Lu Patrick Y, Xie Frank, Woodle Martin C
Intradigm Corporation, Rockville, Maryland 20852, USA.
Adv Genet. 2005;54:117-42. doi: 10.1016/S0065-2660(05)54006-9.
RNAi has rapidly become a powerful tool for drug target discovery and validation in cell culture, and now has largely displaced efforts with antisense and ribozymes. Consequently, interest is rapidly growing for extension of its application to in vivo systems, such as animal disease models and human therapeutics. Studies on RNAi have resulted in two basic methods for its use for gene selective inhibition: 1) cytoplasmic delivery of short dsRNA oligonucleotides (siRNA), which mimics an active intermediate of an endogenous RNAi mechanism and 2) nuclear delivery of gene expression cassettes that express a short hairpin RNA (shRNA), which mimics the micro interfering RNA (miRNA) active intermediate of a different endogenous RNAi mechanism. Non-viral gene delivery systems are a diverse collection of technologies that are applicable to both of these forms of RNAi. Importantly, unlike antisense and ribozyme systems, a remarkable trait of siRNA is a lack of dependence on chemical modifications blocking enzymatic degradation, although chemical protection methods developed for the earlier systems are being incorporated into siRNA and are generally compatible with non-viral delivery systems. The use of siRNA is emerging more rapidly than for shRNA, in part due to the increased effort required to construct shRNA expression systems before selection of active sequences and verification of biological activity are obtained. In contrast, screens of many siRNA sequences can be accomplished rapidly using synthetic oligos. It is not surprising that the use of siRNA in vivo is also emerging first. Initial in vivo studies have been reported for both viral and non-viral delivery but viral delivery is limited to shRNA. This review describes the emerging in vivo application of non-viral delivery systems for RNAi for functional genomics, which will provide a foundation for further development of RNAi therapeutics. Of interest is the rapid adaptation of ligand-targeted plasmid-based nanoparticles for RNAi agents. These systems are growing in capabilities and beginning to pose a serious rival to viral vector based gene delivery. The activity of siRNA in the cytoplasm may lower the hurdle and thereby accelerate the successful development of therapeutics based on targeted non-viral delivery systems.
RNA干扰已迅速成为细胞培养中药物靶点发现和验证的强大工具，目前在很大程度上已取代了反义核酸和核酶的相关研究。因此，将其应用扩展到体内系统（如动物疾病模型和人类治疗）的兴趣迅速增长。关于RNA干扰的研究产生了两种用于基因选择性抑制的基本方法：1）将短双链RNA寡核苷酸（siRNA）胞质递送，其模拟内源性RNA干扰机制的活性中间体；2）将表达短发夹RNA（shRNA）的基因表达盒核递送，其模拟不同内源性RNA干扰机制的微小干扰RNA（miRNA）活性中间体。非病毒基因递送系统是适用于这两种RNA干扰形式的多种技术的集合。重要的是，与反义核酸和核酶系统不同，siRNA的一个显著特点是不依赖于阻止酶促降解的化学修饰，尽管为早期系统开发的化学保护方法正在被纳入siRNA，并且通常与非病毒递送系统兼容。siRNA的使用比shRNA出现得更快，部分原因是在获得活性序列并验证生物活性之前构建shRNA表达系统需要付出更多努力。相比之下，使用合成寡核苷酸可以快速完成许多siRNA序列的筛选。siRNA在体内的使用也首先出现也就不足为奇了。关于病毒和非病毒递送的体内初步研究均有报道，但病毒递送仅限于shRNA。本综述描述了用于功能基因组学的RNA干扰非病毒递送系统在体内的新兴应用，这将为RNA干扰疗法的进一步发展奠定基础。有趣的是，基于配体靶向质粒的纳米颗粒作为RNA干扰剂正在迅速得到应用。这些系统的能力不断增强，开始对基于病毒载体的基因递送构成严重挑战。siRNA在细胞质中的活性可能会降低障碍，从而加速基于靶向非病毒递送系统的治疗方法的成功开发。
