Microparticle Therapy for Cerebral Amyloid Angiopathy

项目总结 脑淀粉样血管病是一种微血管病变，其中β-淀粉样蛋白(Aβ)积聚在脑壁中 脑血管病变与血管脆性和继发于血管壁的出血有关 崩溃了。CAA对血管平滑肌细胞(VSMC)尤其有害。70%-90%的人存在CAA 阿尔茨海默病(AD)病例，增加出血性中风的风险，并因活性淀粉样蛋白而加重 免疫疗法因此损害了这一前景光明的AD治疗方法。CAA目前尚无有效的治疗方法。 尽管CAA在AD中普遍存在，而且AD和CAA是不同的疾病，但CAA往往 在AD研究中被忽视，因为Aβ被广泛推定为导致VSMC在 一艘装载着β的船只。CAA中VSMC的丢失是由于补体系统的细胞膜的形成 含β的CAA血管中膜攻击复合体。基于我们的这一发现 科学团队，我们的目标是开发出有史以来第一种基于抑制MAC形成的CAA治疗方法 我们假设可以预防CAA诱导的血管脆弱性。我们计划以抑制MAC形成为目标 与全身性β抑制相反的是，由于MAC对 针对微生物感染的免疫保护。我们提出了一种鼻对脑纳米颗粒疗法，它由 二乙乙胺(DEAE)修饰的脱热壳聚糖携带CD59载体 它的表达废除了MAC的形成。我们已经开发了一种脱热壳聚糖的技术 这使得内部放置成为可能，因为商业上可获得的壳聚糖被内毒素污染。这些 污染物干扰了质粒的导入和基因的表达。我们的壳聚糖脱热技术 是基于氮等离子体的应用--这是用来去污电势的相同技术 9/11事件后寄给国会的炭疽信件。我们完成了第一阶段的SBIR，在这一阶段我们成功地建立了 含壳聚糖基因纳米粒转染原代人血管平滑肌细胞诱导其表面表达 通过纳米颗粒转染法在VSMCs中表达CD59，并成功地建立了对CD59转基因细胞的保护作用 来自MAC启动的细胞裂解。在这个第二阶段的SBIR中，我们打算(1)提高均匀、可重复的生产 DEAE壳聚糖纳米粒随时间稳定，以便在体内实现一致、可预测的 转染脑微血管并提供实质保护，(2)表征治疗 我们的CS-CD59纳米粒鼻对脑CAA治疗对相关转基因小鼠CAA模型的疗效 以及(3)检测转基因CAA小鼠模型的组织学、生理学和认知效应。 CD59的表达。该项目的成功完成将有助于实现我们的DEAE的知情翻译 壳聚糖纳米粒治疗慢性再生障碍性贫血进入临床。

PROJECT SUMMARY Cerebral Amyloid Angiopathy (CAA), a microvasculopathy in which beta-amyloid (Aβ) accumulates in the walls of cerebral blood vessels, is associated with vascular fragility and bleeding secondary to blood vessel wall breakdown. CAA is especially deleterious to vascular smooth muscle cells (VSMC). CAA is found in 70-90% of Alzheimer's disease (AD) cases, increases hemorrhagic stroke risk, and is exacerbated by active amyloid immunotherapy thereby compromising this promising AD therapeutic. There is no effective therapy for CAA. Despite the prevalence of CAA in AD and the fact that AD and CAA are different diseases, CAA is often overlooked in AD studies as Aβ has been widely presumed to be responsible for the VSMC loss in the walls of Aβ-laden vessels. VSMC loss in CAA occurs due to formation of the complement system's cytolytic membrane attack complex (MAC) in the tunica media of Aβ-laden CAA blood vessels. Based on this discovery by our scientific team, our goal is to develop a first-ever therapeutic for CAA based on inhibition of MAC formation which we hypothesize will prevent CAA-induced vascular fragility. We plan to target inhibition of MAC formation in the walls of Aβ-laden CAA blood vessels, as opposed to systemic MAC inhibition, due to the importance of MAC for immune protection against microbial infection. We propose a nose-to-brain nanoparticle therapy made of depyrogenated chitosan modified with diethylethylamine (DEAE) that delivers an encapsulated CD59 plasmid whose expression abrogates the formation of MAC. We have developed a technique to depyrogenate chitosan that enables internal placement since commercially available chitosans are contaminated with endotoxins. These contaminants interfere with plasmid transfection and gene expression. Our chitosan depyrogenation technique is based on the application of nitrogen plasma – the same technology employed to decontaminate potential anthrax letters sent to Congress after 9/11. We completed a Phase I SBIR in which we established successful transfection of primary human VSMCs with chitosan gene-containing nanoparticles, induced surface expression of CD59 in VSMCs via nanoparticle transfection, and established successful protection of CD59 transfected cells from MAC-initiated cell lysis. In this Phase II SBIR, we intend to (1) improve production of uniform, reproducible DEAE chitosan nanoparticles that are stable over time so as to achieve consistent, predictable in vivo transfection of brain microvasculature and provide parenchymal protection, (2) characterize the therapeutic efficacy of our nose-to-brain CS-CD59 nanoparticle CAA therapy in a relevant transgenic mouse model of CAA, and (3) determine histologic, physiologic, and cognitive effects in the transgenic CAA mouse model of enhanced CD59 expression. Successful completion of this project will help enable informed translation of our DEAE chitosan nanoparticle therapy for CAA into the clinic.