Gene Therapy of a Mouse Model of Stargardt's Disease Using Adeno-Associated Viral Vectors Packaging Large Genes

Abstract

Purpose: : Stargardt's disease (STGD) is the most common macular degeneration in humans and is caused by mutations in the ABCA4 gene. No therapies for STGD are available to date. Vectors derived from the small adeno-associated virus (AAV) are particularly amenable for ocular gene transfer and the identification of dozens of AAV serotypes allows efficient targeting of retinal cells following in vivo vector delivery. However, AAV2 cargo capacity is restricted to 4.7 kb which represents its major limitation when treating diseases due to mutations in large genes as ABCA4. The goal of our study was to generate an AAV vector for gene therapy of STGD.

Methods: : We tested the ability of various AAV serotypes (AAV2/1 to 9) to package an expression cassette containing Abca4 and whether AAV-mediated Abca4 retinal gene transfer can correct the phenotype of Abca4-/- mice.

Results: : We found that vectors with AAV5 capsids (AAV2/5) incorporate a genome of 8.9 kb in size including Abca4 more efficiently than the other AAV serotypes tested. Similar results were obtained with other large genes showing that the AAV2/5 packaging capacity is independent of the genome-packaged sequence. AAV2/5 encoding ABCA4 efficiently transduce cells in vitro and the retina in vivo resulting in translation of proteins of the appropriate size and function. Intraocular administration of AAV2/5 encoding ABCA4 in the Abca4-/- mouse model results in proper protein localization to rod outer segments and in stable retinal morphological and functional improvement.

Conclusions: : Our data support AAV-mediated Abca4 gene transferas a therapeutic strategy for recessive Stargardt’s disease. In addition, the possibility of packaging large genes and/or regulatory sequences in AAV enormously expands the therapeutic potential of this vector system.

Keywords: gene transfer/gene therapy • retinal degenerations: hereditary • macula/fovea