Novel Adeno-Associated Viral Therapy for X-linked Retinitis Pigmentosa

描述(申请人提供)：这个项目的目标是开发一种治疗视网膜色素变性(RP)的基因疗法。RP是最常见的遗传性失明形式，以光感受器丧失为特征，导致外周视力和夜视的进行性丧失。RP是一组疾病，由导致相似表型的多个基因中的一个突变引起，目前还没有治疗方法。X-连锁视网膜色素变性(XLRP)是视网膜色素变性(RP)的一种类型，最常见的原因是光感受器蛋白RPGR(视网膜色素变性GTP酶调节因子)基因的突变，是相对常见的一种，约占所有RP病例的10%。RPGR的大多数突变是隐性的，这表明基因替换可能是一种有效的策略。重组腺相关病毒(AAV)载体在视网膜疾病动物模型中的基因传递方面显示出巨大的前景，可以在没有炎症的情况下长期、稳定地表达基因。包括AAV8在内的以视网膜色素上皮(RPE)或光感受器层为靶点的新型AAV血清型已被鉴定，并以比已知的AAV血清型更高的效率转导这些细胞类型。本项目建议利用AAV8这种擅长向光感受器运送基因的药物来开发一种治疗XLRP的药物。最初，将进行优化研究，以确定RPGR基因替换的最有效临床候选，然后将对建议的临床候选进行安全性和毒理学研究，目标是在项目期结束前提交IND。首先，将在RPGR基因敲除小鼠XLRP的小鼠模型中测试RPGR基因的几种替代亚型，以确定哪种亚型在防止视网膜退化方面最有效。当选择最佳的cdna后，启动子将被优化，同时测试构成(CMV)启动子和光受体特异性(视紫红质)启动子，以确定其中一个启动子是否比另一个启动子具有疗效优势。这些启动子也将与正常小鼠和非人类灵长类动物的报告基因一起使用，以确认两者对长期转基因表达都是稳定的。最终的载体构建也将用于确定在RPGR基因敲除小鼠中产生生物效应所需的最小载体剂量。最后，具有AAV8血清型、优化的启动子和RPGR转基因的临床候选将在小鼠和狗身上进行生物分布和安全性/毒理学研究，以完成提交IND所需的一揽子数据。建议的XLRP基因疗法将服务于RP患者未得到满足的医疗需求，并将是一种新的“一流”疗法，将为治疗其他形式的RP和其他视网膜疾病提供一个平台。 与公共卫生相关：视网膜色素变性是最常见的遗传性失明形式，据估计，美国有10万名患者。它的特点是外周视力和夜间视力逐渐丧失，最终导致失明。这项提议旨在开发一种基因疗法，将为患者提供显著的医疗缓解，开发适用于眼睛其他适应症的技术，并将医疗系统的负担降至最低。

DESCRIPTION (provided by applicant): The goal of this project is to develop a gene therapeutic for retinitis pigmentosa (RP). RP is the most common inherited form of blindness, and is characterized by photoreceptor loss, leading to a progressive loss of peripheral as well as night vision. There is no treatment for RP, which is a group of diseases caused by mutations in one of multiple genes that lead to similar phenotypes. X-linked retinitis pigmentosa (XLRP) is a form of RP that is most often caused by mutations in a gene encoding the photoreceptor protein RPGR (retinitis pigmentosa GTPase regulator), and is relatively common, accounting for approximately 10% of all cases of RP. Most mutations in RPGR are recessive, indicating that gene replacement can be an effective strategy. Recombinant adeno-associated virus (AAV) vectors have been showing great promise in delivering genes in animal models of retinal disease, giving long-term, stable gene expression without inflammation. Novel AAV serotypes including AAV8 have been identified which target the retinal pigment epithelium (RPE) or the photoreceptor layer, and transduce those cell types with higher efficiency than previously known serotypes of AAV. This project proposes to use AAV8, which excels at delivering genes to photoreceptors, to develop a therapeutic for XLRP. Initially, optimization studies will be performed to identify the most effective clinical candidate for RPGR gene replacement, and then safety and toxicology studies will be performed with the proposed clinical candidate, with the goal of filing an IND by the end of the project period. Initially, several alternative isoforms of the RPGR cDNA will be tested in the mouse model of XLRP, the RPGR knockout mouse, to determine which is the most effective in preventing retinal degeneration. When the optimal cDNA is selected, the promoter will then be optimized, testing both a constitutive (CMV) promoter and a photoreceptor- specific (rhodopsin) promoter to determine if one promoter has an efficacy advantage over the other. The promoters will also be used with a reporter gene in normal mice and non-human primates to confirm that both are stable for long-term transgene expression. The final vector construct will also be used to determine the minimal dose of vector required for biologic effect in the RPGR knockout mouse. Finally, the clinical candidate, with the AAV8 serotype, and optimized promoter and RPGR transgene will be put through biodistribution and safety/toxicology studies in mice and in dogs to complete the package of data needed for filing an IND. The proposed gene therapeutic for XLRP will serve an unmet medical need in RP patients, and will be a novel "first-in-class" therapy that will provide a platform for treating additional forms of RP and other retinal diseases. PUBLIC HEALTH RELEVANCE: Retinitis pigmentosa is the most common inherited form of blindness, with an estimated 100,000 patients in the US. It is characterized by progressive loss of peripheral as well as night vision, eventually resulting in blindness. This proposal is designed to develop a gene therapeutic that will provide significant medical relief to patients, develop technologies applicable to other indications in the eye, and minimize the burden on the healthcare system.