ALBUMIN CONJUGATED DETOXIFICATION DEPOT FOR BETA AMYLOID PEPTIDES

描述(申请人提供)：阿尔茨海默病(AD)的特征是大脑中存在斑块，斑块是由2-淀粉样肽中心沉积组成的复杂的细胞外病变。遗传学、神经病理学和生化证据表明，这些沉积在AD的发病机制中起着重要作用。我们已经证实了这样的假设，即阿尔茨海默病(AD)可以使用我们的专利化学制剂有效地治疗，该制剂可以将有毒的2-淀粉样多肽A21-40和A21-42隔离在外围。在AD小鼠模型(APPswe-2576)中，免疫组织学分析表明，我们的多肽类化合物每两周注射3次，大脑中的斑块含量降低了50%。这一数据进一步得到了阿尔茨海默病治疗小鼠脑提取液中AB多肽数量与对照组相比的减少的支持。然而，我们还没有解决皮下解毒库的移除问题。在拟议的研究中，我们将检查一个流动解毒库。在第一阶段，这一新系统将针对有毒A2多肽的强烈和特异性结合以及从AD模型小鼠体内清除的特性进行优化。综上所述，本研究的目的是：1.开发一种具有足够亲和力和特异性的“汇”，在体内低浓度的A2多肽和高水平的外来蛋白质和脂类中作为结合元件发挥作用。2.开发一个过程，该过程将导致降解和/或消除该水槽捕获的有毒A2多肽。白蛋白可以作为载体来减少斑块，而不需要任何额外的去除步骤吗？这项拟议研究的结果预计将对AD的治疗产生更大的影响，并可能导致在随后的研究中应用于人类。与公共卫生相关：阿尔茨海默病现在已知是由一组特定的多肽(40或42个氨基酸)在大脑中积累引起的，这些多肽被称为β-淀粉样多肽(A2)。这些A2多肽往往聚集成不溶的团块，在大脑中以斑块的形式出现。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肽中的五肽序列KLVFF诱导的。在我们的早期研究中，我们合成了“逆转录”(Retro-Inverso，RI)形式的A2多肽(由反向序列中的D-氨基酸组成，ffvlk)，作为潜在的治疗剂，旨在干扰聚集过程。RI多肽对肽酶消化有抵抗力，我们发现它们保留了KLVFF天然的结合/聚集特性。此外，还发现与RI多肽形成的聚集体可以降低结合的A2多肽的毒性。我们还观察到，携带多个RI多肽拷贝的结合物增加了与A2多肽的亲和力。我们目前的研究源于一项已发表的观察，该观察展示了一种治疗AD的新方法[DeMattos，等人(2002)]，脑到血浆-2淀粉样蛋白外流：一种测量阿尔茨海默病小鼠模型脑淀粉样蛋白负荷的方法。科学295,2264-2267]。不是干扰A2多肽的合成或聚集过程，而是有可能隔离(即捕获)A2多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多肽可以穿越血脑屏障。虽然DeMattos成功地使用了抗A2多肽的单抗，但这不适用于必须在患者的余生中使用的治疗剂。然而，我们的RI多肽似乎符合隔离剂的所有要求。RI多肽与A2多肽的结合是特异的，几乎是不可逆转的，它们降低了结合的A2多肽的毒性，而且它们是稳定的(不可消化)。使用结合实验筛选一系列RI多肽，我们选择了一个候选的先导，并在AD小鼠模型上进行了测试。6周后，对照组(4只未治疗的小鼠)出现广泛的脑损伤，而治疗(3只)的小鼠则没有。显然，RI多肽方法可能导致一种有效的治疗产品。

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.