UNLABELLED: Background: Warfarin, an antagonist of vitamin K, is an oral coumarin anticoagulant widely used to control and prevent thromboembolic disorders. Warfarin is clinically available as a racemic mixture of R- and S-warfarin. The S-enantiomer has three to five times greater anticoagulation potency than its optical congener. Recently, vitamin K2 function has been proposed via the pregnane X receptor (PXR) in osteocytes. PXR acts as a xenobiotic sensor that controls expression of many genes involved in drug/xenobiotic metabolic clearance. OBJECTIVE: The aim was to examine whether enantiomers of warfarin stereoselectively interact with PXR to up-regulate main drug/xenobiotic-metabolizing enzymes of the cytochrome P450 superfamily. METHODS: Interactions of warfarin enantiomers with PXR were tested by gene reporter assays and time-resolved fluorescence resonance energy transfer technology (TR-FRET) ligand binding assay. Up-regulation of PXR-target gene mRNAs by warfarin enantiomers was studied using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) in primary human hepatocytes. RESULTS: We found that R-warfarin interacts with the PXR nuclear receptor. Consistently, R-warfarin significantly induced CYP3A4 and CYP2C9 mRNAs in cultures of primary human hepatocytes or in LS174T intestinal cells. On the other hand, S-warfarin is a less potent inducer of PXR-target genes in human hepatocytes and activates PXR only at supraphysiological concentrations. In addition, we showed that racemic 10- and 4'-hydroxywarfarins are also highly potent PXR ligands and inducers of CYP3A4 and CYP2C9 mRNA in human hepatocytes. CONCLUSION: We showed that R-warfarin can significantly up-regulate major drug-metabolizing enzymes CYP3A4 and CYP2C9 in the liver and thus may cause drug-drug interactions (DDI) with co-administered drugs. The results warrant reconsideration of racemic warfarin usage in clinics.

Department of Cell Biology and Genetics Faculty of Science Palacky University in Olomouc Slechtitelu Olomouc

Department of Pharmacology and Toxicology Faculty of Pharmacy in Hradec Kralove Charles University Prague Heyrovskeho Hradec Kralove

Laboratory of Molecular Pharmacology Institute of Molecular Genetics Academy of Sciences of the Czech Republic Videnska Prague Czech Republic

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