The Development of Chemical Tool Technology To Modulate Immune Molecular Recognition

免疫系统对肿瘤的识别和消除是现代肿瘤免疫治疗的基础，也是2018年诺贝尔医学奖的主题。我们化学免疫学研究计划的长期目标是设计新的工具来阐明抗体（Ab）和免疫细胞受体（ICR）对肿瘤分子识别的基本机制。这借鉴了我在开发称为“二聚体”(D)的合成工具方面的丰富经验，二聚体通过与肿瘤配体(L)相互作用在肿瘤表面形成可逆三元复合物（Ab:D:L）来调节肿瘤的免疫识别。二聚体通过同时结合肿瘤细胞表面的蛋白质配体L和抗体形成三元复合物。然后结合抗体通过ICR激活免疫细胞，通过吞噬（吞噬）或细胞毒性（释放细胞毒性颗粒）破坏肿瘤细胞。虽然控制针对靶细胞的免疫激活强度的因素尚不清楚，但很明显，免疫反应取决于结合的三元复合物的数量。形成具有高亲和力的Ab:D:L三元配合物以占据靶细胞上所有可能的配体是一个基本的分子识别问题。二聚体设计的一个主要限制是Ab和L的体内浓度很低（nM到pM），因此很难设计D来支持所需的三元配合物而不是无活性的二元配合物Ab:D和D:L。体内循环中D的快速清除进一步限制了三元络合物的形成。我们的目标是设计、合成和表征二聚体，通过在体内最大限度地提高靶细胞表面三元或二元复合物的稳定性和形成，有效增强免疫系统对靶细胞的分子识别。这将通过两个短期目标来实现：(1)我们将设计与抗体不可逆连接的“共价二聚体”（CD）和“动态捕获”三元复合物Ab-CD:L（其中虚线表示共价连接）。如果低浓度的Ab削弱了目标表面上最大的三元配合物形成，我们将开发额外的双反应性CDs，除了与Ab形成共价键外，还与L形成不可逆的三元配合物Ab- cd -L。(2)我们将开发荧光极化动力学分析，以发现和表征“结合诱导”抗体标记反应。拟议研究项目产生的新知识和技术将提供必要的多学科培训，同时为免疫系统靶向外来病原体和细胞的新策略提供技术平台。

The recognition and elimination of tumors by the immune system is the foundation of modern tumor immunotherapy, the topic of the 2018 Nobel Prize in medicine. The longterm goal of our chemical immunology research program is to design new tools to elucidate the fundamental mechanisms underlying molecular recognition of tumors by antibodies (Ab) and immune cell receptors (ICR). This draws on my extensive experience in the development of synthetic tools called "dimerizers",(D), that modulate the immune recognition of tumors by forming reversible ternary complexes, (Ab:D:L) on the tumor surface through interactions with tumor ligands (L). Dimerizers form ternary complexes by simultaneously binding the protein ligand, L, on the tumor cell surface and an antibody. The bound antibody then activates immune cells through their ICR to destroy the tumor cell through phagocytosis (engulfment) or cytotoxicity (release of cytotoxic granules). Although the factors that control the strength of immune activation against a target cell are not well understood, it is clear that the immune response depends on the number of bound ternary complexes. Forming Ab:D:L ternary complexes with high affinity to occupy all possible ligands on a target cell is a fundamental molecular recognition problem. A major constraint in dimerizer design is that the in vivo concentrations of Ab, and L are so low (nM to pM) that it is difficult to design D to favor the desired ternary complexes over inactive binary complexes Ab:D and D:L. Ternary complex formation is further limited by the fast clearance of D from the circulation in vivo. We aim to design, synthesize and characterize dimerizers that can effectively enhance molecular recognition of target cells by the immune system, by maximizing the stability and formation of ternary vs. binary complexes on the target cell surface in vivo. This will be accomplished through two short term objectives: (1) We will design "covalent dimerizers" (CD) that irreversibly link to antibodies and "kinetically trap" ternary complexes Ab-CD:L (where the dash denotes a covalent linkage). In the event low concentrations of Ab attenuate maximal ternary complex formation on the target surface, we will develop additional bis-reactive CDs that form a covalent linkage to L in addition to Ab to form irreversible ternary complexes Ab-CD-L. (2) We will develop fluorescence polarization kinetic assays to discover and characterize "binding induced" antibody labelling reactions. The new knowledge and technology generated from the proposed research program will provide essential multidisciplinary training while providing technological platforms for new strategies to target foreign pathogens and cells by the immune system.