title: Studies on the role of metabolic activation in tyrosine kinase inhibitor-dependent hepatotoxicity: induction of CYP3A4 enhances the cytotoxicity of lapatinib in HepaRG cells.
abstract: Idiosyncratic hepatotoxicity has been associated with the oral tyrosine kinase inhibitor lapatinib, which is used in metastatic breast cancer therapy. Lapatinib is extensively metabolized by cytochrome P450 3A4/5 to yield an O-debenzylated metabolite, which can undergo further oxidation to a reactive quinone imine. A recent clinical study reported that concomitant use of lapatinib with dexamethasone increased the incidence of hepatotoxicity in metastatic breast cancer patients treated with lapatinib, and so we hypothesized that induction of CYP3A enhances the bioactivation of lapatinib to reactive intermediates that contribute to hepatotoxicity. Therefore, we examined the effect of CYP3A4 induction on the cytotoxicity and metabolism of lapatinib in the HepaRG human hepatic cell line. Differentiated HepaRG cells were pretreated with dexamethasone (100 μM) or the prototypical CYP3A4 inducer rifampicin (4 μM) for 72 hours, followed by incubation with lapatinib (0-100 μM) for 24 hours. Cell viability was monitored using WST-1 assays, and metabolites were quantified by liquid chromatography coupled to tandem mass spectrometry. Induction of CYP3A4 by dexamethasone or rifampicin enhanced lapatinib-induced cytotoxicity, compared with treatment with lapatinib alone. A direct comparison of the cytotoxicity of lapatinib versus O-debenzylated lapatinib demonstrated that the O-debenzylated metabolite was significantly more cytotoxic than lapatinib itself. Furthermore, pretreatment with 25 μM l-buthionine sulfoximine to deplete intracellular glutathione markedly enhanced lapatinib cytotoxicity. Cytotoxicity was correlated with increased formation of O-debenzylated lapatinib and cysteine adducts of the putative quinone imine intermediate. Collectively, these data suggest that CYP3A4 induction potentiates lapatinib-induced hepatotoxicity via increased reactive metabolite formation.
authors.affiliation: Departments of Medicinal Chemistry (K.D.H., M.D.W., A.E.R, S.D.N.) and Pharmaceutics (I.P., J.D.U.), University of Washington School of Pharmacy, Seattle, Washington.
authors.forename: Klarissa D
authors.initials: KD
authors.lastname: Hardy
authors.forename: Michelle D
authors.initials: MD
authors.lastname: Wahlin
authors.forename: Ioannis
authors.initials: I
authors.lastname: Papageorgiou
authors.forename: Jashvant D
authors.initials: JD
authors.lastname: Unadkat
authors.forename: Allan E
authors.initials: AE
authors.lastname: Rettie
authors.forename: Sidney D
authors.initials: SD
authors.lastname: Nelson
mesh_terms: D001711:Biotransformation / Q000187:drug effects*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003545:Cysteine / Q000378:metabolism; D051544:Cytochrome P-450 CYP3A / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D000077341:Lapatinib; D008099:Liver / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011799:Quinazolines / Q000494:pharmacology; D012293:Rifampin / Q000494:pharmacology
chemical_list: D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D000077341:Lapatinib; D003907:Dexamethasone; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D011505:Protein-Tyrosine Kinases; D005978:Glutathione; D003545:Cysteine; D012293:Rifampin