Development of an AAV vector for treatment of inherited retinal dystrophy caused by RPE65 deficiency

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Leber congenital amaurosis (LCA) is a severe inherited retinal dystrophy with a population frequency around 1/50,000, characterised by loss of vision in childhood. Approximately 5% of LCA is caused by mutations in the RPE65 gene, which encodes a retinal pigment epithelium (RPE) specific protein essential for recycling of retinoids to the photoreceptor cells. The results of our trial, and 2 similar trials in the US, show for the first time the impact of this novel therapy on a condition that was previously untreatable. Whilst some subjects in all the trials have benefited from this new intervention, the extent of response in human subjects does not match the improvements in mice and dogs with the same genetic defect. Our data indicates that this difference reflects a greater requirement for RPE65 in humans than in rodents and dogs, and this is not fully met with the current version of the vector. The aim of this project is to improve the efficacy of gene therapy for LCA2 by optimising the RPE65 construct through a number of specific modifications. Since we have reached the maximal tolerated vector dose in these subjects, we intend to improve the efficiency of transgene expression levels by optimising the construct through a number of specific patentable modifications. By improving the efficiency of d protein production we aim to achieve a more than 100-fold reduction in the vector dose required for the optimal treatment in animal models of the disease. The RPE65 protein levels obtained from the improved vector will be tested initially in vitro. One or two constructs with a proven ability to drive higher levels of expression in vitro will be used in animal studies, first in mice and subsequently in dogs in order to test whether lower doses of the new vector can achieve equivalent rescue of retinal function and rescue of vision. Successful completion of this project will enable us to develop a clinical trial of an improved AAV vector for the treatment of LCA2. A more effective construct should enable us to develop a highly effective gene therapy vector that has the potential to be one of the world's first first licenced gene therapy products for clinical application. This would have considerable impact on the quality of life of the patients, with benefits ranging from improved nightvision to improved visual acuity and preservation of vision over time, depending on the condition of the retina at the time of treatment. As the best outcome is expected after treatment early in life, most commonly the end users would be pre-teen LCA2 patients, although ongoing clinical trials have shown that patients up to twenty years of age can benefit from treatment. The young age of the subjects means that a optimally successful treatment outcome could result in years of increased visual acuity and/or a wider visual field, with concomitant improvements in self-sufficiency and potentially other economic benefits. LCA is a relatively rare condition with a population frequency of approximately 1/50,000. In the UK around 5% of all forms of LCA are caused by RPE65 mutations and thus potential end-users for AAV-RPE65 gene therapy. Only patients under the age of 20 are likely to benefit from treatment; however, in this group a near-complete take-up of an established therapy is expected. Retinal dystrophy is a highly diverse group of disorders that can be caused by mutations in as many as 200 different genes. The core technology used in this application - subretinal injection of AAV vectors carrying a transgene - will be suitable for the development of gene therapy protocols for the majority of retinal dystrophies. Robust proof-of-concept studies have been reported for AAV-mediated gene therapy for ten forms of inherited retinal degeneration, of which we have reported five. The results obtained in this project will also facilitate the progression of these therapies to clinical application.