Journal: Cell Reports Medicine

Article Title: Pan-cancer mapping of single CD8 + T cell profiles reveals a TCF1:CXCR6 axis regulating CD28 co-stimulation and anti-tumor immunity

doi: 10.1016/j.xcrm.2024.101640

Figure Lengend Snippet: CXCR6 and CXCL16 expression in human and murine tumors tracks with dysfunctional T cells and myeloid cells, respectively (A) Mean expression (color bar) and fraction of expressing cells (dot size) for CXCR6 and CXCL16 (columns) across cell types (rows) in different human tumor studies (panels). (B) GSEA plots obtained for the KEGG_ANTIGEN_PROCESSING_AND_PRESENTATION gene set when compared against the ranking of genes based on their co-expression with CXCL16 in macrophages in the indicated tumors. (C) Frequency of CXCR6 + cells (y axis, mean ± SEM) in the indicated subsets (x axis) of total (top) or Ova-Dextramer + (bottom) CD8 + TILs harvested from B16F10, B16Ova, or Mc38Ova hi tumors (top: B16F10, n = 8, 2 experiments combined. B16Ova, n = 10, 2 experiments combined. Mc38Ova hi , n = 9, 2 experiments combined. Bottom: B16Ova, n = 3, 1 experiment. Mc38Ova hi , n = 15, 2 experiments combined). (D) Representative distributions of expression levels (x axis, fluorescence intensity) in CXCR6 + and CXCR6 − CD8 + TILs from B16Ova tumors ( n = 4, 1 experiment). (E) Representative distributions of CXCL16 surface (top) or intracellular (bottom) expression (x axis, fluorescence intensity) in myeloid cells from Mc38Ova hi , B16Ova, or B16F10 tumors ( n = 4 per tumor, 1 experiment). (F) Left: B16Ova tumor area (y axis, mean ± SEM) of early-stage or late-stage tumors. Middle: frequency (y axis, mean ± SEM) of PD1- and TIM3-expressing CD8 + TILs (x axis) in early- or late-stage tumors. Right: frequency of CXCR6 + cells (y axis, mean ± SEM) in the indicated subsets (x axis) of CD8 + TILs from early- or late-stage B16Ova tumors ( n = 12, 2 experiments combined). Statistical significance was determined by Student’s unpaired t test (C, F). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: Anti-Mouse Tim3 APC Clone 5D12 , BD , Cat:# 567164 RRID: AB_2916481.

Techniques: Expressing, Fluorescence

Journal: Cell Reports Medicine

Article Title: Pan-cancer mapping of single CD8 + T cell profiles reveals a TCF1:CXCR6 axis regulating CD28 co-stimulation and anti-tumor immunity

doi: 10.1016/j.xcrm.2024.101640

Figure Lengend Snippet: CXCR6 expression increases upon ICB and is repressed by TCF1 (A and B) Left: tumor area (y axis, mean ± SEM) over time (x axis) of Mc38Ova hi (A) or B16Ova (B) implanted in wild-type (WT) mice and treated with anti-PD1 (A), anti-PD-L1 + anti-TIM3 (B), or isotype control. Right: frequency of CXCR6 + cells (y axis, mean ± SEM) in the indicated subsets (x axis) of CD8 + TILs harvested from tumors treated as above (A: n = 4–6 per group, 1 experiment. B: n = 3–4 per group, 1 experiment). (C) Expression (row-normalized TPM, color bar) of top differentially expressed genes (rows) between WT OTI (E8i-Cre - , Tcf7 FL/FL ) and Tcf7 cKO OTI (E8i-Cre + , Tcf7 FL/FL ) cells at different time points after T cell activation (columns) ( n = 3, 1 experiment). (D) Mean expression (color bar) and fraction of expressing cells (dot size) of key genes (rows) in different CD8 + T cell clusters (columns) as determined by scRNA-seq of cells from B16Ova tumors implanted in WT or Tcf7 cKO mice ( n = 3 per group combined, 1 experiment). (E) Frequency of CXCR6 + cells (y axis, mean ± SEM) in the indicated subsets (x axis) of CD8 + TILs from B16Ova tumors from WT (black) or Tcf7 cKO (green) mice ( n = 3–7 per group, representative of 2 experiments). (F) Percent input (y axis, mean ± SEM) following chromatin immunoprecipitation (ChIP) PCR of the Cxcr6 locus with anti-TCF1 or rabbit IgG control antibodies (x axis) in WT or Tcf7 cKO CD8 + T cells ( n = 8, 5 experiments combined). (G–I) Luciferase activity (RLU, relative light unit, y axis, mean ± SEM) in HEK293T cells transfected with Cxcr6 locus-containing pGL4.10 luciferase reporters together with either empty vector (control) or vectors encoding the indicated transcription factors (x axis). Firefly luciferase activity is presented relative to constitutive Renilla luciferase activity ( n = 3, representative of 2 experiments). Statistical significance was determined by linear mixed model (A left, B left), Student’s unpaired t test (A right, B right, E, F comparing WT to Tcf7 cKO anti-TCF1 samples), Student’s paired t test (F comparing WT samples), or one-way ANOVA with Tukey’s multiple comparisons test (G, H, I). NS = not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: Anti-Mouse Tim3 APC Clone 5D12 , BD , Cat:# 567164 RRID: AB_2916481.

Techniques: Expressing, Control, Activation Assay, Chromatin Immunoprecipitation, Luciferase, Activity Assay, Transfection, Plasmid Preparation

Journal: Cell Reports Medicine

Article Title: Pan-cancer mapping of single CD8 + T cell profiles reveals a TCF1:CXCR6 axis regulating CD28 co-stimulation and anti-tumor immunity

doi: 10.1016/j.xcrm.2024.101640

Figure Lengend Snippet:

Article Snippet: Anti-Mouse Tim3 APC Clone 5D12 , BD , Cat:# 567164 RRID: AB_2916481.

Techniques: Virus, Recombinant, Purification, SYBR Green Assay, Staining, Reporter Assay, In Vitro, Activation Assay, Generated, Luciferase, Plasmid Preparation, Sequencing, Software, CRISPR