Regioselectivity significantly impacts microsomal glucuronidation efficiency of R/S-6, 7-, and 8-hydroxywarfarin

Abstract

1. Coumadin (R/S-warfarin) metabolism plays a critical role in patient response to anticoagulant therapy. Several cytochrome P450s oxidize warfarin into R/S-6-, 7-, 8-, 10, and 4′-hydroxywarfarin that can undergo subsequent glucuronidation by UDP-glucuronosyltransferases (UGTs); however, current studies on recombinant UGTs cannot be adequately extrapolated to microsomal glucuronidation capacities for the liver.

2. Herein, we estimated the capacity of the average human liver to glucuronidate hydroxywarfarin and identified UGTs responsible for those metabolic reactions through inhibitor phenotyping. There was no observable activity toward R/S-warfarin, R/S-10-hydroxywarfarin or R/S-4′-hydroxywarfarin.

3. The observed metabolic efficiencies (V_max/K_m) toward R/S-6-, 7-, and especially 8-hydroxywarfarin indicated a high glucuronidation capacity to metabolize these compounds.

4. UGTs demonstrated strong regioselectivity toward the hydroxywarfarins. UGT1A6 and UGT1A1 played a major role in R/S-6- and 7-hydroxywarfarin glucuronidation, respectively, whereas UGT1A9 accounted for almost all of the generation of the R/S-8-hydroxywarfarin glucuronide.

5. In summary, these studies expanded insights to glucuronidation of hydroxywarfarins by pooled human liver microsomes, novel roles for UGT1A6 and 1A9, and the overall degree of regioselectivity for the UGT reactions.

Keywords:

• Warfarin
• hydroxywarfarin
• UGT1A1
• UGT1A6
• UGT1A9
• metabolism
• regioselectivity

Acknowledgements

Moreover, we thank Michael Court (Tufts University) for helpful discussions on UGT phenotyping assays.

Disclosure statement

The authors declare no conflicts of interest.

Additional information

Funding

This work was supported by a bridging grant from the University of Arkansas for Medical Sciences to G.P.M. and Chonnam National University Second Stage BK21 Project from the Ministry of Education, Science and Technology of the Republic of Korea to S.-Y.K. Additional support provided by through a Grant-in-Aid from the SouthWest Affiliate of the American Heart Association [13GRNT16960043] and a pilot study grant provided by the National Institutes of Health [UL1 TR000039] awarded to the University of Arkansas for Medical Sciences along with grants from the National Institutes of Health National Library of Medicine [R01LM012222 and R01LM012482]. The content of this manuscript is solely the responsibility of the authors and does not necessarily represent the official views of the University of Arkansas for Medical Sciences, the American Heart Association, or National Institutes of Health.