CYP-catalysed Cycling of Clozapine and Clozapine- N -oxide Promotes the Generation of Reactive Oxygen Species in vitro .

Clozapine is an effective atypical antipsychotic indicated for treatment-resistant schizophrenia but is under-prescribed due to the risk of severe adverse drug reactions such as myocarditis. A mechanistic understanding of clozapine cardiotoxicity remains elusive. This study aimed to investigate the contribution of selected CYP isoforms to cycling between clozapine and its major circulating metabolites, N -desmethylclozapine and clozapine- N -oxide, with the potential for reactive species production. CYP supersome™-based in vitro techniques were utilised to quantify specific enzyme activity associated with clozapine, clozapine- N -oxide and N -desmethylclozapine metabolism. The formation of reactive species within each incubation were quantified, and known intermediates detected. CYP3A4 predominately catalysed clozapine- N -oxide formation from clozapine and was associated with concentration-dependent reactive species production, whereas isoforms favouring the N -desmethylclozapine pathway (CYP2C19 and CYP1A2) did not produce reactive species. Extrahepatic isoforms CYP2J2 and CYP1B1 were also associated with the formation of clozapine- N -oxide and N -desmethylclozapine but did not favour one metabolic pathway over another. Unique to this investigation is that various CYP isoforms catalyse clozapine -N -oxide reduction to clozapine. This process was associated with the concentration-dependent formation of reactive species with CYP3A4, CYP1B1 and CYP1A1 that did not correlate with known reactive intermediates, implicating metabolite cycling and reactive oxygen species in the mechanism of clozapine-induced toxicity.