Validation of non-animal derived products for stem cell-derived breast cancer organoid models

Cinzia will assess the utility of animal-free products – including serum free-media, synthetic matrices and non-animal derived antibodies – in a 3D breast organoid model using human induced-pluripotent stem cells for studying cancer development and progression. The funding builds on work carried out during an NC3Rs PhD studentship.

This award was made as part of the 2024 non-animal derived product validation grants supported with funding from the Department for Science, Innovation and Technology (DSIT).

This project aims to validate the use of non-animal derived products used for the differentiation of organoids derived from induced pluripotent stem cells (iPSCs) to model breast cancer in vitro. To advance the transition to animal replacement in cancer research, we have developed a human stem cell model to study Triple Negative Breast Cancer (TNBC), a cancer of unmet clinical need and lacking targeted therapies. This close-to patient model enables the study of cancer driver mutations in TNBC, without the need of using genetically modified mice. In our model, we gene-edited human induced pluripotent stem cells to carry the most frequent cancer mutations found in TNBC patients and then differentiated them into 3D mammary organoids to allow disease modelling. The differentiation of the breast cancer organoids requires culture in complex conditions in the presence of extracellular matrix, supplements and growth factors, which are typically of animal origin. Additionally, animal-derived antibodies are used to characterise the organoids to assess expression of breast lineage markers, receptor status and disease biomarkers. The vision of this project is to drive innovation in this research field by developing a completely animal-free system that will use non-animal derived culture media components, extracellular matrices and animal free antibodies. Its scope is to perform a full validation of the use of synthetic matrices for the differentiation of organoids cultured in animal-free conditions. The performance of these animal-free products will be compared to our existing standard protocol to allow a systematic validation. We estimate that the validation will reduce the use of at least 120 animals/year locally and 3,200 animals/year globally. These figures estimate the replacement of animal-derived matrices, highlighting an ever-greater impact on the replacement of other animal-derived reagents that will be also used in the project. With this validation, we aim to demonstrate that transitioning to animal-free models is feasible and that it significantly improves the quality and reproducibility of results. Moreover, we plan to share knowledge and resources to allow a wide uptake of this animal replacement model and animal-free alternatives within the scientific community, thereby creating a substantial project legacy and impact on the 3Rs.