Álava Casado, Enrique deGómez Herreros, Fernando2025-07-182025-07-182025-04-10Olmedo Pelayo, J. (2025). Understanding Genome Instability in Ewing Sarcoma: from Molecular Mechanisms to Novel Therapeutic Approaches. (Tesis Doctoral Inédita). Universidad de Sevilla, Sevilla.https://hdl.handle.net/11441/175447Ewing sarcoma (EwS) is an aggressive mesenchymal neoplasm mainly occurring in children, adolescents and young adults. It is characterized by fusion proteins involving EWSR1 and a member of the ETS family of transcription factors, mainly FLI1 (85% of the cases). Although 5-year survival for patients with localized disease is ~70-80%, the prognosis remains poor for those with metastatic disease, with long-term survival rates around 30%. Current EwS treatment remains based on combining traditional chemotherapy, radiotherapy, and surgery. Despite EwS being highly sensitive to genotoxic agents, therapy failure and tumor relapse occur in 30-35% of patients with primary localized disease and 50-80%of the cases with metastasis. Thus, it is crucial to develop novel therapeutic strategies against EwS. In this doctoral thesis, we describe a novel mechanism of sensitivity to DNA topoisomerase 1 poisons in EwS. We found that EWS::FLI1 prevents the resolution of R-loops induced by these drugs via sequestering DHX9 helicase, ultimately resulting in R-loop accumulation, replication stress, and genome instability. In turn, excessive DHX9 or reduced EWS::FLI1 levels render EwS cells resistant to the active metabolite of irinotecan (SN-38) independent of proliferation and global transcription rates. This resistance helps explain how elevated DHX9 levels predict unfavorable clinical outcomes. Moreover, our results highlight the combination of TOP1 poisons and ATR inhibitors as a novel therapeutic approach for EwS treatment.application/pdf229 p.engAttribution-NonCommercial-NoDerivatives 4.0 Internationalhttp://creativecommons.org/licenses/by-nc-nd/4.0/info:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/embargoedAccess