Use of electrospray ionization liquid chromatography-mass spectrometry to study the role of CYP2D6 in the in vitro metabolism of 5-hydroxytryptamine receptor antagonists.
An electrospray ionization liquid chromatographic-mass spectrometric (ESI-LC-MS) method has been developed to study the involvement of the cytochrome P450 isoenzyme CYP2D6 in the in vitro metabolism of the indole containing 5-hydroxytryptamine (5-HT(3)++) receptor antagonists tropisetron, ondansetron and dolasetron in human liver microsomes. Compounds were eluted using linear gradients of acetonitrile-20 mM ammonium acetate, solvent A, (10:90, v/v) (pH 6.0) and solvent B, (60:40, v/v) (pH 6.0) and a Nucleosil C(4) column. Microsomal incubations were analysed using selected ion monitoring of the molecular ion of parent drug and the molecular ion of hydroxylated metabolites. The involvement of CYP2D6 in drug metabolism was assessed by inhibition studies using quinidine (5 mu M), a specific inhibitor of human CYP2D6, as well as by incubating compounds with microsomes prepared from cells transfected with cDNA encoding human CYP2D6. Results showed that the oxidation of all three compounds involved CYP2D6, but only that of tropisetron was inhibited by over 90% in the presence of quinidine. The present method can be applied to pre-clinical compounds, at an early stage of drug discovery, to assess the involvement of CYP2D6 in their metabolism and to screen for those compounds where CYP2D6 is the only isoenzyme implicated in the formation of major metabolites.
Sanwald
P
P
Department of Drug Metabolism, Marion Merrell Dow, Strasbourg, France.
David
M
M
Dow
J
J
D002853:Chromatography, Liquid / Q000379:methods*; D019389:Cytochrome P-450 CYP2D6 / Q000378:metabolism*; D018076:DNA, Complementary; D006801:Humans; D007211:Indoles / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D011807:Quinolizines / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D012702:Serotonin Antagonists / Q000378:metabolism*
D018076:DNA, Complementary; D007211:Indoles; D011807:Quinolizines; D011994:Recombinant Proteins; D012702:Serotonin Antagonists; C060344:dolasetron; D019389:Cytochrome P-450 CYP2D6