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。特别重要的是肿瘤细胞和免疫细胞之间的关系，它们介导肿瘤破坏或促进肿瘤生长。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尚不清楚。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，以优化免疫治疗策略。这类研究的一个主要障碍是，迄今为止，我们无法在空间和时间上以足够高的分辨率对完整组织深处的细胞相互作用进行成像。本提案的目的是在体内单细胞水平上使用实时成像来深入了解肿瘤和免疫细胞之间复杂的相互作用。我们建议使用活体双光子显微镜（2P-IVM）来直接跟踪T细胞和树突状细胞在自然环境中的活动。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协调，决定性地决定了肿瘤是被破坏还是继续不受阻碍地增殖。我们将利用具有荧光标记的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视觉跟踪其在植入肿瘤内的相互作用。目的1将探索T细胞在肿瘤基质内的迁移，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。内源性T细胞的产生是为了响应肿瘤的挑战，将与过继转移，肿瘤相关的抗原特异性T细胞在体外产生进行比较。Aim 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细胞建立接触。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。这项研究的意义在于，从长远来看，对免疫细胞-肿瘤细胞通讯的深入了解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.