Long-term cone functional rescue was achieved in dogs with achromatopsia AAV vectors containing codon-optimized human CNGB3 and an improved promoter, which may be a stepping stone to treating achromatopsia in humans.
Take-home message: Long-term cone functional rescue was achieved in dogs with achromatopsia AAV vectors containing codon-optimized human CNGB3 and an improved promoter, which may be a stepping stone to treating achromatopsia in humans.
By Lynda Charters; Reviewed by András Komáromy, DrMedVet, PhD
East Lansing, MI-Adeno-associated virus (AAV) vectors containing codon-optimized human CNGB3 (hCNGB3co) driven by the PR1.7 cone-specific promoter achieved long-term cone functional rescue in dogs with CNGB3 mutant achromatopsia.
These results may pave the way for treating all cone subclasses in patients with CNGB3-related achromatopsia.
In humans, achromatopsia is an autosomal recessive disease that affects at least six genes: CNGA3, CNGB3, GNAT2, PDE6C, PDE6H, and ATF6, with about 27% and 50% of cases, respectively, having mutations in the first two of those genes, according to András Komáromy, DrMedVet, PhD, associate professor of ophthalmology, Michigan State University’s College of Veterinary Medicine, East Lansing, MI.
The condition is characterized by the absence of cone photoreceptor function resulting in rod monochromacy. The clinical signs include total color blindness, daytime blindness, photophobia, low visual acuity, and nystagmus. Individuals with achromatopsia can have varying degrees of color perception ranging from only white, black, and grey, to a limited ability to see some colors.
Dr. Komáromy recounted that an early study in Human Molecular Genetics (2002;11:1823-1833) established autosomal recessive cone degeneration in dogs as orthologous to the human achromatopsia locus ACHM3, and caused by autosomal recessive mutations of CNGB3, specifically a genomic deletion (CNGB3-/-) and a D262N missense mutation exon 6 (CNGB3m/m).
In 2010, while at the University of Pennsylvania School of Veterinary Medicine, Dr. Komáromy â together with Gustavo Aguirre, VMD, PhD, professor of medical genetics and ophthalmology, University of Pennsylvania, and colleagues â reported that cone function was rescued by gene therapy in dogs with achromatopsia (Human Molecular Genetics. 2010;19:2581-2593).
The primary study findings were that AAV5 expressing hCNGB3 driven by a PR2.1 promoter achieved long-term red/green cone functional rescue and that shorter versions of PR2.1 promoter (PR0.5 or 3LCR-PR0.5) achieved only transient functional rescue.
In the current study, the Michigan State-University of Pennsylvania collaboration continued as Drs. Komáromy and Aguirre and their teams evaluated the efficacy of an AAV vector, designed and produced by Applied Genetics Technology Corporation, expressing hCNGB3co cDNA driven by a PR1.7 promoter and compared the results with other cone-specific promoters for rescue of cone function in CNGB3 mutant dogs.
Dr. Komáromy explained that the efficiency of cone functional rescue by AAV vectors expressing hCNGB3 or hCNGB3co cDNAs and driven by PR1.7 or PR2.1 promoters and packaged in AAV5, AAV9, or AAV2tYF mutant capsids was evaluated in CNGB3 mutant dogs by subretinal injection.
Photopic flicker electroretinography (ERG) amplitudes were recorded at baseline and 8 and 12 weeks after vector injection in all animals, and then once every 3 months after that in a subset of animals. Objective behavioral testing of day vision also was performed at different time points.
The results showed that ERG cone responses were rescued with both hCNGB3 and hCNGB3co in three eyes treated with AAV5-PR2.1-hCNGB3co, five eyes treated with AAV5-PR2.1-hCNGB3, four eyes treated with AAV9-PR1.7-hCNGB3co, five eyes treated with AAV2tYF-PR1.7-hCNGB3co, and three eyes treated with AAV5-PR1.7-hCNGB3co vector. Hence, rescue of cone function and day vision was not only comparable for both hCNGB3 and hCNGB3co, but also for all three AAV serotypes evaluated, i.e., AAV5, AAV9, and AAV2tYF.
“Two animals treated with AAV2tYF-PR1.7-hCNGB3co in one eye and AAV9-PR1.7-hCNGB3co in the other eye continued to show robust ERG responses for at least 8 months after treatment with no decrease in the ERG amplitude and continued restoration of behavioral day vision,” Dr. Komáromy said.
AAV vectors expressing codon-optimized hCNGB3 driven by the PR1.7 cone-specific promoter achieved safe, long-term cone functional rescue in CNGB3-mutant achromatopsia dogs, he noted.
“Because the PR1.7 promoter directs robust and specific gene expression not only in red and green cones, but also in blue cones in non-human primates, these results support development of an AAV-PR1.7-hCNGB3co vector for treating patients with CNGB3-achromatopsia,” Dr. Komáromy said.
András Komáromy, DrMedVet, PhD
Dr. Komáromy has no financial interest in any aspect of this report.