Work

1161 Novel IL1RAP expression gradient modulates the myeloid cell phenotype and T-cell infiltration patterns of the Ewing sarcoma immune microenvironment

Year: 2026

Type: conference-paper

Abstract: Background Ewing sarcoma (EWS) is a rare, aggressive soft-tissue and bone sarcoma with overall survival below 30% for metastatic or relapsed cases. 1 Despite advances in cancer immunotherapy, success in solid tumors such as EWS has been limited. Deeper investigation of the immune tumor microenvironment (TME) in EWS may reveal barriers to immunotherapy and identify targetable pathways. One promising target is IL1RAP, an IL-1 family co-receptor induced by the pathognomonic EWS-FLI1 fusion oncoprotein and implicated in tumor progression2 and immune modulation.3 However, the heterogeneity of tumor IL1RAP expression and its effects on the immune TME are not understood.Methods We maintain a biorepository of 51 human EWS samples processed fresh for various applications, plus an additional 80 archival fixed samples. We analyzed these cases using multiparametric flow cytometry (FC) and immunohistochemistry (IHC) to identify and characterize the immune components of the EWS TME and correlate these findings to the level of IL1RAP expression in the tumor and myeloid cells. We further confirmed these findings via analysis of publicly available single-cell RNA data.Results IHC and FC analyses reveal a gradient of IL1RAP expression across EWS tumor tissue ( figure 1). This gradient was agnostic to chemotherapy exposure and disease status, but was significantly lower in soft tissue compared to bony sites (IHC H-score 30 vs. 109, n = 16, p = 0.005, figure 2). Further, low tumor IL1RAP expression significantly correlated to increased cytotoxic T-cell infiltration of the tumor (53 vs. 19 CD8+ cells/mm2 tissue, n = 14, p = 0.03, figure 2). Interestingly, the myeloid cell compartment (CD11b+) in both blood and tumor samples consistently express low levels of IL1RAP regardless of expression level in tumor cells. Single-cell RNA analyses confirmed variable IL1RAP transcript levels, and heterogeneous immune populations with a predominance of myeloid cells. Further, decreased CD3e and CD8b transcripts correlate to increased tumor IL1RAP transcript levels.Conclusions EWS’ poor response to immunotherapy was initially attributed to an immune ‘cold’ microenvironment, but our studies show an abundance of myeloid infiltration and variable infiltration of lymphocytes, suggesting dysfunctional recruitment and activation of anti-tumor immune cells. Via multiple techniques, we show that IL1RAP is variably expressed in EWS tumor cells and directly correlates with the local tissue environment. Further, we show that higher levels of tumor IL1RAP expression correlate with decreased lymphocytic infiltration. Altogether, our data support an axis of tumor IL1RAP expression, immunosuppressive macrophage phenotype, and cytotoxic CD8 + T-Cell infiltration that may mediate response to immunotherapy.Acknowledgements Thank you to our collaborators, prior laboratory members, and most importantly the patients and their families.References Miser JS, Goldsby RE, Chen Z, Krailo MD, Tarbell NJ, Link MP, Fryer CJ, Pritchard DJ, Gebhardt MC, Dickman PS, Perlman EJ, Meyers PA, Donaldson SS, Moore SG, Rausen AR, Vietti TJ, Grier HE. Treatment of metastatic Ewing sarcoma/primitive neuroectodermal tumor of bone: evaluation of increasing the dose intensity of chemotherapy--a report from the Children’s Oncology Group. Pediatr Blood Cancer. 2007;49(7):894–900.Zhang HF, Hughes CS, Li W, He JZ, Surdez D, El-Naggar AM, Cheng H, Prudova A, Delaidelli A, Negri GL, Li X, Ørum-Madsen MS, Lizardo MM, Oo HZ, Colborne S, Shyp T, Scopim-Ribeiro R, Hammond CA, Dhez AC, Langman S, Lim JKM, Kung SHY, Li A, Steino A, Daugaard M, Parker SJ, Geltink RIK, Orentas RJ, Xu LY, Morin GB, Delattre O, Dimitrov DS, Sorensen PH. Proteomic Screens for Suppressors of Anoikis Identify IL1RAP as a Promising Surface Target in Ewing Sarcoma. Cancer Discov. 2021;11(11):2884–2903.Zhu W, Peng G, Wu Y, Zhang L, He M, Xin B, Jin W, Sun H. Integrated Single-Cell Analysis Identifies IL1RAP as a Master Regulator of TAMs and a Prognostic Biomarker in Breast Cancer. Int J Mol Sci. 2026;27(4):1894.Consent Patients who provided fresh biobank tissue were consented under an IRB approved protocol. An IRB-approved protocol is utilized for accessing patient information and Pathology Archive samples.Abstract 1161 Figure 1Immunohistochemical analysis of IL1RAP in human EWS tumor sections. Gradient of IL1RAP (brown) expression across 10 cases of human EWS. Nuclei counterstained with Hematoxylin (blue)Abstract 1161 Figure 2HALO digital analysis quantification of IL1RAP IHC by Cytonuclear module (A, B) separated by tissue, and lymphocyte infiltrate density by Immune Cell module (C) divided by tumor IL1RAP expression level. P-values unpaired t-test (B) and 2-way ANOVA (C)

Source: Journal for ImmunoTherapy of Cancer

DOI: 10.1136/jitc-2026-sitc2026.1161