Figure 4.

The Presence of Anti-PD-1 Antibody Enhances T Cell Activation and Function during SV-NYESO1 Therapy

(A) Treatment schema. Tumor-bearing mice were left untreated or were treated with SV with or without anti-PD-1. Mice were sacrificed on day 7, 14, or 21 to analyze the T cell immune response in spleen. (B and C) Percentage of CD44 and Ki-67 expression by CD4⁺ T cells (B) and CD8⁺ T cells (C) in naive mice as well as control and treated tumor-bearing mice using flow cytometry (n = 8 mice per group). Top graphs, representative flow cytometry plots from day 14. Bottom graphs, symbols summarizing data from two independent experiments. Statistical significance between groups treated with SV in presence or absence of anti-PD-1 was determined with the Mann-Whitney test. (D) Correlation of splenic CD4⁺ T cells or CD8⁺ T cells CD44 expression against tumor growth on day 14 by the Spearman-rank correlation test. (E) Interferon-γ (IFN-γ) enzyme-linked immunospot analysis of splenocytes harvested on day 14 from control and treated mice (n = 8 mice per group). (F) Cytotoxic activity of splenocytes harvested on day 14 from control and treated mice (n = 5 mice per group). Splenocytes were co-cultured with either CT26.Fluc.NYESO1 (left column) or CT26.Fluc.LacZ (right column) at various ratios for 2 days. Cytotoxic activity was assessed based on the viability of CT26 cells, which was determined by measuring the luciferase activity and is shown relative to naive splenocytes. Results are representatives from two independent experiments. (B, C, E, and F) Bars or symbols represent means ± SEM, and statistical significance was determined with the Mann-Whitney test (B, C, and E) or with the Kurskal-Wallis test followed by the Dunns’ test (F). n.s. > 0.05, *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001.