ABSTRACT

RNA interference (RNAi) by small interfering RNAs (siRNAs) is a promising therapeutic approach. Because siRNA has limited intracellular access and is rapidly cleared in vivo, the success of RNAi depends on efficient delivery technologies. Particularly, polyion complexation between block catiomers and siRNA is a versatile approach for constructing effective carriers, such as unit polyion complexes (uPIC), core-shell polyion complex (PIC) micelles and vesicular siRNAsomes, by engineering the structure of block catiomers. In this regard, the flexibility of block catiomers could be an important parameter in the formation of PIC nanostructures with siRNA, though its effect remains unknown. Here, we studied the influence of block catiomer flexibility on the assembly of PIC structures with siRNA using a complementary polymeric system, i.e. poly(ethylene glycol)-poly(L-lysine) (PEG-PLL) and PEG-poly(glycidylbutylamine) (PEG-PGBA), which has a relatively more flexible polycation segment than PEG-PLL. Mixing PEG-PGBA with siRNA at molar ratios of primary amines in polymer to phosphates in the siRNA (N/P ratios) higher than 1.5 promoted the multimolecular association of uPICs, whereas PEG-PLL formed uPIC at all N/P ratios higher than 1. Moreover, uPICs from PEG-PGBA were more stable against counter polyanion exchange than uPICs from PEG-PLL, probably due to a favorable complexation process, as suggested by computational studies of siRNA/block catiomer binding. In in vitro experiments, PEG-PGBA uPICs promoted effective intracellular delivery of siRNA and efficient gene knockdown. Our results indicate the significance of polycation flexibility on assembling PIC structures with siRNA, and its potential for developing innovative delivery systems.

 抽象的

小干扰RNA (siRNA) 的RNA 干扰(RNAi) 是一种有前途的治疗方法。由于 siRNA 进入细胞内的途径有限，并且在体内很快被清除，因此 RNAi 的成功取决于高效的递送技术。特别是，嵌段阳离子聚合物和 siRNA 之间的聚离子络合是一种通过改造嵌段阳离子聚合物的结构来构建有效载体的通用方法，例如单位聚离子复合物 (uPIC)、核壳聚离子复合物 (PIC) 胶束和囊泡 siRNA 体。在这方面，嵌段阳离子异构体的灵活性可能是用 siRNA 形成 PIC 纳米结构的一个重要参数，尽管其效果仍然未知。在这里，我们研究了嵌段阳离子异构体灵活性对使用互补聚合系统（即siRNA）组装 PIC 结构的影响。聚乙二醇-聚L-赖氨酸(PEG-PLL)和PEG-聚(缩水甘油基丁胺)(PEG-PGBA)，其具有比PEG-PLL相对更灵活的聚阳离子链段。将 PEG-PGBA 与 siRNA 以聚合物中伯胺与 siRNA 中磷酸盐的摩尔比（N/P 比）高于 1.5 混合，促进了 uPIC 的多分子缔合，而 PEG-PLL 在所有 N/P 比高于 1 时形成 uPIC此外，来自 PEG-PGBA 的 uPIC 比来自 PEG-PLL 的 uPIC 对反聚阴离子交换更稳定，这可能是由于有利的络合过程，如 siRNA/阻断阳离子异构体结合的计算研究所表明的。在体外实验中，PEG-PGBA uPIC 促进了 siRNA 的有效细胞内递送和有效的基因敲除。 我们的结果表明聚阳离子灵活性对于用 siRNA 组装 PIC 结构的重要性，及其开发创新递送系统的潜力。