Polycations With Random Coil And Micellar Structures For Gene Delivery

Gene therapy has been proved to be a promising approach to treat inherited and acquired disease. The key of clinical application in gene therapy is to obtain an efficient gene delivery vector. Cationic polymers(polycations) as an important type of nonviral vectors are believed to be able to protect genes from enzymatic degradation and facilitate their cellular uptake. Although a lot of polycations exhibit attractive properties in many aspects, such as low immune, easy to prepare and low cost, the low gene transfection efficiency is still a bottleneck for their application as gene carrier. In order to improve the gene delivery efficiency, a lot of methods have been proposed through chemical modification of polycations.In this study, we synthesized hydrophilic poly(vinyl pyrrolidone)-graft-poly[2-(N,N-dimethylamino)ethylmethacrylate](PVP-g-PDMAEM A) with two types of PVP, where PVP can play synergistic effect on the gene delivery with the outstanding delivery performance of PDMAEMA. We studied the effect of chain length of hydrophilic segment on gene delivery and optimize the length of hydrophilic segments. And then we introduced the hydrophobic segments to the better one, namely, amphiphilic copolymer(PVP-g-PDMAEMA-b-PMMA), which was synthesized via atom transfer radical polymerization(ATRP) method, and their properties as gene vectors were investigated subsequently. PVP-g-PDMAEMA formed random coil in water and PVP-g-PDMAEMA-b-PMMA self-assembled into spherical core-shell micelle with a very low critical micelle concentration of only 6.3 × 10-3 mg/m L. Compared with PVP-g-PDMAEMA, the zeta potential and particle size of PVP-g-PDMAEMA-b-PMMA/pDNA were lower. PVP-g-PDMAEMA-b-PMMA/pDNA polyplexes performed excellent gene transfection efficiency, which showed not only much higher gene transfection efficiency than PVP-g-PDMAEMA/pDNA polyplexes, but obviously surpassed 25 k PEI at low N/P ratio around 3 on 293 T cell lines. Hence, the results suggested that PVP-g-PDMAEMA-b-PMMA could be a highly efficient gene vector.Although polycations exhibit high gene transfection efficiency in vitro, the transfection efficiency in vivo is relatively low. To address this issue, we developed the charge reversal process and reversible broken-formation of disulfide bond method based on cysteamine modified bovine serum albumin(SH-BSA). SH-BSA can combine with the PVP-g-PDMAEMA-b-PMMA/pDNA polyplexes through electrostatic attraction under physiological conditions and further enhance the stability of the polyplexes by the disulfide bond. The PI of SH-BSA was enhanced from 4.7 to 5.3. SH-BSA possessed negative charges at physiological pH and SH-BSA turned to positive charges when pH was decreased to 5.0. SH-BSA will occur the charge reversial process with the variation of pH and produced electrostatic repulsion with the cationic binary polyplexes. However, it will not get out of the polyplexes due to the cross-linker of disulfide bond. It is observed that SH-BSA can disassociate from the polyplexes through electrostatic repulsion and the broken of disulfide bond when pH adjusted to 5.0 with adding the glutathione. It is promising that the appropriate stability of SH-BSA can do favor for the gene transfer and release.