Abstract
3′-Azido-3′-deoxythymidine (AZT) is frequently prescribed to patients infected with the human immunodeficiency virus. After absorption, AZT is rapidly metabolized into 3′-azido-3′-deoxy-5′-glucuronylthymidine by UDP-glucuronosyltransferase (UGT) enzymes. Using labeled [14C]UDP-glucuronic acid and microsomal preparations from human kidney 293 cells stably expressing the different human UGT2B isoenzymes, it was demonstrated that AZT glucuronidation is catalyzed specifically by human UGT2B7. The identity of the metabolite formed was confirmed as AZT-G by liquid chromatography coupled with mass spectrometry. UGT2B7 is encoded by a polymorphic gene and kinetic analysis of AZT glucuronidation by the two allelic variants UGT2B7(H268) and UGT2B7(Y268), yielded apparent Km values of 91.0 and 80.1 μM, respectively. Normalization to protein levels yielded glucuronidation efficiency ratios (Vmax/Km) of 21.3 and 11.0 μl · min−1 · mg protein−1for UGT2B7(H268) and UGT2B7(Y268), respectively. It remains possible that other UGT enzymes are also involved in AZT conjugation; however, the glucuronidation of AZT by UGT2B7, which is a UGT2B protein expressed in the liver, is consistent with previous findings and supports the physiological relevance of this enzyme in AZT conjugation.
Footnotes
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Send reprint requests to: Dr. Alain Bélanger or Dr. Dean W. Hum, Laboratory of Molecular Endocrinology, CHUL Research Center, 2705 Laurier Blvd., Quebec G1V 4G2, Canada. E-mail:Alain.Bélanger{at}crchul.ulaval.ca orDean.Hum{at}crchul.ulaval.ca
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↵1 These authors contributed equally to this work.
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This work was supported by the Medical Research Council of Canada (A.B. and D.W.H.), the Fonds de la Recherche en Santé du Québec (A.B. and D.W.H.), Endorecherche and the National Institutes of Health (GM26221 to T.R.T.). Olivier Barbier and David Turgeon are holders of scholarships from La Fondation de l'Université Laval, and Caroline Girard is holder of a Summer Research Fellowship from the American Endocrine Society.
- Abbreviations used are::
- HIV-RT
- HIV-reverse transcriptase
- AZT
- 3′-azido-3′-deoxythimidine
- AZT-G
- 3′-azido-3′-deoxy-5′-glucuronylthymidine
- AZT-TP
- AZT-5′-triphosphate
- UDPGA
- UDP-glucuronic acid
- HK293
- human kidney 293
- UGT
- UDP-glucuronosyltransferase
- LC-MS/MS
- liquid chromatography coupled with mass spectrometry
- Received December 10, 1999.
- Accepted February 16, 2000.
- The American Society for Pharmacology and Experimental Therapeutics
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