Inhibition of Cytochrome P450 Activities by Propoxazepam: Safety Assessment in Context for Potential Drug Interactions

Vitalii Larionov, Mykola Golovenko, Iryna Valivodz, Anatoliy Reder — Wed, 05 Feb 2025 00:00:00 +0200

Background. Propoxazepam is a new anelgetic agent of the benzodiazepine group, chemically known as 7-bromo-5-(o-chlorophenyl)-3-propyloxy-1,2-dihydro-3H-1,4-benzodiazepine-2-one. Propoxazepam is considered a possible substrate of the CYP system, so its effect of on the CYP3A4 enzyme activity was investigated in vitro using human liver microsomes.

Objective. To evaluate the effects of propoxazepam on CYP3A4 activity in vitro using testosterone and midazolam as markers of metabolic activity for CYP3A4 in human liver microsomes.

Methods. Midazolam (1′-hydroxylation reaction) and testosterone (6β-hydroxylation reaction) were used as markers for CYP3A4-mediated activity. Ketoconazole (0.2 μM) was used as a positive control for reversible inhibition, and troleandomycin (50 μM) for metabolism-dependent inhibition. For reversible inhibition, propoxazepam was added together with the corresponding substrate and cofactor (NADPH), while for metabolism-dependent inhibition, it was incubated with microsomes and cofactor for 30 minutes prior to substrate addition.

Results. Propoxazepam at various concentrations (0 to 100 μM) consistently inhibited CYP3A4 activities for both substrates, showing a similar "concentration–activity inhibition" dependence, with IC₅₀ values of 52.3 ± 4.9 μM for midazolam and 46.1 ± 9.2 μM for testosterone. For metabolism-dependent inhibition, IC₅₀ values were 36.6 ± 8.6 μM for midazolam and 28.3 ± 7.4 μM for testosterone. Given that the binding of propoxazepam to microsomal protein under the experimental conditions, which reflected those in the IC₅₀ experiments, was low, no microsomal binding correction factor was applied to the reported IC₅₀ values.

Conclusions. The highest predicted unbound C_max plasma concentration of propoxazepam, above which interactions can occur, is between 0.462 and 0.524 μM, or 462 and 524 nM. This corresponds to concentrations of 188 to 214 ng/mL (based on the molecular weight of propoxazepam, 414.73 g/mol). According to pharmacokinetic data, these concentrations are not achievable after a single oral administration. Further studies are required for multiple-dose administration.

Innovative Biosystems and Bioengineering

Inhibition of Cytochrome P450 Activities by Propoxazepam: Safety Assessment in Context for Potential Drug Interactions