Wnt/β-catenin activated Ewing sarcoma cells promote the angiogenic switch.

Wnt/beta-catenin signaling is active in small subpopulations of Ewing sarcoma cells, and these cells display a more metastatic phenotype, in part due to antagonism of EWS-FLI1-dependent transcriptional activity. Importantly, these beta-catenin-activated Ewing sarcoma cells also alter secretion of extracellular matrix (ECM) proteins. We thus hypothesized that, in addition to cell-autonomous mechanisms, Wnt/beta-catenin-active tumor cells might contribute to disease progression by altering the tumor microenvironment (TME). Analysis of transcriptomic data from primary patient biopsies and from beta-catenin-active versus-nonactive tumor cells identified angiogenic switch genes as being highly and reproducibly upregulated in the context of beta-catenin activation. In addition, in silico and in vitro analyses, along with chorioallantoic membrane assays, demonstrated that beta-catenin-activated Ewing cells secreted factors that promote angiogenesis. In particular, activation of canonical Wnt signaling leads Ewing sarcoma cells to upregulate expression and secretion of proangiogenic ECM proteins, collectively termed the angiomatrix. Significantly, our data show that induction of the angiomatrix by Wnt-responsive tumor cells is indirect and is mediated by TGF-beta. Mechanistically, Wnt/beta-catenin signaling antagonizes EWS-FLI1-dependent repression of TGF-beta receptor type 2, thereby sensitizing tumor cells to TGF-beta ligands. Together, these findings suggest that Wnt/beta-catenin-active tumor cells can contribute to Ewing sarcoma progression by promoting angiogenesis in the local TME.