title: Molecular characterization of chordoma xenografts generated from a novel primary chordoma cell source and two chordoma cell lines.
abstract: OBJECTIVE
  Chordoma cells can generate solid-like tumors in xenograft models that express some molecular characteristics of the parent tumor, including positivity for brachyury and cytokeratins. However, there is a dearth of molecular markers that relate to chordoma tumor growth, as well as the cell lines needed to advance treatment. The objective in this study was to isolate a novel primary chordoma cell source and analyze the characteristics of tumor growth in a mouse xenograft model for comparison with the established U-CH1 and U-CH2b cell lines.
  
  
  METHODS
  Primary cells from a sacral chordoma, called "DVC-4," were cultured alongside U-CH1 and U-CH2b cells for more than 20 passages and characterized for expression of CD24 and brachyury. While brachyury is believed essential for driving tumor formation, CD24 is associated with healthy nucleus pulposus cells. Each cell type was subcutaneously implanted in NOD/SCID/IL2Rγ(null) mice. The percentage of solid tumors formed, time to maximum tumor size, and immunostaining scores for CD24 and brachyury (intensity scores of 0-3, heterogeneity scores of 0-1) were reported and evaluated to test differences across groups.
  
  
  RESULTS
  The DVC-4 cells retained chordoma-like morphology in culture and exhibited CD24 and brachyury expression profiles in vitro that were similar to those for U-CH1 and U-CH2b. Both U-CH1 and DVC-4 cells grew tumors at rates that were faster than those for U-CH2b cells. Gross tumor developed at nearly every site (95%) injected with U-CH1 and at most sites (75%) injected with DVC-4. In contrast, U-CH2b cells produced grossly visible tumors in less than 50% of injected sites. Brachyury staining was similar among tumors derived from all 3 cell types and was intensely positive (scores of 2-3) in a majority of tissue sections. In contrast, differences in the pattern and intensity of staining for CD24 were noted among the 3 types of cell-derived tumors (p < 0.05, chi-square test), with evidence of intense and uniform staining in a majority of U-CH1 tumor sections (score of 3) and more than half of the DVC-4 tumor sections (scores of 2-3). In contrast, a majority of sections from U-CH2b cells stained modestly for CD24 (scores of 1-2) with a predominantly heterogeneous staining pattern.
  
  
  CONCLUSIONS
  This is the first report on xenografts generated from U-CH2b cells in which a low tumorigenicity was discovered despite evidence of chordoma-like characteristics in vitro. For tumors derived from a primary chordoma cell and U-CH1 cell line, similarly intense staining for CD24 was observed, which may correspond to their similar potential to grow tumors. In contrast, U-CH2b tumors stained less intensely for CD24. These results emphasize that many markers, including CD24, may be useful in distinguishing among chordoma cell types and their tumorigenicity in vivo.
authors.affiliation: Department of Surgery, Division of Neurosurgery, and 
authors.forename: Isaac O
authors.initials: IO
authors.lastname: Karikari
authors.forename: Christopher L
authors.initials: CL
authors.lastname: Gilchrist
authors.forename: Liufang
authors.initials: L
authors.lastname: Jing
authors.forename: David A
authors.initials: DA
authors.lastname: Alcorta
authors.forename: Jun
authors.initials: J
authors.lastname: Chen
authors.forename: William J
authors.initials: WJ
authors.lastname: Richardson
authors.forename: Mostafa A
authors.initials: MA
authors.lastname: Gabr
authors.forename: Richard D
authors.initials: RD
authors.lastname: Bell
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Kelley
authors.forename: Carlos A
authors.initials: CA
authors.lastname: Bagley
authors.forename: Lori A
authors.initials: LA
authors.lastname: Setton
mesh_terms: D000368:Aged; D000818:Animals; D014408:Biomarkers, Tumor / Q000378:metabolism*; D051927:CD24 Antigen / Q000378:metabolism; D045744:Cell Line, Tumor; D002817:Chordoma / Q000378:metabolism* / Q000473:pathology*; D004195:Disease Models, Animal; D005326:Fetal Proteins / Q000378:metabolism; D005434:Flow Cytometry; D064593:Heterografts; D006801:Humans; D007150:Immunohistochemistry; D007633:Keratins / Q000378:metabolism; D008297:Male; D051379:Mice; D016133:Polymerase Chain Reaction; D012447:Sacrum / Q000473:pathology*; D020825:T-Box Domain Proteins / Q000378:metabolism; D014407:Tumor Cells, Cultured; D000098908:Brachyury Protein
chemical_list: D014408:Biomarkers, Tumor; D051927:CD24 Antigen; D005326:Fetal Proteins; D007633:Keratins; D020825:T-Box Domain Proteins; D000098908:Brachyury Protein
keywords: BSA = bovine serum albumin; CD24; FBS = fetal bovine serum; FITC = fluorescein isothiocyanate; IMDM = Iscove's modified Dulbecco's medium; MFI = mean fluorescence intensity; NSG = NOD/SCID/IL2Rγnull; PBS = phosphate-buffered saline; PCR = polymerase chain reaction; U-CH1; U-CH2b; brachyury; chordoma; oncology; xenograft