Transcriptomic profiling of a novel gastric implantation model identifies mechanisms and pathways that drive implantation into explanted human peritoneum
Epithelial cancers such as stomach and ovary cancer tend to metastasize to the peritoneum, often leading to intractable disease and poor survival. Currently, the mechanisms that enable gastric cancer cells to penetrate the mesothelium, and to implant, invade, and survive in the peritoneal niche are poorly understood. To investigate these mechanisms, we developed a novel human peritoneal explant model. Briefly, fresh peritoneal tissue samples from abdominal surgery patients were cultured on top of a layer of GFP-labeled human gastric adenocarcinoma cells (AGS); 2% of these cells implanted into the peritoneum. The transcriptomic profile of the implanted AGS cells was compared to the profile of AGS cells that failed to implant using RNA sequencing. Differentially expressed genes were enriched significantly (fold change>2) for genes that enable cell adhesion, motility, and membrane depolarization. We compared this list of genes with a previously identified peritoneal metastasis whole exome sequencing dataset (SRP043661). Upon further analysis based on subcellular localization, cell adhesion and cytoskeletal organization, we found nine core "peritoneal implantation" genes. We functionally validated these genes with CRISPR knockout and assessed peritoneal implantation and invasion using the human peritoneal explant model described above. From these data we identified ADAM12 as a key player of peritoneal metastasis. Knock out of ADAM12 significantly impaired peritoneal metastasis in vivo and ex vivo. Exploration of three publicly available independent datasets indicated that ADAM12 is indeed clinically relevant in peritoneal metastasis. To explore the role of ITGA{beta}1 in mediating cell-matrix interactions in the presence of ADAM12, we performed ITGA{beta}1 pull-down assays followed by mass spectrometry analysis in ADAM12 WT cells. ADAM12 KO cells show a marked disruption of the ITGA{beta}1 interactome in GCa cells. Key cytoskeletal proteins such as MYH14, MYH10, MYH9, ACTA2, SPTN1 and TPM1-TPM4 were found to be interactors with ITGA{beta}1 in ADAM12 WT cells but not ADAM12 KO cells. Our approach and the new data identify a distinct peritoneal metastasis gene set that facilitates implantation and invasion of gastric cancer cells within the peritoneum. Disruption of these pathways with peritoneal-directed therapies has the potential to improve survival in patients with high-risk primary gastric cancer.