Enabling technologies to study how mechanics influence T cell function at the molecular and cellular levels

研究力学如何在分子和细胞水平上影响 T 细胞功能的技术

• 批准号: 9567971
• 负责人: Pui-Yan Victor Ma
• 金额: $ 4.45万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-20 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9567971
• 关键词: Adverse effects; Affect; Antigens; Antitumor Response; Architecture; Binding; Biochemical; Biological Process; Biotinylation; Cancer Biology; Cancer Control; Cell Proliferation; Cell Survival; Cell physiology; Cells; Characteristics; Chemicals; Clinical; Coculture Techniques; Complex; Coupled; Cues; Cytotoxic T-Lymphocytes; DNA; Data; Doctor of Philosophy; Enzyme-Linked Immunosorbent Assay; Event; Extracellular Matrix; Flow Cytometry; Genetic Transcription; Goals; Growth; Heterogeneity; Hydrogels; Immune; Immune Evasion; Immune checkpoint inhibitor; Immunosuppression; Immunosuppressive Agents; Immunotherapy; Integrins; Intercellular Junctions; Label; Ligation; Malignant Neoplasms; Maps; Measures; Mechanics; Mediating; Modeling; Molecular; Monitor; Names; Neoplasm Metastasis; Outcomes Research; PDCD1LG1 gene; Peptide/MHC Complex; Phase; Phosphorylation; Play; Postdoctoral Fellow; Production; Proteins; Proteomics; Receptor Activation; Receptor Signaling; Research; Research Personnel; Research Project Grants; Role; Series; Signal Transduction; Spectrum Analysis; Stimulus; Surface; T cell response; T-Cell Activation; T-Cell Receptor; T-Lymphocyte; Technology; Translating; Up-Regulation; Work; Wound Healing; antitumor effect; base; cancer cell; cancer immunotherapy; career; cell behavior; cell killing; cytokine; design; immunological synapse; innovation; interdisciplinary approach; mechanical force; mechanical properties; mimetics; molecular scale; nanosensors; neoplastic cell; oncology; overexpression; patient response; patient subsets; physical property; physical science; receptor function; response; scaffold; single molecule; success; systemic autoimmunity; tissue regeneration; training opportunity; transcriptome sequencing; tumor; tumor immunology; tumor microenvironment; tumor progression

Recent evidence has showed that the tumor microenvironment (TME) may form a sanctuary for immune suppression and evasion. Although Immunotherapies based on checkpoint inhibitors and chemical modulation of the TME have garnered success, many challenges remain, including the heterogeneity of patient response and the serious side effects resulting from systemic autoimmunity. These observations suggested that biochemical stimuli are not the only factor that suppresses T cell functions within the TME. One emerging concept in the field is that the physical properties of the TME such as extracellular matrix stiffness, composition, and architecture also contributes to cancer cell proliferation and survival. However, whether the mechanical properties of the TME specifically modulates T cell activity, and thus contributing to immune evasion, remains unclear. There are two overarching goals for this proposal. First, to better understand the role of mechanical forces in T cell receptor activation, and T cell functional responses. Second, to understand how the physical aspects of TME affect T cell/tumor interactions and T cell function. My PhD work is focused on developing enabling technologies to study mechanobiology at the molecular scale, with a particular focus on the roles of mechanical forces in T cell activation. My work has shown that i) the T cell receptor transmits pN forces to its antigen during initial recognition, and in immunological synapse and ii) T cells use mechanical energy to discriminate antigens during the earliest step of T cell activation. For my remaining F99 phase, I will focus on investigating whether mechanical forces are important for T cell function. To achieve this goal, I will use recently developed proximity labeling technologies to identify mechanosensitive proteins that mediate T cell signaling. This strategy will allow for using proteomic analysis to determine the mechanical interactome (mechanome) in T cells. Then, I will determine whether mechanical forces affect long term T cell biological functions using ELISA and flow cytometry coupled with RNA-SEQ. Overall, the results from this integrative -omics study will offer better understanding of how T cells use mechanical energy to potentiate their biological functions. For my postdoc studies (K00 phase), I aim to understand how the physical aspects of TME affect cytotoxic T cell function (cancer killing). The goal is to quantify how ECM mechanics alter T cell function. This data will support the hypothesis that TME contributes to metastasis by enhancing immune evasion through physical mechanisms. The significance of this work pertains to developing new strategies for promoting T cell anti-tumor response. I propose to work in a lab that employs 3D matrices that mimic the TME and to use these scaffolds for co-culture of T cells and tumor grafts. The work will better define the physical parameters of the matrix (e.g. the extent of hydrogel stiffness, physical cues composition, and architecture) that affect T cell/tumor interaction and T cell function. Collectively, the work from both F99 and K00 phases will provide new fundamental understanding on the physical basis of T cell functions at molecular level and cellular levels. The outcome of this research may offer new design principles for targeted cancer immunotherapies.

最近的证据表明，肿瘤微环境（TME）可能形成免疫抑制的避难所 和逃避。尽管基于检查点抑制剂和 TME 化学调节的免疫疗法已经 尽管取得了成功，但仍然存在许多挑战，包括患者反应的异质性和严重的一面 系统性自身免疫造成的影响。这些观察表明生化刺激并不是 唯一抑制 TME 内 T 细胞功能的因素。该领域的一个新兴概念是物理 TME 的特性（例如细胞外基质硬度、成分和结构）也会导致癌症 细胞增殖和存活。然而，TME 的机械性能是否特异性调节 T 细胞 活性，从而有助于免疫逃避，仍不清楚。该提案有两个总体目标。 首先，更好地了解机械力在 T 细胞受体激活和 T 细胞功能反应中的作用。 其次，了解 TME 的物理方面如何影响 T 细胞/肿瘤相互作用和 T 细胞功能。 我的博士工作重点是开发在分子水平上研究机械生物学的支持技术 规模，特别关注机械力在 T 细胞激活中的作用。我的工作表明 i) T 细胞受体在初始识别过程中以及在免疫突触和 ii) T 细胞中将 pN 力传递至其抗原 在 T 细胞激活的最早步骤中使用机械能来区分抗原。为了我剩下的F99 在此阶段，我将重点研究机械力对于 T 细胞功能是否重要。为了实现这一目标， 我将使用最近开发的邻近标记技术来识别介导 T 的机械敏感蛋白 细胞信号传导。该策略将允许使用蛋白质组分析来确定机械相互作用组 T 细胞中的（机械组）。然后，我将确定机械力是否影响长期 T 细胞生物学功能 使用 ELISA 和流式细胞术结合 RNA-SEQ。总体而言，这项综合组学研究的结果将 更好地了解 T 细胞如何利用机械能来增强其生物功能。 对于我的博士后研究（K00 阶段），我的目标是了解 TME 的物理方面如何影响细胞毒性 T 细胞功能（杀死癌症）。目标是量化 ECM 力学如何改变 T 细胞功能。该数据将支持 TME 通过物理机制增强免疫逃逸从而促进转移的假设。 这项工作的意义在于开发促进 T 细胞抗肿瘤反应的新策略。我 提议在一个采用模仿 TME 的 3D 矩阵的实验室中工作，并使用这些支架进行共培养 T 细胞和肿瘤移植物。这项工作将更好地定义基质的物理参数（例如水凝胶的范围） 硬度、物理线索组成和结构）影响 T 细胞/肿瘤相互作用和 T 细胞功能。 总的来说，F99 和 K00 阶段的工作将为物理提供新的基本理解。 T 细胞在分子水平和细胞水平上的功能基础。这项研究的结果可能会提供新的设计 靶向癌症免疫治疗的原则。