Hypoxia/HIF-1 regulation of mammary morphogenesis and oncogenesis

Epithelial attachment to substratum is critical for survival as cells that do not establish proper adhesion undergo apoptosis. Detachment-induced cell death, termed anoikis, is a fundamental regulator of mammary tissue architecture. Moreover, anoikis resistance is a common feature of cancer cells that has been linked to the disruption of tissue organization associated with breast cancer progression. Premalignant breast cancer lesions such as ductal carcinoma in situ (DCIS), characterized by the presence of cells in ductal lumens, often display hypoxic marker expression. Hypoxia, a common feature of solid tumors, has been linked to tumor-promoting cellular programs including survival, migration and invasion; however, the contribution of hypoxia to cancer-associated morphological alterations has yet to be explored. Here, using a three dimensional (3D) tissue culture model of mammary morphogenesis, we find that hypoxia promotes formation of filled, disorganized acini reminiscent of DCIS-like phenotypes found in vivo. We demonstrate that induction of the pro-anoikis proteins Bim and Bmf during detachment is inhibited by hypoxia. We find that hypoxia-mediated anoikis suppression is dependent upon maintenance of MAPK signaling and the hypoxia-inducible transcription factor HIF-1. Further, hypoxia-mediated acinar disorganization, MAPK signaling and anoikis suppression are sustained after reoxygenation. These data support a model wherein hypoxia, via HIF-1, alters anoikis-associated signaling, allowing epithelial cell survival in the absence of matrix adhesion and potentially contributing to development of early breast cancer lesions. The oncogene ErbB2 is commonly overexpressed breast cancer and can stabilize HIF-1[alpha] expression in a hypoxia-independent manner. Like hypoxia, ErbB2 promotes anoikis resistance through MAPK maintenance and suppression of Bim. To extend our studies, we investigated the role of HIF-1 in ErbB2-mediated anoikis resistance. We find that depletion of HIF-1 in ErbB2-overexpressing cells inhibits anchorage-independence, 3D growth and MAPK and Akt signaling. HIF-1-depleted ErbB2-overexpressing cells also display restoration of anoikis sensitivity that correlates with Bim accumulation and induction of oxidative stress. We propose HIF-1 stabilization as a novel means through which ErbB2 can co-opt signaling pathways associated with hypoxia-mediated survival to establish anoikis resistance. Collectively, these studies define a role for HIF-1 in regulating detachment-induced cell death and tissue architecture in the context of breast carcinogenesis.