使用Lewis x系列寡糖改善嵌合抗原受体T细胞在实体瘤组织中的浸润和 粘附的研究

Lewis x （Le x）系列寡糖在众多生命过程中发挥重要的作用，是重要的肿瘤生物标志物。研究发现，炎症和肿瘤部位附近的内皮细胞表面会上调表达选择素E，而选择素E可与Le x系列寡糖结合，介导淋巴细胞在炎症、肿瘤的粘附和浸润。本项目拟使用Le x系列寡糖对嵌合抗原受体T（CAR-T）细胞进行修饰，增强其向实体瘤组织的迁移和浸润，以提高CAR-T细胞对实体瘤的治疗效果，减少不良反应的发生几率。我们在前期Le x单体及其唾液酸化单体合成的基础上，通过高效的酶法模块化组装策略，完成Le x单体到五聚体及其两种不同唾液酸化形式的Le x系列寡糖化合物库的构建；继而运用糖芯片技术，系统地研究Le x系列寡糖与多种选择素和C型凝集素结合的构效关系；依据得到的构效关系信息，选择合适的寡糖对CAR-T细胞进行表面修饰，观察修饰对CAR-T细胞的迁移、浸润和治疗效果的影响。

Lewis x (Le x) series oligosaccharides are associated with various physiological processes. Le x series oligosaccharides are over-expressed in many malignant tissues, which make them biomarkers for tumor diagnosis and treatment. Studies have revealed that E-selectin, one ligand of Le x series oligosaccharides, is markedly up-regulated in endothelial cells near inflamed and some tumor tissues, promoting the adhesion and infiltration of leukocytes into these affected tissues. This project intends to modify the cell surface of the chimeric antigenic receptor (CAR) T cells with Le x series oligosaccharides, aiming to direct the trafficking and infiltration of CAR-T cells to the solid tumor sites to enhance the therapeutic efficacy and reduce the adverse effects of CAR-T cell therapy in solid tumor treatment..In our previous work, we have successfully synthesized monomer Le x and sialyl Le x. On the basis of the well-established glycosyltransferase-based enzymatic modular assembly (EMA) strategy, we will apply several rational designed modules to construct a library of Le x series oligosaccharides, comprising of monomeric to pentameric Le x, with or without two different terminal sialic acid types (Neu5Ac or Neu5Gc). Subsequently, we will use glycan-array techniques to study the binding affinity of these oligosaccharides to E, P, L-selectin and two C-type lectins (DC-SIGN and L-SIGN). Based on the information provided by glycan-array, suitable Le x series oligosaccharide will be conjugated to phospholipids for CAR-T cell surface modification. The surface glycan-modified CAR-T cells will be tested in vivo to study the migration and tumor infiltration properties, as well as their therapeutic outcome.