Abstract
A thienopyridine antiplatelet agent, prasugrel, is rapidly hydrolyzed to a thiolactone metabolite (R-95913, 2-[2-oxo-6,7-dihydrothieno[3,2-c]pyridin-5(4H)-yl]-1-cyclopropyl-2-(2-fluorophenyl)ethanone). R-95913 is oxidized by hepatic cytochromes P450 to the pharmacologically active metabolite R-138727 (2-[1–2-cyclopropyl-1-(2-fluorophenyl)-2-oxoethyl]-4-mercapto-3-piperidinylidene]acetic acid). One possible intermediate in the in vitro bioactivation pathway is a glutathione conjugate, R-133490, which could be reduced to generate R-138727 in the presence of a reducing agent such as glutathione. In this study, enzymes in human liver cytosols were found to accelerate reduction of R-133490 leading to the formation of R-138727. To explore the possible reductive enzymes, we separated the various proteins in human liver cytosol based on size using gel filtration chromatography. Two active peaks were detected and found to contain thioredoxin and glutaredoxin, respectively. In addition, recombinant human glutaredoxin and thioredoxin promoted the formation of R-138727 from R-133490 with much higher activity for glutaredoxin than for thioredoxin. This study is the first in vitro observation indicating that glutaredoxin and thioredoxin in human liver are active in reducing the mixed disulfide formed between xenobiotics and glutathione.
Footnotes
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
doi:10.1124/dmd.110.035196.
-
ABBREVIATIONS:
- R-95913
- 2-[2-oxo-6,7-dihydrothieno[3,2-c]pyridin-5(4H)-yl]-1-cyclopropyl-2-(2-fluorophenyl)ethanone
- R-138727
- 2-[1–2-cyclopropyl-1-(2-fluorophenyl)-2-oxoethyl]-4-mercapto-3-piperidinylidene]acetic acid
- LC-MS/MS
- liquid chromatography equipped with tandem mass spectrometry
- HPLC
- high-performance liquid chromatography
- FPLC
- fast protein liquid chromatography
- PAGE
- polyacrylamide gel electrophoresis
- TBS
- Tris-buffered saline
- HRP
- horseradish peroxidase
- FA
- formic acid
- KPB
- potassium phosphate buffer
- CLint
- intrinsic clearance
- prasugrel
- 2-acetoxy-5-(α-cyclopropylcarbonyl-2-fluorobenzyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine.
- Received June 29, 2010.
- Accepted October 25, 2010.
- Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics
DMD articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|