Department of Chemical and Biomolecular Engineering, Yonsei University, Seoul, Korea.
Gene Ther. 2012 Mar;19(3):329-37. doi: 10.1038/gt.2011.84. Epub 2011 Jun 9.
The successful development of efficient and safe gene delivery vectors continues to be a major obstacle to gene delivery in stem cells. In this study, we have developed an elastin-like polypeptide (ELP)-mediated adeno-associated virus (AAV) delivery system for transducing fibroblasts and human neural stem cells (hNSCs). AAVs have significant promise as therapeutic vectors because of their safety and potential for use in gene targeting in stem cell research. ELP has been recently employed as a biologically inspired 'smart' biomaterial that exhibits an inverse temperature phase transition, thereby demonstrating promise as a novel drug carrier. The ELP that was investigated in this study was composed of a repetitive penta-peptide with [Val-Pro-Gly-Val-Gly]. A novel AAV variant, AAV r3.45, which was previously engineered by directed evolution to enhance transduction in rat NSCs, was nonspecifically immobilized onto ELPs that were adsorbed beforehand on a tissue culture polystyrene surface (TCPS). The presence of different ELP quantities on the TCPS led to variations in surface morphology, roughness and wettability, which were ultimately key factors in the modulation of cellular transduction. Importantly, with substantially reduced viral quantities compared with bolus delivery, ELP-mediated AAV delivery significantly enhanced delivery efficiency in fibroblasts and hNSCs, which have great potential for use in tissue engineering applications and neurodegenerative disorder treatments, respectively. The enhancement of cellular transduction in stem cells, as well as the feasibility of ELPs for utilization in three-dimensional scaffolds, will contribute to the advancement of gene therapy for stem cell research and tissue regenerative medicine.
高效且安全的基因传递载体的成功开发仍然是基因传递到干细胞中的主要障碍。在这项研究中，我们开发了一种弹性蛋白样多肽（ELP）介导的腺相关病毒（AAV）传递系统，用于转导成纤维细胞和人神经干细胞（hNSC）。AAV 作为治疗性载体具有很大的潜力，因为它们具有安全性和在干细胞研究中用于基因靶向的潜力。ELP 最近被用作一种受生物启发的“智能”生物材料，具有逆温相变特性，因此有望成为新型药物载体。本研究中研究的 ELP 由重复的五肽[Val-Pro-Gly-Val-Gly]组成。一种新型 AAV 变体 AAV r3.45 先前通过定向进化工程设计以增强大鼠 NSCs 的转导效率，非特异性地固定在先前吸附在组织培养聚苯乙烯表面（TCPS）上的 ELPs 上。TCPS 上不同 ELP 数量的存在导致表面形态、粗糙度和润湿性的变化，这些最终是调节细胞转导的关键因素。重要的是，与单次注射相比，ELP 介导的 AAV 传递大大减少了病毒数量，从而显著提高了成纤维细胞和 hNSC 的传递效率，这对于组织工程应用和神经退行性疾病治疗分别具有很大的应用潜力。ELP 介导的 AAV 传递可增强干细胞中的细胞转导效率，并且 ELP 适用于三维支架，这将有助于推进干细胞研究和组织再生医学中的基因治疗。
