Novel RGD peptide derivatives with enhanced pharmacokinetics for PET imaging

Objectives To demonstrate that suitably derivatized monomeric RGD peptide-based PET tracers (targeting integrin α_vβ₃) can offer advantages in image contrast, time for imaging, and low uptake in non-target tissues.

Methods Two c(RGDfk) derivatives, (PEG)₂-RGD and PEG₄-SAA₄-RGD, were constructed using solid-phase synthesis, and conjugated to NOTA for ⁶⁴Cu-labeling. Competitive cell binding assays (¹²⁵I-echistatin as the ligand) were performed to determine/compare integrin α_vβ₃ binding affinity of these peptides. Athymic nude mice bearing U87MG human glioblastoma tumors (integrin α_vβ₃+) were intravenously injected with each tracer, and dynamic PET scans were acquired during the first 30 min post-injection (p.i.). Static scans were performed at later time points to study long-term biodistribution of the tracers. Blocking and ex vivo biodistribution studies were carried out to validate the PET data and confirm the specificity of the tracers.

Results Three tracers, ⁶⁴Cu-NOTA-c(RGDfk) (1), ⁶⁴Cu-NOTA-(PEG)₂-RGD (2), and ⁶⁴Cu-NOTA-PEG₄-SAA₄-RGD (3), were prepared with high purity and radiolabeling yield (>95%). Integrin α_vβ₃ binding affinities for 1, 2, and 3 were comparable at IC₅₀ values of 507, 444, and 288 nM respectively. Dynamic PET data unveiled similar circulation t_1/2 for 2 and 3 (4.8 and 4.1 min respectively), and peak U87MG tumor uptake of ~4 %ID/g at 10 min p.i. for both tracers. Due to its marked hydrophilicity, 3 had faster clearance from tumor and normal tissues while maintaining excellent tumor-to-background ratios at 30 min p.i. (~ 16). Static PET scans at later time points corroborated the enhanced excretion of 3, especially from abdominal organs. Ex vivo biodistribution and receptor blocking studies confirmed the accuracy of the PET data and the specificity of 3 for integrin α_vβ_3.

Conclusions We developed 2 novel RGD peptide derivatives with optimized pharmacokinetic properties that allow for fast, high contrast PET imaging of integrin α_vβ₃. These tracers may facilitate the imaging of abdominal malignancies, normally precluded by high background uptake of other PET tracers.