Reduction of choroidal neovascularization in mice by adeno-associated virus-delivered anti-vascular endothelial growth factor short hairpin RNA
Article first published online: 22 NOV 2012
Copyright © 2012 John Wiley & Sons, Ltd.
The Journal of Gene Medicine
Volume 14, Issue 11, pages 632–641, November 2012
How to Cite
Askou, A. L., Pournaras, J.-A. C., Pihlmann, M., Svalgaard, J. D., Arsenijevic, Y., Kostic, C., Bek, T., Dagnæs-Hansen, F., Mikkelsen, J. G., Jensen, T. G. and Corydon, T. J. (2012), Reduction of choroidal neovascularization in mice by adeno-associated virus-delivered anti-vascular endothelial growth factor short hairpin RNA. J. Gene Med., 14: 632–641. doi: 10.1002/jgm.2678
- Issue published online: 22 NOV 2012
- Article first published online: 22 NOV 2012
- Accepted manuscript online: 18 OCT 2012 03:24PM EST
- Manuscript Accepted: 15 OCT 2012
- Manuscript Revised: 4 OCT 2012
- Manuscript Received: 29 JUN 2012
- adeno-associated virus;
- anti-VEGF gene therapy;
- choroidal neovascularization;
- short hairpin RNA
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.
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.
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.
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.