Laser-Induced Choroidal Neovascularization Model to Evaluate Anti-Vegf Gene Transfer Therapy

Abstract

Purpose: : Pathologic choroidal neovascularizations (CNV) are implicated in the wet form of age-related macular degeneration (ARMD). In CNV the pathological growth of vessels is mediated of CNV in most ocular conditions. The cause may be hypoxia and/or inflammation. Our goal is to establish a standard protocol to induce CNV, and to evaluate the therapeutic value of an anti-VEGF factor.

Methods: : CNV was induced in C57/Bl6 by 4 burns (at the 3; 6; 9 and 12 o’clock positions around the optic disk, using a green argon laser (100 µm diameter spot size; 0,05 sec. duration). We have tested different intensities and the disruption of the Bruch’s membrane was identified by the appearance of a bubble at the site of photocoagulation. Fluorescein angiographies (FA) were undertaken 14 days after laser photocoagulation, and CNV area was determined by quantitaive immunohistochemistry in cryostat sections. For the anti-VEGF gene transfer, mice received subretinal injections of vectors encoding an anti-VEGF (LV-V65) single chain antibody (scFv) (Wenzel A. and al., ARVO 2004, abstract #779) or control vectors (LV-NL and LV-GFP), and were subjected to the laser injury 1 week later. Animals were sacrified 14 days after the laser lesion, and CNV was analysed in each group.

Results: : We first evaluated the proportion of CNV induced by laser and assessed the correlation between the intensity of the laser and the area of CNV in order to ascertain the efficacy of the method. While 180mW and 200mW laser intensity do not induce reliable CNV, 260mW is required to induce CNV and the following retinal degeneration. Higher doses lead to major injury of the retina. Preliminary results after V65 gene transfer suggest an inhibitory effect on CNV that is under quantitative evaluation.

Conclusions: : The identification of the laser intensity inducing both CNV and subsequent photoreceptor degeneration is of prime importance for the use of the laser-induced CNV model. These conditions will allow to evaluate the therapeutic potency of agents to prevent rapidly CNV and inhibit retinal degeneration. Moreover, local gene transfer to reduce angiogenic factor levels will represent an alternative to repeated intra-ocular injections of anti-VEGF therapeutic agents.

Keywords: neovascularization • gene transfer/gene therapy • laser