Panel A shows flow cytometry plots divided into LAG3 negative tdTomato negative, LAG3 negative tdTomato positive, LAG3 positive tdTomato negative, and LAG3 positive tdTomato positive groups, together with a bar graph showing the percentage of Thy1.2 positive CD8 positive T cells expressing tdTomato in each group. Panel B shows a schematic diagram outlining the experimental design used to evaluate the differentiation potential of circulating LAG3 negative tdTomato positive cells following tumor rechallenge. Panel C shows flow cytometry plots and bar graphs of LAG3 and tdTomato expression in CD8 positive T cells isolated from B16-F10 and MC38 tumors, with the bar graphs showing the percentage of LAG3 positive tdTomato positive cells among live CD8 positive T cells. Panel D shows corresponding flow cytometry plots and bar graphs for tumors from SHAM control mice. Panel E shows a schematic diagram outlining the experimental design in a monoclonal setting. Panel F shows flow cytometry plots and bar graphs of LAG3 and tdTomato expression in CD8 positive T cells isolated from B16-gp100 and MC38 tumors, with the bar graphs showing the percentage of LAG3 positive tdTomato positive cells among live CD8 positive T cells. Panel G shows corresponding flow cytometry plots and bar graphs for tumors from SHAM control mice.
Antigen-driven differentiation of circulating LAG3–tdT+ CD8+ T cells upon tumor rechallenge . (A) LAG3 and tdT expression was assessed on the CD8+ T cells in peripheral blood before rechallenge on d42. (B) Scheme to evaluate the differentiation potential of circulating LAG3−tdT+ cells upon tumor rechallenge was designed to be antigen driven. (C and D) LAG3 and tdT expression were subsequently assessed in (C) tumors of experimental mice and (D) tumors of SHAM controls. (E) Scheme to evaluate the differentiation potential of circulating LAG3−tdT+ cells upon tumor rechallenge was designed to be antigen driven in the monoclonal setting. (F and G) LAG3 and tdT expression were subsequently assessed in (F) tumors of experimental mice and (G) tumors of SHAM controls. Data in A are representative of three independent experiments with n = 8 mice. Data in C, D, F, and G are from n = 5–8 mice per group, pooled from two independent experiments. *P < 0.05; **P < 0.01; ****P < 0.0001 by (A) one-way ANOVA, by (C) paired t test, and by (D, F, and G) unpaired t test.