Combination of radiation and suicide gene therapy with radio-sensitized promoter/Endotherial precursor cell in glioma

2003 Fiscal Year Final Research Report Summary

• Project/Area Number: 14571350
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Cerebral neurosurgery
• Research Institution: Chiba Cancer Center Research Institute
• Principal Investigator: OGA Masaru Chiba Cancer Center Research Institute, Division of Neurological Surgery/Division of Chemotherapy, 脳神経外科・化学療法研究部, 医長・兼務研究員 (10251159)
• Co-Investigator(Kenkyū-buntansha): HATANO Kazuo Chiba Cancer Center Research Institute, Division of Radiation Oncology, Head, 放射線治療部, 部長 ; TAKENAGA Keizo Chiba Cancer Center Research Institute, Division of Chemotherapy, Chief Scientist, 化学療法研究部, 主席研究員 (80260256)
• Project Period (FY): 2002 – 2003
• Keywords: Radio-sensitization / Egr-1 / CArG element / gene therapy / Glioma / Suicide gene

Research Abstract

In the first year we verified that the gene expression located at the downstream of radio-sensitized promoter would actually amplify by radiation. We constructed the EGFP&HSV-tk vectors containing E_4/CMV chimeric radio-sensitized promoter (E_4: four-repeated structure of CArG element located at Egr-1 gene). E_4/EGFP was transferred into U251-MG/U373-MG human glioma cell line respectively and the expression of EGFP was evaluated by optical observation under fluorescent microscopy and Western blotting after 3Gy irradiation. As a result, we verified that the expression of EGFP was amplified about 1.3〜1.4 times after 6〜54 hours compared to pre-irradiation stage. Then E4/HSV-tk and HSV-tk were transferred into U251-MG/U373-MG human glioma cell line respectively, and the degree of cell proliferation was evaluated by MTT assay after irradiation (1,3,5Gy) following GCV administration. The results were 1)the radio-sensitized promoter also could be activated by physical/chemical stimuli. 2)the radiation dose-dependent apoptosis was observed only in the E_4/HSV-tk-administrative group but not statistically significant.
In the second year, we replaced the promoter with E_<9ns-2>, stronger than E_4 in radio-sensitization and did the same trial. Contrary to the expectation, however, the expression of EGFP was not significantly amplified in the E_<9ns-2>/EGFP -administrative groups. In the MTT assay, we also could not verified the significant inhibition of the cell proliferation among E_<9ns-2>/HSV-tk-administrative glioma cell lines after irradiation. We are going to promote our previously scheduled plan by using E_4/CMV as the radio-sensitized promoter, transferring E_4/HSV-tk into C6 and endothelial precursor cell line.