Contribution of PAG to Immune Synapse Organization and PD-1 Function

PAG 对免疫突触组织和 PD-1 功能的贡献

• 批准号: 10538164
• 负责人: Emily Kathryn Moore
• 金额: $ 5.18万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-15 至 2026-07-14
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10538164
• 关键词: Actins; Adhesions; Adoptive Cell Transfers; Adverse effects; Anatomy; Antibodies; Antibody Therapy; Antigen-Presenting Cells; Antitumor Response; Architecture; Autoimmune Diseases; Automobile Driving; Binding; Biological Assay; C-terminal; CD28 gene; Cause of Death; Cell Line; Cell membrane; Cell physiology; Cells; Clinical; Complex; Confocal Microscopy; Cytoskeleton; Enzyme-Linked Immunosorbent Assay; Enzymes; Event; Fellowship; Flow Cytometry; Fluorescence Resonance Energy Transfer; Fostering; Future; Glycosphingolipids; Goals; Human; Imaging Techniques; Immune; Immune Targeting; Immune checkpoint inhibitor; Immune response; Immunology; Immunotherapy; Integral Membrane Protein; Knock-out; Ligand Binding; Ligation; Link; Lipids; MAP Kinase Gene; Malignant Neoplasms; Measures; Mediating; Medicine; Membrane; Membrane Microdomains; Mentorship; Methods; Mind; Modeling; Mus; Mutate; Palmitic Acylation Site; Pathway interactions; Patients; Pattern; Peripheral; Pharmacology; Phosphoproteins; Phosphorylation; Physicians; Process; Production; Proteins; Regimen; Research; Research Personnel; Resources; Role; Shapes; Signal Pathway; Signal Transduction; Signaling Protein; Structure; Synapses; T-Cell Activation; T-Cell Receptor; T-Lymphocyte; Technology; Testing; Therapeutic; Therapeutic antibodies; Total Internal Reflection Fluorescent; Tumor-infiltrating immune cells; Tyrosine Phosphorylation Site; Work; anti-PD-1; anti-PD1 antibodies; antibody immunotherapy; cancer cell; cancer immunotherapy; cancer therapy; career; cytokine; cytotoxicity; immune function; immune-related adverse events; immunological synapse; improved; in vivo; innovation; interest; mortality; new therapeutic target; novel; novel therapeutics; polymerization; prevent; programmed cell death protein 1; protein protein interaction; recruit; response; skill acquisition; skills; synaptic function; synaptogenesis; targeted cancer therapy; targeted treatment; therapeutic target; tumor

PROJECT SUMMARY/ABSTRACT Cancer remains the second leading cause of death in the US. Immunotherapy seeks to bolster immune cells’ ability to target malignant cells and has brought immense improvements in the field. One important inhibitory protein in T cells, Programmed Cell Death Protein 1 (PD-1), has become an invaluable target for cancer immunotherapy. While anti-PD-1 antibody therapy is extremely successful in some patients, in many others, it fails to help or causes complications, including cancer hyper-progression and immune-related adverse events. Study of the inhibitory transmembrane protein Phosphoprotein Associated with Glycosphingolipid Rich Microdomains 1 (PAG), a downstream target of PD-1 signaling, will help us better understand the PD-1 pathway, and offer another, perhaps more nuanced, target to potentially improve response rates and/or avoid immune- related adverse events. As a link between lipid-rich/signaling-protein-rich membrane regions and the actin cytoskeleton, PAG is an exciting and novel target for manipulating immune function. Prior therapeutic methods of immune manipulation all disrupt ligand binding or enzyme function. In contrast, innovative use of an anti-PAG antibody to simply disrupt appropriate PAG localization within the synapse could disturb immune synapse architecture. Synapse organization is tightly regulated to prevent inappropriate immune responses, but the precise interaction between cytoskeletal dynamics and synaptic organization is not fully understood. Investigating the role of PAG in this process could provide added clarity. To test the hypothesis that PAG works with actin to control T cell synapse organization and facilitate T cell receptor (TCR) and PD-1 signaling, PAG will be studied in a human T cell line and a murine PAG-knockout model. Methods will include confocal and TIRF microscopy, FRET and proximity ligation assay, flow cytometry, ELISA, adhesion and cytotoxicity assays, and murine tumor models. To understand the contribution of the actin binding domain of PAG on its localization, protein-protein interactions, and T cell activation and effector functions, the primary hypothesis will be examined through the following specific aims: Aim 1. Define the role of the PAG-actin link in driving T cell synapse architecture and stability. Aim 2. Determine the impact of PAG-actin interactions on T cell functions downstream of TCR and PD-1 signaling. Aim 1 will demonstrate the role of PAG in T cells synapse anatomy, and the physical impacts of a PAG-targeting therapy. Aim 2 will illuminate which PD-1 downstream targets are dependent on the PAG-actin link, and provide evidence for whether PAG and PD-1 could serve as good co-targets in cancer therapy regimens. Ultimately, this study will illuminate crucial control mechanisms associated with T cell synapse organization, opening more avenues of targeting the immune synapse. Furthermore, the mentorship and resources in Dr. Adam Mor’s lab and the Columbia MSTP, will foster invaluable technical and professional skill development for a career in medicine and immunology research.

项目摘要/摘要 癌症仍然是美国第二大死因。免疫疗法寻求增强免疫细胞 针对恶性细胞的能力，并在该领域带来了巨大的改进。一个重要的抑制因素 T细胞中的蛋白质，程序性细胞死亡蛋白1(PD-1)，已经成为癌症的宝贵靶点 免疫疗法。虽然抗PD-1抗体疗法在一些患者中非常成功，但在许多其他患者中，它 没有帮助或导致并发症，包括癌症过度进展和免疫相关的不良事件。 富含鞘糖脂的抑制性跨膜蛋白磷脂的研究 微域1(PAG)是PD-1信号的下游靶点，有助于我们更好地了解PD-1通路。 并提供另一个可能更细微的目标，以潜在地提高应答率和/或避免免疫- 相关不良事件。作为富脂/富含信号蛋白的膜区和肌动蛋白之间的纽带 细胞骨架，PAG是一个令人兴奋的新的免疫功能调控靶点。既往治疗方法 所有的免疫操作都会扰乱配体结合或酶功能。相比之下，创新地使用抗PAG 简单地破坏突触内适当的PAG定位的抗体可以干扰免疫突触 建筑。突触组织受到严格监管，以防止不适当的免疫反应，但 细胞骨架动力学和突触组织之间的精确相互作用尚不完全清楚。 调查PAG在这一过程中的作用可以提供更多的明确性。来检验PAG的假设 与肌动蛋白一起控制T细胞突触组织并促进T细胞受体(TCR)和PD-1 信号，PAG将在人类T细胞系和小鼠PAG基因敲除模型中进行研究。方法将包括 共聚焦显微镜和TIRF显微镜、FRET和邻近连接试验、流式细胞术、ELISA、黏附和 细胞毒性试验和小鼠肿瘤模型。为了了解肌动蛋白结合域的作用 PAG对其定位、蛋白质相互作用、T细胞激活和效应功能的初步研究 假说将通过以下具体目标进行检验：目标1.确定PAG-肌动蛋白连接的作用 在驱动T细胞突触的构筑和稳定性方面。目的2.确定PAG-肌动蛋白相互作用的影响 On T细胞功能位于TCR和PD-1信号的下游。目标1将展示PAG在T细胞中的作用 突触解剖，以及PAG靶向治疗的物理影响。目标2将阐明PD-1 下游靶点依赖于PAG-肌动蛋白连接，并为PAG和PD-1 在癌症治疗方案中可以作为良好的联合靶点。最终，这项研究将阐明关键的控制 与T细胞突触组织相关的机制，开辟了更多靶向免疫的途径 Synapse。此外，亚当·摩尔博士实验室和哥伦比亚大学MSTP的指导和资源将促进 在医学和免疫学研究的职业生涯中，宝贵的技术和专业技能发展。