Native chemical ligation for conversion of sequence-defined oligomers into targeted pDNA and siRNA carriers.
Native chemical ligation for conversion of sequence-defined oligomers into targeted pDNA and siRNA carriers.
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DOI：
10.1016/j.jconrel.2014.02.015
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发表时间：
2014-04
期刊：
影响因子：
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通讯作者：
C. Zhang;P. Kos;Katharina Müller;Waldemar Schrimpf;Christina Troiber;Ulrich Lächelt;C. Scholz;D. Lamb;E. Wagner
中科院分区：
文献类型：
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作者：
C. Zhang;P. Kos;Katharina Müller;Waldemar Schrimpf;Christina Troiber;Ulrich Lächelt;C. Scholz;D. Lamb;E. Wagner
天然化学连接(NCL)被建立用于将不同拓扑结构的序列定义的低聚物转化为靶向和聚乙二醇屏蔽的PDNA和siRNA载体。从已有的非靶向低聚乙氨基酸库中，选择6个N-端半胱氨酸齐聚物为阳离子核心，以NCL为靶向配体(或末端丙氨酸为靶向阴性对照配体)与单分散聚乙二醇单分散叶酸(或末端丙氨酸)连接。对连接的偶联物和对照组(总共18个寡聚体)进行了PDNA或siRNA基因传递的评估。测定生物物理特性，包括在无血清或无血清存在时的核酸结合，以及在细胞摄取和基因转移(或基因沉默)方面的生物活性。在大多数情况下，与丙氨酸-聚乙二醇组相比，叶酸-聚乙二醇连接的寡聚体表现出强烈的细胞结合、摄取和基因转移到受体阳性的KB细胞。通过增加阳离子臂的数量来改变拓扑结构，添加酪氨酸三聚体作为多链稳定结构域，或促进内体逃逸的组氨酸，都会产生有益的基因转移特性。筛查显示了对PDNA和siRNA递送的不同要求。叶酸-聚乙二醇组二基化四臂低聚物对pDNA的传递最有效，但对siRNA无效，而叶酸-聚乙二醇三聚体修饰的三臂低聚物在siRNA介导的基因沉默中最有效。结果表明，位置选择性NCL反应是一种有效的低聚物修饰方法。因此，可以容易地获得多功能靶向载体，并用于识别随后的活体递送的前导结构。
Native chemical ligation (NCL) was established for the conversion of sequence-defined oligomers of different topologies into targeted and PEG shielded pDNA and siRNA carriers. From an existing library of non-targeted oligoethanamino amides, six oligomers containingN-terminal cysteines were selected as cationic cores, to which monodisperse polyethylene glycol (PEG) containing terminal folic acid as targeting ligand (or terminal alanine as targeting negative control ligand) were attached by NCL. Ligated conjugates plus controls (in sum 18 oligomers) were evaluated for pDNA or siRNA gene delivery. Biophysical characteristics including nucleic acid binding in the absence or presence of serum, as well as biological activities in cellular uptake and gene transfer (or gene silencing, respectively) were determined. In most cases, the folic acid-PEG-ligated oligomers displayed a strongly improved cellular binding, uptake and gene transfer into receptor-positive KB cells as compared to the alanine-PEG controls. Changing the topological structures by increasing the number of cationic arms, adding tyrosine trimers as polyplex stabilizing domains, or histidines facilitating endosomal escape resulted in beneficial gene transfer characteristics. The screen revealed different requirements for pDNA and siRNA delivery. A folate-PEG ligated histidinylated four-arm oligomer was most effective for pDNA delivery but inactive for siRNA, whereas a folate-PEG-ligated three-arm oligomer with tyrosine trimer modifications was most effective in siRNA mediated gene silencing. The results demonstrate the site-selective NCL reaction as powerful method to modify existing oligomers. Thus multifunctional targeted carriers can be obtained with ease and used to identify lead structures for subsequentin vivodelivery.