Takahashi Hidenori, Tamaki Yasuhiro, Ishii Nobuya, Oikawa Nobuhiro, Mizuguchi Eisaku, Francis Jasmine H, Inoue Yuji, Iriyama Aya, Obata Ryo, Yanagi Yasuo
Department of Ophthalmology, University of Tokyo School of Medicine, Bunkyo-ku, Tokyo, Japan.
Curr Eye Res. 2008 Nov;33(11):1002-10. doi: 10.1080/02713680802492440.
To select a novel orally administered VEGFR-2 (KDR/flk-1) specific tyrosine kinase inhibitor in a murine model of choroidal neovascularization (CNV).
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.
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.
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.
在脉络膜新生血管（CNV）小鼠模型中筛选一种新型口服VEGFR-2（KDR/flk-1）特异性酪氨酸激酶抑制剂。
从化合物库中，通过VEGF诱导的VEGFR-2和RAF激酶磷酸化以及人脐静脉内皮细胞（HUVEC）培养的增殖分析和体外管形成试验，筛选出有效的VEGFR2抑制剂。使用二极管激光光凝在C57/BL6小鼠中诱导CNV。通过血管造影检查评估所选化合物的抗血管生成作用，其中对荧光素渗漏程度进行评分，并进行组织学分析，以便在光学显微镜下测量CNV膜。此外，对C57/BL6小鼠每日口服所选化合物，持续14天，并测量体重。
筛选出6种有效抑制VEGFR-2的化合物进行进一步研究。所选化合物处理条件显示对VEGFR-2酪氨酸激酶磷酸化具有剂量依赖性抑制，IC50为0.0022至0.098微摩尔。所选化合物不抑制HCT116增殖，但对VEGFR-依赖的HUVEC表现出强烈抑制作用（IC50 = 0.0018至0.058微摩尔）。所选化合物处理还导致体外管形成的剂量依赖性减弱。在小鼠CNV模型中，#0451是最有效的化合物。与对照组相比，12.5、25、50和100毫克/千克#0451处理的眼睛中荧光素渗漏强度明显更低。组织学上，#0451处理的眼睛中CNV膜体积以剂量依赖性方式显著减少。在100毫克/千克或更低的治疗剂量下，治疗组和未治疗组之间没有明显的体重减轻。
口服新型VEGFR-2（KDR/flk-）特异性酪氨酸激酶抑制剂#0451在我们的小鼠CNV模型中显示出抗血管生成作用#0451可能对治疗与年龄相关性黄斑变性（AMD）相关的脉络膜新生血管有用。
