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 quasispherical 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-mu F) 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-mu F was easily tailored by modelling droplet deposition time on the superhydrophobic bed. Interestingly, in heterotypic breast cancer-stroma beads a selfarrangement of different cellular populations was observed, independent of dECM-mu F inclusion, with CAFs clustering overtime within the fabricated models. Drug screening assays showed that the inclusion of CAFs and dECM-mu F also impacted the overall response of these living constructs when incubated with gemcitabine chemotherapeutics, with dECM-mu F 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-stromamatrix platforms for drug screening.

keywords

EXTRACELLULAR-MATRIX; CANCER-CELLS; RESISTANCE

subject category

Materials Science

authors

Ferreira, LP; Jorge, C; Henriques-Pereira, M; Monteiro, MV; Gaspar, VM; Mano, JF

Groups

G5 - Biomimetic, Biological and Living Materials

Projects

Produção de Plataformas Microcapsulares 3D+ para Avaliação Laboratorial em Larga Escala de Novas Terapias Combinatórias para o Cancro do Pâncreas (PANGEIA)

CICECO - Aveiro Institute of Materials (UIDB/50011/2020)

CICECO - Aveiro Institute of Materials (UIDP/50011/2020)

acknowledgements

This work was also supported by the Programa Operacional Competitividade e Internacionalizacao (POCI) , in the component FEDER, and by national funds (OE) through FCT/MCTES, in the scope of the projects PANGEIA (PTDC/BTM-SAL/30503/2017) . The authors acknowledge the financial support by the Portuguese Foundation for Science and Technology (FCT) through Doctoral Grants (SFRH/BD/141718/2018, Luis P. Ferreira, and DFA/BD/7692/2020, Maria V. Monteiro) and an assistant researcher contract (2022.02106.CEECIND) . This work was developed within the scope of the project CICECO-Aveiro Institute of Materials, UIDB/50011/2020 & UIDP/50011/2020, financed by national funds through the FCT/MEC and when appropriate co-financed by FEDER under the PT2020 Partnership Agreement.