Abstract 3968: Spatial transcriptomic features associated with response to neoadjuvant therapy in triple-negative breast cancer

Abstract Background: The KEYNOTE-522 trial established pembrolizumab plus neoadjuvant chemotherapy as a standard for early-stage triple-negative breast cancer (TNBC) by significantly improving pathologic complete response (pCR) and event-free survival. Yet biomarkers predicting response and the biology underlying residual disease—particularly the divergent outcomes observed among patients with residual cancer burden class 2 (RCB2) from chemotherapy only and chemotherapy plus immunotherapy—remain poorly defined. We hypothesized that spatial cellular architectures at diagnosis, and their longitudinal remodeling with therapy, govern TNBC treatment response and resistance. Methods: We performed single-cell resolution spatial transcriptomic profiling on 131 diagnostic biopsies, 161 resection specimens, and 38 metastatic lesions from patients treated with neoadjuvant chemotherapy with or without pembrolizumab, including 50 paired pre/post-treatment samples for longitudinal analysis. A 60-marker spatial proteomic panel provided cross-modality validation. Spatial features, including cell-type abundance, co-localization, tumor-immune interface structure, and niche organization, were evaluated across pCR versus non-pCR groups, RCB classes (0-3), and longitudinal transitions within each therapeutic arm. Results: Across 6 pathologist-annotated histologic regions, we resolved 17 major cell types, 72 gene-defined subtypes, and 96 spatial communities. Early analyses highlight contrasting CD8+ T-cell spatial states: intra-tumoral CD8+ infiltration characterizes responders, whereas stromal-restricted CD8+ localization, together with CD33+ myeloid programs, marks non-response and T-cell exclusion. Additional cohort-specific variation was observed in fibroblast density, stromal architecture, and myeloid-lymphoid proximities. Integrating diagnostic and longitudinal spatial features, we are developing an interpretable spatial risk score to distinguish treatment response and RCB class. Conclusions: This study represents one of the largest integrated spatial transcriptomic and proteomic analyses of TNBC neoadjuvant therapy to date. Emerging patterns suggest that cytotoxic T-cell access to tumor nests marks effective anti-PD-1 response, whereas fibroblast-driven stroma and myeloid-mediated exclusion underlie resistance. These insights provide a roadmap for designing combination therapies to overcome stromal and myeloid barriers to neoadjuvant immunotherapy. Citation Format: Shiqing Liao, Teia Noel, Joseph Lownik, Pavithra Nedumaran, Yoona Yang, Richard Mebane, Aagam Shah, Andrew Martinez, Akil A. Merchant, Simon Knott. Spatial transcriptomic features associated with response to neoadjuvant therapy in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 3968.