Microenvironmental Cues That Regulate Cancer Cell Dormancy in Estrogen-Receptor Positive and Triple-Negative Breast...

The processes regulating cellular dormancy in tumor cells remain inadequately characterized. Further, there are very few models available that can recapitulate a dormancy-like phenotype in vitro. In this study, we investigated the role of microenvironmental cues in inducing a dormancy-like phenotype in estrogen receptor-positive (MCF-7) and triple-negative (MDA-MB-231) breast cancer cell lines. We characterized an in vitro model in distinct conditions defined hierarchically by the following: (a) normoxia or true hypoxia, (b) dishes coated with fibronectin or laminin or none, and (c) presence or absence of 10 ng/mL basic fibroblast growth factor (FGF-2) in the culture medium, added on day 0 of culture. Cells were cultured at clonogenic densities for 9 days (day -1, day 0, up to day +7). Cells were characterized for dormancy- like behavior using increased p-p38 and decreased p-ERK expression (dormant, p-p38_High and p-ERK_Low expression), reduced Ki67 expression, elevated p21 and p27 levels by immunofluorescence, absence of senescence using β-galactosidase staining and resistance to doxorubicin. Fibronectin or laminin were sufficient for the induction of a dormancy-like phenotype in MCF-7 cells under hypoxic conditions, whereas the addition of FGF-2 to fibronectin (but not laminin) could induce a dormancy-like phenotype under normoxic conditions. FGF-2 was required with laminin to induce dormancy in MDA-MB-231 cells under hypoxic conditions (FGF-2 plus fibronectin were unable to induce dormancy), whereas laminin or fibronectin alone were sufficient under normoxic conditions. Overall, we established culture conditions that recapitulate an in vitro breast cancer dormancy-like phenotype and reveal subtype-specific differences in dormancy regulation.