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Evaluation of Polymeric Gene Delivery Nanoparticles by Nanoparticle Tracking Analysis and High-throughput Flow Cytometry
Published on: March 1, 2013
Poly(2-oxazoline)-Based Polyplexes as a PEG-Free Plasmid DNA Delivery Platform
Dina N Yamaleyeva1,2, Naoki Makita2,3, Duhyeong Hwang2,4
1Joint UNC-CH and NC State Department of Biomedical Engineering, University of North Carolina at Chapel Hill, Chapel Hill, NC, 27599-7575, USA.
This study developed a polyethylene glycol (PEG)-free gene delivery platform using cationic poly(2-oxazoline) (POx) copolymers for immune cells. A specific DET- and pEtOx-based diblock copolymer showed superior transfection efficiency in macrophages, outperforming targeted versions and commercial reagents.
Area of Science:
- Polymer chemistry and biomaterials science
- Gene therapy and drug delivery
- Cell biology and immunology
Background:
- Polyethylene glycol (PEG) is widely used in gene delivery but can elicit immune responses.
- Developing PEG-free gene delivery systems is crucial for improving safety and efficacy.
- Cationic poly(2-oxazoline) (POx) copolymers offer a versatile platform for gene delivery applications.
Purpose of the Study:
- To expand the utility of cationic POx copolymers as a PEG-free gene delivery platform for immune cells.
- To synthesize and characterize novel POx-based block copolymers with varying hydrophilic, cationic, and hydrophobic blocks.
- To evaluate the transfection efficiency of these copolymers in monocytes and macrophages, including targeted and non-targeted formulations.
Main Methods:
- Synthesis of POx block copolymers with pMeOx or pEtOx hydrophilic blocks and modified MestOx cationic blocks (using DET or TREN).
- Incorporation of a hydrophobic iPrOx block in triblock copolymers.
- Functionalization with mannose targeting ligand via click chemistry.
- Preparation of polyplexes and evaluation of transfection efficiency in macrophages and monocytes.
- Assessment of cell internalization and comparison with a commercial transfection reagent (GeneJuice).
Main Results:
- DET-containing copolymers showed significantly higher macrophage transfection than TREN-based ones.
- Nontargeted pEtOx-based diblock copolymers were more effective than pMeOx-based counterparts.
- Triblock copolymers with hydrophobic iPrOx blocks performed less effectively than diblock copolymers.
- Mannose ligand attachment unexpectedly inhibited transfection and reduced cell internalization.
- The optimized PEG-free, nontargeted DET- and pEtOx-based diblock copolymer achieved transfection levels comparable to GeneJuice.
Conclusions:
- Cationic POx copolymers represent a promising PEG-free platform for gene delivery to immune cells.
- The choice of hydrophilic block (pEtOx > pMeOx) and cationic side chain (DET > TREN) significantly impacts transfection efficiency.
- Hydrophobic blocks can negatively affect performance, suggesting diblock structures are preferable.
- Surface functionalization with mannose for targeting can be detrimental to transfection in this system.
- A specific PEG-free, nontargeted POx diblock copolymer demonstrates high potential for macrophage gene delivery.
