Spatial Profiling Reveals Distinct Immune Microenvironment Specific to Premenopausal Triple Negative Breast Cancer

Background Triple-negative breast cancer (TNBC) is characterized by a lack of targeted therapies, leading to poor prognosis. Premenopausal TNBC often has different clinical outcomes compared to postmenopausal patients, influenced by their hormonal milieu and its impact on the tumor microenvironment (TME). This study used Digital Spatial Profiling (DSP) to explore these differences comprehensively. Methods We analyzed 11 treatment-naive TNBC tumors (premenopausal, n=5; postmenopausal, n=6) with long-term follow-up, using DSP with the NanoString GeoMx Cancer Transcriptomics Atlas (CTA) panel (1812 genes). Tumor and non-tumor areas were selectively analyzed to generate the transcriptomic data, followed by pathway analysis and TME assessment using the ESTIMATE algorithm and tumor immune phenotype (TIP) signature. Cell deconvolution methods were employed to identify immune subtypes and their association with disease-free survival was examined by survival analysis and further validated by immunohistochemistry (IHC) for CD4, CD8, and CD20 markers. Results Cluster analysis revealed distinct profiles based on menopausal status. In premenopausal tumors, immune regulating pathways such as neutrophil degranulation, interferon signaling, and the complement system were upregulated, while T cell checkpoint, cancer antigen, and B cell pathways were downregulated. Non-epithelial compartment showed upregulation of IL2, BCR signaling, and T cell pathways. Immune cell identification using a deconvolution-based method showed increased CD8+ T cells, endothelial cells, myeloid dendritic cells, and monocytes in premenopausal tumors. Increased intratumoral protein expression of CD8, CD4, and CD20 in premenopausal TNBC was noted, with infiltration of CD20 correlating with a better prognosis. Further transcript-based TIP signature analysis showed increased expression of immune cold signature in premenopausal tumors. Conclusion This study provides a detailed analysis of TME in TNBC, emphasizing immunological differences associated with menopausal status. Targeting these distinct immune environments, potentially by modulating hormonal effects or enhancing immune responses, may improve TNBC treatment efficacy. Further research validating these findings and exploring combination therapies could enhance immunotherapy outcomes in TNBC patients.