AAV-Mediated sFLT-1 Gene Therapy Ameliorates Retinal Lesions in Ccl2/cx3cr1 Deficient Mice

Abstract

Purpose: : Vascular endothelial growth factor (VEGF) induces angiogenesis. The binding of the VEGF receptor (VEGFR) to VEGF triggers cascades of signal transduction pathways. Studies have shown that VEGF antagonists, including adeno-associated virus (AAV)-sFLT-1 (soluble VEGFR-1), suppresses chorioretinal neovascularization (CNV). We tested the effects of subretinal delivery of AAV-sFLT-1 on retinal lesions in Ccl2^-/-/Cx3cr1^-/- deficient mice,a model that spontaneously develops age-related macular degeneration (AMD)-like features, including abnormal retinal pigment epithelium, photoreceptor degeneration, retinal A2E accumulation and over-expression of ocular VEGF-A.

Methods: : One microliter of AAV5-sFLT-1 (5.8X10¹² genome containing viral particles per ml) was injected into the subretinal space of the right eyes of 4-week-old Ccl2^-/-/Cx3cr1^-/- mice (n=50). The left eyes were untreated as controls. Funduscopic photographs were taken sequentially once a month. The mice were sacrificed 3 months after injection. Effective AAV transfection was ensured by PCR amplification of the transcript expressed from the virus package in ocular tissues. Ocular histopathology was performed. Retinal A2E was measured. Data were compared for the injected right eyes and the contralateral (control) eyes.

Results: : The treated eyes developed retinal detachment during subretinal injection, indicating a successful procedure. The right eyes that received AAV5-sFLT-1 showed significant improvements in the retinal lesions as compared to the left eyes 3 months after injection. The average lesion scores were 0.23±0.14 in the treated right eyes versus 0.85±0.19 in the left eyes (p=0.01). The histological scores between the right and left eyes from 10 pairs of eyes revealed a decrease in score in 6 pairs, no change in score in 2 pairs, and an increase in score in 2 pairs. Retinal ultrastructures also illustrated fewer lipofuscin granules and better preserved photoreceptors in the right eyes compared with the left eyes. A2E was lower in the right eyes than the left eyes.

Conclusions: : Subretinal injection of AAV5-sFLT-1 can either reverse or stabilize the progression of retinal lesions in Ccl2/Cx3cr1 deficient mice. Lowering retinal expression of VEGF-A by trapping excess VEGF-A using sFLT-1 may result in an amelioration of the damaged retina. The findings support the beneficial effects of sFLT-1 gene therapy for AMD.

Keywords: age-related macular degeneration • gene transfer/gene therapy • pathology: experimental