Rho 1-2 meganuclease, an allele-specific gene-editing therapy, rejuvenates rod photoreceptor structure and function in a pig model of autosomal dominant Retinitis Pigmentosa (adRP).

Abstract

Purpose : The P23H mutation in the rhodopsin (RHO) gene represents the most common form of adRP in North America. Elimination of the causative mutant allele, while leaving the wild-type (WT) allele intact should eliminate vision loss in adRP patients. To this end, we studied the efficacy of the engineered Meganuclease, Rho1-2, in the transgenic P23H human rhodopsin (TgP23H) pig model of adRP.

Methods : Rho1-2, packaged in a self-complementary AAV5 vector and driven by the GRK1 photoreceptor-specific promoter was delivered subretinally (40 ul) to one eye of TgP23H piglets postnatally (P3-7,n=23). The fellow eye served as a control and either was uninjected or injected with vehicle. The efficacy of 2x10⁹, 6x10⁹, 2x10¹⁰, and 6x10¹⁰ vg were compared at regular post injection intervals. Fundus imaging and optical coherence tomography (OCT) assessed retinal structure. Full-field electroretinography (ffERG) assessed rod and cone function (ISCEV protocol). All pigs were assessed through P140 and a subset was assessed through P300. Immunohistochemistry/confocal assessed retinal morphology.

Results : From birth, untreated TgP23H pigs have no rod isolated ffERG response (Fernandez de Castro et al., IOVS, 2014). Regardless of dose, treatment with Rho1-2 was well tolerated, with no signs of an immune response. Treatment with two Rho1-2 doses (2 or 6x10¹⁰) produced a significant rod isolated ffERG b-wave response as early as P60 ((~14 uV; p =0.04 and 0.0003) n = 6 and 12 pigs, respectively). Mean b-wave amplitudes increased at P120 (~25-30 uV; p = 0.002, <0.0001 respectively) and were maintained through P300. There is a strong correlation between rod isolated function and rod morphology. Rho1-2 treated Tg P23H rods are more numerous, have elongated outer segments, and correctly localized rhodopsin in contrast to untreated areas in the same retina or in untreated Tg retinas. The ONL measured in OCT and retinal sections was significantly thicker in the treated vs. untreated areas/retinas (p<0.001).

Conclusions : Our results show that Rho1-2 meganuclease effectively rejuvenates rod structure and function in this TgP23H pig model of adRP and this gene therapy is well tolerated. This strongly indicates that Rho1-2 has the potential to effectively treat rod degeneration in P23H adRP patients

This is a 2021 ARVO Annual Meeting abstract.