Effective and safe in vivo gene delivery based on polyglutamic acid complexes with heterocyclic amine...

Polyethylenimine (PEI) has been extensively used for non-viral gene delivery. Increasing the molecular weight of PEI often improves transfection efficiency, but enhances cytotoxicity and non-specific interaction with plasma proteins, limiting its use in clinical applications. In this study, poly-l-glutamic acid (L-PGA) as an anionic polymer, was introduced to piperazine-modified PEI to improve its in vivo properties. The physicochemical properties, cytotoxicity, in vitro and in vivo tranfection efficiency of these carriers were evaluated. Conjugation of 50% of primary amines of PEI 25 kDa with piperazine in the presence of PGA_1% (PEI₂₅Pip_50%/PGA_1%) could significantly increase transfection efficiency even in the presence of serum compared to PEI 25 kDa. Increasing the PGA content led to lower cytotoxicity of DNA/PEI₂₅Pip_50%/PGA_1% triplexes. Systemic administration of triplexes in Balb/c mice resulted in significant enhancement of luciferase gene expression in brain, spleen, and liver compared to PEI 25 kDa. In a 30-day survival study, no significant changes were observed in mice body weights in DNA/PEI₂₅Pip_50%/PGA_1% group. Moreover, this group exhibited a survival rate of 100% compared to 0% in mice receiving PEI 25 kDa. This novel PEI₂₅Pip_50%/PGA_1% carrier could be used to overcome the serum inhibitory effects on gene expression in vivo, providing a promising gene delivery system for tissue-specific targeting.