Spatial analyses implicate high stromal tumour-infiltrating CD8⁺ lymphocytes as a negative predictive marker for...

Female patients with ER⁺HER2^- breast cancer have a favourable prognosis for 5-10 years. Later relapses are, however, common, yet predictions of late recurrence risk are suboptimal, particularly for patients with intermediate risk determined by the Oncotype Dx Recurrence Score (RS, 16-25). Here, we analyse tissue samples from patients with ER⁺HER2^- breast cancer using spatial proteomics (multiplex immunofluorescence with 5 markers, n = 440) and spatial transcriptomics (n = 359), and find decoupled immune states between stroma and epithelia. Moreover, inflamed stroma express genes linked to tissue remodelling, immune exhaustion, and inhibitory/checkpoint receptors (CTLA4, TIGIT, CD96); inflamed epithelia similarly express genes associated with checkpoints (CTLA4) and exhaustion (CXCL13), but also genes attributed to antigen presentation. In our randomised, Intermediate RS cohort treated with chemotherapy we observe an association between higher stromal tumour-infiltrating CD8⁺ lymphocyte (sTIL CD8⁺) density and poor outcome (ΔLR-χ²: 6.79, p = 0.009), which we validate using data from whole-resection specimens (ΔLR-χ²: 8.90, p = 0.003). Our data thus provide insights into the immune states in ER⁺HER2^- breast cancer, and propose sTIL CD8⁺ density as candidate biomarker for treatment decisions.