Hematopoiesis in germline RUNX1mutation carriers: impact of inflammation and the bone marrow niche

种系 RUNX1 突变携带者的造血作用：炎症和骨髓生态位的影响

• 批准号: 10626095
• 负责人: Anupriya Agarwal
• 金额: $ 51.69万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10626095
• 关键词: 3-Dimensional; Acute leukemia; Affect; Biogenesis; Bone Marrow; Bone Marrow Cells; Bone Marrow Diseases; Bone Marrow Transplantation; CCL2 gene; CRISPR/Cas technology; Candidate Disease Gene; Cell Differentiation process; Cell Survival; Cells; Chronic; Classification Scheme; Climacteric; Clinical; Clonal Expansion; Coculture Techniques; Collection; DNA Damage; Data; Defect; Development; Diagnostic; Differentiation and Growth; Disease; Disease Progression; Early identification; Elements; Environment; Event; Evolution; Exposure to; Extracellular Fluid; Familial Platelet Disorder; Family; Family member; First Degree Relative; Gene Expression; Genetic; Genetic Predisposition to Disease; Genetic Transcription; Germ-Line Mutation; Goals; Growth; Guidelines; Hematopoiesis; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic stem cells; IL8 gene; Impairment; In Vitro; Individual; Inflammation; Inflammatory; Knowledge; Leukemic Cell; Malignant Neoplasms; Mediating; Megakaryocytes; Modeling; Molecular; Monitor; Mus; Mutant Strains Mice; Mutate; Mutation; Myelogenous; Pathway interactions; Patient-Focused Outcomes; Patients; Phenotype; Platelet aggregation; Predisposition; Preleukemia; Prevention; Prevention strategy; Production; Prognostic Marker; RUNX1 gene; Regulation; Resources; Ribosomes; Risk; Role; Sampling; Screening for cancer; Signal Transduction; Stress; Stromal Cells; Syndrome; Techniques; Testing; Therapeutic; Thrombocytopenia; Time; Transgenic Mice; Translations; Up-Regulation; World Health Organization; Xenograft Model; Xenograft procedure; biomarker identification; cytokine; early detection biomarkers; erythroid differentiation; experience; hematopoietic differentiation; improved; improved outcome; in vivo; induced pluripotent stem cell; leukemia; mesenchymal stromal cell; monocyte; mouse model; mutant; mutation carrier; novel; novel therapeutics; pharmacologic; premalignant; prevent; progenitor; programs; proto-oncogene protein Spi-1; rational design; response; self-renewal; stem; stem cells; therapeutic biomarker; therapeutically effective; tool; transcriptomic profiling; transcriptomics; transplant model

Hematopoiesis impact in germline RUNX1 mutation carriers: of inflammation and the bone marrow niche PROJECT SUMMARY/ABSTRACT Germline variants that confer risk for leukemia have been recognized increasingly with 15-20% of acute leukemia patients having at least 1 first-degree relative afflicted with leukemia. Germline variants are now included for the first time in the revised leukemia classification scheme outlined by the World Health Organization, and new clinical guidelines now include testing for inherited susceptibility as a critical element of patient diagnostics. It is devastating for these patients and their family members to live with the knowledge of having increased risk for developing cancer in their lifetimes. Thus, for these patients, early cancer detection, disease monitoring, and prevention would be life-changing, but require a comprehensive understanding of molecular changes that occur prior to overt leukemia. The long-term goal of this study is to identify the mechanisms that regulate hematopoiesis at the premalignant stage in patients with germline RUNX1 mutations [also called familial platelet disorder (FPD)]. These individuals have life-long thrombocytopenia, qualitative platelet aggregation defects, and a risk of developing a variety of hematopoietic malignancies. The acquisition of secondary mutations occurs over time in FPD patients leading to leukemia development. However, the intrinsic and/or extrinsic factors that render pre-leukemic cells vulnerable to acquire secondary mutations are unknown. To fill this knowledge gap, we performed single-cell transcriptome profiling of primary FPD bone marrow samples and identified unique transcriptional changes in FPD progenitors compared to healthy controls. Consistent these changes, FPD progenitors have impaired megakaryocytic but enhanced myeloid differentiation. Cytokine profiling of bone marrow hematopoietic and mesenchymal stromal cells (MSCs) show that there is increased cytokine production within the bone marrow, leading to chronic inflammatory stress, which may confer a growth advantage to FPD progenitors. Moreover, FPD stromal cells also show defective differentiation, clonogenic capacity, and deregulated gene expression that may further promote an inflammatory state. Thus, we hypothesize that early transcriptomic changes in FPD stem/progenitors cooperate with inflammatory microenvironmental signals to provide a growth advantage to these pre-leukemic cells and alter their differentiation. Wewill test our hypothesis using the following aims: 1)Identify how early transcriptomic changes alter hematopoiesis of FPD cells. 2) Determine the impact of inflammatory cytokines and their blockade in FPD evolution. 3) Assess the role of the FPD stromal microenvironment on growth and differentiation of FPD and healthy stem/progenitors. To achieve our goals, we have assembled several tools, including primary stem/progenitor/and stromal cells, iPSCs, xenograft and transgenic mouse models, CRISPR/Cas9 hematopoiesis targeting and editing technique, and 3D co-culture models. This project will advance our understanding of in FPD patients and will provide the molecular basis for the rational design of new therapeutics of RUNX1-mediated changes and the inflammatory microenvironment to normalize hematopoiesis delay or prevent leukemia in FPD patients.

造血 影响 生殖系RUNX 1突变携带者： 炎症和骨髓龛 项目总结/摘要 越来越多的人认识到，15-20%的急性白血病患者存在生殖系变异， 至少有一个一级亲属患有白血病的白血病患者。现在， 首次被纳入世界卫生组织（WHO）修订的白血病分类方案。 组织，新的临床指南现在包括测试遗传易感性作为一个关键因素， 患者诊断。这是毁灭性的，这些病人和他们的家人生活的知识， 一生中患癌症的风险增加。因此，对于这些患者，早期癌症检测， 疾病监测和预防将改变生活，但需要全面了解 发生在白血病之前的分子变化这项研究的长期目标是确定 生殖系RUNX 1突变患者在癌前阶段调节造血的机制 [also称为家族性血小板疾病（FPD）。这些人有终身血小板减少症，定性 血小板聚集缺陷和发展成各种造血系统恶性肿瘤的风险。收购 继发性突变的发生随着时间的推移，在FPD患者导致白血病的发展。但 使前白血病细胞易于获得继发性突变内在和/或外在因素， 未知为了填补这一知识空白，我们对原发性FPD骨进行了单细胞转录组分析， 与健康人相比，在FPD祖细胞中鉴定出独特的转录变化 对照与这些变化一致，FPD祖细胞具有受损的巨核细胞但增强的髓样细胞 分化骨髓造血和间充质基质细胞（MSC）的细胞因子谱显示， 骨髓中细胞因子的产生增加，导致慢性炎症应激， 这可赋予FPD祖细胞生长优势。此外，FPD基质细胞也表现出缺陷， 分化、克隆形成能力和失调的基因表达可能进一步促进肿瘤的发生。 炎症状态因此，我们假设FPD干/祖细胞中的早期转录组学变化 与炎症微环境信号合作，为这些白血病前期细胞提供生长优势， 细胞并改变其分化。我们将使用以下目标来测试我们的假设：1）确定 转录组学变化改变FPD细胞的造血。2)确定炎性细胞因子的影响 以及它们对FPD进化的阻碍。3)评估FPD基质微环境对生长的作用， FPD和健康干/祖细胞的分化。为了实现我们的目标，我们已经组装了几个工具， 包括原代干细胞/祖细胞/和基质细胞、iPSC、异种移植物和转基因小鼠模型， CRISPR/Cas9 造血 靶向 和 编辑技术和3D共培养模型。这个项目将促进我们对 并将为合理设计新疗法提供分子基础 RUNX 1介导的变化和炎症微环境使造血正常化 延迟或预防FPD患者的白血病。

项目成果

Validation and clinical application of transactivation assays for RUNX1 variant classification.

• DOI: 10.1182/bloodadvances.2021006161
• 发表时间: 2022-06-14
• 期刊: BLOOD ADVANCES
• 影响因子: 7.5
• 作者: Decker, Melanie;Agarwal, Anupriya;Benneche, Andreas;Churpek, Jane;Duployez, Nicolas;Duvall, Adam;Ernst, Martijn P. T.;Foerster, Alisa;Hoberg-Vetti, Hildegunn;Hofmann, Inga;Nash, Michelle;Raaijmakers, Marc H. G. P.;Tvedt, Tor H. A.;Vlachos, Adrianna;Schlegelberger, Brigitte;Illig, Thomas;Ripperger, Tim
• 通讯作者: Ripperger, Tim