Synthesis of a barbell-like triblock copolymer, poly(L-lysine) dendrimer-block-poly(ethylene glycol)-block-poly(L-lysine) dendrimer, and its self-assembly with plasmid DNA

A barbell-like ABA-type triblock copolymer, poly(L-lysine) dendrimer-block- poly(ethylene glycol) block-poly(L-lysine) dendrimer (PLLD-PEG-PLLD), was s ynthesized by the liquid-phase peptide synthesis method. The self-assemblin g complex formation of the third and fourth generation of the copolymer wit h plasmid DNA was studied. H-1 NMR and matrix-assisted laser desorption/ion ization-time-of-flight mass spectrometry (MALDI-TOF MS) were used for the c haracterization of the synthesized copolymer. The self-assembling behavior of the 3rd and 4th generations of the copolymer with plasmid DNA was invest igated by electrophoretic mobility shift assay, DNase I protection assay, a nd ethidium bromide exclusion assay. We observed great differences in the s elf-assembling ability of the 3rd and 4th generations of the polymer. This suggests that the number of positively charged amines per polymer molecule should be an important factor for the potential for self-assembling complex formation with DNA. Atomic force microscopy (AFM) and zeta potentials were used for evaluating the shape, size distribution, and surface charge of th e complexes at various charge ratios. From AFM images, it was observed that the shape of the complex was nearly spherical and its size was about 50-15 0 nm in diameter. The in vitro cytotoxicity of the copolymer was compared w ith that of poly(L-lysine), poly(D-lysine), and polyethylenimine.