Pre-metastatic niche drives breast cancer invasion by modulating MSC homing and CAF differentiation

The extent to which cancer-associated alterations in extracellular matrix stiffness influences the crosstalk between cancer cells and mesenchymal stem cells (MSCs) remains unclear. By analyzing multiple singlecell RNA sequencing datasets, we establish the existence of a cell sub-population co-expressing MSC and cancer associated fibroblast (CAF) markers in highly aggressive triple-negative breast cancers in primary tumor, secondary sites, and in circulatory tumor cell clusters. Using hydrogels of varying stiffness corresponding to different stages of cancer progression, we show that on pre-metastatic stroma mimetic 2 kPa gels, MDA-MB-231 breast cancer cell secreted conditioned media drives efficient MSC chemotaxis and induces stable CAF differentiation in a TGF{beta}/contractility-dependent manner. In addition to enhancing cancer cell proliferation, 2 kPa CAFs maximally boost local invasion and confer resistance to flow-induced shear stresses. Together, our results suggest that homing of MSCs at the pre-metastatic stage and their differentiation into CAFs actively drives breast cancer invasion and metastasis.