CHEMICAL TRANSFORMATION NEOPLASTIC PROGESSION & ONCOGENE

化学转化肿瘤进展

• 批准号: 3174945
• 负责人: LEILA DIAMOND
• 金额: $ 16.27万
• 依托单位: WISTAR INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-04-01 至 1990-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3174945
• 关键词: athymic mouse; benzopyrenes; chemical carcinogen; chemical carcinogenesis; gel electrophoresis; genetic translation; hamsters; oncogenes; radiotracer; tissue /cell culture; transfection

Cancer is the end result of a multistage, multifactorial process. The assays for chemical transformation of Syrian hamster embryo fibroblasts (HE) and hamster epidermal cells are excellent models in which to study various factors that may be associated with specific stages in transformation, from the first morphologic changes induced by a carcinogen through progression to the neoplastic state. The objective of this proposal is to determine whether altered expression or activation of cellular ras genes plays a role in chemical transformation of hamster cells, and whether such alterations are associated with any specific stage during the progression from morphologic to neoplastic transformation. The c-ras gene which is most frequently activated in neoplastic transformants of HE and hamster epidermal cells initiated with benzo(a)pyrene (B(a)P) will be determined, using DNA transfection into NIH/3T3 cells and restriction enzyme analysis of transforming activity and of hybridization patterns with specific ras probes. The normal hamster homolog of the activated ras gene will be molecularly cloned, and the coding regions containing the codons for amino acids 12 and 61 subcloned and sequenced. The normal gene sequences subcloned into a single-stranded DNA vector will be used as probes for genomic sequencing of the activated genes in the neoplastic transformants and of homologous regions in their preneoplastic progenitors. It will then be determined when, after treatment of primary cells with B(a)P and clonal isolation of morphologically altered HE cell transformants or differentiation-resistant epidermal cell transformants, a mutation in a ras gene can first be detected by a biochemical test for restriction polymorphism or genomic sequencing of the DNA. We will then determine when, in relation to the progression from morphologic to neoplastic transformation, that mutated ras gene is expressed, as measured by its transfectability into 3T3 cells and/or altered production of p21. These experiments can determine if ras gene activation is or is not a primary event in chemical transformation of hamster cells.

癌症是一个多阶段、多因素过程的最终结果。 的 叙利亚仓鼠胚胎成纤维细胞的化学转化试验 (HE)和仓鼠表皮细胞是研究 可能与特定阶段相关的各种因素， 由致癌物引起的第一次形态学变化 发展到肿瘤状态。 的目的 建议是确定是否改变表达或激活 细胞ras基因在仓鼠化学转化中的作用 细胞，以及这些变化是否与任何特定阶段有关 在从形态学到肿瘤转化的过程中。 的 在肿瘤转化体中最常被激活的c-ras基因 苯并（a）芘（B（a）P）引发的HE和仓鼠表皮细胞 将使用DNA转染到NIH/3 T3细胞中来确定， 转化活性和杂交的限制酶分析 用特异性ras探针检测。 正常的仓鼠同源体 激活的ras基因将被分子克隆， 包含亚克隆和测序的氨基酸12和61的密码子。 亚克隆到单链DNA载体中的正常基因序列将 作为探针，用于对细胞中的活化基因进行基因组测序， 肿瘤转化体和其肿瘤前的同源区域 祖先 然后将确定在治疗原发性 B（a）P细胞和形态学改变的HE细胞的克隆分离 转化体或分化抗性表皮细胞转化体， ras基因中的突变可以首先通过生化试验检测， DNA的限制性多态性或基因组测序。 然后我们将 确定何时，与从形态学到 肿瘤转化，突变的ras基因表达，如测量的 通过其可转染到3 T3细胞和/或改变p21的产生。 这些实验可以确定ras基因激活是否是 仓鼠细胞化学转化的主要事件。