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干扰在本世纪初被概念化并得到证实，并于2006年获得诺贝尔生理学和医学奖。此后，出现了利用RNA干扰用于治疗应用的巨大热潮。RNA干扰是一种天然存在的高度保守的生物学机制，其通过降解一类重要的细胞分子（称为信使核糖核酸（mRNA））来调节细胞功能。mRNA的降解发生在细胞内的胞质溶胶中，胞质溶胶是流体细胞隔室。在胞质溶胶中，mRNA的降解是通过与小干扰RNA（siRNA）的相互作用介导的，小干扰RNA是天然存在的分子。它们能够干扰大量不同的mRNA，从而干扰细胞功能，这使得RNA干扰对多种疾病具有巨大的治疗潜力，例如高胆固醇血症、病毒感染或癌症。对于治疗应用，将合成的siRNA整合到靶向的纳米尺寸的运输媒介物中，其促进运输媒介物与靶细胞的特异性结合。内吞进入细胞后，载体被递送到内体转运系统中，内体转运系统是一个复杂的、孤立的管状-囊泡隔室网络。这些管状-囊泡隔室被胞质溶胶包围。为了启动RNA干扰，siRNA必须从这些区室逃逸到胞质溶胶中，这一过程被称为内体逃逸。siRNA的内体逃逸不足是RNA干扰治疗成功的限制因素之一，并阻碍了基于siRNA的药物的总体功效。因此，研究可以促进siRNA内体逃逸的特异性细胞内递送策略将是siRNA递送领域的一大进步。在预期的项目中，将研究植物来源的递送系统（称为协同原理）用于将siRNA靶向递送到胞质溶胶中的潜力。作为一种进化优化的过程，协同原理是文献中描述的最有效的递送调节系统之一。该系统的基础是某些植物次级代谢物（例如皂苷SA 1641）以高度特异性的方式特异性地触发特定类别的植物衍生酶（皂苷）的内体逃逸的能力。因此，在所提出的项目中，设想产生掺入具有SA 1641的纳米载体中的siRNA，并研究SA 1641介导的siRNA内体逃逸。另一种实验策略是产生皂草素（SapKQ）的无酶活性变体作为siRNA的药物分子，并将SapKQ-siRNA缀合物和SA 1641整合到靶向纳米载体中，然后分析SA 1641介导的SapKQ-siRNA缀合物内体逃逸到胞质溶胶中。