Genomic Instability in Ageing and Cancer

衰老和癌症中的基因组不稳定性

• 批准号: 8195850
• 负责人: David J. Araten
• 金额: --
• 依托单位: VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-10-01 至 2013-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8195850
• 关键词: Address; Age; Aging-Related Process; Antioxidants; Biological; Blood specimen; Caring; Cell Line; Cell division; Cells; Chemoprevention; Clinical Chemoprevention; DNA biosynthesis; Data; Dehydroascorbic Acid; Demographic Aging; Drug Delivery Systems; Elderly; Erythrocytes; Farnesyl Transferase Inhibitor; Flow Cytometry; Frequencies; Genes; Genomic Instability; Human; Incidence; Individual; Intervention; Investigation; Lamin Type A; Life; Link; Lymphoid Cell; Malignant Neoplasms; Measures; Membrane Proteins; Methods; Mutation; Neonatal; Normal Cell; Patients; Pharmaceutical Preparations; Phenotype; Population; Predisposition; Premature aging syndrome; Probability; Process; Progeria; Reactive Oxygen Species; Recruitment Activity; Screening procedure; Sentinel; Somatic Mutation; Syndrome; T-Lymphocyte; Umbilical Cord Blood; United States; Veterans; X Chromosome; age group; age related; base; cancer cell; cancer risk; cohort; granulocyte; high risk; lymphoblastoid cell line; mutant; older patient; prevent; public health relevance; trend; volunteer

Background: In humans, measuring the frequency (f) of cells harboring a mutation is possible for a very small number of sentinel genes and historically has been technically difficult. Measuring the mutation rate (¿)--which represents the probability of new mutations occurring in a gene per cell division-- has been virtually impossible. Nevertheless, f and ¿ may be key parameters in ageing. For example, it is hypothesized: (i) that ¿ increases with age; (ii) that ¿ is elevated in certain premature ageing syndromes; and (iii) that it might be possible to reduce ageing related cancers by decreasing ¿. We have now developed a robust method to measure these parameters using the PIG-A gene, which we hope will greatly facilitate the investigation of these hypotheses. PIG-A is on the X-chromosome- therefore a single inactivating mutation can produce the mutant phenotype. PIG-A mutants lack all GPI-linked membrane proteins, facilitating screening for rare cells with the GPI-null phenotype by flow cytometry. By this approach, we have been able to measure f in granulocytes, and both ¿ and f in B lymphoblastoid cell lines and expanding cultures of ex vivo T lymphocytes from blood samples. Recently, we have shown that it is possible to measure f in human red cells in an additional sentinel gene, XK, which has similar advantages as PIG-A. Specific Aims (a): to determine whether ¿ increases in humans as they age; (b): to determine whether it would be feasible to prevent ageing related cancers with pharmacologic agents that could decrease ¿. For aim (a), volunteer subjects in different age groups will be recruited, and ¿ will be measured directly using PIG-A as a sentinel gene in B lymphoblastoid cell lines and T lymphocyte cultures, in comparison with neonatal cord blood samples. ¿ will also be assessed indirectly, based on analysis of the frequency of granulocytes with PIG-A mutations and red cells with XK mutations as a function of age. For aim (b), using cells from normal older individuals and patients with progeria, the effect on the mutation rate will be determined for three pharmacologic approaches: quenching reactive oxygen species, modulating lamin A using farnesyltransferase inhibitors, and activating SIRT1. Preliminary data: ¿ in cultures of lymphoid cells from normal individuals ranges from 3 to 53 x 10-7 mutations per cell division and is positively correlated with age. ¿ is increased in cells from patients with cancer predisposition syndromes, premature ageing syndromes, and in malignant cell lines. A reduction in the spontaneous mutation rate in human cells has now been demonstrated using dehydroascorbic acid to reduce intracellular reactive oxygen species. Implications: Apart from addressing very important fundamental biological questions regarding ageing and cancer risk, studies under aim (a) are important for planning clinical chemoprevention studies. If ¿ is constant throughout life, then any strategy to reduce mutations may need to start early. If however, as suspected, ¿ starts to increase with age, it may be possible to prevent cancer by reducing ¿ at a later age, once it starts to increase. Studies under aim (b) will be critical for predicting the optimal drug targets and pharmacologic strategy for reducing ¿ clinically.

背景：在人类中，可以测量携带突变的细胞的频率（F） 从技术上讲，哨兵基因和历史上很少。测量 突变率（€） - 代表每个细胞基因中新突变的可能性 分裂 - 几乎是不可能的。然而，F和„可能是衰老的关键参数。为了 例如，它是假设的：（i）随着年龄的增长而增加； （ii）某些过早的 衰老综合征； （iii）有可能通过减少与衰老相关的癌症来减少。 现在，我们已经开发了一种强大的方法来使用PIG-A基因测量这些参数，该参数 我们希望将极大地支持这些假设的投资。 Pig-A在X染色体上 因此，单个灭活突变会产生突变表型。 Pig-A突变体缺乏所有 与GPI连接的膜蛋白，通过通过GPI-NULL表型对稀有细胞进行筛查 流式细胞仪。通过这种方法，我们能够测量粒细胞中的F，以及 B淋巴细胞细胞系和血液样本的过体淋巴细胞的培养物扩展。 最近，我们已经证明可以在额外的前哨中测量人类红细胞中的F Gene，XK，具有与PIG-A相似的优势。具体目的（a）：确定是否增加 随着年龄的增长，在人类中； （b）：确定防止与衰老相关的可行 癌症患者可能会降低药物。为了目标（a），不同的志愿者 将招募年龄组，并将使用PIG-A作为b中的前哨基因直接测量。 与新生儿血液相比，淋巴母细胞系和T淋巴细胞培养物 样品。基于对粒细胞频率的分析，也将间接评估 猪A突变和带有XK突变的红细胞随着年龄的函数。用于AIM（B），使用来自 正常的老年人和促销患者，对突变率的影响将是 确定三种药物方法：淬灭活性氧，调节 层lamin A使用Farneylsylansfer酶抑制剂并激活SIRT1。初步数据：在文化中 来自正常个体的淋巴样细胞每个细胞分裂3至53 x 10-7突变，IS 与年龄正相关。癌症患者的细胞中有所增加 综合征，过早衰老综合征和恶性细胞系。减少 现在已经证明了人类细胞中的赞助突变率使用脱氢酸到 减少细胞内活性氧。含义：除了解决非常重要的问题之外 有关衰老和癌症风险的基本生物学问题，目标（a）下的研究是 对于计划临床化学预防研究的重要性。如果一生都是恒定的，那么 减少突变的策略可能需要尽早开始。但是，如有怀疑，„开始增加 随着年龄的增长，一旦开始增加，就可以通过减少癌症来预防癌症。 AIM（B）下的研究对于预测最佳药物靶标和药理至关重要 临床上减少的策略。