Biomedical Research Center, Korea Institute of Science and Technology, 39-1 Hawolgok-dong, Seongbuk-gu, Seoul 136-791, South Korea.
J Control Release. 2010 Feb 15;141(3):339-46. doi: 10.1016/j.jconrel.2009.10.007. Epub 2009 Oct 14.
Small interfering RNA (siRNA) is a promising biological strategy for treatment of diverse diseases, but the therapeutic application of siRNA has been limited by its instability and poor cellular uptake efficiency. Although the development of various gene delivery systems has increased the siRNA delivery efficiency, many problems still remain to be resolved before the clinical application of siRNA. In this study, we suggest reducible polymerized siRNA a possible solution for low delivery efficiency of siRNA. Dithiol-modified red fluorescent protein (RFP) siRNAs at the 5'-ends of both sense and anti-sense strands were disulfide-polymerized. Polymerized siRNA (poly-siRNA) was composed of 30% oligomeric siRNA (50 approximately 300 bps) and 66% polymeric siRNA (above approximately 300 bps) as fractions, and was reducible in reducing solution through disulfide bond cleavage. Poly-siRNA formed more condensed and nano-sized complexes with low molecular weight polyethylenimine (PEI) by strong electrostatic interaction based on the higher charge density of poly-siRNA, compared with siRNA (mono-siRNA). The compact poly-siRNA/PEI complexes prevented the loss and degradation of siRNA from a polyanion competitor and RNases in serum. Furthermore, poly-siRNA/PEI complexes exhibited superior intracellular uptake by murine melanoma cells (B16F10), and was accompanied with RFP gene silencing efficiency of about 80%, compared to untreated cells. These results sufficiently support that strong polyanionic and reducible poly-siRNA can be utilized as a novel powerful therapeutic strategy for human diseases.
小干扰 RNA（siRNA）是一种有前途的治疗多种疾病的生物策略，但由于其不稳定性和细胞摄取效率低，siRNA 的治疗应用受到限制。尽管各种基因传递系统的发展提高了 siRNA 的传递效率，但在 siRNA 的临床应用之前，仍有许多问题需要解决。在这项研究中，我们提出了还原性聚合 siRNA 作为提高 siRNA 递送效率的一种可能的解决方案。二硫键修饰的红色荧光蛋白（RFP）siRNA 在正义链和反义链的 5'-端被二硫键聚合。聚合 siRNA（poly-siRNA）由 30%的寡聚 siRNA（50 至 300 个碱基对）和 66%的聚合 siRNA（大于 300 个碱基对）组成，在还原溶液中通过二硫键断裂可还原。与 siRNA（mono-siRNA）相比，由于 poly-siRNA 的电荷密度更高，poly-siRNA 通过强静电相互作用与低分子量聚乙烯亚胺（PEI）形成更致密和纳米大小的复合物。与多阴离子竞争物和血清中的 RNases 相比，紧密的 poly-siRNA/PEI 复合物防止了 siRNA 的损失和降解。此外，与未经处理的细胞相比，poly-siRNA/PEI 复合物表现出对小鼠黑色素瘤细胞（B16F10）更高的细胞内摄取能力，并伴随着 RFP 基因沉默效率约为 80%。这些结果充分支持强聚阴离子和还原性 poly-siRNA 可作为一种新的强大的人类疾病治疗策略。
