88. Safety and Biodistribution Study of rAAV2tYF-PR1.7-hCNGB3 in Nonhuman Primates
Authors: Guo-jie Ye, Ewa Budzynski, Peter Sonnentag, T. Michael Nork, Paul E. Miller, Alok K. Sharma, James N. Ver Hoeve, Leia M. Smith, Tara Arndt, Roberto Calcedo, Chantelle Gaskin, Paulette M. Robinson, David R. Knop, William W. Hauswirth, Jeffrey D. Chulay
Journal: Molecular Therapy (2016)

Background: AGTC is developing a recombinant AAV 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. Here we report results of a toxicology and biodistribution study of this vector administered by subretinal injection in cynomolgus macaques. Methods: 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 × 1011 or 4 × 1012 vg/mL) and were evaluated for safety and biodistribution over a 3-month period prior to euthanasia. Toxicity assessment was based on mortality, clinical observations, body weights, ophthalmic examinations, intraocular pressure (IOP) measurements, electroretinography (ERG), visual evoked potentials (VEP), and clinical and anatomic pathology. Vector shedding and biodistribution was assessed by qPCR analyses. Immune responses to AAV and hCNGB3 were measured by ELISA, Elispot, or neutralization antibody assay for AAV2tYF. Results: There was no evidence of local or systemic toxicity and no changes in IOP, VEP responses, or hematology, coagulation or clinical chemistry parameters and no clinically important changes in ERG responses. Aqueous cells, sometimes with aqueous flare, were observed at the Day 3 evaluation in all groups and generally resolved or were at the mild (1+) levels by Week 4 and absent on Week 8 and thereafter except in one high dose animal. Posterior segment findings consisted of varying degrees of dose-related white vitreous cell, vitreous haze, white retinal perivascular sheathing, and white to grey-white subretinal infiltrates within and outside of the injection site. Vitreous haze resolved by Day 8 in eyes given vehicle control, by Week 4 in the low dose group and by Week 13 in the high dose group. Vitreous cells were observed at the mild (trace or 1+) level in the vehicle control group and resolved by Study Weeks 9 or 13 but persisted through Week 13 in a dose-related fashion in the low and high dose groups. Serum neutralizing antibodies against AAV2tYF were detected in all animals given vector. There were no T cell responses to AAV capsid peptides and no antibody or T cell responses to hCNGB3. Mononuclear cell infiltrates in the vitreous body/optic disc, of minimal intensity, in the vector-injected eye of all animals at both dose levels. All other tissues collected for histopathological examination showed no abnormalities. Results of biodistribution studies demonstrated that the vector did not spread widely or consistently outside the injected eye. High levels of vector DNA were found in vector-injected eyes but minimal or no vector DNA was found in any other tissue. Conclusions: Subretinal injection of rAAV2tYF-PR1.7-hCNGB3 at concentrations of 4 × 1011 or 4 × 1012 vg/mL was associated with a dose-related anterior and posterior segment inflammatory response that improved over time. There was no evidence of systemic toxicity and no changes in IOP, VEP responses, or hematology, coagulation or clinical chemistry parameters and no clinically important changes in ERG responses. These results support the use of rAAV2tYF-PR1.7-hCNGB3 in clinical studies in patients with achromatopsia. A Phase 1/2 clinical trial evaluating rAAV2tYF-PR1.7-hCNGB3 in patients with achromatopsia is scheduled to begin in 2016.