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

诱导克隆扩增的造血干细胞中的体细胞突变通常在人类衰老期间获得。不确定潜能的克隆性造血（CHIP）是恶性血液病和心血管疾病发生的重要危险因素，并与全因死亡率增加相关。越来越多的证据表明，失调的炎症有助于克隆扩张和相关的合并症，如来自先前感染的炎症、预先存在的合并症或随着衰老发生的低度炎症（炎症）。CHIP特异性分子特征可能是诊断和开发特异性治疗的重要靶点。 在这个项目中，我们的目标是确定与先进的多组学单细胞分析工具CHIP相关的信号通路和表面标记物的表达模式。在这项建议的初步工作中，我们分析了来自600多名接受干细胞白细胞分离术的患者的动员干细胞产物中的CHIP突变。此外，我们建立了一个特定的生物信息学工具集，在这个项目中的分析。 多组学单细胞分析将用于鉴定不同CHIP突变和细胞类型中的失调信号传导途径和表面标志物模式（CITE-Seq）。随后，我们将在来自脐带血的CD 34+造血干细胞（HSC）中过表达和敲除CHIP相关的表达模式/信号通路，并在体外和体内分析信号/炎症基因的修饰是否与候选驱动突变表型相似。对于可获得特异性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，以评估相应药物优先根除CHIP克隆的潜力。总之，该项目将揭示可能用于诊断和潜在治疗应用的途径/调控基因，以抑制CHIP克隆。

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.