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

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.