Synthesis and Characterization of Polyphenylalanine Grafted Low Molecular Weight PEI as Efficient Gene Carriers

To search safe and evaluate non-viral nucleic acids carrier, a series of polyphenylalanine and phenylalanine grafted low molecular weight PEI with a molecular weight of 1.8 x 10(3) (PEI1.8k-g-PPhe and PEI1.8k-g-Phe) were prepared by NCA ring opening polymerization initiated by PEI-1.8k and phenylalanines conjugation to PEI-1.8k, respectively. The polymers and the complexes of PEI1.8k-g-PPhe/DNA and PEI1.8k-g-Phe/DNA were characterized by nuclear magnetic resonance (NMR) analysis, particle size analysis, zeta potential analysis, luciferase analysis, flow cytometry (FCM) analysis and confocal laser scan microscopy (CLSM). In order to compare the property for compacting DNA into nanoparticles, the particle size and zeta potential analysis were carried out. Both PEI1.8k-g-PPhe and PEI1.8k-g-Phe showed suitable particle size and zeta potential for gene delivery. The particle size of PEI1.8k-g-PPhe10/DNA complexes were about 150 nm and the zeta potentials were about +16 mV, which were suitable for the in vitro experiments. Moreover, cell viability, after treating with different copolymers at various concentrations, was studied by an MTT assay. The reduced cytotoxicity of PEI1.8k-g-PPhe and PEI1.8k-g-Phe may be because of the introduction of neutral hydrophobic phenylalanine moieties. And both of the PEI1.8k-g-PPhe and PEI1.8k-g-Phe had lower cytotoxicity (above 70% viability at a higher concentration 1 mg/mL) than that of PEI with a molecular weight of 2.5 x 10(4) (PEI-25k) in HeLa cells. The in vitro gene transfection of PEI1.8k-g-PPhe10 and PEI1.8k-g-Phe10 was conducted in human cervical cancer (HeLa) and breast cancer (MCF-7) cells. In both of the cells, PEI1.8k-g-PPhe10 exhibited much higher gene transfection efficiency. PEI1.8k-g-PPhe10/DNA complexes showed remarkable gene transfection efficiency, which was about twelve times higher than that of PEI-25k. The endocytosis efficiency of PEI1.8k-g-PPhe10/DNA and PEI1.8k-g-Phe10/DNA were quantified using flow cytometry. Due to their regular polymer chain, PEI1.8k-g-PPhe10/Cy5-DNA showed better internalization efficiency than PEI1.8k-g-Phe10/Cy5-DNA, PEI-25k/Cy5-DNA and PEI-1.8k/Cy5-DNA. The CLSM assay was carried out to verify the internalization efficiency together which also indicated that PEI1.8k-g-PPhe10/Cy5-DNA induced higher intracellular uptake efficiency than the others.