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Please use this identifier to cite or link to this item: http://hdl.handle.net/10119/15875

Title: Freezing Assisted Gene Delivery Combined with Polyampholyte Nanocarriers
Authors: Ahmed, Sana
Nakaji-Hirabayashi, Tadashi
Watanabe, Takayoshi
Hohsaka, Takahiro
Matsumura, Kazuaki
Keywords: gene delivery
freeze concentration
polyampholytes
nanoparticles
endosome escape
Issue Date: 2017-06-25
Publisher: American Chemical Society
Magazine name: ACS Biomaterials Science and Engineering
Volume: 3
Number: 8
Start page: 1677
End page: 1689
DOI: 10.1021/acsbiomaterials.7b00176
Abstract: Physical methodologies such as electroporation and the gene-gun technology have been widely used for transfection; however, their applicability is limited because they lead to cell damage and low cell viability. Therefore, to address these limitations we developed a new freeze concentration-based gene transfection system that provides enhanced in vitro gene delivery compared to that provided by the commercially available systems. The system employs a facile freeze concentration step, whereby cells are simply frozen to very low temperatures in the presence of polymer-pDNA complexes. As part of system development, we also synthesized a low toxicity polyethyleneimine (PEI)-based polyampholyte prepared through succinylation with butylsuccinic anhydride. In aqueous solution, this modified polyampholyte self-assembles to form small (20 nm diameter), positively charged (net surface charge of 35 mV), nanoparticles through a combination of hydrophobic and electrostatic interactions. Agarose gel electrophoresis analysis indicated that the polyampholyte nanoparticle was able to form a complex with pDNA that provided stability against nuclease degradation. Using transfection of HEK-293T cells, we demonstrated that a combination of polyampholyte: pDNA, at an appropriate ratio, and the freeze concentration method resulted in significant enhancement of GFP and luciferase expression compared to commercially available carriers. Endosomal escape of pDNA was also found to be increased when using the modified polyampholyte compared to branched PEI. This study suggests that the efficient combination of freeze concentration and the modified polyampholyte described here has great potential for in vitro gene therapy.
Rights: Sana Ahmed, Tadashi Nakaji-Hirabayashi, Takayoshi Watanabe, Takahiro Hohsaka, Kazuaki Matsumura, ACS Biomaterials Science and Engineering, 2017, 3(8), pp.1677-1689. This document is the author's version of a work that was subsequently accepted for publication in ACS Biomaterials Science and Engineering, copyright (c) American Chemical Society after peer review. To access the final edited and published work, see http://dx.doi.org/10.1021/acsbiomaterials.7b00176
URI: http://hdl.handle.net/10119/15875
Material Type: author
Appears in Collections:c10-1. 雑誌掲載論文 (Journal Articles)

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