Branched Polyethylenimine Derivatives with Reductively Cleavable Periphery for Safe and Efficient In Vitro Gene Transfer

作者：Wang, Yuhe; Zheng, Meng; Meng, Fenghua; Zhang, Jing; Peng, Rui; Zhong, Zhiyuan^*

来源：Biomacromolecules, 2011, 12(4): 1032-1040.

DOI：10.1021/bm101364f

• PLASMID DNA
• POLY(ETHYLENE IMINE)
• POLY(AMIDO AMINE)S
• DELIVERY
• POLYMERS
• DESIGN
• PEI
• NANOCARRIERS
• COPOLYMERS
• VECTOR

Twenty-five kDa polyethylenimine (PEI) is one of the most efficient nonviral gene transfer agents currently applied as a golden standard for in vitro transfection. In this study, novel 25 k Da PEI derivatives with reductively cleavable cystamine periphery (PEI-Cys) were designedto reduce carrier-associated cytotoxicity and to enhance further the transfection activity. The Michael-type conjugate addition of 25 kDa PEI with N-tert-butoxycarbonyl-N'-acryloyl-cystamine (Ac-Cys-Boc) and N-tert-butoxycarbonyl-a-methacryloyl-cystamine (MAc-Cys-Boc) followed by deprotection readily afforded PEI-Cys derivatives, denoted as PEI-(Cys)x(Ac) and PEI-(Cys)x(MAc), with degree of substitution (DS) ranging from 14 to 34 and 13 to 38, respectively. All PEI-Cys derivatives had higher buffer capacity than the parent 25 kDa PEI (21.2 to 23.1% versus 15.1%). Gel retardation and ethidium bromide exclusion assays showed that cystamine modification resulted in largely enhanced interactions with DNA. PEI-(Cys)x(Ac) could condense DNA into small-sized particles of 80-90 nm at and above am N/P ratio of 5/1, which were smaller than polyplexes of 25 kDa PEI (100-130 nm). in comparison, PEI-(Cys)x(MAc) condensed DNA into somewhat larger particles (100-180 nm at N/P ratios from 30/1 to 5/1). Gel retardation and dynamic light scattering (DLS) measurements showed that PEI-Cys polyplexes were quickly unpacked to release DNA in response to 10 mM dithiothreitol (DTT). These PEI-Cys derivatives revealed markedly decreased cytotoxicity as compared with 25 kDa PEI with IC50 values of > 100 mg/L and 50-75 mg/L for HeLa and 293T cells, respectively (corresponding IC50 data of 25 kDa PEI are ca. 11 and 3 mg/L). The in vitro transfection experiments in HeLa and 293T cells using pGL3 as a reporter gene showed that-gene transfection activity of PEI-Cys derivatives decreased with increasing DS and PEI-(Cys)x(MAc) exhibited higher transfection activity than PEI-(Cys)x(Ac) at similar IDS: Notably, polyplexes of PEI-(Cys) 14(Ac) and PEI-(Cys) 13(MAc) showed significantly enhanced gene transfection efficiency (up to 4.1-fold) as compared with 25 IcDa PEI formulation at an N/P ratio of 10/1 in both senum-free and 10% serum-containing conditions. The modification of PEI with reductively cleavable periphery appears to be a potential approach to develop safer and more efficient nonviral gene vectors.

• 出版日期2011-4
• 单位苏州大学