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 在种系Runx1突变载体中： 炎症和骨髓生态位 项目摘要/摘要 赋予白血病风险的种系变体已越来越多，急性为15-20％ 白血病患者至少有1级相对患有白血病。种系变体现在 世界卫生概述的修订的白血病分类计划中首次包括 组织和新的临床指南现在包括测试遗传易感性作为关键要素 患者诊断。这些患者及其家人的了解是毁灭性的 一生中增加了患癌症的风险。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 明显白血病之前发生的分子变化。这项研究的长期目标是确定 种系Runx1突变患者的造血阶段调节造血的机制 [也称为家族血小板疾病（FPD）]。这些人有终生的血小板减少症，定性 血小板聚集缺陷，并有发展各种造血恶性肿瘤的风险。收购 FPD患者随着时间的流逝发生了继发性突变，导致白血病发育。但是， 使易于获得次级突变的固有和/或外在因素是 未知。为了填补这一知识空白，我们进行了主要FPD骨的单细胞转录组分析 与健康相比，骨髓样品并确定了FPD祖细胞的独特转录变化 控件。一致的这些变化，FPD祖细胞损害了巨核细胞，但增强了髓样 分化。骨髓造血和间充质基质细胞（MSC）的细胞因子分析显示 骨髓内的细胞因子产生增加，导致慢性炎症应激， 这可能会给FPD祖细胞带来增长优势。此外，FPD基质细胞也显示出缺陷 分化，克隆生成能力和失调的基因表达，可以进一步促进 炎症状态。因此，我们假设FPD茎/祖细胞的早期转录组变化 与炎症性微环境信号合作，为这些美食前提供增长优势 细胞并改变其分化。我们将使用以下目的检验我们的假设：1）确定如何早期 转录组变化改变了FPD细胞的造血。 2）确定炎症细胞因子的影响 以及他们在FPD进化中的封锁。 3）评估FPD基质微环境在生长和 FPD和健康的茎/祖细胞的分化。为了实现我们的目标，我们组装了几种工具， 包括主要的茎/祖细胞/和基质细胞，IPSC，异种移植和转基因小鼠模型， CRISPR/CAS9 造血 定位 和 编辑技术和3D共文化模型。这个项目将提高我们对 在FPD患者中，将为新疗法的合理设计提供分子基础 Runx1介导的变化和炎症微环境，以使造血归一化 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