Therapeutic Effect of a Novel Antioxidant on Degenerative Eye Disorders

描述（由申请方提供）：拟定研究将在相关动物模型中测试新型硫醇抗氧化剂N-乙酰半胱氨酸酰胺（NACA）滴眼液制剂在预防和治疗白内障和视网膜变性方面的有效性。本申请的广泛、长期目标是开发用于治疗年龄相关性眼病（ARED）的NACA滴眼液制剂，并为进一步探索治疗各种年龄相关性疾病的药理学抗氧化剂方法提供动力。氧化应激损伤在ARED发病机制中的重要性已经在白内障和干性年龄相关性黄斑变性（AMD）中得到证实。透镜和黄斑都受到显著的光氧化应激，而黄斑是代谢活动速率高的部位，并且存在多不饱和脂肪的浓度。透镜和视网膜细胞通过产生足够的抗氧化酶或小分子量抗氧化剂如谷胱甘肽（GSH）来对抗氧化应激。他们产生这种抗氧化剂的能力随着年龄的增长而下降，导致氧化损伤增加。在透镜中，结果是结晶蛋白的溶解度逐渐降低，这表现为透镜的浑浊化。在视网膜中，它导致线粒体损伤和玻璃疣的积累，这是干性AMD的特征。抗氧化剂如N-乙酰半胱氨酸（NAC）已显示出改善氧化应激损伤的前景。我们的数据表明，NAC的稳定药物类似物NACA在降低氧化应激方面比NAC好得多。初步的动物研究表明，腹膜内NACA减少白内障形成，而NACA应用于视网膜上皮细胞防止细胞活力丧失。建议的具体目标 研究是评估1）接受亚硒酸盐注射的雄性Wistar大鼠幼仔中的白内障形成和2）在早期（rd 10 +/+）或晚期（Ccl 2-/-）自发发展进行性感光细胞死亡的rd 10 +/+和Ccl 2-/-小鼠中的视网膜变性。我们还将使用化学诱导的模型，其中将碘酸钠注射到C57/BL 6小鼠中以诱导RPE变性。NACA的保护作用将在动物模型中通过对白内障进行分期并测量各个实验组中的不透明度指数来评估，而在rd 10 +/+和Ccl 2-/-小鼠中，NACA的保护作用将通过基于外核层（ONL）密度和视杆和视锥视网膜电图（ERG）的测量来测量视觉电位和感光器功能来评估。在所有动物模型中，我们将通过去除靶组织（透镜或视网膜）并测量许多氧化应激参数、抗氧化酶的活性以及促凋亡和凋亡蛋白（如细胞色素c和半胱天冬酶）的表达来评估NACA的抗氧化作用。我们还将研究晶体蛋白的蛋白质氧化，脂质过氧化，以及蛋白质结合的GSH和半胱氨酸。 公共卫生相关性：在美国，与年龄相关的眼病导致的视力丧失现在影响着超过3000万人，预计在未来几十年内，这一数字将翻一番。白内障和老年性黄斑变性等疾病严重影响生活质量，并造成相当大的医疗费用。美国每年用于白内障手术的总费用超过90亿美元，而与这种视力问题相关的所有服务的总费用超过200亿美元。一些实验表明，一种称为N-乙酰半胱氨酸酰胺（NACA）的抗氧化剂可能会延迟这些类型疾病的发作或停止其进展。这项研究将测试眼药水形式的NACA是否可以在动物模型中预防和治疗白内障和黄斑变性等眼部疾病。这项研究的成功结果将支持这种药物进入人类使用的进展。NACA滴眼液将成为昂贵的白内障手术的替代方案，降低与年龄相关眼病相关的医疗保健成本，并大大改善受这些疾病影响的人的生活质量。

DESCRIPTION (provided by applicant): The proposed research will test the effectiveness of an eye drop formulation of a new thiol antioxidant, N- acetylcysteine amide (NACA), in preventing and treating cataracts and retinal degeneration in relevant animal models. This application's broad, long-term objectives are to develop an eye drop formulation of NACA for treatment of age-related eye diseases (AREDs) and to provide impetus for greater exploration of pharmacologic antioxidant approaches to the treatment of a variety of age-related diseases. The importance of oxidative stress damage in the pathogenesis of AREDs has been established for cataracts and the dry form of age-related macular degeneration (AMD). The lens and macula are both subjected to significant photo-oxidative stress, while the macula is the site of high rate of metabolic activity and the presence of concentrations of polyunsaturated fats. Lens and retinal cells combat oxidative stress by generating sufficient antioxidant enzymes or small molecular weight antioxidants like glutathione (GSH). Their ability to produce such antioxidants decreases with age, leading to increased oxidative damage. In the lens, the result is a progressive reduction in solubility of crystalline proteins that manifests in an opacification of te lens. In the retina, it results in mitochondrial damage and the accumulation of drusen that characterizes dry AMD. Antioxidants like N- acetyl cysteine (NAC) have shown promise in ameliorating oxidative stress damage. Our data indicates that a stable pharmaceutical analogue of NAC, NACA works much better than NAC in reducing oxidative stress. Preliminary animal studies have shown that intraperitoneal NACA reduces cataract formation while application of NACA to retinal epithelial cells prevents loss of cell viability. The Specific Aims of the proposed research are to evaluate 1) cataract formation in male Wistar rat pups receiving selenite injections and 2) retinal degeneration in rd10+/+ and Ccl2-/- mice that spontaneously develop progressive photoreceptor cell death either early (rd10+/+) or later (Ccl2-/-). We will also use a chemically-induced model, in which sodium iodate will be injected into C57/BL6 mice to induce RPE degeneration. The protective effects of NACA will be assessed in animal models by staging cataracts and measuring opacity indices in various experimental groups, while in the rd10+/+ and Ccl2-/- mice the protective effects of NACA will be assessed by measuring the visual potential and photoreceptor function based on measurements of outer nuclear layer (ONL) density and rod and cone electroretinograms (ERGs). In all animal models, we will assess the antioxidant effects of NACA by removing the target tissue (lens or retina) and measuring a number of oxidative stress parameters, the activities of antioxidant enzymes, and the expression of pro-apoptotic and apoptotic proteins such as cytochrome c and caspases. We will also study protein oxidation of crystallins, lipid peroxidation, and protein bound GSH and cysteine. PUBLIC HEALTH RELEVANCE: The loss of vision from age-related eye diseases now affects over 30 million people in the United States-a number that is expected to double in the coming decades. Diseases such as cataracts and age-related macular degeneration significantly affect quality of life and represent sizable medical costs. The annual total outlay for cataract surgery each year in the U.S. is over $9 billion while the total costs of all services related to such visin problems is over $20 billion. Several experiments show that an antioxidant called N-acetylcysteine amide (NACA) may delay the onset or halt the progression of these types of diseases. This research will test whether NACA in eye drop form can prevent and treat eye diseases such as cataracts and macular degeneration in animal models. Successful results from this study will support the advance of this medication into human use. NACA eye drops would represent an alternative to costly cataract surgery, reduce health care costs related to age-related eye diseases and greatly improve the quality of life of people affected by these diseases.