Abstract
Sirolimus is an immunosuppressive drug currently used alone or in combination with cyclosporine. Both drugs undergo extensive metabolism by the CYP 3A enzymes. This study aimed at comparing the activity of recombinant CYP (rCYP) 3A4 and 3A5 toward sirolimus, investigating the effect of cyclosporine on the metabolic rate of these two cytochromes P450 (P450s), as well as the impact of the CYP 3A5*3 polymorphism on that of human liver microsomes (HLMs). Two distinct approaches were used; i.e., the measurement of (1) hydroxy-sirolimus and desmethyl-sirolimus production, and (2) sirolimus depletion by the in vitro half-life method. rCYP 3A5 exhibited a lower intrinsic clearance (CLint) for both hydroxylation (0.11 versus 0.24 μl/pmol P450/min) and depletion of sirolimus (0.64 versus 2.36 μl/pmol P450/min) than rCYP 3A4. Similar CLint values for hydroxylation, demethylation, and depletion were found when comparing a pool of HLMs carrying at least one CYP 3A5*1 (active) allele with a pool of HLMs not expressing CYP 3A5. This was further confirmed for sirolimus depletion using individual microsome preparations (p = 0.42). A deeper inhibitory effect of cyclosporine on the CLint of sirolimus depletion was found for rCYP 3A4 than for rCYP 3A5 (i.e., –44% versus –8% at 0.62 μM, 750 μg/l cyclosporine), and sirolimus metabolism was slightly less inhibited for HLMs expressing CYP 3A5 than not (–38% versus –56%). In the absence of cyclosporine, the CYP 3A5*3 polymorphism may not influence significantly sirolimus metabolism at the hepatic level. However, strong CYP 3A4 inhibition by cyclosporine could unveil the influence of this polymorphism.
Footnotes
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This study was funded by the Limousin Regional Council.
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.106.012161.
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ABBREVIATIONS: HLM, human liver microsome; P450, cytochrome P450; rP450 recombinant cytochrome P450; TFC-MS/MS: turbulent flow chromatography-tandem mass spectrometry; CLint, intrinsic clearance; MRM, multireaction monitoring; SNP, single nucleotide polymorphism.
- Received July 21, 2006.
- Accepted December 5, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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