Figure 1. Cdh1 loss with RHOA hotspot mutation induces diffuse gastric cancer in vivo.

A, Schematic for the generation of mice with distinct genotypes, including the tomato-GFP reporter allele; bottom: representative stack confocal image of gastric organoids with Mist1CreERT2-R26mTmG 48 hours after tamoxifen (2 μM) induction in vitro. Representative images of (B) phase contrast and (C) H&E for gastric organoids with annotated genotypes after three weeks following in vitro tamoxifen induction. Scale bar = 100 μm. D, Representative higher-magnification image showing signet ring cells in Cdh1^−/−RHOA^Y42C/+organoids following tamoxifen induction. Scale bar = 50 μm. E, Procedure of orthotopic injection into gastric wall with green arrow noting location following injection. F, Representative gross images of mouse with gastric tumor, ascites, peritoneal spread with liver metastases at eight weeks following orthotopic injection of Cdh1^−/−RHOA^Y42C/+ organoids. G, Representative H&E images of liver and lung metastases following orthotopic implantation of Cdh1^−/−RHOA^Y42C/+ organoids. Scale bar = 100 μm. H, Kaplan-Meier survival curve following orthotopic implantation of organoids of noted genotypes. Log-rank (Mantel-Cox) test, P=0.0047 (Cdh1^−/−RHOA^Y42C/+ versus other genotypes).