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This study developed a novel dendrimer-like carrier for plasmid DNA delivery. Functionalization with polyethylene glycol (PEG) and specific ligands enhanced cellular uptake and gene transfer efficacy via receptor-mediated endocytosis.

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Area of Science:

  • Biomaterials Science
  • Gene Delivery Systems
  • Nanotechnology

Background:

  • Developing efficient and targeted non-viral gene delivery vectors is crucial for gene therapy.
  • Oligo(aminoamide) dendrimers offer potential as DNA carriers due to their cationic nature and tunable structure.
  • Surface modification strategies are essential to overcome challenges like shielding and targeted cellular uptake.

Purpose of the Study:

  • To synthesize and characterize a novel cationic dendrimer-like carrier for plasmid DNA delivery.
  • To investigate the impact of polyethylene glycol (PEG) shielding and ligand functionalization on carrier performance.
  • To evaluate the targeted cellular uptake and gene transfection efficacy of modified carriers mediated by specific receptors.

Main Methods:

  • Synthesis of a four-arm oligo(aminoamide) dendrimer functionalized with azido-lysines.
  • Copper-free click chemistry for attaching polyethylene glycol (PEG) chains and/or GalNAc or mannose ligands.
  • Formation of polyplexes with plasmid DNA and characterization of nanoparticle properties (size, stability).
  • In vitro assessment of cellular uptake and gene transfection efficiency in various cell lines (HepG2, Huh7, DC2.4).

Main Results:

  • Prefunctionalization of the carrier with PEG and ligands before polyplex formation was superior to postfunctionalization.
  • Optimal shielding was achieved with 24 ethylene oxide units of PEG and ≥50% azide functionalization.
  • PEG shielding reduced DNA transfer, but this was restored by GalNAc or mannose ligand functionalization.
  • GalNAc-functionalized polyplexes showed ASGPR-mediated uptake in HepG2 and Huh7 cells.
  • Mannose-functionalized polyplexes demonstrated enhanced cellular uptake and transfection in mannose receptor-expressing DC2.4 cells.

Conclusions:

  • The developed dendrimer-like carrier system allows for versatile surface modification via click chemistry.
  • Targeted functionalization with GalNAc and mannose ligands can overcome PEG-induced shielding and enhance receptor-mediated endocytosis.
  • This strategy enables efficient gene delivery to specific cell types expressing ASGPR or mannose receptors.