A trapping approach for semi-quantitative assessment of bioactivation potential has been established for new chemical entities by using [³⁵S]cysteine and [¹⁴C]sodium cyanide as trapping reagents. Reactive metabolites were trapped as radioactive adducts with the trapping reagents to be analyzed by radio-LC(/MS). As a reference, hepatotoxic drugs (clozapine, diclofenac, R-(+)-pulegone and troglitazone) were tested in the [³⁵S]cysteine trapping assay and the proposed structures of the cysteine adducts were consistent with glutathione adducts previously reported. The accuracy of this methodology to predict bioactivation potential of structurally diverse non-radiolabeled test compounds was evaluated by comparing the radiochromatographic peak area obtained in this assays with the extent of covalent binding to protein assessed by the conventional method using radiolabeled test compounds. The value obtained from the [³⁵S]cysteine trapping assay in human liver microsomes predicted potential for covalent binding of the test compounds to proteins with reasonable accuracy. A combination of trapping reagents ([³⁵S]cysteine and [¹⁴C]cyanide) improved the accuracy for prediction of bioactivation potential by simultaneously trapping both types of electrophilic reactive metabolites. This method is expected to be a useful to prioritize compounds for further development based on the bioactivation liability, especially at the lead optimization stage.

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