Angiogenesis is critical for tumor progression. This process is initiated by angiogenic factors produced by tumor cells. As a result, endothelial cells (EC) leave a state of quiescence and surprisingly become more sensitive to apoptosis. This raises the fundamental question of how can EC circumvent the pro-apoptotic forces that are initiated during the angiogenic switch? For many years it has been reported that the levels of plasminogen activator inhibitor-1 (PAI-1) are paradoxically elevated in more advanced stages of cancer and that PAI-1 is an indicator of poor clinical outcome. That PAI-1 could play a stimulatory rather than an inhibitory role in angiogenesis has also been suggested. In our laboratory we have made two observations indicating that PAI-1 may provide a critical pathway to protect EC from apoptosis. First we reported that PAI-1 prevents EC attachment to vitronectin and stimulates migration toward fibronectin in the absence of apoptosis. Second, we recently demonstrated that by its extracellular and anti-proteolytic activity, PAI-1 prevents the cleavage by plasmin of cell membrane-associated Fas ligand (mFasL) and the release of a 21.5 kDa soluble pro-apoptotic FasL fragment that induces apoptosis in EC. On the basis of these observations, the central hypothesis that will be tested in this research proposal is that an increase in PAI-1 expression in stimulated EC is critical to protect EC from FasL/Fas-mediated apoptosis. This hypothesis will be tested by 3 specific aims.
In Aim 1 we will study the mechanism by which the 21.5 kDa plasmin-generated sFasL induces apoptosis, explore whether PAI-1 could also protect EC from apoptosis independently of Fas and test whether the up-regulation of PAI-1 by hypoxia (HIF-11) or VEGF is critical for the survival of stimulated EC.
In Aim 2, we will test in vivo that PAI-1 controls angiogenesis by preventing the release of sFasL and how this process is affected by hypoxia. We will also determine the contribution of PAI-1 produced by bone marrow-derived cells and tumors cells to angiogenesis and metastasis.
In Aim 3, we will use a pharmacological approach to examine the effect of PAI-1 inhibition on tumor growth and angiogenesis. From these studies we anticipate a fundamental understanding of a critical mechanism used by EC to escape FasL/Fas mediated apoptosis during angiogenic stimulation.
Plasminogen activator inhibitor-1 (PAI-1) is a protein that controls the dissolution of blood clots. This protein has been consistently found to be elevated in patients with cancer and in particular breast cancer, and is an indicator of poor clinical outcome. This application will study how PAI-1 positively contributes to tumor angiogenesis and examine whether inhibition of PAI-1 by antibodies or small molecules can be of therapeutic value. Because PAI-1 plays a contributory role in other conditions than cancer, like diabetic retinopathy and thrombotic diseases, our studies will provide valuable information in regard to targeting PAI-1 in the treatment not only of cancer but of several other human diseases.
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