Stargardt disease (STGD1) is the most common form of inherited juvenile macular degeneration, leading to progressive loss of central vision. STGD1 is caused by autosomal recessive mutations in the ATP Binding Cassette Subfamily A Member 4 (ABCA4) gene, which encodes a membrane transporter, ABCA4, that facilitates the removal of all-trans-retinals (atRALs) from photoreceptor outer segment disc membranes as part of the visual cycle. Mutations in ABCA4 result in the accumulation of toxic atRALs and atRAL condensation products (A2E) and the subsequent degeneration of retinal pigment epithelial cells and photoreceptors. Despite its high prevalence, STGD1 has no treatment at present. While gene replacement therapy using Adeno-Associated Virus (AAV) has emerged as a promising therapy for inherited retinal diseases, the 4.7 kb gene capacity of the AAV vector has hindered treatment of mutations in larger genes such as ABCA4. However, the recent advent of CRISPR/Cas9 technology has expanded the application potential of AAV for treating genetic diseases. The CRISPR/Cas9 technology is an efficient genome editing technology, which can promote genomic modification at specific sites. Several studies have successfully demonstrated the modification of disease-causing mutations and rescue of the pathological phenotype in animal models by delivering CRISPR/Cas9 via AAV. These promising results raise the prospect of using AAV and CRISPR/Cas9 to target ABCA4 mutations in the photoreceptors of STGD1 patients. As proof-of-principle, I propose to demonstrate in vivo correction of ABCA4 mutations in the photoreceptors of an STGD1 mouse model using CRISPR/Cas9 technology. The STGD1 mouse model carries two point mutations found in STGD1 patients and displays the corresponding phenotype.
In Aim 1, I will validate the gene-editing efficacy of the CRISPR/Cas9 system in an explant culture derived from STGD1 mice.
In Aim 2, I will package the validated CRISPR/Cas9 system into AAV vectors and deliver them by subretinal injection in STGD1 mice.
In Aim 3, I will assess the correction of STGD1 mutations and phenotype in mice following the subretinal delivery. I envision that this proof-of-concept study will explore the efficacy of CRISPR/Cas9 technology as a new therapeutic approach for STGD1 and lay a solid foundation for the development of AAV and CRISPR/Cas9 combined treatment against any genetic ocular disease driven by a known pathogenic mutation.

Public Health Relevance

Stargardt disease (STGD1), the most common form of inherited juvenile macular dystrophy, results in progressive vision loss that currently cannot be effectively treated. In this study, we propose to develop a novel in vivo therapy to correct STGD1-associated mutations and demonstrate its efficacy in a STGD1 mouse model. This study will explore the clinical potential of the new therapy and lay the foundation for development of a general strategy for treatment of ocular diseases driven by known pathological mutations.

National Institute of Health (NIH)
National Eye Institute (NEI)
Individual Predoctoral NRSA for M.D./Ph.D. Fellowships (ADAMHA) (F30)
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Special Emphasis Panel (ZRG1)
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Agarwal, Neeraj
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Case Western Reserve University
Schools of Medicine
United States
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