Defining the formation and function of carcinoma-associated mesenchymal stem cells in the ovarian cancer microenvironment

定义卵巢癌微环境中癌相关间充质干细胞的形成和功能

• 批准号: 10006503
• 负责人: Lan Coffman
• 金额: $ 16.61万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-15 至 2022-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10006503
• 关键词: Acidosis; Activities of Daily Living; Adipocytes; Bone Marrow; Bone Morphogenetic Proteins; Carcinoma; Cell Hypoxia; Cell physiology; Cells; Chemicals; Coculture Techniques; Communication; Complex; DNA Methylation; Data; Development; Diffuse; Education; Elements; Epigenetic Process; Exhibits; Expression Profiling; Fibroblasts; Foundations; Future; Genotype; Goals; Growth; HIF1A gene; Hyperactive behavior; Hypoxia; Hypoxia Inducible Factor; Hypoxia-Inducible Factor Pathway; In Vitro; Intra-abdominal; Knock-out; Knockout Mice; Knowledge; Leadership; Learning; Malignant - descriptor; Malignant Female Reproductive System Neoplasm; Malignant Neoplasms; Malignant neoplasm of ovary; Mediator of activation protein; Mentors; Mentorship; Mesenchymal Stem Cells; Modification; Molecular; Multipotent Stem Cells; Mutation; Myofibroblast; Neoplasm Metastasis; Non-Malignant; Normal Cell; Normal tissue morphology; Phenotype; Physicians; Play; Population; Property; Research; Research Training; Resistance; Role; Scientist; Signal Pathway; Signal Transduction; Site; Stromal Cells; Stromal Neoplasm; System; Testing; Training; Transgenic Mice; Treatment outcome; Tumor Promotion; Woman; Work; behavior influence; cancer cell; cancer stem cell; career; cell growth; cell type; chemotherapy; conditioning; epigenetic regulation; genome editing; genome-wide; improved outcome; in vivo; innovation; intraperitoneal; migration; molecular phenotype; mortality; mouse model; multipotent cell; neoplastic cell; novel; novel therapeutic intervention; novel therapeutics; ovarian neoplasm; protein expression; skills; small molecule; stem-like cell; stemness; success; tumor; tumor growth; tumor microenvironment; tumorigenic

ABSTRACT: Defining the formation and function of carcinoma-associated mesenchymal stem cells in the ovarian cancer microenvironment Ovarian cancer is the most deadly US gynecologic malignancy with a mortality rate that exceeds 50% at 5 years. Ovarian cancer is characterized by early intraperitoneal metastasis and the development of a complex microenvironment which supports tumor cell growth, survival and spread. Understanding and eventually targeting this cancer-promoting tumor microenvironment offers the potential for powerful new therapeutic approaches. My ultimate goal is to become a world-class independent physician scientist studying the ovarian cancer microenvironment in order to develop new treatments and improve outcomes for women with ovarian cancer. This proposal describes important and innovative research which will lay the foundation for my future career in addition to providing the necessary skills and mentorship vital for my success. The ovarian tumor microenvironment (TME) is a diverse system of cellular and chemical components. The cellular TME includes tumor cells and non-malignant stromal cells. The chemical TME is marked by acidosis and hypoxia. Carcinoma-associated mesenchymal stem cells (CA-MSCs) are multi-potent stromal cells within the cellular TME that can differentiate into multiple pro-tumorigenic stromal cell types including fibroblasts, myofibroblasts, and adipocytes. CA-MSCs are genotypically normal without malignant potential but are functionally different than normal tissue or bone marrow derived MSCs. Compared to normal MSCs, CA-MSCs demonstrate a unique molecular phenotype with very high expression of bone morphogenetic proteins (BMPs). Due to this unique phenotype, these CA-MSCs strongly promote ovarian cancer growth, enhance chemotherapy resistance and enrich the cancer stem cell-like population. How CA-MSCs develop their unique phenotype remains unclear. My preliminary data indicate that tumor secreted factors induce some of the molecular changes associated with CA-MSCs. Another potential mediator of the CA-MSC phenotype is hypoxia. Hypoxia is a hallmark of the chemical TME known to impact normal MSC function. In cancer, hypoxia influences tumor:stromal interactions and hypoxia is a key regulator of BMP expression—high levels of which characterize ovarian cancer CA-MSCs. Preliminary data indicates that hypoxia enhances the ability of tumor cells to induce a CA-MSC expression profile in normal MSCs. While the mechanism of this induction is unknown, given CA-MSCs are genetically normal yet maintain their unique phenotype across multiple passages, tumor-induced epigenetic regulation may be critical to the formation of the CA-MSC phenotype. Indeed, preliminary data indicates CA-MSCs exhibit significant hypomethylation compared to normal MSCs. In addition to influencing the formation of a CA-MSC, hypoxia may also critically regulate the function of CA- MSCs already established in the ovarian TME. My preliminary data suggests that hypoxia maintains the “stemness” of CA-MSCs slowing growth and maintaining differentiation capacity. Further, my data suggests that the hypoxia inducible factor pathway, the main hypoxia signaling pathway, is hyper-active in CA-MSCs compared to normal MSCs. Thus hypoxia may be a critical modulator of CA-MSCs within the ovarian TME. My main research goal is to understand how CA-MSCs obtain their unique phenotype and subsequently interact with and influence the function of the ovarian cancer microenvironment. To achieve this goal, I propose two specific aims: 1) Determine the ability of normal MSCs to acquire a CA-MSC-like phenotype 2) Determine the impact of hypoxia on established CA-MSCs within the tumor microenvironment. In aim 1, I hypothesize that tumor cell conditioning under hypoxia induces normal MSCs to become CA-MSCs. To test this I will perform cancer cell: normal MSC co-cultures under normoxia and hypoxia to determine if cancer cells can functionally turn a normal MSC into a CA-MSC. I will also explore differential DNA methylation as a mechanism for the creation of a CA-MSC. In aim 2, I focus on already established CA-MSCs. I hypothesize that hypoxia enhances the pro-tumorigenic effects of established CA-MSCs within the tumor microenvironment. To test this, I will utilize conditional HIF pathway knockout mice and CRISPER/CAS9 genome editing to assess the impact of hypoxia and HIF signaling on established CA-MSCs. In addition to furthering our understanding of CA-MSCs in ovarian cancer, the proposed research and training will cultivate expertise necessary for an independent career studying the ovarian TME. Through the support of Dr. Laird, I will master the assessment of genome-wide epigenetic modifications and the analysis of large scale “omics” data. Dr. Schipani will facilitate my education in hypoxia and HIF signaling. Through Dr. Schipani and Dr. Cho, I will learn to generate and manipulate transgenic mouse models. Dr. Buckanovich's ongoing mentorship will further my expertise in the function of the ovarian TME and, together with my mentoring committee, will help develop my leadership, team-building and communication skills. By the end of the training period, I will have developed a novel skill set which merges the expertise of multiple scientific leaders yielding a uniquely trained physician scientist ideal for the study of the ovarian cancer microenvironment.

摘要：定义与癌相关间充质干细胞的形成和功能 卵巢癌微环境 卵巢癌是美国最致命的妇科恶性肿瘤，死亡率为50％，在5 卵巢癌的特征是早期 支持肿瘤细胞生长，生存和扩散的微环境。 针对这种癌症 - 促进肿瘤微环境为有力的新治疗提供了潜力 方法。我的最终目标是成为世界一流的独立医生 癌症微环境以开发新的治疗方法并改善卵巢女性的结果 癌症。 职业除了提供对我成功至关重要的必要技能和指导。 卵巢肿瘤微环境（TME）是细胞和化学成分的多样化系统 细胞TME包括肿瘤细胞和非恶性基质细胞。 和缺氧。 可以分化为多形的促肿瘤基质细胞类型的细胞，包括成纤维细胞， 肌成纤维细胞和脂肪细胞在基因型上是正常的，没有恶性潜力 与正常的MSC相比，与正常组织或骨髓的功能不同。 展示具有骨形态发生蛋白（BMP）高表达的独特分子表型。 由于独特的表型，这些CA-MSC强烈促进卵巢癌的生长，增强 化学疗法耐药性并富含癌症干细胞样群体。 CA-MSC如何发展其独特的表型尚不清楚。 分泌的因素诱导了与CA-MSC相关的一些分子变化 CA-MSC表型的缺氧是缺氧的标志 癌症中的MSC功能，缺氧影响肿瘤：基质相互作用和缺氧是BMP的关键调节剂 表达最高水平的特征是卵巢癌CA-MSC。 缺氧增强了肿瘤细胞在正常MSC中诱导CA-MSC表达谱的能力。 鉴于CA-MSC是固定的，但保持其独特 跨多个段落的表型，肿瘤诱导的表观遗传调节可能对形成至关重要 CA-MSC表型的确实，初步数据表明CA-MSC表现出明显的低甲基化 与正常的MSC相比。 除了影响CA-MSC的形成外，缺氧还可以批评调节CA-的功能 MSC已经在卵巢TME中建立。 CA-MSC的“茎”速度慢，并保持区分能力。 在CA-MSC中，低氧诱导因子途径（主要的缺氧信号通路）非常活跃 与正常的MSC相比，缺氧可能是卵巢TME内CA-MSC的关键模块。 我的主要研究目标是了解CA-MSC如何获得其独特的表型，然后再获得其独特的表型 与卵巢癌微侵略性的功能相互作用并影响该目标。 两个具体的目标： 1）确定正常MSC获得CA-MSC样表型的能力 2）确定缺氧对肿瘤微观传染内已建立的CA-MSC的影响。 在AIM 1中，我假设缺氧下的肿瘤细胞调节会诱导正常的MSC成为CA-MSC。 为此，我将执行癌细胞：正常氧和缺氧下的正常MSC共培养 癌细胞在功能上可以将正常的MSC变成CA-MSC。 作为在AIM 2中创建CA-MSC的机制，我专注于已经建立的CA-MSC 假设缺氧增强了肿瘤内建立的CA-MSC的促肿瘤学 微环境要测试，我将使用有条件的HIF途径淘汰小鼠和Crisper/cas9 基因组编辑以评估缺氧和HIF信号对已建立的CA-MSC的影响。 除了进一步了解卵巢癌中的CA-MSC之外，拟议的研究和培训 将通过支持TME的独立职业来培养专业知识。 Laird博士，我将掌握全基因组表观遗传修饰的评估和大规模分析 “ OMICS”数据是我在缺氧和HIF信号的教育。 Cho博士，我将学会生成和操纵转基因鼠标模型。 指导将进一步提高我在卵巢TME功能方面的专业知识，以及我的指导 委员会将在培训结束时帮助我发展领导力，团队建设和沟通能力。 时期，我将开发出一种新颖的技能，该技能融合了多个领导者的实验 一位无权的医师科学家，非常适合研究卵巢癌微效率。