ENZYMATIC OXIDATION OF TYROSINE KINASE INHIBITOR VANDETANIB: A DETAILED STUDY
Stiborová M.1, Martínek V.1, Pompach P.1, Takácsová P.1, Vavrová K.1 , Heger Z.2, Adam V.2, Arlt V.M.3
1 Department of biochemistry, Charles University, Albertov 6, 128 43 Prague 2; 2Department of Chemistry and Biochemistry, Mendel University in Brno, Zemedelska 1, 61300 Brno; 3Analytical and Environmental Sciences Division, MRC-PHE Centre for Environment and Health, King’s College London, Franklin-Wilkins Building, 150 Stamford Street, London, SE1 9NH, UK.
Metabolism affects the pharmacological efficiency of the tyrosine kinase inhibitor vandetanib. Here, we investigated the in vitro metabolism of vandetanib using (i) hepatic subcellular systems rich in drug-metabolizing enzymes (microsomes) isolated from livers of humans and several animal models and (ii) human and/or rat recombinant biotransformation enzymes such as cytochromes P450 (CYPs) and flavin-containing monooxygenases (FMOs). In addition to the structural characterization of vandetanib metabolites, individual human and rat enzymes capable of oxidizing this drug were identified. Two vandetanib metabolites, N-desmethylvandetanib and vandetanib N-oxide, were formed in incubations with hepatic microsomes. The generation of N-desmethylvandetanib was attenuated by inhibitors of CYP3A and 2C subfamilies in both human and rat microsomes, while an inhibitor of CYP2D6 only decreased formation of this metabolite in human microsomes. The FMO inhibitor methimazol decreased the formation of vandetanib N-oxide in both rat and human microsomes. These results indicated that in the microsomal systems studied, CYP3A, 2C and/or 2D are mainly responsible for the formation of N-desmethylvandetanib and FMO1 and 3 mainly for the generation of vandetanib N-oxide. Human recombinant CYP3A4>>>2D6>1A1>2C8>3A5 oxidized vandetanib to N-desmethylvandetanib, while rat recombinant CYP2C11>>3A1>3A2>1A1>1A2>2D1>2D2 were effective in catalyzing this reaction. Cytochrome b5 which serves as electron donor to CYP enzymes influenced the CYP-catalyzed formation of N-desmethylvandetanib; CYP3A4 was most affected. Human CYP3A4 is not only the most efficient enzyme oxidizing vandetanib to N-desmethylvandetanib, but also most important due to its high expression in human liver. Molecular modeling indicated that binding of more than one molecule of vandetanib into the CYP3A4-active center can be responsible for the high efficiency of CYP3A4 N-demethylating vandetanib to N-desmethylvandetanib. Indeed, the CYP3A4-mediated reaction was allosterically modulated exhibiting kinetics of positive cooperativity, which corresponds well to the in-silico docking model, where two bound vandetanib molecules were found in the active center of CYP3A4.
Supported by the Czech Science Foundation (grant 18-10251S).