Background: Hypoxia and the subsequent activation of hypoxia-inducible factor-2 α (HIF2 α ) contribute to the progression of a variety of cancers. However, their role in the generation of renal cell carcinoma-derived stem cells has not been fully addressed. Methods: A sphere formation assay, cell proliferation, RT–PCR, western blot, FACS, immunohistochemistry and tumour xenograft were used to study the role of HIF2 α . Results: Propagation of four renal cell carcinoma (RCC) cell lines (Caki-1, Caki-2, 786-O, 769-P) in anchorage-independent floating spheres led to the expansion of cells bearing the CXCR4 (CD184) surface marker. Inhibition of the CXCR4 pathway reduced sphere expansion. The enhanced self-renewal activity of the CXCR4-positive spheres was preceded by the upregulation of HIF2 α . Knockdown of HIF2 α abrogated CXCR4 expression and sphere formation. Finally, RCC-derived spheres showed an undifferentiated phenotype in vivo and formed subcutaneous tumours that highly expressed HIF2 α and CXCR4. Inhibition of HIF2 α abolished tumour growth in animal models. Conclusions: These results suggest that the generation of RCC-derived CSCs involves the activation of HIF2 α and may provide a foundation for the development of new strategies to prevent the induction of CSCs in RCC.
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