Received for publication January 13, 2006.
Revised June 2, 2006.
Accepted for publication June 7, 2006.

WHICH HYDROXY? EVIDENCE FOR SPECIES DIFFERENCES IN THE REGIOSELECTIVITY OF GLUCURONIDATION IN RAT, DOG AND HUMAN IN VITRO SYSTEMS AND DOG IN VIVO

Abstract

The glucuronidation of (1S,2R,3R,5R)-3-(hydroxymethyl)-5-[7-{[(1R,2S)-2-phenylcyclopropyl]amino}-5-(propylthio)-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-yl]cyclopentane-1,2-diol (AZ11939714) was studied in UDPGA-supplemented hepatic microsomes from rat, dog and human liver. The major biliary metabolite of this compound following intraduodenal administration to a beagle dog was also studied. The techniques of HPLC, HPLC-MS and HPLC-NMR were used to characterise the glucuronides. An analysis of the proton NMR chemical shift differences between parent and metabolites was sufficient to deduce the sites of glucuronidation, though these were confirmed by 2D ROESY experiments. In dog microsomes, AZ11939714 was O-glucuronidated exclusively at the 1-position of the cyclopentanediol. This glucuronide was also the major metabolite in dog bile. In human microsomes, AZ11939714 was O-glucuronidated almost exclusively at the 3-hydroxymethyl position. Rat microsomes produced a mixture of glucuronides at the 2-position of the cylopentanediol (major) and at the 3-hydroxymethyl position (minor). A clear qualitative species difference in the glucuronidation of AZ11939714 has been demonstrated in vitro. This may have implications for the choice of laboratory species to study the pharmacokinetics and safety of this compound.

Key words: glucuronidation, mass spectrometry, metabolite identification