Interaction between Tissue Factor, Junctional Adhesion Molecule-A, and Integrin B1 to drive self-renewal in glioblastoma

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PROJECT SUMMARY Glioblastoma (GBM) is the most common cancer arising in the adult brain, and is lethal in nearly all cases. A key contributing factor to poor outcomes for GBM patients is a subpopulation of cells, known as cancer stem cells (CSCs), that are highly resistant to routinely used genotoxic/cytotoxic therapies, and ultimately manifest as recurrent tumor. Inhibiting tumor recurrence from CSCs that survive therapy would therefore improve GBM patient outcomes. Our preliminary and recently published work show that CSC survival is dependent on Tissue Factor (TF), a conserved transmembrane and secreted protein involved in blood clotting. TF activates protease- activated receptor 2 (PAR2), a G-protein-coupled receptor on GBM cells, which promotes CSC maintenance and expansion, as indicated by analysis of marker expression, self-renewal capacity, and in vivo growth of CSCs. TF suppression greatly reduces CSC subpopulations, in some cases even leading to complete tumor eradication in vivo. Protective and proliferative effects of TF-PAR2 signaling on CSCs appears to be through activation of multiple classes of oncogenic receptor tyrosine kinases (RTKs) like EGFR, although the mechanism by which TF-PAR2-RTKs stimulate CSCs is unclear. Our preliminary data also show that TF positively correlates with expression of Junctional Adhesion Molecule-A (JAM-A), a protein that promotes cell-cell adhesion by stabilizing integrin β1. CSCs depend on such cell-cell adhesion, and our data show that JAM-A is necessary for CSC behavior in GBM. Because TF can also signal through integrin β1, our overarching hypothesis is that TF upregulates JAM-A expression, which stabilizes integrin β1 and enhances the ability of TF to act on integrin β1 and promote self-renewal in GBM. This has therapeutic relevance, because the pro-CSC effects of TF are independent of its role in hemostasis, and blocking JAM-A could potentially reduce the pro-tumor effects of TF, without causing bleeding that would result from targeting TF directly. In Specific Aim 1, we will test the hypothesis that TF drives JAM-A expression through PAR2-RTK signaling. We will identify components of the TF-PAR2 complex that are essential for TF-PAR2 pro-tumor activities, and will test the ability of TF to trigger JAM-A expression while inhibiting each major downstream pathway of RTK signaling. In Specific Aim 2, we will determine whether JAM-A requires serpin B3, a serine-protease inhibitor that we have shown binds to JAM-A, for its pro-CSC activities. In Specific Aim 3, we will use molecular knockouts to determine whether JAM-A is an effective therapeutic target against aggressive, high TF-expressing GBM. Completion of this project will advance our understanding of how CSC subpopulations are maintained in GBM, could well be applicable to a wide range of cancers, and would demonstrate a compelling new therapeutic target in treating numerous malignancies, including GBM.