Metabolomic profiling of neurodevelopmental programs in healthy brain which promote dysregulated neural growth

Background:
Recent evidence from neuroscientific studies have revealed that biochemical and electrochemical signalling from healthy neural cells in the brain, can foster a microenvironment which promotes dysregulated growth of neural progenitor cells. This is most strikingly exemplified by synaptic neurotransmission between neurons and immature cancerous cells in the brain which resemble neural stem/progenitor cells.
Whilst neurotransmitters such as glutamate released from healthy neuronal structures in the brain, can enhance the survival and infiltration of cancerous cells into the surrounding brain parenchyma, the underlying functional relationship remains unclear. Metabolomics is the study of chemical processes involving metabolites and is an integral technology for understanding the function of biological systems. To date however, no metabolomic studies to elucidate fully the biochemical communication between healthy and dysregulated brain cells have been conducted.

Importance of study:
For malignant brain cancers such as glioblastoma, patients are asymptomatic during early stages of disease, likely explained by cancer cells associating functionally with healthy neural cells. This means that for improved therapy, we need to study how neuroscience and aberrant neural stem cell/progenitor biology promotes phenotypes (migration and invasion) characteristic of brain cancer.

Hypotheses:
Neural communication between healthy astrocytes and dysregulated, transformed neural cells is characterised by aberrant metabolomic signatures.

Workplan:
(Months 1-24): Aim 1: Develop 3D co-cultures of human astrocytes and infiltrative glioblastoma cells which can be interrogated to reveal the underlying metabolomes.
We have established a complex 3D cellular model which recapitulates the interaction between normal brain and post-surgical glioblastoma cells. Four patient-derived glioblastoma cell lines stably transduced with the GFP transgene will be co-cultured with human astrocyte progenitors as 3D spheroids. To ensure physiologically-accurate signalling from the tumour microenvironment, 3D spheroids will be placed onto decellularized human brain extracellular matrix (ECM) to retain signalling from brain ECM ligands.
To investigate whether astrocytic-cancerous communication varies during distinct neurodevelopmental states, patient-derived glioblastoma cells will be maintained in conditions which enrich either neural stem cells or lineage-committed progenitor cells. Based on GFP transgene expression, cancer stem/progenitor cells and astrocytes will be isolated by fluorescence-activated cell sorting (FACS) upon periods of co-culture to promote biochemical cross-talk and neurotransmission. Liquid-chromatography mass spectrometry (LC-MS) will be conducted on sorted astrocytes and cancerous cells to determine the metabolome of each cell type
(Months 25-42): Aim 2: Validate results by analysing the metabolome of astrocytes and cancer stem cells isolated from surgically resected tissue of human glioblastoma patients.
5-aminolevulinic acid (5-ALA), a metabolic substrate given to glioblastoma patients prior to surgery, is converted to a fluorescence-emitting product in cancer cells only and permits the isolation of infiltrative diseased tissue which blends into healthy brain parenchyma. Tissue isolated in this manner, will be subjected to FACS to separate brain cancer and healthy astrocytic cells. LC-MS will determine metabolomic profiles which reflect a molecular snapshot when residual cancer cells interact directly with normal neurodevelopmental signals in the surrounding brain.
Outputs:
-Graduate immersed with inter-disciplinary cellular biology and analytical bioscience expertise.
-Two high-impact publications.
-Presentation at global neurobiology conference.