EPITOPE-SPECIFIC ANTIBODIES FOR ALZHEIMER'S DISEASE

描述（由申请人提供）：最近用β -淀粉样蛋白疫苗治疗阿尔茨海默病患者的尝试在试验组中导致了大量不良的脑炎反应。在该研究中，全长42个氨基酸的β -淀粉样肽被用作疫苗。考虑到这种分子的大小，它有望呈现出许多潜在的抗原表位，抗体可以针对这些表位产生。据推测，抗体与其中一个或几个表位的相互作用在治疗上是有益的，而与其他位点的结合可能导致不幸的副作用。拟议的研究将确定分子上的哪些位点是最好的治疗目标，哪些是最好的避免，以避免任何不利的副作用。通过表位特异性抗-淀粉样蛋白scFv人抗体进行被动免疫，阿尔茨海默病的免疫治疗将变得更加集中和一致。已经合成了代表β -淀粉样蛋白分子上不同位点的大量β -淀粉样蛋白抗原。这些将用于筛选酵母重组人单链Fv抗体展示库，其中包含超过10亿种不同的抗体。将鉴定出与β -淀粉样蛋白分子上的特定表位紧密结合的独特克隆。通过诱导表达这些选择的抗-淀粉样蛋白抗体的酵母，并通过其his标记纯化分泌的单链Fv。几个精确的结合试验将被用来表征每个不同抗体的特异性和亲和力。功能分析将用于测试不同单链Fv抗体在体外溶解β -淀粉样蛋白聚集体的能力。治疗潜力将使用转基因小鼠模型进行评估。有效的治疗将通过防止脑斑块形成和/或消散脑内预先形成的斑块的能力来评估。最后，这个酵母展示系统将提供一个独特的机会，通过基因工程来改进这些治疗剂，使它们精确地适应手头的任务。例如，可以对单链抗体进行修饰，以获得更高的结合亲和力、更大的功能、二价结合、延长血清半衰期或进入中枢神经系统。

DESCRIPTION (provided by applicant): Recent attempts at treating Alzheimer's patients with a beta-amyloid vaccine have resulted in an adverse brain inflammation response for a significant number in the trial group. The full-length 42 amino-acid beta-amyloid peptide was used as a vaccine in that study. Given the size of that molecule, it is expected to present many potential epitopes against which antibodies can be generated. It is hypothesized that antibody interaction with one or a few of those epitopes would be therapeutically beneficial while binding to other sites could have resulted in the unfortunate side effects. The proposed research will define which sites on the molecule are best to target therapeutically and which are best to avoid so that any untoward side effects are circumvented. The immunotherapy of Alzheimer's disease will become sharply focused and more consistent by using epitope specific anti-beta-amyloid scFv human antibodies for passive immunization. A large panel of beta-amyloid antigens that represent different sites on the beta-amyloid molecule has been synthesized. These will be used to screen a yeast recombinatorial human single chain Fv antibody display library of over one billion different antibodies. Unique clones, which tightly bind to specific epitopes on the beta-amyloid molecule, will be identified. Yeast expressing those select anti-beta-amyloid antibodies will be induced and the secreted single chain Fv purified via its his tag. Several precise binding assays will be used to characterize the specificity and affinity of each distinct antibody. Functional assays will be used to test the ability of the different single chain Fv antibodies to dissolve beta-amyloid aggregates in vitro. Therapeutic potential will be evaluated using a transgenic mouse model. Effective treatment will be assessed by an ability to prevent cerebral plaque formation and/or dissipate preestablished plaques in the brain. Lastly this yeast display system will provide a unique opportunity to refine those therapeutic agents through genetic engineering so they become precisely tuned to the task at hand. For example the single chain antibodies can be modified to achieve higher binding affinities, greater functionality, bivalent binding, a prolonged serum half-life or passage into the CNS.