ZM323881, a novel inhibitor of vascular endothelial growth factor-receptor-2 tyrosine kinase activity.

Objective: Vascular endothelial growth factor (VEGF) increases vascular permeability and angiogenesis in many pathological conditions including cancer, arthritis, and diabetes. VEGF activates VEGF-Receptor 1(VEGF-R1) and VEGF-Receptor 2 (VEGF-R2), which autophosphorylate to initiate a signaling cascade resulting in angiogenesis and increased microvascular permeability. Here we describe a novel VEGF-R2 selective inhibitor, ZM323881 (5-[[7-(benzyloxy) quinazolin-4-yl]amino]-4-fluoro-2-methylphenol), that is a potent and selective inhibitor of VEGF-R2 tyrosine kinase in vitro (IC(50) < 2 nM), compared with other receptor tyrosine kinases, including VEGF-R1 (IC(50) > 50 microM).

Methods: Endothelial cell proliferation was assayed by (3)H-thymidine incorporation in response to VEGF-A +/- ZM323881. The effect of ZM323881 on VEGF-mediated permeability was measured in frog microvessels using the Landis Michel technique. To ensure that ZM323881 was effective in frogs, western analysis was performed on protein extracted from frog lungs incubated in the presence or absence of VEGF-A or VEGF-A with ZM323881.

Results: ZM323881 inhibits VEGF-A-induced endothelial cell proliferation (IC(50) = 8 nM) and VEGF-R2 tyrosine phosphorylation in vitro. VEGF-A-mediated increases in vascular permeability in perfused mesenteric microvessels in vivo were reversibly abolished by both ZM323881 and the class III receptor tyrosine kinase inhibitor PTK787/ZK222584.

Conclusions: These data suggest that VEGF-R2 phosphorylation is necessary for VEGF-A-mediated increases in microvascular permeability in vivo.