Design and synthesis of novel radiolabelled peptide-based Eph receptor inhibitors for PET

Objectives:

Eph receptors and their ephrin ligands belong to the largest receptor tyrosine kinase family of transmembrane proteins. Since Eph receptors are found to be dysregulated in certain tumour species [1, 2] many efforts have been made to inhibit such receptors in order to stop tumourigenesis and tumour growth [3]. To date, several promising candidates of small molecules exist [4, 5], which inhibit Eph receptors at the intracellular domain. Furthermore, peptides with the key amino acid sequence SNEW show the highest affinity for the EphB2 receptor and, in this regard, block the extracellular receptor domain [6]. In this case, the development of a labelling strategy for the radiofluorination of SNEW peptides to design new PET-tracers is promising for specific tumour diagnosis.
Methods:
Several novel SNEW peptides were synthesized via an automated Fmoc-protecting group strategy. The key sequence at the N-terminus has to be unmodified in order to sustain the affinity to EphB2-receptor. The C-terminus was modified with cysteine, lysine and azidoproline for a radiofluorination with [¹⁸F]FBAM, [¹⁸F]SFB [7] and by use of bioorthogonal reactions like the Staudinger ligation or the Huisgen cycloaddition.
Results:
Since the key sequence of SNEW peptides is essential for the affinity to the EphB2-receptor, the SNEW peptides were modified at the C-terminus for the bioorthogonal labelling. In order to compare the new bioorthogonal reactions with established labelling methods, the cysteine modified SNEW peptide was reacted with [¹⁸F]FBAM (60 °C, 60 min, Sørensen-buffer (pH = 7.2)/acetonitrile (vv = 1:1)). Additionally, [¹⁸F]BFP as new and
versatile labelling building block for the copper-catalyzed Huisgen-cycloaddition was developed. Therefore, the azidoproline modified SNEW peptide was labelled with this click building block under milder conditions (r.t., 30 min, Sørensen-buffer (pH = 7.2)/ethanol (vv = 1:1)).
Conclusions:
New labelling building blocks were synthesized for the bioorthogonal radiofluorination of novel azide-functionalized peptides with a SNEW sequence and compared to standard labelling methods like [¹⁸F]FBAM. The SNEW peptides show a high affinity to the EphB2 receptor and represent promising radiotracers for a specific tumour diagnosis.
References:
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