CELL HETEROGENEITY AND TUMOR THERAPY

细胞异质性与肿瘤治疗

• 批准号: 2087011
• 负责人: MARK L MEUTH
• 金额: $ 30.12万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 1977
• 资助国家: 美国
• 起止时间: 1977-09-30 至 1994-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2087011
• 关键词: ADP ribosylation; DNA damage; DNA directed DNA polymerase; DNA repair; DNA replication; DNA topoisomerases; X ray; antineoplastics; breast neoplasms; camptothecin; cell growth regulation; cellular oncology; chromatin; clone cells; combination cancer therapy; conformation; cytotoxicity; drug adverse effect; drug interactions; drug metabolism; drug resistance; enzyme activity; genetic transcription; ionizing radiation; laboratory mouse; messenger RNA; multidrug resistance; natural gene amplification; neoplasm /cancer chemotherapy; neoplasm /cancer genetics; neoplasm /cancer radiation therapy; neoplastic growth; northern blottings; physical chemical interaction; radiation sensitivity; radiobiology; southern blotting; tissue /cell culture; transfection

The chromatin/DNA topology enzymes [i.e., poly(ADP-ribose) polymerase, and topoisomerases I and II] are considered to have important roles in DNA synthesis, RNA transcription, mitosis, and DNA-damage repair, and their functions are probably interactive. However, their specific role(s) in the repair of DNA damage from irradiation and drugs is uncertain and indeed controversial. The long term goals of this proposal are the elucidation of their interactive roles in the repair of DNA damage from ionizing radiation and mitomycin-C. The parental V79 hamster cells and two x-ray-sensitive mutants (irs-1 and irs-2) are being used and drug-resistant sublines have been developed to relatively specific inhibitors of these three enzymes [i.e., 3-aminobenzamide (3AB), camptothecin (Cp), and m-AMSA (mA) respectively]. The cellular radiation phenotype (cell survival) of the irs-2(3AB)R and irs-2(Cp)R cells has returned to the parental V79 phenotype while that of the V79(3AB)R is unchanged. The endogenous and x-ray-induced activity (incorp. of 3H-NAD) of the polymerase enzyme in the irs-2(3AB)R and irs-2(Cp)R have been elevated markedly above the parental-cell activity. Studies are being conducted on both the activity and enzyme content (Western blots) of the polymerase enzyme as well as that of the topo enzymes (the activity assays for the topos are alkaline and neutral filter elution respectively for Topo I and II after drug-induced DNA strand breaks). The question of gene-amplification (Southern blot analysis) and increased mRNA transcription or stabilization (Northern blot analysis and specific mRNA transcription rates) are being explored as a mechanism for the induced level of the polymerase enzyme. Also DNA damage induction and repair from mitomycin-C are being compared to the radiobiological data in these drug resistant lines to elevate the generality vs specificity of these altered phenotypes (i.e. both long-patch and short-patch repair). In summary, the critical metabolic aspects of these three nuclear enzymes are being studied "in concert" as they relate to (possibly explain) the altered cellular radiation phenotype (resistance) in the (3AB)R and (Cp)R irs-2 sublines.

染色质/DNA 拓扑酶 [即聚 (ADP-核糖) 聚合酶，和 拓扑异构酶 I 和 II]被认为在 DNA 中具有重要作用 合成、RNA转录、有丝分裂和DNA损伤修复及其它们 功能可能是交互式的。 然而，他们在 辐射和药物造成的 DNA 损伤的修复尚不确定，而且确实如此 有争议的。 该提案的长期目标是阐明 它们在修复电离辐射造成的 DNA 损伤中的相互作用 和丝裂霉素-C。 亲本 V79 仓鼠细胞和两个 X 射线敏感细胞 突变体（irs-1 和 irs-2）正在被使用，耐药亚系已经出现 已开发出这三种酶的相对特异性抑制剂 [即 3-氨基苯甲酰胺 (3AB)、喜树碱 (Cp) 和 m-AMSA (mA) 分别]。 细胞辐射表型（细胞存活） irs-2(3AB)R 和 irs-2(Cp)R 细胞已恢复亲本 V79 表型 而 V79(3AB)R 则没有变化。 内源性和 X 射线诱导的 irs-2(3AB)R 中聚合酶的活性（包含 3H-NAD） 和 irs-2(Cp)R 已显着升高至亲代细胞以上 活动。 正在对活性和酶进行研究 聚合酶的含量（蛋白质印迹）以及 拓扑酶（拓扑酶的活性测定为碱性和中性 药物诱导DNA链后分别过滤洗脱Topo I和II 休息）。 基因扩增问题（Southern印迹分析）和 增加 mRNA 转录或稳定性（Northern blot 分析和 特定的 mRNA 转录率）正在被探索作为一种机制 聚合酶的诱导水平。 还有 DNA 损伤诱导和 丝裂霉素-C 的修复与放射生物学数据进行比较 这些耐药系提高了通用性与特异性 这些改变的表型（即长补丁和短补丁修复）。 在 总之，这三种核酶的关键代谢方面是 被“一致地”研究，因为它们与（可能解释）改变的 (3AB)R 和 (Cp)R irs-2 中的细胞辐射表型（抗性） 子线。