Epigenomic Mapping in Human Tumor Stem Cells

描述（由申请人提供）：肿瘤干细胞是一种能够再生肿瘤的自我更新细胞，它的概念揭示了人类癌症的持久性，并为转化研究提供了一个新的焦点。我们的目标是了解区分肿瘤干细胞与其他肿瘤细胞和其他正常的自我更新细胞的表观遗传机制。表观遗传机制，如染色质修饰、DNA甲基化和小非编码rna是细胞转录潜力的稳定、长期（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决定了细胞的身份和细胞潜在命运的界限。因此，了解细胞的表观遗传状态可以识别肿瘤干细胞的身份和潜力。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、亚硫酸盐转化和RNA文库生成以及全基因组测序来绘制。为了深入了解肿瘤干细胞，我们将利用先前开发的模型系统，提供具有肿瘤干细胞能力的一致细胞来源。拟议的实验将检查一组细胞之间的差异，每个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，将从同一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。其他的实验将被设计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随后的肿瘤表型。由此产生的知识可能会带来实质性的新见解，包括确定早期诊断、预后或肿瘤治疗监测的假定标记。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方法，对正常细胞的毒性也更小。操纵表观基因组状态并检查致瘤性结果将为这些特征如何转化为临床相关表型提供直接见解。

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.