WGA as a Novel Vehicle for Intranasal Delivery of an Anti-A-beta Antibody in AD

本申请是对PAS-10-151(阿尔茨海默病药物发现补助金(R21))的响应。 需要新的载体来有效地将免疫疗法直接输送到大脑，以便 最大限度地发挥被动免疫治疗、延迟或预防阿尔茨海默病(AD)的效果，a 目前困扰着500万美国人的疾病。凝集素、小麦胚芽凝集素(WGA)的结合， 已被证明可以增强候选药物的鼻腔吸收，并通过(I)促进直接给药到大脑 吸附或受体介导的内吞作用进入嗅觉神经元(OSN)，然后是细胞内 转运至嗅球；(Ii)非特异性液相内吞进入嗅神经细胞，然后进入细胞内 传输到嗅球；和/或(Iii)沿着嗅觉上皮裂隙的细胞外扩散直接到 嗅球和/或脑脊液。静脉注射Ass单抗的被动免疫 (MAb)已显示出有希望的结果，如降低大脑ASS和稳定或防止认知 阿尔茨海默病转基因小鼠模型和早期人类临床试验的临床前研究存在缺陷。然而， 抗体在大脑中的渗透水平受到血脑屏障(BBB)的限制。此外，在小鼠和 患有血管淀粉样蛋白的人，血液中高水平的抗Ass抗体导致微出血和 血管源性水肿。该项目旨在通过最大限度地发挥鼻腔的功效来克服这些限制。 使用WGA载体蛋白的抗Ass抗体传递途径。该项目将测试WGA作为一种 一种新型载体，不仅将增强与单/二聚体和纤维ASs结合的抗Ass单抗3A1的输送 (由Lemere-Consulting博士提供)由于其内吞摄取偏好，但也将促进 3A1因其鼻黏膜样生化成分而被动扩散，最大限度地发挥免疫治疗作用 鼻腔被动免疫的效果，将在阿尔茨海默病5XFAD模型中进行测试。假设：鼻腔 WGA偶联的3A1单抗与未标记的3A1抗体相结合将使 鼻腔被动免疫改善AD样病理认知改变的免疫治疗潜力 在5XFAD转基因小鼠中。具体目标：[1]WGA-3A1抗体鼻腔给药后的贩运 野生型小鼠脑内3A1-WGA与5XFAD小鼠和人类AD小鼠脑内Ass斑块的结合[2] WGA偶联3A1单抗与未标记3A1联合鼻腔被动免疫的研究 抗体将防止幼年斑块前5XFAD小鼠的AD样病理认知改变。[3]确定是否 WGA标记的3A1单抗与未标记的3A1抗体联合鼻腔被动免疫将 减轻老年晚期斑块5XFAD小鼠的AD样病理认知改变。所有资源， 包括转基因小鼠，以及所需的专业知识已经准备好开始这项研究。意义：该项目将 评估一种新的递送系统，将免疫疗法直接靶向大脑，从而促进 最有希望延缓或预防阿尔茨海默病进展的治疗方法。

This application is in response to PAS-10-151 {Grants for Alzheimer's Disease Drug Discovery (R21)}. Novel vehicles are needed to effectively target direct delivery of immunotherapeutics to the brain in order to maximize the efficacy of passive immunization for treating, delaying or preventing Alzheimer's disease (AD), a disease that currently afflicts >5 million Americans. Conjugation of the lectin, Wheat Germ Agglutinin (WGA), has been shown to enhance intranasal uptake of candidate drugs and promote direct delivery to brain via (i) Adsorptive or receptor-mediated endocytosis into the olfactory sensory neurons (OSN) followed by intracellular transport to olfactory bulb; (ii) Non-specific fluid phase endocytosis into the OSNs followed by intracellular transport to olfactory bulb; and/or (iii) Extracellular diffusion along the inter-olfactory epithelial clefts directly to the olfactory bulb and/or CSF. Passive immunization by intravenous injection of Ass monoclonal antibodies (mAb) has shown promising results such as lowering of cerebral Ass and stabilization or prevention of cognitive deficits in preclinical studies in transgenic mouse models of AD and in early human clinical trials. However, the level of antibody penetration in the brain is limited by the blood-brain-barrier (BBB). In addition, in mice and humans with vascular amyloid, high levels of anti-Ass antibodies in blood has led to microhemorrhage and vasogenic edema. This project aims to overcome such limitations by maximizing the efficacy of the intranasal route of anti-Ass antibody delivery using a WGA carrier protein. This project will test the efficacy of WGA as a novel vehicle that will not only enhance delivery of anti-Ass mAb, 3A1, that binds to mono/dimers and fibrillar Ass (To be provided by Dr. Lemere-Consultant) by virtue of its endocytic uptake preference, but will also facilitate passive diffusion of 3A1 due to its nasal mucosa-like biochemical composition, maximizing immunotherapeutic efficacy of intranasal passive immunization, to be tested in Alzheimer's 5XFAD model. Hypothesis: Intranasal delivery of WGA-conjugated 3A1 mAb combined with unlabeled 3A1 antibody will maximize the immunotherapeutic potential of intranasal passive immunization ameliorating AD-like patho-cognitive changes in 5XFAD transgenic mice. Specific Aims: [1] Trafficking of WGA-3A1 antibody after intranasal delivery to the brain in wild type mice and binding of 3A1-WGA to Ass plaques in the brains of 5XFAD mice and human AD. [2] Determining if intranasal passive immunization with WGA-conjugated 3A1 mAb combined with unlabeled 3A1 antibody will prevent AD-like patho-cognitive changes in young pre-plaque 5XFAD mice. [3] Determining if intranasal passive immunization with WGA-conjugated 3A1 mAB combined with unlabeled 3A1 antibody will attenuate AD-like patho-cognitive changes in old late-stage plaque-bearing 5XFAD mice. All resources, including transgenic mice, and required expertise are ready to begin this study. Significance: This project will evaluate a novel delivery system to target immunotherapeutics directly to the brain, and thus advance one of the most promising therapies for delaying the progression or prevention of AD.