Synthesis and Evaluation of Novel 68 Ga-Labeled GRPR-Targeted PET Tracers Derived from d -Phe 6 ,Pro 14 Bombesin(6–14) and d -Phe 6 ,des-Met 14 Bombesin(6–14) Sequences

Gastrin-releasing peptide receptor (GRPR) is a G-protein-coupled receptor that is overexpressed in several malignancies, rendering it an ideal target for cancer imaging and therapy. In this report, we conducted structure-activity relationship studies by optimizing the substitution of Pro14 based on an agonist sequence, d-Phe6,Pro14Bombesin(6-14), and optimizing the substitution of the C-terminal Leu13-NH2 based on an antagonist sequence, d-Phe6,des-Met14Bombesin(6-14). PEP-1-PEP-5 derived from the agonist sequence and PEP-6-PEP-14 derived from the antagonist sequence were synthesized by a solid-phase approach and were obtained in 10-60% yield. In vitro competition binding assays showed that the Ki values of PEP-1-PEP-14 were in the range of 1.16 nM to 266 nM. PEP-3, PEP-4, PEP-8, PEP-9, and PEP-10 with Ki N-terminus. Calcium release assays confirmed the agonist-antagonist nature of Ga-DOTA-Pip-conjugated peptides. Ga-DOTA-Pip-PEP-4, Ga-DOTA-Pip-PEP-9, and Ga-DOTA-Pip-PEP-10 retaining high GRPR binding affinity (Ki 68Ga labeling for further evaluation. 68GaGa-DOTA-Pip-PEP-4, 68GaGa-DOTA-Pip-PEP-9, and 68GaGa-DOTA-Pip-PEP-10 were obtained in 16-44% decay-corrected radiochemical yields with ≥64 GBq/μmol molar activity and ≥ 97% radiochemical purity. PET imaging and ex vivo biodistribution studies were conducted in PC-3 tumor-bearing mice at 1 h postinjection. All 68GaGa-DOTA-Pip-PEP-4, 68GaGa-DOTA-Pip-PEP-9, and 68GaGa-DOTA-Pip-PEP-10 were excreted mainly via the renal pathway and enabled clear visualization of tumor xenografts in PET images with good tumor-to-background contrast. The pancreas uptake of the agonist tracer, 68GaGa-DOTA-Pip-PEP-4, was much lower than that of the clinically evaluated agonist tracer, 68GaGa-AMBA. Similarly, the pancreas uptake of the antagonist tracers, 68GaGa-DOTA-Pip-PEP-9, and 68GaGa-DOTA-Pip-PEP-10, was also much lower than that of the clinically evaluated antagonist tracers, 68GaGa-RM2, 68GaGa-SB3, and 68GaGa-NeoB. Our data demonstrate that the sequences of Ga-DOTA-Pip-PEP-4, Ga-DOTA-Pip-PEP-9, and Ga-DOTA-Pip-PEP-10 are promising templates for the development of radiopharmaceuticals targeting GRPR-expressing cancer.