Nanocarriers for delivery of antioxidants to endothelium

描述（由申请人提供）： 血管氧化应激在高氧、炎症、缺血、肺和心血管疾病等多种病理状态中起关键作用。然而，目前的抗氧化剂疗法是无效的，部分原因是次优递送至内皮细胞（EC）。以前的研究表明，抗氧化酶（AOE）过氧化氢酶（解毒自由扩散的H2 O2）结合抗体内皮细胞粘附分子（CAM）积累EC静脉注射后，保护动物免受急性氧化性肺损伤。不幸的是，抗CAM/AOE缀合物在几小时内从血液中消除并在EC中降解，仅产生短暂作用。为了解决这一问题并实现更高的治疗持续时间和效用，我们提出将过氧化氢酶加载到H2 O2可渗透的、抗CAM靶向的二嵌段PEG-PLGA聚合物纳米载体（PNC）中，这将延长AOE循环并保护免于溶酶体蛋白水解。中试数据显示，隐形抗CAM/PNC内装载的过氧化氢酶降解H2 O2，结合并保护EC免受氧化剂损伤，在动物体内循环很长一段时间，并在肺血管系统中积累。该补助金的目标是探索，测试和优化这一战略，追求以下具体目标：1。优化PNC的设计，以便AOE交付给EC。PEG-PLA PNC制剂、组成、尺寸、AOE负载、活性和蛋白酶抗性、pH依赖性降解速率、抗CAM偶联、EC结合、摄取和代谢以及EC保护将在体外进行研究和优化。2.表征抗CAM/PNC/AOE的体内行为和靶向。它们的血液清除率、全身效应、生物分布、EC结合、肺靶向、最佳给药途径和在肺中的持久性将在未处理动物和具有氧化剂血管应激的动物中进行测试。3.评价抗CAM/PNC/AOE在动物中的作用。将采用肺血管中急性H2 O2产生和肺部亚急性氧化应激（高氧）的小鼠模型来评估保护的程度和持续时间，并完善AOE靶向方案和保护机制。这些目标的完成将是朝着我们研究的长期目标迈出的重要一步，即将这种有前途的新技术平台转化为临床领域。

DESCRIPTION (provided by applicant): Vascular oxidant stress plays a key role in many pathologic states including hyperoxia, inflammation, ischemia, pulmonary and cardiovascular diseases. However, current antioxidant therapies are not effective, in part due to sub-optimal delivery to endothelial cells (EC). Previous studies have shown that the antioxidant enzyme (AOE) catalase (that detoxifies freely diffusing H2O2) conjugated with antibodies to endothelial Cell Adhesion Molecules (CAM) accumulates in EC after intravenous injection and protects animals against acute oxidant lung injury. Unfortunately, anti-CAM/AOE conjugates are eliminated from the blood and degraded in the EC within a few hours, affording only transient effects. In order to solve this problem and to achieve higher therapeutic duration and utility, we propose to load catalase into H2O2-permeable, anti-CAM targeted di-block PEG-PLGA Polymer Nano-Carriers (PNC) that would prolong the AOE circulation and protect from lysosomal proteolysis. Pilot data show that catalase loaded inside stealth anti-CAM/PNC degrades H2O2, binds to and protects EC from oxidant injury, circulates in animals for a prolonged time, and accumulates in the lung vasculature. The goal of this grant is to explore, test and optimize this strategy, pursuing the following Specific Aims: 1. Optimize the design of PNC for AOE delivery to EC. PEG-PLA PNC formulation, composition, size, AOE loading, activity and protease resistance, rate of pH-dependent degradation, coupling of anti-CAM, binding, uptake and metabolism by EC, and protection of EC will be studied and optimized in vitro. 2. Characterize the behavior and targeting of anti-CAM/PNC/AOE in vivo. Their blood clearance, systemic effects, biodistribution, EC binding, pulmonary targeting, optimal administration routes and persistence in lungs will be tested in naive animals and animals with oxidant vascular stress. 3. Evaluate the effects of anti-CAM/PNC/AOE in animals. Mouse models of acute H2O2 generation in the pulmonary vasculature and sub-acute oxidant stress in the lungs (hyperoxia) will be employed to evaluate the degree and duration of protection, and refine regimens of AOE targeting and mechanisms of protection. Completion of these Aims will be a major step towards the long-term goal of our research, which is translation of this promising new technology platform into the clinical domain.