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特征，也可能是其原因 黄斑变性（AMD）是老年人失明的主要原因。炎症可能是一种 这是年龄相关的氧化应激或毒素在RPE和视网膜内积累的结果。这是一 通过预防由强效细胞因子发出信号的炎症过程来开发AMD基因疗法的项目 如IL-1 β。我们的假设是，通过减少与这些信号分子相关的炎症， 我们可以阻止被诊断为早期疾病的患者的AMD进展。有两种形式的 晚期疾病、湿性（渗出性）AMD和地图状萎缩，这两者都导致中心视力丧失。 虽然存在对湿性AMD的有效治疗，但是没有对地图状萎缩的治疗，地图状萎缩导致 视网膜色素上皮细胞和黄斑感光细胞的死亡。此外，萎缩过程 在湿性AMD中，即使脉络膜新生血管得到控制，也可能持续存在。 这个项目是两个小组之间的合作，这两个小组已经为地理环境产生了不同的小鼠模型。 萎缩其中一种模型通过删除视网膜色素上皮细胞（RPE）中的 一种保护酶锰超氧化物歧化酶的基因。第二个模型删除了 芳香烃受体（AhR），引起与AMD相关的几种途径的眼部失调，包括 胆固醇体内平衡和清除氧化应激诱导毒素，例如在香烟中发现的那些 抽烟两种模型都导致干性AMD的关键特征，包括RPE中基底沉积物的积累， Bruch膜损伤、RPE萎缩和功能障碍以及光感受器死亡。 我们建议在这些模型中使用腺相关病毒（AAV）开发基因治疗， 被证明是安全的基因转移到视网膜。重要的是，这种病毒不会引起严重的 炎症反应。我们在这两种模型中的治疗方法是将分泌型，细胞型， 穿透蛋白，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NLRP 3的活性来抑制炎性细胞因子 炎性小体和阻止caspase-1的活化。我们将验证分泌的抗- 炎性蛋白质向玻璃体的释放产生渗透视网膜和RPE的肽。在决赛中 作为该项目的一部分，我们将使用细胞类型特异性病毒载体， 以确定在我们的小鼠模型中作为炎性细胞因子的主要生产者的细胞。我们的目标是 开发可以在患者身上测试的基因治疗载体，我们相信一次性递送 将治疗性载体转移到玻璃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.