Department of Biosciences & Bioengineering, Indian Institute of Technology Bombay, Mumbai, Maharashtra 400076, India.
Biomater Sci. 2018 Nov 20;6(12):3241-3250. doi: 10.1039/c8bm00916c.
For diseases related to genetic disorders or cancer, many cellular therapies rely on the ex vivo modification of cells for attaining a desired therapeutic effect. The efficacy of such therapies involving the genetic modification of cells relies on the extent of gene expression and subsequent persistence of modified cells when infused into the patient's body. In situ gene delivery implies the manipulation of cells in their in vivo niche such that the effectiveness can be improved by minimizing post manipulation effects like cell death, lack of persistence, etc. Furthermore, material-based in situ localized gene delivery can reduce the undesired side effects caused by systemic modifications. Here, we have used polyethylene (glycol) diacrylate (PEGDA) based cryogels to genetically modify cells in vivo with a focus on immunotherapy. PEGDA cryogels were either blended with gelatin methacrylate (GELMA) or surface modified with poly-l-lysine (PLL) in order to improve cell adhesion and/or retain viruses for localized gene delivery. On using the lentiviruses encoding gene for green fluorescent protein (GFP) in in vitro experiments, we found higher transduction efficiency in HEK 293FT cells via PEGDA modified with poly-l-lysine (PEGDA-PLL) and PEGDA-GELMA cryogels compared to PEGDA cryogels. In vitro release experiments showed improved retention of GFP lentiviruses in PEGDA-PLL cryogels, which were then employed for in vivo gene delivery and were demonstrated to perform better than the corresponding bolus delivery of lentiviruses through an injection. Both physical and biological characterization studies of these cryogels show that this material platform can be used for gene delivery as well as other tissue engineering applications.
对于与遗传疾病或癌症相关的疾病，许多细胞疗法依赖于细胞的体外修饰以达到预期的治疗效果。涉及细胞遗传修饰的此类疗法的疗效取决于基因表达的程度以及将修饰后的细胞注入患者体内后的后续持久性。原位基因传递意味着在体内生态位中对细胞进行操作，从而通过最小化操作后效果（如细胞死亡、缺乏持久性等）来提高有效性。此外，基于材料的原位局部基因传递可以减少由全身修饰引起的不必要的副作用。在这里，我们使用聚乙二醇（二醇）二丙烯酸酯（PEGDA）基冷冻凝胶在体内对细胞进行基因修饰，重点是免疫疗法。PEGDA 冷冻凝胶与明胶甲基丙烯酰胺（GELMA）混合或用聚-l-赖氨酸（PLL）表面改性，以提高细胞黏附性和/或保留病毒以进行局部基因传递。在体外实验中使用编码绿色荧光蛋白（GFP）基因的慢病毒，我们发现通过用聚-l-赖氨酸（PEGDA-PLL）和 PEGDA-GELMA 冷冻凝胶修饰的 PEGDA 获得了更高的 HEK 293FT 细胞转导效率，与 PEGDA 冷冻凝胶相比。体外释放实验表明 GFP 慢病毒在 PEGDA-PLL 冷冻凝胶中的保留得到了改善，然后将其用于体内基因传递，并证明其性能优于通过注射进行相应的慢病毒推注。对这些冷冻凝胶的物理和生物学特性研究表明，这种材料平台可用于基因传递以及其他组织工程应用。
