Hepatic Metabolism of MK-0457, a Potent Aurora Kinase Inhibitor: Interspecies Comparison and Role of Human Cytochrome P450 and Flavin-Containing Monooxygenase

Abstract

ABSTRACT:MK-0457 (N-[4({4-(4-methylpiperazin-1-yl)-6-[(3-methyl-1H-pyrazol-5 -yl)amino]pyrimidin-2-yl}thio)phenyl]cyclopropanecarboxamide), an Aurora kinase inhibitor in development for the treatment of cancer, was evaluated for its in vitro metabolism in different species. This compound primarily underwent N-oxidation and Ndemethylation in human, monkey, dog, and rat liver preparations. However, N-demethylation was less significant in dogs. The formation of minor metabolites varied with species, but all metabolites generated in human hepatocytes were observed in animals. Results of immunoinhibition, selective chemical inhibition, thermal inactivation, and metabolism by recombinant cytochromes P450 and flavin-containing monoogygenases (FMOs) strongly suggest that CYP3A4 and FMO3 comparably contributed to MK-0457 N-oxidation in human liver microsomes, where the reaction conformed to Michaelis-Menten kinetics. These studies indicate a major role of CYP2C8 in the N-demethylation reaction, whereas CYP3A4 only made a minor contribution. However, significant substrate inhibition was observed with MK-0457 N-demethylation at high substrate concentrations (>10 M) in human liver microsomes relative to the anticipated therapeutic exposure. A multienzyme metabolic pathway such as this may mitigate the potential of drug interactions in clinical treatment with MK-0457.MK-0457 is a potent and selective Aurora kinase inhibitor in clinical development for the treatment of solid tumors and hematopoietic cancers. Aurora kinases are involved in the regulation of chromosome segregation and cytokinesis during mitosis, and overexpression of these proteins is likely to contribute to tumorigenesis (Katayama et al., 2003;Jiang et al., 2006). Inhibition of Aurora kinases disrupts progression of the cancer cell cycle and blocks proliferation in vitro, whereas noncycling cells remain unaffected. Rodent in vivo studies have shown that MK-0457 inhibits tumor growth, resulting in regression (Harrington et al., 2004). MK-0457 is the first kinase inhibitor to exhibit clinical activity in patients with T315I BCR-ABL mutated chronic myeloid leukemia or acute lymphocytic leukemia (Giles et al., 2007).To gain an understanding of the species differences and similarities in the disposition of MK-0457 for the purpose of predicting human disposition and determining appropriate species for toxicology studies, metabolism studies were conducted in rat, dog, monkey, and human liver preparations. For the purpose of predicting potential drug-drug interactions, the drug-metabolizing enzymes involved in the biotransformation of MK-0457 in human liver microsomes were identified. In the present studies, the role of cytochrome P450 (P450) and flavin-containing monooxygenase (FMO), two major enzyme systems that catalyze oxidative metabolic reactions, have been investigated using a combination of in vitro reaction phenotyping strategies. These methods include the use of 1) monoclonal antibodies for immunoinhibition of specific P450 families, 2) selective chemical in...

Effect of Human Flavin-Containing Monooxygenase 3 Polymorphism on the Metabolism of Aurora Kinase Inhibitors

“…When the ratios of k cat / K m are compared, both enzymes seem to be equally efficient in transforming Tozasertib into the N -oxide product. The K m values measured are slightly higher when compared to the data published by Ballard and co-workers [16], probably because they specifically address the substrate binding to hFMO3 and do not consider the role of cytochrome P450 3A4 that also contributes to the overall N -oxide metabolite of Tozasertib. Moreover, for Tozasertib the V max of wild type hFMO3 was 536 pmol/min/mg FMO3 while V257M hFMO3 V max resulted in 248 pmol/min/mg FMO3 (Table 1) in good agreement with the data published by Ballard and co-workers [16].…”

“…Tozasertib is metabolised into the N -desmethyl product by CYP3A4 and CYP2C8, and into the N -oxide product by CYP3A4 and hFMO3, as previously demonstrated by Ballard and co-workers [16]. Their study highlighted the combined contribution of hFMO3 and P450 3A4 to the formation of the N -oxide metabolite in human liver microsomes.…”

“…In 2007, Ballard and co-workers [16] identified the metabolites of Tozasertib generated in human hepatocytes that were mainly the N -oxide and the desmethyl products. They went on to characterize the hepatic enzymes responsible and found that hFMO3 appreciably (50%–60%) contributed to the N -oxidation of Tozasertib in human liver microsomes together with cytochrome P450 3A4.…”

Role of Cytochrome P450 2C8 in Drug Metabolism and Interactions

“…The contributions of CYP3A4 and CYP2C8 to the in vivo elimination of ponatinib are estimated to 34% and 19%, respectively (FDA, 2012e). Furthermore, CYP2C8 plays a major role in the N-demethylation of the aurora kinase inhibitor tozasertib (Table 4; Ballard et al, 2007). With the exception of dabrafenib and imatinib, the in vivo importance of CYP2C8 in the metabolism of protein kinase inhibitors seems to have been poorly studied.…”

Escherichia coli Overexpressing a Baeyer-Villiger Monooxygenase from Acinetobacter radioresistens Becomes Resistant to Imipenem

“…As mentioned earlier, human FMO3, Ar-BVMO, and M. tuberculosis EtaA can metabolize a common substrate, ethionamide. Based on this, we investigated the ability of Ar-BVMO to use as a substrate two potent and selective aurora kinase inhibitors, danusertib and tozasertib, which are used for the treatment of solid tumors and hematopoietic cancers, respectively, and metabolized by human FMO3 in liver microsomes (52,53).…”