Impact of PEG Chain Length on the Physical Properties and Bioactivity of PEGylated Chitosan/siRNA Nanoparticles in Vitro and in Vivo.
PEGylation of cationic polyplexes is a promising approach to enhance the stability and reduce unspecific interaction with biological components. Herein, we systematically investigate the impact of PEGylation on physical and biological properties of chitosan/siRNA polyplexes. A series of chitosan-PEG copolymers (CS-PEG2k, CS-PEG5k and CS-PEG10k) were synthesized with similar PEG mass content but with different molecular weight. PEGylation with higher molecular weight and less grafting degree resulted in smaller and more compacted nanoparticles with relatively higher surface charge. PEGylated polyplexes showed distinct mechanism of endocytosis, which was macropinocytosis and caveolae-dependent and clathrin-independent. In vitro silencing efficiency in HeLa and H1299 cells was significantly improved by PEGylation and CS-PEG5k/siRNA achieved the highest knockdown efficiency. Efficient silence of ribonucleotide reductase subunit M2 (RRM2) in HeLa cells by CS-PEG5k/siRRM2 significantly induced cell cycle arrest and inhibited cell proliferation. In addition, PEGylation significantly inhibited macrophage phagocytosis and unspecific interaction with red blood cells (RBCs). Significant extension of in vivo circulation was achieved only with high molecular weight PEG modification (CS-PEG10k), whereas all CS/siRNA and CS-PEG/siRNA nanoparticles showed similar pattern of biodistribution with major accumulation in liver and kidney. These results imply that PEGylation with higher molecular weight PEG and less grafting rate is a promising strategy to improve chitosan/siRNA nanocomplexes performance both in vitro and in vivo.
Interdisciplinary Nanoscience Center (iNANO), Aarhus University , DK-8000 Aarhus C, Denmark.|Department of Molecular Biology and Genetics, Aarhus University , DK-8000 Aarhus C, Denmark.
Chuanxu
http://orcid.org/0000-0003-0884-6943
C
Yang
Interdisciplinary Nanoscience Center (iNANO), Aarhus University , DK-8000 Aarhus C, Denmark.|Department of Molecular Biology and Genetics, Aarhus University , DK-8000 Aarhus C, Denmark.|Suzhou Ribo Life Science Co., Ltd. , Beijing, China.
Shan
S
Gao
Department of Biomedicine, Aarhus University , Bartholin Building Building 1240, Wilhelm Meyers Alle 4,8000 Aarhus C, Denmark.
Frederik
F
Dagnæs-Hansen
Interdisciplinary Nanoscience Center (iNANO), Aarhus University , DK-8000 Aarhus C, Denmark.|Department of Molecular Biology and Genetics, Aarhus University , DK-8000 Aarhus C, Denmark.
Maria
M
Jakobsen
Interdisciplinary Nanoscience Center (iNANO), Aarhus University , DK-8000 Aarhus C, Denmark.|Department of Molecular Biology and Genetics, Aarhus University , DK-8000 Aarhus C, Denmark.
Jørgen
J
Kjems
D045744:Cell Line, Tumor; D048271:Chitosan; D006801:Humans; D053758:Nanoparticles; D011092:Polyethylene Glycols; D034741:RNA, Small Interfering; D014018:Tissue Distribution
D034741:RNA, Small Interfering; D011092:Polyethylene Glycols; D048271:Chitosan
PEGylation; biodistribution; blood circulation; chitosan; nanoparticles; siRNA delivery