Fig 2: PD-1/PD-L1 binding activity related to untreated control.

Fig 3: PAI-1 promotes the production of soluble PD-L1, whereas blockade of PAI-1 suppresses the expression of PD-L1 expression both in vitro and in vivo. (A) Production of soluble PD-L1 in culture supernatants from MC38, B16F10, ES2, MOLM14, and K562 cells (n = 5/cell type) after 4 days of incubation with 100 nM rPAI-1 with or without 100 μM of the PAI-1 inhibitor TM5614. (B) Representative flow cytometry graphs of PD-L1 expression and MFI were obtained from 32Dp210 cells engrafted in the spleen (n = 6) and ES2 cells (n = 6) engrafted in subcutaneous tissue at 14 days post-inoculation. Bars are expressed as means ± SD of three independent experiments. **p < 0.01.

Fig 4: Effect of galectine-3 (Gal-3) on binding between programmed cell death 1 (PD-1)/programmed death-ligand 1 (PD-L1) and anti-PD-1/PD-L1 antibodies. (A) Surface plasmon resonance sensorgrams of the binding between Gal-3:PD-1 and antibodies; pembrolizumab, (B) nivolumab, and Gal-3:PD-L1 binding to antibodies; (C) atezolizumab and (D) durvalumab. Preincubated rhPD-1/PD-L1 and Gal-3 (100–6400 nM) were injected over immobilized antibodies.

Fig 5: Expression of PD-L1 and PD-L2 mRNA in paired CAFs in primary culture from tumor and tumor-adjacent normal samples obtained from patients with endometrial cancers: PD-L1 (A) and PD-L2 (B) mRNA expression (relative ratios to GAPDH) of NCAFs and TCAFs were carried out by qRT-PCR. HUF and tumor cell lines were used as controls. Melting curves for the respective mRNAs and GAPDH are presented as insets. A heatmap of PD-L1 and PD-L2 mRNA expression is presented as ratios of TCAFs/NCAFs pairs (C) of different patients with endometrial cancers. The five-rating icon sets (bars) with a three-color scale (light blue as a minimum, yellow as the midpoint, and violet as the maximum) are used.