Abstract 1172: An estrogen receptor signaling transcriptional program linked to immune evasion in human hormone receptor-positive breast cancer

T cells are sparse in hormone receptor-positive (HR+) breast cancer, potentially due to limited antigen presentation, but the mechanisms remain unclear. This study aimed to elucidate how estrogen receptor (ER) signaling (ERS) modulates immune pathways and the tumor microenvironment using bulk and single-cell RNA sequencing (scRNA-seq) data and spatial analysis via multiplexed cyclic immunofluorescence (CyCIF). Analysis of HR+/HER2- patients from TCGA and METABRIC datasets identified co-expressed gene modules linked to ERS and three key immune pathways—TNFα-induced NF-κB (TNFα/NF-κB), type I interferon response (IFN-I), and antigen presentation machinery/T cell signaling (APM/TC)—all strongly inversely correlated with ERS. scRNA-seq of 34 breast cancer samples revealed ERS activity primarily in cancer cells, while immune pathways were enriched in non-cancer cells. A subset of immune-related modules expressed in CCLE cancer cell lines was inversely correlated with ERS, suggesting a cancer cell-intrinsic mechanism of immune suppression. Bulk and single-nucleus RNA sequencing and CyCIF in 29 HR+ BC samples showed that ERS-high tumors excluded all non-cancer cell types, including T cells, linked to reduced intrinsic IFN-I activity in cancer cells. In ERS-low tumors, TNFα/NF-κB activity was associated with angiogenesis, promoting sprouting microvessels in stroma distant from the tumor core, while IFN-I activity correlated with CD163+ macrophages. These macrophages were co-localized with CD4+/CD8+ T cells, predominantly in the peritumoral stroma, where elevated PD-1 expression indicated T cell exhaustion. T cell density positively correlated with MHC class I and II expression, while ERα expression in cancer cells inversely correlated with adjacent MHC expression. ERα-dependent regulation of MHC varied by cell type, with suppression in cancer cells and macrophages but not fibroblasts or T cells. Analysis of pre- and post-treatment samples from the ACOSOG Z1031B trial showed aromatase inhibitor therapy reduced ERS and increased TNFα/NF-κB and APM/TC activity in responsive tumors, though its effect on IFN-I was less pronounced. Altogether, our study suggests ERS suppresses immune activation and excludes T cells through cancer cell-intrinsic and microenvironmental mechanisms. Combining ER inhibition with therapies targeting TNFα/NF-κB and IFN-I pathways, along with immunotherapy to activate exhausted T cells, may offer a promising strategy to overcome immune suppression and improve outcomes in HR+ breast cancer. Citation Format: Kenichi Shimada, Daniel E. Michaud, Yvonne X. Cui, Kelly Zheng, Jonathan Goldberg, Zhenlin Ju, Stuart J. Schnitt, Ricardo Pastorello, Lukas D. Kania, John Hoffer, Jeremy L. Muhlich, Nhan Hyun, Robert Krueger, Alexander Gottlieb, Adam Nelson, Carlos W. Wanderley, Gabriella Antonellis, Sandra S. McAllister, Sara M. Tolaney, Adrienne G. Waks, Rinath Jeselsohn, Peter K. Sorger, Judith Agudo, Elizabeth A. Mittendorf, Jennifer L. Guerriero. An estrogen receptor signaling transcriptional program linked to immune evasion in human hormone receptor-positive breast cancer [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 1172.