EWS-FLI1 low Ewing sarcoma cells demonstrate decreased susceptibility to T-cell-mediated tumor cell apoptosis

Abstract

Metastatic and relapsed Ewing sarcoma typically afflicts the adolescent population and is largely fatal. These bone tumors are most commonly driven by the fusion oncoprotein EWS-FLI1. Ewing tumors demonstrate significant intra-tumoral heterogeneity, and individual tumor cells can express highly variable and dynamic levels of EWS-FLI1. Recent studies revealed that the EWS-FLI1 oncoprotein level (high versus low expression) greatly influences the behavior of Ewing tumor cells. As compared to cells with high EWS-FLI1, Ewing cells in the EWS-FLI1 low state demonstrate an increased propensity for metastasis. In light of these observations, we sought to determine how tumor cell EWS-FLI1 level influences the anti-tumor cell immune response. Since ICAM-1, which can promote tumor cell/T-cell interaction and T-cell activation, is highly expressed on EWS-FLI1 low cells, we hypothesized that EWS-FLI1 low cells would be more susceptible to T-cell mediated tumor cell apoptosis as compared to cells with high EWS-FLI1. Unexpectedly, we found that EWS-FLI1 low cells are more resistant to T-cell mediated apoptosis than EWS-FLI1 high cells. We investigated the potential mechanisms by which EWS-FLI1 level might influence the T-cell anti-tumor response, and discovered that low EWS-FLI1 expression results in upregulation of PD-L1 and PD-L2, both important ligands for the PD-1 immune checkpoint receptor on T-cells. We demonstrated that blocking PD-1 results in a greater increase of T-cell mediated killing of EWS-FLI1 low tumor cells as compared to cells with higher EWS-FLI1 expression. Our studies suggest that Ewing cells in the EWS-FLI1 low expression state may serve as a niche of tumor immune-evasion.