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

描述(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(T1D)，是由逃脱负选择的自身反应性T细胞引起的，并且可能与突变或外来蛋白发生交叉反应。同样，抗肿瘤免疫可能针对非突变自身蛋白的抗原，或将肿瘤相关突变整合到已识别表位的抗原。直到最近，对TCR/多肽/MHC复合体的结构研究一直局限于抗外源TCR。然而，在2005年，我们报道了一种与髓鞘碱性蛋白(MBP)和人类白细胞抗原DR2a结合的自身免疫性TCR的结构。随后，我们确定了一个人类黑色素瘤特异性TCR的三分子结构，该TCR在人类白细胞抗原DR1的背景下识别糖酵解酶磷酸丙糖异构酶(MuTPI)的自然发生的体细胞突变。值得注意的是，这些结构揭示了非传统的结合拓扑结构，对于TCR结合似乎不是最优的，这可能反映了与抗外源TCR相比，对自身反应施加的不同的选择压力。这些结果为系统研究自身免疫和癌症中自我识别的结构和生物物理原理开辟了道路，并利用这些知识来设计药物来特异性地调节(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。我们将扩展我们之前的工作，以确定来自MS和T1D患者的DR4限制性TCR是否也与具有改变的拓扑和次优相互作用的自体多肽/MHC结合。2.肿瘤抗原TCR识别的基础。为了比较非突变和突变肿瘤抗原的识别，并在自身免疫性疾病的自身抗原识别的背景下解释这一点，我们将检查非突变的HLA-DR4限制性黑色素瘤抗原酪氨酸酶(Ty)和gp100的TCR识别。3.作为MS潜在免疫治疗药物的高亲和力TCRs将被用于设计高亲和力MBP特异性TCRs，作为将免疫抑制细胞因子靶向自身抗原提呈部位的新型药物。4.改变多肽配体以增强抗黑色素瘤T细胞反应。基于Ty和gp100的TCR识别的结合和结构信息，这些共享的黑色素瘤表位将被修改以提高免疫原性。5.配体诱导的TCR二聚化可能是T细胞信号转导机制之一。如溶液中所示，黑色素瘤特异性TCRG4在结合muTPI/DR1时二聚化。为了验证(G4/muTPI/DR1)2复合体代表一个基本的T细胞信号单位的假设，我们将确定其结构，并通过诱变和T细胞激活的相关功能分析来评估其结构。综上所述，这些研究将为自身免疫性疾病和癌症中抗自身免疫的生物物理基础提供一个全面的视角。公共卫生相关声明：虽然在理解TCR识别微生物抗原的分子基础方面取得了显著进展，但对自身免疫性疾病和癌症中TCR识别自我或改变自我的原理知之甚少。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的分析来阐明这些原理，并利用这些知识来设计药物来特异性地抑制或增强T细胞对自身的反应。

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.