CD44 down-regulation reduces self-renewal of oral carcinoma stem cells

Background: The CD44 adhesion molecule acts as a marker of “tumour initiating cells”, the putative “cancer stem cells” (CSCs) in oral squamous cell carcinomas (OSCC), and there is increasing evidence that such cells drive the growth of a wide range of malignancies. However, little is known about the role of CD44 in CSC maintenance or about differential expression of CD44 splice variants on CSCs. Objective: To investigate the function of CD44 in maintaining the self-renewal properties of CSCs in cell lines derived from OSCC. Materials and Methods: CD44 expression was down-regulated in the UK1, 5PT, Ca1, and H357 cell lines by butyrate or by siRNAs. Down regulation was confirmed by immunofluorescence, flow cytometry and QPCR. The effects of CD44 down-regulation were assessed by quantification of stem cell markers (Oct4 and Bmi1) and assays of growth and clonogenicity. Results: For all cell lines, both 1mM sodium butyrate treatment and siRNAs against CD44 resulted in a significant decrease of CD44 at both protein and mRNA levels. CD44 down-regulation reduced expression of mRNAs for Oct4 and Bmi1, decreased in cell yield (p<0.01), and altered colony forming patterns with a significant (p<0.01 ) decrease in holoclones, the colony type containing CSCs. Conclusion: CD44 appears to have a functional role in maintaining the self-renewal of CSCs in OSCC-derived cell lines. In addition to bringing novel knowledge about the role of CD44 in maintenance of the stem cell properties, the present study highlights the possibility of developing novel and more efficient treatment strategies for oral cancer by depletion of stem cell populations through down regulation of CD44. Work continues to examine whether differential expression of CD44 isoforms on CSCs can be exploited to enable specific targeting of malignant stem cell populations.

Supported by the Fanconi Anemia Research Foundation and the Norwegian Research Council.