title: Genetic and epigenetic analysis of putative breast cancer stem cell models.
abstract: BACKGROUND
  Cancer stem cell model hypothesizes existence of a small proportion of tumor cells capable of sustaining tumor formation, self-renewal and differentiation. In breast cancer, these cells were found to be associated with CD44⁺CD24-low and ALDH⁺ phenotype. Our study was performed to evaluate the suitability of current approaches for breast cancer stem cell analyses to evaluate heterogeneity of breast cancer cells through their extensive genetic and epigenetic characterization.
  
  
  METHODS
  Breast cancer cell lines MCF7 and SUM159 were cultured in adherent conditions and as mammospheres. Flow cytometry sorting for CD44, CD24 and ALDH was performed. Sorted and unsorted populations, mammospheres and adherent cell cultures were subjected to DNA profiling by array CGH and methylation profiling by Epitect Methyl qPCR array. Methylation status of selected genes was further evaluated by pyrosequencing. Functional impact of methylation was evaluated by mRNA analysis for selected genes.
  
  
  RESULTS
  Array CGH did not reveal any genomic differences. In contrast, putative breast cancer stem cells showed altered methylation levels of several genes compared to parental tumor cells.
  
  
  CONCLUSIONS
  Our results underpin the hypothesis that epigenetic mechanisms seem to play a major role in the regulation of CSCs. However, it is also clear that more efficient methods for CSC enrichment are needed. This work underscores requirement of additional approaches to reveal heterogeneity within breast cancer.
authors.affiliation: Division of Oncology, Department of Internal Medicine, Medical University of Graz, Auenbruggerplatz 15, A-8036 Graz, Austria.
authors.forename: Marija
authors.initials: M
authors.lastname: Balic
authors.forename: Daniela
authors.initials: D
authors.lastname: Schwarzenbacher
authors.forename: Stefanie
authors.initials: S
authors.lastname: Stanzer
authors.forename: Ellen
authors.initials: E
authors.lastname: Heitzer
authors.forename: Martina
authors.initials: M
authors.lastname: Auer
authors.forename: Jochen B
authors.initials: JB
authors.lastname: Geigl
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Cote
authors.forename: Ram H
authors.initials: RH
authors.lastname: Datar
authors.forename: Nadia
authors.initials: N
authors.lastname: Dandachi
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology*; D051927:CD24 Antigen / Q000032:analysis / Q000378:metabolism; D045744:Cell Line, Tumor; D055028:Comparative Genomic Hybridization; D019175:DNA Methylation / Q000235:genetics*; D044127:Epigenesis, Genetic; D005260:Female; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D014411:Neoplastic Stem Cells; D060888:Real-Time Polymerase Chain Reaction; D059467:Transcriptome
chemical_list: D051927:CD24 Antigen