Abstract Embryonal Rhabdomyosarcoma (ERMS) stands as the prevailing subtype of Rhabdomyosarcoma (RMS), the most prevalent pediatric soft tissue sarcoma. With a survival rate below 20% in the metastatic stage, RMS lacks targeted drug therapies, emphasizing the need for novel treatment avenues. Genomic analyses have unveiled a low incidence of somatic mutations in RMS, highlighting the substantial role of epigenetic modifiers in oncogenesis.This investigation focuses on EHMT1's involvement in ERMS oncogenesis. Our findings illuminate EHMT1 as a pivotal contributor to the migratory and invasive capabilities of ERMS cells, both in vitro and in vivo. Transcriptomic analysis following EHMT1 depletion unveils significant alterations in pathways crucial for cell migration and invasion. Specifically, the transcription factor SOX8, essential for cellular motility, experiences a substantial reduction upon EHMT1 loss. Notably, the phenotypic effects of EHMT1 loss are mirrored upon SOX8 depletion.Further RNA-Sequencing of SOX8-depleted cells exposes the downregulation of multiple integrin genes. Mechanistically, we delineate EHMT1's role in the upregulation of SOX8 through the regulation of BRD4 expression, influencing its occupancy at the promoter. This study unravels a novel EHMT1-SOX8 axis as a key driver of metastasis in ERMS, providing valuable insights for potential therapeutic interventions in this challenging clinical landscape. Citation Format: Dipanwita Das, Upasana Bajaj, Jia Yu Leung, Nurul Fateha Binti Abdul Rashi, Vinay Tergaonkar, Reshma Taneja. EHMT1 mediates cellular motility in embryonal rhabdomyosarcoma by activating SOX8 expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB204.