| Co-Investigator(Kenkyū-buntansha) |
OHTA Yoshihisa Faculty of Medicine, Department of Neurosurgery, Tokyo Medical and Dental University Assistant, 医学部・附属病院, 助手 (70311652)
AOYAGI Masaru Faculty of Medicine, Department of Neurosurgery, Tokyo Medical and Dental University Instructor, 医学部・附属病院, 講師 (40134704)
HAMADA Hirofumi Sapporo Medical University, Faculty of Medicine, Professor, 医学部, 教授 (00189614)
YAMAMOTO Shinji Faculty of Medicine, Department of Neurosurgery, Tokyo Medical and Dental University Assistant, 医学部・附属病院, 助手
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| Research Abstract |
Recombinant adenoviral vectors are widely used in clinical and experimental studies to treat malignant tumors. Recently, host immune responses have been proposed as a major limitation in using adenoviral vectors for repeated gene delivery. We have demonstrated another limitation unrelated to host immunity. The authors established U373R and U373RC, adenovirus-resistant cell lines, by repeated transfection of an adenoviral vector expressing the human p53 gene(AxCIhp53)to U373MG, a glioma cell line susceptible to p53. The transduction efficiencies of the vectors for U373R and U373RC cells were decreased due to the reduced expression levels of integrins_v_3 and _v_5(that is, the penton base receptors)in U373R and CAR(the fiber knob receptor)in U373RC cells. The fiber mutant vector carrying the p53 gene(AxCAhp53-F/K20)improved the transduction efficiency and dramatically induced apoptosis in both cell lines. These results indicate that reduced transduction efficiency due to reduced expressi
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on levels of viral receptors will be a cause of acquired resistance, and that altering the tropism of vectors is one of the effective methods to overcome the resistance. We have also proved that the fiber mutant vector(F/K 20)is integrated into target cells irrespective of expression levels of CAR.
The proteasome, a protease composed of multiple subunits, degrades ubiquitinated proteins including cell cycle related proteins. Inhibition of the proteasome induces apoptosis in tumor cells. In our experiments, inhibition of chymotrypsin-like activities of proteasome strongly induced apoptosis to glioma cells. We found that mitochondria and a wide spectrum of caspases are associated with apoptosis induced by inhibition of chymotrypsin-like activities of proteasome. PSI also induced accumulation of cyclins A and B1 in the treated cells, resulting in the cell cycle arrest at G2/M phase. Inhibition of the proteasome augmented the cytocidal effects of AxCIhp53. Taken together, chymotrypsin-like activities of proteasome and mitotic cyclins might be promising target for the treatment of gliomas, and inhibition of proteasome activities can be applied to gene therapy.
Hamada et al. developed a number of technologies for gene therapy using adenoviral vectors. For example, they applied Fas, NF-KappaB, BAX, caspases etc to experiments for the treatment of gliomas.
In addition to these experimental studies for gene therapy, we have studied biological characteristics of malignant brain tumors including gliomas. We found that gangliosides play very important roles in metastasis of caucers to the brain. Our experiments demonstrated that mutations in p53 gene were very critical for the malignant progression of meningiomas and hemamgiopericytomas. These findings will be a great help to gene therapy using p53 gene. Less
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