title: Anchorage-independent growth of Ewing sarcoma cells under serum-free conditions is not associated with stem-cell like phenotype and function.
abstract: Novel treatment strategies for Ewing sarcoma aim to eliminate residual tumor cells that have maintained the capacity to reinitiate tumor growth after intensive conventional therapy. Preclinical models that more closely mimic in vivo tumor growth than standard monolayer cultures are needed. Sphere formation under anchorage-independent, serum-free conditions has been proposed to enrich for cells with tumor-initiating, stem cell-like properties in various solid cancers. In the present study, we assessed the phenotype and functional stem cell characteristics of Ewing sarcoma spheres. Spheres were generated under serum-free culture conditions from four Ewing sarcoma cell lines and four relapse tumor biopsies. Standard monolayer cultures were established as controls. Median levels of surface expression of the Ewing sarcoma marker CD99 as well as the supposed stem cell marker CD133 and the neural crest marker CD57 were comparable between spheres and monolayers. Ewing sarcoma spheres from individual tumors failed to continuously self-renew by secondary sphere formation. They contained variable proportions of side populations (SPs). Sphere culture did not enhance the in vivo tumorigenicity of Ewing sarcoma cells in a murine xenograft model. We conclude that sphere formation under serum-free conditions is not a reliable tool to enrich for cells with stem cell characteristics in Ewing sarcoma. By mimicking the anchorage-independent, multicellular growth of Ewing sarcoma micrometastases, in vitro sphere growth may still add value as a preclinical tool to evaluate the efficacy of novel therapeutics.
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Leuchte
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Bianca
authors.initials: B
authors.lastname: Altvater
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Simeon
authors.initials: S
authors.lastname: Hoffschlag
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Jenny
authors.initials: J
authors.lastname: Potratz
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Jutta
authors.initials: J
authors.lastname: Meltzer
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Dagmar
authors.initials: D
authors.lastname: Clemens
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Andrea
authors.initials: A
authors.lastname: Luecke
authors.affiliation: Department of Orthopedic Surgery, University Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Jendrik
authors.initials: J
authors.lastname: Hardes
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Uta
authors.initials: U
authors.lastname: Dirksen
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Heribert
authors.initials: H
authors.lastname: Juergens
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Sareetha
authors.initials: S
authors.lastname: Kailayangiri
authors.affiliation: Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, D-48149 Muenster, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Rossig
mesh_terms: D000818:Animals; D014408:Biomarkers, Tumor / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D045744:Cell Line, Tumor; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D006801:Humans; D051379:Mice; D016688:Mice, Inbred NOD; D009362:Neoplasm Metastasis; D009374:Neoplasms, Experimental; D014411:Neoplastic Stem Cells / Q000378:metabolism / Q000473:pathology*; D012512:Sarcoma, Ewing / Q000473:pathology*; D018874:Spheroids, Cellular / Q000378:metabolism*; D014407:Tumor Cells, Cultured
chemical_list: D014408:Biomarkers, Tumor; D016895:Culture Media, Serum-Free