Molecular Alterations Associated with Cancer-Associated Fibroblast-Induced Epithelial-to-Mesenchymal Transition in Estrogen Receptor Positive Breast Cancer

Breast cancer remains the most prevalent cancer in women globally, with increasing incidences despite improved survival rates, emphasizing the need for enhanced diagnostics and treatments. Its heterogeneity is in part explained by a variety of genetic and epigenetic alterations, where particularly estrogen receptor (ER) positive breast tumors display epigenetic dysregulations. This underscores the necessity to understand the molecular mechanisms driving tumorigenesis. The dynamic interplay between cancer cells and the tumor microenvironment (TME) plays a pivotal role in tumorigenesis. As a predominant component of the TME, tumor-activated stromal fibroblasts turn into cancer-associated fibroblasts (CAFs) through the stimulation of paracrine growth factors. In turn, CAFs enhance metastasis in cancerous cells through epithelial-to-mesenchymal transition (EMT) via paracrine signaling. EMT the process in which polarized epithelial cells detach from the epithelium and acquire motile mesenchymal-like features. EMT is a severe turning point in disease progression and represents a major clinical challenge. EMT is also accompanied by epigenetic changes, and our group’s previous in silico study revealed that loss of enhancer methylation is negatively associated with expression of EMT-related genes and fibroblast infiltration. Concurrently, other in vitro studies demonstrated that CAF-derived conditioned media (CM), containing prominent EMT-inducers such as TGF-ꞵ, and potentially other essential cofactors can induce EMT in luminal breast cancer cells. In this thesis, we studied the molecular alterations arising from growing the luminal A breast cancer cell lines MCF7 and T47D in CAF-derived CM, and assessed whether this could induce EMT. Our results revealed subtle EMT induction. Epithelial genes remained mainly unaltered, with low EMT-associated gene upregulations, of a few established EMT-associated genes (SNAI1, ZEB1, FN1). This suggested the presence of a potential hybrid EMT state, where both epithelial and mesenchymal phenotypes are simultaneously exhibited. Additionally, our results showed that the CM treatment altered expression patterns, enriched in EMTassociated and immune-related pathways, with a notable robust activation in the latter. These results underscore the plasticity of EMT and CAFs’ diverse roles in the TME, including inflammation. It also suggests that CAFs influence breast cancer cells through multiple pathways, particularly immune-related ones. Potentially, these could be alternative routes to induce a partial EMT. These findings elucidate CAFs' regulatory influence on breast cancer, and how CAFs affect the heterogeneity of the disease. Further research using epigenetic applications could offer insights into previous in silico findings of our group, linking epigenetic and transcriptional mechanisms, and shedding light on how CAFs influence enhancer methylation during EMT.