Abstract
The substrate scope of fluorinase enzyme mediated transhalogenation reactions is extended. Substrate tolerance allows a peptide cargo to be tethered to a 5'-chloro-5'-deoxynucleoside substrate for transhalogenation by the enzyme to a 5'-fluoro-5'-deoxynucleoside. The reaction is successfully extended from that previously reported for a monomeric cyclic peptide (cRGD) to cargoes of dendritic scaffolds carrying two and four cyclic peptide motifs. The RGD peptide sequence is known to bind upregulated αVβ3 integrin motifs on the surface of cancer cells and it is demonstrated that the fluorinated products have a higher affinity to αVβ3 integrin than their monomeric counterparts. Extending the strategy to radiolabelling of the peptide cargoes by tagging the peptides with [(18)F]fluoride was only moderately successful due to the poor water solubility of these higher order peptide scaffolds although the strategy holds promise for peptide constructs with improved solubility.
Highlights
The fluorinase enzyme (E.C. 2.5.1.63) was originally isolated[1] from the soil bacterium Streptomyces cattleya and has recently been identified in a number of other bacterial species.[2]
The fluorinase catalyses the combination of fluoride ion and S-adenosylL-methionine 1 (SAM) to generate 5′-fluoro-5′-deoxyadenosine 2 (FDA) and L-(S)-methionine 3 (L-Met) (Scheme 1A).[1,7]
Following deprotection to the free terminal alkynes, a multi-CuAAC reaction with an azide bearing cRGD peptide, similar to that reported by Liskamp et al.,[34] should furnish multimeric cRGD substrates
Summary
The fluorinase enzyme (E.C. 2.5.1.63) was originally isolated[1] from the soil bacterium Streptomyces cattleya and has recently been identified in a number of other bacterial species.[2]. While two step strategies involving prosthetic groups remain widely used for radiolabelling of peptides and proteins with fluorine-18,8 it is attractive to consider the development of straightforward, “last step” radiolabelling protocols. The fluorinase has the capacity to catalyse C–F bond formation at ambient temperature and near neutral pH Within this context, we have previously demonstrated[5] a fluorinase enzyme-based system for “last step” radiolabelling of a small cRGD peptide 7. The peptide was tethered to a 5′-chloro-5′-deoxynucleoside substrate through a PEG linker prior to enzymatic radiolabelling with [18F]fluoride, as shown in Scheme 1B This proved successful and the radiolabelled construct [18F]-8 was shown to be stable to defluorination in vivo (in a rat), and the cRGD moiety retained high affinity for αVβ3 integrins. Fewer fluorine-18-based multivalent radiotracers have been developed, as incorporation of fluorine[18] into such scaffolds often requires a multi-step, prosthetic group-based strategy.[25,26,27,28] two multimeric cRGDbased radiotracers have been reported recently, synthesised
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