CD133+ colon cancer cells are more interactive with the tumor microenvironment than CD133-cells

Experimental data indicate that colorectal cancer cells with CD133+ expression exhibit enhanced tumorigenicity over CD133+-negative (CD133+) cells. We hypothesized that CD133+-positive (CD133+) cells, compared with CD133+, are more tumorigenic because they are more interactive with and responsive to their stromal microenvironment. Freshly dissected and dissociated cells from a primary colon cancer were separated into carcinoma-associated fibroblasts (CAF) and the epithelial cells; the latter were further separated into CD133+ and CD133+ cells using fluorescence-activated cell sorter. The CD133+ cells formed large tumors in non-obese diabetic-severe combined immunodeficient (NOD-SCID) mice, demonstrating the phenotypic cellular diversity of the original tumor, whereas CD133+ cells were unable to sustain significant growth. Affymetrix gene array analyses using t-test, fold-change and multiple test correction identified candidate genes that were differentially expressed between the CD133+ vs CD133+ cells. RT-PCR verified differences in expression for 30 of the 46 genes selected. Genes upregulated ( vs-cells) included CD133+ (9.3-fold) and CXCR4 (4-fold), integrin Β8 and fibroblast growth factor receptor 2. The CAF highly express the respective ligands: stromal-derived factor-1 (SDF-1), vitronectin and FGF family members, suggesting a reciprocal relationship between the CD133+ and CAF cells. SDF-1 caused an increase in intracellular calcium in cells expressing both CD133+ and CXCR4, confirming functional CXCR4. The CD133 +CXCR4 phenotype is increased to 32% when the cells are grown in suspension compared with only 9% when the cells were allowed to attach. In Matrigel 3-D culture, the CD133 +CXCR4 group treated with SDF-1 grew more colonies compared with vehicle, as well as significantly larger colony sizes of tumor spheres. These data demonstrate proof of principle that the enhanced tumorigenic potential of CD133+, compared with CD133+, cells is due to their increased ability to interact with their neighboring CAF.