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Reduction of choroidal neovascularization in mice by adeno‐associated virus‐delivered anti‐vascular endothelial growth factor short hairpin RNA
Author(s) -
Askou Anne Louise,
Pournaras JeanAntoine C.,
Pihlmann Maria,
Svalgaard Jesper D.,
Arsenijevic Yvan,
Kostic Corinne,
Bek Toke,
DagnæsHansen Frederik,
Mikkelsen Jacob Giehm,
Jensen Thomas Gryesten,
Corydon Thomas J.
Publication year - 2012
Publication title -
the journal of gene medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.689
H-Index - 91
eISSN - 1521-2254
pISSN - 1099-498X
DOI - 10.1002/jgm.2678
Subject(s) - gene knockdown , choroidal neovascularization , small hairpin rna , vascular endothelial growth factor , gene silencing , intravitreal administration , macular degeneration , in vivo , adeno associated virus , angiogenesis , neovascularization , endogeny , rna interference , genetic enhancement , vascular endothelial growth factor a , biology , medicine , retinal , cancer research , ophthalmology , rna , vector (molecular biology) , cell culture , gene , endocrinology , vegf receptors , genetics , recombinant dna
Abstract Background Strategies leading to the long‐term suppression of inappropriate ocular angiogenesis are required to avoid the need for repetitive monthly injections for treatment of diseases of the eye, such as age‐related macular degeneration (AMD). The present study aimed to develop a strategy for the sustained repression of vascular endothelial growth factor (VEGF), which is identified as the key player in exudative AMD. Methods We have employed short hairpin (sh)RNAs combined with adeno‐associated virus (AAV) delivery to obtain the targeted expression of potent gene‐regulatory molecules. Anti‐VEGF shRNAs were analyzed in human retinal pigment epithelial (RPE) cells using Renilla luciferase screening. For in vivo delivery of the most potent shRNA, self‐complementary AAV vectors were packaged in serotype 8 capsids (scAAV2/8‐hU6‐sh9). In vivo efficacy was evaluated either by injection of scAAV2/8‐hU6‐sh9 into murine hind limb muscles or in a laser‐induced murine model of choroidal neovascularization (CNV) following scAAV2/8‐hU6‐sh9 subretinal delivery. Results Plasmids encoding anti‐VEGF shRNAs showed efficient knockdown of human VEGF in RPEs. Intramuscular administration led to localized expression and 91% knockdown of endogenous murine (m)VEGF. Subsequently, the ability of AAV2/8‐encoded shRNAs to impair vessel formation was evaluated in the murine model of CNV. In this model, the sizes of the CNV were significantly reduced (up to 48%) following scAAV2/8‐hU6‐sh9 subretinal delivery. Conclusions Using anti‐VEGF vectors, we have demonstrated efficient silencing of endogenous mVEGF and showed that subretinal administration of scAAV2/8‐hU6‐sh9 has the ability to impair vessel formation in an AMD animal model. Thus, AAV‐encoded shRNA can be used for the inhibition of neovascularization, leading to the development of sustained anti‐VEGF therapy. Copyright © 2012 John Wiley & Sons, Ltd.