Role of COX-2 in Tumorospheres Derived from a Breast Cancer Cell Line

Abstract

Cyclooxygenase-2 (COX-2) expression in primary breast cancer predicts tumor cell dissemination to bone marrow, which is a risk factor for recurrence and distant metastasis. "Stem-like" phenotype may be important in cancer metastasis. To investigate the role of COX-2 protein in breast cancer stem-like cells, we analyzed it by co-immunofluorescence in tumorospheres derived from the MCF7 estrogen receptor-positive breast cancer cell line. To evaluate COX-2 function we utilized a COX-2 inhibitor in a clonogenicity assay performed with tumorospheres-derived cells. We detected rare cells in tumorospheres (one cell per tumorosphere) with very high COX-2 expression (COX-2(high)). COX-2 transfected MCF7 cells were able to generate long-term tumorospheres culture, even though transfection efficiency was only one in a million cells. We detected expression of OCT4 in some COX-2(high) cells, supporting the hypothesis that these cells could be cancer stem-like cells. It is important that COX-2(high) cells showed less expression of Ki-67 than did neighboring cells, indicating that COX-2(high) cells may be progenitors of tumorospheres. Celecoxib inhibited the growth of tumorosphere cultures and the ability of tumorosphere-derived cells to form colonies in vitro, indicating an active role of COX-2 in these processes. However, 2 μM celecoxib failed to eradicate tumorosphere-initiating cells. Finally, we detected rare COX-2(high) cells among SUM149 inflammatory breast cancer cells growing on plastic in serum-containing medium; the SUM149 cell line produces a very high level of COX-2 protein. Our results support a role for COX-2 in stem-like breast cancer cells and suggest a mechanism behind a role for COX-2 in disseminated tumor cells, which are known to exhibit characteristic biomarkers and functional properties of stem-like cells.