Immunotherapy with anti-programmed cell death protein-1 (PD-1)/programmed cell death ligand-1 (PD-L1) monoclonal antibodies has become routine in the treatment of many kinds of human cancers, such as lung cancer, intestinal cancer, and melanoma. The PD-1/PD-L1 pathway inhibits T cell activation in the micro-environment, making it an attractive target against cancer. Wild-type (WT) PD-1 ectodomain has been shown to have difficulty blocking PD-1/PD-L1 mixture formation due to its low affinity. The present work uses three-dimensional (3D) crystal complex structures to analyze the interaction by which PD-1 binds to PD-L1 or PD-L2. It also reports on a theoretical study of the binding mode between PD-1 and its clinical antibody Opdivo. Based on the theoretical binding analysis of PD-1 and its ligands (i.e., PD-L1 and PD-L2) or antibody (Opdivo), a small-content, epitope-oriented mammalian cell library was established for PD-1. After three rounds of cell sorting, the decoy PD-1 mutant 463, which presented a higher affinity than WT PD-1 to the PD-L1 (the affinity has increased by almost three orders of magnitude) was screened out. It exhibited an inhibitory effect against PD-1 to prevent it from forming mixtures with PD-L1, which was similar to the effect of the commercial anti-PD-L1 antibody atezolizumab (ATE). The median effective concentration (EC₅₀) value of the decoy mutant was 0.031 μg·mL⁻¹ in comparison with 0.063 μg·mL⁻¹ for ATE; both values were much lower than that of WT PD-1, at 2.571 μg·mL⁻¹. The 463 decoy mutant reversed the inhibitory function of PD-1 in T cell activation; furthermore, 10 mg·kg⁻¹ of 463 inhibited about 75% of tumor growth in vivo in a MC38 transgenic xenograft mice model, which was similar to the activity of ATE. More interestingly, an even lower dose of 463 (2 mg·kg⁻¹) showed a better effect than 10 mg·kg⁻¹ of WT PD-1. This work offers the decoy 463 with an improved curative effect, which holds potential to become a good option against PD-1/PD-L1-related cancers.

抗程序性细胞死亡蛋白-1（PD-1）/程序性细胞死亡配体-1（PD-L1）单克隆抗体免疫治疗已成为多种人类癌症治疗的常规治疗，如肺癌、肠癌、和黑色素瘤。PD-1/PD-L1 通路抑制微环境中的 T 细胞活化，使其成为抗癌的有吸引力的靶点。已显示野生型 (WT) PD-1 胞外域由于其低亲和力而难以阻断 PD-1/PD-L1 混合物的形成。目前的工作使用三维 (3D) 晶体复合结构来分析 PD-1 与 PD-L1 或 PD-L2 结合的相互作用。它还报告了 PD-1 与其临床抗体 Opdivo 之间结合模式的理论研究。基于PD-1及其配体（即PD-L1和PD-L2）或抗体（Opdivo）的理论结合分析，为 PD-1 建立了一个小内容、以表位为导向的哺乳动物细胞文库。经过三轮细胞分选，筛选出诱饵PD-1突变体463，其对PD-L1表现出比WT PD-1更高的亲和力（亲和力提高了近三个数量级）。它表现出对 PD-1 的抑制作用，防止其与 PD-L1 形成混合物，这类似于商业抗 PD-L1 抗体 atezolizumab (ATE) 的作用。中位有效浓度（EC 它表现出对 PD-1 的抑制作用，防止其与 PD-L1 形成混合物，这类似于商业抗 PD-L1 抗体 atezolizumab (ATE) 的作用。中位有效浓度（EC 它表现出对 PD-1 的抑制作用，防止其与 PD-L1 形成混合物，这类似于商业抗 PD-L1 抗体 atezolizumab (ATE) 的作用。中位有效浓度（EC₅₀ ) 诱饵突变体的值为 0.031 μg·mL ^-1，而 ATE 的值为0.063 μg·mL ^-1；这两个值都远低于 WT PD-1 的值，为 2.571 μg·mL ^-1。463诱饵突变体逆转了PD-1在T细胞活化中的抑制作用；此外，在 MC38 转基因异种移植小鼠模型中，463 中的10 mg·kg ^-1抑制了约 75% 的体内肿瘤生长，这与 ATE 的活性相似。更有趣的是，甚至更低剂量的 463 (2 mg·kg ^-1 ) 显示出比 10 mg·kg ^-1更好的效果WT PD-1。这项工作为诱饵 463 提供了更好的疗效，有可能成为对抗 PD-1/PD-L1 相关癌症的良好选择。