ABSTRACT
OBJECTIVE: Genomic studies of gastric cancer have identified highly recurrent genomic alterations impacting RHO signalling, especially in the diffuse gastric cancer (DGC) histological subtype. Among these alterations are interchromosomal translations leading to the fusion of the adhesion protein CLDN18 and RHO regulator ARHGAP26. It remains unclear how these fusion constructs impact the activity of the RHO pathway and what is their broader impact on gastric cancer development. Herein, we developed a model to allow us to study the function of this fusion protein in the pathogenesis of DGC and to identify potential therapeutic targets for DGC tumours with these alterations. DESIGN: We built a transgenic mouse model with LSL-CLDN18-ARHGAP26 fusion engineered into the Col1A1 locus where its expression can be induced by Cre recombinase. Using organoids generated from this model, we evaluated its oncogenic activity and the biochemical effects of the fusion protein on the RHOA pathway and its downstream cell biological effects in the pathogenesis of DGC. RESULTS: We demonstrated that induction of CLDN18-ARHGAP26 expression in gastric organoids induced the formation of signet ring cells, characteristic features of DGC and was able to cooperatively transform gastric cells when combined with the loss of the tumour suppressor geneTrp53. CLDN18-ARHGAP26 promotes the activation of RHOA and downstream effector signalling. Molecularly, the fusion promotes activation of the focal adhesion kinase (FAK) and induction of the YAP pathway. A combination of FAK and YAP/TEAD inhibition can significantly block tumour growth. CONCLUSION: These results indicate that the CLDN18-ARHGAP26 fusion is a gain-of-function DGC oncogene that leads to activation of RHOA and activation of FAK and YAP signalling. These results argue for further evaluation of emerging FAK and YAP-TEAD inhibitors for these deadly cancers.
Subject(s)
Claudins , GTPase-Activating Proteins , Mice, Transgenic , Signal Transduction , Stomach Neoplasms , Transcription Factors , YAP-Signaling Proteins , rhoA GTP-Binding Protein , Animals , Stomach Neoplasms/genetics , Stomach Neoplasms/pathology , Stomach Neoplasms/metabolism , GTPase-Activating Proteins/genetics , GTPase-Activating Proteins/metabolism , Mice , rhoA GTP-Binding Protein/metabolism , rhoA GTP-Binding Protein/genetics , Claudins/genetics , Claudins/metabolism , YAP-Signaling Proteins/metabolism , YAP-Signaling Proteins/genetics , Transcription Factors/genetics , Transcription Factors/metabolism , Humans , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Focal Adhesion Protein-Tyrosine Kinases/metabolism , Focal Adhesion Protein-Tyrosine Kinases/genetics , TEA Domain Transcription Factors , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Disease Models, Animal , Organoids/metabolism , Organoids/pathologyABSTRACT
Cell state plasticity is carefully regulated in adult epithelia to prevent cancer. The aberrant expansion of the normally restricted capability for cell state plasticity in neoplasia is poorly defined. Using genetically engineered and carcinogen-induced mouse models of intestinal neoplasia, we observed that impaired differentiation is a conserved event preceding cancer development. Single-cell RNA sequencing (scRNA-seq) of premalignant lesions from mouse models and a patient with hereditary polyposis revealed that cancer initiates by adopting an aberrant transcriptional state characterized by regenerative activity, marked by Ly6a (Sca-1), and reactivation of fetal intestinal genes, including Tacstd2 (Trop2). Genetic inactivation of Sox9 prevented adenoma formation, obstructed the emergence of regenerative and fetal programs, and restored multilineage differentiation by scRNA-seq. Expanded chromatin accessibility at regeneration and fetal genes upon Apc inactivation was reduced by concomitant Sox9 suppression. These studies indicate that aberrant cell state plasticity mediated by unabated regenerative activity and developmental reprogramming precedes cancer development.
Subject(s)
Adenoma , Colorectal Neoplasms , Mice , Animals , Intestines , Colorectal Neoplasms/genetics , Colorectal Neoplasms/pathology , Cell Differentiation , Adenoma/genetics , Adenoma/pathologyABSTRACT
Dysregulated Wnt/ß-catenin signaling is implicated in the pathogenesis of many human cancers, including colorectal cancer (CRC), making it an attractive clinical target. With the aim of inhibiting oncogenic Wnt activity, we developed a high-throughput screening AlphaScreen assay to identify selective small-molecule inhibitors of the interaction between ß-catenin and its coactivator BCL9. We identified a compound that consistently bound to ß-catenin and specifically inhibited in vivo native ß-catenin/BCL9 complex formation in CRC cell lines. This compound inhibited Wnt activity, down-regulated expression of the Wnt/ß-catenin signature in gene expression studies, disrupted cholesterol homeostasis, and significantly reduced the proliferation of CRC cell lines and tumor growth in a xenograft mouse model of CRC. This study has therefore identified a specific small-molecule inhibitor of oncogenic Wnt signaling, which may have value as a probe for functional studies and has important implications for the development of novel therapies in patients with CRC.