Genetic therapy for EYS- and USH2A-associated retinal dystrophy

遗传性视网膜疾病(IRD)是一组患病率约为每3000人中有1人的疾病，约占欧盟内20万名患者。由于视觉系统的复杂性，数百种蛋白质在视网膜中唯一表达，200多个不同基因的突变与IRD有关。在过去的十年里，眼睛的特殊优势使这个器官处于基因治疗开发的前沿，包括使用病毒载体进行基因增强治疗，以转移突变基因的正确cdna拷贝。最近临床试验的成功证明了基于基因的疗法在IRD治疗中的实用性，整个欧洲的患者现在正在等待针对每一种IRD基因亚型的基因特异性方法。然而，对于大基因突变的患者，包括IRD中最常见的两个基因突变，即EYS和USH2A，由于腺相关病毒(AAV)载体的载量有限，临床翻译受到阻碍，AAV载体是目前视网膜基因治疗的首选载体。我们建议的目的是通过两种方式解决这一限制。首先，我们将开发替代的治疗方法，其中包括调节剪接机制，以及使用最先进的方法实施治疗性基因组编辑方法，如基于AON的剪接调制、Prime编辑和新发现的CaS蛋白质变体。其次，我们将优化能够确保EYS和USH2A编码序列正确输送到视网膜的新型大型载体，包括基于内含素的多重AAV方法、新发现的腺病毒载体和最新版本的纳米颗粒。为了尽可能接近患者群体，我们将主要使用人类细胞模型系统，即成熟的细胞系和诱导的基于多能干细胞的视网膜器官。在有需要和有充分理据的情况下，我们亦会利用斑马鱼和猪作为两个与疾病病理学有关的体内系统。由于准备联盟的合作伙伴处于其职业生涯的不同阶段，彼此之间的密切互动可能会使新想法的建立成为可能，并导致颠覆性的科学发展。与许多伙伴国家的患者倡导组织的密切联系将确保传播结果，以提高患有EYS和USH2A突变的患者对这些进展的认识。Get-Ready的成功结果将极大地推动为不可忽视的患者群体开发新的治疗方法，从而有助于提高这些患者的生活质量，并减轻由于进行性视力丧失而造成的社会经济负担。

Inherited retinal diseases (IRDs) are a group of disorders with a prevalence of approximately 1 in 3,000 people, accounting for about 200,000 patients within the European Union. Due to the complexity of the visual system, several hundred proteins are uniquely expressed in the retina, and mutations in over 200 different genes are associated with IRD. Over the last decade, specific advantages of the eye have positioned this organ at the forefront of gene therapeutic development, including gene augmentation therapy using viral vectors for the transfer of correct cDNA copies of mutated genes. Recent success in clinical trials has demonstrated the utility of gene-based therapies in the treatment of IRDs, and patients throughout Europe are now awaiting gene-specific approaches for each genetic subtype of IRD. However, for patients with mutations in large genes, including two genes most frequently mutated in IRD, i.e. EYS and USH2A, clinical translation is hampered due to the limited cargo capacity of adeno-associated virus (AAV) vectors, the current vector of choice in retinal gene therapy. The aim of our proposal is to address this limitation in two ways. First, we will develop alternative therapeutic approaches, which includes the modulation of the splicing machinery and the implementation of therapeutic genome editing approaches using state-of-the-art methods like AON-based splicing modulation, PRIME-editing and newly identified Cas protein variants. Second, we will optimize novel larger vehicles that can ensure a proper delivery of EYS and USH2A coding sequences to the retina, including an intein-based multiple AAV approach, newly identified adenoviral vectors and latest versions of nanoparticles. To remain as close as possible to the patient population, we will mostly employ human cellular model systems, i.e. well-established cell lines and induced pluripotent stem cell-based retinal organoids. Where necessary and justified, we will also employ the zebrafish and the pig as two in vivo systems relevant to disease pathology. Since the partners in the GET READY consortium are at different stages of their career, the close interaction with each other will likely enable the establishment of new ideas and lead to disruptive scientific developments. A close connection to patient advocacy organizations in many partner countries will ensure dissemination of the results to raise awareness of these developments for patients with EYS and USH2A mutations. A successful outcome of GET-READY will significantly advance the development of novel treatment approaches for a non-negligible patient population, and consequently help to improve the life-quality of these patients as well as reduce the socio-economic burden due to progressive vision loss.