Recent studies by many investigators have identified representatives of all four major classes of proteases in breast cancer. However, the exact roles of these proteases in tumor angiogenesis growth, and metastasis are not yet clarified. We hypothesize that the matrix metalloproteases (including MMP-1, MMP-2, and MMP-9) play a pivotal role in tumor invasion and metastasis. We also propose that the serine/thyronine protein kinase family termed PKC is also important in this process due, at least in part, to its regulation of MMP synthesis and secretion. To test these ideas we will continue our study of a high affinity hydroxylamate inhibitor of MMP catalytic activity (termed BB-94, Batimastat) and a high affinity macrocyclic lactone inhibitor of PKC-induced MMP syntheses (bryostatin-1). We will carry out integrated studies of these two drugs in early phase clinical trials and in several experimental animal model systems of breast tumor growth, angiogenesis, invasion, lymph node and lung metastasis, and ascites invasion-late stage survival. In these studies, MMP levels and degree of catalytic activation will be monitored in plasma, pleural/ascites fluids, and in tumor biopsies for biochemical evidence of drug effectiveness. We will also further develop studies demonstrating a favorable interaction of BB-94 with bryostatin-l and of both agents with certain chemotherapeutic agents in our animal models. Interactions of BB-94 and bryostatin-l with retinoids and certain antiangiogenic drugs will also be examined as collaborations with two other projects in this application; retinoids are also known to suppress MMP synthesis. Evidence of favorable interactions in vivo with any of these strategies will lead to design of additional early phase clinical trials in the latter years of this proposal. Finally, we will explore proteolysis-associated mechanisms of treatment failure in animal models and patient trials. Plasma, ascites, and tumor biopsies in breast cancers undergoing treatment-associated failure will be evaluated for upregulated or activated MMPs, upregulation of urokinase (UPA) and increases in a novel 80kDa MMP-like enzyme which we have recently described. Regulation of protease inhibitors will also be evaluated. In summary, this project focusses on MMPs as therapeutic targets in breast cancer, establishes the potential for favorable interactions of antiMMP therapies with conventional chemotherapy and with other new biological therapies, and evaluates the role of further disregulated production of proteases in treatment failure.