Targeted nanoparticle mediated delivery of nucleic acids into muscle stem cells for prevention of critical illness myopathy (B06#)

靶向纳米颗粒介导的核酸递送至肌肉干细胞以预防危重病肌病（B06

• 批准号: 397975747
• 金额: --
• 依托单位: Jena Center for Soft Matter (JCSM)
• 依托单位国家: 德国
• 项目类别: Collaborative Research Centres
• 财政年份: 2017
• 资助国家: 德国
• 起止时间: 2016-12-31 至 2020-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/397975747?language=en
• 关键词: Targeted; nanoparticle; mediated; delivery; nucleic

The genetic manipulation of cells in vivo is still challenging although numerous approaches are well established and investigated in vitro. One reason is the fact that targeted nanoparticles have to overcome different barriers subsequent to intravenous administration. Immunogenic recognition must be avoided, crossing of the endothelial barrier and internalization into the targeted cells are hurdles to be taken before the nanoparticles can transfer their payload to the site of action. In the case of nucleic acids as payload, further difficulties have to be taken into account: They are easily degraded and unable to cross membranes because of their hydrophilicity. Nanoparticles protect the genetic material and promote its endosomal uptake and release to the cytoplasm of the targeted cell in an active form. To import new information into cells, messenger RNA (mRNA) can be more sufficient as mRNA does not have to enter the cell nucleus. Thereby, delivery across the nuclear membrane is not necessary. Polymers with cationic charges can be used to bind, protect and transfer the negatively charged genetic material. This principle has been known since decades. However, a lot of gene carrier failed in vivo although being efficient in cell culture experiments. For an efficient gene delivery to targeted cells, multi-functional nanoparticles for the delivery of mRNA encoding GFP will be developed. The used polymers not only bind the genetic material by electrostatic interactions, due to their hydrophobic nature they also form particles. For in vivo applications, a second functional layer will be introduced to prevent side reactions and clearance. The cationic particles will be shielded by these block copolymers where one block contains pH value dependent anionic charges to bind to the cationic particle and the second block introduces stealth moieties. Functional end groups allow the introduction of targeting moieties to enhance the targeted gene delivery. The particles will be tested in an advanced cell culture setup using a fluidic approach combined with co-cultivation of immune as well as endothelial cells. Different aspects, such as cellular targeting and uptake of nanoparticles, endosomal release and intercellular delivery of mRNA as well as efficiency of nanoparticle mediated gene delivery will be addressed. Design parameters like size, surface charge, particle density, and the balance between active and passive targeting will be investigated within the project. “

尽管在体外已经建立并研究了许多方法，但体内细胞的遗传操作仍然具有挑战性。原因之一是靶向纳米颗粒在静脉注射后必须克服不同的障碍。必须避免免疫原性识别，穿过内皮屏障和内化到靶细胞中是纳米颗粒将其有效负载转移到作用位点之前需要克服的障碍。在核酸作为有效负载的情况下，必须考虑更多的困难：它们很容易降解，并且由于其亲水性而无法跨膜。纳米颗粒保护遗传物质并促进其内体摄取并以活性形式释放到靶细胞的细胞质中。要将新信息导入细胞，信使 RNA (mRNA) 就足够了，因为 mRNA 不必进入细胞核。因此，不需要穿过核膜的递送。带阳离子电荷的聚合物可用于结合、保护和转移带负电荷的遗传物质。这一原则几十年来就为人所知。然而，许多基因载体虽然在细胞培养实验中有效，但在体内却失败了。为了将基因有效递送至靶细胞，将开发用于递送编码 GFP 的 mRNA 的多功能纳米粒子。所使用的聚合物不仅通过静电相互作用结合遗传物质，由于其疏水性，它们还形成颗粒。对于体内应用，将引入第二个功能层以防止副反应和清除。阳离子颗粒将被这些嵌段共聚物屏蔽，其中一个嵌段含有与阳离子颗粒结合的pH值依赖性阴离子电荷，第二个嵌段引入隐形部分。功能性端基允许引入靶向部分以增强靶向基因递送。这些颗粒将在先进的细胞培养装置中使用流体方法结合免疫细胞和内皮细胞的共培养进行测试。将讨论不同的方面，例如纳米粒子的细胞靶向和摄取、mRNA 的内体释放和细胞间递送以及纳米粒子介导的基因递送的效率。该项目将研究尺寸、表面电荷、颗粒密度以及主动和被动靶向之间的平衡等设计参数。 “