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Abstract

Cell migration is not only crucial for a range of physiological processes, such as embryonic development or wound healing, but also a major determinant for pathological processes, including cancer dissemination and metastasis. Cancer cell migration relies on two coordinated and interdependent functions, adhesion and proteolysis. While cell adhesion receptors of the integrin family orchestrate interactions between cells and the extracellular matrix (ECM), matrix metalloproteinases (MMPs) degrade matrix barriers and subsequently create space for cell movement. Most studies on cancer cell migration are based on two model systems: collagen gels and reconstituted basement membrane extracts. The ECM however, is a highly complex structure that consists of several components whose function and contribution to cell migration in physiological and pathological conditions is still poorly understood. Hence, new model systems are required to gain a more profound understanding about the interactions between tumor cells and their microenvironment, which in turn drive malignancy. Accordingly, fibronectin (FN), an adhesive fibrillar ECM protein, came into focus, since its expression levels are upregulated in several tumors and it has been associated with the formation of premetastatic niches. In this thesis, cell migration of human fibrosarcoma cells was investigated in two-dimensional (2D) and three-dimensional (3D) FN environments. In 2D environments, FN molecules in their globular form were physisorbed onto surfaces to form a thin, homogenous layer. 3D environments were assembled via a cell-driven process leading to the formation of fibrillar FN networks. Cell migration behavior in these two types of FN environments was examined using integrin blocking approaches and FN cell-binding site mutation. Here, it was demonstrated that on 2D FN coatings, cell migration is strongly dependent on α5β1 integrin, whereas within 3D FN matrices neither α5β1 nor αvβ3 mediate cancer cell migration. Furthermore, the impact of proteolytic activity on cancer cell migration within both FN environments was investigated. The results of this thesis suggest that general inhibition of MMPs does not influence fibrosarcoma cell migration on FN, regardless of its topography. However, as demonstrated by RNA interference, silencing of a membrane-type MMP, namely MT1-MMP, had opposite effects on cancer cell migration behavior in both FN environments. Depletion of MT1-MMP on 2D FN resulted in reduced migration speed and loss of directionality through inactivation of cofilin activity, which is associated with reduced actin dynamics. On 3D FN matrices, migration speed and cofilin activity was increased upon MT1-MMP silencing.

Since cancer cells are able to switch between proteolysis-driven and actomyosin-based migration modes, the influence of the myosin II inhibitor blebbistatin on cancer cell locomotion was further investigated. In this thesis, fibrosarcoma cell migration in 3D fibrillar FN was highly dependent on myosin II contractility, whereas blebbistatin treatment did not influence the migratory behavior on 2D FN coatings. Therefore, cell migration on both substrates showed remarkable differences regarding adhesion, protease activity and myosin II mediated contractility. These results highlight the importance of substrate topography for regulating cell migration and the need for more physiological model systems to investigate cancer cell migration. They further suggest that targeting of matrix molecules, rather than cellular receptors or proteolytic enzymes, is a promising approach to inhibit metastatic processes.