Last step' enzymatic [18F]-labelling of peptides for Positron Emission Tomography (PET)



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 specific 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 specifically into peptides and proteins.

In this project 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. However we have found a weakness in the substrate specificity. We find that at a very specific location we can attach a linker to the substrate, and it will be accepted by the enze, this linker provides an anchor point to run a molecular line (poly ethylene glycol) to a peptide molecule of choice. The chosen peptides are those that identify cancer cells in the body, known as homing peptides, or small antibodies called 'affibodies' that identify tumour cells. In this way we can use the enzyme to attach the fluorine-18 isotope. The important advantage is that the fluoride-18 is generated in water, and the enzyme functions in water at neutral pH. Also peptides are nicely soluble in water, so the labelling can take place without the difficulty of using organic solvents for these biomolecules. This presents attractive possibilities. The fluorinase is the only example of an enzyme used in fluorine-18 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 enzymatic methods for labelling the petides and proteins will be developed in St Andrews and the radiolabeling protocols carried out at the Aberdeen PET Centre, situated in the Aberdeen Royal Infirmary. The major focus of the research will concentrate on rapid labelling of peptides under neutral ambient conditions.

The research aims to establish new methods for much wider applications by the growing international research community of PET radiochemists and we have ambitions to translate the methods to the clinic through interactions with PET based companies such an Imanova and our established interactions with the Beatson Cancer Institute in Glasgow.
Date made available1 Mar 2016
PublisherUniversity of Aberdeen
Temporal coverage1 Mar 2015 - 28 Feb 2018

Funder and Grant Reference number

  • Engineering and Physical Sciences Research Council (EPSRC)
  • EP/M012573/1

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