Abstract B35: The effect of extracellular matrix proteins from lung on the migration of breast cancer cells

Abstract

Abstract The lung is a frequent site of breast carcinoma metastasis, yet little is known about the mechanism of breast carcinoma metastasis to lung. The purpose of this study was to determine if extracellular matrix components from the pulmonary microenvironment had the potential to encourage metastasis by inducing motility in breast carcinoma cells. Previous studies provided evidence that products of conditioned media of pulmonary epithelial cell co-cultures were able to induce dose-dependent motility in breast carcinoma cells. The deposited material was analyzed by mass spectrometry, revealing extracellular matrix proteins, including thrombospondin 1 (TSP), tenascin C, transforming growth factor-beta-induced protein ig-h3 (βig-h3), and the α3 and β3 chains of laminin 332 (LN332). Motility assays using either recombinant preparations of TSP, βig-h3 or tenascin C, or purified LN332 revealed that LN332 induced greater motility at almost all concentrations of each protein tested. Since pulmonary cells grown on membranes at a liquid air interface more closely resemble the structure of lung tissue in vivo, cells grown in this manner were tested for their ability to produce LN332 and induce motility. After 7-8 days in culture, pulmonary epithelial cells provided a monolayer at the liquid air interface. Membranes with cells on the surface exposed to air were fixed in formalin, paraffin embedded and sectioned for hematoxylin and eosin staining and immunohistochemistry. Immunohistochemistry revealed that the cells produced LN332, which was both cytoplasmic and in a layer on the membrane surface beneath the cells. Lung cells growing at the liquid air interface were removed from the membrane, and MCF-7 cells were added to the same membrane from which lung cells had been removed, or to membranes which had been incubated with pulmonary cell growth media but no lung cells. Whereas MCF-7 cells placed on membranes without prior lung cells grew in tight clusters, MCF-7 cells added to membranes on which lung cells had been removed showed scattering and pseudopodia formation. This response was blocked by antibodies to α3 and β1 integrins, which are receptors of LN332. Finally, western blots of human lung tissue revealed the presence of LN332. These experiments together showed that pulmonary LN332 is a likely motility factor for breast carcinoma cells. These findings provide a mechanism for the migration of metastatic tumor cells from their arrest in the pulmonary vasculature into the surrounding tissue, allowing their establishment as a metastatic colony. Citation Format: Philip M. Carpenter, Cally Xiao, Tuan Ngo, Paul D. Gershon. The effect of extracellular matrix proteins from lung on the migration of breast cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr B35.