Ethyl (6R)-6-[N-(2-chloro-4-fluorophenyl)sulfamoyl]cyclohex-1-ene-1-carboxylate (TAK-242) was metabolized to cyclohexene and phenyl ring moieties in non-clinical pharmacokinetic studies and it was suggested that the cyclohexene ring moiety of TAK-242 is tightly bound to endogenous macromolecules. After incubation of TAK-242 and glutathione (GSH) in phosphate buffer (pH 7.4) at 37 °C, TAK-242 reacted with GSH to produce a glutathione conjugate of the cyclohexene ring moiety of TAK-242, which had been observed as a metabolite (M-SG) in non-clinical pharmacokinetic studies. Formation of M-SG was time dependent with a first order reaction and M-I, a metabolite from the phenyl ring moiety of TAK-242, was also produced in parallel. The formation of M-SG was accelerated with increasing pH, therefore it was indicated that TAK-242 reacted with GSH by a nucleophilic substitution reaction. Because glutathione transferase (GST) enhanced M-SG formation in vitro, it is expected that the conjugation of TAK-242 with GSH is also facilitated by GST in vivo in addition to a spontaneous chemical reaction. When radio-labeled TAK-242 ([cyclohexene ring-U-¹⁴C]TAK-242) was incubated with rat serum albumin (RSA) or human serum albumin (HSA) in vitro, the radioactive material was covalently bound to RSA and HSA, and M-I was generated simultaneously in the reaction mixture. The chemical structure of the TAK-242 adduct covalently bound to HSA was characterized by the accurate mass spectra that cyclohexene ring moiety of TAK-242 was covalently bound to the lysine residue in HSA. The adduct was also detected in the plasma of rats and humans after single i.v. dosing of TAK-242 (in vivo).
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