Abstract C049: Inhibition of FOXM1 as an immunomodulatory therapeutic strategy for pediatric sarcomas

Abstract Purpose: We aim to evaluate inhibition of the oncogenic transcription factor, FOXM1, in rhabdomyosarcoma (RMS) and Ewing sarcoma (EWS), focusing on its therapeutic potential and role in immunomodulation. Methods: FOXM1 expression profiles of RMS and EWS tumors were examined using publicly available microarray and bulk RNA sequencing datasets with integrated clinical metadata. Survival associations were assessed using Kaplan–Meier analyses, log-rank tests, and Cox regression. FOXM1 expression profiles across cellular subtypes and disease states were characterized by leveraging single-cell RNA sequencing analyses of patient tumors and PDXs processed using the Seurat workflow with clinical metadata integration. Immunomodulatory effects of FOXM1 were assessed using MAST (Model-based Analysis of Single-cell Transcriptomics) to enable identification of differentially expressed genes and pathways in tumors exhibiting high FOXM1 expression. FOXM1 knockdown cell lines were generated by engineering RMS cells to express FOXM1-targeting shRNA under a doxycycline-inducible promoter. Knockdown was assessed via qRT-PCR and western blotting. Cellular viability and proliferation upon FOXM1 inhibition were evaluated via cell counting kits and CFU assays. Effects of FOXM1 inhibition in-vivo were assessed via tumor studies in C57BL/6 mice with and without FOXM1 inhibition. Results: Elevated FOXM1 expression in tumors correlates with worse overall survival in RMS and EWS. Significantly increased FOXM1 expression is also found in metastatic vs primary tumors, and in recurrent vs diagnostic biopsies. In RMS, FOXM1 expression is elevated in PAX3::FOXO1 fusion positive tumors, a disease subtype associated with poor prognosis, compared to fusion negative tumors. In EWS, FOXM1 expression appeared higher in proliferating vs nonproliferating tumor subsets. Gene set enrichment analyses revealed immunosuppressive signatures in tumor cells with high FOXM1 expression, including downregulation of unfolded protein response as well as TNFa and IFNg signaling pathways. The engineered FOXM1 knockdown cell lines exhibited a substantial decrease in FOXM1 mRNA and protein levels upon doxycycline induction of shRNA expression. Genetic inhibition of FOXM1 resulted in a significant reduction in cellular viability and proliferation compared to controls in vitro, and mediated a decrease in tumor size compared to controls in-vivo. Conclusions: We demonstrate that elevated FOXM1 expression in RMS and EWS tumors is associated with poor clinical outcomes, including worse survival, tumor metastases, and disease recurrence. We also show that tumors with high FOXM1 expression exhibit immunosuppressive signatures conducive to tumor survival. We then establish an inducible genetic knockdown model, illustrating that FOXM1 inhibition resulted in decreased cellular viability and proliferation in-vitro and tumor size in-vivo. Altogether, we provide support for targeting FOXM1 and its role on immunomodulation as a promising therapeutic approach for pediatric sarcomas such as RMS and EWS. Citation Format: Tiffany Y. Juan, Jonathan Do, Tanya Kalin, Vladimir Kalinichenko, Ying-Wei Lan, Xiaomei Xia. Inhibition of FOXM1 as an immunomodulatory therapeutic strategy for pediatric sarcomas abstract. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr C049.