Testing Gene Therapy in Models of Geographic Atrophy

慢性炎症是与年龄相关的视网膜损伤的一个特征，也可能是一个原因。 相关性黄斑变性(AMD)是老年人失明的主要原因。炎症可能是一种 与年龄相关的氧化应激或RPE和视网膜内毒素积聚的后果。这是一个 通过阻止由强有力的细胞因子发出信号的炎症过程来开发AMD基因治疗的项目 如IL-1β。我们的假设是，通过减少与这些信号分子相关的炎症， 我们可以在被诊断为早期疾病的患者中阻止AMD的进展。有两种形式的 晚期疾病、湿性(渗出性)AMD和地理性萎缩，这两种疾病都会导致中心视力丧失。 虽然湿性AMD有有效的治疗方法，但没有治疗地理萎缩的方法，这会导致 黄斑内视网膜色素上皮和感光细胞死亡。此外，萎缩的过程 即使脉络膜新生血管得到控制，湿性AMD也可能持续存在。 这个项目是两个小组之间的合作，他们已经为地理环境生成了不同的鼠标模型 萎缩。其中一个模型通过删除视网膜色素上皮(RPE)中的 一种保护酶的基因，锰超氧化物歧化酶。第二个模型删除了 芳香烃受体(AhR)，导致与AMD相关的几个通路的眼部调节失调，包括 胆固醇稳态和氧化应激诱导毒素的清除，如香烟中发现的那些 烟。这两种模型都导致了干性AMD的关键特征，包括在RPE中积累基底物， Bruchs膜受损，RPE萎缩，光感受器功能障碍和死亡。 我们建议使用腺相关病毒(AAV)来开发这些模型的基因治疗，这种病毒已经被 证明基因转移到视网膜是安全的。重要的是，这种病毒不会引发严重的 炎症反应。我们在这两个模型中的治疗方法是为分泌的细胞输送基因 穿透蛋白质，增加对氧化应激的保护，或减少 通过刺激抗氧化酶的合成或阻断NLRP3的活性来产生炎症细胞因子 炎症小体，阻止caspase-1活化。我们会确认秘密交付的抗- 玻璃体中的炎性蛋白产生的多肽渗透到视网膜和RPE中。在决赛中 作为该项目的组成部分，我们将使用细胞类型的特定病毒载体在 以确定在我们的小鼠模型中主要产生炎性细胞因子的细胞。我们的目标是 开发可以在患者身上进行测试的基因治疗载体，我们相信一次性交付的 玻璃体的治疗载体将可转化为人类基因治疗。

Chronic inflammation is a characteristic, and possibly a cause, of the retinal damage associated with age related macular degeneration (AMD), a leading cause of blindness among the elderly. Inflammation may be a consequence of age related oxidative stress or accumulation of toxins within the RPE and retina. This is a project to develop gene therapy for AMD by preventing inflammatory processes signaled by potent cytokines such as IL-1β. It is our hypothesis that, by reducing inflammation associated with these signaling molecules, we can halt the progression of AMD in patients diagnosed with early stage disease. There are two forms of the advanced disease, wet (exudative) AMD and geographic atrophy, both of which lead to loss of central vision. While there are effective treatments for wet AMD, there is no treatment for geographic atrophy, which leads to death of the retinal pigment epithelium and photoreceptor cells in the macula. Furthermore, atrophic processes may persist in wet AMD, even when choroidal neovascularization is under control. This project is a collaboration between two groups that have generated distinct mouse models for geographic atrophy. One of the models increases oxidative stress in the retinal pigment epithelium (RPE) by deleting the gene for a protective enzyme, manganese superoxide dismutase. The second model deletes the gene for the aryl hydrocarbon receptor (AhR), causing ocular dysregulation of several pathways relevant to AMD including cholesterol homeostasis and clearance of oxidative stress-inducing toxins such as those found in cigarette smoke. Both models result in key features of dry AMD including accumulation of basal deposits in the RPE, damage to Bruch's membrane, RPE atrophy and dysfunction and death of photoreceptors. We propose to develop gene therapy in these models using adeno-associated virus (AAV) which has been demonstrated to be safe for gene transfer to the retina. Importantly, this virus does not provoke a severe inflammatory response. Our approach to therapy in the two models is to deliver genes for secreted, cell penetrating proteins that increase protection from oxidative stress or that reduce the production of inflammatory cytokines by stimulating synthesis of anti-oxidant enzymes or blocking the activity of the NLRP3 inflammasome and preventing the activation of caspase-1. We will validate that delivery of the secreted anti- inflammatory proteins to the vitreous produces peptides that permeate the retina and RPE. In the final component of the project, we will use cell-type specific viral vectors that produce non-secreted peptides in order to identify the cells that are major producers of inflammatory cytokines in our mouse models. Our goal is to develop gene therapy vectors that could be tested in patients, and we believe that one-time delivery of a therapeutic vector to the vitreous will be translatable to human gene therapy.