Wu Yang, Qin Zhu, Ji Jinkai, Yang Ran, Zhang Xiaoqiang, Li Yuanhui, Yin Lihong, Pu Yuepu, Li Xingsong
Biomaterials and Drug Delivery Laboratories, School of Chemistry and Chemical Engineering, Southeast University , Nanjing , P. R. China .
J Drug Target. 2014 May;22(4):352-64. doi: 10.3109/1061186X.2013.877466. Epub 2014 Jan 9.
In this report, galactosylated poly(ethylene glycol) methacrylate-st-3-guanidinopropyl methacrylamide copolymers (galactosylated PEGMA-st-GPMA, GGP) are developed as siRNA carriers to inhibit Survivin mRNA expression. GGPs are combined with Survivin siRNAs to form siRNA/GGP polyplexes. The polyplexes particles were examined by a dynamic light scattering. It showed that GGP copolymers could condense siRNA to form particles with diameter from 128 to 423 nm and zeta potential value in the range from +2.4 to +14.9 mV at various charge ratios (N/P). The MTT assay data of siRNA/GGP polyplexes on human hepatocellular liver carcinoma cells (HepG2) and human cervix epithelial carcinoma cells (HeLa) indicated that GGP copolymer had better cell viabilities than polyethyleimine (PEI). The transfection of siRNA/GGP polyplexes was detected by real-time quantitative PCR (RT-qPCR) in HepG2 cell line. We found that the siRNA/GGP polyplexes could effectively silence Survivin mRNA expression in the serum-free media (p < 0.01). In the presence of 10% serum medium, the Survivin mRNA expressed has significant difference between siRNA/GGP polyplexes and blank (p < 0.05). The galactose competition assay showed that galactosylated PEGMA-st-GPMA (GGP) may provide the targeting to HepG2 cells mediating by asialoglycoproteins receptors (ASGP-R). Furthermore, Survivin siRNA/GGP polyplexes could significantly (p < 0.01) inhibit both HepG2 tumor growth and Survivin protein expression in vivo studies in a xenograft mouse model.
在本报告中，开发了半乳糖基化聚甲基丙烯酸乙二醇酯-苯乙烯-3-胍基丙基甲基丙烯酰胺共聚物（半乳糖基化聚乙二醇甲基丙烯酸酯-苯乙烯-胍基丙基甲基丙烯酰胺，GGP）作为小干扰RNA（siRNA）载体，以抑制生存素mRNA表达。GGP与生存素siRNA结合形成siRNA/GGP多聚体。通过动态光散射检测多聚体颗粒。结果表明，在不同电荷比（N/P）下，GGP共聚物可使siRNA凝聚形成直径为128至423 nm、ζ电位值在+2.4至+14.9 mV范围内的颗粒。siRNA/GGP多聚体对人肝癌细胞（HepG2）和人宫颈上皮癌细胞（HeLa）的MTT检测数据表明，GGP共聚物比聚乙烯亚胺（PEI）具有更好的细胞活力。在HepG2细胞系中通过实时定量PCR（RT-qPCR）检测siRNA/GGP多聚体的转染情况。我们发现，siRNA/GGP多聚体在无血清培养基中可有效沉默生存素mRNA表达（p < 0.01）。在含有10%血清的培养基中，siRNA/GGP多聚体与空白组之间生存素mRNA表达存在显著差异（p < 0.05）。半乳糖竞争试验表明，半乳糖基化聚甲基丙烯酸乙二醇酯-苯乙烯-胍基丙基甲基丙烯酰胺（GGP）可能通过去唾液酸糖蛋白受体（ASGP-R）介导对HepG2细胞的靶向作用。此外，在异种移植小鼠模型的体内研究中，生存素siRNA/GGP多聚体可显著（p < 0.01）抑制HepG2肿瘤生长和生存素蛋白表达。
