Objectives: Imaging of angiogenic processes is of great interest in preclinical research as well as in clinical settings. One target structure family involved are the integrins. Most radiopharmaceutical development yet is focused on the integrin αvβ3. Here we describe the synthesis and evaluation of [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH, a radiolabelled peptide designed to selectively target the integrin α5β1.Methods: Synthesis of the peptides was based on solid phase peptide synthesis protocols using Fmoc-strategy. For radiolabelling 4-nitrophenyl-(RS)-2-[18F]fluoropropionate was used. For in vitro characterization, partition coefficient, protein binding properties, human se... More
Objectives: Imaging of angiogenic processes is of great interest in preclinical research as well as in clinical settings. One target structure family involved are the integrins. Most radiopharmaceutical development yet is focused on the integrin αvβ3. Here we describe the synthesis and evaluation of [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH, a radiolabelled peptide designed to selectively target the integrin α5β1.Methods: Synthesis of the peptides was based on solid phase peptide synthesis protocols using Fmoc-strategy. For radiolabelling 4-nitrophenyl-(RS)-2-[18F]fluoropropionate was used. For in vitro characterization, partition coefficient, protein binding properties, human serum stability, binding affinity for integrins α5β1, αvβ3, and αIIbβ3 as well as cell uptake were determined. To characterize the in vivo properties, biodistribution studies were carried out. For both, in vitro and in vivo evaluation, human melanoma M21 (αvβ3-positive, α5β1-negative), human melanoma M21-L (αvβ3-negative, α5β1-negative), and human prostate carcinoma DU145 (αvβ3-negative, α5β1-positive) cells were used.Results: Conjugation of the prosthetic group to provide [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH was achieved in high radiochemical purity (>95%) and a radiochemical yield of approx. 55%. Overall synthesis time was comparable with that of the reference RGD peptide [18F]Galacto-RGD. In vitro evaluation showed α5β1 binding affinity in the nanomolar range, whereas binding to αvβ3 and αIIbβ3 was >50 μM. Cell uptake studies confirmed receptor specific binding. The radiotracer was stable in human serum and showed low protein binding tendency (approx. 4% after 120 min incubation). Biodistribution studies showed uptake in the tumour ranging from 2.5 to 3.5% ID/g between 30 and 120 min p.i.. Uptake in blood and the organs investigated was only marginally lower. However, blocking studies using increasing amounts of the lead structure as well as uptake studies using mice bearing the α5β1-negative M21 tumour xenograft did not confirm receptor-specific uptake of [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH.Conclusion: Despite [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH revealed high affinity and substantial selectivity to α5β1 in vitro, the in vivo data could not confirm receptor-specific targeting of integrin α5β1. Further experiments are needed to study the in vivo metabolism of this peptide and to develop improved radiopeptide candidates suitable for PET imaging of α5β1 expression in vivo.
