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Bioactivation of GPR40 Agonist MK-8666: Formation of Protein Adducts in Vitro from Reactive Acyl Glucuronide and Acyl CoA Thioester

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posted on 2019-10-14, 19:40 authored by Jackie Shang, Richard Tschirret-Guth, Mark Cancilla, Koppara Samuel, Qing Chen, Harry R. Chobanian, Ann Thomas, Wei Tong, Hubert Josien, Alexei V. Buevich, Kaushik Mitra
MK-8666, a selective GPR40 agonist developed for the treatment of type 2 diabetes mellitus, was discontinued in phase I clinical trials due to liver safety concerns. To address whether chemically reactive metabolites played a causative role in the observed drug induced liver injury (DILI), we characterized the metabolism, covalent binding to proteins, and amino acid targets of MK-8666 in rat and human hepatocytes or cofactor-fortified liver microsomes. MK-8666 was primarily metabolized to an acyl glucuronide in hepatocytes of both species and a taurine conjugate in rat hepatocytes. Similar levels of covalent binding to proteins were observed in rat and human hepatocytes following incubation with [3H]­MK-8666. After protease digestion of hepatocyte pellets, amino acid adducts A1, A2, and A3 were identified as transacylated products with lysine, serine, and cysteine residues, respectively. Amino acid adducts A4a–c were identified as glycation adducts resulting from rearrangement of MK-8666–1-O-β-acyl glucuronide to ring-opened aldehydes which further condensed with lysine residues of proteins into imine adducts. Adducts A1–A3 and A4a–c were detected in rat and human liver microsomes fortified with UDPGA. Adducts A1–A3 were detected in rat and human liver microsomes fortified with CoA and ATP. Additionally, a trace amount of CoA thioester metabolite of MK-8666 and its transacylated GSH adduct were detected in human liver microsomes fortified with CoA, ATP, and GSH. Higher levels of covalent binding to protein were observed when [3H]­MK-8666 was incubated in liver microsomes supplemented with CoA and ATP compared to UDPGA. Addition of GSH attenuated levels of CoA thioester-mediated covalent binding by 41–45%. Collectively, these studies indicated that metabolism of the −COOH moiety of MK-8666 can form a reactive acyl glucuronide and an acyl CoA thioester, which covalently modifies proteins and may represent one causative mechanism of the observed DILI.

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