Flow-on-repellent biofabrication of fibrous decellularized breast tumor-stroma models

Abstract

On-the-fly biofabrication of reproducible 3D tumor models at a pre-clinical level is highly desirable to level-up their applicability and predictive potential. Incorporating ECM biomolecular cues and its complex 3D bioarchitecture in the design stages of such in vitro platforms is essential to better recapitulate the native tumor microenvironment. To materialize these needs, herein we describe an innovative flow-on-repellent (FLORE) 3D extrusion bioprinting technique that leverages expedited and automatized bioink deposition onto a customized superhydrophobic printing bed. We demonstrate that this approach enables the rapid generation of quasi-spherical breast cancer-stroma hybrid models in a mode governed by surface wettability rather than bioink rheological features. For this purpose, an ECM-mimetic bioink comprising breast tissue-specific decellularized matrix in the form of microfiber bundles (dECM-μF) and photocrosslinkable hyaluronan (HAMA), was formulated to generate triple negative breast tumor-stroma models. Leveraging on the FLORE bioprinting approach, a rapid, automated, and reproducible fabrication of physiomimetic breast cancer hydrogel beads was successfully demonstrated. Hydrogel beads size with and without dECM-μF was easily tailored by modelling droplet deposition time on the superhydrophobic bed. Interestingly, in heterotypic breast cancer-stroma beads a self-arrangement of different cellular populations was observed, independent of dECM-μF inclusion, with CAFs clustering overtime within the fabricated models. Drug screening assays showed that the inclusion of CAFs and dECM-μF also impacted the overall response of these living constructs when incubated with gemcitabine chemotherapeutics, with dECM-μF integration promoting a trend for higher resistance in ECM-enriched models. Overall, we developed a rapid fabrication approach leveraging on extrusion bioprinting and superhydrophobic surfaces to process photocrosslinkable dECM bioinks and to generate increasingly physiomimetic tumor-stroma-matrix platforms for drug screening.