Enhanced APOE2 Expression into Brain for Therapeutic Strategy for Alzheimer's Disease

增强 APOE2 在大脑中的表达,用于阿尔茨海默病的治疗策略

基本信息

  • 批准号:
    10208342
  • 负责人:
  • 金额:
    $ 143.76万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2021
  • 资助国家:
    美国
  • 起止时间:
    2021-05-15 至 2024-04-30
  • 项目状态:
    已结题

项目摘要

SUMMARY/ABSTRACT: Alzheimer’s disease (AD) is a progressive neurodegenerative disease that has emerged as the most prevalent form of late-life dementia in humans. The accumulation, aggregation, and deposition of amyloid-β (Aβ) in the brain are central events in AD pathogenesis. Despite intense effort, an effective therapy for AD has yet to be established. While multiple genetic and environmental factors are involved in AD pathogenesis, the ε4 allele of the APOE gene encoding apolipoprotein E (APOE) is the strongest genetic risk factor for late-onset AD among the three human APOE genotypes (ε2, ε3, ε4). In humans, Aβ deposition is more pronounced in APOE4 carriers compared with non-carriers in both AD patients and aged healthy individuals. APOE plays a critical role in maintaining synaptic plasticity and neuronal function by controlling lipid homeostasis, with the APOE2 allele having a superior function. The ε2 allelic variant has been found to be more prevalent among centenarians and associated with decreased susceptibility to AD. Studies on the role of the APOE2 in relation to AD suggest that APOE2 is neuroprotective and positively associated with cognitive functions in aging. Therefore, increasing APOE2 levels in the brain is predicted to be an effective therapeutic strategy for AD. Development of successful strategies for treating these disorders is limited due to the protective function of blood brain barrier (BBB). Gene therapy possesses a broad potential for the treatment of numerous neurological diseases, including AD. However, the major challenge in the field of gene therapy is the design of safe non-viral vectors that can cross the BBB. The transferrin (Tf) receptors are present on the surface of brain endothelial cells. The lipid nanoparticles can be surface modified with Tf protein for targeting the brain endothelial receptors and conjugated to brain specific cell penetrating peptide (CPP) for improving their internalization into brain by overcoming receptor saturation. Therefore, we propose to design near neutral, PEGylated liposomal nanoparticles encapsulating gene and modifying the surface of nanoparticles with Tf and CPP. Furthermore, the transfection properties of chitosan will be utilized for improving the transfection of gene by facilitating endosomal escape via the proton-sponge mechanism inside the cells. The long-term goal of the proposed research is to design a non- viral gene delivery carrier for efficient delivery of plasmid DNA encoding APOE2 (pAPOE2) to brain for prevention and treatment of AD. We propose three specific aims to accomplish the long-term goal of the proposed research: Aim 1. Synthesize and characterize liposomal nanoparticles loaded with chitosan-pAPOE2 polyplexes: The brain specific CPP-liposomes will be synthesized using thin film hydration technique followed by insertion of Tf coupled micelles using post-insertion technique. We propose to use five BBB specific CPPs: (i) CGN (d- CGNHPHLAKYNGT); (ii) RDP (KSVRTWNEIIPSKGCLRVGGRCHPHVNGGGRRRRRRRRR; (iii) Rabies Virus Glycoprotein RVG-9R, (iv) a non-toxic fragment of tetanus toxin, tetanus toxin C fragment (TTC), and (v) penetratin. The liposomal nanoparticles will be evaluated for particle size, zeta potential, encapsulation efficiency, cell uptake and uptake mechanism(s), transfection efficiency, cell cytotoxicity, and hemolysis assay. The transport efficacy of APOE2 loaded liposomal nanoparticles will be evaluated across an in vitro BBB model designed by combining human cerebral microvascular endothelial cells (hCMEC/D3), human astrocytes and APP Swe/Ind- or MAPT P301L-overexpressing human neuroblastoma cells (SHSY5Y). We will also determine the effect of liposomal nanoparticles on Aβ levels and Tau phosphorylation in the medium and cell lysates from the co-culture system. Aim 2. Evaluate the in vivo biocompatibility, organ toxicity, pharmacokinetics and APOE2 expression in mice of varying ages: To establish successful gene therapies for AD, we will validate the Tf-CPP-liposomal nanoparticles for their biocompatibility, organ toxicity, and pharmacokinetics (biodistribution) in wild type mice at 3 months of age. In addition, the APOE2 gene delivery will be further validated in APOE-knockout mice at 3 and 24 months of ages. Aim 3. Assess the therapeutic effects of the functionalized-liposome-mediated APOE2 gene delivery on cognitive impairment and Aβ and tau pathology in vivo: To establish successful gene therapies for AD-related phenotypes and age-related cognitive decline, we will examine effects of APOE2 gene therapy through the functionalized-liposomes on neurobehaviors, synaptic functions and/or amyloid and tau pathology. The liposomes will be injected into amyloid model APP/PS1 mice (3 and 6 months old), tau model PS19 mice (3 and 6 months old) and aged wild-type mice (12 and 24 months old), and the effects will be assessed 3 months after the administration. For clinical relevance, we will measure neurofilament light chain (NfL) levels in their plasma samples to assess effects on neurodegeneration. In addition, we will also measure plasma concentrations of Aβ and p-tau in respective mouse models. In addition, interstitial fluid Aβ clearance will be analyzed using in vivo microdialysis in the APP/PS1 mice at 3 months of age 1 month after the administration. Collectively, we anticipate that the proposed study will contribute towards the development of high efficiency non-viral gene delivery system to cross the BBB for successful gene therapy for neurological disorders and determine protective effects of increasing brain APOE2 on AD-related conditions.
总结/摘要: 阿尔茨海默病(AD)是一种进行性神经退行性疾病, 老年痴呆症的一种。β淀粉样蛋白(Aβ)在大鼠脑内的积聚、聚集和沉积, 脑是AD发病机制中的中心事件。尽管付出了巨大的努力,但AD的有效治疗方法还没有找到。 确立了习虽然AD的发病机制涉及多种遗传和环境因素,但AD患者的ε4等位基因 编码载脂蛋白E(APOE)的APOE基因是晚发性AD最强的遗传危险因素, 三种人类APOE基因型(ε2,ε3,ε4)。在人类中,Aβ沉积在APOE 4携带者中更明显 与非携带者相比,AD患者和老年健康人。APOE在 APOE 2等位基因通过控制脂质稳态维持突触可塑性和神经元功能 具有上级功能。ε2等位基因变异在百岁老人中更为普遍, 与AD易感性降低有关。关于APOE 2在AD中的作用的研究表明, APOE 2具有神经保护作用,并与衰老中的认知功能呈正相关。因此提高 预测脑中的APOE 2水平是AD的有效治疗策略。发展成功的 由于血脑屏障(BBB)的保护功能,治疗这些疾病的策略受到限制。基因 该疗法具有治疗包括AD在内的多种神经系统疾病的广泛潜力。 然而,基因治疗领域的主要挑战是设计安全的非病毒载体, 的BBB。转铁蛋白(Tf)受体存在于脑内皮细胞表面。脂质 纳米颗粒可以用Tf蛋白进行表面修饰,用于靶向脑内皮受体, 涉及脑特异性细胞穿透肽(CPP),用于通过克服 受体饱和因此,我们建议设计近中性的聚乙二醇化脂质体纳米粒, 用转铁蛋白和CPP修饰纳米粒表面。此外,转染 壳聚糖的性质将用于通过促进内体逃逸来改善基因的转染, 细胞内的质子海绵机制。这项研究的长期目标是设计一个非 将编码APOE 2的质粒DNA(pAPOE 2)有效递送到脑中用于预防的病毒基因递送载体 AD的治疗。我们提出了三个具体目标,以实现拟议研究的长期目标: 目标1。合成和表征负载有壳聚糖-pAPOE 2聚合复合物的脂质体纳米颗粒: 脑特异性CPP-脂质体将使用薄膜水合技术合成,然后插入 后插入法制备转铁蛋白偶联胶束。我们建议使用五种BBB特异性CPP:(i)CGN(d- CGNHPHLAKYNGT);(ii)RDP(KSVRTWNEIIPSKGCLRVGGRCHPHVNGGGRRRRRRRR);(iii)狂犬病病毒 糖蛋白RVG-9 R,(iv)破伤风毒素的无毒片段,破伤风毒素C片段(TTC),和(v) 穿透素将评价脂质体纳米颗粒的粒度、zeta电位、包封 效率、细胞摄取和摄取机制、转染效率、细胞毒性和溶血测定。 将在体外BBB模型中评价载APOE 2的脂质体纳米颗粒的转运功效 通过将人脑微血管内皮细胞(hCMEC/D3)、人星形胶质细胞和 APP Swe/Ind或MAPT P301 L过表达的人神经母细胞瘤细胞(SHSY 5 Y)。我们还将确定 脂质体纳米颗粒对来自人胰腺癌的培养基和细胞裂解物中的Aβ水平和Tau磷酸化的影响 共同文化体系。目标二。评价体内生物相容性、器官毒性、药代动力学和 APOE 2在不同年龄小鼠中的表达:为了建立成功的AD基因疗法,我们将验证 Tf-CPP-脂质体纳米粒的生物相容性、器官毒性和药代动力学 (生物分布)。此外,APOE 2基因的递送将进一步 在3个月和24个月大的APOE敲除小鼠中验证。目标3。评估治疗效果 功能化脂质体介导的APOE 2基因治疗认知功能障碍及对Aβ和tau蛋白的影响 体内病理学:为AD相关表型和年龄相关认知功能障碍建立成功的基因疗法。 下降,我们将检查APOE 2基因治疗的影响,通过功能化脂质体, 神经行为、突触功能和/或淀粉样蛋白和tau病理学。脂质体将被注入淀粉样蛋白 模型APP/PS1小鼠(3和6月龄)、tau模型PS19小鼠(3和6月龄)和老年野生型小鼠 (12和24月龄),并在给药后3个月评估效果。对于临床相关性, 我们将测量他们血浆样本中的神经丝轻链(NfL)水平,以评估对 神经变性此外,我们还将测量相应小鼠中Aβ和p-tau的血浆浓度, 模型此外,将在APP/PS1中使用体内微透析分析间质液Aβ清除率 给药后1个月3月龄的小鼠。总的来说,我们预计拟议的研究将 有助于开发高效的非病毒基因递送系统,以穿过BBB, 神经系统疾病的成功基因治疗和确定增加脑APOE 2的保护作用 与AD有关的疾病

项目成果

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Takahisa Kanekiyo其他文献

Takahisa Kanekiyo的其他文献

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{{ truncateString('Takahisa Kanekiyo', 18)}}的其他基金

Neuronal ABCA7 loss of function and Alzheimer’s disease
神经元 ABCA7 功能丧失与阿尔茨海默病
  • 批准号:
    10629715
  • 财政年份:
    2023
  • 资助金额:
    $ 143.76万
  • 项目类别:
Therapeutic Strategy to Treat Alzheimer's Disease by VGF Delivery into Brain
通过将 VGF 输送至大脑来治疗阿尔茨海默病的治疗策略
  • 批准号:
    10738951
  • 财政年份:
    2023
  • 资助金额:
    $ 143.76万
  • 项目类别:
Biomarker Core
生物标志物核心
  • 批准号:
    10667447
  • 财政年份:
    2021
  • 资助金额:
    $ 143.76万
  • 项目类别:
Biomarker Core
生物标志物核心
  • 批准号:
    10407939
  • 财政年份:
    2021
  • 资助金额:
    $ 143.76万
  • 项目类别:
Administrative Core
行政核心
  • 批准号:
    10667436
  • 财政年份:
    2021
  • 资助金额:
    $ 143.76万
  • 项目类别:
Enhanced APOE2 Expression into Brain for Therapeutic Strategy for Alzheimer's Disease
增强 APOE2 在大脑中的表达,用于阿尔茨海默病的治疗策略
  • 批准号:
    10514954
  • 财政年份:
    2021
  • 资助金额:
    $ 143.76万
  • 项目类别:
Impact of vascular apoE in aging and AD
血管apoE对衰老和AD的影响
  • 批准号:
    10407947
  • 财政年份:
    2021
  • 资助金额:
    $ 143.76万
  • 项目类别:
Administrative Core
行政核心
  • 批准号:
    10407936
  • 财政年份:
    2021
  • 资助金额:
    $ 143.76万
  • 项目类别:
Impact of vascular apoE in aging and AD
血管apoE对衰老和AD的影响
  • 批准号:
    10667475
  • 财政年份:
    2021
  • 资助金额:
    $ 143.76万
  • 项目类别:
Pathogenic mechanisms of ABCA7 in Alzheimer's disease
ABCA7在阿尔茨海默病中的发病机制
  • 批准号:
    9221000
  • 财政年份:
    2017
  • 资助金额:
    $ 143.76万
  • 项目类别:

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环境铜暴露及其对小胶质细胞 Abeta 清除的影响
  • 批准号:
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Environmental copper exposure and its impact on microglial Abeta clearance
环境铜暴露及其对小胶质细胞 Abeta 清除的影响
  • 批准号:
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  • 财政年份:
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人 ApoE 在体内通过 LDLR 清除可溶性 Aβ 中的作用
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人 ApoE 在体内通过 LDLR 清除可溶性 Aβ 中的作用
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    20590697
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    $ 143.76万
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
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