A Non-Covalent Peptide-Based Strategy for siRNA Delivery
A Non-Covalent Peptide-Based Strategy for siRNA Delivery
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DOI：
10.1007/978-1-60761-919-2_25
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发表时间：
2011-01-01
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影响因子：
--
通讯作者：
Divita, Gilles
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文献类型：
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作者：
Crombez, Laurence;Divita, Gilles
短链干扰RNA(SiRNA)的发展为靶向治疗致病基因提供了巨大的希望。然而，细胞对si RNA的摄取不足，以及细胞膜对负电荷分子的低通透性，仍然是临床发展的主要障碍。到目前为止，还没有通用的siRNA传递方法，因为它们都存在一些限制。已经提出了几种非病毒策略来改善合成的si RNA在培养细胞和体内的传递。细胞穿透性多肽(CPP)或蛋白转导结构域(PTD)是非侵入性细胞导入siRNA的非常有前景的工具，基于非共价CPP/PTD的策略已成功地应用于体外和体内治疗性si RNA分子的递送。我们最近描述了一种新的基于多肽的系统，CADY，用于在原代和悬浮细胞系中高效地传递siRNA。Cady是一种二级两亲性多肽，能够与siRNA形成稳定的非共价复合体，并独立于内体途径提高细胞摄取能力。本章介绍了使用Cady-纳米颗粒技术将siRNA输送到贴壁和悬浮细胞系中的易于处理的方案。它还将突出不同关键点的肽/siRNA复合体的制备和转染方案，以便在低纳摩尔浓度下获得siRNA相关的干扰反应。
The development of short-interfering RNA (siRNA) has provided great hope for therapeutic targeting of specific genes responsible for pathological disorders. However, the poor cellular uptake of si RNA together with the low permeability of the cell membrane to negatively charged molecules, remain major obstacles to clinical development. So far there is no universal method for siRNA delivery as they all present several limitations. Several non-viral strategies have been proposed to improve the delivery of synthetic si RNAs in both cultured cells and in vivo. Cell-penetrating peptides (CPPs) or protein transduction domains (PTD) constitute very promising tools for non-invasive cellular import of siRNA and non-covalent CPP/PTD-based strategies have been successfully applied for ex vivo and in vivo delivery of therapeutic si RNA molecules. We recently described a new peptide-based system, CADY, for efficient delivery of siRNA in both primary and suspension cell lines. CADY is a secondary amphiphatic peptide able to form stable non-covalent complexes with siRNA and to improve their cellular uptake independently of the endosomal pathway. This chapter describes easy to handle protocols for the use of the CADY-nanoparticle technology for the delivery of siRNA into both adherent and suspension cell lines. It will also highlight different critical points in the peptide/siRNA complex preparation and transfection protocols, in order to obtain si RNA-associated interfering response at low nanomolar concentration.