Investigating the Mechanobiology of Cancer Cell-ECM Interaction: The Impact of Substrate Stiffness in Breast Cancer Progression

The loss of tissue homeostasis and mechanoreciprocity are considered one of the hallmarks of cancer. Here we have applied a porous type I collagen-based three-dimensional (3D) scaffold to study how breast cancer cells interact with and alter extracellular collagen. We assessed the modifications induced by tumor cells on the micro- and macro- characteristic of extracellular collagen and on its compressive stiffness. Mechanical testing was conducted both with an in-house built low-force compressive device and by Dynamic Mechanical Analysis. The stiffness of single collagen fibers was assessed by Atomic Force Microscopy. When cultured on collagen scaffolds, the two cell lines generated coherent tissue-like structures. MCF7 displayed an epithelial morphology with a tightly cohesive cobblestone appearance, while MDA-MB-231 showed a mesenchymal phenotype with lower cell-to-cell contact. MDA-MB-231, which belongs to the aggressive basal-like subtype, increased scaffold stiffness from 46.9 kPa, to 57.9 kPa, and overexpressed the matrix-modifying enzyme, lysyl oxidase (LOX), whereas luminal A MCF-7 cells did not alter the mechanical characteristics of extracellular collagen. When the activity of LOX was blocked, MDA-MB-231 were unable to alter the scaffold stiffness: the compressive modulus increased by 8.9%, in contrast to the increase observed without LOX inhibition (23%). No significant changes were found between the Young’s modulus of fibers taken from control scaffolds compared to fibers taken from scaffolds after culture with MDA-MB-231. Overall this work provides evidence that invasive, mesenchymal-like breast cancer cells produce high levels of the crosslinking enzyme LOX, and are able to increase the stiffness of extracellular collagen and to alter its structural characteristics. A causal relationship between this behavior of cancer cells and expression of the enzyme LOX was also provided. Our model offers a relevant in vitro tool to reproduce and investigate the biomechanical interplay subsisting between cancer cells and the surrounding ECM.