Identification of Human Liver Cytochrome P450 Isoforms Involved in Autoinduced Metabolism of the Antiangiogenic Agent (Z)-5-((1,2-Dihydro-2-oxo-3H-indol-3-ylidene)methyl)-2,4-dimethyl- 1H-pyrrole-3-propanoic Acid (TSU-68)

( Z )-5-[(1,2-Dihydro-2-oxo-3 H -indol-3-ylidene)methyl]-2,4-dimethyl-1 H -pyrrole-3-propanoic acid (TSU-68) is a new anticancer drug that inhibits angiogenic receptor tyrosine kinases, which play a crucial role in tumor-induced vascularization. TSU-68 undergoes hepatic oxidation and glucuronidation. Incubation of TSU-68 with human liver microsomes in the presence of NADPH resulted in the formation of three major metabolites: 5-, 6-, and 7-hydroxyindolinone derivatives. The 5-, 6-, and 7-hydroxylation followed simple Michaelis-Menten kinetics with V max/ K m values (an indicator of intrinsic clearance) of 13, 25, and 6 μl/min/mg, respectively. Of the 10 cDNA-expressed human cytochrome P450 isoforms examined, only CYP1A1 and CYP1A2 exhibited appreciable TSU-68 hydroxylation activity. Inhibition studies with α-naphthoflavone (a selective CYP1A2 inhibitor) and anti-CYP1A2 antibody also indicated the almost exclusive role of CYP1A2 in microsomal TSU-68 hydroxylation. Treatment of human hepatocytes with 10 μM TSU-68 resulted in a 28- to 140-fold increase in CYP1A1/2-mediated ethoxyresorufin O -deethylase activity. The protein levels of CYP1A2 were increased in TSU-68-treated hepatocytes, and those of CYP1A1, which were undetectable in control hepatocytes, were also increased to detectable levels in the TSU-68-treated hepatocytes. Thus, TSU-68 was shown to induce CYP1A1/2 expression, which was responsible for its hydroxylation. The observation that TSU-68 treatment resulted in a 10- to 45-fold increase in 5-, 6-, and 7-hydroxylation directly demonstrated the autoinduced hydroxylation of TSU-68. In conclusion, TSU-68 has the potential to cause induction of its own CYP1A1/2-mediated oxidative metabolism in humans. This autoinductive effect provides a clear explanation for the clinically observed decrease in TSU-68 plasma concentrations during repeated administration of the drug.