Breast cancer is the second leading cause of cancer death in women in the United States. Metastasis accounts for the death of ~90% of these patients, yet the mechanisms underlying this event remain poorly defined. Transforming growth factor-beta (TGF-beta) is a multifunctional cytokine that functions to suppress tumorigenesis in mammary epithelial cells (MECs). Interestingly, mammary tumorigenesis converts TGF-beta from a tumor suppressor to a tumor promoter through molecular mechanisms that remain incompletely understood. Changes in tissue compliance promote the acquisition of malignant phenotypes in MECs in part through the activity of the extracellular crosslinking enzyme, lysyl oxidase (LOX), which regulates desmoplastic reactions and metastasis. We show that TGF-beta induces the synthesis and secretion of LOX from normal and malignant MECs, and in breast cancers produced in mice. Additionally, antagonizing LOX activity reduces TGF-beta-mediated invasion and epithelial-mesenchymal transition (EMT) in breast cancer cells. We further show that increased extracellular matrix (ECM) rigidity promotes the proliferation of malignant MECs, a cellular reaction that is abrogated by inhibiting TGF-beta signaling or by antagonizing LOX activity.
As a possible mechanism for the influence of matrix rigidity on TGF-beta signaling, we sought to define the microRNA expression profiles induced by TGF-beta and ECM rigidity during metastatic progression. Through global profiling analyses, we identified microRNAs whose expression was induced by TGF-beta and associated with metastatic potential. In doing so, we identified miR-181a as a TGF-beta-regulated “metastamir” that enhanced the metastatic potential of breast cancers by promoting their acquisition of EMT, migratory, and invasive phenotypes. Mechanistically, inactivating miR-181a elevated the expression of the pro-apoptotic molecule, Bim, which sensitized metastatic cells to anoikis. Along these lines, miR-181a activity was essential in driving pulmonary micrometastatic outgrowth and enhancing the lethality of late-stage mammary tumors in mice. Finally, miR-181a expression is dramatically and selectively upregulated in metastatic breast tumors, particularly triple-negative breast cancers, and is highly predictive for decreased overall survival in human breast cancer patients.
Collectively, our findings implicate LOX and miR-181a expression as novel diagnostic markers for metastatic progression, as well as an innovative therapeutic targets to treat metastatic breast cancers.