INSERM U968, Institut de la Vision, 75012 Paris, France; UMRS968, Institut de la Vision, Sorbonne Universités, Pierre et Marie Curie University (UPMC) University Paris 06, 75012 Paris, France; Centre National de la Recherche Scientifique (CNRS) UMR7210, Institut de la Vision, 75012 Paris, France.
Sanofi Ophthalmology Unit, 17 rue Moreau, 75012 Paris, France.
Mol Ther. 2017 Nov 1;25(11):2546-2560. doi: 10.1016/j.ymthe.2017.07.011. Epub 2017 Jul 20.
The majority of inherited retinal degenerations converge on the phenotype of photoreceptor cell death. Second- and third-order neurons are spared in these diseases, making it possible to restore retinal light responses using optogenetics. Viral expression of channelrhodopsin in the third-order neurons under ubiquitous promoters was previously shown to restore visual function, albeit at light intensities above illumination safety thresholds. Here, we report (to our knowledge, for the first time) activation of macaque retinas, up to 6 months post-injection, using channelrhodopsin-Ca-permeable channelrhodopsin (CatCh) at safe light intensities. High-level CatCh expression was achieved due to a new promoter based on the regulatory region of the gamma-synuclein gene (SNCG) allowing strong expression in ganglion cells across species. Our promoter, in combination with clinically proven adeno-associated virus 2 (AAV2), provides CatCh expression in peri-foveolar ganglion cells responding robustly to light under the illumination safety thresholds for the human eye. On the contrary, the threshold of activation and the proportion of unresponsive cells were much higher when a ubiquitous promoter (cytomegalovirus [CMV]) was used to express CatCh. The results of our study suggest that the inclusion of optimized promoters is key in the path to clinical translation of optogenetics.
大多数遗传性视网膜退行性疾病的表型都集中在光感受器细胞死亡上。这些疾病中第二级和第三级神经元得以幸免，这使得使用光遗传学恢复视网膜光反应成为可能。先前已经证明，在普遍启动子的作用下，第三级神经元中通道型视紫红质的病毒表达可以恢复视觉功能，尽管是在超过光照安全阈值的光强下。在这里，我们报告（据我们所知，首次）使用钙通透性通道型视紫红质（CatCh）在安全光强下激活猕猴视网膜，时间可长达注射后 6 个月。由于基于γ-突触核蛋白（SNCG）基因调节区的新型启动子允许在跨物种的神经节细胞中实现高水平的 CatCh 表达，因此实现了高水平的 CatCh 表达。我们的启动子与经过临床验证的腺相关病毒 2（AAV2）相结合，在人眼的光照安全阈值下，在近黄斑的神经节细胞中提供对光反应灵敏的 CatCh 表达。相反，当使用普遍启动子（巨细胞病毒 [CMV]）表达 CatCh 时，激活的阈值和无反应细胞的比例要高得多。我们的研究结果表明，包含优化启动子是光遗传学向临床转化的关键。
