Intranasal gene delivery for Alzheimer’s disease

项目概要/摘要 铁超负荷疾病，包括遗传性血色病（HH）和输血性血红蛋白病（例如， 地中海贫血、骨髓增生异常综合征、镰状细胞性贫血）影响着全世界数千万人。铁 超负荷是几种代谢疾病的发展和进展的明确的危险因素， 包括心肌病、肝硬化、关节炎、糖尿病和高脂血症。更重要的是，增加铁 大脑中的储存与神经退行性疾病（例如阿尔茨海默氏症、帕金森氏症和 亨廷顿氏病）。虽然铁螯合剂有效地从体内去除多余的铁，但它们表现出 显著毒性，包括胃肠道出血、粒细胞缺乏症、感染、心动过速、肾衰竭 和肝纤维化此外，没有螯合剂可以有效地恢复大脑中不适当的高铁， 神经退行性疾病患者。因此，存在对通过以下方法的新治疗策略的未满足的需求： 控制铁在大脑中的运输。膜铁转运蛋白（FPN）是主要的铁转运蛋白， 细胞内铁的输出。由于FPN对于肠道从饮食中摄取铁以及铁释放也是必不可少的， 从巨噬细胞回收金属用于红细胞生产，FPN的组织特异性调节可以 是一个很好的治疗目标，以改变脑铁转运，最小的全身效应。基因疗法可以 潜在地防止一些神经退行性疾病，包括阿尔茨海默氏症，帕金森氏症， 亨廷顿氏病，通过递送编码治疗分子的核酸。相反，基因沉默 选择性地降低不需要的分子的水平，如致癌蛋白和促炎蛋白。 细胞因子我们最近已经证明，鼻内给药纳米颗粒中的mRNA， 报告基因如荧光素酶和GFP在脑中的蛋白表达上调。这些结果 表明体内基因传递可以在铁紊乱领域中利用，并进一步促使我们 询问通过鼻内途径将FPN转基因直接递送至脑（作用部位）是否可以动员脑 铁储存，同时避免脱靶效应。因此，我们的假设是，鼻内给予FPN mRNA 在阳离子脂质体（CL）中，增强脑FPN表达，增加铁流出脑， 改善铁诱导的神经元损伤。具体目标集中在1）开发和验证 FPN转基因/CL以增加脑FPN水平和铁流出，以及2）评估FPN的治疗功效 使用铁相关的阿尔茨海默病的小鼠模型的转基因/CL。总的来说，这一战略提供了一个 铁催化的神经退行性变和 其他类型的神经系统疾病。

PROJECT SUMMARY/ABSTRACT Iron overload disorders, including hereditary hemochromatosis (HH) and transfusional hemoglobinopathies (e.g. thalassemia, myelodysplastic syndrome, sickle cell anemia), affect tens of millions of people worldwide. Iron overload is a well-defined risk factor for the development and progression of several metabolic diseases, including cardiomyopathy, liver cirrhosis, arthritis, diabetes and hypertriglyceridemia. Importantly, increased iron stores in the brain are closely associated with neurodegenerative diseases (e.g. Alzheimer’s, Parkinson’s and Huntington’s diseases). Although iron chelators are efficient to remove excess iron from the body, they exhibit significant toxicities, including gastrointestinal bleeding, agranulocytosis, infection, tachycardia, kidney failure and liver fibrosis. Moreover, there is no chelator that effectively restores inappropriately high iron in the brain of patients with neurodegenerative diseases. Hence, there is an unmet need for a new therapeutic strategy by controlling the transport of iron in the brain. Ferroportin (FPN) is the primary iron transporter responsible for the export of intracellular iron. Since FPN is also essential for intestinal iron uptake from diet as well as iron release from the macrophages to recycle the metal for red blood cell production, tissue-specific modulation of FPN can be an excellent therapeutic target to modify brain iron transport with minimal systemic effects. Gene therapy can potentially protect against a number of neurodegenerative diseases, including Alzheimer’s, Parkinson’s, and Huntington’s diseases, by delivering nucleic acid encoding for therapeutic molecules. Conversely, gene silencing selectively decreases the levels of unwanted molecules, such as oncogenic proteins and pro-inflammatory cytokines. We have recently demonstrated that intranasal administration of mRNA in nanoparticles significantly up-regulated protein expression of the reporter genes, such as luciferase and GFP, in the brain. These results suggested that in vivo gene delivery can be exploited in the area of iron disorders, and further prompted us to inquire if a direct delivery of FPN transgene to the brain (site of action) via the intranasal route can mobilize brain iron stores, while avoiding off-target effects. Thus, our hypothesis is that intranasal administration of FPN mRNA in cationic liposomes (CL) enhances brain FPN expression, increases efflux of iron out of the brain and ameliorates iron-induced neuronal impairments. The specific aims are focused on 1) developing and validating FPN transgene/CL to increase brain FPN levels and iron efflux and 2) evaluating the therapeutic efficacy of FPN transgene/CL using a mouse model of iron-associated Alzheimer’s disease. Overall, this strategy provides a selective, effective and safe approach for gene therapy in the area of iron-catalyzed neurodegenerative and other types of neurological disorders.