Nimesh Surendra, Chandra Ramesh
Dr. BR Ambedkar Center for Biomedical Research, University of Delhi, Delhi, India.
Eur J Pharm Biopharm. 2008 Mar;68(3):647-55. doi: 10.1016/j.ejpb.2007.09.008. Epub 2007 Sep 25.
Polyethylenimine (PEI) is one of the most efficient polycationic non-viral gene delivery vectors. Its efficiency and cytotoxicity depends on molecular weight, with the 25-kDa PEI being most efficient but accompanied with cytotoxicity. In the present study, enhancement in gene delivery efficiency along with reduction in cytotoxicity by attachment of guanidinium side group was explored. The hypothesis was that the guanidination would lead to the delocalization of charge present on primary amines of the polymer thereby leading to enhancement in gene delivery efficiency along with reduction in cytotoxicity. The polymer was guanidinated using O-methylisourea hemisulfate and the chemical linkage characterized by FTIR spectroscopy. The hydrodynamic diameter of guanidinated PEI-DNA complexes was determined using DLS. Subsequently, these complexes were used for DNA binding assay and zeta-potential measurements, taking native PEI as reference. Further, guanidinated PEI-DNA complexes were investigated for their gene delivery efficacy on HEK 293 cells. The hydrodynamic diameter of guanidinated PEI-DNA complexes was found to be in the range of 176-548 nm. As expected, the zeta potential values increased, on increasing the N/P ratios. It was found that guanidinated PEI had higher transfection efficiency at the majority of the N/P ratios tested as compared to commercially available transfecting agent lipofectin and native PEI itself. The toxicity of guanidinated PEI-DNA complexes was also reduced considerably in comparison to PEI polymer, as determined by MTT colorimetric assay. Out of the various derivatives prepared, gPEI 56% was found to be the most efficient in in vitro transfection.
聚乙烯亚胺（PEI）是最有效的聚阳离子非病毒基因传递载体之一。其效率和细胞毒性取决于分子量，25 kDa的PEI效率最高，但伴有细胞毒性。在本研究中，探索了通过连接胍基侧基提高基因传递效率并降低细胞毒性。假设是胍基化会导致聚合物伯胺上的电荷离域，从而提高基因传递效率并降低细胞毒性。使用半硫酸O-甲基异脲对聚合物进行胍基化，并通过傅里叶变换红外光谱对化学键进行表征。使用动态光散射法测定胍基化PEI-DNA复合物的流体动力学直径。随后，以天然PEI为参照，将这些复合物用于DNA结合测定和zeta电位测量。此外，研究了胍基化PEI-DNA复合物对HEK 293细胞的基因传递效果。发现胍基化PEI-DNA复合物的流体动力学直径在176 - 548 nm范围内。正如预期的那样，随着N/P比的增加，zeta电位值升高。结果发现，与市售转染试剂脂质体和天然PEI本身相比，在大多数测试的N/P比下，胍基化PEI具有更高的转染效率。通过MTT比色法测定，与PEI聚合物相比，胍基化PEI-DNA复合物的毒性也显著降低。在所制备的各种衍生物中，发现gPEI 56%在体外转染中效率最高。
