Fig. 2

The pAC65 peptide restore the activation of Jurkat-ECs cell line and primary human T cells. A Dose-dependent reactivation of Jurkat-ECs with hPD-L1-blocking agents: pAC65 (left panel) and atezolizumab (right panel) in the ICB assay. Graphs show fold luminescence induction relative to either untreated (for atezolizumab) or DMSO-treated (for pAC65) cells. Data points represent mean ± SD values from 4–6 independent experiments. B The long-term (48 h) cytotoxicity of the peptide pAC65 towards Jurkat-ECs. The graph shows Jurkat-ECs survival relative to DMSO-treated control cells. Triton X-100-treated cells and cells without the addition of a Redox Dye were used as baseline controls. C The blockade of the human PD-L1 (hPD-L1). D The blockade of mouse PD-L1 (mPD-L1) in the ICB assay. In the assay, three antibodies (atezo. – atezolizumab, ave. – avelumab, durva. – durvalumab) and the peptide pAC65 were used. Graphs show fold luminescence induction relative to either untreated (for antibodies) or DMSO-treated (for pAC65) cells

The expression of PD-1 on either CD4⁺ (E) or CD8⁺ (F) cells was determined by flow cytometry. PBMCs from healthy donors were exposed to either CHO/TCRAct/PD-L1, CHO/TCRAct, or CHO cells for two days in the presence of durvalumab (durva), atezolizumab (atezo), avelumab (avelu), or increasing concentrations of the peptide pAC65 (the concentrations: 2.5 nM, 25 nM, or 250 nM). ø indicates untreated cells. The graphs show fractions of positive cells and represent cumulative data from 3–4 donors. Posthoc test *, p < 0.05, **, p < 0.01, ***, p < 0.001, or DMSO-treated cells: ###, p < 0.001