Recognition of Self and Mutated Self by Autoimmune and Tumor-Specific TCRs

描述（由申请人提供）：自身免疫性疾病，如多发性硬化症（MS）和1型糖尿病（T1 D），是由逃避阴性选择的自身反应性T细胞引起的，并且可能与突变或外源蛋白交叉反应。类似地，抗肿瘤免疫可针对作为非突变自身蛋白或将肿瘤相关突变并入识别表位中的抗原。直到最近，TCR/肽/MHC复合物的结构研究一直限于抗外源TCR。然而，在2005年，我们报道了与来自髓鞘碱性蛋白（MBP）和HLA-DR 2a的自身肽结合的自身免疫TCR的结构。随后，我们确定了人类黑色素瘤特异性TCR的三分子结构，该TCR在HLA-DR 1背景下识别糖酵解酶磷酸丙糖异构酶（mutTPI）中天然存在的体细胞突变。值得注意的是，这些结构揭示了非常规的结合拓扑结构，其对于TCR结合似乎是次优的，并且与抗外源TCR相比，其可以反映对自身反应性施加的不同选择压力。这些结果开辟了一条道路，系统调查的结构和生物物理学原则，管理自身免疫和癌症的自我识别，并利用这些知识工程代理，以具体调节（抑制或增强）T细胞对自身抗原的反应。我们的目标是：1. TCR识别自身抗原的基础。我们将扩展我们以前的工作，以确定来自MS和T1 D患者的DR 4限制性TCR是否也结合具有改变的拓扑结构和次优相互作用的自身肽/MHC。2. TCR识别肿瘤抗原的基础。为了比较非突变与突变肿瘤抗原的识别，并在自身免疫性疾病中的自身抗原识别的背景下解释这一点，我们将检查非突变HLA-DR 4限制性黑素瘤抗原酪氨酸酶（Ty）和gplOO的TCR识别。3.高亲和力TCR作为MS的潜在免疫治疗剂。定向进化（酵母展示）将用于工程化高亲和力MBP特异性TCR作为用于将免疫抑制细胞因子靶向至自身抗原呈递位点的新型试剂。4.用于增强抗黑素瘤T细胞应答的改变的肽配体。基于Ty和gp 100的TCR识别的结合和结构信息，这些共有的黑素瘤表位将被修饰以提高免疫原性。5.配体诱导的TCR二聚化作为可能的T细胞信号传导机制。如溶液中所示，黑素瘤特异性TCR G4在结合mutTPI/DR 1后二聚化。为了验证（G4/mutTPI/DR 1）2复合物代表基本T细胞信号传导单位的假设，我们将确定其结构，并通过T细胞活化的诱变和相关功能分析来评估结构。总之，这些研究将提供一个全面的生物物理基础的抗自身免疫性疾病和癌症的看法。公共卫生相关性声明：虽然在理解TCR识别微生物抗原的分子基础方面取得了显着进展，但对自身免疫性疾病和癌症中TCR识别自身或改变的自身的原理知之甚少。我们的目标是通过对自身免疫和肿瘤特异性TCR的结构、生物物理和相关功能分析来阐明这些原理，并利用这些知识来设计特异性抑制或增强T细胞对自身的反应的试剂。

DESCRIPTION (provided by applicant): Autoimmune diseases, such as multiple sclerosis (MS) and type 1 diabetes (T1D), are caused by autoreactive T cells that have escaped negative selection, and which are potentially cross-reactive with mutant or foreign proteins. Similarly, anti-tumor immunity may be directed against antigens which are non-mutated self proteins, or which incorporate tumor-associated mutations into the recognized epitope. Until recently, structural studies of TCR/peptide/MHC complexes had been limited to anti- foreign TCRs. In 2005, however, we reported the structure of an autoimmune TCR bound to a self- peptide from myelin basic protein (MBP) and HLA-DR2a. Subsequently, we determined the trimolecular structure of a human melanoma-specific TCR recognizing a naturally-occurring somatic mutation in the glycolytic enzyme triosephosphate isomerase (mutTPI) in the context of HLA-DR1. Remarkably, these structures revealed unconventional binding topologies that appear suboptimal for TCR binding, and that may reflect the distinct selection pressures exerted on autoreactive as compared to anti-foreign TCRs. These results open the way to a systematic investigation of the structural and biophysical principles governing self-recognition in autoimmunity and cancer, and to using this knowledge to engineer agents to specifically modulate (suppress or enhance) T cell responses to self-antigens. Our objectives are: 1. Basis for TCR recognition of self-antigens. We will extend our previous work to determine whether DR4-restricted TCRs from MS and T1D patients also bind self-peptide/MHC with altered topologies and suboptimal interactions. 2. Basis for TCR recognition of tumor antigens. To compare recognition of non- mutated versus mutant tumor antigens, and interpret this in the context of self-antigen recognition in autoimmune diseases, we will examine TCR recognition of the non-mutated HLA-DR4-restricted melanoma antigens tyrosinase (Ty) and gp100. 3. High-affinity TCRs as potential immunotherapeutics for MS. Directed evolution (yeast display) will be used to engineer high-affinity MBP-specific TCRs as novel agents for targeting immunosuppressive cytokines to sites of autoantigen presentation. 4. Altered peptide ligands for enhancing anti-melanoma T cell responses. Based on binding and structural information on TCR recognition of Ty and gp100, these shared melanoma epitopes will be modified for heightened immunogenicity. 5. Ligand-induced TCR dimerization as a possible T cell signaling mechanism. As demonstrated in solution, the melanoma-specific TCR G4 dimerizes upon binding mutTPI/DR1. To test the hypothesis that the (G4/mutTPI/DR1)2 complex represents a basic T cell signaling unit, we will determine its structure, and assess the structure through mutagenesis and correlative functional analyses of T cell activation. Taken together, these studies will provide a comprehensive view of the biophysical basis for anti-self immunity in autoimmune diseases and cancer. Public Health Relevance Statement: While remarkable progress has been made in understanding the molecular basis for TCR recognition of microbial antigens, much less is known about the principles governing TCR recognition of self or altered self in autoimmune diseases and cancer. Our objective is to elucidate these principles through structural, biophysical and correlative functional analyses of autoimmune and tumor-specific TCRs, and to use this knowledge to engineer agents to specifically suppress or enhance T cell responses to self.