Construction Of Tumor Gene Delivery Systems Using PEI Derivatives As Carriers: Modification Using Small Molecules And Macromolecules

Polyethyleneimine (PEI), as one of the most typical cationic polymer, has been widelyused as gene transfer vector. However, high cell toxicity and poor blood compatibilityhampered its further application in clinical. Natural compounds such as amino acids, proteinsand glycosaminoglycans, tend to have good biocompatibility, low immunogenicity andcytotoxicity.Someof natural compounds (e.g. chondroitin sulfate) are capable of specificallybinding to tumor cell surface antigens molecules (CD-44), which makes these naturalcompounds become a good choice of PEI modification to achieve targeting delivery. In viewof advantages of natural compounds, we designed and synthesized three natural compoundsmodified b-PEI25K derivatives, respectively. For these PEI derivatives, their biocompatibility,cytotoxicity and transfection efficiency were investigatedexperimentally, then the effects ofthe gene transfection on cells were evaluated. The contents of the thesis were divided asfollows:The N-acetyl-leucine was coupling to the primary amino groupsof the b-PEI25K asthe raw material via EDC/NHS catalyzed to synthesized different proportions of thehydrophobic modified PEI derivatives, gene vector N-Ac-L-Leu-PEI. The structure andphysicochemical properties of the synthetic carrier and the cytotoxicity andbiocompatibilityof the carrier were detected. The intracellular behavior of complexanditsinfluence on cell proliferation, migration and invasion were evaluated. Relatedexperimental results showed that derivatives have low toxicity, good biocompatibility andhigh transfection efficiency.By means of chemically bonding, the low toxicity of thermophilic histones and hightransfection efficiency PEI25K were bondedwith genipin, a natural crosslinking agentwith ared fluorescence, to synthesize a protein-polymer hybrid gene transfer vector which hassynergisticeffect. The structure and property of the PEI hybrid histone gene vectors was characterized, and verified the effective bonding of proteins and PEI and ability of the carrierto assembly with pDNA. Furtherexperiments examined cell toxicity and transfectionefficiency of the carrier. The experimental results showed that the new hybrid gene vector hasa better gene transfection efficiency and lower cytotoxicity.Natural glycosaminoglycan chondroitin sulfate was coupling to the b-PEI25K throughMichael addition reaction. Chondroitin sulfate was capable of specifically recognizing theCD44receptor expressed by tumor cells. A tumor suppressor, miR-34a, was targeting deliveryto CD44overexpressed tumor cells. The physical and chemical property of the carrier and thecytotoxicity and targeting capabilities were evaluated. Intracellular localization of the gene,related protein expression and cell migration were determined. Experimental results showedthat the novel targeting gene carrier could effectively delivery miR-34a into tumor cells viaCD44mediated endocytosis, inducing apoptosis of the tumor cells and inhibiting migration.In summary, wesynthesizeda variety of gene vector system based on b-PEI25K andnatural compounds. The carriers hadbetter biocompatibility, lower cytotoxicity whilemaintaining high transfection efficiency, and were capable of effectively mediating gene toperform biological function in the cells. Therefore, these carriers are expected to become anew type of efficient and safe gene carrier.