title: Glioblastoma-derived spheroid cultures as an experimental model for analysis of EGFR anomalies.
abstract: Glioblastoma cell cultures in vitro are frequently used for investigations on the biology of tumors or new therapeutic approaches. Recent reports have emphasized the importance of cell culture type for maintenance of tumor original features. Nevertheless, the ability of GBM cells to preserve EGFR overdosage in vitro remains controversial. Our experimental approach was based on quantitative analysis of EGFR gene dosage in vitro both at DNA and mRNA level. Real-time PCR data were verified with a FISH method allowing for a distinction between EGFR amplification and polysomy 7. We demonstrated that EGFR amplification accompanied by EGFRwt overexpression was maintained in spheroids, but these phenomena were gradually lost in adherent culture. We noticed a rapid decrease of EGFR overdosage already at the initial stage of cell culture establishment. In contrast to EGFR amplification, the maintenance of polysomy 7 resulted in EGFR locus gain and stabilization even in long-term adherent culture in serum presence. Surprisingly, the EGFRwt expression pattern did not reflect the latter phenomenon and we observed no overexpression of the tested gene. Moreover, quantitative analysis demonstrated that expression of the truncated variant of receptor-EGFRvIII was preserved in GBM-derived spheroids at a level comparable to the initial tumor tissue. Our findings are especially important in the light of research using glioblastoma culture as the experimental model for testing novel EGFR-targeted therapeutics in vitro, with special emphasis on the most common mutated form of receptor-EGFRvIII.
authors.affiliation: Department of Molecular Pathology and Neuropathology, Medical University of Lodz, Czechoslowacka 8/10 str, 92-216 Lodz, Poland. monika.witusik-perkowska@umed.lodz.pl
authors.forename: Monika
authors.initials: M
authors.lastname: Witusik-Perkowska
authors.forename: Piotr
authors.initials: P
authors.lastname: Rieske
authors.forename: Krystyna
authors.initials: K
authors.lastname: Hułas-Bigoszewska
authors.forename: Magdalena
authors.initials: M
authors.lastname: Zakrzewska
authors.forename: Robert
authors.initials: R
authors.lastname: Stawski
authors.forename: Dominika
authors.initials: D
authors.lastname: Kulczycka-Wojdala
authors.forename: Michał
authors.initials: M
authors.lastname: Bieńkowski
authors.forename: Ewelina
authors.initials: E
authors.lastname: Stoczyńska-Fidelus
authors.forename: Sylwia M
authors.initials: SM
authors.lastname: Grešner
authors.forename: Sylwester
authors.initials: S
authors.lastname: Piaskowski
authors.forename: Dariusz J
authors.initials: DJ
authors.lastname: Jaskólski
authors.forename: Wielisław
authors.initials: W
authors.lastname: Papierz
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Zakrzewski
authors.forename: Maciej
authors.initials: M
authors.lastname: Kolasa
authors.forename: James W
authors.initials: JW
authors.lastname: Ironside
authors.forename: Paweł P
authors.initials: PP
authors.lastname: Liberski
mesh_terms: D000818:Animals; D001932:Brain Neoplasms / Q000473:pathology*; D001973:Bromodeoxyuridine / Q000378:metabolism; D002448:Cell Adhesion / Q000502:physiology; D002453:Cell Cycle / Q000502:physiology; D049109:Cell Proliferation; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D005909:Glioblastoma / Q000473:pathology*; D006801:Humans; D023421:Models, Animal; D012333:RNA, Messenger / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000378:metabolism; D018874:Spheroids, Cellular / Q000473:pathology; D013997:Time Factors; D014407:Tumor Cells, Cultured
chemical_list: D012333:RNA, Messenger; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; C410723:epidermal growth factor receptor VIII; C512478:EGFR protein, human; D066246:ErbB Receptors; D001973:Bromodeoxyuridine
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