Figure 5.

CD8⁺ T‐cell depletion does not impact 3M‐052 antiprimary tumor effects but attenuates enhanced long‐term survival. (a) Experimental intervention timeline. BALB/c mice were injected with 1 × 10⁵ 4T1.2 cells into the 4th mammary fat pad, and palpable tumors were i.t. injected with vehicle or 3M‐052 on day 7 post‐inoculation. Mice were treated with five doses of isotype or anti‐CD8 mAb, and primary tumors were resected on day 13. (b) Day 5 flow cytometry analysis of peripheral blood (PB) TCRβ⁺CD8⁺ cell proportions. Primary tumors were assessed for (c) weight (mg); (d) IFN‐γ⁺TCRβ⁺CD8⁺ T cells; (e) CD206⁺F4/80⁺ macrophages; and (f) CD11b⁺ and (g) CD103⁺ DC MHC‐I and (h) F4/80⁺ macrophage MHC‐I (H2‐K^d) expression. (i) Kaplan–Meier survival curve comparing metastasis‐free survival. (j) A proportion of lungs were taken at endpoint for H&E staining to assess metastatic burden; tumor regions are denoted by dotted line and ‘T’ (4× and 20× magnification, scale bars = 200 µm, representative images shown). (k) Lungs were taken at endpoint and assessed for tumor‐specific CD103⁺CD8⁺ T cells in an ICS for IFN‐γ by flow cytometry. MΦ: macrophage. MFI: mean fluorescence intensity. Data are representative of one experiment. (b, c, i) n = 8 mice/group; (d–h, k) n = 6 mice/group; and (j) n = 2 mice/group. Statistical analysis was performed by (b) the Student's two‐tailed t‐test; (c–h, k) one‐way ANOVA with post hoc Tukey's multiple comparison test; and (i) the Mantel–Cox log‐rank test. Error bars are SEM. *P < 0.05; **P < 0.01; ***P < 0.001; and ****P < 0.0001.