ECM-dependent gene expression in normal and tumor-derived mammary organoids

A critical step in metastasis is cancer cell dissemination. Dissemination and metastasis are associated with specific genetic changes and changes in extracellular matrix (ECM), but how these changes interact to enable dissemination remains unclear. Here we tested the importance of ECM to dissemination in both normal and malignant mammary epithelium. By time-lapse imaging, we observed collective invasion and dissemination directly in 3D culture. Our results reveal that the pattern of epithelial migration and local dissemination are constrained by the local ECM microenvironment. To identify RNA expression changes that could regulate these changes in cell behavior, we conducted whole genome RNA expression profiling from normal and malignant mammary epithelium in 3D culture. We collected RNA from normal and malignant epithelium during active growth at 4 days in culture in either Matrigel or collagen I. We hybridized the resulting RNA to Agilent single color microarrays with a minimum of three biologically independent microarray replicates per condition. We observed significant gene expression differences between normal and malignant epithelium, even when cultured in the same ECM. In contrast, the ECM microenvironment had a relatively small impact on RNA expression, despite its large effects on migratory strategy and local dissemination. Gene expression was measured in normal and malignant mammary epithelial fragments cultured in one of two 3D matrices (laminin-rich basement membrane gel or collagen I) and collected at day 4, which we observed to have peak invasion and dissemination. At least three independent experiments were performed at each time using different mice for each experiment.