Progressive retinal atrophy (PRA) is an inherited retinal dystrophy that affects over 100 breeds of dogs. It is characterized by a bilateral retinal degeneration commonly resulting in blindness. Affected dogs typically present with loss of dim light vision, attenuation of retinal blood vessels and tapetal hyperreflectivity. The purpose of this study was to identify the underlying cause of PRA in Papillon dogs and to characterize the phenotype at the cellular and molecular levels. I identified a mutation in the gene CNGB1 which accounts for 70% of the PRA in the Papillon dogs. The CNGB1 mutation involves a 6 base pair insertion and a 1 base pair deletion resulting in exon skipping and a premature stop codon cause by a frameshift. CNGB1 encodes the β-subunit of a cyclic nucleotide-gated ion (CNG) channel. CNGB1 has multiple splice variants expressed in rod photoreceptors, olfactory sensory neurons and other tissues. CNG channels are directly involved in rod phototransduction and olfactory transduction. The retinal phenotype of the CNGB1 affected dogs was characterized by in vivo and ex vivo analyses. Electroretinograms (ERGs) and behavioral vision testing were conducted to assess retinal function throughout the course of the disease. The CNGB1 affected dogs had decreased and abnormal rod function at the earliest age tested but cone function was preserved until 5.5 years of age. Histological analyses showed that the morphology of rod photoreceptors deteriorate slowly over the first ~1.5 years of life while cone photoreceptor morphology is preserved for longer.Adeno-associated viral (AAV) vector therapy was used to treat five CNGB1 affected dogs with a wild-type copy of canine CNGB1 cDNA. One eye was injected with a low titer (1x1012) of an AAV vector delivering CNGB1 cDNA, six eyes were injected with a higher titer (5x1012) and one eye was injected with a GFP-expressing construct as a vehicle and procedural control. All dogs treated with the AAV vector containing the wild-type copy of CNGB1 showed rescue of rod function that was maintained throughout the time course of the study (9 months).The CNGB1 affected dog olfactory phenotype was investigated using in vivo and ex vivo techniques. The olfactory epithelium and the olfactory bulbs were abnormal in the CNGB1 affected dogs when compared to control dogs. I developed a behavioral test that could assess olfactory function in the CNGB1 dogs. The CNGB1 affected dogs had decreased but not absent olfactory function. The detailed descriptions of the retinal and olfactory phenotypes, in addition to the successful gene replacement therapy trial, in the CNGB1 affected dogs have laid the ground work for future studies including working with clinicians to advance gene replacement therapy trials in human patients with mutations in the CNGB1 gene.
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