CHEMICAL TRANSFORMATION NEOPLASTIC PROGESSION & ONCOGENE

癌症是一个多阶段、多因素过程的最终结果。这个 叙利亚仓鼠胚胎成纤维细胞的化学转化方法 (He)和仓鼠表皮细胞是很好的研究模型 可能与以下特定阶段相关的各种因素 从致癌物引起的第一次形态变化开始的转化 通过进展到肿瘤状态。这样做的目的是 建议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，以及这种改变是否与任何特定的阶段有关 在从形态向肿瘤转化的过程中。这个 肿瘤转化中最常被激活的c-ras基因 苯并(A)芘(B(A)P)诱发的HE和仓鼠表皮细胞 通过将DNA导入NIH/3T3细胞和 转化活性和杂交活性的限制性内切酶分析 带有特定ras探针的图案。的正常仓鼠同源物 激活的ras基因将被分子克隆，编码区 包含氨基酸12和61的密码子，并进行亚克隆和测序。 将正常基因序列亚克隆到单链DNA载体中将 被用作基因组测序的探针 肿瘤转化子及其癌前病变中的同源区 祖先。然后将确定何时，在治疗完原发 B(A)P细胞与形态改变的HE细胞的克隆分离 转化子或耐分化表皮细胞转化子，a Ras基因的突变可以首先通过生化测试来检测 DNA的限制性多态或基因组测序。到时候我们会的 确定与形态演变相关的时间 肿瘤转化，根据测量，突变的ras基因被表达。 通过其对3T3细胞的可转染性和/或改变p21的产生。 这些实验可以确定ras基因的激活是否是一种 仓鼠细胞化学转化中的主要事件。

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.