(247d) Chemo-Mechanically Tunable Antibiotics-Based Hydrogel for High Throughput Generation of 3D Tumor Microenvironments (3DTMs)

Despite their popularity, two dimensional cell cultures fail to capture the inherent complexity within a 3D tumor. Three dimensional co-culture systems are effective in vitro models of the tumor microenvironment, since they can better represent cell-cell contacts and interactions, existence of metabolic and nutrient gradients, necrotic core regions, and cellular resistance to therapeutics. Specifically, co-culturing epithelial cancer cells with their stromal/stellate counterparts provides insights into tumor-stromal interactions, cell-ECM interactions, epithelial-mesenchymal transition, and their role in tumor proliferation and metastasis. Available techniques to generate high throughput 3DTMs include hanging drop technique, rotary suspension and non-adhesive substrates for liquid overlay including Agarose, and Matrigel. However, these methods require continuous refreshing of media, and result in spheroids with highly polydisperse sizes.

Here, we describe the discovery and extensive characterization of an antibiotic-based hydrogel whose formation kinetics and chemo-mechanical properties were tunable based on temperature and on the molar ratios of the monomers.  These hydrogels extensively facilitated 3DTM formation across multiple cell lines including those of prostate, breast, and pancreatic cancer cells co-cultured with stromal or stellate cells. Tunable chemo-mechanical properties resulted in the formation of one 3DTM per well, thereby providing exquisite control over size and heterogeneity. Further, 3DTM size was easily controlled by simply plating different cell numbers. These 3DTMs were characterized for cell viability using fluorescence stains and H & E staining, morphology using scanning electron microscopy, cell cycle profiles using flow cytometry, and response to chemotherapeutic drugs and their combinations. Taken together, these novel antibiotic-based hydrogels facilitate high-throughput generation of custom designed tumor microenvironments that can be used as in vitro tumor models for screening drugs and delivery systems, and for fundamental studies on tumor biology.