Fluiter Kees, Mook Olaf R F, Baas Frank
Department of Neurogenetics, AMC, Amsterdam, The Netherlands.
Methods Mol Biol. 2009;487:189-203. doi: 10.1007/978-1-60327-547-7_9.
Post-transcriptional gene silencing mediated by double-stranded RNA represents an evolutionarily conserved cellular mechanism. Small dsRNAs, such as microRNAs (miRNAs), are part of the main regulatory mechanisms of gene expression in cells. The possibilities of harnessing this intrinsic natural mechanism of gene silencing for therapeutic applications was opened up by the discovery by Tom Tuschl's team a few years ago that chemically synthesized small 21-mers of double-stranded RNA (small interfering RNA, siRNA) could inhibit gene expression without induction of cellular antiviral-like responses, siRNAs are especially of interest for cancer therapeutics because they allow specific inhibition of mutated oncogenes and other genes that aid and abet the growth of cancer cells. However, recent insights make it clear that siRNA faces some major hurdles before it can be used as a drug. Some of these problems are similar to those associated with classic antisense approaches, such as lack of delivery to specific tissues (other than the liver) or tumors, while other problems are more specific for siRNA, such as stability of the RNA molecules in circulation, off-target effects, interference with the endogenous miRNA machinery, and immune responses toward dsRNA. Chemical modifications of siRNA may help prevent these unwanted side effects. Initial studies show that minimal modifications with locked nucleic acids (LNA) help to reduce most of the unwanted side effects. In this chapter we will explore the limitations and possibilities of LNA-modified siRNA that may be used in future therapeutic applications.
由双链RNA介导的转录后基因沉默代表了一种进化上保守的细胞机制。小双链RNA，如微小RNA（miRNA），是细胞中基因表达主要调控机制的一部分。几年前，汤姆·图斯尔的团队发现化学合成的21个碱基对的小双链RNA（小干扰RNA，siRNA）能够抑制基因表达而不诱导细胞抗病毒样反应，这为利用这种内在的天然基因沉默机制进行治疗应用开辟了可能性。siRNA在癌症治疗中特别受关注，因为它们能够特异性抑制突变的癌基因以及其他促进癌细胞生长的基因。然而，最近的研究表明，siRNA在用作药物之前面临一些重大障碍。其中一些问题与经典反义方法相关的问题类似，例如难以递送至特定组织（肝脏以外）或肿瘤，而其他问题则是siRNA特有的，例如RNA分子在循环中的稳定性、脱靶效应、干扰内源性miRNA机制以及对双链RNA的免疫反应。对siRNA进行化学修饰可能有助于防止这些不良副作用。初步研究表明，用锁核酸（LNA）进行最小程度的修饰有助于减少大多数不良副作用。在本章中，我们将探讨LNA修饰的siRNA在未来治疗应用中可能存在的局限性和可能性。
