bipolar cell; Retinitis pigmentosa; ophthalmology; gene therapy; metabotropic gluatmate receptors; optogenetics; melanopsin; Age-related macular degeneration; recombinant adeno-associated virus; vision recovery; retinal physiology; photoreceptor degeneration; molecular engineering
van Wyk Michiel, Hulliger EC, Girod L, Ebneter A, Kleinlogel S (2017), Present Molecular Limitations of ON-Bipolar Cell Targeted Gene Therapy., in Frontiers in Neuroscience
, 11, 116.
Kleinlogel Sonja (2017), Optogenetics for Vision Recovery: From Traditional to Designer Optogenetic Tools., in Appasani Krishnarao (ed.), 327-355.
Perny Michael, Muri Lukas, Dawson Heather, Kleinlogel Sonja (2016), Chronic activation of the D156A point mutant of Channelrhodopsin-2 signals apoptotic cell death: the good and the bad., in Cell Death and Disease
, 7, e2447.
Tscherter Anne, Heidemann Martina, Kleinlogel Sonja, Streit Jürg (2016), Embryonic Cell Grafts in a Culture Model of Spinal Cord Lesion: Neuronal Relay Formation Is Essential for Functional Regeneration., in Frontiers in Cellular Neuroscience
, 10, 220.
Kleinlogel Sonja (2016), Optogenetic user’s guide to Opto-GPCRs, in Frontiers in Biosciences
, 21, 794-805.
van Wyk Michiel, Schneider Sabine, Kleinlogel Sonja (2015), Variable phenotypic expressivity in inbred retinal degeneration mouse lines: A comparative study of C3H/HeOu and FVB/N rd1 mice., in Molecular Vision
, 21, 811-827.
van Wyk Michiel, Pielecka-Fortuna Justyna, Löwel Siegrid, Kleinlogel Sonja (2015), Restoring the ON-switch in blind retinas: Opto-mGluR6, a next-generation optogenetic tool., in PLoS Biology
, 13(5), e1002143.
Background: Optogenetics is the combination of genetic and optical methods to control the activity of genetically targeted cells of living tissue remotely by light1. The traditional optogenetic tools are microbial ion channels (i.e. Channelrhodopsin-2, ChR2) and ion pumps (i.e. Halorhodopsin, eNphR), which are heterologously introduced into the target cell types using molecular and genetic methods. Optogenetics has in the last few years revolutionized the field of neuroscience, being elected as Method of the Year by “Nature Methods” in 2010. The clinical potential of a handful of existing optogenetic tools is actively investigated at many fronts. Having a profound knowledge in this field, I am aware of their limitations, which will hinder their use in the clinic. It is thus imperative to develop next-generation light-activatable proteins, engineered to be safe, specialized and potent enough to be employed clinically. About one in three hundred people suffer from complete or partial blindness as a result of photoreceptor loss associated with diseases such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD), for which presently very limited treatment exists. Due to the light-accessibility and the low immunogenicity of the retina, optogenetics is a prime candidate technology for treating blindness. The goal of this investigation is to develop a gene therapy with a next-generation, custom-made optogenetic tool for retinal bipolar cells, which can render them directly light-sensitive to replace the function of the degenerated photoreceptors. Hypothesis:1)Opto-mGluR6 fulfills all the clinical requirements: non-immunogenic, non-toxic and function in photopic light conditions (major improvement compared to traditional optogenetic tools)2)Opto-mGluR6 recovers the visual function in the retina as well as the cortex with high visual resolution (major improvement compared to retinal implants)3)The Opto-mGluR6 gene therapy is transferrable into human tissue Methods: Recovery of retinal visual function will be determined in Opto-mGluR6 treated mice suffering from Retinitis pigmentosa by patch-clamp and electroretinogram recordings. Functional integrity of higher visual centers will be evaluated by behavioral (optomoter reflex, Morris water maze) functional cortical imaging experiments. In parallel, optimized rAAVs will be engineered to reach near-complete infection of the patient’s ON bipolar cells. The methodology will then be transferred into human post-mortem tissue. To suffice clinical requirements, toxicology and immunology studies on Opto-mGluR6 gene-therapeutically treated mice will be performed.Specific Aims:1.To prove that Opto-mGluR6 recovers the natural visual signal transduction cascade within the retina - and thus nearly perfect contrast vision - by simply replacing the missing light-sensor (photoreceptor cells).2.To engineer an optimal rAAV for retinal ON bipolar cells3.To optimize Opto-mGluR6 for human tissue (codon-optimization, specific promoter identification,…)4.To prove that the Opto-mGluR6 gene therapy is non-toxic and non-immunogenic5.To develop a product so ready for clinical trials.Significance: Opto-mGluR6 could bring back high-quality vision to blind patients suffering from photoreceptor degenerative diseases allowing for tasks such as driving a car or reading a book, whilst the only available treatment presently, retinal implants, only allow patients to distinguish high contrast edges to for example avoid an obstacle. This project combines two highly promising and emerging fields in regenerative medicine and in particular ophthalmology, which are rAAV gene therapy and optogenetics. The here proposed project optimizes the two technologies and aims to generate in just 3 years a product suited for clinical trials.