Event Abstract Back to Event Triggering cancer cell invasion at extracellullar matrix interfaces Jiranuwat Sapudom1, Stefan Rubner1, Franziska Ullm1 and Tilo Pompe1 1 Universität Leipzig, Institute of Biochemistry, Germany In malignant tumors single cancer cells leave the primary tumor and invade other tissues. Important factors promoting cancer cell invasion and metastasis rely on altered expression of matrix degrading enzymes, cell adhesion, cell motility as well as properties of the surrounding extracellular matrix (ECM). In particular, local changes of the ECM and ECM properties at tissue boundaries are discussed as promoting or inhibitory stimuli in the determination of cancer cell fate. However, many studies of formation, function and pathology of cancer cells are often conducted in two-dimensional (2D) cell culture or in complex animal systems. Biomimetic three-dimensional (3D) in vitro models are envisioned to reduce the complexity of native tissues, allowing a dissection of microenvironmental regulation of tumor cells, but still providing a nearly physiological situation. We engineered topologically and mechanically defined matrices based on collagen I by varying collagen concentration, pH value during fibril formation as well as optional post-modification using cross-linking, fibronectin or glycosaminoglycans functionalization[1],[2]. We used a sequential fibril formation strategy to form two individual 3D matrices of defined characteristics to mimic ECM interfaces. In situ analysis of the fibril formation process using confocal laser scanning microscopy was applied to tightly control interface formation. Breast cancer cell lines (MDA-MB-231, MCF-7) and melanoma cells derived from primary tumors were studied concerning cell morphology, cluster formation and invasion in the different compartments. Long-term single cell tracking inside the 3D matrices was used for in situ studies of cell migration and proliferation directly at the matrix interface. We successfully established ECM interfaces based on collagen I networks. Step-gradients in topology, mechanics and composition of the ECM could be formed avoiding any artifacts (gap, density increase) at the interface. By using 3D single cell tracking, we found the migration characteristics of invasive cancer cells to be triggered by the matrix interface. Their migration behavior was found to change from random-walk to highly directed migration while crossing the matrix interface from dense to open networks. No migration trigger was found in the opposite direction or at interfaces of similar matrices. The behavior was also found to be independent of an increase in network elasticity by crosslinking. Furthermore, the crossing of the matrix interface was found to persistently change cellular phenotype in dividing cells. The results contradict the frequently found durotaxis on 2D surfaces. They suggest, that ECM pore size, but not stiffness, predominantly influence the migratory phenotype of breast cancer cells and that matrix interfaces can instruct persistent changes of cancer cell phenotype. Furthermore, the ECM interfaces are suggested to be valuable tools in other studies of cell and tissue dynamics at tissue boundaries.