Large-scale functional interrogation of evolutionary forces in pre-germinal center B cell lymphoma development

套细胞淋巴瘤(MCL)是一种侵袭性的恶性肿瘤，其特征是在B细胞发育早期获得的染色体易位导致Cyclin D1过表达。基因组测序为MCL的突变格局提供了重要的见解，但也揭示了许多层次的未探索的复杂性，这一点还远未被理解。由非遗传机制引起的基因失调，包括表观遗传和转录变化，是普遍存在的，区分相关的驱动因素是具有挑战性的。识别关键的调控节点，定位它们在通路、网络和过程中的位置，了解遗传相互作用和通路重新连接，剖析信号输出的广泛上下文依赖性，以及了解肿瘤微环境中的细胞/细胞通信，只是尚未解决的主要挑战的一部分。在我们开发的小鼠模型、遗传工具、方法学和数据资源的基础上，我们建议在不同层面上应对这些挑战。在AIM-1和AIM-2中，我们将进行全基因组筛选和全面的大规模调查，以表征淋巴瘤演变背后的细胞内在和外在过程。通过将体内功能筛选与高分辨率染色质图谱和超级增强子图谱相结合，我们的目标是发现驱动小鼠肿瘤进化不同阶段的核心细胞内在调控电路(AIM-1)。我们还将描述淋巴瘤微环境的组成和结构，并监测其在淋巴瘤发生和发展过程中的纵向变化(AIM-2)。单细胞测序和多光谱成像将有助于对MCL利基内参与细胞/细胞通信的细胞和分子参与者进行系统调查。在AIM-3中，我们将从功能上描述源自AIM-1/2的关键发现。其中一个焦点是我们最近对CD40激活的研究，CD40是MCL中的一个标志性事件，发现IL9/IL9R介导的细胞/细胞通讯在淋巴瘤发病机制中发挥了意想不到的作用。我们的数据表明，连续的前馈循环涉及多个分子和细胞成分。我们将通过表征涉及的细胞类型、基因、信号通路以及与它们的扰动相关的组织表型，在小鼠中对这一模型进行功能性询问。MCL进化中关键调控节点、过程和相关性的系统发现和机制表征有望为MCL的精确肿瘤学工作提供信息。此外，这里创造的丰富数据资源将推动假设生成和下游功能研究远远超出这一提议的框架。

Mantle cell lymphoma (MCL) is an aggressive malignancy, characterized by Cyclin D1 overexpression through a chromosomal translocation acquired during early B cell development. Genome sequencing has given important insights into the mutational landscapes of MCL, but also revealed many layers of unexplored complexity, which is far from being understood. Gene dysregulation by non-genetic mechanisms, including epigenetic and transcriptional changes, is widespread, and discriminating the relevant drivers is challenging. Identifying the key regulatory nodes, positioning them in pathways, networks and processes, understanding genetic interactions and pathway rewiring, dissecting the extensive context-dependency of signaling outputs and understanding cell/cell communication in the tumor microenvironment are only some of the major unresolved challenges. Building on mouse models, genetic tools, methodologies and data resources developed by us, we propose to address these challenges at different levels. In AIM-1 and 2 we will perform genome wide screens and comprehensive large-scale surveys to characterize cell-intrinsic and extrinsic processes underlying lymphoma evolution. By combining in vivo functional screening with high-resolution chromatin profiling and super-enhancer mapping, we aim to discover core cell-intrinsic regulatory circuitries driving different stages of tumor evolution in mice (AIM-1). We will also characterize the composition and architecture of the lymphoma microenvironment and monitor its changes longitudinally during lymphoma initiation and progression (AIM-2). Single cell sequencing and multispectral imaging will aid systematic surveys for cellular and molecular players involved in cell/cell communication within the MCL niche. In AIM-3 we will functionally characterize key discoveries emanating from aim-1/2. One focus is triggered by our recent work on CD40 activation, a hallmark event in MCL, which discovered an unexpected role of IL9/IL9R mediated cell/cell communication in lymphoma pathogenesis. Our data suggest consecutive feed-forward loops involving multiple molecular and cellular constituents. We will functionally interrogate this model in mice by characterizing the involved cell types, genes, signaling pathways, as well as organismal phenotypes related to their perturbation.The systematic discovery and mechanistic characterization of key regulatory nodes, processes and dependencies in MCL evolution is promising to inform precision oncology efforts in MCL. Moreover, the rich data resource created here will drive hypothesis generation and downstream functional studies far beyond the framework of this proposal.