Investigating the role of the JAK/STAT3 Pathway in Clonal Proliferation and Immune Dysfunction in Myelodysplastic Syndrome

项目摘要 骨髓增生异常综合征(MDS)是一组不同类型的白血病前期骨髓衰竭疾病，具有 在美国每年的发病率约为13,000。大约三分之一的MDS患者将继续 发展为急性髓系白血病(AML)，存活率低达5-10%。标准护理治疗仅延长 那些需要治疗的人的中位寿命减少了9个月，而且除了茎外还没有治愈的疾病 细胞移植。随着人口老龄化，这种疾病只会变得更加常见，因此有一个紧迫的问题 需要对MDS进行更有效的治疗。MDS的发病机制是以克隆性髓系增殖为标志的 和骨髓免疫功能障碍。在MDS骨髓中，抑制性免疫微环境 对最初的炎症作出反应，并促进发育中的母细胞的增殖，这些母细胞是 可以自由生长，不受免疫系统的限制。事实上，髓系来源的抑制细胞(MDSCs)，髓系细胞 它们对T细胞功能具有抑制作用，与疾病的发生发展密切相关。这个 JAK/STAT3通路在髓系分化中起关键作用，在成髓细胞和成髓细胞中均异常激活 MDSC，两者都随着MDS的进展而增加。为了更好地理解MDS中的JAK/STAT3信号， 我们将使用单细胞和功能基因组分析来分析初级患者样本中的STAT3，特别是 重点关注成髓细胞和MDSCs--MDS白血病进展中的两种独立但中心的细胞类型。vt.给出 我们在研究AML的单细胞表型和细胞内信号转导方面的经验，我们将调整我们的方法 研究MDS，特别关注p-STAT3。这种方法的优点是能够测量所有单元格 同时在样本内并为每个样本创建单细胞信令配置文件，并且响应于 抑制力。我们提出了三个特定的目标来研究MDS中的JAK/STAT3信号。在目标I中，基本的和 将在储存的MDS样本中测量细胞因子诱导的STAT3信号反应，以了解 STAT3信号在白血病进展中的作用。AIM II将测试对各种 JAK/STAT3途径的抑制剂。最后，AIM III将确定标准疗法对JAK/STAT3的影响 抗性MDS中的信号转导和STAT3靶点。染色质免疫沉淀(CHIP-SEQ)将在这里用于 目的研究MDS中STAT3的基因靶点。这些研究将对我们的基础工作做出重大贡献 了解MDS和JAK/STAT3通路在这种疾病中的治疗潜力。 这项建议的最终目的是让费雷尔博士发展更多的技能，在质量细胞分析，磷酸- Flow、免疫学技术和芯片，并为他提供必要的关键指导和培训 成为一名独立的内科医生兼科学家。

Project Summary Myelodysplastic syndrome (MDS) is a heterogeneous group of pre-leukemic bone marrow failure disorders with a yearly incidence of approximately 13,000 in the U.S. Approximately one third of MDS patients will go on to develop acute myeloid leukemia (AML) with a dismal survival of 5-10%. Standard of care therapy only extends life by a median of 9 months in those requiring treatment and there is no cure for the disease outside of stem cell transplant. As the population ages, this disease will only become more common, thus there is an urgent need for more effective treatments in MDS. MDS pathogenesis is marked by both clonal myeloid proliferation and bone marrow immune dysfunction. In MDS bone marrow, a suppressive immune microenvironment develops in response to initial inflammation and contributes to the proliferation of developing blasts, which are free to grow unchecked by the immune system. In fact, myeloid-derived suppressor cells (MDSCs), myeloid cells which are suppressive to T cell function, have been implicated in disease pathogenesis and progression. The JAK/STAT3 pathway is critical in myeloid differentiation and is aberrantly activated in both myeloblasts and MDSCs, both of which increase as MDS progresses. In order to better understand JAK/STAT3 signaling in MDS, we will use single cell and functional genomic assays to profile STAT3 in primary patient samples, with particular focus on myeloblasts and MDSCs – two separate but central cells types in leukemic progression of MDS. Given our experience studying single cell phenotype and intracellular signaling in AML, we will adapt our methods to study MDS, with particular focus on p-STAT3. An advantage of this approach is the ability to measure all cells within a sample simultaneously and create single cell signaling profiles for each sample and in response to inhibition. We propose three specific aims to investigate JAK/STAT3 signaling in MDS. In Aim I, the basal and cytokine-induced responses of STAT3 signaling will be measured in banked MDS samples to understand the role of STAT3 signaling in leukemic progression. Aim II will test functional and signaling responses to various inhibitors of the JAK/STAT3 pathway. Finally, Aim III will determine the impact of standard therapy on JAK/STAT3 signaling and STAT3 targets in resistant MDS. Chromatin immunoprecipitation (ChIP-seq) will be used here in order to profile the gene targets of STAT3 in MDS. These studies will significantly contribute to our basic understanding of MDS and the therapeutic potential of the JAK/STAT3 pathway moving forward in this disease. This proposal is ultimately designed to allow Dr. Ferrell to develop further skills in mass cytometry, phospho- flow, immunology techniques and ChIP and to provide him with the critical mentoring and training necessary to become an independent physician-scientist.