A Brain Penetrating Bi-functional Transferrin Receptor Antibody-TNF-alpha Decoy Receptor Fusion Protein for Alzheimer's Disease

项目摘要/摘要 阿尔茨海默病(AD)的临床试验主要是以抗Aβ和抗tau为中心的，然而这 到目前为止，单一靶点治疗方法尚未成功。认为AD可以被阻止的想法，或者 因针对单个病理事件而延迟，需要修改和多层次的治疗方法 AD所需的。有限的多层次方法已经针对AD进行了测试，但失败的主要原因是 治疗时间不当，副作用大，缺乏脑穿透力。前景看好的多层次治疗 方法是以肿瘤坏死因子-α为靶点，它在AD病理的多个阶段发挥核心作用。现存生物 因此，肿瘤坏死因子-α抑制剂(TNFI)是一种潜在的治疗AD的方法，然而，它不能跨越血脑屏障 (Bbb)。到目前为止，还没有研究开发出针对AD的穿透血脑屏障的生物TNFi，该应用旨在 填补这一空白，并专注于为AD开发穿透BBB的生物TNFi。模型生物学 目前应用的TnFi为依那西普，它是一种II型肿瘤坏死因子-α诱骗受体(TnFR)-Fc融合蛋白。 我们已经将TNFR与抗小鼠转铁蛋白受体的嵌合抗体(CTfRMAb)融合，其中 CTfRMAb结构域充当分子特洛伊木马，通过 经血管途径。我们最近发现这种穿透血脑屏障的cTfRMAb-TNFR融合蛋白导致 在AD小鼠中比依那西普(非BBB穿透性TNFi)更好的治疗指标，强调了需要 用于脑内传递生物的TNFi。我们的中心假设是全身注射cTfRMAb- 肿瘤坏死因子受体同时调节外周和神经肿瘤坏死因子-α，在AD病理的多个水平上进行干预，并且是 因此对AD有治疗作用。在拟议的研究中，我们将使用两种不同的AD小鼠(APPswe，PSEN1dE9 双和3xTg三重转基因小鼠)，4个月和12个月龄模拟早期和晚期AD，并治疗 用赋形剂、cTfRMAb-TNFR、依那西普或cTfRMAb系统治疗，每周3天，疗程12周。以下是 具体目标将涉及：目标1：优化血脑屏障的剂量反应和治疗时间窗口 穿透生物TNFi。对神经病理指标(Aβ和Tau)及临床指标的影响 (认知缺陷)将使用三种剂量(1 mg/kg、3 mg/kg和6 mg/kg)进行研究。目标2：描述 穿透血脑屏障的生物肿瘤坏死因子的治疗作用机制。对以下方面的影响 将确定炎症、血脑屏障功能、氧化应激和突触/神经元丢失的标志。目标3： 评估长期服用穿透血脑屏障的生物肿瘤坏死因子的不良反应。两者都有 将研究cTfRMAb和肿瘤坏死因子-α抑制相关的不良反应。这些研究将有助于优化 导致高疗效毒性比的最低剂量，概述了肿瘤坏死因子-α的治疗时间窗口 抑制，确定参与cTfRMAb-TNFR治疗作用的机制可能使 确定新的靶点，并将提供必要的临床前结果，以推动 人类阿尔茨海默病cTfRMAb-TNFR研究进展

PROJECT SUMMARY / ABSTRACT Alzheimers disease (AD) clinical trials have primarily been anti-Aβ and anti-tau centric, however this single-target therapeutic approach has not been successful so far. The idea that AD can be stopped, or delayed by targeting a single pathological event, needs revision and a multi-level therapeutic approach is needed for AD. Limited multi-level approaches have been tested for AD but have failed primarily due to inappropriate time of treatment, side-effects and lack of brain penetration. A promising multi-level therapeutic approach is to target TNF-α which plays a central role at multiple stages of AD pathology. Existing biologic TNF-α inhibitors (TNFIs) are thus a potential treatment for AD, however, do not cross the blood-brain barrier (BBB). No study still date has developed a BBB-penetrating biologic TNFI for AD and this application aims to fill this gap, and focuses on the development of a BBB-penetrating biologic TNFI for AD. The model biologic TNFI in the current application is etanercept, which is a type-II TNF-α decoy receptor (TNFR)-Fc fusion protein. We have fused TNFR to a chimeric antibody against the mouse transferrin receptor (cTfRMAb), wherein the cTfRMAb domain acts as a molecular Trojan horse to ferry the TNFR into the brain non-invasively via the transvascular route. We have recently shown that this BBB-penetrating cTfRMAb-TNFR fusion protein results in better therapeutic indices than etanercept (non-BBB-penetrating TNFI) in AD mice, underscoring the need for brain delivery of the biologic TNFI. Our central hypothesis is that systemic administration of cTfRMAb- TNFR, which modulates both peripheral and neural TNF-α, intervenes at multiple levels of AD pathology and is thus therapeutic in AD. In the proposed studies, we will utilize two different AD mice (APPswe, PSEN1dE9 double and 3xTg triple transgenic mice), ages 4- and 12-months to mimic early- and late-stage AD, and treat with vehicle, cTfRMAb-TNFR, etanercept or cTfRMAb systemically 3 days/week for 12-weeks. The following specific aims will be addressed: Aim 1: Optimize dose-response and therapeutic time-window of the BBB- penetrating biologic TNFI. The effect on neuropathological hallmarks (Aβ and tau), and clinical hallmark (cognitive deficit) will be studied using three doses (1mg/kg, 3mg/kg and 6 mg/kg). Aim 2: Characterize the mechanisms involved in the therapeutic effects of the BBB-penetrating biologic TNFI. The effects on markers of inflammation, BBB function, oxidative stress and synaptic/neuronal loss will be determined. Aim 3: Evaluate the adverse effects associated with chronic dosing of the BBB-penetrating biologic TNFI. Both cTfRMAb- and TNF-α inhibition-relevant adverse-effects will be studied. These studies will help optimize the lowest dose that results in a high efficacy to toxicity ratio, outline the therapeutic time-window for TNF-α inhibition, identify mechanisms involved in the therapeutic effects of cTfRMAb-TNFR which may enable the identification of new targets, and will provide the necessary pre-clinical results that will advance the development of cTfRMAb-TNFR for AD in humans.