RADIORESISTANCE, APOPTOSIS AND THE CELL CYCLE

辐射抗性、细胞凋亡和细胞周期

基本信息

批准号：
2443090
负责人：
WILLIAM G. MCKENNA
金额：
$ 23.25万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
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

项目摘要

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.

肿瘤的辐射措施差异很大，一种重要该变化的成分是肿瘤的固有放射敏性细胞本身。癌基因可以改变细胞的放射敏性，但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凋亡。我们发现Oncogenes Hras和Myc 当用于转化大鼠胚细胞（REC）时，会导致细胞系比本身或与Rec一起进行的放射线比大大要大得多要么单独使用过癌基因。我们已经表明，这种差异与辐射引起的双链DNA断裂的程度；也不是与这些断裂重新加入的程度或动力学有关。这放射线抗性表型与较长的G2延迟相关辐射。我们最近表明，抗性细胞对通过降低有丝分裂细胞周期蛋白之一的合成来辐射。 B1，而敏感细胞没有辐射对有丝分裂的影响细胞周期蛋白的表达。 MYC转染的细胞在血清抽出和辐射后。凋亡被认为较少相同剂量的RAS转染细胞培养物的程度辐射尚未在血清戒断后仍会凋亡。因此 RA似乎可以保护细胞免受辐射诱导的凋亡，但不能保护细胞免受血清戒断引起的凋亡。一个在HeLa细胞中也可以看到类似的表型辐射诱导的凋亡，同样的辐射延长诱导的G2延迟降低了细胞周期蛋白B1表达。这个目的应用是探讨辐射诱导G2的假设延迟是凋亡反应的调节因子。自从过渡以来通过G2/m的部分调节，需要细胞周期蛋白的表达 B1，我们建议G2延迟，因此可能是凋亡通过细胞周期蛋白B1表达调节。我们打算调查癌基因确定凋亡是否发生的贡献，通过操纵癌基因表达或活动以及转染其他可能对抗癌基因对的基因凋亡途径。我们将使用的抑制剂抑制RAS作用法尼化。我们将使用反义技术改变有丝分裂细胞周期蛋白的表达，因此改变了G2的时机测试对细胞凋亡和放射性的影响。最后，我们有证据表明辐射诱导的G2延迟可能是易于使用咖啡因和星形孢菌素。我们希望探索是否可以使用这种改变凋亡诱导或放射敏感性的方法细胞。如果这些研究开放了新的，则可能具有临床实用性可以探索以改变肿瘤或正常组织。