Department of Pharmaceutics, School of Pharmaceutical Science, Shandong University, 44 West Culture Road, Ji'nan, China.
J Control Release. 2011 Apr 30;151(2):162-75. doi: 10.1016/j.jconrel.2011.02.027. Epub 2011 Mar 3.
The efficient delivery of therapeutic gene into cells of interest is a critical challenge to broad application of non-viral vectors. The approach of introducing ligands that lead gene vectors to target caveolae-mediated endocytosis on nanoparticle surface might serve as a promising strategy for the effective gene transfection. Recently, in an attempt to enhance the possibility of caveolae-mediated endocytosis, we fabricated a peptide-targeted gene vector for highly efficient receptor-mediated intracellular delivery. Cyclic Asn-Gly-Arg (cNGR) peptide was used to target gene loaded poly(lactic acid)-poly(ethylene glycol) nanoparticles (PLA-PEG NPs) to HUVEC over-expressing CD13. Using 6-lauroxyhexyl lysinate (LHLN) as cationic surfactant, cNGR modified PLA-PEG NPs (cNGR-PEG-PLA NPs) were capable of complexing and compacting DNA into homogeneous small-sized complexes (<200nm) with positive charge (~10mV). Fortunately, the results of in vitro cellular uptake tests and mechanism studies were consistent with our original hypothesis. The cNGR peptide presented on nanoparticles' surface could specifically mediate the fast and efficient internalization of cNGR-PEG-PLA NPs into HUVEC. Moreover, free cNGR inhibited their intracellular uptake into HUVEC revealing the mechanism of receptor-mediated endocytosis. Furthermore, the inspiring results of the mechanism studies and transfection assays demonstrated that caveolae-mediated endocytosis was indeed mainly involved in the internalization of cNGR-PEG-PLA NPs into HUVEC and led to significant gene transfection efficiency in contrast with cNGR non-modified PLA-PEG NPs. Given such encouraging and favorable properties including biocompatibility, high transfer efficiency, low cytotoxicity, and fast uptake by nondestructive endocytic pathways, cNGR-PEG-PLA NPs could be a promising carrier for the intracellular delivery of therapeutic agents.
将治疗基因有效递送至靶细胞是实现非病毒载体广泛应用的关键挑战。在纳米粒子表面引入配体引导基因载体靶向小窝蛋白介导的内吞作用的方法可能成为实现有效基因转染的有前途的策略。最近，为了增强小窝蛋白介导的内吞作用的可能性，我们构建了一种肽靶向基因载体，用于高效受体介导的细胞内递药。环天冬氨酰-甘氨酰-精氨酸（cNGR）肽被用于将负载基因的聚乳酸-聚乙二醇纳米粒（PLA-PEG NPs）靶向递送至过表达 CD13 的 HUVEC。使用 6-月桂酰基赖氨酸（LHLN）作为阳离子表面活性剂，cNGR 修饰的 PLA-PEG NPs（cNGR-PEG-PLA NPs）能够将 DNA 复合并压缩成具有正电荷（~10mV）的均匀小尺寸复合物（<200nm）。幸运的是，体外细胞摄取试验和机制研究的结果与我们的原始假设一致。纳米粒表面呈现的 cNGR 肽能够特异性介导 cNGR-PEG-PLA NPs 快速高效地内化进入 HUVEC。此外，游离的 cNGR 抑制了它们在 HUVEC 中的内化，揭示了受体介导的内吞作用的机制。此外，机制研究和转染试验的鼓舞人心的结果表明，小窝蛋白介导的内吞作用确实主要参与了 cNGR-PEG-PLA NPs 进入 HUVEC 的内化，并与 cNGR 非修饰的 PLA-PEG NPs 相比导致了显著的基因转染效率。鉴于其具有生物相容性、高转染效率、低细胞毒性和快速被非破坏性内吞途径摄取等令人鼓舞的特性，cNGR-PEG-PLA NPs 可能成为治疗药物细胞内递药的有前途的载体。
