Oxidative Bioactivation and Toxicity of Leflunomide in Immortalized Human Hepatocytes and Kinetics of the Non-Enzymatic to Its Major etabolitie, A77 1726

We used immortalized human hepatocytes to study the bioactivation of leflunomide and the metabolic degradation to its major metabolic,A77 1726. Both leflunomide and A77 1726 caused a time-and concentration-dependent increase in LDH release. The cytotoxicity of leflunomide,but not that of A77 1726, was prevented by the pan -CYP inhibitor, 1-aminobenzotriazole,indicating that an oxidative metabolite(s) was responsible for the cell injury.LC/MS/MS analysis revealed that leflunomide was rapidly degraded in hepatocytes biphasically(t1/2a=1.5 h,t1/2b>24h),but much slower in cell-free medium(t1/2>24 h).In contrast,the generation of A77 1726 occured at a similar rate in cells and cell-free system s.In conclusion,leflunomide wsa rapidly metabolized in human hepatocytes to A77 1726, but its toxicity was dependent on other ,CYP-dependent intermedietly metabolized in human hepatocytes to A77 1726, but its toxicity was dependent on other ,CYP-dependent intermediidly metabolized in human hepatocytes to A77 1726, but its toxicity was dependent on other,CYP-dependent intermediidly metabolized in human hepatocytes to A77 1726,but its toxicity was dependent intermediates.