Abstract 1179: Epigenetic subtypes and cellular plasticity are driven by an interplay between RUNX2 and FOSL1 in osteosarcoma

Abstract Chromatin accessibility is a crucial driver of cellular plasticity, priming gene expression and defining distinct cellular states. Variability in chromatin accessibility is a significant determinant of epigenetic transcriptional regulation and can reveal clinically relevant subtypes and biomarkers of therapy response. In osteosarcoma (OS), we identified two epigenetic subtypes, EC1 and EC2, using ATAC-seq analysis. These subtypes are defined by differential chromatin accessibility and transcriptional circuitry, with EC1 characterized by developmental transcription factors such as RUNX2, MEF2C, and homeobox proteins, while EC2 is dominated by AP1 transcription factors, including FOSL1 and FOSL2. Our findings reveal intricate regulatory mechanisms underpinning these states. RUNX2 in EC1 is tightly controlled by superenhancers that promote osteogenic transcriptional programs, while in EC2, chromatin remodeling supports FOSL1-driven activation of stress-response and proliferation pathways. CUT&RUN assays demonstrated that distinct enhancer landscapes marked by H3K27ac regulate these subtypes. By integrating ATAC-seq and superenhancer mapping using the ROSE algorithm, we identified core regulatory circuits (CRC) unique to each subtype, demarcating transcription factor binding regions and refining cis-regulatory site predictions. OS cells can transition between EC1 and EC2 states driven by CRC members, causing alterations in chromatin accessibility. This plasticity is accompanied by RUNX2 and FOSL1 activity shifts, with transitions regulated by feedback loops in their respective CRCs. Therapeutic screening of the PDX-derived cell line panel revealed distinct vulnerabilities between subtypes. EC2, marked by AP1 upregulation and ERK activation, was highly responsive to MEK inhibitors, while EC1 demonstrated sensitivity to AURKB inhibitors. Single-cell RNA-seq analysis further suggested that these subtypes may coexist within a single tumor, highlighting intratumoral heterogeneity. In summary, our study uncovers two epigenetically distinct subtypes of OS, defined by intricate regulation of RUNX2 and FOSL1, and highlights their capacity for state transitions. These findings offer a framework for subclassifying OS and identifying targeted therapies, paving the way for precision medicine approaches in this aggressive disease. Citation Format: Eunice Lopez Fuentes, Andrew S. Clugston, Alex Lee, Leanne Sayles, Alejandro Sweet-Cordero. Epigenetic subtypes and cellular plasticity are driven by an interplay between RUNX2 and FOSL1 in osteosarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1179.