Abstract Background Invasive Lobular carcinomas (ILC) comprise a histologically and biologically diverse group of breast cancers. While classic invasive lobular carcinomas (ICLCs) are typically characterized by cohesive, infiltrative growth and immune-excluded microenvironments, pleomorphic invasive lobular carcinomas (IPLCs) display more aggressive behavior. Based on our previous findings of differential CAF cell densities in five phenotypes between CLCs and PLCs, we aimed to investigate the spatial relationships and distribution patterns within these phenotypes, as the spatial organization of fibroblasts and immune cells in these histologic subtypes remains underexplored. Methods We performed high-dimensional multiplex immunofluorescence (mIF) spatial analysis on 12 cases of ILC (IPLCs: cases 7–12; ICLCs: cases 13–18). A comprehensive fibroblast-focused panel (including FAP, S100A4, Thy1, A-SMA) and immune markers (CD45) were used to characterize tumor (CK+), fibroblast, and immune compartments. Spatial clustering and neighborhood analysis were conducted using the Spatial Image Analysis of Tissues (SPIAT) R package v 4.5.1, to define the microenvironmental architecture. Results IPLCs demonstrated marked spatial heterogeneity, with fragmented CK+ tumor regions interspersed among stromal and immune elements. Fibroblast landscapes in IPLCs were notably diverse, featuring abundant hybrid subtypes (e.g., S100+Thy1+, A-SMA+FAP+) and frequent tumor-stroma integration within shared neighborhoods. These dynamic fibroblast populations supported active extracellular matrix (ECM) remodeling, potentially facilitating invasion and contributing to partial immune engagement. CD45+ immune cells were consistently more prominent in IPLCs, forming discrete immune-enriched clusters and mixed neighborhoods, suggesting an immune-intermediate or partially inflamed phenotype. In contrast, ICLCs displayed large, cohesive CK+ tumor regions with minimal fragmentation and low spatial heterogeneity. Fibroblast subtypes were simpler and predominantly peripherally distributed, and they rarely formed hybrid phenotypes. The stroma primarily acted as structural support, with limited ECM remodeling and minimal immune modulation. CD45+ cells were sparse and diffusely scattered, consistent with an immune-excluded (“cold”) microenvironment. Conclusions While descriptive, our data reveal profound differences in the spatial microenvironments of IPLC and ICLCs. IPLCs exhibit a dynamic, heterogeneous stroma characterized by integrated fibroblast-tumor interactions and increased immune infiltration, aligning with their aggressive clinical behavior. In contrast, ICLCs maintain a cohesive tumor architecture with simplified, peripheral stroma and sparse immune presence. These insights highlight the significance of spatial context as an innovative and informative strategy in comprehending ILC biology and suggest that stromal and immune features may inform therapeutic strategies, including potential vulnerability to stroma- or immune-targeted interventions in IPLCs Citation Format: E. N. Barrientos-Toro, R. Pandurengan, H. Batra, Q. Ding, A. Contreras, A. A. Sahin, C. Haymaker, M. G. Raso. A Novel Spatial Profiling Approach Reveals Distinct Fibroblast and Immune Architectures in Invasive Pleomorphic versus Classic Lobular Carcinomas abstract. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS3-12-21.
Barrientos-Toro et al. (Tue,) studied this question.