Genotoxicity & Emergence of Genome Instability in Humans

遗传毒性

• 批准号: 7226963
• 负责人: BARRY Alan FINETTE
• 金额: $ 31.96万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7226963
• 关键词: Acute Lymphocytic Leukemia; Address; Antineoplastic Agents; Base Excision Repairs; Biological Assay; Biological Markers; Chromosomal Instability; Chromosome abnormality; Chronic; Clinical; Clonality; Comet Assay; DNA Repair; DNA Repair Pathway; Defect; Development; Disease; Double Strand Break Repair; Event; Evolution; Frequencies; Gene Expression; Gene Expression Microarray Analysis; Gene Mutation; Gene Rearrangement; Genetic; Genetic Predisposition to Disease; Genomic Instability; HPRT1 gene; Hodgkin Disease; Human; Hypoxanthine Phosphoribosyltransferase; Inherited; Lead; Luciferases; Malignant - descriptor; Malignant Neoplasms; Measures; Messenger RNA; Microsatellite Instability; Mismatch Repair; Mutation; Neoplastic Cell Transformation; Non-Malignant; Nucleotide Excision Repair; Pathway interactions; Patient Monitoring; Patients; Plasmids; Population; Predisposition; Proliferating; Protocols documentation; Reporter Genes; Research; Screening procedure; Second Primary Cancers; System; T-Lymphocyte; Testing; Therapeutic; Translating; Ulcerative Colitis; base; beta Chain Antigen T Cell Receptor; drug development; gene environment interaction; gene function; genotoxicity; in vivo; insight; mutant; neoplastic; novel therapeutics; repaired

DESCRIPTION (provided by applicant): The objective of this research is to investigate the association between genotoxic exposure and the emergence of mutagenic mechanisms involved with the development of primary and secondary malignancies in humans. Our hypotheses are: 1) Genotoxic exposure leading to clonal proliferative bursts will result in the in vivo evolution of genomic instability in nonmalignant T cell clones from subjects with cancer or predisposed to cancer; and 2) T cell clones with genomic instability captured from these subjects are the result of altered gene function/expression in DNA repair pathways. Our specific aims are: 1) to isolate and characterize proliferating T cell clones with genomic instability from subjects with acute lymphocytic leukemia, Hodgkin's disease and chronic ulcerative colitis following therapeutic genotoxic exposure; and 2) to perform four functional assays for DNA repair pathways on T cell mutants with genomic instability to determine DNA repair pathway integrity. T cell clones with genomic instability will be isolated and characterized using the HPRT T cell biomarker system. T cell clonality will be determined by T cell receptor beta gene rearrangement mRNA analysis. Mechanistic studies of clones with genomic instability will initially include functional screening assays for DNA repair pathway defects. These assays include: a) microsatellite instability to test for mismatch repair defects; b) single cell gel electrophoresis and c) a plasmid based luciferase DNA repair assay to test for double strand break repair, nucleotide excision repair, and base excision repair capacity following repair specific genotoxic exposures; and d) chromosome instability by measuring frequency of chromosomal aberrations. Subsequent studies will include microarray gene expression and genotypic analysis of DNA repair defects in clones in which a functional DNA repair defect has been established. This research will provide insight into gene-environment interactions, genomic instability and malignant transformation in humans, In addition, these studies could eventually be translated to the clinical setting by leading to: 1) the direct screening of patients for inherited and acquired genetic defects associated with the initiation and progression of malignant transformation; 2) new therapeutic protocols and drug development that decreases the genotoxicity and the development of proliferative clones with genomic instability as a consequence of antineoplastic therapy; and 3) patient monitoring/screening for genetic mutations that are associated with the development of second malignant neoplasms.

描述（由申请人提供）：本研究的目的是调查基因毒性暴露与与人类原发性和继发性恶性肿瘤发展有关的致突变机制之间的关系。我们的假设是：1)基因毒性暴露导致克隆增殖性爆发将导致来自癌症或癌症易感受试者的非恶性T细胞克隆的基因组不稳定性的体内进化；2)从这些受试者身上捕获的基因组不稳定的T细胞克隆是DNA修复途径中基因功能/表达改变的结果。我们的具体目标是：1)从治疗性基因毒性暴露后的急性淋巴细胞白血病、霍奇金病和慢性溃疡性结肠炎患者中分离和表征具有基因组不稳定性的增殖T细胞克隆；2)对基因组不稳定的T细胞突变体进行DNA修复途径的四项功能测定，以确定DNA修复途径的完整性。具有基因组不稳定性的T细胞克隆将被分离并使用HPRT T细胞生物标记系统进行表征。通过T细胞受体β基因重排mRNA分析来确定T细胞的克隆性。基因组不稳定克隆的机制研究将首先包括DNA修复途径缺陷的功能筛选试验。这些检测包括：a)微卫星不稳定性测试错配修复缺陷；b)单细胞凝胶电泳和c)基于质粒的荧光素酶DNA修复试验，以测试修复特异性基因毒性暴露后的双链断裂修复、核苷酸切除修复和碱基切除修复能力；d)通过测量染色体畸变频率来测定染色体不稳定性。后续的研究将包括微阵列基因表达和DNA修复缺陷的克隆基因型分析，其中功能性DNA修复缺陷已经建立。本研究将深入了解基因-环境相互作用、基因组不稳定性和人类恶性转化。此外，这些研究最终可以转化为临床环境，导致：1)直接筛选与恶性转化的开始和进展相关的遗传和获得性遗传缺陷的患者；2)新的治疗方案和药物开发，以降低遗传毒性和由于抗肿瘤治疗导致的基因组不稳定的增殖性克隆的发展；3)患者监测/筛查与二次恶性肿瘤发展相关的基因突变。