Functional Analysis of MET Exon 14 Skipping Alteration in Cancer Invasion and Metastatic Dissemination

Abstract

MET exon 14 skipping alteration (METΔ14Ex) is an actionable oncogenic driver that occurs in 2% to 4% of non–small cell lung cancer (NSCLC) cases. The precise role of METΔ14Ex in tumor progression of NSCLC is poorly understood. Using multiple isogenic METΔ14Ex cell models established with CRISPR editing, we demonstrate that METΔ14Ex expression increases receptor kinase activity and downstream signaling by impairing receptor internalization and endocytic degradation, significantly boosting cell scatter, migration, and invasion capacity in vitro as well as metastasis in vivo. RNA sequencing analysis revealed that METΔ14Ex preferentially activates biological processes associated with cell movement, providing novel insights into its unique molecular mechanism of action. Activation of PI3K/Akt/Rac1 signaling and upregulation of multiple matrix metallopeptidases (MMP) by METΔ14Ex induced cytoskeleton remodeling and extracellular matrix disassembly, which are critical functional pathways that facilitate cell invasion and metastasis. Therapeutically, MET inhibitors dramatically repressed METΔ14Ex-mediated tumor growth and metastasis in vivo, indicating potential therapeutic options for METΔ14Ex-altered NSCLC patients. These mechanistic insights into METΔ14Ex-mediated invasion and metastasis provide a deeper understanding of the role of METΔ14Ex in NSCLC. Significance: These findings reveal the mechanistic function of METΔ14Ex alteration in driving metastasis and define novel metastasis-related pathways that could be targeted for more effective treatment of lung cancer with METΔ14Ex alterations.

Characterization of MET Exon 14 Skipping Alterations (in NSCLC) and Identification of Potential Therapeutic Targets Using Whole Transcriptome Sequencing

“…Last, consistent with our previous preclinical studies, 18 an up-regulation of genes related to cellular adhesion, extracellular matrix disassembly, and angiogenesis, mechanisms of invasion and metastases in MET ex14 were also up-regulated in our MET ex14 cohort by pathway analysis. Tumor angiogenesis is hypothesized to promote an immunosuppressive environment 45 and may serve as an additional factor in immunotherapy resistance in MET ex14.…”

“…10 ). mRNA expression of individual markers involved in invasion and metastases of MET , including PLAU , SERPINE1 , CSF2 , MMP3 , EFNB2 , PLAUR , IL1A , CXCL2 , and VEGFC , which were up-regulated in our previously established preclinical cell-based models of MET ex14 18 was also found to be significantly overexpressed in MET ex14 patients (q < 0.00001 for all) ( Fig. 5 B ).…”

“…An up-regulation in tumor necrosis factor in METex14 was observed, which was not observed in our previous preclinical model. 18 On 70 cases with data available for logistic regression on multivariate analysis, apical junction pathway was observed to be associated with METex14 (p < 0.05). These up-regulated pathways and genes highlight several potential vulnerabilities that can be explored for potential therapeutic benefit.…”

“…We have conducted preclinical assays on clustered, regularly interspaced, short palindromic repeat-modified cellular models of MET ex14-driven lung cancer and have found that MET ex14 NSCLC cells were associated with pathways related to cytoskeletal remodeling, cellular adhesion, epithelial to mesenchymal transition (EMT), and angiogenesis. 18 The in vivo up-regulation and biological role of these pathways in MET ex14-driven lung cancers remain to be understood and may open novel avenues for more effective and durable therapeutic interventions.…”

Transforming properties of MET receptor exon 14 skipping can be recapitulated by loss of the CBL ubiquitin ligase binding site

“…The receptor lacking the juxtamembrane domain has been produced by ectopic expression in several fibroblast cell lines, but these do not allow proper reproduction of biological responses [44,45]. MET exon 14 skipping has also been mimicked by genome editing in human kidney embryonic (HEK) cells and a human lung cell line [46][47][48]. Taken together, these studies assess that MET deprived of its juxtamembrane domain triggers stronger activation of the downstream signaling pathway, promotes proliferation and anchorage-independent growth, and favors growth of experimental tumors.…”