Tumor and Immune Programming of Tumor-Associated Endothelium

项目摘要/摘要 血管内皮细胞(BEC)通过血管生成和分化调控肿瘤生长 支持血管，并通过调节免疫细胞募集来引导宿主对癌症的免疫反应 鲜血。最近的研究强调了bec的特殊亚群在肿瘤生物学中的重要性，包括 高内皮微静脉(HEV，专用于淋巴细胞募集的血管)，有助于直接有效 肿瘤免疫。尽管它们在肿瘤生物学中起着核心作用，但在分子水平上对bec知之甚少。 水平，调节肿瘤血管生成的机制，以及引起肿瘤血管生成的前体 血管生成的肿瘤EC和肿瘤相关的白细胞募集血管仍有待确定。 在目标1下，我们将应用最先进的单细胞高维质量标记(CyTOF)流式细胞术 以及单细胞RNAseq分析，以揭示肿瘤及其环境中EC亚群的多样性， 明确肿瘤血管生成过程中增殖反应的动力学和亚群特异性，并监测 功能性HEV和其他用于免疫细胞运输的毛细血管后小静脉(PCV)的出现和成熟。 轨迹分析将揭示识别的子集的发展关系，包括候选 祖细胞，免疫荧光组织学和共聚焦组织成像将定义它们在 肿瘤及相关血管系统。单细胞BEC信号将被映射到肿瘤血管系统 定量组织免疫组织学。在目标2下，创新的命运测绘方法将阐明 BEC子集之间的前体-产物关系，并将定义前体对 血管生成肿瘤EC、支持血管的特化EC、高内皮细胞。目标3将会是我的地雷 诱导EC亚群的转录谱及应用PAN-EC和EC亚群特异性诱导基因 靶向系统确定控制祖细胞和放大肿瘤EC的途径和机制 激活，并指导肿瘤相关HEV的分化。将应用新的生物信息学工具 揭示在免疫治疗环境中诱导招募血管的转录程序和途径。 制作一份全面的血液内皮细胞亚群、分子表型和 对肿瘤发生的反应将开辟癌症生物学和免疫学研究的新领域。 阐明内皮细胞特化和内环境稳定的机制，包括机制 调节控制淋巴细胞进入肿瘤的内皮细胞，可能会导致新的靶点和 加强癌症免疫治疗的方法。

PROJECT SUMMARY/ABSTRACT Blood vascular endothelial cells (BEC) control tumor growth through angiogenesis and differentiation of support vessels, and direct the host immune response to cancer by regulating immune cell recruitment from the blood. Recent studies emphasize the importance of specialized subsets of BEC in tumor biology, including high endothelial venules (HEV, vessels specialized for lymphocyte recruitment) which help direct effective tumor immunity. In spite of their central role in tumor biology, little is known about the BEC at the molecular level, mechanisms that regulate tumor angiogenesis are poorly understood, and the precursors that give rise to angiogenic tumor EC and to tumor-associated leukocyte-recruiting vessels remain to be determined. Under Aim 1 we will apply state-of-the-art single cell high dimensional mass label (CyTOF) flow cytometry and single cell RNAseq analyses to uncover the diversity of EC subsets in tumors and their environment, define the kinetics and subset-specificity of proliferative responses during tumor angiogenesis, and monitor the emergence and maturation of functional HEV and other post capillary venules (PCV) for immune cell traffic. Trajectory analyses will reveal developmental relationships of identified subsets including candidate progenitors, and immunofluorescence histology and confocal tissue imaging will define their location within the tumor and associated vasculature. Single cell BEC signatures will be mapped to the tumor vasculature using quantitative tissue immunohistology. Under Aim 2, innovative fate mapping approaches will elucidate precursor-product relationships among BEC subsets and will define clonal contributions of precursors to angiogenic tumor EC, to specialized EC of support vessels, and to high endothelium. Aim 3 will mine transcriptional profiles of induced EC subsets and apply pan-EC and EC subset-specific inducible gene targeting systems to define pathways and mechanisms that control progenitor cell and amplifying tumor EC activation, and that direct the differentiation of tumor-associated HEV. Novel bioinformatics tools will be applied to uncover transcriptional programs and pathways that induce recruiting vessels in settings of immunotherapy. Generation of a comprehensive atlas of blood endothelial cell subsets, molecular phenotypes and responses to tumorigenesis will open up new areas of investigation in cancer biology and immunology. Elucidation of the mechanisms of endothelial cell specialization and homeostasis, including mechanisms regulating endothelial cells that control lymphocyte traffic into tumors, may lead to novel targets and approaches to enhance cancer immunotherapies.