Single-cell profiling reveals a novel CAF subpopulation linking stromal heterogeneity to immune suppression in breast cancer subtypes

Vigiano Benedetti, Virginia; Conte, Federica; Santoni, Daniele; Pentimalli, Tancredi Massimo; Bei, Roberto; Cifaldi, Loredana; Barillari, Giovanni; Spallotta, Francesco; Cencioni, Chiara; Melaiu, Ombretta

Single-cell profiling reveals a novel CAF subpopulation linking stromal heterogeneity to immune suppression in breast cancer subtypes

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Item Type:	Article
Title:	Single-cell profiling reveals a novel CAF subpopulation linking stromal heterogeneity to immune suppression in breast cancer subtypes
Creators:	Vigiano Benedetti, Virginia, Conte, Federica, Santoni, Daniele, Pentimalli, Tancredi Massimo ORCID: https://orcid.org/0000-0002-8461-7918, Bei, Roberto, Cifaldi, Loredana, Barillari, Giovanni, Spallotta, Francesco, Cencioni, Chiara and Melaiu, Ombretta
Abstract:	BACKGROUND: The tumor microenvironment critically influences breast cancer (BC) progression, immune surveillance, and therapeutic response. Cancer-associated fibroblasts (CAFs), a heterogeneous stromal population, are key regulators of these processes, yet their subtype-specific contributions in BC remain insufficiently defined. METHODS: We integrated three single-cell RNA sequencing datasets from 29 BC patients to characterize stromal populations. Bulk RNA-seq data from The Cancer Genome Atlas (TCGA) were analyzed to assess correlations between CAF subsets and immune infiltration. Gene signatures were derived to identify subtype-specific CAF-immune interactions, prognostic markers, and potential predictors of chemotherapy response. RESULTS: Three conserved stromal populations (iCAFs, myCAFs, and pericytes) were identified, along with a previously unrecognized subset, the cluster 3 (CL3) CAF-like cells, referred as metabolic stressed CAF (msCAF). msCAF cells displayed transcriptional programs associated with antigen presentation, stress response, glycolysis, and extracellular matrix remodeling. Their abundance was inversely correlated with T-cell infiltration and function, in a subtype-specific manner: triple negative breast cancer (TNBC) was enriched for msCAFs in immune-infiltrated but functionally constrained microenvironments, whereas Luminal A tumors exhibited weaker immune infiltration with heterogeneous CAF-immune associations. msCAFs were characterized by a conserved gene signature (HLA-A, HLA-C, IL32, EMP3) and subtype-specific genes related to T-cell exhaustion. Several genes demonstrated prognostic relevance with distinct patterns in Luminal A (IER3, TIMP1, TBX3, SEC61G) and TNBC (ADM, C4orf3, LDHA) tumors, as well as shared biomarkers (FN1, LOXL2, P4HA1). Multiple msCAF genes also predicted chemotherapy response, suggesting utility as treatment stratification biomarkers. CONCLUSION: msCAFs represent a clinically relevant CAF subset that drives immune suppression, impacts subtype-specific prognosis, and influences therapy response in BC. These findings highlight msCAFs as promising targets for enhancing immunotherapy and personalizing treatment strategies.
Keywords:	Breast Cancer, Cancer-Associated Fibroblasts, T Cells, Immunomodulation, Single-Cell RNA-seq
Source:	Journal of Translational Medicine
ISSN:	1479-5876
Publisher:	BioMed Central
Date:	25 June 2026
Official Publication:	https://doi.org/10.1186/s12967-026-08446-x
PubMed:	View item in PubMed

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