Prolonged Inhibition of Pathologic Neovascularization by Catalytic Antioxidants

描述（由申请人提供）：在某些形式的糖尿病视网膜病变（DR）和黄斑变性（AMD）中，失明是由于新血管的病理发展造成的，这些血管是不完整的、脆弱的和多孔的。这些新生血管疾病的进展被认为是通过产生毒性分子活性氧（ROS）而发生的。对于这些疾病没有长期成功的疗法，这些疾病对患者具有破坏性影响，并且每年花费美国超过500亿美元。我们的长期目标是开发一种治疗方法，以保护视网膜细胞的健康和功能，从而延长DR或AMD患者的视力并改善其生活质量。由于ROS的过度升高发生在大多数其他视网膜病变的“上游”，因此它代表了可以被抗氧化剂和增加“II期”抗氧化酶表达的其他分子靶向的共同节点。我们发表的和初步的数据表明，氧化铈纳米颗粒，催化破坏活性氧，可以防止病理性脉络膜和视网膜新生血管病变的发展，并导致现有的病理性新生血管在非常低密度脂蛋白受体无效的视网膜通过调节许多视网膜基因，包括血管内皮生长因子（VEGF）的表达消退。我们的中心假设是，氧化铈纳米颗粒，因为它们的催化抗氧化活性和长期保留在视网膜中，将持续清除ROS和抑制病理性新血管形成的时间延长-长达12个月。具体目标1将确定纳米氧化铈在视网膜中的持续时间以及它们保留抗新血管形成活性的程度。电感耦合等离子体质谱法将定量铈在十亿分之几的水平。眼底镜检查、视网膜电图和光学相干断层扫描将用于同一动物的纵向研究，以评价新生血管形成、视网膜功能和外核层厚度。纳米氧化铈对参与氧化应激、炎症和新生血管形成的特定基因的影响将使用共聚焦显微镜、蛋白质印迹和PCR阵列进行分析。具体目标2将证明纳米氧化铈通过减少氧化应激对光感受器和视网膜色素上皮（RPE）细胞的影响来保护视网膜。将评价指示光感受器-和/或RPE-氧化应激的基因活性、蛋白质和结构。具体目标3将检验以下假设：纳米氧化铈和萝卜硫素（II相抗氧化酶的诱导剂）的组合使用将在Vldlr视网膜中产生累加或协同效应。预期成果-所提出的工作预计将证明纳米氧化铈抑制视网膜中病理性新生血管形成的寿命、效力和机制。预期结果将具有重要的积极影响，因为纳米氧化铈的长期有效性的证明将最有可能支持其治疗用途，并且所鉴定的基因的活性的变化将提供对于治疗DR、AMD和涉及氧化应激的其他疾病重要的额外靶标。

DESCRIPTION (provided by applicant): In some forms of Diabetic Retinopathy (DR) and Macular Degeneration (AMD), blindness results from the pathologic development of new blood vessels which are incomplete, weak and porous. The progression of these neovascular diseases is thought to occur through the production of toxic molecules, Reactive Oxygen Species (ROS). There are no long term successful therapies for such diseases which have devastating effects on patients and cost the USA over $50 billion/yr. Our long term goal is to develop a therapeutic treatment to protect the health and function of retinal cells and thereby prolong vision and improve the quality of life for patients with DR or AMD. Because the excessive rise in ROS occurs "upstream" of most other retinal pathologies, it represents a common node which can be targeted by antioxidants and other molecules which increase the expression of "Phase II" antioxidant enzymes. Our published and preliminary data show that cerium oxide nanoparticles, which catalytically destroy ROS, can prevent development of pathologic choroidal and retinal neovascular lesions and cause the regression of existing pathologic neovessels in the Very Low Density Lipoprotein Receptor null retina by modulating the expression of many retinal genes including Vascular Endothelial Growth Factor (VEGF). Our central hypothesis is that cerium oxide nanoparticles, because of their catalytic antioxidant activity and long term retention in the retina, will continuously scavenge ROS and inhibit pathologic neovascularization over prolonged times -up to 12 months. Specific aim 1 will determine duration of nanoceria in the retina and the extent to which they retain activity against neovascularization. Inductively coupled plasma mass spectrometry will quantitate cerium at the parts per billion levels. Fundoscopy, electroretinography and optical coherence tomography will be used for longitudinal studies on the same animal to evaluate neovascularization, retinal function and thickness of the outer nuclear layer. Nanoceria effects on specific genes involved in oxidative stress, inflammation and neovascularization will be analyzed using confocal microscopy, Western blots and PCR arrays. Specific Aim 2 will demonstrate that nanoceria provide protection to the retina by reducing the effects of oxidative stress on photoreceptors and Retinal Pigment Epithelial (RPE) cells. Gene activity, proteins and structures indicative of photoreceptor- and/or RPE- oxidative stress will be evaluated. Specific Aim 3 will test the hypothesis that the combinatorial use of nanoceria and sulforaphane, an inducer of Phase II antioxidant enzymes, will result in an additive or synergistic effects in the Vldlr retina. Expected outcomes - the work proposed is expected to demonstrate the longevity, potency and mechanisms by which nanoceria inhibit pathologic neovascularization in the retina. The results are expected to have an important positive impact because the demonstration of the long term effectiveness of the nanoceria will most likely support their therapeutic use and changes in activity of identified genes should provide additional targets important for treating DR, AMD and other diseases which involve oxidative stress.