Author of

• 17629

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  P2.03b - Poster Session with Presenters Present (ID 465)

  • Event: WCLC 2016
  • Type: Poster Presenters Present
  • Track: Advanced NSCLC
  • Presentations: 2
  • Moderators:
  • Coordinates: 12/06/2016, 14:30 - 15:45, Hall B (Poster Area)

  • 17699

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    P2.03b-070 - Establishment of Organoid Cell Lines from Lung Squamous Cell Carcinoma (ID 5362)

    14:30 - 14:30  |  Author(s): R. Shi
    • Abstract

    Background:
    The limited number of available lung squamous cell carcinoma (LUSC) cell lines poses significant challenge for biological, experimental therapeutic and biomarker research in LUSC. Novel approaches to establish new preclinical models are urgently needed. We have previously established patient-derived xenografts (PDX) from resected tumours of LUSC patients and characterized them on the genomic, transcriptomic, and proteomic levels. We have used these PDX models to develop a method for establishment of 3D organoid cultures and cell lines as new in vitro preclinical models of LUSC.
    
    Methods:
    PDX models were established and propagated from resected primary non-small cell lung cancer (NSCLC) in NOD/SCID mice; they were molecularly profiled by exome sequencing, SNP array for copy number analysis, and immunohistochemistry (IHC). PDX tissue harvested from mice was dissociated into single cells and plated in 100% matrigel dome, with overlaying media on top. Organoids were characterized by H&E, and IHC of p63, CK5/6, TTF-1, and CK7. Organoid growth rate and drug screening were assessed using Celltiter glo cell viability assay.
    
    Results:
    A total of 17 LUSC PDX models have been used for this study. All organoids were able to initiate in culture at passage 1, and the organoid establishment rate (beyond passage 4) is 50% (6/12). 4/12 (33%) LUSC organoids were able to be propagated beyond 10 passages for over 60 days with an average doubling rate of 2-3 days. Organoid tumour cells recapitulated the histological features of LUSC and were positive for p63 and CK5/6, and negative for TTF-1 and CK7 by IHC. Molecular characterization of LUSC PDX models revealed PIK3CA mutations, amplifications, and PTEN loss. Over 40% (4/9) of PI3K altered LUSC organoids were sensitive to PI3K inhibitor BKM120.
    
    Conclusion:
    LUSC organoids can be established for long term culture and recapitulate the phenotypic features of the PDX. The culture protocol is currently being tested on primary patient LUSC tumours. Organoid cultures and cell lines may be useful as additional preclinical models for functional validation of novel therapeutic targets in LUSC.
    
    

  • 17700

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    P2.03b-071 - Therapeutic Targeting of the Phosphatidylinositol-3 Kinase Pathway in Lung Squamous Cell Carcinoma (ID 5369)

    14:30 - 14:30  |  Author(s): R. Shi
    • Abstract

    Background:
    The phosphatidylinositol-3 kinase (PI3K) belongs to a family of lipid kinases involved in the regulation of cell proliferation and survival and is often dysregulated in cancer. Comprehensive molecular profiling by The Cancer Genome Atlas (TCGA) has identified PIK3CA mutations, amplifications, and the tumor suppressor PTEN loss in 30-40% of lung squamous cell carcinoma (LUSC) patients. Inhibitors of PI3K such as BKM120 have been initiated in BASALT-1 trial (NCT01820325) of PI3K activated LUSC, however with modest response rate (40% of patients with stable disease and 3.3% with partial response). We aim to assess the efficacy of PI3K inhibition in LUSC patient-derived xenografts (PDX) harboring different PI3K pathway alterations to identify potential mechanisms of innate resistance.
    
    Methods:
    PDX models were established from early stage LUSC patients and molecularly characterized via exome sequencing, SNP array for copy number variation (CNV) and gene expression analysis. PIK3CA mutations were validated by direct sequencing, amplifications by fluorescence in situ hybridization (FISH), and PTEN loss by immunohistochemistry (IHC). For in vivo drug screening, each PDX model was implanted in two mice; one treated with BKM120 (50mg/kg) and the other with vehicle control by daily oral gavage. Tumors were monitored twice weekly with caliper measurement. A responder is a tumor that regresses completely, shrinks more than 30%, or remains a stable size according to the RECIST criteria.
    
    Results:
    Of the 75 LUSC PDX models that our laboratory has established, 11 (14%) harbored PIK3CA E545K and E542K mutations, 36 (47%) harbored PIK3CA amplifications, and 23 (30%) showed loss of PTEN protein expression. Using the RECIST criteria, BKM120 screening in selected PDX models revealed stable disease and progressive disease in 4/9 (46%) and 5/9 (54%) of the PDX models, respectively, after 21 days of treatment. Of the 9 PDX models tested, 3/5 PIK3CA mutant models were responsive to BKM120, whereas none of the other 4 PIK3CA amplified and/or PTEN deleted models were responsive to BKM120. Additionally, downregulation of pErk1/2 and pS6 in a responder model and no change in phosphorylated proteins in non-responding models were observed. Pharmacodynamics studies, validation of responders with more mouse replicates, and testing on the remaining models are ongoing and the results will be reported.
    
    Conclusion:
    60% of LUSC PDXs with PIK3CA mutation demonstrate high sensitivity to pan-PI3K inhibitor. Understanding innate resistance mechanisms of PI3K inhibition may provide important insights on tractable targets and therapeutic strategy for LUSC patients with aberrant PI3K pathway.