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Identification of Uridine 5′-Diphosphate-Glucuronosyltransferases Responsible for the Glucuronidation of Mirabegron, a Potent and Selective β3-Adrenoceptor Agonist, in Human Liver Microsomes

  • Kentaro Konishi
  • Daisuke Tenmizu
  • Shin Takusagawa
Original Research Article

Abstract

Background and Objectives

Mirabegron is cleared by multiple mechanisms, including drug-metabolizing enzymes. One of the most important clearance pathways is direct glucuronidation. In humans, M11 (O-glucuronide), M13 (carbamoyl-glucuronide), and M14 (N-glucuronide) have been identified, of which M11 is one of the major metabolites in human plasma. The objective of this study was to identify the uridine 5′-diphosphate (UDP)-glucuronosyltransferase (UGT) isoform responsible for the direct glucuronidation of mirabegron using human liver microsomes (HLMs) and recombinant human UGTs (rhUGTs).

Methods

Reaction mixtures contained 1–1000 μM mirabegron, 8 mM MgCl2, alamethicin (25 μg/mL), 50 mM Tris–HCl buffer (pH 7.5), human liver microsome (HLM) or rhUGT (1.0 mg protein/mL), and 2 mM UDP-glucuronic acid in a total volume of 200 μL for 120 min at 37 °C. HLMs from 16 individuals were used for the correlation study, and mefenamic acid and propofol were used for the inhibition study.

Results

Regarding M11 formation, rhUGT2B7 showed high activity among the rhUGTs tested (11.3 pmol/min/mg protein). This result was supported by the correlation between M11 formation activity and UGT2B7 marker enzyme activity (3-glucuronidation of morphine, r 2 = 0.330, p = 0.020) in individual HLMs; inhibition by mefenamic acid in pooled HLMs (IC50 = 22.8 μM); and relatively similar K m values between pooled HLMs and rhUGT2B7 (1260 vs. 486 μM). Regarding M13 and M14 formation, rhUGT1A3 and rhUGT1A8 showed high activity among the rhUGTs tested, respectively.

Conclusions

UGT2B7 is the main catalyst of M11 formation in HLMs. Regarding M13 and M14 formation, UGT1A3 and UGT1A8 are strong candidates for glucuronidation, respectively.

Notes

Acknowledgements

The authors would like to sincerely thank Mr. Tadashi Hashimoto, Dr. Kiyoshi Noguchi, and Dr. Takashi Usui for their useful suggestions about the experiments and Dr. Toshifumi Shiraga, Dr. Tsuyoshi Minematsu, Dr. Yasuhisa Nagasaka, Dr. Takafumi Iwatsubo, Dr. Yoichi Naritomi, Mr. Aiji Miyashita, and Dr. Kenji Tabata for their contribution to this study.

Author Contributions

Participated in research design Mr. Kentaro Konishi, Dr. Daisuke Tenmizu, and Dr. Shin Takusagawa. Conducted experiments Mr. Kentaro Konishi. Performed data analysis Mr. Kentaro Konishi, Dr. Daisuke Tenmizu, and Dr. Shin Takusagawa. Wrote or contributed to the writing of the manuscript Mr. Kentaro Konishi, Dr. Daisuke Tenmizu, and Dr. Shin Takusagawa

Compliance with Ethical Standards

Conflict of interest

Mr. Kentaro Konishi, Dr. Daisuke Tenmizu and  Dr. Shin Takusagawa are employees of Astellas Pharma, Japan.

Funding

This study was sponsored by Astellas Pharma, Japan. Editorial support was funded by Astellas Pharma, Japan.

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Copyright information

© Springer International Publishing AG, part of Springer Nature 2017

Authors and Affiliations

  1. 1.Analysis and Pharmacokinetics Research Laboratories, Drug Discovery ResearchAstellas Pharma Inc.Tsukuba-shiJapan
  2. 2.Clinical Pharmacology, DevelopmentAstellas Pharma Inc.TokyoJapan

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