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Takeshi Mizutani, Nagaraj Kerur, Shengjian Li, Younghee Kim, Reo Yasuma, Tetsuhiro Yasuma, Ana Bastos-Carvalho, Benjamin Fowler, Bradley D Gelfand, Jayakrishna Ambati; Dual anti-angiogenic activity of Bevacizumab from its IgG1-Fc domain and VEGF neutralization. Invest. Ophthalmol. Vis. Sci. 2014;55(13):1936.
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© ARVO (1962-2015); The Authors (2016-present)
Bevacizumab has been reported to inhibit angiogenesis in various mouse models, even though it does not recognize mouse Vegfa. The purpose of this study is to evaluate the relative anti-angiogenic activity of Fc and Fab antibody fragments by determining the efficacy of multiple anti-VEGF drugs in mice in the presence of human VEGFA.
Serum starved Py4 mouse blood endothelial cells were stimulated with mouse Vegfa or human VEGFA to assess Vegfr2 phosphorylation. Bevacizumab or neutralizing anti-mouse Vegfa mAb was added to medium at the same time. We employed C57BL/6J, and hum-X VEGF mice which express a humanized form of VEGFA. Human VEGFA was injected intravitreously in C57BL/6J mice before laser injury. Choroidal neovascularization (CNV) was induced by laser injury in C57BL/6J mice, and volumes were measured 7 days later by confocal evaluation of Griffonia simplicifolia Isolectin B4 staining of RPE-choroid flatmounts. Bevacizumab, ranibizumab, bevacizumab-Fab, bevacizumab-Fc, or human IgG1 was injected into the vitreous humor of mice immediately after laser injury.
Bevacizumab and ranibizumab inhibited Vegfr2 phosphorylation induced by human VEGFA but not by mouse Vegfa. In C57BL/6J mice, CNV was suppressed by bevacizumab, not by ranibizumab (p<0.05 compared with PBS). CNV, augmented by administration of human VEGFA, was suppressed to similar extents by ranibizumab and bevacizumab-Fab and bevacizumab-Fc and human IgG1, and, to a greater extent, by bevacizumab (p<0.05 compared with PBS + human VEGFA). In humanized VEGF mice, CNV volume was suppressed by bevacizumab-Fc, and also by ranibizumab and bevacizumab-Fab, and, to a greater extent, by bevacizumab (p<0.05 compared with PBS).
Our study conclusively demonstrates an unexpected anti-angiogenic function of the Fc fragment of bevacizumab. The angioinhibitory effects by targeting human VEGFA and endogenous mouse Fc receptor pathways, are additive. These data form the basis for novel Fc-based, non-VEGF targeting, therapeutic strategies for treating neovascular AMD.
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