Figure 5. YAP and β-catenin pathways are both required for the Cdh1^−/−RHOA^Y42C/+-induced transformation.

A, Tumor volume of Cdh1^−/−RHOA^Y42C/+ organoids with ectopic expression of either TCF4-DN (del aa 1–31) or YAP-DN (S94A) followed by flank implantation, with representative images (bottom) of tumors from each group (n=4 for each). Data are mean ± S.E.M. ****P<0.0001, two-way ANOVA, (TCF4-DN or YAP-DN versus EV group). B, Representative images of Ki67 staining of the tumors from (A). Scale bar = 50 μm. C, Representative H&E images for the tumors from (A). Scale bar = 100 μm. D, Tumor volume of Trp53^−/−Kras^G12D/+ organoids with ectopic expression of TCF4-DN (del aa 1–31) or YAP-DN (S94A) or combination, implanted into flanks, with representative images of tumors from each group (n = 4 for each). Data are mean ± S.E.M. E, In vitro proliferation of Cdh1^−/−RHOA^Y42C/+ and Trp53^−/−Kras^G12D/+ organoids treated with DMSO or verteporfin (YAP inhibitor, 5 μM) combined with ICG-001 (antagonist of β-catenin/TCF4 binding, 5 μM) or MK-2206 (AKT inhibitor, 2 μM) for 48 h. Data are mean ± S.E.M. *P<0.05, ***P<0.001, unpaired two-tailed Student’s t-test. F, Representative phase contrast images of Cdh1^−/−RHOA^Y42C/+ or Trp53^−/−Kras^G12D/+ organoids treated for 48 h with DMSO or verteporfin (5 μM), combined with ICG-001 (5 μM) or MK-2206 (2 μM). Scale bar = 100 μm. G, Tumor volume of Cdh1^−/−RHOA^Y42C/+ organoids injected into flanks of NSG mice and treated with DMSO, pictilisib (PI3K inhibitor, 75 mg/kg), verteporfin (100 mg/kg) or the combination (n = 8 tumors for each). Data are mean ± S.E.M. **P<0.01, ****P<0.0001, two-way ANOVA (treatment versus DMSO).