First pilot study of intrastromal rAAV-PAX6 gene therapy suggests improved corneal thickness and transcription correction in aniridic mouse.

Aniridia is a rare congenital vision-loss disorder caused primarily by loss-of-function variants of the paired box 6 (PAX6) gene. There is currently no cure. Augmentation gene therapy has emerged as a successful treatment for inherited ocular diseases. Here, we conducted the first pilot preclinical intrastromal augmentation gene therapy for aniridic cornea. It was undertaken in an aniridia mouse model, using adeno-associated virus 9 (AAV9), with a ubiquitous promoter driving PAX6, delivered by injection at 3 months, and harvest at 5 months post-injection (PI). The primary endpoint was histologically quantified epithelial thickness. The secondary endpoints were the slit lamp assessment of keratopathy and the RT-ddPCR measurement of four key signaling genes in the cornea. At 5 months PI, we demonstrated virally delivered PAX6 protein in the aniridic mouse corneal stroma. Critically, we found a statistically significant, albeit suggestive due to low n values, structural improvement in the corneal epithelial thickness. When the data were pooled, we also demonstrated a significant and complete transcription correction of four key genes-Notch1, Hes1, Wnt5a, and Mtor. We explore a variety of hypothesizes to explain these initial results, including that the virally delivered PAX6 is acting non-cell autonomously, moving from the non-dividing stroma into the epithelium, causing structural and molecular improvement.