Fig. 4

Reduced suppressive function of MDSCs lacking Siglec-E upon sialidase or Siglec-E blocking antibody treatment. A Experimental setup: Depletion of Ly6G-positive cells in SigE^ΔLysM mice and SigE^WT littermates bearing B16F10 tumors using a depleting antibody. Mice were injected up to 6 times (gray arrow) with the anti-Ly6G depletion antibody starting 1 day before subcutaneous B16F10 tumor injection (black arrow). Tumor growth and survival were monitored. B Kaplan‒Meier survival curves or (C) tumor growth curves from pooled experiments from (A). n = 9–11 mice per group. D Experimental setup for assessing the effect of the DR5 antibody on SigE^ΔLysM mice and SigE^WT littermates subcutaneously injected with B16F10 tumors. E Kaplan‒Meier survival curves and (F) tumor growth curves from (D) with n = 6–7 mice per group. G Experimental setup for assessing the suppressive effect of Gr1⁺CD11b⁺ (MDSC) cells on naïve CD3⁺ (T cell) cells. MDSCs were isolated from the spleens of B16F10 tumor-bearing SigE^ΔLysM mice and SigE^WT littermates. T cells were isolated from naïve littermates and stained with CellTrace Violet (CTV) to track T-cell proliferation by flow cytometry. Stained T cells were cocultured with MDSCs for 48 h in the presence of aCD3, aCD28 and IL-2. MDSCs were used immediately or pretreated with sialidase. A Siglec-E blocking antibody or rat IgG2a, κ isotype control was added to the cocultures as indicated. H Percentage of proliferating CD8⁺ T cells cocultured without MDSCs, (I) with MDSCs from SigE^WT mice or (J) MDSCs from SigE^ΔLysM mice. Exemplary results for each untreated condition are shown on the right, indicating untreated T cells alone (J, gray), SigE^WT (K, green), and SigE^ΔLysM (L, pink). n = 3–9 mice per condition. The data are presented as the mean ± SD or SEM (C, F). Multiple paired t tests were used. For survival analysis, the log-rank test was used, followed by the Šidák correction for multiple comparisons. Tumor growth was compared by mixed-effects analysis followed by Bonferroni’s multiple comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001