ELA COOPERATIVE ONCOGENE PRODUCTS IN HUMAN TUMOR CELLS

描述：鉴定改变的生长调节基因 通过碱基突变、基因扩增或重排是一个主要的焦点 癌症研究。 对于大多数人类实体肿瘤，包括 乳腺癌，没有发现这样的癌基因改变。 细胞遗传学和染色体都强烈地表明了基因的存在 杂交研究 对于乳腺癌，17 q22位点， 20 q13、1 q32、6 q21和19 q似乎特别有可能孕育新的 致癌基因 以前的体外研究表明，许多癌基因在两个 的方式 有些基因（如RAS）足以转化永生化的 细胞系，而其它细胞系（例如，C-MYC形成致瘤病灶， 转化细胞只与其他癌基因合作。大 第一类基因的数量已经被直接鉴定， 用肿瘤DNA转染永生化NIH 3 T3细胞（例如，RAS， HER-2/neu）。 然而，没有检测存在直接分离协同 致癌基因 一个主要的限制是转染的效率 用于合适的细胞类型，例如原代啮齿动物细胞。 并不是所有的第一类基因都可以用NIH 3 T3细胞分离 是由几个已知的癌基因的行为，包括GIP 2; BCR-ABL、GL 1和Wilm肿瘤基因WT-1的激活等位基因。 这些基因不能转化NIH 3 T3细胞，但能有效地转化 永生化大鼠细胞，包括用腺病毒永生化的那些细胞 E1a 这种特别有效的癌基因通过结合几种 细胞蛋白质，包括肿瘤抑制因子p105 Rb， 与其他基因，包括RAS，GL 1，WT-1或E1 b， 原代啮齿动物细胞的转化。 在这项提案中，研究人员描述了一种有效的方法， 使用腺病毒-聚赖氨酸缀合物作为载体的稳定转染 转染试剂 研究人员引入了质粒乳房 癌症cDNA表达文库中的一个良好的特点，E1 a- 永生化大鼠肾细胞系。 与对照质粒相比， 该文库诱导了几个转化细胞的大病灶。 对这些克隆的基因组DNA的初步鉴定表明， 不涉及已知的癌基因RAS或HER-2/neu。 的 研究人员建议确定和描述新的E1 A合作社， 在这些细胞系中的致癌基因，并测试基因的遗传 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.