Identification of cytochrome P450 enzymes involved in the metabolism of 3′,4′-methylenedioxy-α-pyrrolidinopropiophenone (MDPPP), a designer drug, in human liver microsomes

Abstract

The metabolism of 3′,4′-methylenedioxy-α-pyrrolidinopropiophenone (MDPPP), a novel designer drug, to its demethylenated major metabolite 3′,4′-dihydroxy-pyrrolidinopropiophenone (di-HO-PPP) was studied in pooled human liver microsomes (HLM) and in cDNA-expressed human hepatic cytochrome P450 (CYP) enzymes. CYP2C19 catalysed the demethylenation with apparent Km and Vmax values of 120.0 ± 13.4 µM and 3.2 ± 0.1 pmol/min/pmol CYP, respectively (mean ± standard deviation). CYP2D6 catalysed the demethylenation with apparent Km and Vmax values of 13.5 ± 1.5 µM and 1.3 ± 0.1 pmol/min/pmol CYP, respectively. HLM exhibited a clear biphasic profile with an apparent Km,1 value of 7.6 ± 9.0 and a Vmax,1 value of 11.1 ± 3.6 pmol/min/mg protein, respectively. Percentages of intrinsic clearances of MDPPP by specific CYPs were calculated using the relative activity factor (RAF) approach with (S)-mephenytoin-4′-hydroxylation or bufuralol-1′-hydroxylation as index reactions for CYP2C19 or CYP2D6, respectively. MDPPP, di-HO-PPP and the standard 4′-methyl-pyrrolidinohexanophenone (MPHP) were separated and analysed by liquid chromatography-mass spectrometry in the selected-ion monitoring (SIM) mode. The CYP2D6-specific chemical inhibitor quinidine (3 µM) significantly (p < 0.001) inhibited di-HO-PPP formation by 75.8% ± 1.7% (mean ± standard error of the mean) in incubation mixtures with HLM and 2 µM MDPPP. It can be concluded from the data obtained from kinetic and inhibition studies that polymorphically expressed CYP2D6 and CYP2C19 are almost equally responsible for MDPPP demethylenation.