Targeting aberrant enhancer landscapes in pancreatic cancer

项目总结/摘要 该项目的中心目标是开发表观遗传学重编程胰腺癌细胞的策略， 减少转移扩散。这一目标是基于我们最近的证明，胰腺癌 转移伴随着增强子激活的典型模式。我们暗示先驱者 转录因子FOXA 1作为增强子重编程和转移扩散的驱动因素。在 在这个提议中，我们试图定义增强子-转移连接。在此过程中，我们寻求提名一个新的 一类表观遗传靶点，这可能是唯一能够消除转移潜力。在第一个目标 在这项提议中，我们将采用功能基因组学方法来干扰每一个FOXA 1调节基因， 增强子，并确定FOXA 1依赖性转移是否可以抑制。这种方法将采取 我们最近在以领域为重点的CRISPR筛选方面的创新优势，并将加深我们的理解 这个表观遗传程序的促转移成分。在本建议的第二个目标中，我们将 研究在FOXA 1上调之前发生的增强子重编程过程的最早步骤。 这一努力建立在我们意想不到的观察基础上，即转移特异性增强子已经存在于 转移前胰腺肿瘤细胞中的可接近染色质状态。这表明额外的分子 事件发生在FOXA 1上调之前，从而为转移期间的增强子重编程设定阶段。我们 将研究转录因子FOXA 2，我们的实验表明它是关键的书签， 在转移前癌细胞中打开了转移特异性增强子。此外，我们将确定如何 抑制性Polycomb复合物用于在FOXA 1上调之前抑制增强子激活。通过评估 FOXA 2/Polycomb扰动的后果，我们将提供转移的概念验证- 相关的增强子可以在胰腺癌进展的早期阶段被消除。最终目的是 建议将我们的增强子定位研究扩展到胰腺癌的鳞状亚型， 这是一种最近定义的疾病实体，与特别令人沮丧的预后相关。我们将雇用一名 新表征的患者源性胰腺癌类器官的补体以比较增强子谱 鳞状细胞亚型与更经典的这种疾病的比较。我们将确定这方面的主要监管机构 鳞状转录程序，并进行遗传实验，以确定这些因素的作用， 促进肿瘤进展和转移扩散。我们还将确定是否鳞状细胞的身份， PDA与独特的表观遗传依赖性有关。总的来说，拟议的研究将提供一个 机制框架，用于开发靶向独特增强子构型的表观遗传疗法， 转移性胰腺癌细胞。

Project Summary/Abstract The central goal of this project is to develop strategies to epigenetically reprogram pancreatic cancer cells to diminish metastatic spread. This objective is based on our recent demonstration that pancreatic cancer metastasis is accompanied by a stereotypical pattern of enhancer activation. We implicated the pioneer transcription factor FOXA1 as a driver of enhancer reprogramming and of metastatic spread in this context. In this proposal, we seek to define the enhancer-metastasis connection. In doing so, we seek to nominate a new class of epigenetic targets, which might be uniquely capable of eliminating metastatic potential. In the first Aim of this proposal, we will employ a functional genomics approach to perturb every FOXA1-regulated gene and enhancer and determine whether FOXA1-dependent metastasis can be suppressed. This approach will take advantage of our recent innovations in domain-focused CRISPR screening and will deepen our understanding of the pro-metastatic components of this epigenetic program. In the second Aim of this proposal, we will investigate the earliest steps of the enhancer reprogramming process that occur prior to FOXA1 upregulation. This effort builds from our unexpected observation that metastasis-specific enhancers are already present in an accessible chromatin state in pre-metastatic pancreatic tumor cells. This suggests that additional molecular events occur prior to FOXA1 upregulation to set the stage for enhancer reprogramming during metastasis. We will investigate the transcription factor FOXA2, which our experiments suggest is the critical bookmark that opens up metastasis-specific enhancers in pre-metastatic cancer cells. In addition, we will determine how repressive Polycomb complexes act to restrain enhancer activation prior to FOXA1 upregulation. By evaluating the consequences of FOXA2/Polycomb perturbation, we will provide a proof-of-concept that metastasis- associated enhancers can be effaced at early stages of pancreatic cancer progression. The final Aim of this proposal will be to extend our enhancer mapping studies into the squamous-subtype of pancreatic cancer, which is a recently defined disease entity associated with a particularly dismal prognosis. We will employ a newly characterized complement of patient-derived pancreatic cancer organoids to compare enhancer profiles of squamous-subtype versus the more classical form of this disease. We will identify master-regulators of this squamous transcriptional program, and perform genetic experiments to determine the role of such factors in promoting tumor progression and metastatic spread. We will also determine whether squamous cell identity in PDA is associated with unique epigenetic dependencies. Collectively, the proposed research will provide a mechanistic framework for developing epigenetic therapies that target the unique enhancer configuration of metastatic pancreatic cancer cells.