DELINEATING INTRA-TUMOUR METABOLOMIC AND PHOSPHO-PROTEOMIC HETEROGENEITY IN PATIENT GLIOBLASTOMAS THROUGH ADVANCED ANALYTICAL METHODS

BACKGROUND: Molecularly targeted approaches informing next-generation glioblastoma (GBM) chemotherapy must account for the intra-heterogeneous genome landscape and aim to eradicate invasive residual sub-populations. We outline an analytical strategy that forms a functional genomic corollary to existing genome-wide mutation and gene expression studies. METHODS: Biopsies from spatially distinct intra-GBM regions were surgically resected via Stealth and 5-ALA fluorescence guidance, including core, enhancing and invasive margin regions. Cellular intra-tumour heterogeneity (ITH) was confirmed by immunohistochemistry and qPCR arrays, whilst molecular analytics were conducted using liquid chromatography-mass spectrometry (metabolomic/lipidomic) magnetic resonance spectroscopy (metabolic) and SDS PAGE phosphoprotein enrichment (proteomic). RESULTS: Cellular ITH was confirmed via distinct neural stem cell expression in medial and posterior rims relative to the central enhancement region. Metabolites present in the invasive margin (5-ALA-determined), central enhancement and peri-necrotic core were distinct within each GBM. Controlling for the high proportion of normal brain cells surrounding the invasive margin, high tumour-specific glutamine levels were consistently present. Relative abundance of cytosolic phosphoproteins distinguished active protein pathways within intra-GBM regions, including invasive margin. Metabolomic network analyses confirmed that GBM residual disease is distinct from core ITH regions with respect to metabolic dependence. CONCLUSIONS: This innovative concept is the first to address ITH with multi-region biopsies using advanced analytical bioscience techniques. Identification of metabolites, lipids and phospho-proteins that are functional determinants of GBM infiltration reveal putative molecular targets on residual disease cells and represent a route to expedite new chemotherapy agents to clinic within a precision medicine framework.