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Updated: Aug 18, 2026
Enhanced Gene Delivery and Expression using Intraosseous Injection of Chitosan Nanoparticles Encapsulated Adenine Base Editor Plasmids
Published on: May 16, 2025
Targeting delivery of oligonucleotide and plasmid DNA to hepatocyte via galactosylated chitosan vector
Shuying Gao1, Jiangning Chen, Lei Dong
1Department of Biochemistry, State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University, Nanjing, China.
Abstract:
Delivery of oligonucleotide to specific cells and maintenance of its biological function are important for nucleic acid therapy. The objective of this paper is to demonstrate that galactosylated low molecular weight chitosan (gal-LMWC) is a safe and effective vector of antisense oligonucleotide (ASO) and plasmid DNA for the hepatocyte targeting delivery. Gal-LMWC has been successfully prepared and MTT cytotoxic assay shows that cytotoxicity of gal-LMWC is lower than that of high molecular weight chitosan (HMWC) and low molecular weight chitosan (LMWC) in HepG2 cells. Using a complex coacervation process, gal-LMWC can form stable nano-complexes with plasmid DNA or with ASO by the electrostatic interaction. The morphometrics, particle size, and the zeta potential of gal-LMWC/ASO complexes and gal-LMWC/plasmid DNA complexes are very similar. The transfection efficiency by using gal-LMWC vector is significantly higher than that of naked DNA or naked ASO in HepG2 cells. Transfection efficiency of gal-LMWC/ASO complexes and gal-LMWC/plasmid DNA complexes depends on the molar ratio of the positive chitosan amino group and the negative DNA phosphate group (N/P ratio) strongly. Inhibition experiments confirm that the enhanced transfection efficiency is due to the ASGR mediated endocytosis of the gal-LMWC/ASO complexes or gal-LMWC/DNA complexes. These results suggest that gal-LMWC can be used in gene therapy to improve the transfection efficiency in vitro and in vivo.
Insights
Galactosylated low molecular weight chitosan (gal-LMWC) effectively delivers nucleic acids to liver cells. This safe and efficient vector enhances gene therapy by improving transfection efficiency.
Area of Science:
- Biomaterials Science
- Gene Therapy
- Nanotechnology
Background:
- Effective delivery of nucleic acids like antisense oligonucleotides (ASO) and plasmid DNA to target cells is crucial for nucleic acid therapy.
- Chitosan-based vectors offer potential for gene delivery, but optimizing their safety and targeting efficiency remains a challenge.
Purpose of the Study:
- To evaluate galactosylated low molecular weight chitosan (gal-LMWC) as a safe and effective vector for hepatocyte-targeted delivery of ASO and plasmid DNA.
- To investigate the characteristics and transfection efficiency of gal-LMWC/nucleic acid complexes.
Main Methods:
- Preparation and characterization of gal-LMWC.
- MTT cytotoxicity assay in HepG2 cells to assess safety.
- Complex coacervation to form gal-LMWC/ASO and gal-LMWC/plasmid DNA nano-complexes.
- Analysis of complex morphometrics, particle size, and zeta potential.
- Transfection efficiency assessment in HepG2 cells.
- Inhibition studies to confirm the mechanism of cellular uptake.
Main Results:
- Successfully prepared gal-LMWC with lower cytotoxicity than high molecular weight chitosan (HMWC) and low molecular weight chitosan (LMWC) in HepG2 cells.
- Gal-LMWC formed stable nano-complexes with ASO and plasmid DNA, exhibiting similar morphometrics, particle size, and zeta potential.
- Transfection efficiency of gal-LMWC complexes was significantly higher than naked DNA or ASO in HepG2 cells.
- Transfection efficiency was strongly dependent on the N/P ratio.
- Inhibition experiments confirmed ASGPR-mediated endocytosis as the mechanism for enhanced cellular uptake.
Conclusions:
- Gal-LMWC is a safe and effective vector for targeted delivery of ASO and plasmid DNA to hepatocytes.
- Gal-LMWC-based gene delivery systems show promise for improving transfection efficiency in vitro and in vivo applications.
- The ASGPR-mediated endocytosis pathway highlights the potential of gal-LMWC for targeted gene therapy.
