Crosstalk between CYP2E1 and CYP3A enzymes and its Possible Involvement in Alcohol‐Drug Interactions.

Abstract

The core of the drug‐metabolizing ensemble in human liver is formed by the ensemble of different P450 enzymes co‐localized in the membrane of endoplasmic reticulum (ER). There is an emerging recognition of the fact that multiple P450 species interact with each other with the formation of heteromeric complexes, where the catalytic properties of the individual enzymes are modified. Functional consequences of these interactions are likely to be responsible for the lack of a straight correlation between the composition of the P450 ensemble and the profile of drug metabolism. Of particular importance are the changes in P450‐P450 crosstalk induced by alcohol consumption. Although the multi‐fold increase in the content of CYP2E1 in liver of alcohol consumers is well documented, the involvement of CYP2E1 in alcohol‐drug interactions is considered insignificant due to a minor role of this enzyme in drug metabolism. However, the effects of induction of CYP2E1 may also involve its effects on the functional properties of other drug‐metabolizing enzymes.In order to probe the crosstalk between CYP2E1 and other P450 species and elucidate its role in alcohol‐drug interactions we established a model of alcohol‐induced increase in CYP2E1 content that implements incorporation of purified CYP2E1 into human liver microsomes (HLM). The adopted CYP2E1 becomes a fully‐functional member of the drug‐metabolizing ensemble and interacts with other P450 enzymes with the formation of heteromeric complexes (Davydova et al. 2019, https://doi.org/10.1101/685545). Investigating the impact of CYP2E1 on CYP3A enzymes, we found that the enrichment of HLM with CYP2E1 results in an increased rate of metabolism of 7‐benzyloxyquinoline (BQ) and eliminates the homotropic cooperativity observed with this CYP3A‐specific substrate. To probe the physiological relevance of these effects we compared three pulled preparations of HLM from normal donors (N‐HLM) with a preparation obtained from heavy alcohol consumers (A‐HLM). The content of major P450 species in the samples was characterized with mass‐spectroscopic determination of their specific peptides. The content of CYP2E1 in N‐HLM samples was from 2 to 5 lower than that found in A‐HLM. In contrast, the content of CYP3A in A‐HLM was the lowest among all four preparations. Despite the lower content of CYP3A, A‐HLM exhibited much higher rate of metabolism and lower homotropic cooperativity with BQ, similar to what is observed in CYP2E1‐enriched N‐HLM. Incorporation of additional CYP2E1 into A‐HLM results in a further increase in the rate of BQ metabolism that makes it an order of magnitude higher than in the N‐HLM sample with a similar CYP3A content. Importantly, the CYP2E1‐dependent activation observed with BQ, a highly specific substrate of CYP3A, is not observable with 7‐dimethylamino‐4‐trifluoromethyl coumarin (Coumarin 152), a substrate preferentially metabolized by CYP3A, but capable of high‐affinity interactions with several other P450 species. This observation corroborates our hypothesis that the mechanisms of substrate‐dependent activation of a particular P450 enzyme through P450‐P450 crosstalk involve a reorganization of P450 heterooligomers induced by the interactions of the target enzyme with its specific substrate.Support or Funding InformationThis research was supported by the grant R21‐AA024548 from NIH.