MET exon 14 skipping mutations (METΔ14Ex) represent a new class of molecular alterations that occur in approximately 3%-5% of non-small cell lung cancer (NSCLC) cases, and recently emerged as an actionable oncogenic driver with significant responsiveness to MET tyrosine kinase inhibitors, including crizotinib and capmatinib. METΔ14Ex mutations are generally was found to be mutually exclusive with other recognized oncogenic drivers and higher frequency has been noted in a unique, highly aggressive subtype of lung cancer called pulmonary sarcomatoid carcinoma. In the current study, we sought to perform an array of experiments to determine the intrinsic tumorigenic role of METΔ14Ex in NSCLC. Using multiple isogenic METΔ14Ex cell models established with CRISPR, we first determined that METΔ14Ex increased HGF-dependent kinase activity and enhanced HGF/MET downstream signaling through impairing receptor endocytic degradation. In subsequent functional analyses, we identified that METΔ14Ex significantly increases HGF-dependent cell scatter, migration and invasion capacity in vitro as well as metastasis in vivo compared to wild-type MET (MET WT). To explore the intrinsic mechanism in regulation of aforementioned phenotypes, an RNA sequencing study was conducted to evaluate the global changes of dynamic gene expression profile and key functional pathways. Ingenuity pathway analysis (IPA) and gene set enrichment analysis (GSEA) revealed a large cluster of cell movement-related signaling pathways that were remarkably activated in HGF-treated METΔ14Ex models (vs MET WT) in a time-dependent manner. In addition, a number of upregulated DEGs (Differently Expressed Genes) were enriched in functional categories of cell cytoskeleton reorganization and cell-matrix adhesion degradation in METΔ14Ex models (vs MET WT). Based on the information of GSEA, we further found that the activity of the small GTPase Rac1, a crucial regulator of cytoskeleton dynamics, was significantly increased by HGF treatment in METΔ14Ex cells, whereas inhibition of the PI3K/Akt pathway repressed HGF/METΔ14Ex-mediated Rac1 activity and cell movement. Moreover, a group of Matrix metallopeptidases (MMPs) were highly upregulated in our models. Notably, among these MMPs, HGF-dependent upregulation of MMP1 expression (>100 fold in both mRNA and protein level) was very prominent in METΔ14Ex compared to MET WT cells, and silencing MMP1 using siRNA dramatically repressed HGF-triggered METΔ14Ex cell invasion. We finally found that the MET inhibitors, MGCD516 and capmatinib remarkably inhibited HGF/METΔ14Ex signaling and repressed cell invasion in vitro and metastasis in vivo. In conclusion, these data demonstrate that METΔ14Ex plays a critical role in enhancing cell movement and metastasis, at least partially via upregulation of the PI3K/Akt-Rac1 pathway and expression of MMPs, and blockade of HGF/METΔ14Ex signaling using MET inhibitors might be potent therapeutic options for METΔ14Ex-mutated NSCLC patients and recognition of the unique biology of METΔ14Ex-mediated oncogenesis might yield additional synergistic treatment strategies.
Citation Format: Feng Wang, Yang Liu, Elaine Shum, Mariam Alexander, Deyou Zheng, Roman Perez-Soler, Haiying Cheng, Balazs Halmos. A novel functional analysis of MET exon 14 skipping mutation in regulation of tumor cell invasion and metastasis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4907.