Article Figures & Data
Figures
- Figure 1.
Immune profile of tumor-infiltrating CD8+ T cells (TIL) from NSCLC patients. The expression of the inhibitory receptors PD-1, Tim-3, CTLA-4, LAG-3, and BTLA was determined by flow cytometry on tumor-infiltrating CD8+ T cells from tumor digestions. A, the gating strategy is shown for one representative donor. B, indicated cell subsets were analyzed and heat mapped based on the percentage of expression with the use of an Excel conditional formatting program. C, distribution of naïve and memory T-cell subsets, characterized by CCR7 and CD45RA, in CD8+ T cells from lung cancer specimens (TIL) or PBMCs from healthy donors (HD). D, expression of inhibitory receptors on tumor-infiltrating CD8+ T cells.
- Figure 2.
Expression of PD-1 and Tim-3 increases with tumor stage. Expression of inhibitory receptors on CD8+ tumor-infiltrating T cells was correlated to the TNM stage. Each dot represents one patient sample. The P values were calculated using the one-way ANOVA test.
- Figure 3.
Cumulative expression of inhibitory receptors increases with tumor progression. The cumulative expression of the inhibitory receptors PD-1, Tim-3, CTLA-4, LAG-3, and BTLA, as represented by the inhibitory receptor (iR) score, was correlated to the nodal status and the TNM stage. Each dot represents one patient sample. The P values were calculated using the Wilcoxon rank sum test.
- Figure 4.
Coexpression of inhibitory receptors by TILs from NSCLC patients. Tumor-infiltrating CD8+ T cells with expression of the indicated receptor are analyzed for the expression of the additional inhibitory receptors. The results are shown as a heat map (A) or as a radar plot (B). The latter indicates the mean expression (black line) and SD (gray area) of the four other receptors.
- Figure 5.
Cumulative expression of inhibitory receptors defines T-cell dysfunction. Whole tumor digests were stimulated for 24 hours with agonistic anti-CD3 and anti-CD28 or left untreated (control). The expression of CD25 and Granzyme B in CD8+ T cells as well as IL2, IFNγ, and TNFα secretion was determined by flow cytometry (A) or ELISA (B), respectively. The increase of these parameters was correlated to the cumulative expression of the inhibitory receptors PD-1, Tim-3, CTLA-4, LAG-3, and BTLA as reflected by the inhibitory receptor (iR) score (C + D). The P values were calculated using the one-way ANOVA test.
- Figure 6.
Blockade of PD-1 restores T-cell function depending on the level of PD-1 expression. Whole tumor digests were stimulated for 24 hours with agonistic anti-CD3/CD28 in the presence or absence of a blocking anti–PD-1 antibody. A, IL2, TNFα, and IFNγ secretion was determined by ELISA and normalized to 1 × 106 CD3+ T cells. B, the baseline distribution of CD8+ T cells between PD-1hi and PD-1int subsets is shown for a patient where T-cell function can be rescued by addition of anti–PD-1 (BS-268) and a patient with no response to PD-1 blockade (BS-199). C, the increase of cytokine secretion induced by anti–PD-1 [(% expression Ab treated) – (% expression untreated)] was correlated with the percentage of PD-1hi CD8+ T cells from the PD-1–positive population. The P values were calculated using the one-way ANOVA test.
- Figure 7.
PD-1hi and PD-1int T-cell subsets coexpress different levels of other inhibitory receptors. The percentage of PD-1hi or PD-1int CD8+ T cells expressing additional immune checkpoints is shown. Each dot represents one patient sample. The P values were calculated using the Wilcoxon rank sum test.
Additional Files
Supplementary Data
- Supplementary Table 1, Figures 1 - 2 - Supplementary Table 1. Clinical data of NSCLC patients. Supplementary Figure 1. Calculation of the iR score. Supp. Figure 2. Activation of PBMC from healthy donors by polyclonal stimulation.
Volume 3, Issue 12
