Bioengineered Submucosal Organoids for In Vitro Modeling of Colorectal Cancer.

The physical nature of the tumor microenvironment significantly impacts tumor growth, invasion, and response to drugs. Most in vitro tumor models are designed to study the effects of extracellular matrix (ECM) stiffness on tumor cells, while not addressing the effects of ECM's specific topography. In this study, we bioengineered submucosal organoids, using primary smooth muscle cells embedded in collagen I hydrogel, which produce aligned and parallel fiber topography similar to those found in vivo. The fiber organization in the submucosal organoids induced an epithelial phenotype in spheroids of colorectal carcinoma cells (HCT-116), which were embedded within the organoids. Conversely, unorganized fibers drove a mesenchymal phenotype in the tumor cells. HCT-116 cells in organoids with aligned fibers showed no WNT signaling activation, and conversely, WNT signaling activation was observed in organoids with disrupted fibers. Consequently, HCT-116 cells in the aligned condition exhibited decreased cellular proliferation and reduced sensitivity to 5-fluorouracil chemotherapeutic treatment compared to cells in the unorganized construct. Collectively, the results establish a unique colorectal tumor organoid model to study the effects of stromal topography on cancer cell phenotype, proliferation, and ultimately, chemotherapeutic susceptibility. In the future, such organoids can utilize patient-derived cells for precision medicine applications.