Flavin-containing monooxygenase 1-catalysed N,N-dimethylamphetamine N-oxidation

Area of Science:

• Pharmacology
• Enzymology
• Drug Metabolism

Background:

• N,N-dimethylamphetamine (DMA) is a stimulant drug and analogue of methamphetamine.
• DMA is metabolized to DMA N-oxide (DMANO), a process potentially catalyzed by flavin-containing monooxygenases (FMOs).
• The specific enzymes responsible for DMA N-oxidation have not been previously identified.

Purpose of the Study:

• To identify and characterize the specific human enzyme(s) responsible for the N-oxidation of N,N-dimethylamphetamine (DMA).
• To determine the kinetic parameters of DMA N-oxidation mediated by the identified enzyme(s).

Main Methods:

• Incubation of DMA with human recombinant drug-metabolizing enzymes, including FMOs and cytochrome P450s (CYPs).
• Assays to measure the formation of DMA N-oxide (DMANO).
• Kinetic analysis using Michaelis-Menten parameters (Km, Vmax, intrinsic clearance).
• Inhibition studies using an FMO1-specific inhibitor, imipramine.

Main Results:

• Flavin-containing monooxygenase 1 (FMO1) was identified as the predominant enzyme catalyzing DMA N-oxidation.
• Michaelis-Menten kinetic constants for FMO1-mediated DMA N-oxidation were determined (Km = 44.5 µM, Vmax = 7.59 nmol/min/mg protein, intrinsic clearance = 171 µl/min/mg protein).
• FMO1-mediated DMA N-oxidation exhibited approximately twelve-fold higher intrinsic clearance compared to FMO3.
• Imipramine selectively inhibited DMA N-oxidation, confirming FMO1's role.

Conclusions:

• N,N-dimethylamphetamine (DMA) N-oxidation is primarily mediated by the flavin-containing monooxygenase 1 (FMO1) enzyme.
• FMO1 plays a significant role in the biotransformation of DMA.
• The characterized kinetic parameters provide valuable insights into the metabolic pathway of DMA.