ELA COOPERATIVE ONCOGENE PRODUCTS IN HUMAN TUMOR CELLS

描述：鉴定被改变的生长调节基因 碱基突变、基因扩增或重排是一个主要焦点。 癌症研究的最高成就。对于大多数人类实体肿瘤，包括 乳腺癌，目前还没有发现这种癌基因突变。这样的话 基因的存在是由细胞遗传学和染色体强烈提示的 杂交研究。乳腺癌的基因座位于17q22， 20q13、1q32、6q21和19q似乎特别有可能包含新的 致癌基因。 先前的体外研究表明，许多癌基因在两种中的一种中起作用。 方式。一些基因(如RAS)足以转化成永生的 细胞系，而其他(例如，C-MYC)形成致瘤的焦点， 只有在与其他癌基因合作的情况下才能转化细胞。大号 第一类基因的数量已经被直接通过 用肿瘤DNA(例如，RAS， HER-2/neu)。然而，目前还没有直接分离合作社的方法。 致癌基因。一个主要的限制是转基因的效率。 对于合适的细胞类型，如原代啮齿动物细胞。 并不是所有第一类基因都能用NIH3T3细胞分离出来 由几个已知癌基因的行为提示，包括GIP2； 肾母细胞瘤基因WT-1的bcr-abl、GL1和激活等位基因。 这些基因不能转化NIH3T3细胞，但能有效转化 永生化的大鼠细胞，包括用腺病毒永生化的细胞 E1a。这种特别强大的癌基因通过结合几个 细胞蛋白，包括肿瘤抑制基因p105Rb，并与 与包括RAS、GL1、WT-1或E1b在内的其他基因一起提供完整的 原代啮齿动物细胞的转化。 在该提案中，调查人员描述了一种高效、 用腺病毒-多赖氨酸偶联物作为靶向的稳定转染法 转染剂。研究人员引入了一种乳房质粒 将癌基因表达文库构建成功能完善的E1a- 永生化大鼠肾细胞系。与对照质粒不同， 该文库诱导了几个大的转化细胞灶。 对这些克隆的基因组DNA的初步鉴定表明 已知的癌基因RAS或HER-2/neu不参与。这个 调查人员建议识别和表征新的E1a-合作社 这些细胞系中的癌基因，并测试这些基因的 乳腺癌的改变。

DESCRIPTION: Identification of growth regulatory genes that are altered by base mutation, gene amplification, or rearrangement is a major focus of cancer research. For the majority of human solid tumors, including breast cancer, no such oncogene alteration has been found. That such genes exist is strongly suggested by cytogenetic and chromosome hybridization studies. For carcinoma of the breast, loci at 17q22, 20q13, 1q32, 6q21, and 19q seem especially likely to harbor new oncogenes. Previous in vitro studies show that many oncogenes function in one of two ways. Some genes (e.g. RAS) are sufficient to transform immortalized cell lines, whereas others (e.g., C-MYC form foci of tumorigenic, transformed cells only in cooperation with other oncogenes. A large number of genes of the first class have been directly identified by transfection of immortalized NIH3T3 cells with tumor DNA (e.g., RAS, HER-2/neu). However, no assay exists to directly isolate cooperative oncogenes. A major limitation has been the efficiency of transfection for appropriate cell types such as primary rodent cells. That not all genes of the first class can be isolated using NIH3T3 cells is suggested by the behavior of several known oncogenes, including GIP2; BCR-ABL, GL1, and activated alleles of the Wilm's tumor gene, WT-1. These genes fail to transform NIH3T3 cells, but efficiently transform immortalized rat cells, including those immortalized with the adenovirus E1a. This particularly potent oncogene functions by binding several cellular proteins, including the tumor suppressor p105Rb, and cooperates with other genes including RAS, GL1, WT-1, or E1b to give complete transformation of primary rodent cells. In this proposal the investigators describe a method for efficient, stable transfection that uses adenovirus-polylysine conjugates as a transfection reagent. The investigators introduced a plasmid breast cancer cDNA expression library into a well-characterized, E1a- immortalized rat kidney cell line. In contrast to control plasmids, this library induced several large foci of transformed cells. Preliminary characterization of genomic DNA from these clones shows that the known oncogenes RAS or HER-2/neu are not involved. The investigators propose to identify and characterize new E1A-cooperative oncogenes in these cell lines and to test the genes for genetic alteration in breast cancer.