Wnt/Beta-Catenin Signaling and the Tumor Microenvironment in Ewing Sarcoma Progression

Abstract

Metastasis remains the primary cause of death in cancer patients. The invasion-metastasis cascade is dependent on crosstalk between tumor cells and the tumor microenvironment, including stromal cells, extracellular matrix, and blood vessels, at both primary and distal sites. Ewing sarcoma is a bone and soft tissue tumor primarily affecting children and adolescents. Patients with only local disease have high survival rates of >70%. However, current treatment strategies for patients developing metastasis have dismal survival rates of < 20%, making it imperative to better understand the biological processes that drive Ewing sarcoma metastasis. Wnt/beta-catenin activation is correlated with worse overall survival in Ewing sarcoma patients and tumor cell autonomous changes in cytoskeletal organization; however, the tumor cell non-autonomous changes induced by Wnt/beta-catenin remain unexplored. Extracellular matrix encoding genes and genes involved in tumor: tumor microenvironment crosstalk are among the genes activated by Wnt/beta-catenin signaling. Therefore, we hypothesized that Wnt/beta-catenin induces changes in the tumor microenvironment to promote Ewing sarcoma progression. The results of our studies have shown that activation of the Wnt/beta-catenin pathway led to increased secretion of structural collagens and matricellular proteins. Intriguingly, in the secretome of Wnt-activated Ewing sarcoma cells we also detected TGF-beta ligands and proteins that are downstream targets of the TGF-beta signaling cascade. We show that activation of Wnt leads to derepression of TGFBR2, the key mediator of TGF-beta signaling, and detect discrete tumor cell sub-populations, responsive to both Wnt and TGF-beta ligands. Studies of Ewing sarcoma models, in vitro and in vivo, as well as in two independent patient cohorts, confirmed a direct relationship between beta-catenin signaling, TGF-beta activation, and angiogenesis in tumor cells. Mechanistically, this is due, in part, to increased endothelial cell proliferation mediated by increased secretion of the matricellular protein tenascin C. Additionally, we evaluated the other potential functions of the matricellular protein, tenascin C, in Ewing sarcoma progression, and identified stress-mediated activation of tenascin C through activation of Src kinase enhances an invasive phenotype. In conclusion, we have elucidated the importance of Wnt/beta-catenin signaling in altering tumor: tumor microenvironment interactions to promote Ewing sarcoma progression, through secretion of proteins such as tenascin C into the tumor microenvironment. These studies highlight tenascin C as a potential “achilles heel” and therapeutic target in Ewing sarcoma progression. Together these studies illustrate the critical contribution of tumor cell heterogeneity, cell plasticity, and tumor: tumor microenvironment crosstalk to sarcoma progression.