A breast cancer progression model: the importance of three-dimensional tissue architecture and metalloproteinases

Previous studies from our laboratory have shown that non-malignant and malignant cells can be distinguished easily and rapidly by their morphology and growth rate when cultured in three-dimensional (3D) laminin-rich basement membrane but not when cultured on traditional tissue culture plastic (two-dimensional [2D]) [1,2]. In addition, we have shown that cellular responses to signaling inhibitors and apoptotic agents differ in cells cultured in 2D versus 3D [3,4]. This applies also to our finding with reverted tumor cell lines [3-8]. In this presentation, I will address two inter-related topics. First, we asked how the 3D morphology and gene expression profiles for a panel of 60 breast cancer cell lines for which the Gray laboratory has obtained 2D expression as well as CGH profiles may differ, and whether any of the surrogate genes or phenotypes could track with response to therapy. The cell lines examined so far fell into four distinct morphologies of 'round', 'mass', 'grape-like' and 'stellate'. An ANOVA analysis of Affymetrix gene expression profiles for each of these cell lines was used to identify genes, the expression profiles of which could distinguish the other known parameters of the cultured cells. Of the 22,283 genes on the Affymetrix 133A chip, ~5800 genes were identified where expression patterns differed between different cell lines both in 2D and 3D, and ~2000 genes were identified where expression differed between the non-malignant and malignant cell lines. About 700 genes differed between 2D and 3D, and ~800 correlated with the morphological differences seen in 3D. These genes fall into a number of functional classes, which we are currently analyzing to identify common signaling themes and/or morphological regulators that will be tested by manipulation of expression and correlated with therapeutic response of these cell lines in 2D and 3D to Herceptin and other chemotherapeutic drugs. Second, we have also shown previously that loss of basement membrane in both cultured mammary mouse cells [9] and in transgenic animals led to epithelial to mesenchymal transition (EMT) and mammary tumors [10]. We have now determined the molecular pathways induced by MMP-3 to lead to EMT and genomic instability via production of reactive oxygen species [11]. These mechanisms will be discussed.