Human liver cytochrome P450 2D6 genotype, full-length messenger ribonucleic acid, and activity assessed with a novel cytochrome P450 2D6 substrate.

Objective The goal of this study was to develop and validate a cytochrome P450 (CYP) 2D6 probe substrate with improved sensitivity to elucidate the relationship of CYP2D6 ribonucleic acid transcript levels, genotype, and enzyme activity in human liver biopsy samples. Methods CYP2D6 activity in tissue homogenates of liver biopsy specimens collected from control subjects (with no apparent liver disease), liver biopsy subjects, liver transplant subjects, and liver bank specimens was assessed with a calcimimetic, R-568, a high-clearance and specific substrate of CYP2D6. The livers were genotyped for the 6 most common CYP2D6 genetic variants (ie, *3, *4, *5, *6, *7, and *8). The 1.5-kilobase CYP2D6 messenger ribonucleic acid (referred to as full-length) transcripts were estimated with a semiquantitative reverse transcription–polymerase chain reaction assay. Results As a CYP2D6-specific catalytic probe, R-568 offers a 20-fold higher sensitivity compared with that of dextromethorphan. The improved assay sensitivity allowed evaluation of CYP2D6 enzyme activity in a few milligrams of tissue collected from biopsy specimens. The ratio of CYP2D6 enzyme activity to transcript remained relatively constant within each group of subjects, especially within the control group. However, mean activity to transcript varied greatly across the 4 groups of subjects. The liver samples in the control group showed significantly higher enzyme activity but a lower transcript level. Conclusions A combination of genotyping and messenger ribonucleic acid level determination could allow a quantitative estimation of functional CYP2D6 activity in healthy human livers with a reasonable degree of confidence. Kinetic study with R-568 indicates that this compound is probably the most sensitive CYP2D6 probe substrate available. Clinical Pharmacology & Therapeutics (2004) 75, 282–297; doi:10.1016/j.clpt.2003.12.003