Abstract
Carbon monoxide (CO), notorious for its toxicity, is endogenously produced by heme oxygenase and plays a role in diverse signalling pathways with wide-ranging physiological effects. Consequently, CO delivery has been explored as a potential therapy for conditions such as cardiovascular diseases. To address this need, CO-releasing pro-drugs have been developed, offering diverse release kinetics and potencies, making them potentially valuable candidates for clinical application. This research direction led to the development of novel and organic pH-dependent carbon monoxide-releasing molecules (oCOms) which have been demonstrated to exert a wide range of cardioprotective effects at low micromolar concentrations with minimal cardiotoxic injury.
The strong binding of CO to heme-containing proteins suggests the strong possibility that oCOms may inhibit the activity of cytochromes P450 (CYP450), key enzymes in xenobiotic metabolism. This study examines the concentration-dependent inhibitory effects of the fast-releasing CO-donor oCOm-21, with a half-life of 20 mins against that of a non-active debromo oCOm-21 (DB-21) and of a CO-depleted oCOm-21 by-product (BP-21) on in vitro microsomal CYP activity. Rat liver microsomes (RLM) were isolated from male Sprague Dawley rats and incubated with 0.3 - 10 µM of the CO-releasing oCOm-21 and the non-CO-releasing compounds DB-21 and BP-21. The enzymatic activity of hepatic rat microsomal CYP3A was assessed by measuring the formation of formaldehyde as the product of an erythromycin N-demethylation assay.
The results showed that both oCOm-21 and DB-21 significantly inhibited CYP3A activity, whereas BP-21 had no effect. This finding confirms that oCOm-21-derived CO produces a concentration-dependent inhibition of CYP3A activity and suggests that oCOm-21 CO release may be additionally activated by CYP450 oxidation mechanisms apart from the established mechanism of pH-triggered CO release. Furthermore, as hypothesised, the CO-depleted BP-21 had far less effect on CYP3A activity suggesting that BP-21 is a more suitable control for oCOm-21 than DB-21 in microsomal assays. These findings support further investigation into the pharmacokinetics of oCOm-21 and contribute to the pre-clinical safety information on oCOms. This research advances our understanding of the potential for drug interactions in a polypharmacy setting, where drug metabolism occurs through CYP450 mechanisms.