engleski

APPLICATION OF LIQUID CHROMATOGRAPHY FOR CHARACTERIZATION OF CYTOCHROME P450 MECHANISM-BASED INACTIVATION

Lapatinib, an oral breast cancer drug, can cause idiosyncratic hepatotoxicity that has been related to mechanism-based inactivation of cytochrome P450 3A. Chan et al. (2012) used glutathione as trapping agent of a reactive metabolite and confirmed quinoneimine metabolite of O-dealkylated lapatinib as mechanism-based inactivator. Similar approach using GSH related trapping agent was used to demonstrate protein adduct formation of quinone methides by P450 2B6 bioactivation of tamoxifen that was further confirmed by ESI-LC-MS of whole protein. Ritonavir, a protease inhibitor, causes mechanism-based inactivation of P450 3A4 through heme destruction that was attributed to isocyanate intermediate. High performance liquid chromatography represents basis of metabolism studies. Accompanied with other techniques like ion exchange chromatography (used for His-tagged cytochrome P450 purification), thin layer chromatography (determining metabolites of radioactive substrates), 16O2/18O2 incubations with subsequent high resolution MS analysis (mechanism of P450 catalysis, determining substrates of orphan P450s) etc., it represents the most powerful tool for studying of all aspects of cytochromes P450.

P450; inhibition; inactivation

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