Cell signaling as a leukemia biomarker

细胞信号传导作为白血病生物标志物

• 批准号: 8243064
• 负责人: James W Jacobberger
• 金额: $ 20.49万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-15 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8243064
• 关键词: Abnormal Cell; Acute Myelocytic Leukemia; Affect; Age; Apoptosis; Aspirate substance; Biological Assay; Biological Markers; Blast Cell; Bone Marrow; Bone Marrow Cytometry; Cell Proliferation; Cells; Clinical; Clinical Trials; Clonal Evolution; Commit; Cytometry; Data; Data Set; Dependence; Detection; Diagnosis; Differentiation Antigens; Disease; Disease Progression; Dysmyelopoietic Syndromes; Early Diagnosis; Early treatment; Epitopes; Exons; FLT3 gene; Feasibility Studies; Flow Cytometry; Frequencies; Future; Gender; Genes; Genetic; Genome; Genotype; Goals; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic stem cells; Hour; Human; Human Volunteers; IL3 gene; Immunofluorescence Immunologic; Kinetics; Lesion; Leukemic Cell; Ligands; Measurement; Measures; Membrane; Modeling; Molecular Abnormality; Monitor; Mutate; Mutation; Myelogenous; Myeloid Leukemia; Neoplasms; Noise; Normal Range; Pathway interactions; Patient Monitoring; Patients; Pattern; Phase; Phenotype; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Population; Process; Prospective Studies; Proteins; Proto-Oncogene Protein c-kit; Publishing; Receptor Protein-Tyrosine Kinases; Reproducibility; Sample Size; Sampling; Scaffolding Protein; Signal Pathway; Signal Transduction; Signaling Protein; Span 20; Specificity; Statistical Methods; Stem Cell Factor; Surface; Syndrome; System; Testing; Therapeutic; Time; Transcription factor genes; Woman; Work; base; cytokine; enzyme pathway; genetic analysis; healthy volunteer; leukemia; men; novel; outcome forecast; precursor cell; response; sex; stem; success; therapeutic target; tool; transcription factor; tumorigenesis; Abnormal Cell; Acute Myelocytic Leukemia; Affect; Age; Apoptosis; Aspirate substance; Biological Assay; Biological Markers; Blast Cell; Bone Marrow; Bone Marrow Cytometry; Cell Proliferation; Cells; Clinical; Clinical Trials; Clonal Evolution; Commit; Cytometry; Data; Data Set; Dependence; Detection; Diagnosis; Differentiation Antigens; Disease; Disease Progression; Dysmyelopoietic Syndromes; Early Diagnosis; Early treatment; Epitopes; Exons; FLT3 gene; Feasibility Studies; Flow Cytometry; Frequencies; Future; Gender; Genes; Genetic; Genome; Genotype; Goals; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic stem cells; Hour; Human; Human Volunteers; IL3 gene; Immunofluorescence Immunologic; Kinetics; Lesion; Leukemic Cell; Ligands; Measurement; Measures; Membrane; Modeling; Molecular Abnormality; Monitor; Mutate; Mutation; Myelogenous; Myeloid Leukemia; Neoplasms; Noise; Normal Range; Pathway interactions; Patient Monitoring; Patients; Pattern; Phase; Phenotype; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Population; Process; Prospective Studies; Proteins; Proto-Oncogene Protein c-kit; Publishing; Receptor Protein-Tyrosine Kinases; Reproducibility; Sample Size; Sampling; Scaffolding Protein; Signal Pathway; Signal Transduction; Signaling Protein; Span 20; Specificity; Statistical Methods; Stem Cell Factor; Surface; Syndrome; System; Testing; Therapeutic; Time; Transcription factor genes; Woman; Work; base; cytokine; enzyme pathway; genetic analysis; healthy volunteer; leukemia; men; novel; outcome forecast; precursor cell; response; sex; stem; success; therapeutic target; tool; transcription factor; tumorigenesis

DESCRIPTION (provided by applicant): The long term objectives of this proposal are to develop a system to monitor patients with pre-leukemic conditions like myelodysplastic syndrome (MDS) and myeloid proliferative neoplasms (MPN) for early detection of leukemic transition. This general system is stimulation of unfractionated bone marrow aspirates with cytokines and measuring the fine kinetics of modified epitopes on signaling proteins by immunofluorescence flow cytometry. Leukemia is an evolutionary process that works by clonal selection. Cytometry is a measurement sys- tem that works at the level of single cells, therefore, the specificity of detection and signal to noise ratio can be very high. In addition, cytometry is an established clinical tool used for diagnosis of hematopoietic malignancies based on measuring the levels of differentiation antigens. One model for the genesis of acute myeloid leukemia (AML) is that at least two mutations are leukemogenic: 1) in a transcription factor that alters differentiation, and 2) in a signaling pathway protein that affects cell proliferation and/or apoptosis. A significant component of cell signaling involves phosphorylation/de-phosphorylation cycles on pathway proteins, essentially from membrane surfaces to the genome, and between pathways (networks). We have developed the ability to measure cell signaling in committed myeloid precursor cells in unfractionated bone marrow by flow cytometry. Published results on normal, healthy volunteers demonstrates remarkable uniformity of response across age and gender, and published results on 4 (Jacobberger lab) and 14 (Goolsby lab) leukemias demonstrates clear, robust detection of abnormal signaling in eight pathways (Kit?Erk; ?Akt; ?S6; ?Stat5, and Flt3?Erk; ?Akt; ?S6; ?Stat5) that demonstrate classifiable complexity. Thus, we propose that cell signaling can be used as a biomarker to detect leukemic cells with high fidelity. Additionally, since the enzymes of these pathways are focus of targeted therapy, this approach provides information with therapeutic implications. Because a small fraction of abnormal cells can be detected by this approach, this should work well for early detection. Current diagnosis, therapeutic choices, and selection for clinical trials depend to some degree on genetic analysis, and in the future, this dependence will be greater. The signaling patterns, identified by our approach, will associate with specific mutations in signaling protein genes. Thus, cell signaling may provide rapid genetic inferences. To test this idea, we propose to correlate phenotype (signaling) and genotype, determined by high throughput sequencing. Our specific aims are 1) to measure signaling for 2 ligands (Stem cell factor and Flt3-ligand) and 5 endpoints (Erk, Akt, S6, Stat3, and Stat5) on committed myeloid precursor cells in 60 healthy human donors to define the normal ranges for each component of the signal analysis and determine any age or gender effects; 2) to measure signaling in 75 MDS and 75 AML patient blast cells to test the hypothesis that signaling can be classified by patterns and determine any age or gender effects; 3) to sequence 182 gene exons that may should impact on the signaling patterns for 10 MDS and 10 AML patients to demonstrate the feasibility of this approach for associating phenotype to genotype. PUBLIC HEALTH RELEVANCE: Monitoring myeloid dysplastic syndrome (MDS) and related myeloid proliferative neoplasm (MPN) patients by measuring the transition of hematopoietic stem / committed myeloid precursor cell signaling patterns to altered patterns, correlated with genetic change, should be a sensitive, yet robust, early detection system for oncogenesis. Because the measurements monitor signaling pathways that are known to depend on genes that are rarely mutated in MDS and MPN but often mutated in secondary acute myeloid leukemia (sAML) and are the focus of experimental targeted therapeutics, the idea of early intervention, prior to the onset of overt disease, could be entertained. There are three steps toward success: 1) feasibility studies, 2) prospective studies, and 2) clinical trials. This proposal represents the first step.

描述（由申请人提供）：该提案的长期目标是开发一种系统，以监测具有骨髓增生性综合征（MDS）和髓样增殖性肿瘤（MPN）等美质发育性综合征（MDS）（MPN）的患者，以早期发现白血病过渡。该通用系统是用细胞因子刺激未分流的骨髓吸液，并通过免疫荧光流式细胞仪测量信号蛋白上修饰的表位的细胞动力学。白血病是一种进化过程，可通过克隆选择起作用。细胞仪是在单个细胞水平上起作用的测量系统，因此，检测和信号与噪声比的特异性可能非常高。此外，基于测量分化抗原的水平，细胞仪是一种已建立的临床工具，用于诊断造血恶性肿瘤。急性髓样白血病（AML）起源的一种模型是，在转录因子中至少有两个突变具有变化的分化，而2）在影响细胞增殖和/或凋亡的信号通路蛋白中。细胞信号传导的重要组成部分涉及途径蛋白上的磷酸化/去磷酸化周期，从膜表面到基因组以及途径（网络）之间。我们已经开发了通过流式细胞仪中未分离的骨髓骨髓中固定髓样前体细胞中细胞信号传导的能力。关于正常，健康志愿者的已发表结果表明，整个年龄和性别的反应都显着统一，并在4（Jacobberger Lab）和14（Goolsby Lab）白血病上发表的结果表明，在八个途径（Kit？erk;？akt;？akt;？s65;？STAT5;？stat5;？stat5＆lt flt 3; s6;？可分类的复杂性。因此，我们提出可以将细胞信号用作生物标志物，以检测具有高富度性的白血病细胞。此外，由于这些途径的酶是靶向治疗的重点，因此该方法提供了具有治疗意义的信息。由于可以通过这种方法检测到一小部分异常细胞，因此这应该很好地用于早期检测。当前的诊断，治疗选择和临床试验的选择在某种程度上取决于遗传分析，将来，这种依赖性将更大。通过我们的方法识别的信号传导模式将与信号蛋白基因中的特定突变相关联。因此，细胞信号传导可能会提供快速的遗传推断。为了测试这个想法，我们建议通过高吞吐量测序确定的表型（信号）和基因型。我们的具体目的是1）在60位健康的人类供体中承诺的髓样前体细胞上测量2个配体（干细胞因子和FLT3-rigand）和5个终点（ERK，AKT，S6，STAT3和Stat5）的信号传导，以定义60位健康的人类供体中的髓样前体细胞，以定义信号分析的每个成分的正常范围，并确定任何年龄或性别效应的正常范围； 2）测量75 MD和75个AML患者爆炸细胞中的信号传导，以测试可以通过模式对信号进行分类并确定任何年龄或性别效应的假设； 3）序列182基因外显子，可能会影响10个MDS和10名AML患者的信号传导模式，以证明这种方法将表型与基因型关联的可行性。 公共卫生相关性：通过测量造血性茎 /承诺的髓样前体信号传导模式到改变的模式，监测髓样不增生综合征（MDS）和相关的髓样增殖性肿瘤（MPN）患者，应与遗传变化相关，应与遗传变化相关。由于测量值监测已知取决于MDS和MPN中很少突变但经常在继发性急性髓样白血病（SAML）中突变的基因的信号传导途径，并且在公开疾病发作之前是实验性靶向疗法的重点，是实验性靶向疗法的重点，即早期干预的概念。成功有三个步骤：1）可行性研究，2）前瞻性研究和2）临床试验。该提议代表了第一步。