Testing of a therapeutic vector for CNGB1 retinitis pigmentosa

Abstract

Purpose : To test the efficacy of an adeno-associated virus (AAV) vector designed for the treatment of CNGB1-Retinitis pigmentosa using a spontaneous Cngb1 mutant dog model.

Methods : An AAV5 vector packaged with a human CNGB1a cDNA under control of a novel short (194 bp) human rhodopsin promoter with a minimal SV40 polyadenylation signal was developed and produced by a triple transfection method. Ten young Cngb1 mutant dogs were used. Vector was delivered by subretinal injection aiming to create two ~100µl blebs in the superior fundus. Two eyes were untreated, a titer of 1E11vg/ml was injected in 3 eyes and 5E11, 1E12 and 5E12vg/ml were each injected in 5 eyes. All dogs were followed for 6 months and 7 (12 treated eyes) up to 12 months post injection. In life outcome measures were assessment of rod-mediated vision, electroretinography (ERG) and optical coherence tomography (OCT). Following euthanasia eyes were processed for immunohistochemistry (IHC).

Results : All doses resulted in a similar marked improvement in dim-light vision in all dogs. This was maintained for the duration of the study. ERG showed restoration of rod function in all eyes with a dose-effect. Two out of five eyes in the 1E12 group developed some areas of retinal degeneration in the injected region. In these dogs this was associated with a reduction in rod-mediated ERG amplitudes, although amplitudes remained higher than in the control eyes. In one of these eyes OCT had suggested the presence of inflammation 3 months post injection, which was treated with systemic steroids, but the changes were mild and not apparent at that time on fundus examination. The ERG rescue in all eyes was maintained for the duration of the study. OCT in all eyes showed improved appearance of the outer retina representing the inner and outer segments (e.g. presence and continuity of the ellipsoid zone) and preservation of retinal structure in the treated areas. The two 1E12 eyes with patchy degeneration in the treated region also showed regions of structural rescue within the injected area. IHC showed expression of the transgene and its co-channel subunit, Cnga1 within the treated retinal regions and reversal of accumulation of cGMP that occurs in this model.

Conclusions : This new AAV5.CNGB1 vector was shown to effectively rescue the Cngb1 mutant dog phenotype. The dose escalation study suggests a safe maximal titer that could be used in a future clinical trial in CNGB1-RP patients.

This is a 2020 ARVO Annual Meeting abstract.