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

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.

Metabolic Activation of Mefenamic Acid Leading to Mefenamyl-S-Acyl-Glutathione Adduct Formation In Vitro and In Vivo in Rat

“…This concentration was much lower than the C max observed for S-acyl-CoA metabolites of (R)-ibuprofen (2600 nM; Grillo and Hua, 2008) and phenylacetic acid (1300 nM; Grillo and Lohr, 2009) measured under similar incubation conditions with rat hepatocytes; however, it was similar to the C max of flunoxaprofen-S-acyl-CoA (42 nM) detected in recent studies with rat hepatocytes incubated with 100 M (R)-flunoxaprofen . In addition, the length of incubation time leading to C max (T max ) of MFA-SCoA was 30 min in the present studies, whereas the reported T max of formation of ISCoA, phenylacetyl-S-acyl-CoA, and flunoxaprofen-S-acyl-CoA was 3-to 7.5-fold shorter at 10, 4, and 6 min, respectively (Grillo, 2011). The formation of MFA-SG thioester in rat hepatocyte incubations with MFA (100 M) was more rapid than MFA-SCoA formation and reached a 22-fold higher C max of 330 Ϯ 22 nM at the 10-min time point (Fig.…”

“…In addition to reactive acyl glucuronides, thioester-linked acylCoA derivatives ( Fig. 1) are also electrophilic and can transacylate biological nucleophiles, including protein and GSH, forming S-acyl-GSH thioester adducts (Boelsterli, 2002;Li et al, 2002;Skonberg et al, 2008;Grillo, 2011). Before the present studies, experiments on the detection of S-acyl-CoA and S-acyl-GSH metabolites formed in vivo or in vitro of fenamic acid-type NSAIDs, including MFA, have not been performed.…”

Interaction ofγ-Glutamyltranspeptidase with Ibuprofen-S-Acyl-Glutathione In Vitro and In Vivo in Human

“…Carboxylic acid-containing drugs such as ibuprofen can be metabolized to reactive acyl glucuronide and S-acyl-CoA metabolites that can mediate the transacylation of nucleophilic residues on protein and GSH in vitro and in vivo Grillo, 2011). For ibuprofen, most attention has focused on the chemical reactivity of its acyl glucuronide metabolite, where reports demonstrated it to mediate the covalent binding of ibuprofen to plasma protein and human serum albumin in vitro .…”

“…Carboxylic acid-containing drugs can be metabolized to chemically reactive 1-O-acyl glucuronide and/or S-acyl-CoA thioester derivatives capable of transacylating cellular nucleophiles including GSH in vitro and in vivo (Faed, 1984;Boelsterli, 2002;Skonberg et al, 2008;Grillo, 2011). Ibuprofen, a nonsteroidal anti-inflammatory drug, is metabolized by both acyl glucuronidation and acyl-CoA formation, leading to ibuprofen-1-b-O-acyl glucuronide (I-1-b-O-G; Kepp et al, 1997) and ibuprofen-S-acyl-CoA (I-SCoA; Knadler and Hall, 1990), respectively ( Fig.…”

Molecular Mechanisms and Physiological Significance of Organelle Interactions and Cooperation

“…This phenomenon, also described as "CoA sequestration" (Brass, 1994), can occur in metabolic disorders (Mitchell et al, 2008) or upon exposure to xenobiotics and drugs (Brass, 1994). In addition, CoA-esters are chemically reactive species that are known to transacylate the cysteinyl-thiol of GSH, and glutathione depletion has been described upon treatment of cells with certain carboxylates (Grillo, 2011). In conclusion, the CoA-SH/CoAester ratio can influence the cellular (mitochondrial) GSH/GSSG balance.…”