DNA REPAIR AND COLON CANCER

DNA 修复和结肠癌

• 批准号: 6344750
• 负责人: MARK L MEUTH
• 金额: $ 10.06万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-08-16 至 2001-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6344750
• 关键词: DNA repair; alleles; apoptosis; cell cycle; cell line; colon neoplasms; complementary DNA; gene expression; gene induction /repression; gene mutation; pathologic process; phenotype; tissue /cell culture

Colon carcinogenesis is a multi-step process dependent upon the accumulation of mutations in tumor suppressors and proto-oncogenes. Recent work indicates that an inherited form of colon cancer (HNPCC) the accumulation of mutations may be driven by the loss of mismatch repair. Cells losing this repair pathway have a distinctive mutator phenotype including microsatellite instability, a high rate of spontaneous mutation, and resistance to DNA alkylating agents. Our understanding of the biological consequences of mismatch repair deficiency stems largely from studies of colon cancer cell lines. However, these lines have many other alterations that could contribute to the mutation phenotype. In Specific Aims 1 and 2 the hypothesis that mismatch repair deficiency is sufficient for induction of the mutation phenotype seen in these colorectal carcinoma cell lines will be tested. In Aim 1 the ability of inducible cDNA expression constructs of the mismatch repair genes to correct the mutator phenotype in repair deficient colon cancer cell lines will be investigated. Dominant negative alleles of the mismatch repair genes that can induce a mutator phenotype in repair proficient cell lines will be sought in Aim 2. Expression constructs of mutant forms of these mismatch repair cDNAs will be introduced into repair proficient cell lines (in particular non-tumorigenic cell lines of epithelioid origin) to determine the effects on spontaneous mutation rate, cell cycle checkpoint controls, and apoptosis. The strength of this approach is the use of isogenic repair proficient and deficient cell lines to determine effects on the determinants of genome stability and cell growth. In Specific Aim 3 the lethal effects of over-expression of the mismatch repair cDNAs will be examined by microinjection of wild type and mutant constructs. This will test the hypothesis that the mismatch repair genes are involved in cell cycle regulation. In Specific Aim 4 the role of the mismatch repair homologs in the repair process will be determined by the manipulation of these genes in cultured cells. Long term objectives are reflected in Specific Aim 5 where we will determine whether mutations of these genes in cultured cells. Long term objectives are reflected in Specific Aim 5 where we will determine whether mutations of other repair genes contribute to inherited forms of colon cancer. These studies should significantly improve our understanding of the role of mismatch repair deficiency in early events of colon carcinogenesis.

结肠癌发生是一个多步骤的过程 肿瘤抑制剂和原始基因中突变的积累。最近的 工作表明遗传形式的结肠癌（HNPCC） 突变的积累可能是由不匹配修复的损失驱动的。 失去此修复途径的细胞具有独特的突变器表型 包括微卫星的不稳定性，自发突变率很高 和对DNA烷基化剂的抗性。我们对 不匹配修复缺乏的生物学后果主要来自 结肠癌细胞系的研究。但是，这些线还有许多其他 可能有助于突变表型的改变。具体 目的1和2的假设是不匹配维修缺乏就足够了 为了诱导这些结直肠癌中看到的突变表型 细胞系将进行测试。在AIM 1中诱导cDNA的能力 不匹配修复基因的表达构造以纠正突变器 修复缺陷的结肠癌细胞系中的表型将是 调查。不匹配修复基因的主要负等位基因 可以在修复熟练的细胞系中诱导突变器表型 在目标2中寻求的。这些不匹配的突变形式的表达构造 维修cDNA将被引入维修熟练的细胞系中（在 上皮细胞起源的特定非肿瘤细胞系以确定 对自发突变率，细胞周期检查点对照的影响， 和凋亡。这种方法的强度是使用等源性修复 熟练且缺陷的细胞系来确定对 基因组稳定性和细胞生长的决定因素。在特定目标3中 不匹配维修cDNA过表达的致命作用将是 通过对野生型和突变构建体的显微注射进行检查。这会 测试了不匹配修复基因与细胞有关的假设 周期调节。在特定目标4中，不匹配维修的作用 维修过程中的同源物将通过操纵 这些基因在培养的细胞中。长期目标反映在 特定目标5我们将确定这些基因的突变是否在 培养的细胞。长期目标反映在特定目标5中 我们将确定其他维修基因的突变是否有助于 结肠癌的遗传形式。这些研究应该显着 提高我们对不匹配维修缺陷作用的理解 结肠癌发生的早期事件。