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ヒトVEGFおよびEpo遺伝子を用いた低酸素応答遺伝子発現ベクターの検討：崔　秉哲¹、柴田　徹¹、秋山暢丈²、野田　亮²、笹井啓資¹、平岡真寛¹（¹京大・医・放、²京大・医・分子腫瘍）

Enhancement of gene expression under hypoxic conditions using fragments of the human vascular endothelial growth factor and the erythropoietin genes :Heitetsu SAI¹,Toru SHIBATA¹, ,Nobutake AKIYAMA²,Makoto NODA²,Keisuke SASAI¹,Masahiro HIRAOKA¹ (¹Dep. of Radiology, Kyoto Univ. Hosp. ²Dep. of Molecular Oncology, Faculty of Med., Kyoto Univ.)

Purpose: Selective gene expression in response to hypoxia may provide a new strategy for cancer therapy. We have assessed the extent of hypoxia-responsiveness of various DNA constructs by luciferase assay to design vectors suitable for cancer therapy.
Materials and Methods: Reporter plasmids were constructed with a pGL3 vector and fragments of the vascular endothelial growth factor (VEGF) and erythropoietin (Epo) genes encompassing the putative hypoxia response element (HRE). Test plasmids and a control pRL-CMV plasmid were co-transfected into tumor cells by calcium phosphate method. After 6 hours of hypoxic stimulus, cells were harvested for assay.
Results: The construct pGL3/VEGF showed an increase in luciferase activity, the hypoxic/aerobic ratios of which were 3-4 and 8-12 for murine and human tumor cells, respectively. Epo fragment had an additive effect on human cell lines but not on murine ones. The insertion of 5 copies of HRE derived from the human VEGF and the adenovirus E1b minimal promoter resulted in a maximal enhancement of hypoxia responsiveness.
Conclusion: The constructs with VEGF and Epo fragments containing HRE are useful for inducing specific gene expression in hypoxic cells. Especially, the use of multiple copies of HRE and the E1b promoter markedly enhances hypoxia-responsiveness.