Transcriptomic Profiling of Immune Cell-Enriched Regions in Triple-Negative Breast Cancer Using Targeted RNA Sequencing

Triple-negative breast cancer (TNBC), characterized by the lack of hormone receptors and HER2 expression, remains a challenging subtype due to the absence of effective targeted therapies. Tumor budding (TB), a histopathological feature reflecting invasive potential, has recently emerged as a valuable prognostic marker across various cancers, including TNBC. Identifying transcriptomic alterations associated with high TB is critical for understanding mechanisms of tumor progression and immune evasion. In this study, we employed TempO-Seq, a high-throughput targeted RNA sequencing technology, to profile gene expression in immune cell-enriched regions of breast cancer tissues. Our objective was to uncover immune-related biomarkers and gene signatures specific to high TB cases, with the ultimate goal of informing prognostic stratification and potential therapeutic targets in aggressive breast cancer subtypes. This study analyzed transcriptomic profiles from 15 formalin-fixed, paraffin-embedded (FFPE) breast cancer tumor specimens, including 13 triple-negative breast cancer (TNBC) cases, one HER2-positive case, and one luminal subtype. Tumor budding (TB) status was assessed histologically and used as a stratification variable. Among all samples, 12 exhibited high TB and 3 showed low TB features. Specifically, within the TNBC subset, 11 samples were categorized as high TB and 2 as low TB. Spatially distinct immune cell–enriched tumor regions were microdissected for targeted RNA sequencing using the TempO-Seq platform, enabling focused evaluation of immune-related gene expression signatures in relation to TB status.