Figure 2 from IFNγ Production by Functionally Reprogrammed Tregs Promotes Antitumor Efficacy of OX40/CD137 Bispecific Agonist Therapy

<p>OX40/CD137 bispecific agonist (FS120m) drives functional fragility and lineage instability of Tregs. <b>A,</b> Schema (top) showing tamoxifen and FS120m treatment schedule. Tumor measurements (bottom) at indicated timepoints after MC38 tumor implantation. (Solid line, mean values; dotted lines, individual mouse tumor curves.) <b>B,</b> Representative plots showing percentages of CD4⁺ GFP⁺ RFP⁺ Tregs and CD4⁺ RFP⁺ GFP⁻ exTregs of total CD4⁺ T cells in the spleens and MC38 tumors of <i>Foxp3</i>^{EGFP-Cre-ERT2}<i>Rosa</i>^flSTOPfl-RFP reporter mice after treatment. <b>C,</b> Quantification of the percentage of RFP⁺ single-positive exTregs (GFP⁻) out of total RFP⁺ cells in spleens and tumors. <b>D,</b> Representative plots showing the production of IFNγ and TNF by indicated cell subsets from the spleens and tumors of reporter mice. <b>E,</b> Quantification of IFNγ and TNF production by indicated cell types from the spleens and tumors of MC38 tumor–bearing mice at day 21 after tumor implantation. <b>F,</b> Schema showing setup of Treg suppression assay (left), representative flow plots (center), and replicate measurements (right) showing percentage of dividing responders in indicated conditions. Data were analyzed by two-way ANOVA with Šídák correction for multiple comparisons (<b>A</b>), unpaired Student <i>t</i> test (<b>C</b>), with Bonferroni–Dunn correction for multiple comparisons (<b>E</b>), and ordinary one-way ANOVA with Tukey correction for multiple comparisons (<b>F</b>). Bars and error are mean and SEM. *, <i>P</i> ≤ 0.05; **, <i>P</i> ≤ 0.01; ***, <i>P</i> ≤ 0.001; ****, <i>P</i> ≤ 0.0001. Ctrl, control.</p>