A path to thrombosis in primary myelofibrosis

原发性骨髓纤维化的血栓形成途径

• 批准号: 10064585
• 负责人: KATYA RAVID
• 金额: $ 47.67万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-12-24 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10064585
• 关键词: Acute; Adhesions; Affect; Aldehydes; Binding; Blood Platelets; Bone Marrow; Cardiovascular system; Carotid Arteries; Cell surface; Cells; Chronic; Chronic Kidney Failure; Cicatrix; Collaborations; Collagen; Collagen Receptors; Collagen Type I; Complement; Data; Deposition; Disease; Dyes; Elastin; Engineering; Enzymes; Event; Extracellular Matrix; Flowmeters; Future; Gene Mutation; Genes; Human; Image; Incidence; Integrin alpha2; Integrins; Investigation; Kidney; Knockout Mice; LOX gene; Laboratories; Lasers; Lead; Light; Link; Lysine; Malignant Neoplasms; Measurement; Mediating; Megakaryocytes; Membrane Proteins; Mus; Mutagenesis; Myelofibrosis; Myeloproliferative disease; Nitrogen; Oxidases; Oxides; Pathologic; Pathology; Pathway interactions; Patients; Pharmacology; Physiologic pulse; Platelet Activation; Platelet-Derived Growth Factor Receptor; Primary Myelofibrosis; Property; Protein-Lysine 6-Oxidase; Proteins; Proteomics; Publishing; Receptor Activation; Research; Risk; Role; Sampling; Serum; Shapes; Stenosis; Structure; Tensile Strength; Testing; Thrombosis; Thrombus; Tissues; Transgenic Mice; Up-Regulation; Western Blotting; Work; calreticulin; cohort; crosslink; drug development; extracellular; ferric chloride; human disease; in vivo; inhibitor/antagonist; innovation; intravital microscopy; knock-down; loss of function; mouse model; novel; overexpression; oxidation; platelet function; receptor; response; thrombotic

Abstract Controlling thrombosis is central to management of various pathologies. Studies proposed here will identify a new, unexpected path to thrombosis, involving the enzyme Lysyl Oxidase (LOX). The proposed work looks at LOX effects on platelet function and thrombosis in primary myelofibrosis (PMF), a pathology hallmarked by proliferation and clustering of megakaryocytes, and myelofibrosis in bone marrow. Data from the Swedish Cancer Register from 1980 to 2009 showed that in a cohort of 11,155 patients with myeloproliferative neoplasms and 44, 620 matched healthy controls, the risk of arterial thrombosis was significantly 4.9-fold increased (4.8-5.0 p<0.001) in the patients compared to matched controls, highlighting the importance of studying regulators implicated in controlling thrombosis associated with this pathology. LOX is known to oxidize peptidyl lysines, leading to cross-linked matrix proteins in the bone marrow niche. Our published studied identified a link between LOX upregulation in megakaryocytes and bone marrow fibrotic progression in a mouse model of myelofibrosis. Importantly, more recently we found LOX expression to be vastly upregulated in platelets of patients with PMF (of which the majority tested carry the JAK2V617F or calreticulin gene mutations), compared to matching controls, and platelets isolated from PMF patients have greater adhesion to type I collagen, compared to controls. Further, transgenic mouse platelets engineered to overexpress LOX, at a level similar to cells from a myelofibrotic mouse model, show increased adhesion to monomeric collagen, and significantly greater propensity for arterial thrombosis in vivo. Considering LOX known activity, we hypothesized that it influences platelets through lysine oxidation of at least one of the collagen receptors, further supported by preliminary studies involving Oxy-Western blot analysis. Thus, we propose two specific aims of research: Aim 1. To explore the mechanism by which LOX affects platelet response to collagen using proteomics approaches, and mouse and human MPN samples; Aim 2. To determine the relevance of LOX- regulated thrombosis in vivo in myelofibrotic mice, and the role of collagen receptors in this effect. Pursuing these aims will be facilitated by: 1) a new transgenic mouse line produced in our lab, in which platelet LOX level is upregulated under wild type background (free of MPN), and 2) the availability of a new LOX inhibitor, and Lox gene loss of function studies. This work is innovative in that we are the first to reveal a link between LOX and collagen receptors activation. Further, these receptors were not suspected before to be regulated by oxidation. Our research is significant in that it will shed new light on basic mechanisms of platelet activation, in general, and implicate LOX in thrombotic events related to PMF, thus, recognizing the LOX pathway as new potential target for future anti-thrombosis drug development.

摘要 控制血栓形成是各种病理管理的核心。这里提出的研究将确定一个 新的，意想不到的途径血栓形成，涉及酶赖氨酰氧化酶（LOX）。拟议的工作着眼于 LOX对原发性骨髓纤维化（PMF）中血小板功能和血栓形成的影响， 巨核细胞的增殖和聚集以及骨髓中的骨髓纤维化。瑞典数据 1980年至2009年的癌症登记显示，在11，155例骨髓增生性疾病患者中， 肿瘤患者和44，620名匹配的健康对照者，动脉血栓形成的风险显著增加4.9倍 与匹配的对照组相比，患者中增加（4.8-5.0 p<0.001），突出了 研究涉及控制与这种病理学相关的血栓形成的调节剂。众所周知，LOX会氧化 肽基赖氨酸，导致骨髓龛中的交联基质蛋白。我们发表的研究 确定了巨核细胞中LOX上调与骨髓纤维化进展之间的联系， 小鼠骨髓纤维化模型。更重要的是，最近我们发现LOX表达在大肠杆菌中显著上调。 PMF患者的血小板（其中大多数检测携带JAK 2 V617 F或钙网蛋白基因 与匹配的对照相比，从PMF患者中分离的血小板对血小板的粘附性更高。 I型胶原蛋白，与对照组相比。此外，转基因小鼠血小板工程过表达LOX，在 与来自骨髓纤维化小鼠模型的细胞的水平相似，显示对单体胶原的粘附增加， 并且体内动脉血栓形成的倾向显著更大。考虑到液氧的已知活性，我们 假设它通过至少一种胶原受体的赖氨酸氧化影响血小板， 进一步得到了涉及Oxy-Western印迹分析的初步研究的支持。因此，我们提出了两个具体的 研究目的：目的1。探讨脂氧合酶影响血小板对胶原反应的机制。 蛋白质组学方法，以及小鼠和人类MPN样品;目的2.为了确定液氧的相关性- 调节骨髓纤维化小鼠体内血栓形成，以及胶原受体在这种作用中的作用。追求 这些目标将有助于：1）我们实验室生产的一种新的转基因小鼠系，其中血小板LOX 水平在野生型背景下上调（不含MPN），和2）新LOX抑制剂的可用性， 和Lox基因功能丧失的研究。这项工作是创新的，因为我们是第一个揭示之间的联系 LOX和胶原蛋白受体激活。此外，这些受体以前没有被怀疑是由 氧化我们的研究意义重大，因为它将揭示血小板活化的基本机制， 一般情况下，并暗示LOX与PMF相关的血栓形成事件有关，因此，将LOX途径视为新途径 是未来抗血栓药物开发的潜在靶点。

项目成果

The role of extracellular matrix stiffness in megakaryocyte and platelet development and function.

• DOI: 10.1002/ajh.25008
• 发表时间: 2018-03
• 期刊: American journal of hematology
• 影响因子: 12.8
• 作者: Leiva O;Leon C;Kah Ng S;Mangin P;Gachet C;Ravid K
• 通讯作者: Ravid K

Lysyl oxidase inhibition in primary myelofibrosis: A renewed strategy.

• DOI: 10.46439/stemcell.1.005
• 发表时间: 2020-12
• 期刊: Archives of stem cell and therapy
• 作者: Piasecki A;Leiva O;Ravid K
• 通讯作者: Ravid K

Cardiovascular Disease in Myeloproliferative Neoplasms: JACC: CardioOncology State-of-the-Art Review.

• DOI: 10.1016/j.jaccao.2022.04.002
• 发表时间: 2022-06
• 期刊: JACC: CARDIOONCOLOGY
• 影响因子: 11.1
• 作者: Leiva, Orly;Hobbs, Gabriela;Ravid, Katya;Libby, Peter
• 通讯作者: Libby, Peter

Novel lysyl oxidase inhibitors attenuate hallmarks of primary myelofibrosis in mice.

• DOI: 10.1007/s12185-019-02751-6
• 发表时间: 2019-12
• 期刊: International journal of hematology
• 影响因子: 2.1
• 作者: Leiva O;Ng SK;Matsuura S;Chitalia V;Lucero H;Findlay A;Turner C;Jarolimek W;Ravid K
• 通讯作者: Ravid K

Characterization of Glycoproteoforms of Integrins α2 and β1 in Megakaryocytes in the Occurrence of JAK2V617F Mutation-Induced Primary Myelofibrosis.

• DOI: 10.1016/j.mcpro.2022.100213
• 发表时间: 2022-04
• 期刊: Molecular & cellular proteomics : MCP
• 作者: Gaye MM;Ward CM;Piasecki AJ;Stahl VL;Karagianni A;Costello CE;Ravid K
• 通讯作者: Ravid K