Fig. 2

Inhibition of BCL9 enhances response to anti-PD-1 antibody in CRC models. a Combination treatment of Bcl9-shRNA and anti-PD-1 antibody resulted in almost complete regression of the CT26 tumor. BALB/c mice were either inoculated with CT26 cells via single flank implantation and treated with immunoglobulin G (IgG), anti-PD-1 antibody [twice-weekly (BIW)], or with CT26 cells that transduced with non-targeting (NT)-shRNA or Bcl9-shRNA. The mice were subsequently treated with anti-PD-1 antibody as indicated after tumor volume reached 40–60 mm³ (n = 5 per cohort). b Image of tumor tissue from (a). c Ratio of CD4⁺CD25⁺FOXP3⁺ cells among CD45⁺ cell populations in the tumors from (a). d Percentage of GZMB⁺ cells among CD45⁺CD8⁺ T cells in the tumor from (a) was analyzed. e Percentage of IFN-γ⁺ T cells among CD45⁺CD8⁺ T cells in the tumor from (a) was analyzed. f Mice inoculated with Bcl9-shRNA-transduced-MC38 cells and treated with anti-PD-1 antibody resulted in almost complete regression of the MC38 tumor. C57BL/6 mice were inoculated with MC38 cells via single flank implantation and treated with immunoglobulin G (IgG) or anti-PD-1 antibody (BIW), or with Bcl9-shRNA-transduced-MC38 cells and treated with anti-PD-1 antibody as indicated after tumor volume reached 40–60 mm³ (n = 5 per cohort). Ab, antibody. Results were denoted as means ± SEM for experiments performed in triplicate. g Kaplan–Meier analysis of survival end point (tumor size >2000 mm³, n = 8 per cohort). h Mice were inoculated with Bcl9-shRNA-transduced-CT26 cells used anti-PD-1 antibody resulted in almost complete regression of the CT26 tumor. BALB/c mice were inoculated with CT26 cells via single flank implantation and treated with anti-PD-1 antibody (BIW), the mice were inoculated with Bcl9-shRNA-transduced-CT26 cells used anti-CD8 antibody (intraperitoneal injection, i.p., 300 μg per mouse at days 12, 15, and 19 after tumor cell inoculation) plus anti-PD-1 antibody as indicated after tumor volume reached 40–60 mm³ (n = 5 per cohort). Each experiment was repeated three times, and the statistical significance of differences between groups was determined by Two-way ANOVA or non-parametric test (Two-tailed). P < 0.05 means statistically significant.