(A) Representative intracellular cytokine staining dot plots of IFNγ (top) and TNFα (bottom) production upon re-stimulation of CD8+ T cells on day six after activation and differentiation in control (DMSO) or BioE-1197 (50μM) conditions. (B) Quantification of the percent of IFNγ (top) and TNFα (bottom) cytokine positive cells represented in A across independent experiments. (C) Quantification of the fold change in geometric mean fluorescence intensity (gMFI) of IFNγ (top) and TNFα (bottom) production by CD8+ T cells represented in A across independent experiments. (D) Quantification of IFNγ (top) and TNFα (bottom) production in the supernatants of re-stimulated CD8+ T cells across independent experiments by ELISA. (E) Representative dot plots of IFNγ (left) and TNFα (right) production upon re-stimulation of WT (top) or PASK knockout (KO) (bottom) CD8+ T cells differentiated in control (DMSO) or BioE-1197 (50μM) conditions. (F) Quantification of the fold change in gMFI of IFNγ (top) and TNFα (bottom) production by WT and PASK KO CD8+ T cells represented in E across independent experiments. IFNγ: Fgenotype p = 0.3640, FBioE-1197 p = 0.0012. TNFα: Fgenotype p = 0.9442, FBioE-1197 p = 0.0133. (G) Schematic representing the experimental design for park and recall co-adoptive transfer of control (DMSO) and BioE-1197 (50μM) differentiated P14 CD8+ T cells. (H) Quantification of the percent of cytokine positive cells for IFNγ and TNFα upon ex vivo peptide re-stimulation and intracellular cytokine staining across ten mice. (I) Quantification of the fold change in gMFI for IFNγ (left) and TNFα (right) upon ex vivo peptide re-stimulation and intracellular cytokine staining across ten mice. Each dot represents values from an independent experiment, summary data are presented as the mean (black line) with SEM error bars (B-D, F). Each dot represents an individual mouse within one independent experiment and presented results are representative of two independent experiments (H, I). Paired t test (B, C, D). Two-way ANOVA, Sidak’s multiple comparisons (F). Paired t test (H, I – IFNγ). Wilcoxon test (I-TNFα). *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001.