Optogenetic engineering of retinal ganglion cells with AAV2.7m8-ChrimsonR-tdTomato (GS030) in combination with an event-based camera photostimulation device

Abstract

Purpose : Ectopic expression of a light-sensitive opsin in retinal ganglion cells (RGCs) is an attractive strategy to restore vision. Here, we evaluated the ability of ChrimsonR-tdTomato (ChrR-tdT), developed by Ed Boyden (MIT), to convert light insensitive RGCs into photoactivable cells both in rd1 mice and normal macaques. A photostimulation device inspired by natural vision has also been developed.

Methods : rd1 mice and Cynomolgus macaques were injected intravitreally with AAV2.7m8 vector encoding ChrR-tdT (GS030). Multielectrode array (MEA) measurements and expression of tdT were assessed in explanted retinas 1 and 2 months after injections in mice and macaques, respectively. Local tolerance was also evaluated in macaques by ophthalmic exams and histology.

Results : In both rd1 mice and normal macaques, ChrR-tdT was essentially expressed in RGCs. In macaques, its expression was restricted to the perifoveal area, and was not associated with any structural modifications, inflammation or necrosis; all ophthalmic exams were normal. Patch-clamp experiments, conducted on monkey retina by targeting ChrR-tdT-expressing RGCs, allowed biophysical characterization of ChrR-tdT-elicited currents. MEA recordings showed that light responses followed the action spectrum of ChrR-tdT. Activity was correlated with TdT fluorescence reaching very high spiking frequencies (above 300Hz). Required light levels were below safety radiation limits. A complete prototype of the photostimulation device was developed with the optical chain, a functional electronic, firmware and software architecture. It captures external scenes through an event-based camera over a high dynamic range to deliver visual stimulations onto the ChrR-tdT-transduced retina.

Conclusions : Our GS030 vector efficiently and safely photo-engineered RGCs in both blind mice and normal macaques. ChrR-tdT-based gene therapy combined with a photostimulation device appears to be a viable treatment option for vision restoration in RP patients.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.