Abstract
Breast cancer is a major health concern worldwide and is the leading cause of cancer-related death among American women. Traditional therapies, such as surgery, chemotherapy, and radiotherapy, are usually ineffective. Furthermore, cancer recurrence following targeted therapy often results from acquired drug resistance. Therefore, more realistic tumor models than monolayer cell culture for drug screening and discovery in an in vitro setting would facilitate the development of new therapeutic strategies. Toward this goal, we first developed a simple, rapid, low-cost, and high-throughput method for generating uniform multi-cellular tumor spheroids (MCTS) with controllable size. Next, biomimetic cryogel scaffolds fabricated from hyaluronic acid (HA) were utilized as a platform to reconstruct breast tumor microtissues with aspects of the complex tumor microenvironment in three dimensions. Finally, we investigated the interactions between the HA-based cryogels and CD44-positive breast tumor cells, individually or as MCTS. We found that incorporating the adhesive RGD peptide in cryogels led to the formation of a monolayer of tumor cells on the polymer walls, whereas MCTS cultured on RGD-free HA cryogels resulted in the growth of large and dense microtumors, more similar to native tumor masses. As a result, the MCTS-laden HA cryogel system induced a highly aggressive and chemotherapy drug-resistant tumor model. RGD-free HA-based cryogels represent an effective starting point for designing tumor models for preclinical research, therapeutic drug screening, and early cancer diagnosis.