Harnessing Platelet-Endothelial Interactions for Exosome Delivery

项目摘要 研究表明，成体干细胞，如间充质干细胞（MSC）， 心肌梗死（MI）或缺血/再灌注（I/R）损伤的间接旁分泌机制，而不是通过 分化和组织替换。在过去的十年中，外泌体已经成为有前途的无细胞 用于治疗缺血性损伤的药物。几家基于外泌体的治疗公司已经推出了早期阶段 临床试验与大多数治疗剂一样，迫切需要有效的递送策略以确保 足够数量的外泌体到达受伤的组织。由于它们是纳米级的天然脂质双层 颗粒，一种选择是血管输送。然而，由于在很大程度上， 脱靶分布到单核吞噬细胞系统和其他器官，如肝、脾和 肺此外，MSC衍生的外泌体（MSC-XO）需要与天然存在的外泌体竞争， 体液MSC-XO的细胞结合和摄取也阻碍了治疗效果。新颖的方法是 需要将治疗性外泌体递送到受损组织中的细胞。理想情况下，修饰的外泌体应该1） 减少单核吞噬细胞系统的清除，2）与受损血管结合，和3） 被损伤组织中的靶细胞类型有效地摄取。已经确定急性MI可以诱导 血管损伤并暴露内皮下基质的成分，包括胶原蛋白、纤连蛋白和血管内皮生长因子。 维勒布兰德因子（vWF）募集血小板。血小板可与受损的血管系统结合并在其上积聚 继MI之后。我们没有对亲本细胞进行一般性修饰或化学工程化外泌体，而是使用了 血小板膜包裹外泌体（制造P-XO），并增加其巨胞饮介导的 细胞内化和它们靶向损伤组织的能力。在这项研究中，我们计划 研究P-XO（AIM 1）的制备、表征和毒性。在那之后，我们将测试 P-XO对具有心脏损伤的小鼠模型（AIM 2）和猪模型（AIM 3）的作用。我们的研究将 为外泌体的细胞内化机制、靶向特性和机制提供了新的见解 P-XO治疗的效果总之，提出的机制和翻译实验将提供一个 科学前提，以了解系统管理的外来体，同时提出新的方法， 促进治疗性外泌体的靶向和治疗效果。

PROJECT SUMMARY Studies have demonstrated that adult stem cells such as mesenchymal stem cells (MSCs) repair myocardial infarction (MI) or ischemia/reperfusion (I/R) injury by indirect paracrine mechanisms rather than by differentiation and tissue replacement. In the past decade, exosomes have emerged as promising cell-free agents for treating ischemic injury. Several exosome-based therapeutic companies have launched early phase clinical trials. Like most therapeutics, there is an urgent need for effective delivery strategy to ensure a sufficient number of exosomes to reach the injured tissue. Since they are nanosized natural lipid bilayer particles, one option is vascular delivery. However, high and repeated dosing is needed due to a large degree of off-target distribution to the mononuclear phagocyte system and other organs, such as the liver, spleen and lungs. Moreover, MSC-derived exosomes (MSC-XOs) need to compete with naturally existing exosomes in body fluids. Cellular binding and uptake of MSC-XOs also hinder the therapeutic effects. Novel approaches are required to deliver therapeutic exosomes to cells in injured tissues. Ideally, modified exosomes should 1) reduce the clearance by the mononuclear phagocyte system, 2) bind to injured blood vessels, and 3) be efficiently uptake by target cell types in injured tissues. It has been established that acute MI can induce vascular damage and expose components of the subendothelial matrix including collagen, fibronectin and von Willebrand factor (vWF) to recruit platelets. Platelets can bind to and accumulate on injured vasculature following MI. Instead of generically modifying the parental cells or chemically engineering exosomes, we used platelet membranes to envelope exosomes (to make P-XOs) and increase their macropinocytosis-mediated cellular internalization, and their ability to target the injured tissue. In this proposed study, we plan to investigate the fabrication, characterization and toxicity of P-XOs (AIM 1). After that, we will test the therapeutic effect of P-XOs on a mouse model (AIM 2) and a porcine model (AIM 3) with cardiac injury. Our study will provide new insights on cellular internalization mechanism of exosomes, targeting properties and mechanism of the P-XOs treatment. Together, the proposed mechanistic and translational experiments will provide a scientific premise to understand system administrated exosomes while suggesting new approaches for promoting targeting and therapeutic effect of therapeutic exosomes.