Journal: Cell Reports Medicine
Article Title: Coupling IL-2 with IL-10 to mitigate toxicity and enhance antitumor immunity
doi: 10.1016/j.xcrm.2025.102257
Figure Lengend Snippet: Coupling and targeting IL-2 and IL-10 together retains broad immune activation and increases anti-tumor potency in mice (A–M) C57BL/6 mice (C57BL/6 and IFNγ-knockout mice in H) were implanted subcutaneously with a human EGFR-expressing B16F10 cell line (B16F10 hEGFR+ ). B16F10 hEGFR+ tumor-bearing mice were treated three times per week with vehicle control or the reagent(s) indicated. (A–F, H–L) Data are representative of the tumor volumes (mm 3 ) from 4 to 15 animals per group. (A) Mice treated with vehicle, IL-2 (0.4 mg/kg), IL-10 (1.0 mg/kg), or both ( n = 10). Statistical analyses were performed by Mann-Whitney test. (B) Mice treated with vehicle, IL-2 (scFv) (0.4 mg/kg), IL-10 (scFv) (1.0 mg/kg), or both ( n = 10). Statistical analyses were performed by two-way ANOVA with Tukey’s multiple comparisons test. (C) Mice treated with vehicle, IL-2 (EGFR) (0.4 mg/kg), IL-10 (EGFR) (1.0 mg/kg), or both ( n = 10). Statistical analyses were performed by two-way ANOVA with Tukey’s multiple comparisons test. (D) Mice treated with vehicle control or varying doses (0–6 mg/kg) of DK2 10 (EGFR) ( n = 7). Statistical analyses were performed by two-way ANOVA with Tukey’s multiple comparisons test. (E) Mice treated with vehicle, DK2 10 (scFv) (2.0 mg/kg), or DK2 10 (EGFR) (2.0 mg/kg) ( n = 9–10). Statistical analyses were performed by Mann-Whitney test. (F) Mice treated with vehicle, DK2 10 (EGFR) (2.0 mg/kg), or varying doses of IL-2 (0–4.0 mg/kg) ( n = 4–7). Statistical analyses were performed by two-way ANOVA with Tukey’s multiple comparisons test. (G) Mice treated with vehicle, DK2 10 (EGFR) (2.0 mg/kg), or IL-2 (2.0 mg/kg) ( n = 4–7). Serum was collected, and cytokine concentrations are reported. Statistical analyses were performed by Student’s t test. Results are reported as the mean ± SD. (H) Wild-type or IFNγ-knockout mice were treated with vehicle or DK2 10 (EGFR) (2.0 mg/kg) ( n = 7–9). Statistical analyses were performed by Welch’s t test. (I–K) Mice treated with vehicle or DK2 10 (EGFR) (2.0 mg/kg) with and without cell ablation of CD8 + (I), CD4 + (J), or NK cells (K) ( n = 5–6). Statistical analyses were performed by Mann-Whitney test. (L) Mice treated with vehicle or DK2 10 (EGFR) (2.0 mg/kg) with and without FTY720 ( n = 11–15). Statistical analyses were performed by Mann-Whitney test. (M–P) Mice were treated with vehicle or DK2 10 (EGFR) (2.0 mg/kg) for 6–8 days ( n = 4). (M) The frequencies of CD4 + and CD8 + T cells among TILs. (N) Following treatment, CD8 + Tem (CD3 + /CD8 + /CD44 + /CD62L − ) TILs were sorted, and gene expression was performed by nCounter analysis. Genes with a fold change of 1.5 from control animals and p value ≤0.05 are reported. (O) The oxygen consumption rate of isolated CD8 + TILs ( n = 9). (P) Isolated CD8 + TILs from treated mice were rested overnight before co-culturing with B16F10 hEGFR+ cells, and an Elispot was performed to detect IFNγ-producing tumor-reactive cells ( n = 5–9). Statistical analyses were performed by Welch’s t test. (Q) Tumor volumes (mm 3 ) from mice bearing LL2 hEGFR+ tumors treated with vehicle or DK2 10 (EGFR) with or without anti-PD-1 (2.5 mg/dose) for 1 week ( n = 9). Statistical analyses were performed by Welch’s t test. (R) A patient-derived xenograft (PDX) model (non-small cell lung cancer) was established by subcutaneous implant of patient tumor cells, and mice were treated with vehicle, anti-PD-1 (10 mg/kg; every 5 days intraperitoneally), or DK2 10 (EGFR) (0.5 or 2.0 mg/kg, three times a week subcutaneously) ( n = 2). Tumor volumes are reported (mm 3 ). Statistical analyses were performed by Welch’s t test. (ns, not significant; p value; ∗ p ≤ 0.05; ∗∗ p ≤ 0.01; ∗∗∗ p ≤ 0.001; ∗∗∗∗ p ≤ 0.0001). Results are reported as the mean ± SEM, unless otherwise noted.
Article Snippet: Spots were quantified using a MabTech IRIS ELISpot reader with Mabtech Apex 1.1 software.
Techniques: Activation Assay, Knock-Out, Expressing, Control, MANN-WHITNEY, Gene Expression, Isolation, Enzyme-linked Immunospot, Derivative Assay
