Palm-Apergi Caroline, Eguchi Akiko, Dowdy Steven F
Department of Cellular and Molecular Medicine, UCSD School of Medicine, Howard Hughes Medical Institute, La Jolla, CA, USA.
Methods Mol Biol. 2011;683:339-47. doi: 10.1007/978-1-60761-919-2_24.
A major hurdle in drug delivery today is for the drug to reach inside the cell to exert its biological effect. Many drug candidates are hydrophilic and are therefore not able to cross the hydrophobic plasma membrane, which serves to protect the cell from foreign molecules and pathogens. One promising drug candidate is the hydrophilic and negatively charged short-interfering RNA (siRNA), known to degrade target mRNA 1,000-fold more efficiently than small molecule drugs. The delivery capacity of small cationic peptides called protein transduction domains or cell-penetrating peptides, suggested them to be suitable delivery vehicles for siRNA. However, it has proven troublesome to utilize the PTD-siRNA conjugates for mRNA degradation due to the characteristics of siRNA, often resulting in precipitation and aggregation. This chapter describes a recently reported delivery strategy, PTD-DRBD fusion protein siRNA delivery, where a double-stranded RNA-binding domain expressed as a fusion protein together with three TAT PTDs binds the siRNA, thus masking the negatively charged backbone and preventing aggregation. This new protocol results in noncytotoxic mRNA degradation even more effective than lipofection.
当今药物递送中的一个主要障碍是药物要进入细胞内部以发挥其生物学效应。许多候选药物具有亲水性，因此无法穿过疏水的质膜，质膜的作用是保护细胞免受外来分子和病原体的侵害。一种有前景的候选药物是亲水且带负电荷的小干扰RNA（siRNA），已知其降解靶mRNA的效率比小分子药物高1000倍。被称为蛋白质转导结构域或细胞穿透肽的小阳离子肽的递送能力表明它们适合作为siRNA的递送载体。然而，由于siRNA的特性，利用PTD-siRNA缀合物进行mRNA降解已被证明很麻烦，常常导致沉淀和聚集。本章描述了一种最近报道的递送策略，即PTD-DRBD融合蛋白siRNA递送，其中与三个TAT PTD一起作为融合蛋白表达的双链RNA结合结构域结合siRNA，从而掩盖带负电荷的主链并防止聚集。这种新方法导致无细胞毒性的mRNA降解，甚至比脂质转染更有效。
