SSEA-3 as a novel amplifying cancer cell surface marker in colorectal cancers

Findings from studies on stem cells have been applied to cancer stem cell (CSC) research, but little is known about the relationship between ES cell-related cell surface markers and CSCs. In this study, we focused on stage-specific embryonic antigen 3 (SSEA-3), a marker of mesenchymal stem cells and Muse cells in colorectal cancer (CRC). Expression of SSEA-3 in human CRC cell lines and clinical specimens, specifically the relationship of SSEA-3 expression and the representative CSC markers (CD44, CD166, ALDH, CD24 and CD26) as well as with mesenchymal stem cell/Muse cell marker (CD105) were assessed. To characterize SSEA-3^-expressing cells, tumorigenicity, sphere formation ability, expression of iPS genes (Oct4, NANOG, SOX2 and c-Myc), cell proliferation and cell cycle status were assessed. SSEA-3 expression was identified in Caco-2, DLD-1, HT-29, SW480 and HCT116, but not in CaR-1 cells. No significant relationship between SSEA-3 and other stem cell markers was detected. SSEA-3⁺ cells showed increased tumorigenicity in vivo, but lower sphere formation ability in vitro than SSEA-3^-. iPS gene expression was not correlated with SSEA-3 expression status. SSEA-3⁺ cells showed higher proliferative ability than SSEA-3^- through enhanced cell cycles by decreased expression of p21^Cip1/Waf1 and p27^Kip1. Immunofluorescence analysis in clinical specimens indicated that expression of SSEA-3 is limited to stromal cells in normal mucosa but broad in poorly differentiated adenocarcinoma. These observations indicated that SSEA-3⁺ cells in CRC have immature phenotype but decreased self-renewal ability and may function as tumor transient amplifying cells or delayed contributing tumor-initiating cells.