Novel, biologically relevant, in vitro model of the blood-brain tumour barrier (BBTB)

Why did we fund this project?

This award aims to establish an in vitro model of the blood-brain tumour barrier to replace some rodent xenograft studies.

Glioblastoma (GBM) is the most common form of brain cancer, but treatment has not improved significantly in the past 20 years and life expectancy for newly diagnosed patients is 12 to 14 months. The blood vessels in the brain (the blood-brain barrier or BBB) tightly regulate molecules crossing from the blood stream. The BBB is often altered by a GBM tumour, creating a blood-brain tumour barrier (BBTB), which can block drugs from reaching cancer cells. The BBTB is not currently well represented in experimental models. Existing in vitro models consist of 2D cultures which do not reflect the complex structure of the BBTB and can use non-brain derived cells, which have limited relevance to human physiology. In vivo experiments involve implanting GBM cells into immunocompromised mice and monitoring tumour growth and pathophysiology, but these may not mimic the clinical disease. The student, with Dr David Dickens, aims to include GBM cells in a previously developed 3D model of the BBB, which also includes stem cell derived brain-like endothelial cells and astrocytes.

Glioblastoma (GBM) is the commonest primary malignant brain tumour. Treatment involves surgery, radiotherapy and chemotherapy, but outcomes remain very poor. In the past 5-10 years, there have been several failed phase III trials of targeted therapies. The reasons for the lack of progress are multifactorial but includes the challenges of delivering drugs across the blood-brain barrier (BBB) / blood-brain tumour barrier (BBTB). GBM tumours alter the vasculature, and this altered barrier is known as the BBTB.

Current in vitro models of the BBTB are limited with very restricted relevance so rodent xenograft models for GBM/BBTB research are the go-to method for investigating drug delivery and efficacy, and basic biology of the BBTB. Novel drugs that work on GBM xenograft models have not been successful in the clinic. This suggests that the current xenograft models do not represent the human disease, in particular the BBTB. Our collaborators at the Open University have recently established a novel BBB model based on deriving brain-like endothelial cells (BECs) from iPSCs and co-culturing with astrocytes on a hydrogel. This model has been fully validated and forms a tight monolayer with expression of key BBB proteins. This validated model of the BBB is key, as the first method for deriving BECs from iPSCs has recently been shown to generate the incorrect cell type. A 3D Hydrogel has been utilised in this model to provide a more realistic microenvironment for the co-cultured cells. The monolayer of BECs on top of the hydrogel transwell culture provides an ideal model where the barrier properties and function can be investigated directly.

The objective of this proposal is to establish and biologically validate a novel in vitro model of the BBTB by inclusion of GBM cells with the BBB model. Our collaborator at University of Edinburgh has provided several well-defined cell lines for this, including neural stem cells (NSCs) and patient derived GBM stem-like cells (GSCs). The inclusion of the GBM cells with astrocytes in the hydrogel component of the model enables the establishment of an in vitro BBTB and a way to study the effects of the GBM cells on the BECs in comparison to the normal BBB with control NSCs. Barrier tightness, astrocyte endfeet, gene expression, drug crossing and tight junction protein expression will be assessed. Finally, using transcriptomics, the novel in vitro BBTB model will be compared to GBM patient datasets of the BBTB to define the biological relevance of the in vitro model to the human disease. This is a key step in the validation of the model. The impact of this model will be to reduce animal usage significantly and in the longer-term have the potential model to replace GBM xenograft models.

Our collaborator uses 1500 mice in GBM xenograft work each year. We estimate an approximate use of at least 4,500 rodents a year for GBM xenograft experiments in the UK. For the worldwide usage of GBM xenograft rodents, a search shows nearly 800 primary publications published in the last year that used GBM xenograft models. From this we estimate at least 60,000 rodents are used per year worldwide for GBM xenograft studies. A well validated in vitro model of the BBTB would lead to a reduction in the use of GBM xenograft studies as this in vitro model would in many cases be used first to study the crossing of new therapeutics and/or to investigate the biology of the BBTB. This would lead to an estimated 20% reduction in animal work which would lead to around 12,000 rodents per year that would not be used in GBM research.

This project will generate a validated BBTB model that can be rolled out to GBM labs thus leading to the reduction in the use of GBM xenograft models and improved translation to the human disease.