Polycation As Non-Viral Vector: Synthesis, Characterization And Design

Polycation as non-viral vectors has many advantages when compared with viral vectors, such as improved safety, greater flexibility and more facile manufacturing. Therefore, it has become more and more popular among gene therapy fields. However, the lack of specificity, accumulation of toxicity for in vivo and relative low transfection efficiency has limited its application as ideal gene delivery vectors. To develop novel degradable polycation with high transfection efficiency and low toxicity may become one of the most vital factors for the success application of gene therapy.The aims of this study is to synthesis and characterize a kind of polyurethane polymer based on polyethylenimine (PEI), which is able to degrade into non-toxic small molecular chemistries and more importantly during its degradation acid is not produced, which is always supposed to prevent the escape of Polymer/DNA from lysosome, and with this polymer as example to testify the feasibility of this design method. With PEI-800 (Mw=800Da) as reactant for its low toxicity and 1, 4-Butanediol bis(chloroformate) as functional linker, PEI derivate (PL) was acquired. Its Mw is 3924Da and half life of degradation at pH=4.5 is about 100 days by GPC assay. PL was able to condense DNA and formed polyplexes effectively at w/w ratio of 1 and above. The mean particle size of the polyplexes was lower than 100 nm and the zeta potential was about 20 mv. The cytotoxicity of PL is lower than PEI-25k (Mw=25000Da) and similar to PEI-800. In vitro transfection assay, the deviate achieved at least one order of magnitude higher than PEI-25k at w/w ratio of 7 and above for luciferase DNA. EGFP plasmid was used to investigate the optimal ratio and transfection time, the expression amount can be observed significantly and reach the highest at w/w 40 after 72 h. In vivo transfection assay, the gene expression of PL/DNA was relatively low after the tail intravenous for mice, but it was still higher than that of PEI-25k and was spontaneously targeted to lung. However, in vivo application, systematic administration of PL/DNA caused death of mice.In conclusion, the development of polyurethane polymer has great potential to solve problems existing in the application of polycation as non-viral gene carrier.