Development of a genetic CNV model for Age-Related Macular Degeneration

描述（由申请人提供）：本提案的目的是开发一种用于治疗黄斑变性（AMD）的化合物的高通量药物筛选服务。在我们的初步研究中，我们已经确定了一种基因修饰的小鼠（Vldlr-/-），其表现出与人类AMD发病机制几乎相同的症状。基于以下观察结果，我们假设Vldlr-/-小鼠是AMD的良好模型1）VLDLR变体最近已显示与人类患者中的AMD相关，表明该模型的发病机制与人类AMD相关。2)该小鼠模型发展了人类AMD的大多数表型，包括视网膜炎症、光感受器变性、视网膜血管渗漏和CNV。3)与激光诱导的新生血管模型相比，Vldlr-/-小鼠是一种遗传模型，CNV具有高度可重复性，易于定量。CNV可以通过计数在RPE/脉络膜平坦安装件的表面上穿透RPE的血管而容易地定量。4)VEGF的过度表达是该模型的主要致病因素，与人类AMD相似，适合于筛选新的抗VEGF药物。4)与其他几种现有的AMD转基因模型相比，该模型发展了早发性AMD，这使其成为筛选新的抗CNV药物的合适模型。5)Vldlr-/-小鼠易于繁殖，不需要额外的基因分型，因此使用该模型进行药物筛选的成本较低。已知视网膜血管形成由血管内皮生长因子（VEGF）蛋白介导，并且目前FDA批准的AMD治疗是该蛋白及其信号传导能力的抑制剂。这些药物是治疗AMD的一大进步，但它们1）未能解决疾病的炎症性质; 2）对所有AMD患者无效; 3）需要每月注射到眼睛中以递送药物; 4）只能在已经发生视力丧失时使用。因此，仍然需要更有效的疗法来治疗AMD，并且许多制药公司正在大力开发新的AMD疗法。为此，需要合适的动物模型来快速筛选和开发可用于治疗AMD的化合物。我们提出的研究的目标是建立在AMD药物筛选中使用Vldlr-/-小鼠的理由。我们计划在Vldlr-/-小鼠中使用几种已建立的抗VEGF化合物来证明与可比化合物在预防AMD表型方面的相似功效。在该I期项目中，我们将在该小鼠模型中测试这些抗VEGF化合物对视网膜血管渗漏和脉络膜新生血管形成的功效。如果这些VEGF抑制剂成功地减轻了在Vldlr-/-小鼠中观察到的AMD症状，这将提供这些小鼠是AMD的优良模型的实质性证据。我们公司积极参与提供合同服务，以筛选眼科药物化合物在我们专有的糖尿病动物模型中的疗效，上述研究的完成将使我们能够为AMD药物筛选提供类似的服务。公共卫生相关性：视网膜相关性黄斑变性（AMD）是全球50岁以上患者失明的主要原因。治疗AMD的疗法的开发在很大程度上依赖于动物模型的临床前测试;然而，不存在用于AMD药物筛选的令人满意的动物模型。我们已经确定了AMD的小鼠模型，本提案的目标是证明这些小鼠在AMD药物筛选中的利用，以便我们可以开发高通量合同服务，用于评估各公司开发的药物的疗效。

DESCRIPTION (provided by applicant): The objective of this proposal is to develop a service for the high-throughput drug screening of compounds to treat Age-Related Macular Degeneration (AMD). In our preliminary studies, we have identified a genetically-modified mouse (Vldlr-/-) that presents with symptoms nearly identical to AMD pathogenesis in humans. Based on the following observations, we hypothesize that Vldlr-/- mice is a good model for AMD 1) VLDLR variants have been recently shown to associate with AMD in human patients, suggesting that the pathogenesis of this model is relevant to human AMD. 2) This mouse model develops most phenotypes of human AMD, including retinal inflammation, photoreceptor degeneration, retinal vascular leakage and CNV. 3) Compared to the laser-induced neovascularization models, Vldlr-/- mice are a genetic model and the CNV is highly reproducible and easy to quantify. The CNV can be readily quantified by counting vessels penetrating the RPE on the surface of the RPE/choroid flat mount. 4) VEGF over-expression is a major causative factor in this model, similar to that in human AMD and suitable for the screening of new anti-VEGF drugs. 4) Compared to several other existing transgenic models of AMD, this model develops early onset AMD, which makes it a suitable model for screening new anti-CNV drugs. 5) Vldlr-/- mice are easy to breed and do not need additional genotyping, so that the costs of drug screening using this model is low. Retinal blood vessel formation is known to be mediated by the vascular endothelial growth factor (VEGF) protein, and current FDA approved treatments for AMD are inhibitors of this protein and its ability to signal. These drugs are a great advancement in the treatment of AMD, but they 1) fail to address the inflammatory nature of the disease; 2) are not effective in all AMD patients; 3) require monthly injections into the eye to deliver the drug; 4) can only be used once visual loss has already occurred. Therefore, more efficacious therapies are still needed to treat AMD, and numerous pharmaceutical companies are heavily engaged in the development of new AMD-therapeutics. Towards this end, suitable animal models are needed to rapidly screen and develop compounds that could be useful in treating AMD. The goal of our proposed research is to establish justification for the utilization of Vldlr-/- mice in AMD-drug screening. We plan to use several established anti-VEGF compounds to demonstrate a similar efficacy in preventing AMD-phenotypes with comparable compounds in Vldlr-/- mice. In this Phase I project, we will test the efficacy of these anti-VEGF compounds on retinal vascular leakage and choroidal neovascularization in this mouse model. If these VEGF-inhibitors are successful in alleviating the AMD-symptoms observed in Vldlr-/- mice, this will provide substantial evidence that these mice are an excellent model of AMD. Our company is actively engaged in providing contractual services to screen the efficacy of ophthalmic pharmaceutical compounds in our proprietary animal models of diabetes, and the completion of the studies described above would enable us to offer similar services for AMD-drug screening. PUBLIC HEALTH RELEVANCE: Age-Related Macular Degeneration (AMD) is the leading cause of blindness worldwide in patients over the age of 50. The development of therapies to treat AMD is heavily reliant on preclinical testing in animal models; however, no satisfactory animal models exist for AMD-drug screening. We have identified a mouse model for AMD, and the goal of this proposal is to justify the utilization of these mice in AMD-drug screening so that we may develop a high-throughput contract service for evaluating the efficacy of pharmaceuticals in development by various companies.