PD-1 blockade enhances T cell activation by reorchestrating CD28 and CTLA4 ligand interactions

The network of interactions comprising CD28, CTLA4 and PD-1 and their ligands CD80, CD86, and PD-L1 have profound implications for T cell activation. Despite their critical role, factors that determine their receptor occupancy when all three ligands are present remains incompletely understood. Using a supported lipid bilayer to replicate the antigen-presenting cell membrane facilitated a quantitative analysis of CD80, CD86, and PD-L1 interactions. Our observations reveal that PD-1 blockade operates through a previously unrecognized competitive mechanism beyond conventional checkpoint inhibition: liberated PD-L1 molecules redistribute to form CD80-PD-L1 heterodimers that strengthen CD28-CD80 interactions and outcompete CD86 for CD28 binding. This preferential binding enhances T cell activation through superior CD80 co-stimulation. Furthermore, the redistribution by PD-1 blockade reduces CD80 homodimer availability, limiting CTLA4 binding to CD80 and enhancing CD28 signaling. These findings provide vital insights into the competitive principles that allow B7-CD28 family receptors to regulate T cell activation through network-level effects.