Epigenomic Mapping in Human Tumor Stem Cells

描述(申请人提供)：肿瘤干细胞的概念，一种能够再生肿瘤的自我更新细胞，揭示了人类癌症的持久性，并为翻译研究提供了一个新的焦点。我们的目标是了解将肿瘤干细胞与其他肿瘤细胞以及其他正常的自我更新细胞区分开来的表观遗传学机制。表观遗传机制，如染色质修饰、DNA甲基化和小的非编码RNA是细胞转录潜力的稳定的、长期的(通常是可遗传的)变化，与潜在基因组序列的变化无关。细胞的表观遗传状态用于定义细胞的身份和该细胞潜在命运的极限。因此，对细胞表观遗传状态的了解可以识别肿瘤干细胞的特征和潜能。组蛋白修饰、DNA甲基化和小非编码RNA的表观遗传状态将通过染色质免疫沉淀、亚硫酸氢盐转化和RNA文库的生成与全基因组测序相结合来绘制。为了深入了解肿瘤干细胞，我们将利用之前开发的模型系统，该系统提供具有肿瘤干细胞能力的一致细胞来源。拟议的实验将检验一组细胞之间的差异，每一组细胞都被分类，以丰富具有肿瘤干细胞能力的亚群。为了进行比较，将从相同的来源分离非肿瘤干细胞，并检查肿瘤干细胞与多能干细胞和多能干细胞之间的差异。还将设计其他实验来扰乱细胞的表观基因组，并直接测试起源细胞的表观遗传状态如何影响后续的肿瘤表型。产生的知识可能带来实质性的新见解，包括识别用于早期诊断、预后或监测肿瘤治疗的假定标记物。了解肿瘤干细胞和其他自我更新细胞之间的差异可能会导致更具体的治疗，同时对正常细胞的毒性也更低。操纵表观基因组状态并检查致瘤性的结果将提供对这些特征如何转化为临床相关表型的直接洞察。公共卫生相关性：我们计划确定与非肿瘤干细胞、多能干细胞和多能干细胞相比，有助于调节肿瘤干细胞中基因表达的独特分子特征。了解肿瘤干细胞基因调控的独特方面将带来用于诊断、预后和监测的新生物标记物，以及对肿瘤干细胞更具特异性且对正常、自我更新细胞毒性较低的新治疗方法。

DESCRIPTION (provided by applicant): The concept of a tumor stem cell, a self-renewing cell capable of regenerating the tumor, has shed light on the persistence of human cancers and offers a new focus for translational research. Our goal is to understand the epigenetic mechanisms that distinguish the tumor stem cell from other cells of the tumor and from other normal, self-renewing cells. Epigenetic mechanisms, such as chromatin modifications, DNA methylation and small noncoding RNAs are stable, long-term (typically heritable) changes in the transcriptional potential of a cell that are independent of changes in the underlying genomic sequence. The epigenetic state of a cell serves to define cell identity and the limits of that cell's potential fates. Thus, knowledge of the epigenetic state of cells may identify signature for both tumor stem cell identity and potential. The epigenetic states of histone modifications, DNA methylation and small noncoding RNAs will be mapped using chromatin immunoprecipitation, bisulfite conversion and generation of RNA libraries coupled with genome-wide sequencing. For insight on tumor stem cells, we will take advantage of a previously developed model system that provides a consistent source of cells with tumor stem cell ability. Proposed experiments will examine the differences between a panel of cells, each sorted to enrich for the subpopulation with tumor stem cell ability. For comparisons, non-tumor stem cells will be isolated from the same source and the differences between tumor stem cells and pluripotent and multipotent stem cells will be examined. Additional experiments will be designed to perturb the epigenome of cells and directly test how the epigenetic state of the cell-of-origin affects subsequent tumor phenotype. The knowledge generated could lead to substantial new insights, including the identification of putative markers for early diagnosis, prognosis or monitoring of tumor therapies. Understanding the differences between tumor stem cells and other self-renewing cells could lead to more specific therapies that were also less toxic to normal cells. Manipulating the epigenomic state and examining the results on tumorigenicity would provide direct insight on how these signatures translate into clinically relevant phenotypes. Public Health Relevance: We plan to identify unique molecular signatures that contribute to the regulation of gene expression in tumor stem cells compared to non-tumor stem cells, pluripotent stem cells and multipotent stem cells. Understanding the unique aspects of tumor stem cell gene regulation will lead to new biomarkers for diagnosis, prognosis and monitoring as well as new therapeutic approaches that are both more specific for tumor stem cells and less toxic to normal, self- renewing cells.