CHARACTERIZATION OF DIFFERENTIAL GENE EXPRESSION IN RESISTANT CELLS

接受放射治疗的癌症患者的细胞系已经被 与临床结果和体外放射反应相关。那 一些组织结构相似的细胞，尤其是来自其患者的 体外实验显示，治疗未能局部控制他们的肿瘤 抗辐射类型的辐射存活参数，而其他类型的辐射存活参数 敏感型，暗示辐射敏感性的遗传基础。 这个项目的总体目标是识别可能是 与人类肿瘤的抗辐射能力有关。使用 蛋白质分析技术(双向凝胶电泳法和 微测序)差异筛选/消减cDNA文库分析， 和载体介导的基因转移，规划了一种多途径策略 在第一个目的是识别和分离与基因相关的基因 耐辐射表型。我们将检验这一假设 细胞中差异基因的表达是辐射的原因 肿瘤细胞的敏感或抗辐射表型。 在该项目的第二个目标中，识别基因的两种策略 与阻力相关的将被用来改变辐射响应 利用载体介导的基因转移技术获得敏感的肿瘤细胞。第一 将使用一种包含来自 抗辐射细胞，第二个将使用粘粒策略 利用抗性细胞中较大的DNA片段进行基因转移。 将对辐射敏感的人类肿瘤细胞进行转基因 并进行表型选择，以允许随后的救援 DNA片段。大豆杉差异表达基因的鉴定 抗辐射细胞将提供对肿瘤机制的洞察 抗辐射能力，可能成为未来基于组织的预测的基础 测试。

Cell lines from cancer patients who undergo radiation therapy have been correlated to clinical outcomes and to in vitro radiation responses. That some cells of similar histologies, particularly from patients whose treatments have failed to locally control their tumors, show in vitro radiation survival parameters of a resistant type, while others are of a sensitive type, suggests a genetic basis for radiation sensitivity. The overall objective of this project is to identify genes that may be associated with the radiation resistance of human tumors. Using the techniques of protein analysis (2-D gel electrophoresis and microsequencing) differential screening/subtraction cDNA library analysis, and vector mediated gene transfer, a multiple approach strategy is planned in the first AIM to identify and isolate genes associated with the radiation-resistant phenotype. We will test the hypothesis that differential gene expressions in cells are responsible for the radiation sensitive or the radiation resistant phenotypes of tumor cells. In the second AIM of this project, two strategies to identify genes associated with resistance will be used to modify the radiation responses of sensitive tumor cells using vector mediated gene transfer. The first will use an episomally maintained vector strategy containing cDNA from radiation resistant cells and the second will use a cosmid strategy for gene transfer using larger fragments of DNA from resistant cells. Transfections will be performed into radiation sensitive human tumor cells and phenotypic selection will be performed to permit subsequent rescue of the DNA fragments. The identification of differentially expressed genes in radiation resistant cells will provide insight into the mechanisms of tumor radiation resistance and may be a future basis for tissue based predictive tests.