From niche dynamics to drug targets: the role of cancer-associated fibroblasts in breast cancer

Cancer-associated fibroblasts (CAFs) are central architects of the tumor microenvironment (TME), influencing cancer progression, therapeutic response, and immune regulation. This thesis explores CAF heterogeneity and functional specialization in breast cancer through integrated single-cell transcriptomics, spatial proteomics, and functional studies.High-resolution analysis of murine and human datasets revealed distinct CAF substates with different transcriptional programs and spatial distributions. Spatial mapping using multiplex immunofluorescence and bioimage analysis demonstrated that myofibroblastic CAFs (myCAFs) form a barrier around the tumor core, acting as a shield that prevents immune cells infiltration, whereas immunomodulatory CAFs (iCAFs) co-localize with immune cells in more peripheral niches. These findings support a model in which CAF identity is shaped by spatial context and paracrine interactions, positioning CAF niches as potential therapeutic targets.Functional studies uncovered a stromal mechanism of endocrine resistance in luminal breast cancer. CAFs selectively rewired estrogen receptor (ER)-α signaling, suppressing canonical estrogen-responsive genes while maintaining pathways linked to invasion, basal-like differentiation, and drug resistance. Co-culture experiments and a xenograft model confirmed that CAFs confer estrogen-independent growth, and pharmacological screening identified TGF-β and JAK signaling as actionable nodes to counteract CAF-induced endocrine resistance.Finally, we identified a tumor-promoting CAF subset expressing PTGFRN (CD315), a regulator of prostaglandin F2α (PGF2α) signaling. PTGFRN+ CAFs exhibited transcriptional programs associated with ECM remodeling, angiogenesis and EMT, and their gene signature correlated with poor prognosis. Genetic and pharmacological modulation of PGF2α signaling suggests that PTGFRN supports metastatic traits and CAF survival, highlighting prostaglandin signaling as a potential therapeutic target.Collectively, we demonstrate that CAF heterogeneity, spatial organization, and signaling plasticity critically shape breast cancer biology and therapeutic response, providing a rationale for precision targeting of stromal components to improve patient outcomes.