Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, G4 0RE, UK.
Cell Analysis Facility, Medical and Veterinary & Life Sciences Shared Research Facilities, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, G12 8QQ, UK.
Int J Nanomedicine. 2024 Nov 21;19:12189-12203. doi: 10.2147/IJN.S490936. eCollection 2024.
Gene therapy is a promising therapeutic approach for treating various disorders by introducing modified nucleic acids to correct cellular dysfunctions or introduce new functions. Despite significant advancements in the field, the effective delivery of nucleic acids remains a challenge, due to biological barriers and the immune system's ability to target and destroy these molecules. Due to their branched structure and ability to condense negatively charged nucleic acids, cationic dendrimers have shown potential in overcoming these challenges. Despite this, standardized scalable production methods are still lacking. This study investigates the use of microfluidics to formulate generation 3-diaminobutyric polypropylenimine (DAB) dendriplexes and compares their characteristics and in vitro gene delivery efficacy to those prepared using conventional manual mixing.
DAB dendriplexes were produced by both microfluidic and manual approaches and characterized. Their cellular uptake and gene expression were evaluated on C6 glioma cancer cells in vitro.
Dendriplexes formed using microfluidics at the optimal flow rate and ratio demonstrated enhanced DNA condensation over time, achieving up to 97% condensation at 24 hours. Both preparation methods produced positively charged dendriplexes, indicating stable formulations. However, dendriplexes prepared through hand mixing resulted in smaller particle sizes, significantly higher cellular uptake and gene expression efficacy compared to those prepared by microfluidics. Nonetheless, microfluidic preparation offers the advantage of standardized and scalable production, which is essential for future applications.
This study highlights the potential of microfluidic technology to improve precision and scalability in gene delivery, paving the way for future advancements in gene therapy. Our findings suggest that, with further optimization, microfluidic systems could provide superior control over dendriplex formation, expanding their potential use in gene therapy applications.
基因治疗是一种有前途的治疗方法，通过引入修饰的核酸来纠正细胞功能障碍或引入新的功能，从而治疗各种疾病。尽管该领域取得了重大进展，但由于生物屏障和免疫系统靶向和破坏这些分子的能力，核酸的有效传递仍然是一个挑战。由于其分支结构和凝聚带负电荷的核酸的能力，阳离子树状聚合物在克服这些挑战方面显示出了潜力。尽管如此，标准化的可扩展生产方法仍然缺乏。本研究调查了使用微流控技术来配方第三代二氨基丁基聚丙亚胺（DAB）树枝状聚合物，并且将其特性和体外基因传递效率与使用常规手动混合制备的进行了比较。
通过微流控和手动方法制备 DAB 树枝状聚合物，并对其进行了表征。在体外评估了它们在 C6 神经胶质瘤癌细胞中的细胞摄取和基因表达。
在最佳流速和比例下使用微流控形成的树枝状聚合物随时间显示出增强的 DNA 凝聚，在 24 小时内达到高达 97%的凝聚。两种制备方法都产生带正电荷的树枝状聚合物，表明是稳定的配方。然而，与通过微流控制备的相比，通过手动混合制备的树枝状聚合物具有更小的粒径、更高的细胞摄取和基因表达效率。尽管如此，微流控制备具有标准化和可扩展生产的优势，这对于未来的应用至关重要。
本研究强调了微流控技术在基因传递中提高精度和可扩展性的潜力，为基因治疗的未来发展铺平了道路。我们的研究结果表明，通过进一步优化，微流控系统可以更好地控制树枝状聚合物的形成，从而扩大其在基因治疗应用中的潜力。
