In Vitro Metabolism of the Novel, Highly Selective Oral Angiogenesis Inhibitor Motesanib Diphosphate in Preclinical Species and in Humans

Abstract

ABSTRACT:Motesanib diphosphate is a novel, investigational, highly selective oral inhibitor of the receptor tyrosine kinases vascular endothelial growth factor receptors 1, 2, and 3, the platelet-derived growth factor receptor, and the stem cell factor receptor (Kit). The in vitro metabolic profiles of [ 14 C]motesanib were examined by using microsomes and hepatocytes from preclinical species and humans. Several oxidative metabolites were observed and characterized by tandem mass spectrometry, nuclear magnetic resonance spectroscopy, and coinjection with authentic standards. Cytochrome P450 (P450) 3A4 is the major isozyme involved in the oxidative biotransformation of motesanib, but the CYP2D6 and CYP1A isozymes also make minor contributions. In hepatocyte incubations, oxidative and conjugative pathways were observed for all species examined, and indoline N-glucuronidation was the dominant pathway. Three less common and novel phase II conjugates of the indoline nitrogen were detected in hepatocytes and in microsomes supplemented with specific cofactors, including N-carbamoyl glucuronide, N-glucose, and N-linked ␤-N-acetylglucosamine. An N-glucuronide metabolite was the most frequently observed phase II conjugate in liver microsomes of all species, whereas the N-acetylglucosamine conjugate was observed only in monkey liver microsomes. Incubations with recombinant human UDP-glucuronosyltransferases (UGTs) and inhibition by the UGT1A4 and UGT1A1 substrates/inhibitors imipramine and bilirubin suggested that UGT1A4 is the major UGT isozyme catalyzing the N-glucuronidation of motesanib, with a minor contribution from UGT1A1. The in vitro metabolic profiles were similar between the human and preclinical species examined. All metabolites found in humans were also detected in other species.Motesanib diphosphate (formerly known as AMG 706) (Fig. 1) is a highly selective kinase inhibitor with both antiangiogenic and direct antitumor activity (Polverino et al., 2006;Rosen et al., 2007). Motesanib is an ATP-competitive inhibitor of vascular endothelial growth factor (VEGF) receptors 1, 2, and 3, platelet-derived growth factor (PDGF) receptor, and stem cell factor receptor (Kit), which have all been implicated in the pathogenesis of human tumors (Dvorak, 2002;Heinrich et al., 2002;Song et al., 2005). In preclinical models of human cancer, oral administration of motesanib potently inhibited VEGF-induced angiogenesis in the rat corneal model and induced regression of established A431 xenografts (Polverino et al., 2006). Furthermore, motesanib has shown antiangiogenic and antitumor activity in patients with advanced solid malignancies (Rosen et al., 2007). Additional studies of motesanib as monotherapy and in combination with various agents are currently ongoing.Definitive characterization of metabolites and examination of species differences in metabolism are becoming increasingly important.

Metabolism and Pharmacokinetics of Novel Selective Vascular Endothelial Growth Factor Receptor-2 Inhibitor Apatinib in Humans

“…The primary routes of apatinib biotransformation involved E-and Z-cyclopentyl-3-hydroxylation, N-dealkylation, pyridyl-25-N-oxidation, 16-hydroxylation, dioxygenation, and Oglucuronidation after cyclopentyl-3-hydroxylation. Apatinib shared some common metabolic pathways with its analog motesanib (Li et al, 2009), such as pyridyl-N-oxidation and N-dealkylation. However, the major metabolic reactions of apatinib occurred on the cyclopentyl group, including hydroxylation and further Oglucuronidation.…”

Reaction Phenotyping

“…In the absence of synthetic metabolite standard, however, the use of LC/MS/MS for quantitation of metabolites needs to be cautioned, because the relative rates of metabolite formation may not be estimated based on its peak response due to different ionization effi ciencies from the drug candidate. Sometimes, combination of detection methods from UV, radiometric, and LC -MS/MS can be very useful in metabolite quantitation and reaction phenotyping (Li et al, 2009 ).…”

“…As shown in Figure 13.2 , the effect of several CYP isoform selective chemical inhibitors on metabolism of a highly selective angiogenesis inhibitor motesanib was evaluated (Li et al, 2009 ). The involvement of several CYP isozymes results in inhibition of up to 80%.…”

“…It should be noted that proper control incubations be also conducted to measure the ability of the antibody to inhibit an established marker P450 activity and the effect of control antibody on the metabolism of drug candidate of interest. An example of the effect of CYP antibodies on the oxidative metabolism of highly selective angiogenesis inhibitor motesanib (Li et al, 2009 ) is shown in Figure 13.3 . A primary limitation of anti -P450 antibodies for reaction phenotyping has been the cross -reactivity with related P450s, especially in the case of polyclonal or antipeptide antibodies.…”

“…Valuable information on which CYP enzymes can and which ones cannot metabolize the drug candidate can be readily obtained (Mankowski, 1999 ). An example is shown in Figure 13.4 , in which the metabolism of motesanib, a highly selective angiogenesis inhibitor, was evaluated in a panel of recombinant CYP enzymes (Li et al, 2009 ), and several recombinant CYP enzymes (CYP3A, CYP2D6, CYP1A1, CYP2B1, and CYP2B6) exhibited activity in catalyzing the formation of three oxidative metabolites of motesanib.…”