biological Formation and Degradation

Mark      P. Grillo

Abstract

Carboxylic acid-containing drugs can be metabolized to chemically-reactive acyl glucuronide, S-acyl-CoA thioester, and/or intermediate acyl-adenylate metabolites that are capable of transacylating the cysteinyl-thiol of glutathione (GSH) resulting in the formation of drug-S-acyl-GSH thioesters detected in vivo in bile and in vitro in hepatocytes. Authentic S-acyl-GSH thioesters of carboxylic acids can be readily synthesized by modifying the cysteinyl-thiol group of GSH with an applicable acylating reagent. Bionanalytical characterization of S-acyl-GSH derivatives has demonstrated enhanced extraction efficiency from biological samples when formic acid is included in appropriate extraction solvents, and that tandem mass spectrometry of S-acyl-GSH conjugates results in fragmentation producing a common MH+-147 Da product ion. Chemical reactivity comparisons have shown that S-acyl-CoA thioester and acyl-adenylate conjugates are more reactive than their corresponding 1-β-O-acyl glucuronides toward the transacylation of GSH forming S-acyl-GSH thioesters. S-Acyl-GSH thioester derivatives are also chemically-reactive electrophiles capable of transacylating biological nucleophiles. Glutathione S-transferases (GSTs) weakly catalyze S-acyl-GSH conjugate formation from S-acyl-CoA, acyl-adenylate, and 1-β-O-acyl glucuronide substrates; however purified-GSTs have also been shown to hydrolyze S-acyl-GSH thioesters. Mechanistic in vitro studies in hepatocytes have revealed the primary importance of the S-acyl-CoA formation pathway leading to S-acyl-GSH-adduct formation. In addition to being hydrolytically-unstable in hepatocytes and plasma, S-acyl-GSH thioesters undergo γ-glutamyltranspeptidase-mediated cleavage of the γ-glutamyl-group leading to N-acyl-cysteinylglycine amide-linked products. In summary, S-acyl-GSH thioesters are indicators of reactive transacylating metabolite formation produced from the biotransformation of carboxylic acids, but since they are also chemically-reactive, perhaps these derivatives can contribute to covalent binding to tissue proteins and potential toxicity.

Keywords: Acyl-adenylate, acyl glucuronide, bioactivation, γ-glutamyltranspeptidase, glutathione, S-acyl-CoA, S-acyl-glutathione, transacylation, ibufenac, arachidonic acids