Metabolic profiling of TRPV1 antagonists of the benzothiazole amide series : implications for in vitro genotoxicity assessment

• 1. In vitro metabolic profiling and in vitro genotoxicity assessment are important aspects of the drug discovery program as they eliminate harmful compounds from further development. In standard in vitro genotoxicity testing, induced rat liver S9 is used as an exogenous bio-activation system for detecting promutagens. In this study we show that rat liver S9 is an insufficient system regarding the conversion of TRPV1 antagonists of the benzothiazole amide series into relevant in vivo metabolites. 2. Human and rat hepatocyte experiments demonstrated generation of an aryl amine metabolite that was subsequently N-acetylated. The hydrolyzed metabolites as well as the parent compound were also metabolized into glutathione (GSH) conjugates. Rat liver S9 exhibited a very low amide hydrolysis capacity and no formation of GSH conjugates when supplemented with NADPH and GSH. 3. The discrepancy in metabolic capability between hepatocytes and rat liver S9 led to confounding results in in vitro genotoxicity assessment for this chemical class as judged by the results of Ames test, mouse lymphoma assay, SOS/umu test and Comet assay in rat hepatocytes. 4. This study highlights the pivotal role that understanding the mechanism of metabolite formation has in interpreting as well as designing reliable and relevant in vitro genotoxicity experiments.

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