Effects of alcohol consumption and tobacco smoking on the composition of the ensemble of drug metabolizing enzymes and transporters in human liver

Abstract

We examined the effect of alcohol consumption and smoking on the abundance of drug-metabolizing enzymes and transporters (DMET) in human liver microsomes (HLM) isolated from liver tissues of 94 donors. Global proteomics analysis was performed and DMET protein levels were analyzed in relation to alcohol consumption levels, smoking history, and sex using non-parametric tests (p-value ≤ 0.05; cutoff of 1.25-fold change, FC). The examination of the alcohol-induced changes was further enforced by correlational analysis, where we used arbitrary alcohol consumption grade (ACG) scaling from 0 to 4 to establish a set of protein markers. We elaborated a provisional index of alcohol exposure (PIAE) based on a combination of relative abundances of four proteins (ER chaperone HSPA5, protein disulfide isomerases PDIA3 and P4HB, and cocaine esterase CES2) best correlating with ACG. The PIAE index was then used to find its correlations with the abundances of DMET proteins. Our results demonstrate considerable alcohol-induced changes in composition of the pool of cytochrome P450 enzymes in HLM. We observed significantly increased abundances of CYP2E1, CYP2B6, CYP2J2, and NADPH-cytochrome P450 reductase. In contrast, CYP1A2, CYP2C8, CYP2C9, CYP4A11, and cytochrome b5 protein levels were downregulated. Significant alteration in abundances of UDP-glucuronosyltransferase (UGT) were also detected, comprising of elevated UGT1A6, UGT1A9, and UGT2A1, and reduced UGT1A3, UGT1A4, UGT2B7, UGT2B10, and UGT2B15 levels. Important alcohol-induced changes were also observed in the expression of non-CYP and non-UGT DMET. Additionally, tobacco smoke was associated with elevated CYP1A2, UGT1A6, UGT2A1, and UGT2B4 and decreased FMO3, FMO4, and FMO5 levels.

Non-additivity of the functional properties of individual P450 species and its manifestation in the effects of alcohol consumption on the metabolism of ketamine and amitriptyline

“…PIAE is the combination of the relative abundances of four protein markers of alcohol exposure the heat-shock 70 protein HSPA5 (GRP75), protein disulfide isomerases P4HB and PDIA3, and carboxylesterase CES2. A strict correlation of PIAE with the level of alcohol consumption reported by liver donors has been demonstrated in our previous study [27]. This index varies from ∼0 to ∼5, and its numeric value roughly approximates the average number of alcoholic drinks per day consumed by the liver donor.…”

“…To probe the effect of alcohol consumption on ketamine metabolism we assessed the dependence of the overall turnover rate ( V max total ) and the distribution between the three individual kinetic components of the ketamine metabolism on the Provisional Index of Alcohol Exposure (PIAE) introduced in our recent study of the effect of alcohol exposure on the proteome of the drug-metabolizing enzymes and transporters in the human liver [27]. PIAE is the combination of the relative abundances of four protein markers of alcohol exposure the heat-shock 70 protein HSPA5 (GRP75), protein disulfide isomerases P4HB and PDIA3, and carboxylesterase CES2.…”

“…The results of our recent study of HLM proteome in a set of 96 HLM samples from donors with different levels of alcohol consumption [27] allowed us to establish a biomarker-based provisional index of alcohol exposure (PIAE). This parameter was used here to explore the impact of alcohol exposure on the metabolism of ketamine and amitriptyline, the drugs used as antidepressants in patients with alcohol withdrawal syndrome.…”

“…The abundances of 11 major P450 species, cytochrome b 5 , and CPR were quantified by a targeted proteomics assay as described earlier [23, 26]. Relative abundance of P450 species and protein markers of alcohol exposure (HSPA5, PDIA3, P4HB, and CES2 [27]) in the individual and pooled HLM preparations were determined based on the global proteomics-based total protein approach as described in [25].…”

“…To find the correlation of the shape and amplitudes of SSP with variations of the abundances of individual P450 enzymes, we calculated the vectors of relative fractional abundance (VFRA) for 11 major P450 in HLMs (CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4, CYP3A5, and CYP4A11). For these calculations, the raw MS intensities (MSI) for each ( i -th) P450 protein in each ( j- th) HLM sample in the dataset of 23 individual HLM samples or the combined data set of 23 individual and 9 pooled preparations were first normalized on the sum of intensities of 75 drug-metabolizing enzymes (DME) and ER-stress-related proteins with known localization in the microsomal membrane or microsomal lumen (see [27] for the list of these proteins): …”