We have previously demonstrated that Ewings sarcomas are exquisitively sensitive to apoptosis via TRAIL receptor agonists. The vast majority of Ewings sarcoma cell lines express TRAIL-Receptor 2 (TR2) and most cells undergo apoptosis within 24 hours of ligation of the TR2 receptor with either monoclonal antibody or the natural TRAIL ligand. Moreover, xenograft models show that anti-TR2 is an active agent against Ewings sarcomas growing in vivo. On this basis, we have initiated a Phase I clinical trial of anti-TR2 in childhood solid tumors and this trial is currently proceeding. Despite this promising preclinical data, we have also noted that a minority of Ewings tumor cell lines are not sensitive to anti-TR2 and we observed that anti-TR2 was unable to cure most animals with established Ewings xenografts. Therefore, we have sought to understand the factors that limit sensitivity of Ewings sarcoma to TR2 targeted agents. It is important to note in the context of this work, that TRAIL based killing is one of the primary pathways through which the immune system eradicates tumors and therefore our interest in this system is based both upon the possibility for direct clinical application with anti-TR2 as well as the possibility that optimizing cell death via TRAIL receptor signaling could also improve the effectiveness of immune based therapies for these tumors. The primary accomplishment for this project during FY2008 was the publication of a manuscript demonstrating the importance of caspase 8 in sensitizing Ewings sarcoma cells TRAIL mediated cell death and identifying that caspase 8 expression in Ewings sarcoma clinical samples is heterogeneous (Lissat et al, Am J Path 2007; 170:1917). This was a collaborative work and our program contributed by providing the analysis of more than 50 Ewings tumors for caspase 8 expression which was a fundamental element of this report. Thus, we have demonstrated that the full benefit of TRAIL mediated killing in Ewings sarcoma, whether delivered via a moAb or via a cytolytic cell, can only be accomplished if caspase 8 expression is assured. We have further demonstrated that interferon gamma is a primary regulator of caspase 8 expression and that exposure of Ewings sarcoma cells to interferon gamma results in substantial caspase 8 upregulation and increased sensitivity to anti-TRAIL receptor therapies. We therefore will seek to incorporate interferon into our ongoing clinical trial of anti-TR2 and will seek to combine TRAIL based therapies with immune therapies that can deliver interferon locally to the tumor site and therefore potentially synergize with each other.
