Identification of a novel vascular endothelial growth factor receptor 2 inhibitor and its effect for choroidal neovascularization in vivo.
OBJECTIVE
To select a novel orally administered VEGFR-2 (KDR/flk-1) specific tyrosine kinase inhibitor in a murine model of choroidal neovascularization (CNV).


METHODS
From a compound library, potent VEGFR2 inhibitors were selected by VEGF-induced phosphorylation of VEGFR-2 and RAF kinases and the proliferation analysis by HUVEC cultures and in vitro tube formation assay. CNV was induced in C57/BL6 mice using diode laser photocoagulation. The antiangiogenic effect of selected compounds was assessed by angiographic examination, in which extent of fluorescein leakage was scored and histological analysis, allowing for measurement of CNV membrane under light microscope. In addition, C57/BL6 mice were treated with daily oral administration of selected compounds for 14 days and body weights were measured.


RESULTS
Six compounds that potently inhibited VEGFR-2 were selected for further investigation. Selected compounds-treated conditions showed a dose-dependent inhibition of phosphorylation of VEGFR-2 tyrosine kinase with an IC50 of 0.0022 to 0.098 microm. Selected compounds did not inhibit the HCT116 proliferation but did demonstrate a strong inhibition effect for VEGFR-2 dependant HUVEC (IC50=0.0018 to 0.058 microm). Selected compounds treatment also resulted in a dose-dependent attenuation of in vitro tube formation. In the murine CNV model, #0451 is the most effective compound. The intensity of fluorescein leakage was significantly lower in doses of 12.5, 25, 50, and 100 mg/kg #0451-treated eyes compared to controls. Histologically, CNV membrane volumes were significantly reduced in #0451-treated eyes in a dose-dependent manner. At therapeutic doses of 100 mg/kg or less, there was no significant weight loss between the treated and untreated groups.


CONCLUSIONS
Oral administration of #0451, a novel VEGFR-2 (KDR/flk-1)-specific tyrosine kinase inhibitor, demonstrates anti-angiogenic effects in our murine model of CNV. #0451 may be useful to treat the choroidal neovascularization associated with AMD.
Takahashi
Hidenori
H
Department of Ophthalmology, University of Tokyo School of Medicine, Bunkyo-ku, Tokyo, Japan.
Tamaki
Yasuhiro
Y
Ishii
Nobuya
N
Oikawa
Nobuhiro
N
Mizuguchi
Eisaku
E
Francis
Jasmine H
JH
Inoue
Yuji
Y
Iriyama
Aya
A
Obata
Ryo
R
Yanagi
Yasuo
Y
D000284:Administration, Oral; D020533:Angiogenesis Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D000792:Angiography; D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D020256:Choroidal Neovascularization / Q000000981:diagnostic imaging / Q000378:metabolism / Q000503:physiopathology*; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000166:cytology; D019793:Fluorescein; D005456:Fluorescent Dyes; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D010766:Phosphorylation / Q000187:drug effects; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000037:antagonists & inhibitors* / Q000378:metabolism
D020533:Angiogenesis Inhibitors; D005456:Fluorescent Dyes; D040301:Vascular Endothelial Growth Factor Receptor-2; D019793:Fluorescein