Abstract

Cytochrome P450 2A6 (CYP2A6) metabolizes several clinically relevant substrates, including nicotine, the primary psychoactive component in cigarette smoke. Smokers vary widely in their rate of inactivation and clearance of nicotine, altering numerous smoking phenotypes. We aimed to characterize independent and shared impact of genetic and non-genetic sources of variation in CYP2A6 mRNA, protein, and enzyme activity in a human liver bank (n=360). We quantified levels of CYP2A6, cytochrome P450 oxidoreductase (POR), and aldo-keto reductase 1D1 (AKR1D1) mRNA, and CYP2A6 and POR protein. CYP2A6 enzyme activity was determined through measurement of cotinine formation from nicotine and 7-hydroxycoumarin formation from coumarin. Donor DNA was genotyped for CYP2A6, POR, and AKR1D1 genetic variants. CYP2A6 phenotype measures were moderately to strongly correlated (r values ranging from 0.47-0.88, P<0.001). Female donors exhibited higher CYP2A6 mRNA expression relative to males (P<0.05). Donor age was weakly correlated with CYP2A6 protein (r=0.12, P<0.05) and activity (r=0.20, P<0.001). CYP2A6 reduce-of-function genotypes, but not POR or AKR1D1 genotype, were associated with lower CYP2A6 protein (P<0.001) and activity (P<0.01). AKR1D1 mRNA was moderately correlated with CYP2A6 mRNA (r=0.57, P<0.001), protein (r=0.30, P<0.001), and activity (r=0.34, P<0.001). POR protein was moderately correlated with CYP2A6 activity (r=0.45, P<0.001). Through regression analyses, 17% (P<0.001), 37% (P<0.001), and 77% (P<0.001) of the variation in CYP2A6 mRNA, protein, and activity, respectively, was accounted for by genetic and non-genetic factors. Overall, several independent and shared sources of variation in CYP2A6 activity in vitro have been identified, which could translate to variable hepatic clearance of nicotine.