Regio- and stereospecific N-glucuronidation of medetomidine:: The differences between UDP glucuronosyltransferase (UGT) 1A4 and UGT2B10 account for the complex kinetics of human liver microsomes

Medetomidine is a chiral imidazole derivate whose dextroenantiomer is pharmacologically active. The major metabolic pathway of dexmedetomidine [(+)-4-(S)-[1-(2,3-dimethylphenyl)ethyl]-1H-imidazole] in humans is N-glucuronidation at the imidazolate nitrogens. We have purified the N-3- and N-1 -glucuronides of dexmedetomidine, termed DG1 and DG2, respectively, according to their elution order in liquid chromatography and determined their structure by H-1 nuclear magnetic resonance (NMR). Studying medetomidine glucuronidation by human liver microsomes (HLMs) and recombinant UDP glucuronosyltransferase (UGT) 1A4 indicated that another human UGT plays a major role in these activities. We now demonstrate that this enzyme is UGT2B10. HLMs catalyzed DG1 and DG2 formation, at a ratio of 3: 1, with two-enzyme kinetics that contain both a high-affinity component, K-m1 values of 6.6 and 8.7 mu M, and a low-affinity component, K-m2 values > 1 mM. The DG1/DG2 ratio in the case of UGT2B10 was lower, 1.4:1, whereas the substrate affinity for both reactions was high, K-m values of 11 and 16 mu M. UGT1A4 produced mainly DG1 (DG1/DG2 ratio of 6.6: 1) at low substrate affinities, K-m values above 0.6 mM, but superior expression-normalized V-max values. Levomedetomidine [( -)-4-( R)-[ 1-(2,3-dimethylphenyl)ethyl]-1H-imidazole] glucuronidation by HLMs yielded mostly the N-3- glucuronide (LG1, structure determined by NMR), with monophasic kinetics and a K-m value of 14 mu M. The activity of UGT1A4 toward levomedetomide was low and generated both LG1 and LG2, whereas UGT2B10 exhibited relatively high activity and sharp regioselectivity, yielding only LG1, with a K-m value of 7.4 mu M. The results highlight the contribution of UGT2B10 to medetomidine glucuronidation and its potential importance for other N-glucuronidation reactions within the human liver.