Determination of the optimal time window for gene replacement therapies for blindness using an inducible rescue mouse model of Bardet-Biedl Syndrome

Genes that disrupt outer segment (OS) formation cause retinal degeneration, including genes that code for components of the BBSome, a complex known to regulate protein transport in the primary cilia. Loss of BBSome function in the retina causes mislocalization of numerous inner segment (IS) proteins to the OS, including syntaxin-3. We previously shown that restoration of BBSome function prior to the maturation of the retina can fully rescue retinal phenotypes. However, it is not known whether remaining photoreceptors in mid- to late- stage degenerating retinas can be successfully rescued by restoration of the target gene. We aim to define the optimal time window for gene replacement therapies by restoring BBSome function at different stages of retinal degeneration.
We used a mutant mouse model in which the Bbs8 gene, coding for a member of the BBSome complex, is inhibited by a gene trap cassette. The gene trap can be eliminated by tamoxifen inducible FLP recombinase to provide precise temporal control over the restoration of BBSome function in these mice. BBSome function was restored by tamoxifen injections at 5 different ages: a) postnatal days 9-15 (P9-P15) when the OS develops, b) P21-P30, c) 6 weeks, d) 3.5 months, and e) 6 months of age. We performed immunohistochemistry, transmission electron microscopy, electroretinography (ERG), and functional vision assay to measure rescue effectiveness.
We found that photoreceptors can be successfully rescued even in mid-stage degenerating retinas. In unrescued mice, syntaxin-3 is mislocalized to the OS, and photoreceptors are gradually lost over time. In mice rescued within the effective time window, photoreceptor cell death is arrested, syntaxin-3 is localized to the IS, and their retinal function is sustained for at least 6 months. Interestingly, re-establishing completely normal outer segment structure is not required for photoreceptor survival in these rescued mice. Even though the outer segment shows an age-dependent morphogenic potential, photoreceptors rescued within the time window show long-term survival and syntaxin-3 localization to the IS despite possessing OSs with notable morphological defects.
Photoreceptors in early- to mid- stages of degeneration are amenable to rescue by restoration of target gene expression in a mutant mouse model of the BBSome.