ANTITUMOR THERAPIES BASED ON INHIBITION OF SUPEROXIDE DISMUTASE

基于抑制超氧化物歧化酶的抗肿瘤疗法

• 批准号: 6203295
• 负责人: LARRY OBERLEY
• 金额: $ 11.34万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-08-31 至 2001-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6203295
• 关键词: MCF7 cell; antineoplastics; antisense nucleic acid; bleomycin; catalase; complementary DNA; doxorubicin; drug resistance; electron spin resonance spectroscopy; enzyme activity; fatty acids; free radical oxygen; glutathione; glutathione peroxidase; glutathione reductase; ionizing radiation; lipid peroxides; neoplasm /cancer pharmacology; northern blottings; oxidoreductase inhibitor; southern blotting; superoxide dismutase; transfection; tumor necrosis factor alpha

The purpose of this proposal is to study the role of active oxygen species in cancer cell cytotoxicity and to determine if lipids are a major target of these species. In order to study the role of active oxygen, superoxide dismutase (SOD) levels of parental and doxorubicin- resistant human breast carcinoma MCF-7 cells will be modulated and the effect on cell killing by selected antitumor agents (doxorubicin, bleomycin, ionizing radiation, tumor necrosis factor) determined. Modulation of superoxide dismutase will be accomplished by transfection of sense and anti-sense SOD cDNA. After transfection, clones with lower and higher than control levels of SOD activity will be isolated. These clones will then be tested for sensitivity to the selected antitumor agents. Both CuZnSOD and MnSOD will be transfected. Verification that superoxide dismutase modulation has occurred will be accomplished using enzyme activity, immunoreactive protein, Northern and Southern blot assays. Catalase, glutathione peroxidase, glutathione reductase, and glutathione levels will also be measured in the stable clones to determine if any of the other common antioxidants have been altered. Similar studies will be performed after glutathione peroxidase (GPX) transfection. GPX will be transferred into wild type MCF-7 cells as well as an SOD-overexpressing clone. The role of oxygen and lipid radicals will also be tested directly by measuring qualitatively and quantitatively the radicals present after treatment of control and SOD-and/or GPX-modified cells with the selected antitumor agents. Measurement of radicals will be accompanied by using the electron paramagnetic resonance technique of spin trapping. Last, the role of lipids in the antitumor effect will be studied. This will be done in two different ways: 1) measuring the amount of lipid peroxidation that occurs during tumor cell killing by the above agents; and 2) modifying the lipids of the cell and determining the effect on tumor cell killing by selected antitumor combinations in control and SOD- and/or GPX-transfected cell lines. This research should provide valuable new information on the mechanisms and targets of antitumor agents. It is hoped this knowledge will eventually lead to more powerful antitumor protocols.

这个建议的目的是研究活性氧的作用 并确定脂质是否是癌细胞毒性的一个重要因素。 这些物种的主要目标。 为了研究积极的作用 氧，超氧化物歧化酶（SOD）水平的父母和阿霉素- 将调节耐药人乳腺癌MCF-7细胞， 通过选择的抗肿瘤剂（阿霉素， 博莱霉素、电离辐射、肿瘤坏死因子）测定。 超氧化物歧化酶的调节将通过转染完成 正义和反义SOD cDNA的表达。 转染后，具有较低的 且高于对照水平的SOD活性将被分离。 这些 然后测试克隆对所选抗肿瘤药物的敏感性 剂. CuZnSOD和MnSOD都将被转染。 将验证是否发生超氧化物歧化酶调节， 使用酶活性、免疫反应蛋白、北方和 Southern印迹分析。 过氧化氢酶、谷胱甘肽过氧化物酶、谷胱甘肽 还原酶和谷胱甘肽水平也将在稳定的 克隆，以确定是否有任何其他常见的抗氧化剂已经 改变了 在谷胱甘肽过氧化物酶（GPX）之后将进行类似的研究 转染 GPX也将被转移到野生型MCF-7细胞中 作为SOD过度表达克隆。 氧和脂质自由基的作用也将直接通过 定性和定量测量存在的自由基后， 用所选化合物处理对照和SOD和/或GPX修饰的细胞 抗肿瘤剂。 自由基的测量将伴随着使用 自旋捕获的电子顺磁共振技术。 最后，将研究脂质在抗肿瘤作用中的作用。 这 将以两种不同的方式进行：1）测量脂质的量 在上述试剂杀死肿瘤细胞期间发生的过氧化; 和2）修饰细胞的脂质并确定对细胞的影响， 在对照组和SOD-1中通过选择的抗肿瘤组合杀死肿瘤细胞。 和/或GPX转染的细胞系。 这项研究应该提供有关这些机制的有价值的新信息。 和抗肿瘤剂的靶点。 希望这些知识能 最终产生更强大的抗肿瘤方案。