The Function of Wnt/beta-catenin Signaling in Ewing Sarcoma and its Contribution to Pathogenesis

Ewing sarcoma is an aggressive bone and soft tissue tumor with a high propensity for metastasis; however, the mechanisms that contribute to this process are poorly understood. The Wnt/beta-catenin signaling pathway is critical for oncogenesis in numerous cancers, and although previous studies implicate a role for this pathway in Ewing sarcoma, its specific function and contribution is unknown. Previous work by our lab revealed that the Wnt-modulatory receptor LGR5 is highly expressed in patients with aggressive disease, and we hypothesized that LGR5 regulates activation of Wnt/beta-catenin signaling. Through investigation of primary tumors, we discovered that focal nuclear beta-catenin is detectable in a subset of Ewing sarcoma patients and strongly associated with LGR5 expression. Patients whose tumors have nuclear beta-catenin or high expression of the downstream Wnt/beta-catenin target LEF1, experienced worse clinical outcomes and overall survival. We next used in vitro and in vivo models to determine the function of Wnt/beta-catenin signaling in Ewing sarcoma. Importantly, we found that LGR5 expression and Wnt activation were highly heterogeneous. We then investigated the downstream effects of Wnt/beta-catenin activation in the most highly Wnt-responsive cells. RNA-sequencing revealed that Wnt/beta-catenin paradoxically inhibits EWS-ETS transcriptional activity, resulting in a phenotypic change from a proliferative to a migratory and metastatic state in vitro and in vivo. In addition, the metastasis-associated molecule Tenascin C was upregulated by Wnt/beta-catenin signaling, and was found to be a mediator of migration in vitro and metastasis in vivo. In the context of the tumor microenvironment, we further found that patient tumors with high Wnt/LEF1 expression had significant correlation with expression of stroma- and angiogenesis-related genes associated with a poor prognosis. Together, these data provide novel avenues of exploration for tumor-microenvironment interactions. In conclusion these findings implicate a critical role for Wnt/beta-catenin-singaling in mediating migration and metastasis. This occurs in part through antagonism of EWS/ETS fusion protein activity and by up-regulation of the metastasis-associated gene Tenascin C. In addition, tumor-microenvironment interactions modulated by Wnt/beta-catenin further contribute to pathogenesis. Together these findings provide exciting new venues for therapeutic investigation.PHDMolecular & Cellular Path PhDUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/144020/1/easp_1.pd