# Molecular Mechanisms Limiting the Therapeutic Window of AAV Gene Therapy in Mouse Models of Blue Cone Monochromacy

## TL;DR

This study explores why gene therapy for a rare eye disorder is less effective in older mice, pointing to molecular and structural changes in retinal cells.

## Contribution

The study identifies molecular mechanisms limiting gene therapy efficacy in older BCM mouse models and suggests strategies to improve treatment.

## Key findings

- AAV8-Y733F capsid outperformed AAV5 in rescuing retinal cones in BCM mouse models.
- Degenerative changes in aged cones, such as mitochondrial mislocalization and ciliary compromise, reduce gene therapy efficacy.
- Cone-specific promoters like Pde6c and Cngb3 remain active in degenerating cones, offering potential for optimized gene therapy.

## Abstract

Blue cone monochromacy (BCM) is an X-linked retinal disorder caused by mutations in the OPN1LW/OPN1MW gene locus, resulting in impaired cone function and structural degeneration. We conducted a comparative analysis of AAV-mediated gene therapy in Opn1lw/Opn1mw double knockout (DKO) and Opn1mwC198R/Opn1sw−/− (C198R) BCM mouse models and evaluated the therapeutic window, efficacy, and longevity. Our results demonstrate that the AAV8-Y733F capsid achieved superior cone rescue compared to AAV5. While both DKO and C198R models showed similar therapeutic windows and rescue longevity, treatment efficacy decreased markedly in older mutant mice. Structural analysis revealed that aged cones in both models displayed degenerative changes, including mislocalized mitochondria and compromised connecting cilia. At the molecular level, we observed reduced AAV-mediated transgene expression in DKO and C198R older cones, which may result from decreased transduction efficiency, decreased circular episome stability, genome-wide transcription/translation downregulation, targeted mRNA/protein degradation, or overall cone degeneration. Notably, the cone-specific promoters for Pde6c and Cngb3 maintained robust activity in degenerating cones. These findings suggest that combining an efficient AAV serotype with an optimized cone promoter could be a viable approach to extend the therapeutic window and enhance treatment longevity for BCM patients.