An innovative strategy for the efficient cytosolic delivery of small interfering RNA (siRNA)

高效胞浆递送小干扰 RNA (siRNA) 的创新策略

• 批准号: 240246050
• 负责人: Professor Dr. Alexander Weng
• 金额: --
• 依托单位: Institut für Pharmazie
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2013
• 资助国家: 德国
• 起止时间: 2012-12-31 至 2013-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/240246050?language=en
• 关键词: innovative; strategy; efficient; cytosolic; delivery

RNA-interference was conceptualized and proven in the beginning of this century and was honored by the Nobel Prize for physiology and medicine in 2006. Thereafter, there has been tremendous upsurge to utilize RNA-interference for therapeutic applications. RNA-interference is a naturally occurring highly conserved biological mechanism that modulates cellular functions by the degradation of an important class of cellular molecules, which are known as messenger ribonucleic acid (mRNA). The degradation of mRNA takes place in the cell within the cytosol, which is a fluidic cellular compartment. In the cytosol the degradation of mRNA is mediated by the interaction with small interfering RNAs (siRNA), which are naturally occurring molecules. Their ability to inactivate a huge number of different mRNA and thus cellular functions makes RNA-interference of great therapeutic potential for diverse diseases, such as hypercholesterolemia, viral infections or cancer. For therapeutic applications, synthetic siRNA is integrated into a targeted nano-sized transport vehicle, which facilitates the specific binding of the transport vehicle to the target cell. After endocytosis into the cells the vehicle is delivered into the endosomal transport system, which is a complex, isolated network of tubular-vesicular compartments. These tubular-vesicular compartments are surrounded by the cytosol. To initiate RNA-interference siRNA has to escape from these compartments into the cytosol, a process, which is designated as endosomal escape. The insufficient endosomal escape of siRNA is one of the restricting factors for the therapeutic success of RNA-interference and hampers the overall efficacy of siRNA based drugs. The investigation of specific intracellular delivery strategies that could facilitate the endosomal escape of siRNA would therefore be a great step forward in the field of siRNA delivery. In the intended project the potential of a plant derived delivery system (termed as synergistic principle) will be investigated for the targeted delivery of siRNA into the cytosol. An evolutionarily optimized process, the synergistic principle is one of the most efficient delivery modulating systems ever described in literature. The basis for this system is the ability of certain plant secondary metabolites (e.g. Saponin SA1641) to specifically trigger the endosomal escape of a particular class of plant-derived enzymes (Saporin) in a highly specific manner. Therefore in the proposed project it is envisaged to generate siRNA incorporated in nano-vehicles with SA1641 and to investigate the SA1641-mediated endosomal escape of siRNA. Another experimental strategy is the generation of an enzymatically inactive variant of Saporin (SapKQ) as a drag molecule for siRNA and the integration of SapKQ-siRNA conjugates and SA1641 into targeted nano-vehicles followed by the analysis of the SA1641-mediated endosomal escape of SapKQ-siRNA conjugates into the cytosol.

RNA干扰在本世纪初概念化和证明，并于2006年获得诺贝尔生理学和医学奖的荣誉。此后，有很大的兴起来利用RNA的治疗应用。 RNA介入是一种天然存在的高度保守的生物学机制，通过降解一类重要的细胞分子来调节细胞功能，这些细胞分子被称为信使核糖核酸（mRNA）。 mRNA的降解发生在细胞质内的细胞中，这是一个液体细胞室。在细胞质中，mRNA的降解是通过与天然存在分子的小干扰RNA（siRNA）的相互作用介导的。它们使大量不同的mRNA失活并因此细胞功能的能力使RNA干扰具有多种疾病的巨大治疗潜力，例如高胆固醇血症，病毒感染或癌症。对于治疗应用，将合成siRNA集成到靶向纳米大小的运输车辆中，从而促进了运输车辆与目标细胞的特定结合。内吞作用到细胞中后，将车辆输送到内体运输系统中，该系统是一个复杂的，孤立的肾小管腔室网络。这些管状腔室被细胞质包围。要启动RNA干扰siRNA必须从这些隔室中逃脱到细胞质中，该过程被指定为内体逃生。 siRNA的内体逃逸不足是RNA干扰成功的限制因素之一，并阻碍了基于siRNA的药物的整体疗效。因此，对可能促进siRNA内体逃逸的特定细胞内输送策略的调查将是siRNA递送领域迈出的一大步。在预期的项目中，将研究植物衍生的输送系统（称为协同原理）的潜力，以将siRNA的靶向递送到细胞质中。一种进化优化的过程，协同原理是文献中描述的最有效的交付调节系统之一。该系统的基础是某些植物二级代谢产物（例如皂苷SA1641）的能力，以高度特异性的方式触发特定类别的植物衍生酶（saporin）的内体逃生。因此，在拟议的项目中，它设想生成具有SA1641的纳米车辆中的siRNA，并研究siRNA的SA1641介导的内体逃生。另一个实验策略是saporin（SAPKQ）作为siRNA的阻力分子的产生酶促的非活性变体，以及将SAPKQ-SIRNA偶联物和SA1641整合到靶向的纳米架中，然后分析SA1641介导的SapkqQ的sa1641-介导的sapkqqQ的分析。 -SiRNA结合到细胞质中。