ALBUMIN CONJUGATED DETOXIFICATION DEPOT FOR BETA AMYLOID PEPTIDES

描述（由申请人提供）：阿尔茨海默病（AD）的标志是大脑中存在斑块，斑块是由2-淀粉样肽中心沉积组成的复杂细胞外病变。遗传学、神经病理学和生物化学证据表明，这些沉积物在阿尔茨海默病的发病机制中起重要作用。我们已经证实，使用我们专有的化学剂可以有效地治疗阿尔茨海默病（AD），该化学剂可以隔离周围的有毒2-淀粉样肽A21-40和A21-42。在AD小鼠模型（appsw -2576）中，免疫组织学分析证明，每两周注射3次我们的肽样化合物可使脑内斑块含量降低50%。与对照组相比，阿尔茨海默病治疗小鼠脑提取物中AB肽的数量减少进一步支持了这一数据。然而，我们还没有解决的问题，去除皮下解毒库。在拟议的研究中，我们将检查一个移动解毒库。在第一阶段，我们将对该新系统进行优化，使其具有强而特异性的毒性A2肽结合和从AD模型小鼠体内清除的特性。综上所述，本研究的目的是：1。开发一种具有足够的亲和力和特异性的“汇”，在低浓度的A2肽和高水平的体内外来蛋白质和脂质中作为结合元件发挥作用。2. 开发一个过程，将导致降解和/或消除有毒的A2肽捕获该汇。白蛋白是否可以作为减少斑块的载体而不需要任何额外的去除步骤？拟议研究的结果预计将对阿尔茨海默病的治疗产生更大的影响，并可能在随后的研究中导致人类应用。公共卫生相关性：现在已知阿尔茨海默病是由大脑中被称为β -淀粉样蛋白（A2）肽的一组特定肽（40或42个氨基酸）的积累引起的。这些A2肽倾向于聚集成不可溶的团块，在大脑中表现为斑块。这些斑块对附近的神经元有毒，从而破坏精神功能。聚集过程是由A2肽内的五肽序列KLVFF诱导的。在我们的早期研究中，我们合成了“逆转录”（RI）形式的A2肽（由反向序列的d -氨基酸组成，ffvlk），作为潜在的治疗剂，旨在干扰聚集过程。RI肽抵抗肽酶消化，我们发现它们保留了天然的KLVFF结合/聚集特性。此外，与RI肽的聚集体被发现可以降低结合A2肽的毒性。我们还观察到，携带多个拷贝的RI肽的缀合物增加了与A2肽相互作用的快速性。我们目前的研究来源于一项已发表的观察，该观察证明了一种新的阿尔茨海默病治疗方法[DeMattos等人（2002）]。脑向血浆- 2淀粉样蛋白外溢：阿尔茨海默病小鼠模型中脑淀粉样蛋白负荷的测量。科学通报，1999,22(2):1 - 4。与其干扰A2肽的合成或聚集过程，还可以隔离（即捕获）A2肽并使其保持非聚集形式，甚至将其从体内移除。这一概念的基本原理是A2肽可以穿过血脑屏障。虽然DeMattos成功地使用了一种针对A2肽的单克隆抗体，但这并不适合作为一种治疗药物，因为它必须在患者的剩余生命中使用。然而，我们的RI肽似乎符合隔离剂的所有要求。RI肽与A2肽特异性且几乎不可逆地结合，它们降低了结合A2肽的毒性，并且它们是稳定的（不可消化的）。通过结合试验筛选一系列RI肽，我们选择了一种主要候选肽，并在AD小鼠模型中进行了测试。6周后，对照组（4只未经治疗的小鼠）出现了广泛的脑损伤，而治疗组（3只小鼠）则没有。显然，RI肽方法可以潜在地导致有效的治疗产品。

DESCRIPTION (provided by applicant): The hallmark of Alzheimer's disease (AD) is the presence in brain of plaques, which are complex extracellular lesions composed of a central deposition of 2-amyloid peptide. Genetic, neuropathological and biochemical evidence have shown that these deposits play an important role in the pathogenesis of AD. We have confirmed the hypothesis that Alzheimer's disease (AD) can be effectively treated using our proprietary chemical agent that can sequester the toxic 2-amyloid peptides A21-40 and A21-42 in the periphery. In a mouse model of AD (APPSWE-2576), 3 biweekly injections of our peptide-like compound resulted in a 50% lower plaque content in the brain, as evidenced by immunohiostological analysis. This data is further supported by a quantitative reduction in the amount of AB peptides in brain extracts from treated AD versus control mice. However, we have not yet addressed the issue of removal of the subcutaneous detoxification depots. In the proposed study, we will examine a mobile detoxification depot. In Phase 1, this new system will be optimized for the properties of strong and specific binding of toxic A2 peptides and clearance from the body of AD model mouse. In summary, the current study aims to 1. Develop a "sink " that has sufficient avidity and specificity to function as a binding element at the low concentration of A2 peptides and at the high levels of extraneous proteins and lipids in the body. 2. Develop a process that will result in degradation and/or elimination of toxic A2 peptides captured by this sink. Can albumin be used as the carrier to provide plaque reduction without any extra removal steps? The outcome of the proposed research is expected to result in a greater impact in the treatment of AD and is likely to lead to human application in subsequent investigations. PUBLIC HEALTH RELEVANCE: Alzheimer's disease is now known to be cause by an accumulation of a particular group of peptides (40 or 42 amino acids), known as beta-amyloid (A2 ) peptides, in the brain. These A2 peptides tend to aggregate into insoluble masses that appear as plaques in the brain. These plaques are toxic to nearby neurons, thereby destroying mental function. The aggregation process is induce by a pentapeptide sequence , KLVFF, within the A2 peptide. In our early studies, we synthesized "retro-inverso" (RI) forms of A2 peptides (composed of D-amino acids in the reverse sequence, ffvlk) as potential therapeutic agents designed to interfere with the aggregation process. RI peptides are resistant to peptidase digestion, and we found that they retain the natural KLVFF binding/aggregation properties. Furthermore, aggregates with RI peptides were found to reduce the toxicity of bound A2 peptides. We also observed that conjugates bearing multiple copies of the RI peptide increase the avidity of interaction with A2 peptides. Our current research is derived from a published observation that demonstrated a novel approach to AD therapy [DeMattos, et al.(2002) Brain to plasma - 2 amyloid efflux: a measure of brain amyloid burden in a mouse model of Alzheimer's disease. Science 295, 2264-2267]. Instead of interfering with the synthesis of A2 peptides or with the aggregation process, it might be possible to sequester (i.e. capture) A2 peptides and keep them in a non-aggregating form or even to remove them from the body. The rationale for this concept is that the A2 peptides can cross the blood-brain barrier. While DeMattos was successful using a monoclonal antibody against A2 peptides, this would not be suitable for a therapeutic agent that would have to be used for the remainder of the patient's life. Our RI peptides however, seemed to fit all the requirements for a sequestering agent. RI peptides bind A2 peptides specifically and virtually irreversibly, they reduce the toxicity of bound A2 peptides and they are stable (non-digestible). Using a binding assay to screen a series of RI peptides, we selected a lead candidate and tested it in a mouse model of AD. After 6 weeks, control (4 untreated mice) had extensive brain damage, whereas treated (3 mice) did not. Clearly, the RI peptide approach can potentially lead to an effective therapeutic product.