Self-Assembly of siRNA/PEG-b-Catiomer at Integer Molar Ratio into 100 nm-Sized Vesicular Polyion Complexes (siRNAsomes) for RNAi and Codelivery of Cargo Macromolecules
Self-Assembly of siRNA/PEG-b-Catiomer at Integer Molar Ratio into 100 nm-Sized Vesicular Polyion Complexes (siRNAsomes) for RNAi and Codelivery of Cargo Macromolecules
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DOI：
10.1021/jacs.8b13641
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发表时间：
2019-02-27
影响因子：
15
通讯作者：
Kataoka, Kazunori
中科院分区：
文献类型：
--
作者：
Kim, Beob Soo;Chuanoi, Sayan;Kataoka, Kazunori
囊泡聚离子复合物（PIC）是通过刚性圆柱形分子、小干扰RNA（siRNA）与具有5-氨基戊基侧链的聚（乙二醇）（2kDa）和阳离子聚丙烯酰胺衍生物（70个单元）的柔性嵌段共聚物的自组装来制备的。100 nm大小的siRNA组装的囊泡PIC，称为siRNAsome，在siRNA和阻断剂之间的特定混合范围内制造。siRNA体膜由满足简单摩尔比（1：2或2：3）的阻断剂和siRNA的PIC单元组成。这些比率对应于最小整数摩尔比以最大限度地补偿PIC的电荷不平衡，因为每个嵌段阳离子聚合物和siRNA的电荷数分别为+70和-40。因此，以1：2和2：3制备的siRNAsome的zeta-电位分别为负和正。截面透射电镜观察表明，1：2和2：3 siRNAsome的膜厚度分别为11.0和17.2 nm。考虑到siRNA的计算长轴长度为5.9nm，这些厚度值对应于分别与一个和两个嵌段聚体缔合的两个（11.8nm）和三个（17.7nm）串联排列的siRNA的膜模型。对于生物学应用，siRNAsome通过与戊二醛的膜交联来稳定。带正电荷和交联的siRNAsome促进siRNA内化到培养的癌细胞中，引起显著的基因沉默，细胞毒性可忽略不计。通过简单的涡旋混合，siRNA体稳定地将作为模型货物大分子的葡聚糖包封在空腔中。共聚焦激光扫描显微镜观察显示，两种有效载荷一起内化到培养的细胞中。这些结果证明了siRNAsome作为siRNA与其他货物大分子共递送的通用平台的潜力。
Vesicular polyion complexes (PICs) were fabricated through self-assembly of rigid cylindrical molecules, small interfering RNAs (siRNAs), with flexible block catiomers of poly(ethylene glycol) (2 kDa) and cationic polyaspartamide derivative (70 units) bearing a 5-aminopentyl side chain. 100 nm-sized siRNA-assembled vesicular PICs, termed siRNAsomes, were fabricated in specific mixing ranges between siRNA and block catiomer. The siRNAsome membrane was revealed to consist of PIC units fulfilling a simple molar ratio (1:2 or 2:3) of block catiomer and siRNA. These ratios correspond to the minimal integer molar ratio to maximally compensate the charge imbalance of PIC, because the numbers of charges per block catiomer and siRNA are +70 and -40, respectively. Accordingly, the zeta-potentials of siRNAsomes prepared at 1:2 and 2:3 were negative and positive, respectively. Cross-section transmission electron microscopic observation clarified that the membrane thicknesses of 1:2 and 2:3 siRNAsomes were 11.0 and 17.2 nm, respectively. Considering that a calculated long-axial length of siRNA is 5.9 nm, these thickness values correspond to the membrane models of two (11.8 nm) and three (17.7 nm) tandemly aligned siRNAs associating with one and two block catiomers, respectively. For biological application, siRNAsomes were stabilized through membrane-cross-linking with glutaraldehyde. The positively charged and cross-linked siRNAsome facilitated siRNA internalization into cultured cancer cells, eliciting significant gene silencing with negligible cytotoxicity. The siRNAsome stably encapsulated dextran as a model cargo macromolecule in the cavity by simple vortex mixing. Confocal laser scanning microscopic observation displayed that both of the payloads were internalized together into cultured cells. These results demonstrate the potential of siRNAsomes as a versatile platform for codelivery of siRNA with other cargo macromolecules.