Invasion and metastasis are the major cause of cancer-related deaths. One of the emerging themes in cancer progression is the cooperative role between cellular transformation and the tumor microenvironment that plays a critical role in promoting invasive and metastatic behavior. Changes in the tumor microenvironment, including reprogramming of stromal cells to secrete cytokines, aid in invasion and dissemination of the tumor cells. The molecular network of such cooperation is poorly understood.
In this report we demonstrate that normal epithelial cells with disruption of polarity genes cooperate with changes in the microenvironment (Matrigel/Collagen-I mixture and stimulation with inflammatory cytokines, IL-6 or TNFα) to gain invasive properties even in the absence of expression of transforming oncogenes. In oncogene expressing cells, loss of expression of cell polarity genes cooperates with transformation to promote invasion in culture and metastasis in vivo.
Our results provide a surprising new insight into the nature of collaboration between ErbB2 and disruption of polarity. Our results on cooperation between disruption of cell polarity and activation of ErbB2 are consistent with recent studies in Zebrafish where loss of Lgl2 cooperated with ErbB2 to induce invasive behavior 
and in human breast cancer cells where overexpression of 14-3-3ζ
,a component of polarity complexes, cooperated with ErbB2 to induce EMT and invasion 
. We show that inhibition of the ErbB2-Par6 pathway was sufficient to block ErbB2-induced invasion, even in the presence of the second polarity gene loss, suggesting that developing ways to inhibit the ErbB2-Par6/aPKC pathway will open new avenues for blocking metastasis in ErbB2 positive cancers.
The observations on the cooperation between ErbB2 and loss of polarity can have significant clinical implications because amplification and overexpression of ErbB2 correlates with poor patient outcome in breast and multiple epithelial malignancies 
. In addition several lines of evidence suggest that amplification of ErbB2 is an early event in tumorigenesis and not directly associated with invasive disease 
. For example, although only 20–25% of human breast cancers fall under the ErbB2 positive subtype, more than 45% of human non-invasive breast carcinomas can possess amplified and overexpressed ErbB2 suggesting that ErbB2 overexpression is more frequently associated with non-invasive disease 
. Results presented in this manuscript, taken together with previously published results, demonstrate that loss of polarity proteins efficiently cooperate with ErbB2 to induce invasion and suggest that dysregulation of polarity proteins can regulate metastatic progression of ErbB2 positive cancers. In addition to cooperating with ErbB2, we show that changes in polarity proteins themselves regulate the signaling by the ErbB family of receptors. Downregulation of cell polarity proteins resulted in an increased expression of ErbB1 (Epidermal growth factor receptor, EGFR) which is likely to play a role in Akt activation. We show that inhibition of Akt blocks the invasive behavior of epithelial cells lacking cell polarity proteins. Thus, a better understanding of the changes in cell polarity proteins in ErbB2 positive cancers can identify novel ways to treat and/or predict cancer progression.
Our results provide a novel perspective on the role played by cell polarity proteins in metastasis. Our results suggest that normal polarity protein function is required for maintenance of a differentiated, epithelial cell phenotype. Disruption of multiple polarity proteins in oncogene naïve, normal epithelial cells induces phenotypic plasticity where the cells behave like invasive mesenchymal cells in response to a tumor-like microenvironment, while they behave like epithelial cells under normal microenvironmental conditions. This phenotypic plasticity has been referred to as ‘partial-EMT’, or ‘metastable’ or ‘hybrid’ state 
. Partial-EMT has been observed in basal-type breast cancers where both epithelial and mesenchymal proteins can be co-localized within single cancer cells 
. A comprehensive analysis of the various mouse models of breast cancer show that cells in spindle cell tumors express both epithelial and mesenchymal markers, suggesting the presence of phenotypic plasticity in vivo
. Human metaplastic breast tumors that are drug resistant and exhibit some of the characteristics of spindle cell tumors also have epithelial, EMT and stem cell like characteristics compatible with the presence of phenotypic plasticity 
The plastic differentiation state will provide a significant advantage for migrating/invading epithelial cells. Migrating cells need to reorganize their cytoskeleton, their vesicle trafficking and vesicle targeting machinery in order to transit from apical-basal polarity to front-rear polarity 
. During this transition polarity proteins function as core signaling proteins that are retooled and rewired 
. For example, in glandular epithelial cells undergoing apical-basal polarization, the Par6/Par3/aPKC complex is recruited to early nascent cell-cell contacts of epithelial cells and plays a critical role during establishment of tight junctions and apical-basal polarity 
. AF-6 regulates tight junctions via interaction with ZO-1 
and Scribble localizes to cell-cell junctions and regulates establishment of lateral membrane polarity 
. In migrating cells with front-rear polarity, the Par complex localizes to the leading edge and regulates localized activation of Cdc42 and orientation of the microtubule organizing center to facilitate directional migration 
. Scribble localizes to the leading edge and regulates assembly of β-pix and PAK1 and induces localized Rac activation and directional migration 
and AF6 relocalizes to the leading edge of migrating fibroblasts 
. While the mechanisms by which loss of a polarity protein regulates cell behavior is not well understood, it is likely that it involves both direct and indirect mechanisms. Loss of polarity proteins may directly affect signaling pathways since many of the polarity proteins function as signaling scaffolds that assemble multiple signaling transduction proteins. In addition, loss of one polarity protein can induce changes in localization and function (not necessarily changes in expression levels) which in turn can indirectly affect cell behavior. A deeper understanding of how changes in polarity proteins induce the phenotypic plasticity is likely to provide novel insights for understanding metastasis.