| Budget Amount *help |
¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 2004: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2003: ¥1,900,000 (Direct Cost: ¥1,900,000)
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| Research Abstract |
Recent y precipitation of powders with supercritical carbon dioxide- (SCF) has been attracting much attention as a method to produce tarry partides with high functionality We have already reported that the gene powders grew cl by the SCF prep bad improved stability and iir aced gene expression in lungs after intratracheal insufflation in mice. In the present study, we prepared chitosan-interferon β (DNA) powders by the SCF process to exannne the therapeutics of there in murinelung metastasis model
An bus :solution of pCMV-MuBβ a plasmid DNA cabling murine intern β, and chitosan, a nonviral vector was dispersed in SCF with ethanol as a modifier to precipitate them. Mannitol was used as a powder vehicle. To establish a lung metasitatis, CT26, mouse colon carcinoma cells, were injected intravenously into mouse tail vein. The DNA powder or solution was administered intratracheally or intravenously to examine the lung weighty, number of metastatic nodules, and survival rate with time. The ge
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ne expression after intratracheal administration of DNA was abserved in normal and cancer tissue in the lung, while no expression was observed in the other organs. The DNA powders suppressed the increase in lung weight and number of nodules and prolonged the survival of the mice with smaller dose of DNA than DNA solutions. Intratracheal administration was more effective than intravenous administration. These findings suggested that the DNA powders prepared by the SCF processs had high therapeutic potential in murine lung metastasis model.
The next study examined the stability of a gene in powers prepared with supercritical carbon dioxide (CO2) from the viewpoints of the ternary structure of DNA and in vivo inch potential. An aqueous chitosan-pCMV-Luc complex solution containing mannitol was injected into the stream of a supercritical CO2/ethanol admixture to precipitate a gene powder. The obtained gem powders and gene solutions were placed in stability chambers at 25 or 40゜C for 4 weeks. The integrity, and transfection potency of the gene were examined by electrophoresis and in van pulmonary transfection study in mice The supercritical CO2 process decreased the sided DNA doting the manufacturing process; however, the decrease in the remaining supercoiled and open circular DNA in the powders during storage was much slower than that in solutions. In addition, the powders had W war w on potency than the solutions containing the same amount of DNA. The effect of chitosan on the stability of DNA in solutions was not obvious in the solutions but it improved the stability of DNA in powders du manufacturing and storage. Thus, a gene powder with a vector is a promising formulation for a ready-to use inhalation therapy of pulmonary diseases. Less
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