NISA: Novel approaches for in situ analysis of biomolecules

The aim of the research is to develop novel approaches for the analysis of biomolecules, and in particular proteins, directly from their natural (or actual) environment, i.e., to develop approaches for in situ biomolecular analysis. Proteins are the work-horses of the cell and perform all the functions required for life. They also find uses as therapeutics and in consumer products. To gain insight into the various and specific roles of proteins in life processes, or to determine the therapeutic efficacy of protein drugs, or to establish the environmental fate of protein additives in consumer products, it is necessary to be able to analyse proteins at a molecular level. Mass spectrometry, in which ionised molecules are characterised according to their mass-to-charge, is ideally suited to this challenge, offering high sensitivity, broad specificity (all molecules have a mass), and the capability for chemical structure elucidation. The ultimate goal is to link molecular analysis directly to molecular environment. Much like a forensics officer tasked with determining the presence of an illicit substance, there is much greater reliability and credibility afforded to an analysis performed at the scene of the crime than to one performed following removal of the sample to a separate location and alternative surroundings.

Growing evidence suggests in situ protein analysis has groundbreaking roles to play in biomarker discovery, diagnosis & early detection of disease, targeting therapeutics (personalised medicine) and assessment of therapeutic efficacy. The benefits of in situ protein analysis can be illustrated by considering a thin tissue section through a drug-treated tumour. In principle, in situ analysis would inform on drug-target interactions (i.e., is the drug binding to the correct protein?). Moreover, with in situ protein analysis the capacity for artefact introduction as a result of sample preparation (e.g., application of a matrix) or sample damage is eliminated.

Nevertheless, a number of challenges exist. Proteins are large molecules associated with a vast array of chemical modifications, and which form loosely-bound complexes with themselves, other proteins and other molecule types. It is not only their chemical structure but also their overall 3-D structure which dictate their function. Other molecular classes that are hugely important in biological processes also have an intricate relationship with proteins. Any in situ mass spectrometry approach needs to be able to meet these analyte-driven challenges, i.e., it must be capable of (a) measuring proteins and characterising any modifications, (b) detecting protein complexes and determining their constituents, (c) providing information on 3-D structure, and (d) detecting other relevant molecular classes. Moreover, there are technique-driven challenges for in situ analysis including inherently high sample complexity and wide ranging concentrations, and opportunities for quantitation.

The research will meet these challenges by developing a newly emerging in situ approach, liquid extraction surface analysis mass spectrometry, in combination with two complementary types of ion mobility spectrometry (which can either provide information on 3-D structure, or separate ionised molecules in the mass spectrometer on the basis of their 3-D shape) and a structural elucidation strategy known as electron-mediated dissociation mass spectrometry.
The research will be undertaken primarily at the University of Birmingham in the Advanced Mass Spectrometry Facility in the School of Biosciences and the School of Chemistry mass spectrometry facility. The programme involves a number of academic and industrial collaborators and additional research will be carried out during scientific visits to National Physical Laboratory (NPL), Thermo Fisher Scientific, Waters, Owlstone, Florida State University, Texas A&M University and Université d'Aix-Marseille.

Who will benefit from this research?

The beneficiaries of this research will be analytical instrument manufacturers, the pharmaceutical industry, the healthcare industry, the NHS and its patients, and one of the UK's National Measurement Institutes (the National Physical Laboratory, NPL).

How will they benefit from this research?

The proposed research has the potential to contribute significantly to the nation's health and wealth:

Analytical instrumentation manufacturers: The work will enable the development of new instrumentation and workflows which can be employed in a broad range of new applications. The 2006 Royal Society of Chemistry Analytical and Measurement Sciences Report reported the UK analytical sector has an annual turnover of £7 billion and that the worldwide laboratory and analysis market was $25 billion. Thermo Fisher Scientific, Waters Corp. and Advion Inc are global companies, all of whom have a strong base in the UK. The proposed work will lead to increased sales thus fostering global economic performance and the economic competitiveness of the UK. Owlstone Ltd. is an SME (small and medium-sized enterprise), established in 2004, developing instruments based on the FAIMS technology. Again, the research will lead to increased sales and thus UK economic competitiveness.

National Measurement Institute: The proposed research will facilitate the UK's national measurement institute NPL, project partner on the fellowship, in achieving its goal of providing 'world-leading measurement solutions which are critical to commercial research and developments and supporting business across the UK and the globe'. NPL is a world leader in mass spectrometry imaging with the largest worldwide MSI research group and a recently established National Centre of Excellence in Mass Spectrometry Imaging (NiCE-MSI). In the first year of the NiCE-MSI, NPL delivered £100K of commercial consultancy to UK pharma. The proposed research will increase this through the development of extra capabilities.

Pharmaceutical industry: The pharmaceutical industry (annual turnover £31.8 bn) is underpinned by analytical science. The proposed research will facilitate the development of new drugs by providing methods for determining drug distribution and associated effects on tissue biochemistry, drug-target engagement and efficacy. AstraZeneca, a global company with a UK base, are partners on the programme meaning the research will have a direct impact on the health and wealth of the nation.

Healthcare industry: The healthcare industry is similarly underpinned by analytical science. The proposed research has direct implications for safety and quality assurance and will feed into the PI's ongoing collaboration with Unilever, a global company with a UK base. Again, the benefits are improved health and increased wealth of the nation.

NHS and its patients: The annual cost to the UK of periodontitis is £2.8 billion, for rheumatoid arthritis is £4 billion and for cancer is greater than £15 billion. Annual costs for kidney dialysis account for 2% of the NHS budget. In each case, there is a need for new analytical approaches for determining risk factors and for monitoring disease onset, progression and therapeutic response. The proposed research addresses that need and therefore has the potential to both reduce costs and improve health. Similarly, screening for haemoglobinopathies and in-born errors of metabolism is carried out for ~700,000 newborns every year in the UK. The combined estimate for Europe and the US is 8 million tests annually. The benefits of the proposed research would be reduced cost per test and improved diagnosis, and the research is supported by Birmingham Children's Hospital - the largest newborn screening centre in the UK.