Identification of Endothelin‐1 and NR4A2 as CD133‐regulated genes in colon cancer cells

Abstract

Several in vitro assays have been proposed to identify cancer stem cells (CSCs), including immunophenotyping, sphere assay and side population (SP) assay. CD133 antigen has been proposed as a CSC marker in colon cancer (CC). However, no functional data are available to date and conflicting results have been reported regarding its role as true CSC marker. Here we set out to identify a molecular signature associated with potential CSC. CD133(+) cells isolated from the CaCo-2 CC cell line were analysed by microarray molecular profiling compared to CD133(-) counterparts. Various differentially expressed genes were identified and the most relevant transcripts found to be over-expressed in CD133(+) cells were evaluated by quantitative RT-PCR in the CD133(+) fractions isolated from several CC cell lines. In the attempt to find a correlation between putative CSCs, isolated by means of CD133 immunophenotyping and the SP approach, we demonstrated a significant enrichment of CD133(+) cells within the SP fraction of CC cells, and comparison of the gene expression profiles revealed that Endothelin-1 (END-1) and nuclear receptor subfamily 4, group A, member 2 (NR4A2) transcripts are highly expressed in both CD133(+) and SP fractions of CC cells. Moreover, depletion of CD133 by siRNA induced a significant attenuation of END-1 and NR4A2 expression levels in CaCo-2 cells, while expression of all three molecules decreased during sodium butyrate-induced differentiation. In conclusion, we have identified a molecular signature associated with potential CSCs and showed for the first time the existence of a functional relationship between CD133, END-1 and NR4A2 expression in colon cancer cells.

Increased expression of CD133 and reduced dystroglycan expression are strong predictors of poor outcome in colon cancer patients

“…In our opinion this can be a highly confusing factor. Indeed, we previously demonstrated, by western blot analysis, that CD133 is expressed at various levels in colon cancers [32,33] and that different results can be obtained by using different antibodies [34] and similar observations have been also reported by other Authors [35,36]. The observation that high CD133 expression has been reported to be a negative prognostic factor for colorectal cancers in several studies using different antibodies strongly suggests an important prognostic significance of its detection [1,2,37].…”

Tumor Initiating Cells and Chemoresistance: Which Is the Best Strategy to Target Colon Cancer Stem Cells?

“…These cells are called tumor initiating stem cells or cancer stem cells (CSCs). Currently, it is believed that only this small fraction of cancer cells is able to drive tumor initiation, proliferation, and spreading [6]. Biologically distinct populations of CSCs have been identified in most solid tumors, including colon cancer [7, 8].…”

“…This ability of self-renewal enables CSCs to perpetuate the tumor [6]. Actually, despite their capacity for self-renewal, CSCs are relatively quiescent; that is, they have low proliferative rate and are often not cycling.…”

Aspects of the endothelin system in colorectal cancer

“…These CD133 positive cancer stem cells show over-expression of EDN-1 and nuclear receptor subfamily 4, group A, member 2 (NR4A2) transcripts, and there is a relation between the expression of CD133, EDN-1 and NR4A2, as shown by siRNA knockdown of CD133 expression in CaCo-2 colorectal cancer cells, which caused a significant reduction in the expression of EDN-1 and NR4A2 (59).…”