(483f) Structurally Decoupled Hyaluronic Acid Hydrogels to Study Matrix Metalloproteinase-Mediated Invasion of Metastatic Breast Cancer Spheroids

Polymeric hydrogels have been extensively employed to study cancer cell-matrix interactions as they provide cancer cells with a relevant three dimensional (3D) context evidenced in vivo and enabling us to maintain the cellular phenotype in vitro. The ability to tune hydrogel properties has enabled recapitulation of several key aspects of the tumor microenvironment in vitro. In the context of breast cancer, it is known that cancer cells invade the tissue at the primary and metastatic site by degrading the native extracellular matrix (ECM) using matrix-metalloproteinases (MMPs) resulting in disease progression. Efforts have been made to model MMP-mediated invasion of cancer cells by incorporating MMP-cleavable crosslinks in the hydrogel structure, however, to our knowledge, model systems enabling effective decoupling between hydrogel mechanical properties and mesh size, while incorporating MMPs into the hydrogel matrix have not been reported.

To address this need, we fabricated a structurally decoupled hyaluronic acid (HA) based 3D biomimetic model to specifically investigate the invasion of metastatic breast cancer cells mediated by MMPs. The hydrogels were fabricated using varying ratios of biologically sensitive (i.e., MMP cleavable peptide) and insensitive crosslinkers (i.e., Dithiothreitol (DTT) or polyethylene glycol dithiol (PEGDT)) to investigate the impact of incorporated MMP-cleavable peptides on the invasion of encapsulated MDA-MB-231Br metastatic breast cancer spheroids. We found that HA hydrogels crosslinked with various ratio of DTT/MMP or PEGDT/MMP exhibited comparable mechanical and physical properties as tested via rheological measurements, swelling ratio analysis, estimated mesh size, and permeability measurements. However, their degradation rate in the presence of collagenase enzyme was significantly altered and directly related to the concentration of MMP-cleavable peptide used to crosslink the hydrogel. Consistent with this, encapsulated MDA-MB-231Br spheroids in HA hydrogels with MMP sensitivity showed more invasiveness than those without MMP after 14 days of culture. Further, F-actin staining revealed invaded cells with a well-developed actin cytoskeleton and presence of invasive protrusions at the periphery of spheroids within HA hydrogels containing MMP cleavable peptides as opposed to those without MMP-cleavable peptide incorporation. Overall, these structurally decoupled HA hydrogels provide a platform to study MMP-mediated invasion of metastatic breast cancer spheroids in vitro.