N-demethylation of levo-alpha-acetylmethadol by human placental aromatase.
Levo-alpa-acetylmethadol (LAAM) is a methadone derivative used to treat the opiate addict. We previously reported on the kinetics for transplacental transfer of LAAM and its levels in the fetal circuit using the technique of dual perfusion of the placental lobule. The aim of this investigation was to identify the enzyme responsible for the biotransformation of LAAM and norLAAM and the metabolites formed in the term human placenta. Placental microsomes exhibited higher activities than the mitochondrial and cytosolic fractions in metabolizing LAAM to norLAAM. None of these subcellular fractions catalyzed the formation of dinorLAAM from either LAAM or norLAAM as determined by HPLC/UV. Evidence obtained from the effects of cytochrome P450 (CYP) inhibitors on the demethylation of LAAM to norLAAM by placental microsomes suggested that CYP 19/aromatase is the major enzyme involved. Out of 10 monoclonal antibodies raised against various CYP isoforms, only that for aromatase caused over 80% inhibition of norLAAM formation. The biotransformation of LAAM to norLAAM exhibited monophasic kinetics with apparent Km and Vmax values of 105 +/- 57 microM and 86.8 +/- 15.6 pmol mg(-1) protein min(-1), respectively. The kinetic profile determined for a cDNA-expressed CYP 19 metabolism of LAAM to norLAAM was similar to that determined for placental microsomes. Taken together, the above data indicate that CYP 19/aromatase is the enzyme responsible for the N-demethylation of LAAM to norLAAM in term human placentas obtained from healthy pregnant women.
Deshmukh
Sujal V
SV
Department of Obstetrics and Gynecology, University of Texas Medical Branch, 301 University Boulevard, Galveston, TX 77555-0587, USA.
Nanovskaya
Tatiana N
TN
Hankins
Gary D V
GD
Ahmed
Mahmoud S
MS
D001141:Aromatase / Q000378:metabolism*; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008692:Methadyl Acetate / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism*; D008745:Methylation; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D013347:Subcellular Fractions
C020783:paracymethadol; D001141:Aromatase; D008692:Methadyl Acetate