Developing non-viral gene therapy vectors that both protect and functionally deliver nucleic acid cargoes will be vital if gene augmentation and editing strategies are to be effectively combined with advanced regenerative medicine approaches. Currently such methodologies utilize high concentrations of recombinant growth factors, which result in toxicity and off-target effects. Herein we demonstrate the use of modified cell penetrating peptides (CPPs), termed Glycosaminoglycan (GAG)-binding Enhanced Transduction (GET) peptides with plasmid DNA (pDNA) encapsulated poly (lactic-co-glycolic acid) PLGA nanoparticles (pDNA-encapsulated PLGA NPs). In order to encapsulate the pDNA, it was first condensed with a cationic low molecular weight Poly L-Lysine (PLL) into 30–60 nm NPs followed by encapsulation in PLGA NPs by double emulsion; yielding encapsulation efficiencies (EE) of ∼30%. PLGA NPs complexed with GET peptides show enhanced intracellular delivery (up to sevenfold) and transfection efficiencies (up to five orders of magnitude). Moreover, the pDNA cargo has enhanced protection from nucleases (such as DNase I) promoting their translatability. As an example, we show these NPs efficiently deliver pBMP2 which can promote osteogenic differentiation in vitro. Gene delivery to human Mesenchymal Stromal Cells (hMSCs) inducing their osteogenic programming was confirmed by Alizarin red calcium staining and bone lineage specific gene expression (Q RT-PCR). By combining simplistic and FDA-approved PLGA polymer nanotechnology with the GET delivery system, therapeutic non-viral vectors could have significant impact in future cellular therapy and regenerative medicine applications.

中文翻译：

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有效传递转导聚合物纳米颗粒，用于基因介导的成骨诱导骨再生

如果基因增强和编辑策略要与先进的再生医学方法有效结合，开发既能保护又可在功能上传递核酸货物的非病毒基因治疗载体将是至关重要的。目前，此类方法利用高浓度的重组生长因子，这会导致毒性和脱靶效应。在此，我们展示了使用修饰的细胞穿透肽 (CPP)，称为糖胺聚糖 (GAG) 结合增强转导 (GET) 肽与质粒 DNA (pDNA) 封装的聚（乳酸-乙醇酸）PLGA 纳米颗粒（pDNA 封装的 PLGA NP）。为了封装 pDNA，首先将其与阳离子低分子量聚 L-赖氨酸 (PLL) 缩合成 30-60 nm 的 NPs，然后通过双乳液封装在 PLGA NPs 中；产生约 30% 的封装效率 (EE)。与 GET 肽复合的 PLGA NPs 显示出增强的细胞内递送（高达七倍）和转染效率（高达五个数量级）。此外，pDNA 货物增强了对核酸酶（如 DNase I）的保护，促进了它们的可翻译性。作为一个例子，我们展示了这些 NPs 有效地传递 pBMP2，它可以在体外促进成骨分化。茜素红钙染色和骨谱系特异性基因表达 (Q RT-PCR) 证实了向人类间充质基质细胞 (hMSC) 传递基因以诱导其成骨编程。通过将简单且经 FDA 批准的 PLGA 聚合物纳米技术与 GET 递送系统相结合，治疗性非病毒载体可能对未来的细胞治疗和再生医学应用产生重大影响。