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L.R. Kong

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    P1.02 - Poster Session with Presenters Present (ID 454)

    • Event: WCLC 2016
    • Type: Poster Presenters Present
    • Track: Biology/Pathology
    • Presentations: 1
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      P1.02-041 - Characterization of MET-N375S as an Activating Mutation in Squamous Cell Carcinoma of the Lung (ID 5705)

      14:30 - 15:45  |  Author(s): L.R. Kong

      • Abstract
      • Slides

      Over the years, significant progress has been made in the treatment of lung carcinoma. While targeting EGFR mutations in the tyrosine kinase domain and EML4-ALK rearrangements have yielded impressive therapeutic gains in lung adenocarcinoma; the rarity of genetic aberration in squamous cell carcinoma (SCC) has restricted the use of molecular-targeting agents. Next-Generation sequencing analysis has identified a high frequency of c-MET mutation in Asian lung SCC specimens, in particular the MET-N375S mutant. This study aims to characterize the functional significance and therapeutic implications of MET-N375S in lung SCC cells.

      MET (N375S) mutation in tumour tissues was verified with droplet digital PCR. c-MET mutant expressing clones were generated through site-directed mutagenesis and single cell clonal selection. The impact of MET-N375S in lung SCC cells was characterized by defining the downstream effectors (Proteome profiling), biological functions (anchorage-independent growth, invasion and migration assays), transciptomic regulation (RNAseq) and protein-protein interaction (SILAC). Novel binding partners of MET-N375S was verified using co-immunoprecipitation (co-IP) and proximity ligation assay (PLA). Sensitivity of c-MET mutant to various kinase inhibitors was determined with CellTiter-Glo assay.

      MET (N375S) mutation was confirmed in 9/45 lung SCC specimens (20%). Ectopic expression of MET-N375S in lung SCC cells significantly elevated the activation of downstream Src, p38 and ERK1/2 kinases, increased hsp27 expression, while enhanced migratory, invasive and anchorage-independent colony forming ability in vitro. Despite so, comparative transcriptomic analysis revealed that epithelial-mesenchymal signature genes were not induced in cells expressing mutant c-MET. On the contrary, SILAC and co-IP analyses on the MET-N375S interactome demonstrated an increase in binding affinity towards receptor tyrosine kinases (RTKs) as compared to wild type c-MET, which was confirmed with in situ PLA. Moreover, MET-N375S augmented in vitro sensitivity to selective MET inhibitors.

      These findings suggest the role of MET-N375S as an activating mutation that strongly enhance malignant transformation and metastatic potential in lung SCC cells. Mechanistically, the dysregulation of various oncogenic events could be associated with the formation of heterodimers with RTKs. Clinically, MET-N375S could be utilized as a potential predictive biomarker for patients with advanced SCC of the lung to be treated with selective MET inhibitors.

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