(PQB6) Accurate Comparative Genome-wide Analysis of Primary Tumors and Metastases

描述（由申请人提供）：大多数癌症死亡不是由原发肿瘤引起的，而是由远处转移引起的。 惰性原发性肿瘤进展为转移性的过程涉及肿瘤细胞的基因表达和染色质修饰模式的戏剧性重编程。 有效地分析整个基因组中的这些变化，并确定强大的促转移基因表达和染色质修饰特征，对于了解肿瘤进展和治疗人类癌症至关重要。 近年来，我们在全基因组范围内分析表观遗传和基因表达模式的能力有了巨大的进步。 然而，由于分离不含污染基质的纯肿瘤细胞群的困难，对小鼠癌症模型的适用性变得复杂。 我们将开发专门分析小鼠实体瘤肿瘤细胞的技术，不含基质和其他污染细胞类型。 这将允许对来自原发性肿瘤和转移瘤的纯细胞核群体进行准确比较。 具体而言，我们将：1）开发用于在人细胞系中生物素标记核膜蛋白和用于从裸鼠中的异种移植肿瘤分离生物素标记的细胞核的方法。 作为原理的证明，我们将测试我们是否可以对这些模型的原发性肿瘤和转移瘤进行全基因组比较。 2)生成并测试基因工程小鼠模型，其中核被膜生物素化盒的表达可由Cre重组酶诱导。 当与人类癌症的小鼠模型结合时，其中肿瘤由Cre介导的癌基因激活或肿瘤抑制因子缺失驱动，这将允许分离和分析纯肿瘤细胞核群体。 这项工作将产生广泛适用于人类癌症小鼠模型中肿瘤进展和转移研究的工具和方法。

DESCRIPTION (provided by applicant): The majority of deaths due to cancer are caused not by the primary tumor but by distant metastases. The process by which an indolent primary tumor progresses to become metastatic involves a dramatic reprogramming of the tumor cell's gene expression and chromatin modification patterns. Effectively analyzing these changes across the entire genome, and identifying robust pro-metastatic gene expression and chromatin modification signatures, will be essential to understanding tumor progression and treating human cancer. Our ability to profile epigenetic and gene expression patterns on a genome-wide scale has advanced enormously in recent years. However, the applicability to mouse models of cancer is complicated by the difficulties in isolating pure populations of tumor cells that are free from contaminating stroma. We will develop technology for specifically profiling tumor cells from solid tumors in mice, free from stroma and other contaminating cell types. This will allow the accurate comparison of pure populations of nuclei from primary tumors and metastases. Specifically, we will: 1) Develop methods for biotin tagging a nuclear envelope protein in human cells lines and for isolating the biotin tagged nuclei from a xenograft tumor in nude mice. As proof of principle, we will test whether we can perform a genome-wide comparison of primary tumors and metastases from these models. 2) Generate and test a genetically engineered mouse model in which expression of a nuclear envelope biotinylation cassette can be induced by the Cre recombinase. When combined with mouse models of human cancer in which the tumor is driven by Cre-mediated activation of an oncogene or deletion of a tumor suppressor, this will allow for the isolation and analysis of pure populations of tumor cell nuclei. This work will generate tools and methods that will be widely applicable to the study of tumor progression and metastasis in mouse models of human cancer.