Motesanib Diphosphate (AMG 706) is the orally bioavailable diphosphate salt of a multiple-receptor tyrosine kinase inhibitor with potential antineoplastic activity.

VEGFR2/Flk1:6 nM, c-Kit:8 nM, VEGFR3:6 nM, VEGFR1:2 nM, VEGFR2:3 nM

In a rat corneal model, administering Motesanib Diphosphate orally twice daily (ED50=2.1 mg/kg) or once daily (ED50=4.9 mg/kg) effectively inhibited vascular endothelial growth factor-induced angiogenesis. Additionally, in a model of transplanted squamous cell carcinoma of the head and neck, the combined use of Motesanib Diphosphate and radiation therapy demonstrated significant anticancer activity.

In human umbilical vein endothelial cells (HUVECs) induced by VEGF (IC50=10 nM), Motesanib Diphosphate significantly inhibits cell proliferation. It also markedly suppresses proliferation prompted by platelet-derived growth factor (IC50=207 nM) and c-kit phosphorylation induced by SCF (IC50=37 nM). Additionally, Motesanib Diphosphate enhances the sensitivity of these cells to radiation and exhibits broad-spectrum activity against the human VEGFR family.

In vitro kinase assays: Optimal enzyme, ATP, and substrate (gastrin peptide) concentrations are established for each enzyme using homogeneous time-resolved fluorescence (HTRF) assays. Motesanib Diphosphate is tested in a 10-point dose-response curve for each enzyme using an ATP concentration of two-thirds Km for each. Most assays consist of enzyme mixed with kinase reaction buffer [20 mM Tris-HCl (pH 7.5), 10 mM MgCl2, 5 mM MnCl2, 100 mM NaCl, 1.5 mM EGTA]. A final concentration of 1 mM DTT, 0.2 mM NaVO4, and 20 μg/mL BSA is added before each assay. For all assays, 5.75 mg/mL streptavidin-allophycocyanin and 0.1125 nM Eu-PT66 are added immediately before the HTRF reaction. Plates are incubated for 30 minutes at room temperature and read on a Discovery instrument. IC50 values are calculated using the Levenberg-Marquardt algorithm into a four-parameter logistic equation.

Cells are preincubated for 2 hours with different concentrations of Motesanib Diphosphate, and exposed with 50 ng/mL VEGF or 20 ng/mL bFGF for an additional 72 hours. Cells are washed twice with DPBS, and plates are frozen at -70 °C for 24 hours. Proliferation is assessed by the addition of CyQuant dye, and plates are read on a Victor 1420 workstation. IC50 data are calculated using the Levenberg-Marquardt algorithm into a four-parameter logistic equatio(Only for Reference)