The AC133 Epitope, but not the CD133 Protein, Is Lost upon Cancer Stem Cell Differentiation

Abstract

Colon cancer stem cells (CSC) can be identified with AC133, an antibody that detects an epitope on CD133. However, recent evidence suggests that expression of CD133 is not restricted to CSCs, but is also expressed on differentiated tumor cells. Intriguingly, we observed that detection of the AC133 epitope on the cell surface decreased upon differentiation of CSC in a manner that correlated with loss of clonogenicity. However, this event did not coincide with a change in CD133 promoter activity, mRNA, splice variant, protein expression, or even cell surface expression of CD133. In contrast, we noted that with CSC differentiation, a change occured in CD133 glycosylation. Thus, AC133 may detect a glycosylated epitope, or differential glycosylation may cause CD133 to be retained inside the cell. We found that AC133 could effectively detect CD133 glycosylation mutants or bacterially expressed unglycosylated CD133. Moreover, cell surface biotinylation experiments revealed that differentially glycosylated CD133 could be detected on the membrane of differentiated tumor cells. Taken together, our results argue that CD133 is a cell surface molecule that is expressed on both CSC and differentiated tumor cells, but is probably differentially folded as a result of differential glycosylation to mask specific epitopes. In summary, we conclude that AC133 can be used to detect cancer stem cells, but that results from the use of this antibody should be interpreted with caution.

Noninvasive positron emission tomography and fluorescence imaging of CD133 + tumor stem cells

“…The AC133 epitope of CD133/prominin is perhaps the most intensely investigated of the known CSC markers (8,13,17,19,20,22). Although of great interest and importance, noninvasive imaging of AC133 + CSCs or other types of CSCs with clinically relevant tracers and imaging modalities has not yet been reported (9)(10)(11).…”

“…AC133 is an N-glycosylation-dependent epitope of the second extracellular loop of CD133/prominin-1, a cholesterol-binding protein of unknown function that locates to plasma membrane protrusions (12)(13)(14). Postnatally, the CD133 protein is expressed by certain epithelial and nonepithelial cells, by stem and progenitor cells of various organs, and by CSCs of many different types of malignant tumors (15).…”

“…Postnatally, the CD133 protein is expressed by certain epithelial and nonepithelial cells, by stem and progenitor cells of various organs, and by CSCs of many different types of malignant tumors (15). With a few exceptions, recognition of the AC133 epitope by the AC133 mAb appears to be limited to cells harboring stem cell properties, and the AC133 epitope-but not necessarily the CD133 protein-is down-regulated upon differentiation, presumably because of changes in glycosylation (12,13,15). AC133 + tumor stem cells have been described for glioblastoma multiforme (the most common and most aggressive primary brain tumor in adults), various pediatric brain and central nervous system tumors (medulloblastoma, ependymoma, pineoblastoma, teratoid/rhabdoid tumors, and retinoblastoma), brain metastases, many different types of carcinomas including colon, pancreatic, lung, liver, and ovarian cancer, melanoma, sarcomas, and different types of leukemia.…”

Understanding cancer stem cell heterogeneity and plasticity

“…Chen et al [66] [75][76][77][78][79][80][81][82][83][84][85][86] using cell surface markers CD133 [75][76][77], CD44 [78,79] or ABCB5 [85], clonal analysis [80], SP phenotype [84], and Aldefluor assays [82]. As in studies of BTSCs, use of CD133 as a positive selection marker for colon CSCs has generated conflicting results [75][76][77]81], but subsequent studies indicate that the AC133 epitope, but not CD133 protein, is differentially and specifically expressed in colon CSCs and its expression is lost upon differentiation [83]. On the other hand, CD44 + cells seem to consistently enrich tumorigenic colon cancer cells and additional subfractionation of the CD44 + EpCAM + cell population with CD166 further enhances the success rate of tumor engraftments [78].…”

Frontiers in Suicide Gene Therapy of Cancer

“…Glioblastoma multiforme (GBM) is the most often and the most deadly brain cancer [7][8][9][10][11][12][13]. It is incurable.…”