Building research capacity with high-throughput Ultra-performance liquid chromatography high resolution Q-TOF

The analysis of biological molecules by mass spectrometry (LC-MS) enables the identification and quantification of many thousands of metabolites, lipids, and proteins in a single measurement. Over the last 10-15 years, there has been a revolution in instrument sensitivity and resolution allowing deeper insight into the molecular processes inside living cells. At Imperial College London, we have world-leading capabilities for metabolite and lipid characterisation from clinical and research samples and have pioneered workflows to image the molecular footprint of clinical tissue (such as cancer biopsies). However, we lack the technical instrumentation for similar proteomics workflows. We wish to establish a state-of-the-art multi-user platform to support health research with cutting-edge proteomics applications and small-molecule detection. Investment in a new LC-MS platform will enable the unique capacity for comprehensive molecular footprinting of clinical samples, leading to a greater understanding of the chemical reactions that give us life, and new ways of treating disease. The instrument that we have identified (EvoSep - 7600 ZenoTOF) has unique attributes to allow us to identify different classes of molecules (such as bile-acid species) and analyse thousands of proteins in a high-throughput manner (~20 minutes per sample) to support research in key areas such as neurodegeneration, cancer, nutrition, experimental medicine and anti-microbials.

Neurodegeneration: The Dementia Research Institute in Imperial College have hundreds of blood and brain samples for studying diseases such as Alzheimer's disease (AD). In particular, understanding how different genetic variants can lead to AD will be determined, in part, through proteomic analysis of >260 brains, leading to greater insight into this currently incurable disease. Other areas of priority areas include Huntington's disease and chronic pain.

Cancer: Imaging mass spectrometry of cancer tissue has been pioneered by our research group, revealing distinct regions within tumours that may cause different responses to chemotherapies. We can use a laser to dissect these distinct regions (sometimes only a few cells) and further characterise the molecular pathways by highly sensitive proteomics and metabolomic profiling. We wish to apply this to better understand cancers and their response to drugs, in particular breast and glioblastoma.

Nutrition and experimental medicine: Detecting changes in nutrient availability is essential to maintaining healthy physiology. Understanding nutrient-sensing requires detecting distinct classes of molecules (bile acids and signalling peptides) at low abundance in biological fluids. The sensitivity of the 7600 ZenoTOF will permit quantification of the low abundance of signalling peptides within blood while the unique EAD fragmentation feature will characterise bile acids to identify new relationships with gut microbiota. Gut microbiotas play a fundamental role in gastrointestinal health, this platform will contribute to understanding the interaction between gut microbiota and ourselves as well as the mechanisms behind the health benefits of intestinal microbiota transplantation.

Antimicrobial resistance: The recent pandemic has highlighted the urgent need for new antivirals. Using LC-MS, we will investigate the relationship between viruses and their host (humans) by applying technologies that "label" proteins coming into contact with the viral genetic material. In this way, we detect what the virus interacts with at a molecular level to replicate itself. Understanding this can lead to the discovery of new ways to treat viruses, by producing antivirals that disrupt these interactions. It will also support research into the development of a new non-antibiotic class of anti-microbials that disrupt how bacteria pathologically interact with our body.

To support ongoing research into neurodegeneration (Alzheimer's disease, Huntington's disease, chronic pain), nutrition (gut microbiota-host interactions, nutrient-sensing, diabetes) cancer tumour heterogeneity (combining MS-imaging-led laser-capture microdissection with deep proteomic profiling) and anti-viral/microbial discovery, we wish to establish a multi-user state-of-the-art liquid chromatography-mass spectrometry system to provide high-throughput proteomic analysis and enhanced post-translational modification and small-molecule detection. The EvoSep high-performance liquid-chromatography system combined with a 7600 ZenoTOF mass spectrometer is a technology platform that will permit proteomic analysis in <20 min per sample with high sensitivity and quantitation. Unique to the 7600 ZenoTOF is EAD fragmentation, which will provide new fragmentation data for the identification of bile acid epimers (of critical importance to understand gut microbiota-host interactions) and lipid acyl-chain double bond location. Furthermore, this soft fragmentation approach permits the analysis of labile post-translational modifications, facilitating the identification of specific amino acid localisation of phosphorylation and glycosylation. Current facilities within the Department of Metabolism, Digestion and Reproduction include transcriptomics, small-molecule (lipid/metabolites) profiling by LC-MS and NMR and pioneering developments in molecular imaging by mass spectrometry. The provision of state-of-the-art proteomics capabilities to support these ongoing activities will provide unprecedented capabilities to analyse this trinity of bio-molecules (transcriptomic, metabolome, proteome) from single samples, giving a complete molecular characterisation of clinical/research samples and driving new discoveries in biomolecular medicine.