The tumor microenvironment (TME) is comprised of cellular and acellular components that dynamically interact with cancer cells. Accumulating evidence shows that these interactions significantly influence critical steps, including migration and extracellular matrix (ECM) remodelling, in the invasion-metastasis cascade of events. The 3D collagenous hydrogel-based system presented in this study provides a platform for studying the cell-cell and cell-matrix interactions in the tumor microenvironment (TME).

Excessive deposition of ECM components such as collagen and hyaluronan (HA) is one of the key indicators of cancer progression. Furthermore, overproduction of hyaluronan in the TME leads to the collapse of the blood vessel and inhibits the transport of drugs to the TME, thereby causing poor prognosis. This study examined the biophysical effects of concentration and molecular weight of HA on cancer cell migration and ECM remodelling to evaluate the potential therapeutic intervention of degrading HA in the TME. The study showed that HA degradation increased cell migration since migration speed was higher in the presence of HAs of lower molecular weights. Additionally, the elastic modulus and pore size of the matrix increased with the concentration and molecular weight of HA. Cell migration on 2D substrates has been shown to follow a biphasic response. Our study showed that the steric and dimensional factors in 3D are dominant, particularly in the presence of larger molecular weight HA, that presented significant hindrance to cell migration. Additionally, ECM compaction increased with HA in the gels. Cell contractility increased with the elastic modulus, and as a result, the degree of compaction was higher in gels with higher concentrations and molecular weights of HA.

Tumor-associated macrophages (TAMs) are immune cells found in abundance in the TME. TAMs have been shown to express proteases involved in ECM remodelling. Our study showed that macrophages enhanced ECM remodelling by cancer cells in co-culture studies. While HA increased remodelling by single-cell mediated compaction by MDA-MB-231 cells, it did not affect compaction among 4T1 cells that formed cell-cell junctions. The spatial organization of multi-cell clusters was studied to elucidate the influence of cell-cell adhesions in force transmission. The end-point proliferation studies showed that J774 macrophages formed small and confined clusters (< 50 mm), whereas 4T1 clusters spanned larger (approximately 150 mm). In co-culture studies, the clusters spanned about 50% larger than conditions with same number of 4T1s. Macrophages enhanced the ability of cancer cells to form cell-cell adhesions. Studies with conditioned media demonstrated that cell-cell interactions between cancer cells and macrophages are dependent on the surrounding matrix and/or dimensionality. Additionally, cell contractility is directly linked to their ability to form long-range cell network in the matrix through which forces are applied.

The composite hydrogel systems presented in this study provide a simple and adaptable platform that recapitulates the complexities of the in vivo conditions. Cell migration and ECM remodelling by cancer cells were influenced by the viscoelastic and microstructural properties of the surrounding matrix. Our study provided evidence for the ability of other cells in the TME to boost remodelling by cancer cells. Collagen-based composite hydrogel networks can be used to examine how cells orchestrate fiber reorganization and matrix remodelling under different ECM compositions. These systems can also be used to evaluate the ability of cells to adapt a pro-remodelling behavior or a pro-migratory behavior, as we observed in the presence of different molecular weights of HA.