Development of a new system for the delivery of protective molecules to the heart

描述（由申请人提供）：心血管疾病（CVD）仍然是美国死亡的主要原因。在属于心血管疾病保护伞下的各种疾病中，心肌梗死（MI），更常见的是心脏病发作，直接导致大量归因于CVD的死亡，并且是归因于其他形式CVD的死亡率的促成因素。每年有120万新发或复发的MI病例报告，其中超过三分之一导致死亡，改善目前的治疗方法并开发新的更有效的MI治疗方法是当务之急。心脏研究人员已经确定了许多保护心脏免受I/R损伤的蛋白质。然而，这种知识的翻译到临床设置是阻碍了我们不能修改蛋白质和它们的表达水平在体内没有转基因。α B-晶体蛋白（1BC）是一种蛋白质，已被鉴定为在模拟心肌梗死的各种模型中具有稳健的心脏保护作用。此外，已经鉴定了增强蛋白质的保护作用的修饰。我们已经在开发一种系统中迈出了第一步，该系统可以在不使用转染，病毒转移或转基因的情况下将1BC蛋白质递送到细胞和组织中。这种独特的系统利用了细胞穿透肽（CPP）的特性，CPP具有使自身和货物穿过细胞膜的能力。我们设计了一种系统，该系统在1CB蛋白和CPP之间产生可逆的连接，从而允许将1BC递送到细胞中。一旦进入细胞内，两个分子之间的连接就被切断，释放出细胞内的1BC蛋白。我们已经证明，该系统在体外有效地工作，并且可以将1BC递送到培养中的细胞。此外，将与CPP肽连接的1BC注射到小鼠心脏的左心室壁中导致驻留心肌细胞摄取1BC蛋白。我们建议在细胞培养和动物模型中进行进一步研究，以继续开发该系统并在已建立的心脏模型中测试其性能。这些研究包括评估递送系统保护培养物中的细胞免受渗透、活性氧和缺氧应激的能力。我们还将检查该系统在离体模型中以提供I/R保护的方式向心脏递送1BC的能力。最后，我们将在MI的体内小鼠模型中检查递送系统将1BC递送到梗塞区内的心肌细胞并保护组织免受损伤和细胞死亡的能力。 公共卫生相关性：在美国，心脏病发作是造成大量死亡的原因，也是其他形式的心脏病相关死亡的主要因素。这项拟议中的研究探索了一种新技术，将保护性分子直接引入心脏细胞，目的是保护心脏免受心脏病发作造成的损害。

DESCRIPTION (provided by applicant): Cardiovascular disease (CVD) continues to be the leading cause of death in the U.S. Of the various conditions that fall under the umbrella of cardiovascular disease, myocardial infarction (MI), known more commonly as a heart attack, is directly responsible for a significant number of the deaths attributed to CVD, and is a contributing factor in mortality attributed to other forms of CVD. With 1.2 million new or recurrent cases of MI reported every year, more than a third of which result in death, improving on current treatments and developing new, more effective treatments for MI are top priorities. Cardiac researchers have identified many proteins that that protect the heart against I/R injury. However, translation of this knowledge to the clinical setting is hampered by our inability to modify proteins and their expression levels in vivo without transgenesis. Alpha B- Crystallin (1BC) is a protein that has been identified as having robust cardioprotective effects in various models which mimic myocardial infarction. Furthermore, modifications have been identified which enhance the protective effects of the protein. We have taken the first steps in developing a system to deliver 1BC proteins to cells and tissue without using transfection, viral transfer or trangenesis. This unique system utilizes the properties of cell penetrating peptides (CPP) which have the ability to move themselves and cargo across cell membranes. We have devised a system that creates a reversible linkage between the 1CB protein and the CPP allowing for the delivery of 1BC into cells. Once inside the cell, the linkage between the two molecules is cleaved, freeing the 1BC protein within the cell. We have demonstrated that this system works effectively in vitro and can deliver 1BC to cells in culture. Furthermore, injecting 1BC linked to the CPP peptide into the left ventricle wall of mouse hearts results in uptake of the 1BC protein by resident cardiomyocytes. We are proposing further studies in both cell culture and animal models, to continue the development of this system and to test its performance in established cardiac models. The studies include assessing the ability of the delivery system to protect cells in culture from osmotic, reactive oxygen species and hypoxic stressors. We will also examine the ability of the system to deliver 1BC to hearts, in an ex vivo model, in a manner that provides protection from I/R. Lastly, we will examine the ability of the delivery system to deliver 1BC to cardiomyocytes within the infarct zone and protect the tissue from damage and cell death in an in vivo mouse model of MI. PUBLIC HEALTH RELEVANCE: Heart attacks are responsible for a significant number of deaths in the U.S. and are a major contributing factor to other forms of heart disease related death. The proposed research explores a new technique to introduce protective molecules directly in to the cells of the heart, with the goal of protecting the heart against damage caused by a heart attack.