Mechanisms of impaired T-cell mechanosensing of melanoma antigens

黑色素瘤抗原 T 细胞机械感应受损的机制

基本信息

批准号：
9899742
负责人：
MICHELLE KROGSGAARD
金额：
$ 94.63万
依托单位：
GEORGIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-04-01 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9899742
关键词：
Address Affinity Animal Model Animals Antigens Autoantigens Avidity Binding Biological Assay Blood Blood Circulation CD8-Positive T-Lymphocytes CD8B1 gene Cell membrane Cell physiology Cell surface Cells Cessation of life Chickens Clinical DNA Data Development Equilibrium Functional disorder Grant Human Image Immune Immunity Immunology Immunosuppression Immunotherapeutic agent Immunotherapy Impairment In Situ Kinetics Ligand Binding Ligands Link Major Histocompatibility Complex Measurement Mechanics Melanoma Cell Methods Microfluidics Molecular Molecular Analysis Mus Myeloid-derived suppressor cells Nature Outcome Ovalbumin Patient-Focused Outcomes Patients Peptide Receptor Peptides Receptor Cell Receptor Signaling Regulatory T-Lymphocyte Reporting Role Sampling Signal Transduction Skin Cancer Spleen System T cell therapy T-Cell Activation T-Cell Receptor T-Lymphocyte TCR Activation Techniques Testing Time Tissues Transforming Growth Factor beta Transgenic Mice Tumor Immunity Tumor-Infiltrating Lymphocytes United States United States National Center for Health Statistics Work advanced disease anti-melanoma immunity base checkpoint receptors design digital drug efficacy effector T cell exhaustion gp100 Antigen high throughput analysis immune checkpoint immunogenic improved improved outcome in vivo in vivo Model innovation mechanotransduction melanoma mouse model neoantigens novel strategies patient response physical science pre-clinical programmed cell death ligand 1 programmed cell death protein 1 protective effect response single molecule success tool tumor tumor microenvironment two-dimensional

项目摘要

Project Summary This project investigates how the tumor microenvironment (TME) impairs in situ interactions of T-cell surface molecules with counter-molecules on the melanoma cells to suppress anti-tumor immunity. Detailed mechanistic understanding will be obtained by an integrated approach that combines physical science (PS) based tools with two complementary pre-clinical mouse models of melanoma T cell immunity, which will be further tested using biospecimens from melanoma patients. The molecular focus is the T-cell receptor (TCR) that initiates the T-cell antigen recognition upon binding to peptide-major histocompatibility complex (pMHC), and the coreceptor CD8 that co-ligates with the pMHC. The first PS tool is quantifying TCR mechanosensing by single-molecule force probes through in situ kinetic analyses of molecular interactions with concurrent imaging of intracellular signals on a single cell. The second PS tool is DNA-based digital tension probes that report cell generated pulling forces on the TCR and CD8 via engaged pMHC. One animal model is a recognized standard that uses melanoma conjugated with a chicken ovalbumin antigen recognized by the OT-I TCR. The other animal model is a melanoma self-antigen gp100 in conjunction with JR209 humanized transgenic mice. By analyzing the mechanically regulated two-dimensional (2D) ligand binding of TCR and/or CD8 at the T-cell membrane, we observed that the TCR avidities for the pMHC of CD8 T cells infiltrating primary murine melanomas grown in vivo are significantly reduced relative to T cells within non-tumor associated tissues (spleen and blood). Such differential avidities were not detected by the conventional assay using pMHC tetramer, attesting to the power of our mechanics-based methods for analyzing TCR–pMHC interactions. We also found melanomas to substantially alter the force-dependent TCR–pMHC bond durability: in tumor-free animals, the TCR and pMHC formed a catch-slip bond whose lifetime first increased and then decreased with increasing force, which we have previously demonstrated to govern T cell signaling and effector function, whereas in melanoma-bearing animals, the TCR–pMHC bond lifetime only decreased with increasing force, i.e., behaved as a slip bond and were associated with reduced T cell effector functions. We hypothesize that deficient CD8 T cell immunity in melanoma results, at least in part, from impaired antigen recognition within the TME, as manifested by the altered TCR mechanosensing of pMHC. Three specific aims are proposed to test our hypothesis: 1) Determine the molecular interactions crucial to T cell antigen recognition that are impaired by the TME; 2) Define the functional consequences of suppressed T cell antigen recognition; and 3) Elucidate the mechanisms underlying the TME suppression of T cell antigen recognition. Completing these aims has the potential to identify new immunotherapeutic targets for the treatment of melanoma to improve the outcomes of patients with advanced disease.

项目摘要该项目研究了肿瘤微环境（TME）如何损害T细胞表面的原位相互作用分子在黑色素瘤细胞上的分子分子抑制抗肿瘤免疫。详细的机理将通过将基于物理科学（PS）工具与两种完整的黑色素瘤T细胞免疫的临床前小鼠模型，将进一步测试黑色素瘤患者的生物测量。分子焦点是启动T细胞的T细胞接收器（TCR）抗原识别与肽 - 莫霍尔组织相容性复合物（PMHC）和cocector cd8结合后识别抗原识别这与PMHC共同绑扎。第一个PS工具是通过单分子力量化TCR机械感应通过原位动力学分析分子相互作用的问题与细胞内信号的同时成像在一个单元格上。第二个PS工具是基于DNA的数字张力问题，该问题报告了细胞产生的拉力。在TCR和CD8上通过订婚PMHC。一种动物模型是使用黑色素瘤的公认标准用ot-i TCR识别的鸡肉卵蛋白抗原共轭。另一个动物模型是黑色素瘤自我抗原GP100与JR209人源性转基因小鼠结合使用。通过分析 TCR和/或CD8在T细胞膜上机械调节的二维（2D）配体结合，We 观察到CD8 T细胞PMHC的TCR狂热浸润在体内生长的原代鼠黑素瘤相对于非肿瘤相关组织（脾脏和血液）中T细胞的T细胞显着降低。这样的使用PMHC四聚体的常规测定未检测到差异狂热，证明了功率我们基于力学的方法用于分析TCR – PMHC相互作用的方法。我们还发现黑色素瘤实质上改变了力依赖性TCR – PMHC键耐用性：无肿瘤动物，TCR和PMHC 形成了一种流滑键，其生命周期首先增加，然后随着力量的增加而减少，我们拥有先前证明是控制T细胞信号传导和效应子功能，而在含黑色素瘤的动物中， TCR – PMHC键寿命仅随着力增加而降低，即表现为滑动键，为与降低T细胞效应子功能相关。我们假设黑色素瘤缺乏CD8 T细胞免疫结果至少部分来自TME内抗原识别受损的结果，如TCR改变所表现出的结果 PMHC的机械感应。提出了三个特定目的来检验我们的假设：1）确定分子 TME损害的T细胞抗原识别至关重要的相互作用； 2）定义功能抑制T细胞抗原识别的后果； 3）阐明TME的基础机制 T细胞抗原识别的抑制。完成这些目标有可能确定新的目标用于治疗黑色素瘤的免疫治疗靶标，以改善晚期患者的结局疾病。