Neuroblastoma is the most common extracranial solid
tumor in children, accounting for up to 10% of
all childhood malignancies. Cellular heterogeneity is a hallmark of this
embryonal cancer, as distinct neural crest lineages can be found within the same
tumor sample. The aim of our study was to investigate the presence of a subpopulation of immature cells with features of
cancer-like stem cells in 10
neuroblastoma cell lines. RT-PCR and flow cytometry were performed in order to analyze different kinds of 'stemness genes' such as:
NESTIN (NES), CD133, SOX-2, BMI1, c-KIT, MELK1, MUSASHI-1 (MSI1), FAS, CD44 and
VIMENTIN (VIM). In addition, glial and neuronal markers such as NCAM1, GFAP and B-
TUBULIN III (TUBB3) were analyzed. Epigenetic changes within the CD133 (Prominin-1) gene promoter were also analyzed.
Neuroblastoma cell lines showed a particular pattern of expression, suggesting the presence of an immature cancer stem cell-like subpopulation. The CD133
protein, commonly used to enrich putative
cancer propagating stem cell-like populations in different kinds of solid
tumors, presented a half-methylated
DNA state in 7 of the 12
neuroblastoma cell lines analyzed. An increase in
RNA and
protein levels of CD133 was achieved following demethylation by assays using
5-aza-2'-deoxycytidine (5-Aza-dC). Since cancer stem cells are believed to be responsible for
tumor metastasis, escape from anticancer
therapies and disease relapse, their therapeutic targeting and analysis is crucial in
neuroblastoma. Moreover, the regulation of CD133 by epigenetic changes may provide an innovative mechanism of CD133 expression as its regulation still remains unclear.