Abstract B020: Establishing roles for the FLI1-EWSR1 reciprocal fusion protein in Ewing sarcoma

Abstract Sarcomas constitute a large group of rare, heterogeneous cancers of mesenchymal lineage arising in the bone and soft tissues of children and young adults. Each sarcoma subtype is characterized by distinct clinical, histological, and genetic characteristics. The heterogeneity of these sarcoma subtypes is a contributing factor to poor survival outcomes observed in patients. Despite their diversity, many subtypes harbor a balanced chromosomal translocation fusing the Ewing Sarcoma RNA binding protein 1 (EWSR1) gene with a gene encoding a transcription factor (TF). Balanced translocations generate two distinct fusions: [1] a canonical fusion consisting of the N-terminal transactivation domain of EWSR1 and the C-terminal DNA-binding domain of a TF and [2] a reciprocal fusion harboring the N-terminus of the TF and C-terminal RNA-binding domain of EWSR1. Ewing sarcoma is one sarcoma subtype that harbors canonical EWSR1-FLI1 and reciprocal FLI1-EWSR1 fusions. The oncogenic roles of EWSR1-FLI1 are well-established in that the fusion protein promotes tumorigenesis in precursor cells through the rewiring of transcriptional programs and modulation of the epigenetic landscape. The role of FLI1-EWSR1 in Ewing sarcoma tumorigenesis, however, is unclear. Here, we begin to characterize the functions of FLI1-EWSR1. We developed lentiviral vectors carrying HA-tagged canonical and FLAG-tagged reciprocal fusions. To date, we have introduced these vectors into the fusion-negative fibrosarcoma cell line HT-1080 and Ewing sarcoma cell lines A673 and TC-32. RNA-seq analyses show that FLI1-EWSR1 expression in A673 leads to significant changes in Notch, Wnt, and Hedgehog signaling pathways, as well as in epithelial-mesenchymal transition and RNA-related processes. These findings suggest roles for FLI1-EWSR1 in extracellular matrix remodeling and potential increases in metastatic potential of cells expressing the reciprocal fusion. Ongoing studies aim to evaluate the effects of the reciprocal FLI1-EWSR1 on Ewing sarcoma tumorigenesis and the epigenome, with hopes of uncovering potentially targetable vulnerabilities, paving the road for the development of novel treatments in a patient population in desperate need of effective therapies. Citation Format: Sarah Gawlak, Emily Isenhart, Joyce Ohm. Establishing roles for the FLI1-EWSR1 reciprocal fusion protein in Ewing sarcoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Fusion-Positive Cancer: From Discovery to Therapy; 2026 Jan 13-15; Philadelphia PA. Philadelphia (PA): AACR; Cancer Res 2026;86(1_Suppl):Abstract nr B020.