ABSTRACT
Vinorelbine (VRL) (IV Navelbine) is a semi-synthetic vinca alkaloid, used in therapeutics for the treatment of non-small-cell lung cancer and advanced breast cancer. The aim of this study was to characterize the cytochrome P450 (CYP) isoenzymes involved in VRL metabolism. VRL was incubated at 1.28 x 10(-5) m for 90 min with human hepatic microsomes prepared from 14 donors (one woman and 13 men aged from 27 to 76 years old) and characterized for CYP1A2, CYP2D6, CYP2E1 and CYP3A4 activities. A four-combined-approach study was performed, including correlation between CYP activities and VRL metabolism among the 14 batches of microsomes, inhibition of VRL biotransformation by isoform-selective substrates and by specific inhibitory antibodies, and incubation with supersomes. Analysis of unchanged VRL and its metabolites was performed using an HPLC method coupled with both radioactive and UV detections. No correlation between CYP1A2 or CYP2E1 and VRL metabolism was observed in the 14 batches of microsomes used. A correlation was shown between VRL metabolism and CYP3A4 activity as determined with the dextromethorphan N-demethylase and nifedipine oxidase activities (r(2)=0.80 and 0.59, respectively). These results were strengthened by a correlation between the metabolism extent of VRL and CYP3A4 protein content determined by immunoblotting (r(2)=0.75). Furthermore, VRL biotransformation was inhibited by troleandomycine, the CYP3A4-specific inhibitor substrate (80% of inhibition) and by anti-CYP3A antibodies (36% of inhibition). On the contrary, a low correlation with CYP2D6 activity as determined by dextrometorphan O-demethylation (r(2)=0.31) was established. CYP2D6 supersomes did not metabolize the drug whereas 63.4% of VRL were metabolized by microsomes overexpressing CYP3A4 isoform. These data indicated that CYP3A4 is the main enzyme involved in the hepatic metabolism of VRL in human, whereas CYP2D6 is not involved.
Subject(s)
Cytochrome P-450 CYP2D6/metabolism , Cytochrome P-450 Enzyme System/metabolism , Vinblastine/analogs & derivatives , Adult , Aged , Animals , Antibodies/pharmacology , Antineoplastic Agents, Phytogenic/metabolism , Blotting, Western , Cytochrome P-450 CYP2D6/immunology , Cytochrome P-450 CYP2D6 Inhibitors , Cytochrome P-450 CYP3A , Cytochrome P-450 Enzyme Inhibitors , Cytochrome P-450 Enzyme System/immunology , Enzyme Inhibitors/pharmacology , Female , Humans , Isoenzymes/antagonists & inhibitors , Isoenzymes/immunology , Isoenzymes/metabolism , Ketoconazole/pharmacology , Male , Microsomes, Liver/drug effects , Microsomes, Liver/metabolism , Middle Aged , Time Factors , Tritium , Troleandomycin/pharmacology , Vinblastine/metabolism , VinorelbineABSTRACT
Milnacipran is an antidepressant drug belonging to the class of serotonin and noradrenaline reuptake inhibitors. A sensitive high performance liquid chromatographic during the development method coupled with a fluorimetric detection was set up, validated and then used routinely of the drug. After liquid-liquid extraction, milnacipran and its internal standard were analyzed by reversed-phase liquid chromatography (LC). The drug was derivatized with fluorescamine for fluorescence detection. The identity of the liquid chromatography peaks was controlled using mass spectrometry. The assay linearity was validated up to 1000 ng/ml. The limit of quantification was set at 5 ng/ml. Precision values (relative standard deviations) were lower than 5.4%, whereas the mean accuracy was higher than 95%. The extraction recoveries were higher than 70% for both milnacipran and the internal standard. In clinics, the LC-fluorescence method was routinely used to investigate the pharmacokinetics of milnacipran in patients and proved to be robust and capable of quantifying milnacipran in plasma for at least 36 h (four- to five-fold the elimination half-life).
