Identification of the human cytochromes P450 catalysing the rate-limiting pathways of gliclazide elimination.
OBJECTIVE
To identify the human cytochrome P450 (CYP) enzymes responsible for the formation of the 6beta-hydroxy (6beta-OHGz), 7beta-hydroxy (7beta-OHGz) and hydroxymethyl (MeOH-Gz) metabolites of gliclizide (Gz).


METHODS
6beta-OHGz, 7beta-OHGz and MeOH-Gz formation by human liver microsomes and a panel of recombinant human P450s was measured using a high-performance liquid chromatography procedure, and the kinetics of metabolite formation was determined for each pathway. Effects of prototypic CYP enzyme selective inhibitors were characterized for each of the microsomal metabolic pathways.


RESULTS
Microsomes from six human livers converted Gz to its 6beta-OHGz, 7beta-OHGz, and MeOH-Gz metabolites, with respective mean (+/- SD) K(m) values of 461 +/- 139, 404 +/- 143 and 334 +/- 75 microm and mean V(max) values of 130 +/- 55, 82 +/- 31 and 268 +/- 115 pmol min(-1) mg(-1), respectively. V(max)/K(m) ratios for the microsomal reactions parallelled relative metabolite formation in vivo. Sulfaphenazole inhibited microsomal 6beta-OHGz, 7beta-OHGz and MeOH-Gz formation by 87, 83 and 64%, respectively, whereas S-mephenytoin caused significant inhibition (48%) of only MeOH-Gz formation. Recombinant CYP2C9, CYP2C18 and CYP2C19 catalysed all hydroxylation pathways, whereas CYP2C8 formed only 6beta-OHGz and 7beta-OHGz.


CONCLUSIONS
Taken together, the results indicate that CYP2C9 is the major contributor to Gz metabolic clearance, although CYP2C19 may also be involved in MeOH-Gz formation (the major metabolic pathway). Factors known to influence CYP2C9 activity will provide the main source of variability in Gz pharmacokinetics.
Elliot
David J
DJ
Department of Clinical Pharmacology, Flinders University and Flinders Medical Centre, Bedford Park, Adelaide, Australia.
Suharjono
Lewis
Benjamin C
BC
Gillam
Elizabeth M J
EM
Birkett
Donald J
DJ
Gross
Annette S
AS
Miners
John O
JO
D001189:Aryl Hydrocarbon Hydroxylases / Q000502:physiology*; D065731:Cytochrome P-450 CYP2C19; D065729:Cytochrome P-450 CYP2C9; D004791:Enzyme Inhibitors / Q000378:metabolism*; D005907:Gliclazide / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation / Q000187:drug effects; D008862:Microsomes, Liver / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000502:physiology*
D004791:Enzyme Inhibitors; D006899:Mixed Function Oxygenases; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C045793:CYP2C19 protein, human; D065731:Cytochrome P-450 CYP2C19; D005907:Gliclazide