Abstract
Lapatinib, an oral tyrosine kinase inhibitor used for breast cancer, has been reported to cause idiosyncratic hepatotoxicity. Recently, it has been found that lapatinib forms a metabolite-inhibitor complex (MIC) with CYP3A4 via the formation of an alkylnitroso intermediate. Because CYP3A5 is highly polymorphic compared with CYP3A4 and also oxidizes lapatinib, we investigated the interactions of lapatinib with CYP3A5. Lapatinib inactivated CYP3A5 in a time-, concentration-, and NADPH-dependent manner using testosterone as a probe substrate with KI and kinact values of 0.0376 mM and 0.0226 min−1, respectively. However, similar results were not obtained when midazolam was used as the probe substrate, suggesting that inactivation of CYP3A5 by lapatinib is site-specific. Poor recovery of CYP3A5 activity postdialysis and the lack of a Soret peak confirmed that lapatinib does not form a MIC with CYP3A5. The reduced CO difference spectrum further suggested that a large fraction of the reactive metabolite of lapatinib is covalently adducted to the apoprotein of CYP3A5. GSH trapping of a reactive metabolite of lapatinib formed by CYP3A5 confirmed the formation of a quinoneimine-GSH adduct derived from the O-dealkylated metabolite of lapatinib. In silico docking studies supported the preferential formation of an O-dealkylated metabolite of lapatinib by CYP3A5 compared with an N-hydroxylation reaction that is predominantly catalyzed by CYP3A4. In conclusion, lapatinib appears to be a mechanism-based inactivator of CYP3A5 via adduction of a quinoneimine metabolite.
Footnotes
This work was supported by the National University of Singapore, Academic Research Fund Tier 1 [Grants R-148-000-13-112, R-148-000-117-133, R-148-000-136-112]; and National University of Singapore Department of Pharmacy [Final Year Project Grant R-148-000-003-001].
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
↵ The online version of this article (available at http://dmd.aspetjournals.org) contains supplemental material.
ABBREVIATIONS:
- P450
- cytochrome P450
- MIC
- metabolite-intermediate complex
- ACN
- acetonitrile
- r
- recombinant
- DMSO
- dimethyl sulfoxide
- IS
- internal standard
- LC
- liquid chromatography
- MS/MS
- tandem mass spectrometry
- ESI
- electrospray ionization
- MS
- mass spectrometry
- MRM
- multiple reaction monitoring
- PIS
- product ion scanning
- MOE
- Molecular Operating Environment
- MBI
- mechanism-based inactivator
- rCYP3A5
- recombinant P450 3A5 Supersomes.
- Received February 6, 2012.
- Accepted April 10, 2012.
- Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics
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