Development of a Hybrid Nanoink for 3D Bioprinting of Heterogeneous Tumor Models

Despite the rapid progress in applying three-dimensional (3D) printing in the field of tissue engineering, fabrication of heterogeneous and complex 3D tumor models remains a challenge. In this study, we report a hybrid nanoink (AGC) composed of alginate, gelatin methacryloyl (GelMA), and cellulose nanocrystal (CNC), designed for multinozzle microextrusion 3D printing of tumor models. Our results show that the ink consisting of 2 wt % alginate, 4 wt % GelMA, and 6 wt % cellulose nanocrystals (AGC246) possesses a superior shear-thinning property and little hysteresis in viscosity recovery. The fabrication of a colorectal cancer (CRC) model is demonstrated by printing a 3D topological substrate with AGC246 and then seeding/printing endothelial (EA-hy 926) and colorectal carcinoma (HCT 116) cells on top. Direct seeding of cells by dropping a cell suspension onto the 3D substrate with distinctive topological features (villi and trenches) deemed inadequate in either creating a monolayer of endothelial cells or precise positioning of cancer cell clusters, even with surface treatment to promote cell adhesion. In contrast, 3D biopinting of a CRC model using cell-laden AGC153, coupled with dual ultraviolet (UV) and ionic cross-linking, is shown to be successful. Hence, this study brings advancements in 3D bioprinting technology through innovative material and methodology designs, which could enable the fabrication of complex in vitro models for both fundamental studies of disease processes and applications in drug screening.