Safety and Biodistribution Evaluation in Cynomolgus Macaques of rAAV2tYF-PR1.7-hCNGB3, a Recombinant AAV Vector for Treatment of Achromatopsia

Abstract

Applied Genetic Technologies Corporation (AGTC) is developing rAAV2tYF-PR1.7-hCNGB3, a recombinant adeno-associated viral (rAAV) vector expressing the human CNGB3 gene, for treatment of achromatopsia, an inherited retinal disorder characterized by markedly reduced visual acuity, extreme light sensitivity, and absence of color discrimination. We report here results of a study evaluating the safety and biodistribution of rAAV2tYF-PR1.7-hCNGB3 in cynomolgus macaques. Three groups of animals (n = 2 males and 2 females per group) received a subretinal injection in one eye of 300 μl containing either vehicle or rAAV2tYF-PR1.7-hCNGB3 at one of two concentrations (4 × 10(11) or 4 × 10(12) vector genomes/ml) and were evaluated over a 3-month period before being euthanized. Administration of rAAV2tYF-PR1.7-hCNGB3 was associated with a dose-related anterior and posterior segment inflammatory response that was greater than that observed in eyes injected with the vehicle control. Most manifestations of inflammation improved over time except that vitreous cells persisted in vector-treated eyes until the end of the study. One animal in the lower vector dose group was euthanized on study day 5, based on a clinical diagnosis of endophthalmitis. There were no test article-related effects on intraocular pressure, visual evoked potential responses, hematology or clinical chemistry parameters, or gross necropsy observations. Histopathological examination demonstrated minimal mononuclear infiltrates in all vector-injected eyes. Serum anti-AAV antibodies developed in all vector-injected animals. No animals developed antibodies to CNGB3. Biodistribution studies demonstrated high levels of vector DNA in the injected eye but minimal or no vector DNA in any other tissue. These results support the use of rAAV2tYF-PR1.7-hCNGB3 in clinical studies in patients with achromatopsia caused by CNGB3 mutations.

Figure 1.

Ocular inflammation findings. Intensity of parameters in individual animals was scored in a standardized fashion as 0, trace (0.5), 1+, 2+, 3+, or 4+ as described in Research Design and Methods. Each color and symbol represents an individual animal, with four animals per dosing group. One animal in group 2 was euthanized on day 5, based on a clinical diagnosis of endophthalmitis. One animal in group 1 developed a cataract after an intraoperative posterior lens capsule tear; vitreous cells and haze could not be scored after study day 3 in this animal.

Figure 2.

Histological findings. Top left: A normal optic disc and retina in an eye injected with the vehicle control article. Top right: A normal retina from an uninjected eye. Inset shows the absence of mononuclear infiltrates around the blood vessel. The lower left panel shows mononuclear infiltrates around the blood vessel in the optic disc of an animal given 4.0 × 10¹¹ vg/mL, and the inset shows a higher magnification of the blood vessel. The lower right panel shows mononuclear infiltrates around a retinal blood vessel and in the vitreous body in an animal given 4.0 × 10¹² vg/mL, and the inset shows a higher magnification of the optic nerve. The low magnification was 10× and high magnification (insets) was 40×.