Effects of conditioned media from breast cancer cell lines on the reprogramming of normal primary fibroblasts into cancer associated fibroblast-like cells

The reprogramming of stromal fibroblasts into cancer-associated fibroblasts (CAFs) contributes to breast cancer (BC) progression through paracrine signaling that modulate extracellular matrix remodeling, angiogenesis, tumor migration and immune escape. However, it remains unclear whether and how the metastatic properties of BC cells affect the transition from normal to activated fibroblasts. We treated primary human fibroblasts with conditioned media (CM) from the highly metastatic MDA-MB-231 or the poorly tumorigenic MCF-7 cell lines. The exposure to MDA-MB-231 CM led to increased secretion of inflammatory cytokines and chemokines, including IL-6, IL-8, ICAM-1, and MCP-1, and production of extracellular matrix proteins. Consistently, we observed the upregulation of FAP, a well-known marker for BC CAFs. These effects were moderate or absent when fibroblasts were treated with MCF-7 CM. In parallel, we conducted 1H-NMR metabolomic profiling of culture media. Fibroblasts grown in MDA-MB-231 CM showed a more pronounced shift from oxidative phosphorylation to aerobic glycolysis than those cultured in MCF-7 CM. We observed increased secretion of alanine and TCA intermediates that support cancer cell feeding and favor immune escape mechanisms. All these metabolic changes are concomitant with c-MYC upregulation. Altogether, our findings suggest that soluble factors secreted by highly aggressive BC cells promote a functional reprogramming of normal fibroblasts toward a pro-tumorigenic, inflammatory phenotype that can eventually lead to immune evasion and drug resistance. The elucidation of the symbiotic crosstalk between BC cells and their tumor microenvironment holds promise to open new avenues for targeted therapeutic interventions.