Development of a reduction-sensitive diselenide-conjugated oligoethylenimine nanoparticulate system as a gene carrier

Gang Cheng,^1,2 Yiyan He,¹ Li Xie,¹ Yu Nie,¹ Bin He,¹ Zhirong Zhang,² Zhongwei Gu<sup>1

1</sup>National Engineering Research Center for Biomaterials, ²Key Laboratory of Drug Targeting and Drug Delivery Systems, West China School of Pharmacy, Sichuan University, Chengdu, Sichuan, People's Republic of China

Background: The reduction-sensitive cationic polymer is a promising nonviral carrier for gene delivery. Until now, disulfide bonds have been the only golden standard for its design. The aim of this research was to develop a novel reduction-responsive cationic polymer as a gene carrier.
Methods: Polycationic carriers were synthesized by addition of branched oligoethylenimine 800 Da (OEI₈₀₀) via an active ester containing diselenide bonds. Disulfide bonds cross-linked with OEI₈₀₀-SS_x and monoselenide bonds linked with OEI₈₀₀-Se_x were synthesized and compared. Their molecular weights and degradation properties were determined using gel permeation chromatography. Changes in particle size, morphology, and DNA binding were investigated by dynamic light scattering, transmission electron microscopy, and electrophoresis assay in a reduction environment. Cytotoxicity and transfection in vitro were evaluated in a murine melanoma cell line (B16F10) and a human cervical epithelial carcinoma cell line (HeLa), while intracellular degradation and dissociation with DNA were studied by confocal laser scanning microscopy with FITC-labeled OEI₈₀₀ derivatives and Cy5-labeled DNA.
Results: Diselenide-conjugated OEI₈₀₀ (OEI₈₀₀-Se Se_x) polymer carriers of high molecular weight were successfully synthesized. After compacting with DNA, the OEI₈₀₀-SeSe_x polymers formed nanoparticles with an average size of 140 nm at an adequate C/P ratio. OEI₈₀₀-Se Se_x showed reduction-responsive degradation properties similar to those of the OEI₈₀₀-SS_x via gel permeation chromatography, dynamic light scattering, and transmission electron microscopy. OEI₈₀₀-SeSe_x showed much lower cytotoxicity than PEI_25k, and significantly higher transfection efficiency than OEI₈₀₀ in both B16F10 and HeLa cells. Transfection of luciferase in the OEI₈₀₀-SeSe_x group was comparable with that of standard PEI_25k and traditional reduction-sensitive polymer OEI₈₀₀-SS_x groups. Furthermore, intracellular degradation of OEI₈₀₀-SeSe_x and dissociation with DNA were also confirmed by confocal laser scanning microscopy.
Conclusion: The OEI₈₀₀-SeSe_x obtained was able to bind plasmid DNA efficiently to yield nanosized particles and had reduction sensitivity which is as efficient as that for OEI₈₀₀-SS_x. In vitro experiments confirmed its low cytotoxicity and high transfection ability. Diselenide bonds can be used as effective and novel reduction-sensitive linkages for gene delivery.

Keywords: diselenide, oligoethylenimine, reduction-sensitive, gene carriers