Induction Of P450 2B Research Articles

Strong Induction of Cytochrome P450 1A/3A, But not P450 2B, in Cultured Hepatocytes from Common Marmosets and Cynomolgus Monkeys by Typical Human P450 Inducing Agents.

Common marmosets (Callithrix jacchus) and cynomolgus monkeys (Macaca fascicularis) are used as non-human primate models in preclinical studies for drug development. The assessment of P450 induction in hepatocytes from marmosets and cynomolgus monkeys was performed using typical P450 inducers. Induction of cytochrome P450 1-4 family enzymes was analyzed in two lots of cultured hepatocytes from common marmosets and cynomolgus monkeys after 24-h treatment with typical human P450 inducing agents by real-time reverse transcription-polymerase chain reaction. Marmoset P450 3A4 mRNA and P450 2C8/2C19 mRNA in hepatocytes were strongly (>10- fold) and weakly (>2) induced by rifampicin, respectively. Marmoset 1A1 and 1A2 mRNA were induced strongly (>200-fold) by β-naphthoflavone and omeprazole. Marmoset P450 2B6 mRNA was induced (~5-fold) by a constitutive androstane receptor agonist, but not by phenobarbital. Cynomolgus monkey P450 3A4 mRNA and P450 1A1 mRNA in cultured hepatocytes were also induced by rifampicin and omeprazole, respectively, but P450 2B6 mRNA was not induced by phenobarbital. These results indicate that P450 1A/3A induction by typical human P450 inducers in hepatocytes from marmosets and/or cynomolgus monkeys are similar to those of humans (except for P450 2B induction by phenobarbital in humans), suggesting that marmosets and cynomolgus monkeys might be suitable models for evaluating the drug interactions in preclinical studies.

Propofol metabolism is enhanced after repetitive ketamine administration in rats: the role of cytochrome P-450 2B induction

In a series of ex vivo and in vivo studies we investigated the ability of repetitive ketamine administration to alter the metabolism and anaesthetic effect of propofol and the role of ketamine-mediated P-450 2B induction in rats. Male Wistar rats were pretreated with 80 mg kg(-1) ketamine i.p. twice daily for 4 days. Pentoxyresorufin O-dealkylation (PROD), P-450 2B protein and mRNA were determined. Residual propofol concentration was measured after incubating hepatic microsomes with 100 muM propofol. Sleeping times induced by i.p. 80 mg kg(-1) propofol were determined. Orphenadrine, a P-450 2B inhibitor, was added in both ex vivo and in vivo studies. Finally, serial whole blood propofol concentrations were determined after i.v. infusion of 15 mg kg(-1) propofol. Ketamine pretreatment produced 5.4-, 3.4- and 1.7-fold increases in hepatic PROD activity, P-450 2B protein and mRNA, respectively. Residual propofol concentration was 46% lower after incubation with microsomes from ketamine-pretreated rats than in the control group. The addition of orphenadrine to ketamine-pretreated microsomes produced an increase in residual propofol concentration in a concentration-dependent manner. Ketamine pretreatment reduced propofol sleeping time to 12% of the control, which was reversed by orphenadrine. The whole blood propofol concentration in ketamine-pretreated rats was significantly lower than that of control rats at 1, 2, 4 and 8 min after cessation of propofol infusion. Repetitive ketamine administration enhances propofol metabolism and reduces propofol sleeping time in rats. We suggest that P-450 2B induction may produce ketamine-propofol interaction in anaesthetic practice.

Absence of enantioselectivity in the pharmacodynamics of P450 2B induction by 5‐ethyl‐5‐phenylhydantoin in the male rat liver or in cultured rat hepatocytes

To explore the enantioselectivity of ligand interaction with the putative phenobarbital receptor, the pharmacodynamics of cytochrome P450 2B (CYP2B) induction by racemic 5-ethyl-5-phenylhydantoin and its two enantiomers were investigated in the male F344/NCr rat and in cultured adult male rat hepatocytes. Steady-state serum drug concentrations, measured following 14 days of administration of the compounds in the diet (0-1320 ppm, n = 3 rats per group), were used as an approximation of intrahepatocellular drug concentration. The serum xenobiotic concentrations associated with half-maximal hepatic CYP2B induction were 5-10 microM, based on measurement of pentoxy- or benzyloxyresorufin O-dealkylation activities, or immunoreactive CYP2B1 protein. The corresponding potency values in the hepatocyte culture experiments were 8-12 microM, based on measurement of total cellular RNA coding for CYP2B1. In both the in vivo and the hepatocyte culture experiments, the potencies for CYP2B induction were essentially equivalent for the racemate and the individual enantiomers of 5-ethyl-5-phenylhydantoin. In the case of this compound, there would appear to be no enantioselectivity for CYP2B induction. This finding may be interpreted as evidence against receptor mediation in the induction of CYP2B activity, although it is also possible that a receptor is involved that does not exhibit enantioselectivity.

Hepatic cytochrome P450 2B induction by ethyl/ phenyl‐substituted congeners of phenobarbital in the B6C3F1 mouse

The abilities of structural congeners of phenobarbital to induce immunoreactive hepatic cytochrome P450 2B (CYP2B) protein and associated catalytic activity (benzyloxyresorufin O-dealkylation) in the male B6C3F1 mouse were examined. Interspecies differences in inducing ability were examined through comparison of the results with induction data obtained previously with the male F344/NCr rat. The congeners were administered in the diet for 2 weeks at concentrations equimolar to 500 ppm of the prototype CYP2B inducer, phenobarbital. Of the series of compounds tested, phenobarbital was the most effective inducer of benzyloxyresorufin O-dealkylation and immunoreactive CYP2B protein, with 2-ethyl-2-phenylsuccinimide, 5-ethyl-5-phenylhydantoin, primidone, and glutethimide being only 19-42% as effective. 5-Ethyl-5-phenyloxazolidinedione and the ring-opened and decarboxylated congeners, N-(2-ethyl-2-phenylacetyl)urea and 2-ethyl-2-phenylmalonamide, displayed minimal induction of these catalytic activities. Dose-response experiments performed with 5-ethyl-5-phenylhydantoin indicated that the intrinsic CYP2B-inducing activity of this congener was as great as that of phenobarbital in the mouse, although a fourfold greater dietary concentration of this hydantoin (2000 ppm) was required to elicit a response equivalent to that caused by 500 ppm phenobarbital. When extent of induction was related to serum total xenobiotic concentration rather than to administered dietary concentration, the potencies of the two congeners were determined to be more similar (58 vs. > or = 78 microM for phenobarbital and 5-ethyl-5-phenylhydantoin, respectively).

Growth Hormone-Dependent and -Independent Sexually Dimorphic Regulation of Phenobarbital-Induced Hepatic Cytochrome P450 2B1 and -2B2

Sexually dimorphic regulation of phenobarbital-induced cytochromes P450 2B1 and 2B2 (collectively referred to as P450 2B) as well as P450 2B-dependent monooxygenase activities was studied in multi-hormone-depleted hypophysectomized rats and in growth hormone (GH)-deficient monosodium glutamate (MSG)-treated rats. Our results indicate that endogenous GH suppresses phenobarbital induction of P450 2B and that the feminine pattern of continuous GH secretion is more suppressive than the masculine profile of episodic secretion. Moreover, we have found that it is the height of the GH pulse, and not necessarily its frequency nor the interpulse trough periods, that signals the suppressive effects of GH on P450 2B expression in male rats. Last, irrespective of the presence or absence of circulating GH, the magnitude of phenobarbital induction of P450 2B and associated monooxygenases was consistently lower in female rats, suggesting the presence of some degree of sex-dependent, but GH-independent regulation.

Pharmacodynamics of cytochrome P450 2B induction by phenobarbital, 5-ethyl-5-phenylhydantoin, and 5-ethyl-5-phenyloxazolidinedione in the male rat liver or in cultured rat hepatocytes.

The pharmacodynamics of rat hepatic cytochrome P450 2B (P450 2B) induction by phenobarbital (PB) and two structural congeners, dl-5-ethyl-5-phenylhydantoin (EPH) and dl-5-ethyl-5-phenyloxazolidinedione (EPO), were investigated. The in vivo induction of P450 2B was probed in F344/NCr rats by measuring immunoreactive hepatic P450 2B1 protein and by assaying the hepatic 16 beta-hydroxylation of testosterone and O-dealkylation of (benzyloxy)- and pentoxyresorufin. The induction of (benzyloxy)resorufin O-dealkylation activity was also measured in adult rat hepatocyte cultures exposed to the three xenobiotics. The concentration of xenobiotic at the putative active site in the in vivo studies was approximated by measuring serum total xenobiotic levels, while in the hepatocyte culture studies, the nominal xenobiotic concentration in the culture medium was used. Concentration-dependent induction of P450 2B activities was observed in the in vivo and hepatocyte culture studies. The in vivo ED50 values for P450 2B induction were approximately 110, approximately 100, and approximately 3000 dietary ppm (14 days administration) for PB, EPH, and EPO, respectively. The in vivo EC50 values for P450 2B induction were approximately 9, approximately 6, and approximately 130 microM (total serum) for PB, EPH, and EPO, respectively. In cultured rat hepatocytes, the ED50 values for induction of (benzyloxy)resorufin O-dealkylation activity were 14.5, 14.2, and 108 microM for PB, EPH, and EPO, respectively. These data indicate that pharmacodynamic results obtained with cultured hepatocytes represent a good qualitative and quantitative approximation of the in vivo hepatic responses in male rats caused by PB-type inducers.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential regulation of cytochrome(s) P450 2B1/2 by phenobarbital in hepatic hyperplastic nodules induced by aflatoxin B 1 or diethylnitrosamine plus 2-acetylaminofluorene in male F344 rats

Phenobarbital (PB) is an effective growth stimulator of hepatic hyperplastic nodules developed with diethylnitrosamine and 2-acetylaminofluorene plus partial hepatectomy (the Solt-Farber model), but it does not apparently stimulate the growth of preneoplastic lesions produced with aflatoxin B 1 (AFB). Some studies have suggested a correlation between the induction of specific cytochrome P450 enzymes and the tumor promoting effects produced by repeated treatment with PB. To examine this hypothesis further, hepatic hyperplastic nodules were produced with AFB (10 ip doses of AFB, 150 μg/kg/day, followed by partial hepatectomy) or by a modified Solt-Farber protocol (DEN/AAF), and the effects of PB on nodule growth and expression of cytochrome(s) P450 2B1 and/or P450 2B2 (P450 2B1/2) were determined. Both treatment protocols (without PB) produced multiple, large nodules within 10–17 weeks of carcinogen administration. These nodules stained intensely for glutathione S-transferase p (GST-p; GST7-7) and γ-glutamyl transpeptidase (GGT) and weakly for P450 2B1/2. Pentoxyphenoxazone dealkylation activity was decreased to less than 50% of the surrounding tissue levels in both types of nodules. PB treatment of animals with DEN/AAF-induced nodules greatly increased P450 2B1/2 expression in surrounding tissues, whereas most, but not all, nodules were not inducible. Pentoxyphenoxazone dealkylation was increased 31- to 35-fold in surrounding tissue, but it was increased only 2-fold in pooled nodular tissue, relative to untreated control liver. In contrast to the DEN/AAF group, immunohistochemical staining and pentoxyphenoxazone dealkylation in the AFB group demonstrated that P450 2B1/2 was equally inducible in nodular and surrounding tissues. Short-term treatment (5 days) with PB produced a 2-fold increase in the number and total area of GGT-positive nodules in the DEN/AAF group, but it had no significant effect on the number, size distribution, or total area of GGT-positive nodules in the AFB group. All large GGT-positive nodules in the DEN/AAF group were nonresponsive to induction of P450 2B 1/2, whereas all of the GGT-positive nodules which were responsive to P450 2B1/2 induction by PB in this group were relatively small. The size and area of AFB-induced GGT-positive nodules was not affected by PB treatment, and P450 2B1/2 in all of these nodules was inducible by PB. Although a causal, inverse relationship between the responsiveness of nodules to PB induction of P450 2B1/2 and their reaction to PB growth stimulation cannot be firmly established, these data are consistent with such a hypothesis.