Received for publication June 5, 2007.
Revised September 25, 2007.
Accepted for publication October 24, 2007.

Interaction of the electrophilic ketoprofenyl-glucuronide and ketoprofenyl-coenzyme A conjugates with cytosolic glutathione S-transferases

Abstract

Carboxylic acid-containing drugs are metabolized mainly through the formation of glucuronide and coenzyme A esters. These conjugates have been suspected to be responsible for the toxicity of several nonsteroidal anti-inflammatory drugs because of the reactivity of the electrophilic ester bond. We investigated in the present study the reactivity of ketoprofenyl-acylglucuronide (KPF-OG) and ketoprofenyl-acyl-Coenzyme A (KPF-SCoA) towards cytosolic rat liver glutathione S-transferases (GST). We observed that KPF-ScoA, but not KPF-OG inhibited the conjugation of 1-chloro-2,4-dinitrobenzene and 4-nitroquinoline N-oxide catalyzed by both purified cytosolic rat liver GST and GST from FAO and H5-6 rat hepatoma cell lines. Photoaffinity labeling with KPF-SCoA suggested that the binding of this metabolite may overlap the binding site of 4-methylumbelliferone sulfate. Furthermore, high performance liquid chromatography and mass spectrometry analysis showed that both hydrolysis and transacylation reactions were observed in the presence of GST and glutathione. The formation of KPF-S-acyl-glutathione could be kinetically characterized (apparent K_M = 196.0 ± 70.6 µM). It is concluded that KPF-SCoA is both a GST inhibitor and a substrate of a GST-dependent transacylation reaction. The reactivity and inhibitory potency of thioester CoA derivatives towards GST may have potential implications on the reported in vivo toxicity of some carboxylic acid-containing drugs.

Key words: bioactivation, enzyme inhibitors, glucuronidation, glutathione transferases, inhibition, phase II drug metabolism, UDP glucuronyltransferases