Multifunctional non-viral gene vectors with enhanced stability, improved cellular and nuclear uptake capability, and increased transfection efficiency.
Multifunctional non-viral gene vectors with enhanced stability, improved cellular and nuclear uptake capability, and increased transfection efficiency.
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DOI：
10.1039/c4nr02395a
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发表时间：
2014-08
期刊：
影响因子：
6.7
通讯作者：
Zhe Yang;Zhaozhong Jiang;Z. Cao;Chao Zhang;Di Gao;Xingen Luo;Xiaofang Zhang;Hui‐yan Luo;Qing Jiang;Jie Liu
中科院分区：
文献类型：
--
作者：
Zhe Yang;Zhaozhong Jiang;Z. Cao;Chao Zhang;Di Gao;Xingen Luo;Xiaofang Zhang;Hui‐yan Luo;Qing Jiang;Jie Liu
我们开发了一种新的多功能，非病毒基因传递平台，包括用于DNA凝聚的阳离子聚胺-共酯（PPMS），用于纳米颗粒稳定的PEG壳，用于加速细胞摄取的聚γ-谷氨酸（γ-PGA）和mTAT（一种细胞穿透肽），以及用于增强DNA到细胞核的细胞内运输的核定位信号肽（NLS）。体外研究表明，用γ-PGA包覆二元PPMS/DNA复合体可促进细胞对复合体颗粒的摄取，特别是γ-谷氨酰转肽酶（GGT）阳性细胞通过GGT介导的内吞途径对复合体颗粒的摄取。将PEG与γ-PGA偶联可形成三元PPMS/DNA/PGA-g-PEG复合物，复合物颗粒表面正电荷减少，在含血清的水介质中稳定性大大提高。通过将mTAT加入三元复合体系，细胞摄取率进一步提高。NLS肽的添加是为了促进质粒在细胞内传递到细胞核，这是基因转染过程中的一个限速步骤。结果表明，与二元PPMS/LucDNA复合物相比，新的mtat -四元PPMS/LucDNA/NLS/PGA-g-PEG-mTAT体系表现出更低的细胞毒性、更快的细胞摄取速率和更强的DNA向细胞核的运输。所有这些有利的功能都有助于mtat -四元复合物在体外和体内的显著基因转染效率，超过了二元复合物和商用Lipofectamine™2000/DNA脂质体。多功能mtat -四元复合物系统具有更高的效率和更低的细胞毒性，是一种有前途的新型非病毒载体，可用于传递治疗基因以治疗肿瘤。
We have developed a new multifunctional, non-viral gene delivery platform consisting of cationic poly(amine-co-ester) (PPMS) for DNA condensation, PEG shell for nanoparticle stabilization, poly(γ-glutamic acid) (γ-PGA) and mTAT (a cell-penetrating peptide) for accelerated cellular uptake, and a nuclear localization signal peptide (NLS) for enhanced intracellular transport of DNA to the nucleus. In vitro study showed that coating of the binary PPMS/DNA polyplex with γ-PGA promotes cellular uptake of the polyplex particles, particularly by γ-glutamyl transpeptidase (GGT)-positive cells through the GGT-mediated endocytosis pathway. Conjugating PEG to the γ-PGA led to the formation of a ternary PPMS/DNA/PGA-g-PEG polyplex with decreased positive charges on the surface of the polyplex particles and substantially higher stability in serum-containing aqueous medium. The cellular uptake rate was further improved by incorporating mTAT into the ternary polyplex system. Addition of the NLS peptide was designed to facilitate intracellular delivery of the plasmid to the nucleus--a rate-limiting step in the gene transfection process. As a result, compared with the binary PPMS/LucDNA polyplex, the new mTAT-quaternary PPMS/LucDNA/NLS/PGA-g-PEG-mTAT system exhibited reduced cytotoxicity, remarkably faster cellular uptake rate, and enhanced transport of DNA to the nucleus. All these advantageous functionalities contribute to the remarkable gene transfection efficiency of the mTAT-quaternary polyplex both in vitro and in vivo, which exceeds that of the binary polyplex and commercial Lipofectamine™ 2000/DNA lipoplex. The multifunctional mTAT-quaternary polyplex system with improved efficiency and reduced cytotoxicity represents a new type of promising non-viral vectors for the delivery of therapeutic genes to treat tumors.