Osteomimetic matrix components alter cell migration and drug response in a 3D tumour-engineered osteosarcoma model.

Osteosarcoma management continues to lack the appropriate prognostic tools to assign personalised treatment. This leaves non-responders to standard care vulnerable to recurring disease and pulmonary metastases. Developing 3D in vitro disease models to serve as a test bed for personalised treatment is a promising approach to address this issue. This study describes the generation of 3D osteosarcoma models termed "tumouroids", which are geometrically compartmentalised to reproduce the bone cancer mass and its surrounding. Although the tumour microenvironment impacts osteosarcoma in many ways, this model focussed on interrogating the influence of a biomimetic matrix on tumour cell behaviour. The 3D matrix was supplemented with the bone-marrow proteins laminin, fibronectin and NuOss® bone granules. This led to increased invasion of osteosarcoma cell aggregates from within the bone-like matrix into the surrounding acellular bone marrow-like ECM. The presence of bone granules also yielded an atypical molecular profile of osteosarcoma cells, suggesting malignant metabolic reprogramming. Changes include decreased MMP-9 (p < 0.05) and increased PTEN (p < 0.05), MCP-1 (p < 0.01) and MCT-4 (p < 0.05) gene expression. This complex 3D biomimetic composition also changed cellular responses to doxorubicin, a common chemotherapeutic agent used to treat osteosarcoma, and reproduced key issues of in vivo treatment like drug penetrance and doxorubicin-induced bone toxicity. This work highlights the importance of a biomimetic matrix in 3D osteosarcoma models for both basic and translational research. STATEMENT OF SIGNIFICANCE: This study describes the generation of 3D osteosarcoma models termed "tumouroids", which are geometrically compartmentalised to reproduce the bone cancer mass and its environment. Utilising this novel model, specific parameters of osteosarcoma growth and invasion were investigated. Osteosarcoma cell lines proliferate at a slower rate, exhibit malignant metabolic reprogramming, and respond to drug intervention at lower concentrations of doxorubicin hydrochloride in matrix-complex compared to basic tumouroids. As such, this study provides evidence that the tumour microenvironment impacts osteosarcoma in many ways. The osteosarcoma tumouroid described herein may form the basis of a personalised-medicine strategy, which will allow the testing of drug effectiveness similar to that used for antibiotic selection for pathogenic bacteria. Crown