Kinetic Control in Assembly of Plasmid DNA/Polycation Complex Nanoparticles

Area of Science:

• Biomaterials Science
• Nanotechnology
• Gene Therapy Delivery

Background:

• Polyelectrolyte complex (PEC) nanoparticles, formed from plasmid DNA (pDNA) and polycations like linear polyethylenimine (lPEI), are key nonviral gene therapy vectors.
• Current methods for controlling nanoparticle properties focus on polycation structure and assembly conditions, but reproducible production requires control over assembly kinetics.

Purpose of the Study:

• To develop a kinetically controlled method for assembling pDNA/lPEI nanoparticles with tunable size, composition, and pDNA payload.
• To demonstrate the scalability and reproducibility of this novel assembly process.

Main Methods:

• Flash nanocomplexation (FNC), a microfluidic technique, was used to rapidly mix pDNA and lPEI solutions, controlling polyelectrolyte complex assembly kinetics.
• A combined experimental and simulation approach was employed to characterize the resulting nanoparticles.

Main Results:

• FNC enabled precise control over nanoparticle size (35-130 nm) and pDNA payload (1.3-21.8 copies per nanoparticle).
• Nanoparticles produced via FNC showed improved uniformity and scalability compared to bulk mixing methods.
• The study demonstrated a correlation between pDNA payload, formulation composition, and *in vivo* transfection efficiency and toxicity.
• The nanoparticles exhibited excellent long-term stability in a lyophilized formulation for at least 9 months at -20 °C.

Conclusions:

• Flash nanocomplexation offers a robust and scalable method for producing well-defined pDNA/lPEI nanoparticles for gene medicine.
• This approach facilitates the development of off-the-shelf gene therapy products with predictable performance.