Reversible Stabilization of Nanofiber-Polyplexes through Introducing Cross-Linkages.

Area of Science:

• Biomaterials Science
• Gene Therapy
• Nanotechnology

Background:

• Non-viral gene delivery vectors require a balance between stability for uptake and instability for cargo release.
• Previous work showed multi-arm PEG-PLL (maPEG-PLL) nanofiber-polyplexes offer effective gene transcription.
• Achieving reversible stabilization without compromising gene expression is a key challenge.

Purpose of the Study:

• To investigate the reversible stabilization of maPEG-PLL nanofiber-polyplexes.
• To introduce disulfide cross-links for controlled destabilization under reducing conditions.
• To evaluate the impact of reversible stabilization on gene delivery efficiency.

Main Methods:

• Utilized dithiobis (succinimidyl propionate) (DSP) to introduce disulfide cross-links into maPEG-PLL polyplexes.
• Assessed polyplex stability and pDNA reactivity using agarose gel electrophoresis and real-time PCR.
• Evaluated gene expression in cultured cells, comparing DSP-crosslinked polyplexes with polyethyleneimine (PEI) polyplexes.

Main Results:

• Successfully achieved reversible stabilization of nanofiber-polyplexes using DSP cross-linking.
• Demonstrated that disulfide bonds can be disrupted under reducing conditions (dithiothreitol).
• DSP-crosslinked polyplexes showed superior gene expression compared to conventional PEI polyplexes in cell-based assays.

Conclusions:

• Reversible stabilization of gene delivery polyplexes is feasible using disulfide cross-linking.
• DSP-crosslinked maPEG-PLL nanofiber-polyplexes offer a promising non-viral gene delivery strategy.
• This approach enhances gene expression while maintaining controlled stability.