Application of Intelligent Targeting System for Cancer Gene Therapy

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1) Application of boron entrapped stealth liposome to tumour cell growth inhibition in in vivo boron neutron capture therapy model : The tumor cell destruction in boron neutron-capture therapy (BNCT) is due to the nuclear reaction between ^<10>B and thermal neutrons. It is necessary for effective BNCT to accumulate of ^<10>B atoms in the tumor cells. We prepare a polyethylene-glycol (PEG) binding liposome (DPPC/cholesterol/DSPC-PEG2000) entrapped ^<10>B compound for the delivery system. We evaluated the cytotoxic effects of intrveneously injected ^<10>B-PEG-liposome on human pancreatic carcinoma xenografts in nude mice with thermal neutron irradiation. After thermal neutron irradiation of mice injected with ^<10>B- bare liposome or ^<10>B-PEG-liposome, AsPC-1 tumour growth was suppressed relative to controls. Injection of ^<10>B-PEG-liposome caused the greatest tumour suppression with thermal neutron irradiation in vivo These results suggest that intraveneous injection of ^<10>B-PEG-li … More posome can increase the retention of ^<10>B atoms by tumor cells, causing tumor growth suppression in vivo upon thermal neutron irradiation.2) Liposomal gene delivery using novel lipoplexes called "Qplex" for cancer therapy in vitro : Non-viral vectors, such as cationic liposomes and cationic polymers have developed in the merits without immunogenicity, potential recombination, nor complementation. It is necessary to increase the transfectional efficiency when we use non-viral vector on cancer gene therapy. We have prepared new typed plasmid DNA-cationic lipoplexes, called quatenary complex ("Qplex"), and evaluated DNA delivery efficiency. Qplex is composed with cationic liposome, pDNA, protamine and transferrin. The lipid of liposome M-(α-trimethylammonioacetyl)-didodecyl-D-glutamatechloride (TMAG)/dilauroyl-phospatidylcholine (DLPC)/dioleoylphospatidylethanolamine (DOPE) (1 : 2 : 2). The transfectional efficiency of lac Z gene is increased to 67% with Qplex from 4% with conventional cationic lipoplexes in Xgal staining. The transfection efficiency is superior in the existence of serum.Tob (Transducer of Erb B-2) is a newly identified tumor suppressor that may interact and interfere with tyrosine kinase receptors including Erb B-2. Introduction of tob gene into NIH3T3 cells results in suppression of growth of the cells. Tob plasmid was entrapped into the "Qplex". The tumor growth suppression of AsPC-1 pancreatic cancer cells was shown in 50% of thymidine uptake regression. These results, suggest that the Qplex has an candidate for non viral vector of gene therapy for treatment of cancer.3) Transfection with Luciferase Plasmid/Chitosan Complex and Analyses of Transfection Mechanism : In this study, we transfected tumor cells (A549, B16 and Hela) with the plasmid/chitosan complex. The complex showed higher transfection activity than the plasmid/lipofectin complex did. Although the polygalactosamine is the same amino polysaccharide as the chitosan, the plasmid/polygalactosamine complex did not show any gene expression. So we analysed the travsfection mechanism between chitosan and polygalactosamine complex. We synthesized FITC-labeled luciferase plasmid and Texas Redlabeled chitosan to evaluate the transfection efficiency, cell uptake and sub-cellular distribution of the plasmid/chitosan complex. Gel shift assay was used to evaluate the stability of the complexes in the presence of Dnase I and anionic surfactant.. Less