HT screens for the disruption of the T cell receptor CD3 interface

描述(由申请人提供)：我们的项目直接响应计划目标，应用高通量技术分离能够扰乱或改变独特型T细胞受体(TCR)与其专属信号伙伴CD3之间的通信的小分子。结构和生物物理研究告诉我们，TCR/CD3相互作用的亲和力很低，对微小的变化非常敏感，使其成为小分子开发的完美靶点。我们建立并优化了一种适用于高通量筛选的荧光偏振分析方法，该方法使用在单个半胱氨酸突变体上标记荧光基团的重组TCR分子。一项概念验证活动分离出6种化合物，这些化合物显示出TCR结合的特异性，并能够特异性地干扰T细胞的TCR/CD3通讯，而不影响T细胞信号转导。所有6个HIT都抑制MHC I类和MHC II类限制性T细胞的T细胞信号，因此可以归类为免疫抑制化合物。我们的目标是识别化学探针，使我们能够进一步了解T细胞激活的基本知识，并在同样的过程中发现新的免疫调节剂家族。事实上，由于目前治疗的毒性，在移植和自身免疫领域非常需要新的免疫抑制药物。由于我们已经开发的策略，我们将分离的一些化学片段也有可能表现出免疫刺激特性。这类药物，如一般免疫刺激剂，目前还不存在，将填补一个巨大的空白，以对抗癌症和慢性传染病的免疫抑制。我们的项目展示了我们研究所通过让化学、结构生物学、生物物理学、化学生物学、细胞免疫学和动物免疫学方面的专家并肩工作，朝着将基础科学转化为新疗法开发的明确目标而提供的整合能力。

DESCRIPTION (provided by applicant): Our project is directly responsive to the program objectives by applying high throughput technologies to isolate small molecules capable of disrupting or modifying the communication between the idiotypic T cell receptor (TCR) and CD3, its obligate signaling partner. Structural and biophysical studies have taught us that the TCR/CD3 interaction was of very low affinity and very sensitive to minor alterations, making it a perfect target for small molecules development. We have created and optimized a fluorescence polarization assay amenable to high throughput screen using recombinant TCR molecules labeled with fluorophore on single cysteine mutants. A proof of concept campaign isolated 6 compounds that showed specificity for TCR binding and were capable of disrupting specifically the TCR/CD3 communication of T cells without affecting T cells signaling. All 6 hits were inhibiting T cell signaling of both MHC class I- and MHC class II-restricted T cells and as such could be categorized as immunosuppressive compounds. Our goals are to identify chemical probes that will allow us to further our basic knowledge of T cell activation and to discover, in that same process, new families of immunomodulators. Indeed, new immunosuppressive drugs are much needed in the field of transplantation and autoimmunity, because of the toxicity of the current treatments. Because of the strategy that we have developed, it is also possible that some chemical fragments that we will isolate, will exhibit immunostimulatory properties. This class of drugs, e.g. general immunostimulants, does not exist yet and would fill a huge gap to combat the immunosuppression of cancer and chronic infectious diseases. Our project illustrates the capabilities of integration that our institute offers by having side-by-side specialists in chemistry, structural biology, biophysics, chemical biology, cellular immunology and animal immunology working together towards a clearly defined goal of translating basic science into the development of new therapeutics.