IN VIVO SOMATIC MUTATIONS IN CHILDREN WITH CANCER

癌症儿童体内体细胞突变

• 批准号: 6173638
• 负责人: BARRY Alan FINETTE
• 金额: $ 16.12万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-30 至 2002-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6173638
• 关键词: DNA damage; Hodgkin's disease; T cell receptor; acute lymphocytic leukemia; adolescence (12-20); cell transformation; child (0-11); clinical research; gene expression; gene frequency; human subject; mutagens; neoplasm /cancer genetics; neuroblastoma; nucleic acid sequence; p53 gene /protein; pediatric neoplasm /cancer; restriction fragment length polymorphism; sarcoma; tumor suppressor genes

DESCRIPTION (Applicant's Description): The primary objective of this proposal is to complete a 3 component career development plan that will result in the applicant's acquiring the necessary skills and experience to develop an independent translational research program investigating the genetic mechanisms responsible for malignant transformation in children. The 3 components of this plan are: 1) a one-year co-mentorship with two senior cancer researcher scientists who will provide him with the guidance and expertise required to complete the academic and research portion of his training: 2) a didactic program that focuses on specific areas of cancer biology; and 3) a transition segment from colleague/mentor to independent research scientist and collaborator during the independent research portion of his proposal. The principal research objectives of this proposal are to investigate the molecular mechanisms and biologic consequences of in vivo somatic genetic events responsible for pediatric malignancies. The hypothesis of the applicants is that somatic mutational events in children with cancer will occur at a higher frequency and with a unique mutational spectra compared to a normal population. The specific aims which will test the proposed hypotheses are: 1) to determine the frequency and mutational spectra of background and t h erapy induced in vivo somatic mutations in children with specific malignancies (acute lymphocytic leukemia [ALL], Hodgkin's disease, neuroblastoma, and sarcomas) and where possible, to correlate these molecular events with subsequent diseases: 2) to determine if "illegitimate" V(D)J recombinase mediated mutations occur at a known cancer gene, p53; and 3) to isolate and identify T-lymphocyte imitator phenotype clones from children with relapsed ALL who have an extremely high frequency of background somatic mutations. The frequency of somatic mutational events will be determined by the hprt T-cell cloning assay. Mutational spectra of hprt and p53 mutations will be determined by a variety of methods including multiplex PCR, RT-PCR, IPCR, Southern blotting and DNA sequencing. Clonality of hypermutable clones will be determined by RFLP analysis of TCR gamma and CDR3 region DNA sequence analysis of TCR-beta. These studies will provide the first data on the frequency and mutational spectrum of in vivo spontaneous or treatment induced somatic mutations in children with specific malignancies. The investigators will also begin to determine the potential mutagenic effects associated [with] background "illegitimate" V(D)J recombinase mediated events in a known cancer gene, p53. In addition, the isolation of mutator phenotype clones will allow for future m e chanistic studies characterizing the genetic and biologic defect(s) associated with leukemogenesis in children.

描述（申请人的描述）： 该提案的主要目标是完成由 3 个部分组成的职业生涯 发展计划将使申请人获得必要的 开展独立转化研究的技能和经验 研究恶性遗传机制的计划 儿童的转变。 该计划的 3 个组成部分是：1) 与两名高级癌症研究科学家进行为期一年的共同指导，他们将 为他提供完成所需的指导和专业知识 他的培训的学术和研究部分：2）一个教学计划 专注于癌症生物学的特定领域； 3) 过渡段 从同事/导师到独立研究科学家和合作者 在他的提案的独立研究部分期间。 本提案的主要研究目标是调查 体内体细胞遗传的分子机制和生物学后果 导致儿科恶性肿瘤的事件。 的假设 申请人认为，癌症儿童的体细胞突变事件将 与相比，发生频率更高且具有独特的突变谱 正常人群。 将测试所提议的具体目标 假设是：1）确定频率和突变谱 背景和治疗诱导的儿童体内体细胞突变 特定恶性肿瘤（急性淋巴细胞白血病 [ALL]、霍奇金病、 神经母细胞瘤和肉瘤），并在可能的情况下将这些关联起来 与后续疾病相关的分子事件：2) 确定是否 “非法”V(D)J 重组酶介导的突变发生在已知癌症中 基因，p53； 3) 分离和鉴定 T 淋巴细胞模仿表型 来自复发率极高的 ALL 儿童的克隆 背景体细胞突变。 体细胞突变事件的频率将由 hprt 决定 T 细胞克隆测定。 hprt 和 p53 突变的突变谱将是 通过多种方法测定，包括多重PCR、RT-PCR、IPCR、 Southern 印迹和 DNA 测序。 超突变克隆的克隆性将 通过 TCR gamma 和 CDR3 区 DNA 序列的 RFLP 分析来确定 TCR-β 分析。 这些研究将提供有关频率和突变的第一批数据 体内自发或治疗诱导的体细胞突变谱 患有特定恶性肿瘤的儿童。 调查人员也将开始 确定与背景相关的潜在诱变效应 已知癌症基因中“非法”V(D)J 重组酶介导的事件， 第 53 页。 此外，突变表型克隆的分离将允许 未来表征遗传和生物学特征的机制研究 与儿童白血病发生相关的缺陷。