Evaluation of a 3D osteosarcoma model: Experimental studies and mathematical modelling of drug response

Osteosarcoma remains a significant clinical challenge, with poor outcomes in metastatic cases and slow progress in the development of novel therapies. In this work, a previously developed 3D osteosarcoma model, comprising bone-mimicking macroporous alginate scaffolds with hydroxyapatite particles and a perfusion bioreactor, was evaluated as a tool for anticancer drug testing. Murine osteosarcoma cells (K7M2-wt) were seeded onto the scaffolds (15× 10⁶ cells per cm³ ) and cultured statically for 1 day. In two experimental series, scaffolds were then exposed to doxorubicin (1μg cm⁻³ ) under perfusion for 1 or 3 days (40 μm s⁻¹ superficial velocity), modelling post-resection conditions with predominantly individual or loosely aggregated cells. In two additional series, the same treatments were applied after 7 days of perfusion, when compact spheroids had formed, mimicking established tumours. Histological analysis and MTT assay were used to assess cell morphology and metabolic activity. Short-term treatment had negligible effects, while prolonged exposure significantly reduced metabolic activity of individual cells, but spheroid-like structures exhibited increased doxorubicin resistance. To rationalize the results from the first two series, a mathematical model was developed describing drug transport, intracellular accumulation, cell proliferation, and cell death. The model captured the initial treatment phase, but overestimated the effect of prolonged treatment, suggesting additional resistance mechanisms. Overall, the results demonstrate the potential of this 3D model for anticancer drug evaluation.