Real-time Imaging of Immune Cell-Tumor Cell Interactions

描述(由申请人提供)：肿瘤细胞-宿主细胞相互作用是肿瘤进展的关键参数。尤其重要的是肿瘤细胞和免疫细胞之间的关系，它们要么介导肿瘤破坏，要么促进肿瘤生长。免疫细胞如何在肿瘤微环境中导航，它们如何与肿瘤细胞相互作用，以及它们对肿瘤微环境的总体贡献还没有被很好地理解。该项目的长期目标是确定负责免疫细胞-肿瘤细胞相互作用的细胞和分子线索，以便优化免疫治疗策略。这类研究的一个主要障碍是我们至今无法在空间和时间上以足够高的分辨率成像完整组织内部的细胞相互作用。这项建议的目的是通过在体内单细胞水平上的实时成像来洞察肿瘤和免疫细胞之间的复杂相互作用。我们建议使用活体双光子显微镜(2P-IVM)直接跟踪T细胞和树突状细胞在其自然环境中的活动。我们的假设是，这些免疫细胞在肿瘤环境中的迁移模式以及它们与肿瘤细胞相互作用的编排决定了肿瘤是被摧毁还是继续顺利增殖。我们将利用带有荧光标记的T细胞或树突状细胞的基因工程小鼠品系，我们将通过2P-IVM直观地跟踪它们在移植肿瘤中的相互作用。目的1将探索T细胞在肿瘤间质中的迁移，分析T细胞与细胞外基质的相互作用，并确定T细胞与肿瘤细胞相互作用的动力学。对肿瘤挑战作出反应的内源性T细胞将与过继转移的、体外产生的肿瘤相关抗原特异性T细胞进行比较。目的2将跟踪肿瘤内树突状细胞亚群的迁移行为，确定树突状细胞是否与肿瘤细胞相互作用，并检查它们是否与T细胞建立联系。这些实验将提高我们对导致肿瘤细胞破坏和/或诱导对肿瘤抗原的免疫耐受的事件的理解。这项研究的含义是，从长远来看，深入了解免疫细胞与肿瘤细胞的沟通将有助于产生改进的细胞免疫治疗方法。

DESCRIPTION (provided by applicant): Tumor cell-host cell interactions are critical parameters for the progression of neoplasms. Of particular importance is the relationship between neoplastic and immune cells that either mediate tumor destruction or promote tumor growth. How immune cells navigate within the tumor microenvironment, how they interact with neoplastic cells, as well as their overall contribution to the tumor micro-milieu is not well understood. The project's long-term goal is to define the cellular and molecular cues responsible for immune cell-tumor cell interaction in order to optimize immuno-therapeutic strategies. A major obstacle to such investigation has been our inability to date to image cellular interactions deep inside intact tissues at sufficiently high resolution in space and time. The objective of this proposal is to gain insight into the complex interplay between tumor and immune cells using real-time imaging at the single cell level in vivo. We propose to employ intravital two-photon microscopy (2P-IVM) to follow the activities of T cells and dendritic cells directly within their natural environment. Our hypothesis is that the migratory pattern of these immune cells within the tumor environment and the orchestration of their interactions with neoplastic cells decisively determine whether a tumor is destroyed or continues to proliferate unimpeded. We will make use of genetically engineered mouse strains having fluorescently tagged T cells or dendritic cells, whose interactions within implanted tumors we will track visually by 2P-IVM. Aim 1 will explore T cell migration within the tumor stroma, analyze T cell interactions with the extracellular matrix, and determine the kinetics of T cell-tumor cell interactions. Endogenous T cells that arise in response to tumor challenge will be compared to adoptively transferred, tumor associated antigen-specific T cells generated in vitro. Aim 2 will follow the migratory behavior of dendritic cell subsets within tumors, determine whether dendritic cells interact with tumor cells, and examine whether they establish contacts with T cells. These experiments will improve our understanding of the events leading to tumor cell destruction and/or the induction of immunologic tolerance to tumor antigens. The implication of this research is that an in-depth understanding of immune cell-tumor cell communication will, in the long-term, help to generate improved cellular immuno-therapeutic approaches.