Abstract A056 Intertumoral epigenetic heterogeneity in response to a novel therapy in Ewing sarcoma

Abstract Like many pediatric cancers, Ewing sarcoma (EwS) is a genomically quiet disease. Driven by an oncogenic fusion protein, usually EWSR1-FLI1, the epigenome of this genetically homogenous cancer is a driver of clinical outcomes, where heightened epigenetic heterogeneity within tumors is associated with metastatic disease. While prognosis for localized disease is moderate, outcomes are dismal for patients who present with advanced, relapsed, or refractory disease and there is a lack of targeted therapies in this setting. We have previously described a targeted combination therapy harnessing DDK and WEE1 inhibition exploits elevated endogenous replication stress in EwS to force cells into mitotic catastrophe in vitro. While our experiments evaluating the efficacy of this combination in vivo have been extremely encouraging, the data have shown that individual mice receiving the same treatment have variable patterns of outgrowth after treatment cessation, with some mice demonstrating moderate increases in tumor volume and some mice demonstrating little to no tumor outgrowth. This differential response to therapy is most likely to occur through epigenetic dysregulation. Thus, we aim to interrogate intertumoral epigenetic heterogeneity and examine epigenetic profiles associated with treatment response using samples from in vivo studies. Samples were analyzed from a CDX model using TC32 cells injected subcutaneously. Reduced representation bisulfite sequencing was performed on a control arm (n = 5) and the following three treatment arms, where the DDK inhibitor is TAK-931 (simurosertib) and the WEE1 inhibitor is MK1775 (adavosertib): 1) Irinotecan + Temozolomide (n=9), 2) Irinotecan (full dose) + DDKi + WEE1i (n=7), 3) Irinotecan (half dose) + DDKi + WEE1i (n=9), for a total of 30 samples. Our analysis of this data evaluates epigenetic changes between treatment conditions and increases our understanding of epigenetic heterogeneity in EwS. Citation Format: Emily Isenhart, Ajay Gupta, Joyce Ohm. Intertumoral epigenetic heterogeneity in response to a novel therapy in Ewing sarcoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pediatric Cancer Research; 2024 Sep 5-8; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl):Abstract nr A056.