Atypical Kinetics and Albumin Effect of Glucuronidation of 5-n-Butyl-4-{4-[2-(1H-tetrazole-5- yl)-1H-pyrrol-1-yl]phenylmethyl}-2,4-dihydro-2-(2,6- dichlorophenyl)-3H-1,2,4-triazol-3-one, a Novel Nonpeptide Angiotensin Type 1 Receptor Antagonist, in Liver Microsomes and UDP-Glucuronosyl-transferase

Abstract

Ib is a new nonpeptide AT1 receptor antagonist, which plays an active role in cardiovascular protection. Ib monoglucuronide has been identified as its main metabolite. A detailed study of Ib glucuronidation is important for predicting potential DDI. Besides, the elucidation of the “BSA effect” in Ib glucuronidation would make obtained kinetic parameters more predictive in IVIVE. “BSA effect” means that there is a significant change in in vitro kinetic parameters when generated from incubations performed in the presence of bovine serum albumin (BSA). Five UGTs (UGT1A3, UGT2B4, UGT2B7, UGT1A9 and UGT1A8) were identified that produced abundant Ib monoglucuronide, especially UGT1A3. We investigated Ib glucuronidation in liver microsomes from different species (rat, dog, human) and in five identified major human UGTs. Ib glucuronidation in liver microsomes and recombinant human UGTs all showed substrate inhibition kinetics. DLM showed the strongest affinity and activity, HLM showed the lowest affinity, and RLM showed the weakest activity. The addition of BSA did not alter the enzyme kinetics, but significantly altered enzyme kinetic parameters resulting in a reduction in Km value and an increase in CLint value. However, high concentrations of BSA could significantly attenuate this positive effect on enzyme affinity and activity, and the effect of BSA on the Vmax of Ib glucuronidation was opposite in different enzyme sources. In conclusion, this study demonstrated the substrate inhibition kinetics of Ib glucuronidation in the liver metabolism and the effect of BSA on its kinetic parameters, in order to provide more accurate in vitro data for in vivo prediction.