Extending fluorinase [C-18F]-bond biocatalysis for Positron Emission Tomography (PET)

Lead Research Organisation: University of St Andrews
Department Name: Chemistry

Abstract

Positron emission tomography (PET) is the most sensitive functional imaging method clinically and it application is growing rapidly through the Western world and developing countries, particularly as a diagnostic imaging tool for cancers and degenerative neurological disorders. Many major hospitals and clinical research centres in the Europe, the US and Asia are now commissioning cyclotrons and developing PET research facilities locally. Fluorine-18 is an important isotope for PET. It has a relatively long half-life (109 mins) and is readily generated in a cyclotron in the form of [18F]-fluoride ion, in very high specificic activity (GBq's) from oxygen-18 water. As a consequence new methods to develop C-[18F]F bond formation for PET labelling are in demand, In general the link between fluorine chemistry and pharmaceutical/medical applications is strong. Approximately 20% of all pharmaceutical, since the 1950s, contain a fluorine atom and the bio-distribution of all new pharmaceutical products are required to be explored by PET, as part of clinical trials. Also new PET tracers are in demand as tools for early diagnosis as indicators of disease states. New fluorine chemistry is required to meet the demands of a growing and dynamic PET research community both in the UK and internationally. This proposal aims to develop a novel methodology for incorporating fluoride-18 into relevant orgaganic molecules.
In this proposal we aim to exploit a novel enzyme which can form C-F bonds from fluoride ion. The fluorinase enzyme was discovered in 2002 (Nature, 2002, 416, 279) in St Andrews and it has been over-expressed and its structure (X-ray) and mechanism elucidated. The enzyme catalyses the reaction of fluoride ion and S-adenosyl-L-methionine (SAM) to generate 5'-FDA and L-methionine. It has proven to be a chemoselective biotransformation method for generating C-18F bonds from inorganic [18F]-fluoride. The fluorinase is the only example of an enzyme used in PET synthesis and in this regard it offers an entirely new method for incorporating fluorine. In practical terms it has emerged to be particularly appropriate, because PET uses picomolar [18F]-fluoride ion, but the over-expressed fluorinase enzyme is present at mg/ml (microM), and therefore the kinetics favour C-18F synthesis due to a large molar excess of enzyme.
This is a research collaboration between the Universities of St Andrews and Aberdeen where the methods will be developed in St Andrews and the radiolabeling protocols carried out at the Aberdeen PET Centre, situated in the Royal Infirmary Hospital. The major focus of the research will concentrate on developing fluorinase based chemistry to prepare novel fluorine-18 small molecules that can be used to attach to proteins through site specific cysteine residues. It is a current objective to find novel methods to radiolabel proteins such as enzymes and antibodies, to monitor their in vivo distribution by imaging. A key strategy will explore substrate analogues for the fluorinase, which carry acetylene residues, such that a 'click' chemistry approach can be adopted.
A secondary objective of the research is to exploit an efficient chemo-enzymatic synthesis of [18F]-fluoroacetyl-CoA, developed between St Andrews and Aberdeen, to prepare [18F]-fluorocitrate, [18F]-fluoromalate and [18F]-N-aceylglutamate, as novelties for in vivo imaging.

In overview the research will present a range of new methodologies for PET chemistry for the incorporation of the fluorine-18 isotope into molecules of biological relevance. The research in generic and aims to expemplify new methods, for much wider applications by the growing international research community of PET radiochemists.

Planned Impact

Who will benefit?
The research will develop enzymatic C-18F bond formation for PET. In particular it will develop a range of novel 18F-labelled molecules with potential as new imaging agents eg for cancer. It will also introduce a novel methodology for radiolabelling proteins in a site specific manner, for particular proteins used to diagnose particular disease states, by PET imaging. The fluorinase has the potential to be utilised by a variety of PET Centres and radio-chemists involved in protein labelling for imaging. These chemists work in PET radiochemistry labs closely associated with the pharmaceuticals industries or directly in clinical research environments. So the research has the potential to impact quickly and directly on translational medicine research environments. Outstanding success, perhaps in 10 years time would see fluorinase methods being used internationally to prepare a specific compound or for labelling a specific protein, that will impact on disease diagnosis and contribute to improving health treatment. In that respect members of the public will benefit.

How will they benefit?
The programme will demonstrate the potential and range of the fluorinase methodology and creative scientists will be able to apply it in other arenas to solve their particular diagnostic challenge. Eg to make a specifically labelled molecule, or label a protein using fluorinase technology applied to a particular disease.

What will be done?
The outcomes of the research will be published in leading journals, following a pattern from our recent research projects. The methodology and how to apply it will be available quickly to the international community. Patenting and IPR protection will be required for societal impact. This is already in place, but can be developed for individual applications, eg specific protein lebelling. Such IPR will most reasonably be pursued at the individual research project level, as developments unfold. The important aspect of this programme is to demonstrate the scope of the methodology to the international community and demonstrate the potential that may lie in fluorinase biocatalysis applied to PET.

Methods of Communication and engagement
The research will be published. Presentations will be made at key international conferences both in fluorine/pharma chemistry environments and among the radiochemistry/PET Imaging community, following a current pattern by DO'H/MZ. Specific targeting of the imaging community will be pursued.
The PI and CoI are on the Executive Committee of SINAPSE (Scottish Imaging Network, a Platform for Excellence). ( http://www.sinapse.ac.uk/ )
They have access to major meetings and congregations of the International Clinical Imaging Community, through that mechanism. The SINAPSE connection enables an ongoing and intensive discussion with the relevant international community. SINAPSE are keen to project their successes and provide an excellent additional vehicle for dissemination.

Collaboration and exploitation
St Andrews has a full patent on the fluorinase (2002). There is a sound platform on which to exploit any new technology. The fluorinase also has BBSRC Follow on Funding (till Nov 2011) to develop the fluorinase as a user friendly catalyst for [18F]-fluorodeoxyribose ([18F]-FDR) synthesis. Through this and SINAPSE we have a close relationship with GSK at the Hammersmith Hospital in London. Key GSK scientists advise on potential applications of the research in clinical imaging perspective. Through such routes the impact of the research will reach front line translational medical laboratories working in the relevant area.

The grpwth of PET research centres now required trained radiochemists with a specialist focus. This programme contributes directly to growing that skill base in the UK

Publications

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Thompson S (2016) Enzymatic transhalogenation of dendritic RGD peptide constructs with the fluorinase. in Organic & biomolecular chemistry

 
Description A very efficient method for fluorine-18 labelling of peptides and proteins was developed using 5- [18F]-fluorodeoxyribose.
This allows cancer relevant peptides to be tagged with a radioactive label, so that tumors can be imaged for diagnosis or disease progression.
Exploitation Route The method has already been used in other clinical research environments in Amsterdam, Turku and the Hammersmith Hospital in London.
We have filed a patent. We continue to exemplify the method.We have also started a new collaboration with King College London, on a potential prostrate cancer imaging technology
Sectors Healthcare

 
Description A patent was filed by the University of St Andrews
First Year Of Impact 2013
Sector Healthcare
Impact Types Societal

 
Description EPSRC Responsive Mode
Amount £341,000 (GBP)
Funding ID EP/M01262X/1 
Organisation Engineering and Physical Sciences Research Council (EPSRC) 
Sector Public
Country United Kingdom
Start 02/2015 
End 01/2018
 
Description A2A receptor agonist labelling 
Organisation Leiden University
Country Netherlands 
Sector Academic/University 
PI Contribution We have collaborated with Leiden University where they have A2A receptor agonist assays
Collaborator Contribution Leiden were able to assay two novel seelctively fluorinated A2A receptor agonists prepare in our lab. These were also successfully labelled with fluorine-18 by out Aberdeen University collaborators, for positron emission tomography studies.
Impact A publication : Phillip T. Lowe, Sergio Dall'Angelo, Thea Mulder-Krieger, Adriaan P. IJzerman, Matteo Zanda and David O'Hagan, A novel class of fluorinated A2A adenosine receptor agonist with application to last step enzymatic [18F]fluorination for PET imaging, ChemBioChem., 2017, 18, 2156 - 2164 Multidicsiplinary - Chemistry and human biology
Start Year 2007
 
Description Aberdeen PET Centre 
Organisation University of Aberdeen
Department Health Services Research Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution We collaborate with Professor Matteo Zanda's group at the Positron Emission Tomography Centre at the Royal Infirmary, University of Aberdeen. They carry out the fluorine-18 experiments.
Collaborator Contribution Aberdeen PET Centre has been a medical imaging unit for over 20 years. It is now led by Professor Matteo Zanda an internationally recognised organo-fluorine chemist and fluorine-18 radiochemist. St Andrews and Aberdeen have collaborated closely applying fluorinase technology to PET. The PET laboratory at Abedeen has three radiochemists chemists who develop research programmes usually in conjunction with pharma. They host St Andrews scientists to carry out fluorinase catalysed radiolabelling experiments
Impact Funding and training of Scientists Ph.D student Stephen Thompson is now employed at the PET Centre at University of Michigan (Ann Arbour) Medical School
Start Year 2012