Abstract Pediatric gliomas are the most common form of malignant brain tumors in children, representing ∼50% of all pediatric brain tumor cases and being the leading cause of cancer-related deaths in children under the age of 15. Aggressive standard treatments offer limited benefit in high-grade gliomas, with low survival rates (25%; 5-year survival rate) and significant long-term cognitive consequences diminishing life quality due to the harsh impact of these interventions on the developing brain. These limitations underscore the urgent necessity of novel therapeutic approaches and a more comprehensive understanding of the disease’s molecular drivers and tumor behavior. To explore the molecular underpinnings of pediatric gliomagenesis, we utilized an advanced non-invasive postnatal electroporation approach (MADR) to create an inducible mouse model and derived organoids in combination with extensive genomic, transcriptomic, and proteomic analyses. Through this platform, we identified several ETS transcription factors as key contributors to tumor development. These factors, including the Pea3 subfamily (ETV1, ETV4, ETV5) and ETV6 (also known as Tel), are known for regulating key cellular functions such as proliferation and cell differentiation which could shape the tumor heterogeneity. We observed significant and overexpression of ETV5 and other ETS factors in pediatric gliomas and in a cell-population dependent manner, where they interact directly and dynamically with glial-specific transcriptional regulators (e. g.: OLIG2) to drive tumor formation. Furthermore, our single-cell analyses of Ets inducible MADR-derived organoids were further unraveling the common and different roles of ETV1, ETV4, ETV5, and ETV6 in determining the different tumor progression, cell identity, and how they regulate transcriptional programs during gliomagenesis showing they importance of its regulations in the heterogeneity shaping for the tumor success. Interestingly, among them, ETV4 showed an exclusive potential to force the glioma cell phenotype to fate them to a unique cell cluster. In addition, our data reveal the central role of ETV5 in glial fate decisions by promoting the differentiation of neural precursor cells toward an oligodendrocyte progenitor cell (OPC) -like phenotype. OPCs are highly proliferative and migratory cells considered potential origins of glioma. Thus, targeted in vivo suppression of ETV5 resulted in substantial reductions in tumor growth, delayed disease progression, and improved survival outcomes, affirming its functional importance in the tumor behaviour. These findings emphasize the central involvement of ETS transcription factors in promoting glioma diversity and illustrate how the distinct activation of individual Ets family members can differentially direct the lineage and functional state of tumor subpopulations. Altogether, this study position ETS factors as key regulators of tumor biology and promising therapeutic targets in high-grade pediatric gliomas. Citation Format: Antonio C. Fuentes-Fayos, Hannah Park, Naomi Kobritz, Moise Danielpour, Joshua J. Breunig. ETS transcription factors as key modulators of heterogeneity in pediatric high-grade gliomas abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Brain Cancer; 2026 Mar 23-25; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86 (6Suppl): Abstract nr B046.
Fuentes-Fayos et al. (Mon,) studied this question.