Measuring Methylation Kinetics in Cancer Cells: Computations and Experiments

测量癌细胞中的甲基化动力学：计算和实验

• 批准号: 8050644
• 负责人: Natalia L Komarova
• 金额: $ 31.24万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8050644
• 关键词: Apoptosis; Cancer cell line; Cell Aging; Cell Line; Cell division; Cells; Colon Carcinoma; Computer Simulation; Coupled; DNA Methylation; DNA Repair; Data; Deletion Mutation; Development; Epigenetic Process; Event; Gene Silencing; Genes; Genetic; Genetic Code; Human; In Vitro; Institution; Kinetics; Knowledge; Learning; Light; Malignant Neoplasms; Measurement; Measures; Methylation; Modeling; Modification; Mutation; Oncogenes; Patients; Phenotype; Point Mutation; Population; Probability; Procedures; Process; Promoter Regions; Series; Site; Speed; Techniques; Testing; Time; Tumor Suppressor Genes; Tumor-Suppressor Gene Inactivation; Validation; base; cancer cell; cancer prevention; carcinogenesis; chromosome loss; mathematical model; promoter; research study; tumor progression

DESCRIPTION (provided by applicant): Methylation is an epigenetic mechanism of gene silencing found to be important in cancer progression. Many tumor suppressor genes are believed to be inactivated by methylation events. Therefore studying the process of methylation can advance our understanding of carcinogenesis. We propose to develop and implement computational and experimental approaches to measuring de novo methylation kinetics in cancer cells. Our overall aim is to quantify the rate of promoter methylation in cell lines with different genetic backgrounds. We will construct mathematical models which describe promoter methylation in a population of cells growing exponentially in vitro. Several assumptions regarding the methylation process are uncertain; therefore, we will consider a variety of different models. Testing these models will allow us to reject certain assumptions, and to determine which model is most consistent with data. We will conduct in vitro experiments to quantify the process of de novo methylation in several cancer cell lines. We will obtain time-series measurements of the colony size as well as methylation data, by using COBRA and pyrosequencing techniques. We will then use data from the experiments in order to distinguish which models are incorrect, and which model is most consistent with data. This will shed light on the mechanisms which underlie the methylation process. We will then use the experimentally validated, best-fitting model in order to calculate the site-specific and average rates of promoter methylation in cells with different genetic backgrounds. Lab summary: This project would help us better understand the speed/rate of colon cancer development that happens as a result of inappropriate DNA methylation, a mechanism that is prevalent in most human cancers. Since DNA methylation is a reversible process, a better knowledge of rate of cancer development in these patients would allow timely institution of methylation-reversing therapies that have implications for cancer prevention.Project Narrative: The initiation and progression of cancers involves both genetic and epigenetic modifications of cells. While genetic modifications (mutations) have been studied in detail, epigenetic changes (methylation) are not well understood, so we propose to study the methylation process of cancer gene promoters. By testing and rejecting various mathematical models we will find which mechanisms are key to the process, and use the best-fitting model to measure the actual methylation rate in different cell lines.

描述（由申请人提供）：甲基化是基因沉默的表观遗传机制，发现对癌症的进展很重要。人们认为许多肿瘤抑制基因被甲基化事件灭活。因此，研究甲基化过程可以提高我们对癌变的理解。我们建议开发和实施计算和实验方法，以测量癌细胞中的从头甲基化动力学。我们的总体目的是量化具有不同遗传背景的细胞系中启动子甲基化速率。 我们将构建数学模型，这些模型描述了在体外成倍增长的细胞群中的启动子甲基化。关于甲基化过程的几个假设尚不确定。因此，我们将考虑各种不同的模型。测试这些模型将使我们能够拒绝某些假设，并确定哪种模型与数据最一致。 我们将进行体外实验，以量化几种癌细胞系中从头甲基化的过程。我们将通过使用眼镜蛇和焦磷酸测序技术获得菌落大小以及甲基化数据的时间序列测量。然后，我们将使用实验中的数据来区分哪些模型不正确，哪些模型与数据最一致。这将阐明甲基化过程的基础的机制。然后，我们将使用经过实验验证的，最合适的模型，以计算具有不同遗传背景的细胞中启动子甲基化的位点特异性和平均速率。 实验室摘要：该项目将有助于我们更好地了解由于不适当的DNA甲基化而发生的结肠癌发展的速度/速度，这种机制在大多数人类癌症中都普遍存在。由于DNA甲基化是一个可逆的过程，因此对这些患者的癌症发展速率的更好了解将允许及时建立对甲基化的替代疗法，这些疗法对预防癌症有影响。项目叙事： 癌症的起始和进展涉及细胞的遗传和表观遗传修饰。尽管已经详细研究了遗传修饰（突变），但表观遗传变化（甲基化）尚不清楚，因此我们建议研究癌基因启动子的甲基化过程。通过测试和拒绝各种数学模型，我们将发现哪些机制是该过程的关键，并使用最合适的模型来测量不同细胞系中的实际甲基化速率。