Cichon, Magdalena A.

Abstract Multinucleation is found in more than one third of tumors and is linked to increased tolerance for mutation, resistance to chemotherapy, and invasive potential. The integrity of the genome depends on proper execution of the cell cycle, which can be altered through mechanotransduction pathways as the tumor microenvironment stiffens during tumorigenesis. Here, we show that signaling downstream of matrix metalloproteinase-3 (MMP3) or TGFβ, known inducers of epithelial–mesenchymal transition (EMT), also promotes multinucleation in stiff microenvironments through Snail-dependent expression of the filament-forming protein septin-6, resulting in midbody persistence, abscission failure, and multinucleation. Consistently, we observed elevated expression of Snail and septin-6 as well as multinucleation in a human patient sample of metaplastic carcinoma of the breast, a rare classification characterized by deposition of collagen fibers and active EMT. In contrast, a soft microenvironment protected mammary epithelial cells from becoming multinucleated by preventing Snail-induced upregulation of septin-6. Our data suggest that tissue stiffening during tumorigenesis synergizes with oncogenic signaling to promote genomic abnormalities that drive cancer progression. Significance: These findings reveal tissue stiffening during tumorigenesis synergizes with oncogenic signaling to promote genomic abnormalities that drive cancer progression. Cancer Res; 78(9); 2277–89. ©2018 AACR.

Abstract Basal subtype breast cancers have a particularly poor prognosis, with high invasiveness and resistance to most targeted therapies. TGFβ and MYC drive central features of basal breast cancer: TGFβ is an autocrine and paracrine signaling factor that drives cell invasion and metastasis, and MYC is a central regulator of cellular proliferation that is upregulated in many cancer types. We show here that genetic or pharmacologic inhibition of MYC in MCF10A basal breast cells results in increased sensitivity to TGFβ-stimulated invasion and metastasis and also show that this signaling loop is dependent on activation of SRC. Analysis of human breast cancer datasets and additional experiments with breast cancer cell lines further suggest the relevance of this signaling loop in basal, but not luminal, breast cancers. Our results imply precaution should be taken when utilizing therapeutic inhibitors of MYC with basal breast cancer patients as this could lead to increased metastasis; however, simultaneous pharmacologic inhibition of SRC and MYC for these patients could facilitate the antiproliferative effects of MYC inhibition while blocking the consequent promotion of metastasis. Cancer Res; 76(12); 3520–30. ©2016 AACR.

Epithelial-mesenchymal transition (EMT) is a physiological program that is activated during cancer cell invasion and metastasis. We show here that EMT-related processes are linked to a broad and conserved program of transcriptional alterations that are influenced by cell contact and adhesion. Using cultured human breast cancer and mouse mammary epithelial cells, we find that reduced cell density, conditions under which cell contact is reduced, leads to reduced expression of genes associated with mammary epithelial cell differentiation and increased expression of genes associated with breast cancer. We further find that treatment of cells with matrix metalloproteinase-3 (MMP-3), an inducer of EMT, interrupts a defined subset of cell contact-regulated genes, including genes encoding a variety of RNA splicing proteins known to regulate the expression of Rac1b, an activated splice isoform of Rac1 known to be a key mediator of MMP-3-induced EMT in breast, lung, and pancreas. These results provide new insights into how MMPs act in cancer progression and how loss of cell-cell interactions is a key step in the earliest stages of cancer development.

Epithelial–mesenchymal transition (EMT) is a form of epithelial plasticity implicated in fibrosis and tumor metastasis. Here we show that the mechanical rigidity of the microenvironment plays a pivotal role in the promotion of EMT by controlling the subcellular localization and downstream signaling of Rac GTPases. Soft substrata, with compliances comparable to that of normal mammary tissue, are protective against EMT, whereas stiffer substrata, with compliances characteristic of breast tumors, promote EMT. Rac1b, a highly activated splice variant of Rac1 found in tumors, localizes to the plasma membrane in cells cultured on stiff substrata or in collagen-rich regions of human breast tumors. At the membrane, Rac1b forms a complex with NADPH oxidase and promotes the production of reactive oxygen species, expression of Snail, and activation of the EMT program. In contrast, soft microenvironments inhibit the membrane localization of Rac1b and subsequent redox changes. These results reveal a novel mechanotransduction pathway in the regulation of epithelial plasticity via EMT.

Abstract Epimorphin is a stromally produced factor that plays a role in normal mammary branching morphogenesis, but deregulated expression of epimorphin can have oncogenic consequences, although the oncogenic processes activated by epimorphin have not been elucidated. We had previously found that treatment of cultured mammary epithelial cells with epimorphin blocked activation of cell death pathways by a variety of apoptotic stimuli. These results implicated a potential role for epimorphin for inhibition of cellular apoptosis, a critical stage of tumor development. To identify potential mediators by which epimorphin could be affecting cell death pathways, we used a transcriptional profiling approach comparing mammary epithelial cells treated with epimorphin versus untreated cells. We found that epimorphin treatment led to significantly increased expression of IKBKE, which inhibits apoptosis through activation of NF-kappaB and which has recently been characterized as a key breast cancer oncogene. We found that epimorphin specifically blocks induction of apoptosis and activation of caspase-3 in cells treated with tumor necrosis factor-alpha, and that this activity was linked to epimorphin-mediated activation of IKBKE. We also found that knockdown of IKBKE blocked epimorphin-mediated induction of branching morphogenesis, implicating activation of IKBKE by epimorphin as a critical function for its normal morphogenic activities as well. These studies provide insight into an additional and unexpected linkage between mammary gland developmental processes and breast cancer progression. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4774. doi:10.1158/1538-7445.AM2011-4774