3D in vitro model of osteosarcoma: advantages and limitations of two distinct approaches

In the realm of anti-cancer drug development, 3D cell culture models in vitro have emerged as promising alternatives to conventional 2D monolayer and animal models due to their physiological relevance and potential for personalized medicine. However, existing models primarily focus on soft tissue cancers, neglecting the complexities of hard tissue cancers such as osteosarcoma. This thesis aims to address this gap by developing a robust, tuneable, and reproducible in vitro model of osteosarcoma that mimics the complexity of bone tissue and the heterogeneity of the tumour microenvironment while integrating mechanical stimuli that are characteristic of the musculoskeletal system. The main objectives include optimizing cell spheroid generation methods, developing a synthetic scaffold mimicking bone tissue, evaluating the scaffold cytocompatibility and performance in dynamic culture systems, and assembling a tri-culture 3D in vitro model applicable for drug testing. Chapter 1 discusses the importance of a cell spheroid-based approach as the first step from 2D to 3D and presents a method for their generation applicable to the osteosarcoma cell line of choice. Chapter 2 focuses on the development of freeze-dried alginate-based porous scaffolds, their characterization, application, and limitations. Chapter 3 investigates 3D-printed tricalcium phosphate scaffolds via material characterization and biological evaluation with emphasis on primary human bone-marrow-derived mesenchymal stem cells, as well as evaluates scaffold performance in various bioreactor systems. Chapter 4 discusses the assembly of the described components into a tri-culture in vitro model in a perfusion bioreactor, culminating in the testing of an anticancer compound. This thesis contributes to the advancement of in vitro modelling of skeletal diseases on the example of osteosarcoma, offering an approach for designing a platform aimed at more accurate drug screening and personalized treatment strategies.