title: Evaluation of stem-like side population cells in a recurrent nasopharyngeal carcinoma cell line.
abstract: BACKGROUND
  Side population (SP) assay identifies cells with dye/drug extrusion ability, a characteristic of stem cells. Here, we determined if SP cells exist in a verified cell line originating from recurrent nasopharyngeal carcinoma (NPC) and a xenograft established from recurrent metastatic NPC. These cells were evaluated for stem-like properties via functional assays as well as for tumourigenicity.
  
  
  METHODS
  We used Hoechst 33342 to identify the SP from non-SP (NSP) cells in HK1 NPC cell line and xeno-284 NPC xenograft. The cells were assayed for in vitro characteristics of cancer stem cells (CSC), gene expression and tumourigenicity ability. Student's t test was used to test for significance.
  
  
  RESULTS
  Five to ten percent and less than 0.5% of HK1 and xeno-284 NPC cells, respectively, were SP cells. Fumitremorgin C (FTC), as opposed to verapamil, was effective in causing the cells to retain Hoechst 33342 dye. HK1 SP cells formed more holoclones, had more aldehyde dehydrogenase (ALDH) activity, divided asymmetrically and contained slow-proliferating cells. ABCG2, SOX2, TERT, MYC, Hedgehog, Notch, TGFβ and Wnt signalling pathway genes were significantly upregulated in the SP cells. However, despite these differences in vitro, both HK1 SP and NSP cells had an overall similar tumourigenic potential in vivo.
  
  
  CONCLUSIONS
  HK1 SP cells were ABCG2-specific as confirmed by FTC inhibition and gene expression data. Despite data from in vitro and gene expression experiments suggesting stem-like features, there was no significant difference in tumourigenic potential between SP and NSP cells. We conclude that SP assay alone is not sufficient to identify CSCs in HK1 cells. Our work also suggests the presence of a stem-cell like population among NPC cells which do not display increased tumourigenicity.
authors.affiliation: Molecular Pathology Unit, Cancer Research Centre, Institute for Medical Research, Jalan Pahang, 50588 Kuala Lumpur, Malaysia ; Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: Susan Ling Ling
authors.initials: SL
authors.lastname: Hoe
authors.affiliation: Molecular Pathology Unit, Cancer Research Centre, Institute for Medical Research, Jalan Pahang, 50588 Kuala Lumpur, Malaysia.
authors.forename: Lu Ping
authors.initials: LP
authors.lastname: Tan
authors.affiliation: Molecular Pathology Unit, Cancer Research Centre, Institute for Medical Research, Jalan Pahang, 50588 Kuala Lumpur, Malaysia ; Current address: Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: Juliana
authors.initials: J
authors.lastname: Jamal
authors.affiliation: Faculty of Medical Sciences, UCSI University, 1 Jalan Menara Gading, UCSI Heights, 56000 Cheras, Malaysia.
authors.forename: Suat Cheng
authors.initials: SC
authors.lastname: Peh
authors.affiliation: Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: Ching Ching
authors.initials: CC
authors.lastname: Ng
authors.affiliation: Genome Institute of Singapore, 60 Biopolis Street, #02-01, Genome, 138672 Singapore.
authors.forename: Wen Cai
authors.initials: WC
authors.lastname: Zhang
authors.affiliation: Molecular Pathology Unit, Cancer Research Centre, Institute for Medical Research, Jalan Pahang, 50588 Kuala Lumpur, Malaysia.
authors.forename: Munirah
authors.initials: M
authors.lastname: Ahmad
authors.affiliation: Molecular Pathology Unit, Cancer Research Centre, Institute for Medical Research, Jalan Pahang, 50588 Kuala Lumpur, Malaysia.
authors.forename: Alan Soo Beng
authors.initials: AS
authors.lastname: Khoo
keywords: Cancer stem cells; Nasopharyngeal carcinoma; Side population; Slow-cycling; Stem-like