Characterizing and Targeting the Hypoxic T Cell Surfaceome to Promote Immune Function in Cancer

项目摘要 最近的努力集中在开发基于T细胞的免疫疗法来治疗癌症。但这些 治疗具有几个局限性，包括对实体瘤的最小功效。的一个特征 实体瘤微环境是低氧可用性或缺氧。一般认为肿瘤缺氧 抑制抗肿瘤免疫反应。这个项目的目标是提高我们对T 细胞对缺氧的反应，并揭示了增加缺氧T细胞抗肿瘤功能的新策略。 这些发现最终将为T细胞疗法的工程设计提供信息，以克服缺氧肿瘤 微环境调节抗肿瘤免疫功能的两种关键T细胞亚型是促炎性T细胞， 效应子（Teff）和抗炎性T调节（Treg）细胞。以前的研究考察了 缺氧对这些细胞类型的影响非常不一致。阐明缺氧对Teff和Treg功能的影响将有助于 扩展我们对基本T细胞生物学的理解，并可能发现增强抗肿瘤T细胞的新方法。 细胞缺氧时的活性。在这里，我们假设缺氧改变了T细胞表面蛋白（“表面组”）， T细胞以与净免疫抑制效应一致的方式发挥功能。为了验证这个假设，我们将 利用原代人T细胞、动物模型和患者样本的研究组合。我们将 利用我们建立的基于表面组学和噬菌体展示的重组抗体工程策略， 检查缺氧对疑似缺氧诱导抗原（HIA）的影响，并鉴定新的HIA。 我们将首先将这些技术应用于在常氧或低氧环境中培养的分离的原代人Teff和Treg细胞。 体外缺氧条件下。然后，我们将检查来自人源化的低氧肿瘤浸润性T细胞， 胰腺癌的小鼠模型以及患者胰腺肿瘤样品。这个表面分析 结合T细胞增殖，活化和迁移研究，将提供一个全面的图片， 缺氧诱导的T细胞表面蛋白和功能变化。此外，我们将设计新的抑制剂， 靶向疑似和新鉴定的HIA的抗体或双特异性构建体。这些工程化 将在低氧培养物和人源化胰腺细胞中测试蛋白质的Teff/Treg调节功能。 癌症小鼠模型，最终目标是鉴定促进缺氧Teff功能的策略， 免疫介导的肿瘤杀伤。从长远来看，基本的生物学见解和抗体工具， 这些拟议中的研究将为正在进行的使用T细胞治疗癌症的努力提供信息，并将为 未来的治疗方法，以提高缺氧肿瘤的免疫靶向。

Project Summary Recent efforts have focused on developing T cell-based immune therapies to treat cancer. However, these therapies have several limitations, including minimal efficacy against solid tumors. One characteristic of the solid tumor microenvironment is low oxygen availability, or hypoxia. It is generally thought that tumor hypoxia suppresses the anti-tumor immune response. The goal of this project is to improve our understanding of how T cells respond to hypoxia and reveal new strategies for increasing the anti-tumor function of hypoxic T cells. These findings will ultimately inform the engineering of T cell therapies to overcome the hypoxic tumor microenvironment. Two key T cell subtypes that modulate anti-tumor immune function are pro-inflammatory T effector (Teff) and anti-inflammatory T regulatory (Treg) cells. Previous studies examining the effects of hypoxia on these cell types are highly discordant. Clarifying the effects of hypoxia on Teff and Treg function will expand our understanding of basic T cell biology and potentially identify new means of enhancing anti-tumor T cell activity in hypoxia. Here, we hypothesize that hypoxia alters T cell surface proteins (the “surfaceome”) and T cell function in a manner consistent with a net immunosuppressive effect. To test this hypothesis, we will utilize a combination of studies with primary human T cells, animal models, and patient samples. We will employ our established surfaceomic and phage display-based recombinant antibody engineering strategies to examine the effect of hypoxia on suspected hypoxia-induced antigens (HIAs) as well as to identify novel HIAs. We will first apply these techniques to isolated primary human Teff and Treg cells cultured in normoxic or hypoxic conditions in vitro. We will then examine hypoxic tumor-infiltrating T cells derived from humanized mouse models of pancreatic cancer as well as patient pancreatic tumor samples. This surfaceomic profiling, combined with T cell proliferation, activation, and migration studies, will provide a comprehensive picture of hypoxia-induced T cell surface protein and functional changes. Furthermore, we will engineer novel inhibitory antibodies or bispecific constructs targeting both suspected and newly-identified HIAs. These engineered proteins will be tested for Teff/Treg modulatory function in hypoxic culture and in a humanized pancreatic cancer mouse model with the ultimate goal of identifying strategies to promote hypoxic Teff function and immune-mediated tumor killing. In the long term, the basic biological insights and antibody tools gained from the proposed studies will inform ongoing efforts to treat cancer using T cells and will lay the foundation for future therapeutics to boost immune targeting of hypoxic tumors.