TPGS-b-PBAE Copolymer-Based Polyplex Nanoparticles for Gene Delivery and Transfection In Vivo and In Vitro

Area of Science:

• Biomaterials Science
• Gene Therapy
• Nanotechnology

Background:

• Poly (β-amino ester) (PBAE) is a promising non-viral gene vector but can cause cytotoxicity due to its high positive charge.
• Hydrophilic d-α-tocopherol polyethylene glycol succinate (TPGS) can improve polymer biocompatibility.

Purpose of the Study:

• To synthesize and characterize TPGS-b-PBAE (TBP) copolymers.
• To evaluate the gene transfection efficiency and cytotoxicity of TBP-based polyplex nanoparticles (NPs).
• To explore the potential of TBP-based CRISPR/cas9 systems for gene therapy.

Main Methods:

• TPGS-b-PBAE copolymers were synthesized with varying molar ratios.
• Polyplex nanoparticles (NPs) were formed by complexing TBP with plasmid DNA.
• Gene transfection efficiency was assessed in HEK 293T cells using GFP plasmid.
• Cytotoxicity was evaluated across multiple cell lines (HEK 293T, HeLa, Me180, SiHa, SCC-7, C666-1).
• CRISPR/cas9 gene editing systems targeting MUC2 and LMP1 were constructed and tested in vitro and in vivo.

Main Results:

• TBP copolymers were successfully synthesized with tunable molecular weights.
• TBP-based polyplex NPs demonstrated effective gene transfection with significantly reduced cytotoxicity.
• The TBP-GFP polyplex NPs showed efficient transfection in a broad range of cancer cell lines.
• Targeted CRISPR/cas9 gene editing systems were successfully constructed and validated for MUC2 and LMP1, showing therapeutic potential.

Conclusions:

• TPGS-b-PBAE copolymers offer enhanced biocompatibility and reduced cytotoxicity for non-viral gene delivery.
• TBP-based polyplex NPs are effective gene carriers with broad applicability in cancer cell lines.
• The developed CRISPR/cas9 gene editing systems show significant potential for future gene therapy applications.