title: Colorectal cancer derived organotypic spheroids maintain essential tissue characteristics but adapt their metabolism in culture.
abstract: BACKGROUND
  Organotypic tumor spheroids, a 3D in vitro model derived from patient tumor material, preserve tissue heterogeneity and retain structural tissue elements, thus replicating the in vivo tumor more closely than commonly used 2D and 3D cell line models. Such structures harbour tumorigenic cells, as revealed by xenograft implantation studies in animal models and maintain the genetic makeup of the original tumor material. The aim of our work was a morphological and proteomic characterization of organotypic spheroids derived from colorectal cancer tissue in order to get insight into their composition and associated biology.
  
  
  RESULTS
  Morphological analysis showed that spheroids were of about 250 μm in size and varied in structure, while the spheroid cells differed in shape and size and were tightly packed together by desmosomes and tight junctions. Our proteomic data revealed significant alterations in protein expression in organotypic tumor spheroids cultured as primary explants compared to primary colorectal cancer tissue. Components underlying cellular and tissue architecture were changed; nuclear DNA/ chromatin maintenance systems were up-regulated, whereas various mitochondrial components were down-regulated in spheroids. Most interestingly, the mesenchymal cells appear to be substantial component in such cellular assemblies. Thus the observed changes may partly occur in this cellular compartment. Finally, in the proteomics analysis stem cell-like characteristics were observed within the spheroid cellular assembly, reflected by accumulation of Alcam, Ctnnb1, Aldh1, Gpx2, and CD166. These findings were underlined by IHC analysis of Ctnnb1, CD24 and CD44, therefore warranting closer investigation of the tumorigenic compartment in this 3D culture model for tumor tissue.
  
  
  CONCLUSIONS
  Our analysis of organotypic CRC tumor spheroids has identified biological processes associated with a mixture of cell types and states, including protein markers for mesenchymal and stem-like cells. This 3D tumor model in which tumor heterogeneity is preserved may represent an advantageous model system to investigate novel therapeutic approaches.
authors: NorLux Neuro-Oncology Laboratory, Department of Oncology, CRP-Santé, Luxembourg, Luxembourg ; Department of Research and Development, Blood Transfusion Center of Slovenia, Ljubljana, Slovenia ; Institute of Pathology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia.
authors: Uros
authors: U
authors: Rajcevic
authors: OncoProteomics Laboratory, Department of Medical Oncology, VUmc-Cancer Center Amsterdam, VU University Medical Center, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands.
authors: Jaco C
authors: JC
authors: Knol
authors: OncoProteomics Laboratory, Department of Medical Oncology, VUmc-Cancer Center Amsterdam, VU University Medical Center, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands.
authors: Sander
authors: S
authors: Piersma
authors: NorLux Neuro-Oncology Laboratory, Department of Oncology, CRP-Santé, Luxembourg, Luxembourg.
authors: Sébastien
authors: S
authors: Bougnaud
authors: NorLux Neuro-Oncology Laboratory, Department of Oncology, CRP-Santé, Luxembourg, Luxembourg.
authors: Fred
authors: F
authors: Fack
authors: Department of Biomedicine, University of Bergen, Bergen, Norway.
authors: Eirik
authors: E
authors: Sundlisaeter
authors: Department of Surgery, Haraldsplass Deaconal Hospital, University of Bergen, Bergen, Norway.
authors: Karl
authors: K
authors: Søndenaa
authors: Haukeland University Hospital, Bergen, Norway.
authors: Reidar
authors: R
authors: Myklebust
authors: OncoProteomics Laboratory, Department of Medical Oncology, VUmc-Cancer Center Amsterdam, VU University Medical Center, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands.
authors: Thang V
authors: TV
authors: Pham
authors: NorLux Neuro-Oncology Laboratory, Department of Oncology, CRP-Santé, Luxembourg, Luxembourg.
authors: Simone P
authors: SP
authors: Niclou
authors: OncoProteomics Laboratory, Department of Medical Oncology, VUmc-Cancer Center Amsterdam, VU University Medical Center, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands.
authors: Connie R
authors: CR
authors: Jiménez
keywords: 3D cell cultures; Bioinformatics; Cancer biomarkers; Cancer stem cells; Colorectal cancer; Label-free proteomics; Mass spectrometry; Proteomics; Tissue spheroids