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Elizabeth M Simpson, Andrea J. Korecki, Siu Ling Lam, Jack W. Hickmott; Intrastromal Delivery of rAAV PAX6 Transiently Rescues Corneal Defects in a Mouse Model of Aniridia. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3417.
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Aniridia is a rare autosomal-dominant blinding disorder, caused by loss-of function mutations in the gene paired box 6 (PAX6). Born with poor vision, vision loss advances in early adulthood. Corneal pannus and keratopathy obscure light entering the eye and are major underlying causes of postnatal vision loss in many patients. Current treatments, such as corneal transplant and prosthetic corneal implants temporarily improve visual acuity, but fail to provide lasting vision, necessitating the need for new treatments. Gene therapy has become a successful treatment avenue for rare disorders, and mouse models are typically used in developing new therapies. The Pax6Sey/+ (Sey) mouse model of aniridia presents with corneal clouding, pannus, and reduced corneal thickness. Here we test the hypothesis that supplementation of PAX6 in the adult Sey cornea can ameliorate corneal thinning using recombinant adeno-associated virus (rAAV) mediated augmentation gene therapy.
A 3xFLAG/PAX6 open reading frame was cloned into a rAAV genome plasmid and commercially packaged into serotype rAAV9. Adult wild type (WT) and Sey mice were administered virus by intrastromal injection of one of three treatments: PBS, rAAV9 smCBA-EmGFP-WPRE, or rAAV9 smCBA-3xFLAG/PAX6-WPRE. Contralateral eyes were used as non-injected controls. Mice were monitored for either one or two weeks and then harvested for histological analysis including measurements of corneal epithelial thickness and immunofluorescent staining for PAX6 and EmGFP. Analysis was performed blinded to treatment group.
Intrastromal injection of rAAV transduces the cornea, with EmGFP expression detectable at both one two weeks. At one week, PAX6 treated Sey mice had a significantly thicker corneal epithelia than untreated control Sey mice (p<0.005), and the epithelia were not significantly thinner than Wt controls. Treated Sey corneas had an intermediate number of epithelial cells: more than Sey (p<0.01) but less than Wt (p<0.05) controls. By two weeks, no significant differences were observed. At both time points, no significant reduction in epithelial thickness or the number of epithelial cells was detected in treated Wt mice.
The transient success of PAX6 gene therapy paves the way for delivery and dose optimization, longitudinal studies, and functional characterization.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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