Modeling PCa Bone Metastasis: Dual Role of E-selectin Ligands and Integrins

PCa 骨转移建模：E-选择素配体和整合素的双重作用

基本信息

批准号：
8474863
负责人：
Steven Russell Barthel
金额：
$ 6.04万
依托单位：
BRIGHAM AND WOMEN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-05-24 至 2013-05-23
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8474863
关键词：
Adherence Adhesions American Animals Avidity Binding Biological Assay Bioluminescence Blocking Antibodies Blood Blood Circulation Blood flow Bone Marrow Cell Adhesion Molecules Cells Cessation of life Comprehension Data Detection E-Selectin Endothelial Cells Goals Hematopoietic Hematopoietic stem cells Homing Image Immigration Immunoprecipitation Integrins Knowledge Lead Life Ligands Link Malignant Neoplasms Malignant neoplasm of prostate Mediating Metabolic Metastatic Neoplasm to the Bone Metastatic Prostate Cancer Metastatic to Methods Migration Assay Mission Modeling Molecular Movement Mus Neoplasm Metastasis Organ PC3 cell line Patients Phospho-Specific Antibodies Physiological Proteins Public Health Publications Published Comment Publishing RNA Interference Research Design Reverse Transcriptase Polymerase Chain Reaction Role Signal Pathway Signal Transduction Solid Neoplasm Suggestion Testing Time United States National Institutes of Health Up-Regulation base bone cancer cell cellular engineering cytokine galactoside 3-fucosyltransferase hemodynamics imaging modality in vivo inhibitor/antagonist innovation men migration neoplastic cell novel public health relevance small molecule treatment strategy

项目摘要

DESCRIPTION (provided by applicant): Metastatic prostate cancer (PCa) claimed the lives of 28,660 American men in 2008. Of note, 80- 90% of these patients presented with bone metastasis. Unfortunately, our comprehension of how bone metastasis is initiated is ill-defined. Our data shows that ligands for endothelial (E)-selectin are upregulated and functional on bone-metastatic PCa cells, while other groups have shown that b1 and b3 integrins correspond to PCa cell avidity for bone marrow endothelial cells (BMEC) and to metastatic potential. Since bone-homing hematopoietic stem cells also utilize E-selectin ligands and integrins for adherence and transendothelial migration (TEM) into bone marrow (BM), we and others believe that a similar molecular repertoire is necessary for TEM of PCa cells. Objective/Hypothesis: We hypothesize that circulating PCa cells adhere to BMEC through adhesion to E- selectin and then firmly adhere to and traverse BMEC via b1 and b3 integrin adhesion molecules. Our objective is to determine how PCa cell rolling on BMEC E-selectin causes PCa cell TEM into BM via b1 and b3 integrins. Specific Aims: (1) To investigate the role of E-selectin ligands and b1 and b3 integrins in TEM of PCa cells; (2) To investigate the role of E-selectin ligands and b1 and b3 integrins in the migration of PCa cells into bone in vivo. Study Design: (1) TEM of PCa cells engineered to express E-selectin ligand and expressing b1 and b3 integrin heterodimers, E-selectin ligand+/b1+/b3+ PCa cells, will be analyzed using state-of-the-art migration assays under physiologic blood flow conditions. Control PCa cell lines, blocking antibodies, RNAi, small molecule antagonists and metabolic inhibitors will be employed to control for E-selectin ligand/integrin function. (2) In an experimental bone metastasis model, we will assay the migration of PCa cells engineered to express E-selectin ligand and expressing b1 and b3 integrins using a highly sensitive PCR detection method and bioluminescence imaging of live animals. Control PCa cell lines, blocking antibodies, RNAi, small molecule antagonists and metabolic inhibitors will also be employed to control for E-selectin ligand/integrin function. Signaling pathways regulating E-selectin ligand expression in PCa cells will be assessed by RT-PCR, phospho-specific antibodies and pharmacologic inhibitors; physical interaction between E-selectin ligands and integrins will be determined by immunoprecipitation; role of an E-selectin ligand regulator, a1,3 fucosyltransferase 7, in integrin activation will be assessed by flow cytometric and E-selectin cell binding assays. Cancer relevance: Determining how PCa cells enter bone is vital to public health and to the mission of the NIH, as results from our studies could lead to treatment strategies that would diminish metastasis-related deaths. PUBLIC HEALTH RELEVANCE: These studies will help define a critical, yet under-appreciated and ill-characterized step in the metastasis of a solid tumor cell into a vital organ. Determining how PCa cells enter bone is vital to public health and to the mission of the NIH as results from our studies could lead to treatment strategies that might greatly diminish metastasis- related deaths.

描述（由申请人提供）：2008 年，转移性前列腺癌 (PCa) 夺去了 28,660 名美国男性的生命。值得注意的是，这些患者中 80-90% 出现骨转移。不幸的是，我们对骨转移如何发生的理解尚不明确。我们的数据显示，内皮 (E)-选择素的配体在骨转移性 PCa 细胞上表达上调并发挥作用，而其他研究组则表明 b1 和 b3 整合素与 PCa 细胞对骨髓内皮细胞 (BMEC) 的亲合力和转移潜力相对应。由于骨归巢造血干细胞也利用 E-选择素配体和整合素进行粘附和跨内皮迁移 (TEM) 进入骨髓 (BM)，因此我们和其他人认为类似的分子库对于 PCa 细胞的 TEM 是必要的。目的/假设：我们假设循环 PCa 细胞通过粘附 E-选择素粘附到 BMEC，然后通过 b1 和 b3 整合素粘附分子牢固粘附并穿过 BMEC。我们的目标是确定 PCa 细胞在 BMEC E-选择素上滚动如何通过 b1 和 b3 整合素导致 PCa 细胞 TEM 进入 BM。具体目的： (1) 研究E-选择素配体和b1、b3整合素在PCa细胞TEM中的作用； (2)研究E-选择素配体和b1、b3整合素在体内PCa细胞向骨迁移中的作用。研究设计：(1) 将使用最先进的迁移测定法对经过工程改造以表达 E-选择素配体并表达 b1 和 b3 整联蛋白异二聚体、E-选择素配体+/b1+/b3+ PCa 细胞的 TEM 进行分析生理血流状况。对照 PCa 细胞系、阻断抗体、RNAi、小分子拮抗剂和代谢抑制剂将用于控制 E-选择素配体/整联蛋白功能。 (2) 在实验性骨转移模型中，我们将使用高灵敏度 PCR 检测方法和活体动物生物发光成像来测定表达 E-选择素配体并表达 b1 和 b3 整合素的 PCa 细胞的迁移。对照 PCa 细胞系、阻断抗体、RNAi、小分子拮抗剂和代谢抑制剂也将用于控制 E-选择素配体/整合素功能。将通过 RT-PCR、磷酸化特异性抗体和药物抑制剂评估调节 PCa 细胞中 E-选择素配体表达的信号通路； E-选择素配体和整合素之间的物理相互作用将通过免疫沉淀测定； E-选择素配体调节剂 a1,3 岩藻糖基转移酶 7 在整合素激活中的作用将通过流式细胞术和 E-选择素细胞结合测定进行评估。癌症相关性：确定 PCa 细胞如何进入骨骼对于公共健康和 NIH 的使命至关重要，因为我们的研究结果可能会导致制定减少转移相关死亡的治疗策略。公众健康相关性：这些研究将有助于确定实体瘤细胞转移至重要器官过程中的一个关键但未被充分认识且特征不明的步骤。确定 PCa 细胞如何进入骨骼对于公共健康和 NIH 的使命至关重要，因为我们的研究结果可能会导致治疗策略的出现，从而大大减少与转移相关的死亡。