Gene therapy rescued cones in an all-cone mouse model of blue cone monochromacy with the most common cone opsin missense mutation

Abstract

Purpose : Blue cone monochromacy (BCM) is a congenital X-linked vision disorder characterized by absence or severely diminished L- and M- cone function. Recently, we demonstrated that gene therapy rescues cone function and structure in Opn1mw^C198ROpn1sw^-/- mice which carry the most common missense mutation (C203R) linked to BCM. As L- and M- cones are concentrated within fovea, we tested gene augmentation therapy in all-cone Opn1mw^C198Opn1sw^--/-Nrl^-/-(C198R-AC) mice, whose retinas mimic the fovea of BCM patients with C203R mutation.

Methods : C198R-AC mice were generated by Opn1mw^C198ROpn1sw^-/- and Nrl^-/- mice. C198R-AC eyes were injected subretinally at 1M and 5M with AAV8-Y733F expressing hOPN1LW under a PR2.1 cone-specific promoter. Photopic electroretinography (ERG) was performed at 1M and 5M post-injection. Structural rescue of cones was examined by immunohistochemistry (IHC) with antibodies against OPN1LW/MW, PDE6H, and GNAT2.

Results : C198R-AC cones lack photopic ERG responses and exhibit shortened cone outer segments (COS), structurally and functionally mimicking BCM foveal cones carrying the corresponding C203R missense mutation. Similar to our previous studies in Opn1mw^C198ROpn1sw^-/- mice, we were unable to detect OPN1MW^C198R mutant protein by IHC. AAV8-Y733F-mediated gene therapy rescued cone function and structure and rescue persisted for at least 5M in both 1M and 5M injected eyes. The average b-wave amplitude in 1M injected C198R-AC mice at 1M and 5M post-injection was 156±18μV (n=6, p<0.0001) and 117±29μV (n=6, p<0.0005), respectively, which was significantly higher than untreated contralateral controls. In 5M treated C198R-AC animals, ERG responses were 71±13μV (n=6, p<0.0001) and 74±23μV (n=6, p<0.005) at 1M and 5M post-injection, respectively. IHC demonstrated that gene therapy restored COS structure, as we detected ample GNAT2 and PDE6H expression to the COS in 1M and 5M treated C198R-AC cones.

Conclusions : We observe robust rescue of cone function and restoration of COS proteins GNAT2 and PDE6H following gene therapy in 1M and 5M treated C198R-AC mice. C198R-AC cones, which mimic the foveal cones of a BCM patient carrying the C203R missense, are an excellent model for improving future gene therapy.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.