High-Affinity Decoy PD-1 Mutant Screened from an Epitope-Specific Cell Library

摘要

Immunotherapy with anti-programmed cell death protein-1(PD-1)/programmed cell death ligand-1(PDL1)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.