Preclinical Safety Evaluation of AAV5-hRORA Gene Therapy for Geographic Atrophy and ABCA4-Related Retinopathies

<title>Abstract</title> <p>Geographic atrophy (GA) secondary to dry age-related macular degeneration (AMD) and ABCA4-related retinopathies, including Stargardt disease (STGD1), represent leading causes of irreversible central vision loss for which no disease-modifying therapies are currently approved. Despite their distinct genetic etiologies and ages of onset, both conditions converge on shared maculopathological mechanisms driven by lipofuscin accumulation within the retinal pigment epithelium (RPE), oxidative stress, complement dysregulation, chronic neuroinflammation, and progressive RPE and photoreceptor degeneration. We previously demonstrated that retinal delivery of RAR-related orphan receptor alpha (RORA) via an adeno-associated virus serotype 5 (AAV5) vector rescues the macular degeneration phenotype in the Abca4⁻/⁻ mouse model. Building on that proof-of-concept, this study evaluates the preclinical safety, tolerability, and biodistribution of OCU410 — an AAV5 vector encoding human RORA (AAV5-hRORA)—following bilateral subretinal administration in cynomolgus monkeys (Macaca fascicularis). Animals received a single 70 µL subretinal injection per eye at one of three dose levels: low (8.6 × 10¹⁰ vg/eye), medium (1.75 × 10¹¹ vg/eye), or high (3.5 × 10¹¹ vg/eye), and were followed for 6 months with comprehensive clinical, ophthalmic, immunological, and biodistribution assessments. Injection-related structural changes detected by optical coherence tomography (OCT) showed partial to complete resolution in most animals by Day 92. Multifocal pigmentary changes within the drug solution coverage area, observed by fundus photography and fluorescein angiography, were more frequent in the high-dose group and were attributed primarily to the subretinal delivery procedure rather than to RORA transgene expression. ERG amplitude changes were maximal at Day 29 and exhibited a recovery trend by Days 92 and 182; no dose-dependent pattern was identified, and implicit times remained largely stable across the observation period. Transgene-specific mRNA was predominantly confined to ocular tissues, with the highest expression levels in the retina, RPE-choroid complex, and iris-ciliary body; systemic biodistribution was negligible. RORA protein was below the lower limit of quantification (5 ng/mL) in all tissues assessed. No RORA-specific T-cell immunoreactivity was detected by ELISpot at any dose or timepoint. Anti-AAV5 antibody titers were dose-dependent in serum and aqueous humor, while anti-RORA antibodies were minimal in serum and absent in aqueous humor, consistent with the high sequence identity (92%) between human and cynomolgus monkey RORA. No systemic toxicity, treatment-attributable changes in body weight, or clinically significant intraocular pressure alterations were observed in any animal. These findings collectively establish the ocular safety and tolerability profile of OCU410.The highest tested dose, 3.5 × 10¹¹ vector genomes per eye, was considered tolerable and was selected as a supported dose for clinical development.</p>