The global objective of this research is to clarify the relevance of ?5?1 integrin-fibronectin interactions and mechanical force in breast tumor malignant progression. Tumor development in the breast is accompanied by significant remodeling and progressive stiffening of the tissue. Transformed mammary epithelial cells (MECs) become sensitized to extracellular matrix (ECM) stiffness, implying that it is the dialogue between a dynamically evolving microenvironment and a progressively aberrant mammary epithelium. Integrin adhesions are the primary means by which a cell senses and transmits mechanical cues from the ECM to affect cell and tissue phenotype. While we and others have demonstrated that integrin signaling is consistently modified with tumor progression and is essential for tumor malignant transformation, the mechanisms by which this occurs and whether there are specific integrins driving tumor development remains unclear. We and others have identified that ?5?1 integrin and its ligand fibronectin (FN) are specifically upregulated with oncogenic transformation and is associated with poor survival in the human breast cancer. Functionally, we have demonstrated that ?5?1 integrin-fibronectin binding potentiates tumor cell malignant phenotype. ?5?1-FN binding is unique in that ?5?1 binds both the RGD domain and PHSRN synergy site of FN in what has been described as a 'catch-bond'effect. Accordingly, I hypothesize that In the stiffened ECM of a tumor, the ?5 integrin-FN binding synergy site catch-bond enhances cell contractility and focal adhesion maturation and signaling, resulting in a pro-invasive and survival phenotype and driving malignant progression. The project will take an interdisciplinary approach, using novel methods to manipulate ECM mechanics and composition combined with orthotopic tumor mouse models to investigate how ?5?1-FN binding promotes tumor progression. For in vitro assays, I will modify mammary epithelial cell integrin expression and determine the specific effect of ?5?1-FN binding and the importance of the synergy site catch-bond in cell contractility, adhesion signaling, pro-invasion signaling, and subsequent effects on tumor cell malignant phenotype. I will demonstrate functional relevance in vivo through an orthotopic tumor mouse model in which I alter ?5?1-FN interactions through treatment with a peptide inhibitor of synergy site binding (ATN-161). These experiments will allow better understanding of the synergy between the tumor ECM and adhesion signaling in tumor evolution. In particular this work aims to elucidate how biophysical characteristics of integrin adhesions can mediate subsequent signaling and influence cell phenotype and consequently disease progression. Importantly, due to the correlation of ?5?1 and poor survival, the results of this proposal focused on tumor cell invasion, migration and metastasis will elucidate key signaling pathways in metastatic progression with direct relevance in human breast cancer.
Mortality from breast cancer is due to tumor metastasis; a multi-step process initiated by tumor cell invasion and subsequent migration through the stroma towards blood and lymphastic vessels permitting tumor cell dissemination. We have demonstrated mammary gland stroma organization and density are dramatically altered with tumor progression; and this remodeling enhances oncogenically transformed epithelial cell invasion and migration. My goal is to understand the interplay between breast tumor cells and their mechanical microenvironment during the transition to invasion and metastasis; and thereby elucidate key signaling pathways and potential therapeutic targets; in particular; we and others have identified ?5?1 integrin as a key driver of tumor progression and correlated with poor patient survival.