title: Spheroid Culture of Head and Neck Cancer Cells Reveals an Important Role of EGFR Signalling in Anchorage Independent Survival.
abstract: In solid tumours millions of cells are shed into the blood circulation each day. Only a subset of these circulating tumour cells (CTCs) survive, many of them presumable because of their potential to form multi-cellular clusters also named spheroids. Tumour cells within these spheroids are protected from anoikis, which allows them to metastasize to distant organs or re-seed at the primary site. We used spheroid cultures of head and neck squamous cell carcinoma (HNSCC) cell lines as a model for such CTC clusters for determining the role of the epidermal growth factor receptor (EGFR) in cluster formation ability and cell survival after detachment from the extra-cellular matrix. The HNSCC cell lines FaDu, SCC-9 and UT-SCC-9 (UT-SCC-9P) as well as its cetuximab (CTX)-resistant sub-clone (UT-SCC-9R) were forced to grow in an anchorage-independent manner by coating culture dishes with the anti-adhesive polymer poly-2-hydroxyethylmethacrylate (poly-HEMA). The extent of apoptosis, clonogenic survival and EGFR signalling under such culture conditions was evaluated. The potential of spheroid formation in suspension culture was found to be positively correlated with the proliferation rate of HNSCC cell lines as well as their basal EGFR expression levels. CTX and gefitinib blocked, whereas the addition of EGFR ligands promoted anchorage-independent cell survival and spheroid formation. Increased spheroid formation and growth were associated with persistent activation of EGFR and its downstream signalling component (MAPK/ERK). Importantly, HNSCC cells derived from spheroid cultures retained their clonogenic potential in the absence of cell-matrix contact. Addition of CTX under these conditions strongly inhibited colony formation in CTX-sensitive cell lines but not their resistant subclones. Altogether, EGFR activation was identified as crucial factor for anchorage-independent survival of HNSCC cells. Targeting EGFR in CTC cluster formation might represent an attractive anti-metastatic treatment approach in HNSCC.
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.|German Cancer Consortium (DKTK), Deutsches Krebsforschungszentrum (DKFZ), Partner Site Berlin, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Diana
authors.initials: D
authors.lastname: Braunholz
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.|German Cancer Consortium (DKTK), Deutsches Krebsforschungszentrum (DKFZ), Partner Site Berlin, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Mohammad
authors.initials: M
authors.lastname: Saki
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.|German Cancer Consortium (DKTK), Deutsches Krebsforschungszentrum (DKFZ), Partner Site Berlin, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Franziska
authors.initials: F
authors.lastname: Niehr
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Merve
authors.initials: M
authors.lastname: Öztürk
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Berta
authors.initials: B
authors.lastname: Borràs Puértolas
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.|German Cancer Consortium (DKTK), Deutsches Krebsforschungszentrum (DKFZ), Partner Site Berlin, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Robert
authors.initials: R
authors.lastname: Konschak
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Volker
authors.initials: V
authors.lastname: Budach
authors.affiliation: Translational Radiooncology and Radiobiology Research Laboratory, Department of Radiooncology and Radiotherapy, Charité University Hospital Berlin, Berlin, Germany.|German Cancer Consortium (DKTK), Deutsches Krebsforschungszentrum (DKFZ), Partner Site Berlin, Charité University Hospital Berlin, Berlin, Germany.
authors.forename: Ingeborg
authors.initials: I
authors.lastname: Tinhofer
mesh_terms: D023102:Anoikis; D002294:Carcinoma, Squamous Cell / Q000378:metabolism / Q000473:pathology*; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D066246:ErbB Receptors / Q000378:metabolism*; D006258:Head and Neck Neoplasms / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D015398:Signal Transduction; D000077195:Squamous Cell Carcinoma of Head and Neck
chemical_list: D066246:ErbB Receptors