Signaling properties of driver mutations in clonal hematopoiesis of indeterminate potential (CHIP) as targets for diagnostics and therapeutic intervention

导致克隆性扩增的造血干细胞的体细胞突变通常是在人类衰老过程中获得的。克隆性造血不确定潜能(CHIP)是血液系统恶性肿瘤和心血管疾病发展的重要危险因素，并与全因死亡率增加有关。越来越多的证据表明，调节失调的炎症有助于克隆性扩张和相关的共病，如既往感染引起的炎症、先前存在的共生疾病或随年龄增长而发生的低度炎症(炎症)。芯片特有的分子特征可能是诊断和开发特定治疗方法的重要靶点。在这个项目中，我们的目标是用先进的多组学单细胞分析工具识别芯片相关的信号通路和表面标记表达模式。在这项建议的前期工作中，我们分析了600多名接受干细胞白细胞分离的患者动员后的干细胞产品中的芯片突变。此外，我们还为本项目的分析建立了一套专门的生物信息学工具。多组学单细胞分析将被用于识别不同芯片突变和细胞类型的非调控信号通路和表面标记模式(CITE-SEQ)。随后，我们将在脐血CD34+造血干细胞(HSC)中过表达和敲除芯片相关的表达模式/信号通路，并在体外和体内分析信号/炎症基因表型的修饰是否可能导致驱动突变。对于具有特定抑制剂的顶级调控途径/靶点，我们将分析体外药物反应，并将进行体内异种移植研究，以评估各自药物优先根除芯片克隆的潜力。综上所述，该项目将揭示可能用于诊断的途径/调控基因，并可能用于抑制芯片克隆的治疗应用。

Somatic mutations in hematopoietic stem cells which induce clonal expansion are commonly acquired during human aging. Clonal hematopoiesis of indeterminate potential (CHIP) constitutes an important risk factor for the development of hematological malignancies and cardiovascular disease and is associated with an increased all-cause mortality. A growing body of evidence suggest that dysregulated inflammation contributes to clonal expansion and associated comorbidities such as inflammation from previous infections, pre-existing comorbidities or low-grade inflammation occurring with aging (inflammaging). CHIP-specific molecular features might be important targets for diagnostics and development of specific therapies. In this project we aim to identify CHIP associated signaling pathways and surface marker expression patterns with advanced multi-omics single cell analysis tools. In preliminary work for this proposal we analyzed CHIP mutations in mobilized stem cell products from more than 600 patients undergoing stem cell leukapheresis. Also, we established a specific bioinformatics tool set for the analyses in this project. Multi-omics single cell analyses will be used to identify deregulated signaling pathways and surface marker patterns (CITE-Seq) across diverse CHIP mutations and cell types. Subsequently, we will overexpress and knockout CHIP-associated expression patterns/signaling pathways in CD34+ hematopoeitic stem cells (HSC) from cord blood and analyse in vitro and in vivo whether modification of signaling/inflammatory genes phenocopies candidate driver mutations. For the top-regulatory pathways/ targets for which specific inhibitors are available we will analyze in vitro- drug responses and will perform in vivo xenograft transplantation studies to evaluate the potential of the respective drugs to preferentially eradicate CHIP clones. Taken together, this project will reveal pathways/regulatory genes which might be used for diagnostics and potentially for therapeutic applications to suppress CHIP clones.