Gene therapy rescues cone function and structure in a novel mouse model of blue cone monochromacy with the most common C203R missense mutation

Abstract

Purpose : Blue cone monochromacy (BCM) is an X-linked inherited vision disorder resulting in loss of functional L- and M- cones. A C203R missense mutation in the OPN1LW/MW gene cluster is one the most common causes of BCM. In this study, we provide the first data demonstrating the effectiveness of AAV-mediated gene therapy in a mouse model of BCM carrying the equivalent of a human C203R missense mutation.

Methods : Opn1mw^C198ROpn1sw^-/- mice were generated and characterized as described previously. Gene therapy utilizing AAV5-mediated expression of human OPN1LW, driven by a cone-specific PR2.1 promoter, was delivered by subretinal injection at 1-, 3-, and 5-months of age. Visual function was evaluated at 1-, 4-, and 7-months post-injection by electroretinography (ERG). Transgene expression and cone outer segments (COS) were analyzed by immunohistochemistry (IHC) using antibodies against L/M-opsin, GNAT2, and PDE6C.

Results : Expression of mutant OPN1MW^C198R protein was undetectable by IHC, suggesting it is degraded efficiently. Gene therapy showed rescue of cone function, rebuilding of COS, and restoration of COS proteins in mice treated at 1- and 3- months of age. Visual function assessed by photopic ERG 1-month post-injection was significantly higher in treated eyes compared to untreated, with a b-wave amplitude of 72±13μV and 61±9μV (avg±SD) in 1- and 3-month injected eyes compared to a flat ERG in untreated eyes. Rescue lasted long-term, with a significantly higher ERG even at 7-months post-injection. In cones expressing L-opsin, we observed replenished expression of GNAT2 and PDE6C, all of which were properly localized to the COS. Alternatively, 5-month treated Opn1mw^C198ROpn1sw^-/- cones showed reduced rescue of function following gene therapy, with a b-wave amplitude of 26±6μV in treated eyes. Transgene expression in 5-month treated eyes was also more difficult to find, with only several cones expressing L-opsin, despite ~50% of viable cones remaining.

Conclusions : Regardless of the potential toxicity of mutant OPN1MW^C198R, Opn1mw^C198ROpn1sw^-/- cones remain viable targets for gene replacement therapy. We demonstrated robust functional and structural rescue, similar to what we have previously reported in a deletion mutation model of BCM. Together, both models demonstrate that early intervention is critical for gene therapy to be advantageous in treating BCM.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.