Non‐Additive Effects of Changing the Composition of the Cytochrome P450 Ensemble in Human Liver Microsomes: the Focus on CYP2E1 and Alcohol‐Drug Interactions

Abstract

Recent studies on a large selection of human liver microsomes (HLM) demonstrated the lack of a straight correlation between the composition of human cytochrome P450 ensemble and the profile of drug metabolism. At the same time, there is an emerging recognition of the functional importance of physical interactions between multiple P450 species co‐localized in the microsomal membrane. Due to complex effects of these interactions any change in the content of a particular P450 enzyme may affect drug metabolism in a complex, hard‐to‐predict manner. Of particular practical importance are the functional effects of the changes in the composition of P450 ensemble induced by alcohol consumption. Although the multi‐fold increase in the content of cytochrome P450 2E1 (CYP2E1) in liver observed in both alcoholics and moderate alcohol consumers is well documented, the role of CYP2E1 in the instances of alcohol‐drug interactions is commonly considered insignificant due to a minor role of this enzyme in drug metabolism. However, the involvement of CYP2E1 in alcohol‐drug interactions may stretch far beyond its immediate effects on the metabolism of its substrates.The aim of this study is to probe the effect of increasing concentrations of CYP2E1 in HLM on the metabolism of drug substrates of other P450 species. In a search for a model system suitable for these studies we introduced a method based on incorporation of purified cytochromes P450 into HLM that allows creating the microsomal preparations with altered composition of the P450 ensemble. Incorporation of CYP2E1 into HLM results in an increase in the rate of CYP2E1‐dependent metabolism of chlorzoxazone, which is proportional to the amount of the incorporated enzyme up to its content of 0.3 nmol per mg protein (or ~50% of CYP2E1 in the total P450 content). Studies with a series of substrates of drug‐metabolizing P450 species showed that CYP2E1 incorporation also results in pronounced alterations in the metabolism of substrates of several other P450 enzymes. The most important effect is observed on CYP2C19. The changes in the effective KM and Vmax values with such probe substrates as 7‐etoxy‐4‐cyanocoumarin (CEC, the substrate of CYP2C19 and CYP1A2), 7‐dimethylamino‐4‐trifluoromethylcoumarin (coumarin‐152, the substrate of CYP2B6, CYP2C19 and CYP2C9) and N‐butyl 7‐methoxy‐4‐(aminomethyl)‐coumarin (BMAMC, the substrate of CYP1A2, CYP2C19 and CYP2D6) reveal a substantial decrease in the involvement of CYP2C19 in their metabolism. Furthermore, increase in CYP2E1 content causes an important changes in the function of CYP3A enzymes, which is revealed in a compromised homotropic cooperativity, decreased affinity and increased Vmax observed with 7‐benzyloxyquinoline, a selective substrate of CYP3A4 and CYP3A5. These observations suggest that ethanol‐dependent induction CYP2E1 may be involved in pronounced pharmacokinetic interactions of alcohol with such drug substrates of CYP2C19 and CYP3A enzymes as benzodiazepines (diazepam), tricyclic antidepressants (amitriptylline, imipramine) and anticonvulsants (phenytoin, carbamazepine).Support or Funding InformationThis research was supported by grant R21‐AA024548 from NIH.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.