Degradation of extracellular matrices (ECM) by proteolytic enzymes is a critical aspect of tumor metastasis. It has been shown that ECM degradation occurs at the cell surface and at the cell-matrix contacts of invading cancer cells. Therefore, cell surface localization of ECM-degrading enzymes facilitates tumor cell migration and invasion. Cell surface association of proteinases has also been shown to play a central role in the control of proteolytic activity by promoting the interaction of enzymes with receptors, activators, and relevant substrates. The matrix metalloproteinase-9 (MMP-9) is a key member of the MMP family of ECM-degrading enzymes that has been associated with malignant progression. Functional studies in the experimental models have shown a direct involvement of MMP-9 in tumor metastasis. Therefore, MMP-9 represents a promising target for novel anti-metastatic approaches. MMP-9 activity is regulated by the activation of the zymogen (proMMP-9) form into an active enzyme species capable of proteolytic activity. Although much information has been gained on the process of proMMP-9 activation in solution, the mechanisms yielding active MMP-9 in tumor cells are still poorly understood. New in vivo and in vitro evidence demonstrates that proMMP-9 binds with high affinity to the surface of tumor cells and localizes at the tumor-basement membrane junctions. Consistently, a surface-associated alpha2(IV) chain of collagen IV has been shown to mediate the binding of proMMP-9 to the surface of various tumor cell lines. Preliminary studies show that surface association of proMMP-9 enhances activation and catalytic activity while precluding interactions with TIMP-1, a specific MMP-9 inhibitor. However, the precise cellular and molecular mechanisms mediating these effects remain to be determined. Studies designed in this application will (1) define the proteolytic mechanisms leading to proMMP-9 activation in tumor cells, (2) establish the structural basis for the cell surface association of proMMP-9, (3) define the role of alpha2(IV) in the cell surface binding and activation of proMMP-9 and (4) investigate the role of cell surface association and activation of proMMP-9 on ECM degradation and tumor cell invasion. The results of these studies will address a new paradigm in the regulation of proMMP-9 activation and may provide novel and specific tools to inhibit MMP-9 activity in human tumors.
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