UC San Diego

The goal of this project was to understand the mechanisms by which hypoxia contributes to breast cancer progression. What emerged is an understanding that migratory pathways activated under hypoxia are cell type specific. In MCF-7 cells, hypoxic culture was found to increase production of Erythropoietin, and thus initiate downstream signaling to ERK/MAP kinase in an autocrine manner. MDA-MB-468 cells, on the other hand, demonstrated increased uPAR expression and signaling to Rac1, Akt, and ERK/MAP kinase. Further, while MCF-7 cells show increased cell migration under hypxia, MDA-MB-468 cells become more migratory, more invasive through extracellular matrix, and undergo phenotypic changes consistent with epithelial-mesenchymal transition. These phenotypic changes were confirmed by internalization of E-cadherin, increased vimentin, and translocation of Snail to the nucleus. Finally, using a novel model of tumor hypoxia on the chick chorioallantoic membrane, we confirm that hypoxia induces increased dissemination from a developing tumor, and increased vimentin and uPAR expression in vivo. Multiple signaling pathways downstream of uPAR are involved in the diverse changes induced under hypoxia. Rac1 is required for increased cell migration, matrigel invasion, and type I collagen remodeling. PI3K/Akt is required for increased Snail expression, internalization of E-cadherin, and the shift from an epithelial to a mesenchymal phenotype. uPAR thus emerges as a coordinator of diverse signaling pathways which are complementary in inducing the full spectrum of cellular changes observed in EMT. Upon reoxygenation of MDA-MB-468 cells, EMT is rapidly reversed. This process of mesenchymal-epithelial transition is significant because a reversion to the primary tumor phenotype may be a critical step in metastasis. Upon return to normoxic cell culture uPAR expression rapidly returns to basal level accompanied by reexpression of E-cadherin at cell junctions, loss of vimentin, and a return to basal levels of cell migration and Matrigel invasion. Thus, hypoxia sets off a wide range of cellular changes which may explain studies linking tumor hypoxia to poor prognosis. A better understanding of the pathways which promote tumor progression in poorly perfused tissues may allow for the discovery of novel drug combinations to complement those targeting angiogenesis