Mechanism /Consequence of Chromosome Aneuploidy/Transloc

染色体非整倍体/易位的机制/后果

• 批准号: 6558744
• 负责人: Thomas E. Ried
• 金额: --
• 依托单位: DIVISION OF CLINICAL SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6558744
• 关键词: adenoma; aneuploidy; animal tissue; carcinoma; cell line; cell transformation; chromosome movement; chromosome translocation; colorectal neoplasms; cytogenetics; gel electrophoresis; genetically modified animals; green fluorescent proteins; karyotype; laboratory mouse; microarray technology; molecular genetics; neoplasm /cancer genetics

The review of CGH analyses performed in our laboratory has revealed that the acquisition of whole chromosomes or chromosome arm gains and losses is a frequent event, particularly at early stages of carcinogenesis. The comparison of 25 cell lines established from solid tumors by SKY and CGH has provided compelling evidence that the vast majority of epithelial cancers have either numerical or structural chromosomal aberrations resulting in genomic imbalances. The comparison of diploid, mismatch repair deficient colorectal carcinomas with aneuploid ones indicates that numerical chromosomal aberrations are 60 times more prevalent in the aneuploid tumors. Abnormalities of the centrosome correlated with chromosomal aneuploidy both in human cancer cell lines and in animal cells deficient for cell cycle regulator genes such as p53 and BRCA1. All these results taken together support the notion that chromosomal aneuploidy is a major theme in epithelial cancers. In order to elucidate mechanisms leading to aneuploidy, to establish the functional relevance of chromosomal aneuploidy, and to identify whether aneuploidy is a cause or a consequence of genetic changes in solid tumors, we have focused on the following projects: - Structural and functional analysis of abnormal centrosomes and their relationship to chromosome segregation fidelity. - Live cell imaging of clonally derived cells after transfection with g-tubulin green fluorescent protein (GFP) and CenPB-GFP vectors. - Sequential inhibition of p53 and Rb-function with E6 and E7-genes from human papilloma virus 16 and subsequent analyses of chromosomal instability. - Microcell mediated chromosome transfer of chromosomes that are frequently gained in colorectal carcinomas into cells derived from normal colorectal epithelium and adenomas followed by cytogenetic analysis and assays for cellular immortalization and transformation. - In vitro induction of chromosomal aneuploidy in primary cultures derived from normal murine cells of different organ origin with aneugens followed by cytogenetic analysis and assays for confirmation of cellular immortalization and transformation. - Comparison of cells carrying chromosomal aneuploidies with their wildtype parental cells using cDNA microarrays and 2D-gel protein electrophoresis.

对我们实验室进行的CGH分析的回顾表明，获得整个染色体或染色体臂的获得和丢失是一种频繁的事件，特别是在癌症发生的早期阶段。Sky和CGH从实体肿瘤建立的25个细胞系的比较提供了令人信服的证据，表明绝大多数上皮性癌症存在数量或结构染色体异常，导致基因组失衡。二倍体错配修复缺陷结直肠癌与非整倍体结直肠癌的比较表明，染色体数目异常在非整倍体肿瘤中的发生率是非整倍体肿瘤的60倍。中心体的异常与染色体非整倍体有关，在人类癌细胞系和缺乏细胞周期调节基因的动物细胞中，如P53和BRCA1。所有这些结果综合起来支持染色体非整倍体是上皮性癌症的一个主要主题的观点。为了阐明导致非整倍体的机制，建立染色体非整倍体的功能相关性，并确定非整倍体是实体瘤遗传变化的原因还是结果，我们集中在以下项目： -异常中心体的结构和功能分析及其与染色体分离保真度的关系。 -将g-微管蛋白绿色荧光蛋白(GFP)和CenPB-GFP载体导入克隆来源的细胞后进行活细胞成像。 -用来自人类乳头瘤病毒16的E6和E7基因顺序抑制P53和Rb功能，以及随后对染色体不稳定性的分析。 -微细胞介导将结直肠癌中经常获得的染色体转移到来自正常结直肠上皮和腺瘤的细胞中，然后进行细胞遗传学分析和细胞永生化和转化的分析。 -体外诱导来源于不同器官来源的正常小鼠细胞的染色体非整倍体，然后进行细胞遗传学分析和确认细胞永生化和转化的试验。 -使用基因芯片和2D-Gel蛋白电泳法比较携带染色体非整倍体的细胞及其野生型亲代细胞。