ABSTRACT
1. We used an in vitro screening procedure and studies with individual human liver microsomes and cDNA-expressed CYP enzymes to investigate the metabolism of the putative neuroprotective drug N-methyl,N-propargyl-2-phenylethylamine (MPPE) to N-methylphenylethylamine (N-methylPEA) and N-propargylphenylethylamine (N-propargylPEA). 2. An electron-capture gas chromatographic procedure previously developed in our laboratories was used to measure the quantities of N-methylPEA and N-propargylPEA formed in the experiments with a single donor human liver microsome panel and cDNA expressed single CYP enzyme systems. The data were fitted to nonlinear regressions using Prism to determine kinetic constants. The results from a fluorogenic screen determined which cDNA-expressed single CYP enzymes were investigated. 3. CYP2B6, CYP2C19, and CYP2D6 all contributed to the formation of N-methylPEA, while only CYP2B6 catalyzed the formation of N-propargylPEA. The K (M) and V (max) values for N-propargylPEA formation were 290 +/- 70 microM and 139+/-16 ng/mL/min. The values for formation of N-methylPEA were not determined from these experiments due to the complexity of fitting the data to a three-variable equation, but data on the time course of N-methylPEA formation are presented. 4. Catabolism of MPPE to N-methylPEA and N-propargylPEA is catalyzed by CYP enzymes. CYP2B6, 2C19 and 2D6 all contribute to the depropargylation of the parent compound, but only CYP2B6 also catalyzes demethylation. CYP2C19 was found to be the most active with respect to generation of N-methylPEA.
Subject(s)
Cytochrome P-450 Enzyme System/genetics , Cytochrome P-450 Enzyme System/metabolism , Microsomes, Liver/enzymology , Neuroprotective Agents/pharmacokinetics , Selegiline/analogs & derivatives , Aryl Hydrocarbon Hydroxylases/genetics , Aryl Hydrocarbon Hydroxylases/metabolism , Binding, Competitive , Cytochrome P-450 CYP2B6 , Cytochrome P-450 CYP2C19 , Cytochrome P-450 CYP2D6/genetics , Cytochrome P-450 CYP2D6/metabolism , DNA, Complementary , Enzyme Activation , Gene Expression Regulation, Enzymologic , Humans , In Vitro Techniques , Inactivation, Metabolic , Mixed Function Oxygenases/genetics , Mixed Function Oxygenases/metabolism , Neuroprotective Agents/chemistry , Oxidoreductases, N-Demethylating/genetics , Oxidoreductases, N-Demethylating/metabolism , Selegiline/chemistry , Selegiline/pharmacokineticsABSTRACT
INTRODUCTION: N-Methyl,N-propargyphenylethylamine (MPPE) is a novel analog of (-)-deprenyl, a drug prescribed for Parkinson's disease and shown to have neuroprotective and neurorescue properties in a wide variety of in vitro and in vivo models. MPPE is also neuroprotective, but has the advantage over (-)-deprenyl of not being metabolized to amphetamine or N-methylamphetamine. METHOD: In this paper, extractive derivatization with pentafluorobenzenesulfonyl chloride (PFBSC) followed by electron-capture gas chromatography was utilized to study the metabolism of MPPE. RESULTS: The procedure is rapid and reproducible, giving derivatives with excellent chromatographic properties. Using this procedure, it has now been shown that beta-phenylethylamine (PEA), N-methylphenylethylamine (N-methylPEA) and N-propargylphenylethylamine (N-propargylPEA) are formed from MPPE during incubation of this drug with human liver microsomes. Levels of all three metabolites were shown to increase with increasing time of incubation with the microsomes. DISCUSSION: Extractive derivatization with PFBSC followed by electron-capture gas chromatography represents an efficient means of separating and quantitating the metabolites of MPPE, a novel neuroprotective agent.
