Bogacheva Mariia, Egorova Anna, Slita Anna, Maretina Marianna, Baranov Vladislav, Kiselev Anton
D.O. Ott Research Institute of Obstetrics, Gynecology and Reproductology, Mendeleevskaya line, 3, Saint-Petersburg 199034, Russia.
D.O. Ott Research Institute of Obstetrics, Gynecology and Reproductology, Mendeleevskaya line, 3, Saint-Petersburg 199034, Russia.
Bioorg Med Chem Lett. 2017 Nov 1;27(21):4781-4785. doi: 10.1016/j.bmcl.2017.10.001. Epub 2017 Oct 3.
The major barriers for intracellular DNA transportation by cationic polymers are their toxicity, poor endosomal escape and inefficient nuclear uptake. Therefore, we designed novel modular peptide-based carriers modified with SV40 nuclear localization signal (NLS). Core peptide consists of arginine, histidine and cysteine residues for DNA condensation, endosomal escape promotion and interpeptide cross-linking, respectively. We investigated three polyplexes with different NLS content (10 mol%, 50 mol% and 90 mol% of SV40 NLS) as vectors for intranuclear DNA delivery. All carriers tested were able to condense DNA, to protect it from DNAase I and were not toxic to the cells. We observed that cell cycle arrest by hydroxyurea did not affect transfection efficacy of NLS-modified carriers which we confirmed using quantitative confocal microscopy analysis. Overall, peptide carrier modified with 90 mol% of SV40 NLS provided efficient transfection and nuclear uptake in non-dividing cells. Thus, incorporation of NLS into arginine-rich cross-linking peptides is an adequate approach to the development of efficient intranuclear gene delivery vehicles.
阳离子聚合物介导细胞内DNA转运的主要障碍在于其毒性、较差的内体逃逸能力和低效的核摄取。因此，我们设计了用SV40核定位信号（NLS）修饰的新型模块化肽基载体。核心肽分别由精氨酸、组氨酸和半胱氨酸残基组成，用于DNA凝聚、促进内体逃逸和肽间交联。我们研究了三种具有不同NLS含量（SV40 NLS的10摩尔%、50摩尔%和90摩尔%）的多聚体作为核内DNA递送载体。所有测试的载体都能够凝聚DNA，保护其免受DNA酶I的降解，并且对细胞无毒。我们观察到羟基脲引起的细胞周期停滞并不影响NLS修饰载体的转染效率，我们通过定量共聚焦显微镜分析证实了这一点。总体而言，用90摩尔%的SV40 NLS修饰的肽载体在非分裂细胞中提供了高效的转染和核摄取。因此，将NLS掺入富含精氨酸的交联肽中是开发高效核内基因递送载体的一种适当方法。
