Fig. 2
SMG1 reduction in mouse breast cancer tumor cells slow tumor growth in an immune-dependent manner. (A) Several NMD factors (SMG1, UPF1 and UPF2) were knocked-down using CRISPR/Cas9 in 4T1 breast cancer mouse model implanted subcutaneously in Balb/c mice and tumor growth was measured over time. n = 10-12/group. (B) Tumor volumes from (A). Each line depicts the growth over time of an individual tumor. n = 10-12/group. (C) Top: Experiment schedule. Bottom: Tumor volume over time of 4T1.gCtrl tumors implanted subcutaneously in Balb/c mice treated with isotype, anti-CD4 (clone GK1.5), anti-CD8a (clone 53-6.7), anti-Asialo GM1 (Poly21460) (NK) depleting or anti-IFNAR blocking antibodies (clone MAR1-5A3). n = 6-8/group. (D) Tumor weights of (C) at end point (day 24). n = 6-8/group. (E) Knockdown of SMG1 in 4T1 cells enhances immune infiltrate. Balb/c mice were subcutaneously implanted with 4T1.gCtrl or SMG1KD cells and on day 14 tumors were resected and analyzed by flow cytometry. n = 7-8/group. (F) Knockdown of SMG1 in Panc02 cells enhances immune infiltrate. C57/BL6 mice were subcutaneously implanted with Panc02.gCtrl or SMG1KD cells and on day 14 tumors were resected and analyzed by flow cytometry. n = 6/group. Left to right: CD8 T cell infiltration, CD4+ T cell infiltration, Treg (FOXP3+CD25+) T-cell infiltration, CD8+/Treg ratio. n = 6/group. (G) Draining lymph node flow immune analysis by cytometry study in 4T1.SMG1KD and 4T1.gCtrl tumor bearing mice at day 14. n = 5 (4T1.gCtrl group); 8 (4T1.SMG1KD group). (H) Draining lymph node flow immune analysis by cytometry study in Panc02.SMG1KD and Panc02.gCtrl tumor bearing mice at day 14. n = 6/group. p-values are shown for 2-way ANOVA with Bonferroni’s post-hoc test for tumor growth experiments. 1-way ANOVA was performed in (D); and 2-tailed t-test was performed in the infiltrate study. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001