The Induction of Macrophage Endoplasmic Reticulum Stress by Irradiated-Tumor Derived Extracellular Vesicles Supports the Adoption of a Pro-Tumor Phenotype

项目摘要 放射治疗启动交替刺激(M2)巨噬细胞向肿瘤的募集 微环境。这些细胞表现出“亲肿瘤表型”，刺激血管生成，抑制抗肿瘤 肿瘤免疫，增强肿瘤细胞的抗辐射能力。这限制了放射治疗的整体疗效 实体瘤，如非小细胞肺癌。目前放射免疫学领域的研究主要集中在 要么抑制这些细胞的招募，要么在它们到达 肿瘤微环境。然而，受辐射的癌细胞如何操纵巨噬细胞的确切方式在很大程度上是 未知。我们发现，照射后的肺癌细胞(IR-EVS)所形成的胞外小泡 在SRA/CD204中诱导内质网应激并诱导巨噬细胞产生促肿瘤细胞因子 依赖的态度。这个项目的总体目标是定义IR-EVS的分子机制 诱导巨噬细胞亲肿瘤细胞因子表达。首先，我们将测试IR-EV诱导的功能影响 CHOP-/-C57BL/6小鼠巨噬细胞内质网应激对巨噬细胞表型的影响及可溶性内质网抑制剂 压力。然后，我们将研究巨噬细胞SRA/CD204在巨噬细胞对IR-EVS反应中的作用。 我们将使用SRA/CD204-/-C57BL/6小鼠以及开发的一种新的SRA/CD204封闭抗体来实现这一点 在我们的实验室里。最后，我们将比较复发和非复发非小细胞的循环IR-EVS 以判断肺癌患者是否能预测放射治疗的结果。来自经常性和非经常性的电动汽车 复发的NSCLC患者在放疗前、中和之后被隔离，将被评估他们刺激 促肿瘤巨噬细胞的体外表型。然后，将使用质谱学来识别潜在的 SRA/CD204结合蛋白在患者和培养细胞的IR-EV表面表达上调。潜力 将肺癌患者分为复发或无病生存组 使用商业上可用的浓缩血清外切体的酶联免疫吸附试验进行评估。

Project Summary Radiotherapy initiates the recruitment of alternatively stimulated (M2) macrophages to the tumor microenvironment. These cells exhibit a “pro-tumor phenotype,” stimulating angiogenesis, suppressing anti- tumor immunity, and enhancing tumor cell radioresistence. This limits the overall efficacy of radiotherapy for solid tumors such as non-small cell lung cancer. Current research in the field of radioimmunology is focused on manipulations that either inhibit the recruitment of these cells or that alter their behavior once they reach the tumor microenvironment. However, exactly how irradiated cancer cells manipulate macrophages is largely unknown. We have discovered that extracellular vesicles elaborated by irradiated lung cancer cells (IR-EVs) induce ER-stress and elicit the production of “pro-tumor” cytokines from macrophages in an SRA/CD204 dependent manner. The overall objective of this project is to define the molecular mechanisms by which IR-EVs induce macrophage pro-tumor cytokine expression. First, we will test the functional impact of IR-EV induced macrophage ER stress on macrophage phenotype using CHOP-/- C57BL/6 mice and soluble inhibitors of ER stress. Then, we will investigate the role of macrophage SRA/CD204 in the response of macrophages to IR-EVs. We will do this using SRA/CD204-/- C57BL/6 mice as well as a novel SRA/CD204 blocking antibody developed in our laboratory. Finally, we will compare the circulating IR-EVs of recurrent and non-recurrent non-small cell lung cancer patients to determine if they can predict the outcome of radiotherapy. EVs from recurrent and non- recurrent NSCLC patients isolated before, during, and after RT will be assessed for their ability to stimulate a pro-tumor macrophage phenotype in vitro. Then, mass spectrometry will be used to identify a potential SRA/CD204 binding protein upregulated on the surface of IR-EVs from patients and cultured cells. The potential of ligand bearing exosomes to segregate lung cancer patients into relapse or disease-free survival groups will be evaluated using a commercially available ELISA on concentrated serum exosomes.