In vitro characterization of the inhibitory effects of ketoconazole on metabolic activities of cytochrome P-450 in canine hepatic microsomes.
OBJECTIVE
To evaluate the inhibitory potency of ketoconazole (KTZ) on the metabolic activities of isozymes of cytochrome P-450 (CYP) in dogs.


METHODS
4 healthy 1-year-old male Beagles.


METHODS
Hepatic microsomes were harvested from 4 dogs after euthanasia. To investigate the effects of KTZ on CYP metabolic activities, 7-ethoxyresorufin, tolbutamide, bufuralol, and midazolam hydrochloride were used as specific substrates for CYP1A1/2, CYP2C21, CYP2D15, and CYP3A12, respectively. The concentrations of metabolites formed by CYP were measured by high-performance liquid chromatography, except for the resorufin concentrations that were measured by a fluorometric method. The reaction velocity-substrate concentration data were analyzed to obtain kinetic variables, including maximum reaction velocity, Michaelis-Menten constant, and inhibitory constant (Ki).


RESULTS
KTZ competitively inhibited 7-ethoxyresorufin O-deethylation and midazolam 4-hydroxylation; it noncompetitively inhibited tolbutamide methylhydroxylation. Bufuralol 1'-hydroxylation was inhibited slightly by KTZ. The mean Ki values of KTZ were 10.6+/-6.0, 170+/-2.5, and 0.180+/-0.131 microM for 7-ethoxyresorufin O-deethylation, tolbutamide methylhydroxylation, and midazolam 4-hydroxylation, respectively.


CONCLUSIONS
In dogs, KTZ at a therapeutic dose may change the pharmacokinetics of CYP3A12 substrates as a result of inhibition of their biotransformation. Furthermore, no influence of KTZ on the pharmacokinetics of CYP1A1/2, CYP2C21, and CYP2D15 substrates are likely. In clinical practice, adverse drug effects may develop when KTZ is administered concomitantly with a drug that is primarily metabolized by CYP3A12.
Kuroha
Masanori
M
Department of Veterinary Medicine, Faculty of Agriculture, Tokyo University of Agriculture and Technology, Fuchu, Japan.
Kuze
Yoji
Y
Shimoda
Minoru
M
Kokue
Eiichi
E
D000818:Animals; D000935:Antifungal Agents / Q000494:pharmacology*; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004285:Dogs / Q000378:metabolism*; D004347:Drug Interactions; D004983:Ethanolamines / Q000378:metabolism; D007527:Isoenzymes; D007654:Ketoconazole / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D008874:Midazolam / Q000378:metabolism; D010078:Oxazines / Q000378:metabolism; D014044:Tolbutamide / Q000378:metabolism
D000935:Antifungal Agents; D065607:Cytochrome P-450 Enzyme Inhibitors; D004983:Ethanolamines; D007527:Isoenzymes; D010078:Oxazines; C007649:ethoxyresorufin; C010831:bufuralol; D003577:Cytochrome P-450 Enzyme System; D014044:Tolbutamide; C475779:cytochrome P-450 CYP3A12 (canine); D008874:Midazolam; D007654:Ketoconazole