S100A16 promotes differentiation and contributes to a less aggressive tumor phenotype in oral squamous cell carcinoma.
BACKGROUND
Altered expression of S100A16 has been reported in human cancers, but its biological role in tumorigenesis is not fully understood. This study aimed to investigate the clinical significance and functional role of S100A16 in oral squamous cell carcinoma (OSCC) suppression.


METHODS
S100A16 mRNA and/or protein levels were examined by quantitative RT-PCR and immunohistochemistry in whole- and laser microdissected-specimens of normal human oral mucosa (NHOM, n = 65), oral dysplastic lesions (ODL, n = 21), OSCCs (n = 132) and positive cervical nodes (n = 17). S100A16 protein expression in OSCC was examined for correlations with clinicopathological variables and patient survival. S100A16 was over-expressed and knocked-down in OSCC-derived (CaLH3 and H357) cells by employing retroviral constructs to investigate its effects on cell proliferation, sphere formation and three dimensional (3D)-organotypic invasive abilities in vitro and tumorigenesis in a mouse xenograft model.


RESULTS
Both S100A16 mRNA and protein levels were found to be progressively down-regulated from NHOM to ODL and OSCC. Low S100A16 protein levels in OSCC significantly correlated with reduced 10-year overall survival and poor tumor differentiation. Analysis of two external OSCC microarray datasets showed a positive correlation between the mRNA expression levels of S100A16 and keratinocyte differentiation markers. CaLH3 and H357 cell fractions enriched for differentiated cells either by lack of adherence to collagen IV or FACS sorting for low p75NTR expression expressed significantly higher S100A16 mRNA levels than the subpopulations enriched for less differentiated cells. Corroborating these findings, retroviral mediated S100A16 over-expression and knock-down in CaLH3 and H357 cells led to respective up- and down-regulation of differentiation markers. In vitro functional studies showed significant reduction in cell proliferation, sphere formation and 3D-invasive abilities of CaLH3 and H357 cells upon S100A16 over-expression. These functional effects were associated with concomitant down-regulation of self-renewal (Bmi-1 and Oct 4A) and invasion related (MMP1 and MMP9) molecules. S100A16 over-expression also suppressed tumorigenesis of H357 cells in a mouse xenograft model and the resulting tumor xenografts displayed features/expression of increased differentiation and reduced proliferation/self-renewal.


CONCLUSIONS
These results indicate that S100A16 is a differentiation promoting protein and might function as a tumor suppressor in OSCC.
Department of Clinical Medicine, The Gade Laboratory for Pathology, University of Bergen, Haukeland University Hospital, N-5021, Bergen, Norway. Dipak.Sapkota@k1.uib.no.|Centre for Cancer Biomarkers (CCBIO), Faculty of Medicine and Dentistry, University of Bergen, N-5021, Bergen, Norway. Dipak.Sapkota@k1.uib.no.
Dipak
D
Sapkota
Center of Medical Genetics and Molecular Medicine, Haukeland University Hospital, University of Bergen, N-5021, Bergen, Norway. ove.bruland@helse-bergen.no.
Ove
O
Bruland
Department of Clinical Medicine, The Gade Laboratory for Pathology, University of Bergen, Haukeland University Hospital, N-5021, Bergen, Norway. Himalaya.Parajuli@k1.uib.no.|Centre for Cancer Biomarkers (CCBIO), Faculty of Medicine and Dentistry, University of Bergen, N-5021, Bergen, Norway. Himalaya.Parajuli@k1.uib.no.
Himalaya
H
Parajuli
Department of Clinical Medicine, The Gade Laboratory for Pathology, University of Bergen, Haukeland University Hospital, N-5021, Bergen, Norway. Tarig.Osman@k1.uib.no.|Centre for Cancer Biomarkers (CCBIO), Faculty of Medicine and Dentistry, University of Bergen, N-5021, Bergen, Norway. Tarig.Osman@k1.uib.no.
Tarig A
TA
Osman
Centre for Clinical and Diagnostic Oral Sciences, Institute of Dentistry, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, England, UK. m.t.teh@qmul.ac.uk.
Muy-Teck
MT
Teh
Department of Clinical Medicine, The Gade Laboratory for Pathology, University of Bergen, Haukeland University Hospital, N-5021, Bergen, Norway. Anne.Johannessen@k1.uib.no.|Centre for Cancer Biomarkers (CCBIO), Faculty of Medicine and Dentistry, University of Bergen, N-5021, Bergen, Norway. Anne.Johannessen@k1.uib.no.|Department of Pathology, Haukeland University Hospital, Bergen, Norway. Anne.Johannessen@k1.uib.no.
Anne C
AC
Johannessen
Department of Clinical Medicine, The Gade Laboratory for Pathology, University of Bergen, Haukeland University Hospital, N-5021, Bergen, Norway. Daniela.Costea@k1.uib.no.|Centre for Cancer Biomarkers (CCBIO), Faculty of Medicine and Dentistry, University of Bergen, N-5021, Bergen, Norway. Daniela.Costea@k1.uib.no.|Department of Pathology, Haukeland University Hospital, Bergen, Norway. Daniela.Costea@k1.uib.no.
Daniela Elena
DE
Costea
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D001706:Biopsy; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000378:metabolism; D049109:Cell Proliferation / Q000235:genetics; D005260:Female; D005822:Genetic Vectors / Q000378:metabolism; D064593:Heterografts; D006801:Humans; D008297:Male; D051379:Mice; D008875:Middle Aged; D023421:Models, Animal; D009061:Mouth Mucosa / Q000473:pathology; D009062:Mouth Neoplasms / Q000235:genetics* / Q000378:metabolism; D009361:Neoplasm Invasiveness / Q000235:genetics; D010641:Phenotype; D060888:Real-Time Polymerase Chain Reaction; D012190:Retroviridae / Q000378:metabolism; D009418:S100 Proteins / Q000378:metabolism*
D009418:S100 Proteins; C000589580:S100A16 protein, human