Hepatic biotransformation of docetaxel (Taxotere) in vitro: involvement of the CYP3A subfamily in humans.

Docetaxel metabolism mediated by cytochrome P450-dependent monooxygenases was evaluated in human liver microsomes and hepatocytes. In microsomes, the drug was converted into four major metabolites resulting from successive oxidations of the tert-butyl group on the synthetic side chain. Enzyme kinetics appeared to be biphasic with a V(max) and apparent K(m) for the high-affinity site of 9.2 pmol/min/mg and 1.1 microm, respectively. the intrinsic metabolic clearance in human liver microsomes (V(max)/K(m), 8.4 ml/min/g protein) was comparable to that in rat and dog liver microsomes, but lower in mouse liver microsomes. Although the metabolic profile was identical in all subjects, a large quantitative variation in docetaxel biotransformation rates was found in a human liver microsome library, with a ratio of 8.9 in the highest:lowest biotransformation rates. Docetaxel biotransformation was correlated significantly (0.7698; P < 0.0001) with erythromycin N-demethylase activity, but not with aniline hydroxylase or debrisoquine 4-hydroxylase. It was inhibited, both in human hepatocytes and in liver microsomes, by typical CYP3A substrates and/or inhibitors such as erythromycin, ketoconazole, nifedipine, midazolam, and troleandomycin. Docetaxel metabolism was induced in vitro in human hepatocytes by dexamethasone and rifampicin, both classical CYP3A inducers. These data suggest a major role of liver cytochrome P450 isoenzymes of the CYP3A subfamily in docetaxel biotransformation in humans. Finally, some Vinca alkaloids and doxorubicin were shown to inhibit docetaxel metabolism in human hepatocytes and liver microsomes. These findings may have clinical implications and should be taken into account in the design of combination cancer chemotherapy regimens.

Predictive performance of two software packages (USC*PACK PC and Abbott PKS system) for the individualization of amikacin dosage in intensive care unit patients.

Many dosing methods (nomogram, pharmacokinetic methods, Bayesian methods) can be used for the individualization of amikacin dosing. Among these methods, it is now well known that the Bayesian method provides a rapid and accurate means for individualizing dosage requirements for patients with diverse pharmacokinetic profiles. However, one problem has not been fully resolved. Should we use population-based parameters reflecting the patient population being monitored or should we used general population parameters? The aim of this study was to answer this question using two widely used software programs (USC*PACK PC and Abbott PKS system) and two different population parameters sets. Predictive performance of these methods was assessed with respect to the prediction of amikacin serum concentrations in intensive care unit (ICU) patients. Our results show that the differences between predicted and measured concentrations were unbiased when the population parameters used were adequate. Precision values were comparable with previously reported values. The predictive performance of the two tested software programs are very comparable in ICU patients. In addition, we demonstrated that performance can be enhanced when using population-based parameters which reflect the patient population being monitored. It is therefore advisable for each user to properly characterize each particular patient population.

Involvement of the cytochrome P-450IID subfamily in minaprine 4-hydroxylation by human hepatic microsomes.

4-Hydroxylation of minaprine was measured on microsomal fractions prepared from 25 different human liver samples. In vitro formation of 4-hydroxyminaprine exhibited a large interindividual variability. Indeed, minaprine 4-hydroxylase activity ranged between 0.033 and 0.421 nmol/min/mg microsomal protein. Two samples presented a particularly low enzyme activity. Minaprine 4-hydroxylation followed Michaelis-Menten kinetics with KM and Vmax values of 5.26 microM and 0.478 nmol/min/mg microsomal protein, respectively, for one particular representative sample. The effects of various compounds (substrates or inhibitors of cytochrome P-450 isoforms) on 4-hydroxyminaprine formation were investigated. Selective substrates for P-450IA [benzo(a)pyrene, theophylline, and phenacetin], IIC (hexobarbital), IIE (aniline), and IIIA (erythromycin, nifedipine, and troleandomycin) cytochrome subfamilies did not inhibit 4-hydroxyminaprine formation. The nonspecific cytochrome P-450 inhibitor, cimetidine, slightly inhibited minaprine 4-hydroxylation. The classical substrates of the P-450IID cytochrome subfamily (debrisoquine, propranolol, and sparteine) inhibited minaprine 4-hydroxylation, as did the known P-450IID specific inhibitor, quinidine. These compounds inhibited minaprine 4-hydroxylase with Ki values of 16.5 (debrisoquine), 14.4 (propranolol), 61.9 (sparteine), and 0.146 microM (quinidine). 4-Hydroxyminaprine formation rate was shown not to be correlated with the activity of both erythromycin N-demethylase (r = 0.29, non-significant) and aniline hydroxylase (r = -0.15, NS). In contrast, minaprine 4-hydroxylase was well correlated with both debrisoquine 4-hydroxylase activity (r = 0.501, p less than 0.05) and immunoquantified cytochrome P-450IID6 (r = 0.579, p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolism of minaprine in human and animal hepatocytes and liver microsomes--prediction of metabolism in vivo.

1. The metabolism of minaprine and its major metabolite p-hydroxyminaprine were studied using hepatocytes and liver microsomes from different species. Metabolism of this drug in vitro was then compared with in vivo data already published. 2. Our results showed that the major metabolic route (4-hydroxylation of the aromatic ring) is the same in the two experimental systems. Other in vivo biotransformation pathways (i.e. N-oxidation, reductive ring cleavage, N-dealkylation, oxidation) were also confirmed in hepatocytes. 3. Similar inter-species variability was observed both in vitro and in vivo. The present study has led to the same conclusion as previous in vivo metabolic investigations, namely, that metabolism in the dog quantitatively differs from that observed in other animal species. 4. These results clearly demonstrate that in vitro models (i.e. isolated hepatocytes and liver microsomes) are powerful tools in predicting the metabolic pathways of a drug in man and animal species.

Use of human and animal liver microsomes in drug metabolic studies.

A bank of readily available well-characterized human and animal hepatic microsomal fractions has been established. By using these "in vitro" models, we evidenced large interspecies variabilities for various compounds including digoxin, minaprine and two vincaalkaloïds (navelbine, vinblastine). Therefore, extrapolation from animal to human appeared limited and we focused our interest on human liver microsomes. Enzymatic characteristics of human microsomes from 35 different livers were determined using specific monooxygenase (i.e. erythromycin, aniline, aminopyrin...) and UDP-glucuronosyltransferase substrates (i.e. p-nitrophenol, monodigitoxoside digitoxigenin...). A wide variability was thus ascertained between individual for both phase I and phase II metabolic processes. Microsomal fractions were also shown to be of great interest for assessing the P450 cytochrome isoform(s) involved in the biotransformation of a given drug. For instance, using inhibitory experiments, we showed the implication of P450IID in minaprine metabolism. We also demonstrated that P450IIIA is probably involved in vindesine biotransformation. Drug metabolic interactions between cyclosporin A and macrolides were studied using the same model. These results demonstrating that erythromycin is a much more potent inhibitor of cyclosporin A biotransformation than spiramycin, agree closely with "in vivo" data. In conclusion, liver microsomes are powerful tools in studying: i) interspecies and interindividual variabilities, ii) metabolic drug interactions.