Application of Brain-Targeted Anti-inflammatory RNA Nanoparticle for Ischemic Stroke Therapy

 描述(申请人提供)：尽管缺血性中风后神经元损伤的一些细胞机制已相当清楚，但由于缺乏有效的治疗方法，缺血性中风后神经元死亡无法得到很好的控制。我研究的长期目标是开发一种有效的缺血性中风治疗策略。这项拟议的研究集中在针对血脑屏障(BBB)的缺血后炎症，以寻找治疗中风的新方法。我们提出了一种新型的RNA纳米颗粒3WJ-PRNA/2siICAM-FB4，它由一个RNA适配子(FB4)、两个ICAM-1 siRNA和噬菌体phi29的包装RNA(3WJ-PRNA)的三向连接组成。FB4是转铁蛋白受体(TFR)的特异性RNA适配子，可以识别血脑屏障上的TFR，并促进RNA纳米颗粒进入脑内皮细胞。一旦3WJ-PRNA/2siICAM-FB4进入细胞，siRNAs将被释放，并下调ICAM-1的表达，从而抑制血脑屏障的炎症反应。PRNA三向连接是将FB4和siRNA结合在一起的核心，而不会破坏连接的有效载荷中的任何一个的折叠。我们将通过体内小鼠短暂性脑缺血模型和体外原代培养脑内皮细胞缺氧缺糖实验来论证这种血管靶向策略治疗缺血性卒中的可行性。我们的中心假设是3WJ-PRNA/2siICAM-FB4通过抑制缺血后炎症而对缺血性中风有治疗作用。具体目的：目的1：测定3WJ-PRNA/2siICAM-FB4的体外药理活性。目的：评价3WJ-PRNA/2siICAM-FB4对短暂性脑梗塞动物模型的保护作用。本项目将开发一种新的脑靶向RNA疗法用于治疗缺血性中风，该方法可广泛应用于影响血脑屏障和中枢神经系统(CNS)的疾病的治疗。

 DESCRIPTION (provided by applicant): Although some of the cellular mechanisms of neuronal injury after ischemic stroke are fairly well understood, the neuronal death after ischemic stroke cannot be well controlled because of a lack of efficient treatments. The long-term goal of my research is to develop an effective strategy for ischemic stroke therapy. The proposed research is focused on targeting the post- ischemic inflammation at blood-brain barrier (BBB) in search of new approaches for stroke therapy. We propose a novel RNA nanoparticle, 3WJ-pRNA/2siICAM-FB4, consisting of one RNA aptamer (FB4), two ICAM-1 siRNA and a 3-way junction of packaging RNA (3WJ-pRNA) of bacteriophage phi29. FB4 is a RNA aptamer specific to transferrin receptor (TfR) that can recognize TfRs at the BBB and facilitate RNA nanoparticle delivery into brain endothelial cells. Once the 3WJ-pRNA/2siICAM-FB4 enters the cell, the siRNAs will be released and knockdown ICAM-1 expression leading to inhibition of inflammatory responses at the BBB. The pRNA 3-way junction serves as the core for holding FB4 and siRNAs together without disrupting the folding of either of the attached payload. We will use mouse transient ischemic stroke model in vivo and oxygen-glucose deprivation in primary brain endothelial cells in vitro to demonstrate the feasibility of this vascular targeting strategy for treatment of ischemic stroke. Our central hypothesis is that the 3WJ-pRNA/2siICAM-FB4 has therapeutic effects on ischemic stroke by inhibition of post-ischemic inflammation. Specific Aims: Aim 1: Determine pharmacological activity of 3WJ-pRNA/2siICAM-FB4 in vitro. Aim 2: Evaluate protective effect of 3WJ- pRNA/2siICAM-FB4 in transient ischemic stroke model in vivo. This project will develop a novel brain-targeted RNA therapeutics for therapy of ischemic stroke, and this approach could be broadly applied for therapy of diseases affecting the BBB and the central nervous system (CNS).