Manufacturing of bone-forming callus organoids by introducing automation and at-line monitoring

Bone fractures are among the most occurring traumas worldwide. The regenerative capacity of our bones is sufficient to heal most fractures. However, 5-10% of all bone fractures do not heal properly and thus form a pressing clinical and societal problem. The field of cell based tissue engineering aims to offer a therapeutic solution for these patients by transplantation of live cells in bioartificial tissues. However, the complexity of the biological system and the empirical nature of traditional tissue engineering research hampers the translation into robust industrial processes. Therefore, the field is shifting towards a biomimetic approach following the developmental engineering paradigm. This is an approach for an in vitro process design where sequential subprocesses mimic in vivo developmental stages. The subprocesses follow a gradual and modular progression of tissue growth and cell differentiation from cells into intermediate tissues. For example, the developmental process of endochondral ossification follows a set of distinct steps from mesenchymal stem cells towards cartilage-intermediate chondrocytes that is gradually vascularized and replaced by bone. Interestingly, a modular developmental engineering strategy allows for a rational robust and reproducible manufacturing process according to quality by design (QbD) principles. The first advantage of this strategy is the possibility for modular upscaling and automation using robotics. During the research phase, protocols are often only performed on a small experimental scale by specialized personnel. Despite promising results, the hurdle towards clinical translation is often too high. This can be adressed by an early integration of robotics, especially in those steps that are highly handler dependent and time consuming. A second important aspect of QbD in cell therapy is the integration of process analytical technology (PAT) for continuous monitoring of biological processes. Currently, monitoring of culture conditions is based on pH, temperature, etc. or general metabolites (CO2, O2, lactate, glucose, etc.). These measurements can only loosely be linked to cell phenotype, meaning that these "low specificity" measurements do not reveal information related to cell identity or maturation status. This information gap can be filled by developing and adopting technologies that can accurately measure target panels of secreted proteins in the culture medium. This project aims to lower the translational hurd