Highly branched poly(β-amino ester)s with narrow molecular weight distribution: Fractionation and gene transfection activity

Highly branched poly(β-amino ester)s (HPAEs) have shown their great promise in gene delivery. However, their broad molecular weight distribution (MWD) poses an additional challenge to the mechanistic understanding of the influence of molecular weight (MW) on their gene transfection activity. Using a stepwise precipitation strategy, HPAEs were fractionated. It is shown that MW has a significant effect on the transfection activity and cytotoxicity of HPAEs. The intermediate MW mediates higher transfection efficiency while maintaining high cell viability. Mechanistic studies show that the intermediate MW confers stronger DNA binding affinity to HPAEs, leading to the formulation of polyplexes with a relatively smaller size and more positive zeta potential. This study not only suggests a simple strategy to fractionate HPAEs with narrow MWD but also provides new insights into understanding the structure-property relationship, which would facilitate the clinical translation of HPAEs in gene therapy.