MOLECULAR GENETIC ANALYSIS OF COLORECTAL CANCER

结直肠肿瘤提供了一个独特的研究分子的机会。 一种常见人类肿瘤发生和发展的相关事件 打字。以前的研究表明，结直肠肿瘤的发生是由 通过癌基因和肿瘤抑制基因的顺序突变。尽管 这些基因的突变已经有了很好的记录，关于 这些突变对结直肠肿瘤生物学和生理学的影响 细胞是原始的。我们计划利用最近开发的技术 在我们的实验室，以及经典的遗传和生化方法， 进一步研究被认为在结直肠癌中重要的三条途径 肿瘤： L。P53-P53基因在大多数结直肠肿瘤中失活，以及 在许多其他类型的人类肿瘤中。P53基因产物被认为是 功能的一部分，是通过激活受 特定的P53结合DNA序列。一种新的基因分析方法 表达模式(SAGE)将用于识别 在经历生长停滞或凋亡的结直肠癌细胞中被激活 以响应P53的表达。测试基因产品的重要性 通过这种方式鉴定，相关基因将被同源基因扰乱 重组和工程化细胞的评估 对P53和其他生长调节剂的反应。 2.转化生长因子-β--大多数结直肠肿瘤对这种阴性反应不敏感 生长调节因子，以及参与这一途径的基因突变 在大肠肿瘤的一个子集中被发现。其他人类基因 可能参与的途径将通过同源克隆来确定 方法：研究方法。然后将对这些基因进行评估，以确定它们是否 在结直肠肿瘤中发生突变。那些被发现被更改的将被中断 通过在适当的结直肠肿瘤细胞系中进行同源重组和 评估它们对转化生长因子-β信号的影响。 错配修复-参与错配修复(MMR)的四个基因 被证明在以下情况下会导致结直肠肿瘤的遗传不稳定 以突变的形式遗传的。这些突变的一个子集与 不寻常的表型或用标准化验无法检测到。我们计划 开发基因系统来评估这种MMR基因变异体的功能。 其中一些系统将涉及对基因的靶向破坏 合适的受体细胞类型。从这项研究中获得的信息 应说明MMR的某些基本方面，并提供新的 诊断机会。

Colorectal tumors provide a unique opportunity to study the molecular events responsible for initiation and progression of a common human tumor type. Previous studies have shown that colorectal tumorigenesis is driven by sequential mutations of oncogenes and tumor suppressor genes. Though mutations in these genes have been well documented, knowledge about the effect of such mutations on the biology and physiology of colorectal tumor cells is rudimentary. We plan to exploit technologies recently developed in our laboratory, as well as classic genetic and biochemical methods, to further investigate three pathways thought to be important in colorectal neoplasia: l. p53 - The p53 gene is inactivated in most colorectal tumors, as well as in many other human tumor types. The p53 gene product is thought to function, in part, by activating the expression of genes controlled by specific p53-binding DNA sequences. A novel method for analyzing gene expression patterns (SAGE) will be used to identify genes that are activated in colorectal tumor cells undergoing growth arrest or apoptosis in response to p53 expression. To test the importance of gene products identified in this way, the relevant genes will be disrupted by homologous recombination and the engineered cells assessed with respect to their response to p53 and other growth-regulating agents. 2. TGF-beta - Most colorectal tumors are insensitive to this negative growth regulator, and mutations in genes participating in this pathway have been identified in a subset of colorectal tumors. other human genes that may participate in the pathway will be identified by homologous cloning methods. Such genes will then be evaluated to determine whether they are mutated in colorectal tumors. Those found to be altered will be disrupted by homologous recombination in appropriate colorectal tumor cell lines and assessed for their impact on TGF-beta signaling. 3. Mismatch repair - Four genes participating in mismatch repair (MMR) have been shown to lead to genetic instability in colorectal tumors when inherited in mutant form. A subset of these mutations are associated with unusual phenotypes or are undetectable by standard assays. We plan to develop genetic systems to evaluate the function of such MMR gene variants. Some of these systems will involve targeted disruption of the genes in suitable recipient cell types. The information derived from this study should illuminate certain basic aspects of MMR as well as provide new diagnostic opportunities.