Fig. 2
The PD-L1 N35A variant perturbs the ability of anti-PD-L1 ICBs to block PD-L1/PD-1 interactions, whereas the lack of glycosylation at all four PD-L1 glycosylation sites enhances blocking. A. Schematic representation of the experimental procedure determining the efficacy of anti-PD-L1 ICBs in blocking the interactions between PD-L1WT/different PD-L1 glycosylation variants and PD-1. Here, PD-L1-expressing cells were incubated with increasing concentrations of anti-PD-L1 antibodies (Atezolizumab, Avelumab or Durvalumab). Then, IcAR-PD-1 cells were added to assess PD-L1/PD-1 binding inhibition, measured by the levels of mIL-2 (determined by ELISA). B. The capacity of anti-PD-L1 antibodies in blocking the interactions of PD-L1 (WT and glycosylation variants) expressed by MDA-MB231 cells with PD-1. Graphs demonstrate the normalized blocking capacity of Atezolizumab (B1), Avelumab (B2), and Durvalumab (B3) on each cell type. Antibody concentrations ranged from 1.5 µg/mL to 20 µg/mL. Data points represent mean ± SEM from triplicate biological repeats. C. The capacity of anti-PD-L1 antibodies in blocking the interactions of PD-L1 (WT and glycosylation variants) expressed by MCF7 cells with PD-1. As in panel B, the graphs display the normalized blocking capacity of Atezolizumab (C1), Avelumab (C2), and Durvalumab (C3). The same antibody concentration range was used