Project 1: Effects of inflammation on clonal competition and malignant transformation

项目总结/摘要 克隆性造血（CH）是一种常见的癌前病变，其中一个或几个造血干细胞和 祖细胞（HSPC）对外周血的贡献不成比例。杂合子突变 约有20个基因与CH相关，其中一些基因在恶性血液病中常见。虽然 有证据表明，CH患者有恶性转化的高风险，其机制是 CH的产生和导致白血病的发生仍然知之甚少。揭示这些机制是一个 疾病预防的优先权和我们计划的总体目标。在我们的初步研究中，我们表明， 炎症可以驱动Dnmt 3a突变型HSPC的扩增。此外，我们开发了一种新型的马赛克鼠标， 通过移植来自五种不同CH相关突变小鼠系的HSPC混合群体作为 供体细胞（5x嵌合体小鼠）。然后，我们将这些小鼠暴露于可改变的外部应激源， 扩增携带某些突变的HSPC克隆。我们观察到不同的胁迫特异性克隆选择 突变型HSPCs。在这些数据的基础上，并利用团队的专业知识，项目1将测试假设 炎症应激导致具有CH相关突变的HSPC的表观遗传和代谢变化， 加速克隆扩张和恶性转化。我们将创造5个突变的嵌合体小鼠， 在表观遗传学、剪接和信号应答中起关键作用的常见CH相关基因：Dnmt 3a， Tet 2、Asxl 1、Jak 2和Sf 3b 1。我们将使用这种新的5倍小鼠模型来研究机制， 其中感染（目的1）和高脂饮食诱导的炎症和代谢变化（目的2）加速了克隆形成。 HSPC的选择、扩增和恶性转化。在这两个目标中，我们将进一步研究 干预炎症和代谢途径可以减弱克隆扩增和白血病 发展在目标3中，我们将探索炎症对Dnmt 3a-和Dnmt 3b-协同扩增的影响。 Tet 2突变体HSPC克隆。我们的团队已经提出了大量的初步研究，以强烈支持 我们提出的创新方法和整体假设的可行性。我们的小鼠模型和定量 在核心A中使用的方法描述了单个HSPC克隆的扩增和恶性转化， 对可改变的炎症应激源的反应是技术创新的。这项研究在概念上 创新，因为它引入了新的见解，如何改变风险因素选择性地驱动克隆扩张， HSPCs和白血病发生在扩大的遗传缺陷范围内。该项目将广泛互动 通过与Project共享5x镶嵌模型和定量检测技术以及转录组数据， 2（遗传毒性应激和DDR基因），并通过机械测试应激源之间的因果关系， 项目3中的药物、蛋白质组学和表观遗传学变化以及CH（大样本中CH和恶性肿瘤的分析） 人类社区群组）。总体方案将阐明CH进展的机制， 恶性肿瘤，并为人类CH相关突变的风险分层提供基础。

Project Summary/Abstract Clonal hematopoiesis (CH) is a common pre-malignant condition in which one or a few hematopoietic stem and progenitor cells (HSPCs) contribute disproportionately to peripheral blood. Heterozygous somatic mutations in about 20 genes are associated with CH, some of which are frequent in hematologic malignancies. Although evidence suggests individuals with CH are at high-risk for malignant transformation, the mechanisms by which CH arises and contributes to leukemogenesis remain poorly understood. Revealing these mechanisms is a priority for disease prevention, and an overall goal of our Program. In our preliminary studies, we showed that inflammation can drive expansion of Dnmt3a-mutant HSPCs. Further, we developed a novel mosaic mouse model by transplanting a mixed population of HSPCs from five different CH-associated mutant mouse lines as donor cells (5x mosaic mice). We then exposed these mice to modifiable external stressors and tracked expansion of HSPC clones harboring certain mutations. We observed stress-specific clonal selection of different mutant HSPCs. Building on these data and leveraging the team’s expertise, Project 1 will test the hypothesis that inflammatory stress leads to epigenetic and metabolic changes in HSPCs with CH-associated mutations to accelerate clonal expansion and malignant transformation. We will create 5x mosaic mice with mutations in five common CH-associated genes that play critical roles in epigenetics, splicing, and signaling responses: Dnmt3a, Tet2, Asxl1, Jak2, and Sf3b1. We will use this novel 5x mouse model to investigate the mechanisms through which infection (Aim 1) and high-fat diet-induced inflammation and metabolic changes (Aim 2) accelerate clonal selection, expansion, and malignant transformation of HSPCs. In both aims, we will further investigate whether intervening in inflammatory and metabolic pathways could attenuate clonal expansion and leukemia development. In Aim 3, we will explore the impact of inflammation on cooperative expansion of Dnmt3a- and Tet2-mutant HSPC clones. Our team has presented a large body of preliminary studies to strongly support the feasibility of our proposed innovative approaches and overall hypothesis. Our mouse models and the quantitative approaches used in Core A to delineate expansion and malignant transformation of individual HSPC clones in response to modifiable inflammatory stressors are technically innovative. This research is conceptually innovative, as it introduces novel insights into how modifiable risk factors selectively drive clonal expansion of HSPCs and leukemogenesis across an expanded range of genetic defects. This Project will interact extensively by sharing 5x-mosaic model and quantitative detection technologies as well as transcriptomic data with Project 2 (genotoxic stress and DDR genes), and by mechanistically testing causal connections between stressors, drugs, proteomic and epigenetic changes and CH from Project 3 (analysis of CH and malignancy in a large human community cohort). The Overall Program will clarify the mechanisms by which CH progresses toward malignancy and provide the groundwork for risk stratification of CH-associated mutations in humans.