Author of

• 25940

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  P2.03 - Biology (Not CME Accredited Session) (ID 952)

  • Event: WCLC 2018
  • Type: Poster Viewing in the Exhibit Hall
  • Track:
  • Presentations: 1
  • Moderators:
  • Coordinates: 9/25/2018, 16:45 - 18:00, Exhibit Hall

  • 26956

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    P2.03-19 - RET-Mediated Activation of Ezrin is Associated with Cell Motility and Survival in a Subset of Lung Adenocarcinomas (ID 13005)

    16:45 - 18:00  |  Author(s): Jenny Zhao
    • Abstract

    Background
    

    Lung cancer is the leading cause of cancer mortality worldwide with 40% of cases diagnosed as lung adenocarcinoma (LADC). Increased expression of RET has been reported in 10-20% of LADCs and is associated with metastasis and reduced survival. Alternative gene splicing of RET generates two co-expressed isoforms; RET9 and RET51, with unique C-terminal tails. The tails confer distinct protein binding opportunities with signaling proteins, known as scaffolds, to promote different signaling events and cell processes. RET9 interacts with the SHANK family scaffolds at a PDZ-binding motif and Enigma at tyrosine (Y) 1062, whereas, RET51 interacts with Grb2 at the unique Y1096. RET9 and RET51 are associated with distinct functions in thyroid cancer, where RET51plays a predominant role in cancer metastasis. Ezrin is a scaffold protein and tyrosine kinase substrate that bridges the plasma membrane to the actin cytoskeleton to promote adhesion, organization and migration. Overexpression of ezrin is associated with invasion and metastasis of osteosarcoma and pancreatic cancer cells. The roles of RET isoform-scaffold interactions in the progression of LADC has never been investigated.

    a9ded1e5ce5d75814730bb4caaf49419 Method
    

    HEK-293 cells were co-transfected with RET isoform or mutant constructs and Ezrin and/or Grb2 dominant negative (DN) constructs. A549 LADC cells were transfected with RET51 construct. SH-SY5Y neuroblastoma and HCC1833 LADC cells endogenously express Ezrin and RET. Cells were treated with RET/ Ezrin inhibitors and/or stimulated with glial-derived neurotrophic factor (GDNF) then fixed or harvested. Co-immunoprecipitation (Co-IP) or pull-down assays coupled to western blotting were used to investigate binding domains required for RET interaction with and activation of Ezrin or downstream effectors. Immunofluorescence confocal imaging was used to determine co-localization of RET and Ezrin and changes to cell morphology. GDNF-impregnated agarose bead live-cell fluorescence assays were used to measure cell growth, survival, spread and motility. SynTarget (cancer gene expression analysis tool) was used to assess co-expression of RET and Ezrin with respect to clinicopathological parameters and survival outcomes in LADC.

    4c3880bb027f159e801041b1021e88e8 Result
    

    Ezrin binds kinase active RET51 with Grb2. GDNF-stimulation promotes the formation of cytoskeletal outgrowths with increased co-localization of Ezrin and RET. GDNF-activation of RET results in scaffold-mediated activation of Rho-family GTPases to promote cell motility. Like RET, overexpression of Ezrin is associated with poor overall survival in LADC patients and overexpression of both RET and Ezrin has a negative synergistic effect on overall survival in a subset of LADC patients.

    8eea62084ca7e541d918e823422bd82e Conclusion
    

    RET interaction with Ezrin identifies a distinct functional role of RET as an oncogenic driver in lung adenocarcinoma.

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