Baral Bineeth, Panigrahi Bijayananda, Kar Avishek, Tulsiyan Kiran D, Suryakant Uday, Mandal Dindyal, Subudhi Umakanta
DNA Nanotechnology & Application Laboratory, Environment & Sustainability Department, CSIR-Institute of Minerals & Materials Technology, Bhubaneswar 751013, Odisha, India.
Academy of Scientific & Innovative Research (AcSIR), Ghaziabad 201002, India.
Mol Ther Nucleic Acids. 2023 Jul 18;33:493-510. doi: 10.1016/j.omtn.2023.07.017. eCollection 2023 Sep 12.
Self-assembled branched DNA (bDNA) nanomaterials have exhibited their functionality in various biomedical and diagnostic applications. However, the anionic cellular membrane has restricted the movement of bDNA nanostructures. Recently, amphiphilic peptides have been investigated as cationic delivery agents for nucleic acids. Herein, we demonstrate a strategy for delivering functional bDNA nanomaterials into mammalian cells using self-assembled linear peptides. In this study, antisense oligonucleotides of vascular endothelial growth factor (VEGF) were inserted in the overhangs of bDNAs. Novel linear peptides have been synthesized and the peptide-bound bDNA complex formation was examined using various biophysical experiments. Interestingly, the W4R4-bound bDNAs were found to be exceptionally stable against DNase I compared to other complexes. The delivery of fluorescent-labeled bDNAs into the mammalian cells confirmed the potential of peptide transporters. Furthermore, the functional efficacy of the peptide-bound bDNAs has been examined through RT-PCR and western blot analysis. The observed results revealed that W4R4 peptides exhibited excellent internalization of antisense bDNAs and significantly suppressed (3- to 4-fold) the transcripts and translated product of VEGF compared to the control. In summary, the results highlight the potential use of peptide-based nanocarrier for delivering bDNA nanostructures to regulate the gene expression in cell lines.
自组装分支DNA（bDNA）纳米材料已在各种生物医学和诊断应用中展现出其功能。然而，阴离子细胞膜限制了bDNA纳米结构的移动。最近，两亲性肽已被研究作为核酸的阳离子递送剂。在此，我们展示了一种使用自组装线性肽将功能性bDNA纳米材料递送至哺乳动物细胞的策略。在本研究中，血管内皮生长因子（VEGF）的反义寡核苷酸被插入到bDNA的突出端。合成了新型线性肽，并使用各种生物物理实验检测了肽结合的bDNA复合物的形成。有趣的是，与其他复合物相比，发现W4R4结合的bDNA对DNase I具有异常的稳定性。将荧光标记的bDNA递送至哺乳动物细胞证实了肽转运体的潜力。此外，通过RT-PCR和蛋白质印迹分析检测了肽结合的bDNA的功能效力。观察结果表明，与对照相比，W4R4肽表现出反义bDNA的优异内化，并显著抑制（3至4倍）VEGF的转录本和翻译产物。总之，结果突出了基于肽的纳米载体在递送bDNA纳米结构以调节细胞系中基因表达方面的潜在用途。
