| Co-Investigator(Kenkyū-buntansha) |
SANO Akihiko Sumitomo Pharmaceuticals Co. Ltd., Formulation Research Laboratories, 製剤技術研究所・創剤研究グループ, 主任研究員
MIRISHITA Ryuichi Osaka University, Graduate School of Medicine, Professor, 医学系研究科, 教授 (40291439)
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| Research Abstract |
We attempted to prolong release of NFkB oligodeoxynudeotid.es without degradation. First, we tested the efficiency of introduction of FITC-labeled oligonucleotides into cultured cells using atelocollagen. No fluorescence was detected at 48 hours after the transfer to HeLa, BHK-21 and human tongue cancer cell line SAS. Then, we developed HVJ (hemagglutinating virus of Japan ; Sendai virus) envelope vector which can introduce synthetic oligonucleotides, proteins, peptides and chemical drugs as well as genes. FITC-labeled oligonucleotides were incorporated into HVJ envelope vector by the treatment of mild detergent and centrifugation. Ten minutes after incubation of those cultured cells described above with the HVJ envelope vector, fluorescence was observed in almost all the nuclei of the cells. When NFkB decoy oligonucleotides were introduced into cancer cells such as SAS and HeLa cells using HVJ envelope vector, apoptosis after irradiation was enhanced by the suppression of NFkB-induced
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gene expression. When FITC-labeled NFkB decoy oligonucleotides were introduced into the joint space of Cynomolgus monkeys using HVJ envelope vector, fluorescence was detected at both synovium and articular cartilage. Next, HVJ envelope vector was embedded with various polymers. First, without polymer, HVJ envelope vector reaches spleen by intravenous injection and FITC-oligonucleotides were detected at the marginal zone of mouse spleen. Then, the vector containing luciferase gene was decorated with protamine suifate and the complex was injected into tail vein of mouse. Gene expression was detected exclusively in lung, not in spleen. Furthermore, when HVJ envelope vector mixed with heparin was injected into muscle or brain directly, gene expression was approximately 5 fold increased than that without heparin. The effect of polymers on slow release of the vector is being investigated, but we have got preliminary data suggesting that cationic polymers may enhance slow release of HVJ envelope vector. Less
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