Characterization of cytochrome P450 isoforms involved in sequential two-step bioactivation of diclofenac to reactive p-benzoquinone imines.
Idiosyncratic drug-induced lever injury (IDILI) is a rare but severe side effect of diclofenac (DF). Several mechanisms have been proposed as cause of DF-induced toxicity including the formation of protein-reactive diclofenac-1',4'-quinone imine (DF-1',4'-QI) and diclofenac-2,5-quinone imine (DF-2,5-QI). Formation of these p-benzoquinone imines result from two-step oxidative metabolism involving aromatic hydroxylation to 4'-hydroxydiclofenac and 5-hydroxydiclofenac followed by dehydrogenation to DF-1',4'-QI and DF-2,5-QI, respectively. Although the contribution of individual cytochrome P450s (CYPs) in aromatic hydroxylation of DF is well studied, the enzymes involved in the dehydrogenation reactions have been poorly characterized. The results of the present study show that both formation of 4'-hydroxydiclofenac and it subsequent bioactivation to DF-1',4'-QI is selectively catalyzed by CYP2C9. However, the two-step bioactivation to DF-2,5-QI appears to be catalyzed with highest activity by two different CYPs: 5-hydroxylation of DF is predominantly catalyzed by CYP3A4, whereas its subsequent bioactivation to DF-2,5-QI is catalyzed with 14-fold higher intrinsic clearance by CYP2C9. The fact that both CYPs involved in two-step bioactivation of DF show large interindividual variability may play a role in different susceptibility of patients to DF-induced IDILI. Furthermore, expression levels of these enzymes and protective enzymes might be important factors determining sensitivity of in vitro models for hepatotoxicity.
['den Braver|Michiel W|MW|', 'den Braver-Sewradj|Shalenie P|SP|', 'Vermeulen|Nico P E|NP|', 'Commandeur|Jan N M|JN|']
D065767:Activation, Metabolic D000894:Anti-Inflammatory Agents, Non-Steroidal D016227:Benzoquinones D065729:Cytochrome P-450 CYP2C9 D051544:Cytochrome P-450 CYP3A D004008:Diclofenac D005978:Glutathione D006801:Humans D006900:Hydroxylation D007097:Imines D007700:Kinetics D008862:Microsomes, Liver D010084:Oxidation-Reduction D011994:Recombinant Proteins D018570:Risk Assessment
Cytochrome P450 Bioactivation Diclofenac Glutathione conjugates Idiosyncratic liver injury Reactive intermediates