Received for publication September 22, 2006.
Revised November 16, 2006.
Accepted for publication December 1, 2006.

The glucuronidation of 4-3-keto C19- and C21- hydroxy-steroids by human liver microsomal and recombinant UDP-glucuronosyltransferases (UGT): 6- and 21- hydroxyprogesterone are selective substrates for UGT2B7

Abstract

The stereo- and regio- selective glucuronidation of ten ⁴-3-keto mono hydroxylated androgens and pregnanes was investigated in order to identify UGT enzyme selective substrates. Kinetic studies were performed using human liver microsomes (HLM) and a panel of 12 recombinant human UGTs as the enzyme sources. Five of the steroids, which were hydroxylated in the 6, 7-, 11- or 17- positions, were not glucuronidated by HLM. Of the remaining compounds, comparative kinetic and inhibition studies indicated that 6- and 21- hydroxyprogesterone (6- and 21- OHP) were glucuronidated selectively by human liver microsomal UGT2B7. 6-OHP glucuronidation by HLM and UGT2B7 followed Michaelis-Menten kinetics, whereas 21-OHP glucuronidation by these enzyme sources exhibited positive cooperativity. UGT2B7 was also identified as the enzyme responsible for the high affinity component of human liver microsomal 11- hydroxyprogesterone (11-OHP) glucuronidation. In contrast, UGT2B15 and UGT2B17 were the major forms involved in human liver microsomal testosterone 17-glucuronidation and the high affinity component of 16- hydroxyprogesterone (16-OHP) glucuronidation. Activity of UGT1A subfamily enzymes towards the hepatically glucuronidated substrates was generally low, although UGT1A4 and UGT1A9 contribute to the low affinity components of microsomal 16-OHP and 11-OHP glucuronidation, respectively. Interestingly, UGT1A10, which is expressed only in the gastrointestinal tract, exhibited activity towards most of the glucuronidated substrates. The results indicate that 6- and 21- OHP may be used as selective 'probes' for human liver microsomal UGT2B7 activity and, taken together, provide insights into the regio- and stereo- selectivity of hydroxy-steroid glucuronidation by human UGTs.

Key words: glucuronidation, in vitro-in vivo prediction, phase II drug metabolism, steroids, UDP glucuronyltransferases

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