RADIORESISTANCE, APOPTOSIS AND THE CELL CYCLE

辐射抗性、细胞凋亡和细胞周期

• 批准号: 2106569
• 负责人: WILLIAM G. MCKENNA
• 金额: $ 21.85万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-07-01 至 1998-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2106569
• 关键词: HeLa cells; X ray; animal tissue; antineoplastics; antisense nucleic acid; apoptosis; biological signal transduction; caffeine; cell cycle; cyclins; dosage; farnesyl compound; fibroblasts; gene expression; inhibitor /antagonist; oligonucleotides; oncogenes; phenotype; radiation genetics; radiation related neoplasm /cancer; radiation resistance; radiation sensitivity; radiosensitizer; staurosporine; transfection

Tumors vary considerably in their radioresponsiveness and one important component of that variation is the intrinsic radiosensitivity of the tumor cells themselves. Oncogenes can alter the radiosensitivity of cells, but the mechanisms through which they affect the cells' response to radiation remain unknown. We have begun to explore whether oncogenes affect apoptosis after radiation. We have found that the oncogenes Hras and Myc when used to transform rat embryo cells (REC) lead to cell lines with substantially greater radioresistance than REC themselves or REC with either oncogene alone. We have shown that this difference is unrelated to the extent of double strand DNA breaks induced by radiation; nor is it related to the extent or kinetics of rejoining of these breaks. The radioresistant phenotype correlates with a longer G2 delay induced by radiation. We have recently shown that the resistant cells respond to radiation by depressing synthesis of one of the mitotic cyclins, cyclin B1, whereas, the sensitive cells show no effect of radiation on mitotic cyclin expression. The Myc transfected cells undergo apoptosis after serum withdrawal and after radiation. Apoptosis is seen to a much lesser degree in the cultures of the ras transfected cells after the same dose of radiation yet they still undergo apoptosis after serum withdrawal. Thus ras appears to protect the cells from radiation induced apoptosis, but does not protect the cells from apoptosis induced by serum withdrawal. A similar phenotype is seen in Hela cells which are also very resistant to radiation induced apoptosis and similarly have a prolonged radiation induced G2 delay with depressed cyclin B1 expression. The purpose of this application is to explore the hypothesis that the radiation induced G2 delay acts as a regulator of the apoptotic response. Since the transition through G2/M is regulated in part by and requires the expression of cyclin B1, we propose that the G2 delay and thus perhaps also apoptosis are regulated through cyclin B1 expression. We intend to investigate the contribution the oncogene makes to determining whether apoptosis occurs, both by manipulating oncogene expression or activity and by transfecting other genes which may counter the effects of the oncogenes on the apoptotic pathway. We will inhibit ras action using inhibitors of farnesylation. We will use antisense technology to alter the kinetics of mitotic cyclin expression and hence alter the timing of G2 in order to test the effect on apoptosis and radioresistance. Finally, we have evidence that the induction of a radiation induced G2 delay may be susceptible to pharmacological manipulation using caffeine and staurosporine. We wish to explore whether it is possible using such methods to alter the induction of apoptosis or of the radiosensitivity of the cells. These studies may have clinical utility if they open up new pathways which may be explored to alter the radiosensitivity of tumors or normal tissues.

肿瘤的放射反应性差异很大， 这种变化的一个组成部分是肿瘤固有的放射敏感性 细胞本身。 癌基因可以改变细胞的放射敏感性， 它们影响细胞对辐射反应的机制 仍然未知。 我们已经开始探索癌基因是否影响 辐射后细胞凋亡。 我们发现癌基因Hras和Myc 当用于转化大鼠胚胎细胞（REC）时， 比REC本身或REC具有更大的辐射抗性， 单独的致癌基因。 我们已经证明，这种差异与 辐射诱导的双链DNA断裂的程度;也不是 与这些断裂的重新连接的程度或动力学有关。 的 辐射抗性表型与由辐射诱导的较长G2延迟相关。 辐射 我们最近发现，抵抗细胞对 通过抑制有丝分裂细胞周期蛋白之一细胞周期蛋白的合成来进行辐射 B_1，而敏感细胞的有丝分裂无辐射效应。 cyclin表达。 Myc转染的细胞在转染后发生凋亡。 血清撤回和辐射后。 细胞凋亡明显减少 在相同剂量的 然而，它们在血清撤回后仍然经历凋亡。 因此 ras似乎保护细胞免受辐射诱导的凋亡，但 不能保护细胞免受血清提取诱导的细胞凋亡。 一 在Hela细胞中观察到类似的表型， 辐射诱导的细胞凋亡和类似的延长辐射 诱导G2延迟并抑制细胞周期蛋白B1表达。 这样做的目的 应用是探讨辐射诱导G2的假设， 延迟作为凋亡反应的调节器。 由于过渡 从G2/M期到G4/M期，部分受细胞周期蛋白的调节，并需要细胞周期蛋白的表达 B1，我们提出G2延迟，因此也可能是细胞凋亡。 通过细胞周期蛋白B1的表达调控。 我们打算调查 癌基因对决定细胞凋亡是否发生的贡献， 既通过操纵癌基因表达或活性， 其他基因可能会抵消癌基因对肿瘤细胞的影响， 凋亡途径 我们将使用以下抑制剂来抑制ras的作用： 法尼基化 我们将使用反义技术来改变 有丝分裂细胞周期蛋白的表达，从而改变G2的时间， 检测对细胞凋亡和辐射抗性的影响。 我们终于有 有证据表明，辐射诱导的G2延迟的诱导可能是 易受使用咖啡因的药理学操作的影响， 星形孢菌素。 我们希望探讨是否有可能利用这种 改变细胞凋亡诱导或辐射敏感性的方法 细胞。 这些研究可能具有临床实用性，如果他们开辟了新的 可以探索改变肿瘤放射敏感性的途径， 正常组织