In Vitro and in Vivo Evaluation of ⁶⁴Cu-Labeled DOTA−Linker−Bombesin(7−14) Analogues Containing Different Amino Acid Linker Moieties

Abstract

The gastrin-releasing peptide receptor (GRPR) is overexpressed on a variety of tumor types and has been targeted with radiolabeled peptides for detection and therapy of these cancers. Analogues of the 14 amino acid bombesin (BN) peptide have been radiolabeled with both gamma- and positron-emitting radionuclides for detection of GRPR-expressing tumors. We have previously evaluated BN analogues radiolabeled with the positron-emitter, copper-64 (64Cu), that contained various aliphatic linkers placed between the BN peptide and the 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator. These studies showed that the analogues could be used for positron-emission tomographic (PET) imaging of GRPR-positive tumors in mice but clinical translation would be hindered by significant uptake in background tissues. Therefore, the purpose of this study was to determine if the use of amino acid linkers placed between the DOTA chelate and the BN peptide would reduce nontarget tissue uptake, while maintaining good prostate tumor uptake. The linkers studied utilized three amino acid combinations of glycine (G), serine (S), or glutamic acid (E). In vitro assays in PC-3 cells showed that the glutamic acid-containing linkers had poor binding and internalization, while the other analogues had IC50 values <100 nM and good internalization. In vivo, these same analogues demonstrated tumor-specific uptake and good imaging characteristics that were comparable to, or better than the previously reported 64Cu-labeled DOTA-BN analogues. Overall, this study shows that BN analogues containing amino acid linkers can be used for the PET imaging of GRPR-expressing prostate cancer and that these linkers lead to lower background tissue uptake.

18F-Labeled Bombesin Analog for Specific and Effective Targeting of Prostate Tumors Expressing Gastrin-Releasing Peptide Receptors

“…The specificities of uptake for the tumor and the pancreas were 82% and 96%, respectively. 64 Cu-DOTA-bombesin peptides exhibited tumor uptake and good imaging characteristics (30). Our laboratories have focused on labeling peptides for PET of both breast cancer and prostate cancer with 18 F because this nuclide has ideal characteristics for peptide receptor imaging studies.…”

Peptide-modified liposomes for selective targeting of bombesin receptors overexpressed by cancer cells: a potential theranostic agent

“…[28][29][30] Many studies demonstrate that the BN(7-14) fragment modified on its N-terminus with radiometal complexes, for diagnostic or therapeutic nuclear medicine applications, preserves its affinity for these receptors. [31][32][33] In-vitro and in-vivo experiments, carried out on radiolabeled liposomes containing the BN (7)(8)(9)(10)(11)(12)(13)(14) peptide fragment, demonstrate that the peptide, exposed on the liposome surface, maintains the capability to selectively target aggregates to GRPR-expressing xenografts. 27 This paper describes new BN-conjugated nanosystems for theranostic purposes with improved properties, both in the synthesis and formulation aspects and for their in-vitro and in-vivo behavior.…”

64Cu-Labeled Peptide for PET of Breast Carcinomas Expressing the Thomsen-Friedenreich Carbohydrate Antigen

“…Improvement in the tumor uptake and pharmacokinetics of 64 Cu-labeled cyclic RGD peptides using Gly-Gly-Gly (GGG) and polyethyleneglycol 4 (PEG4) linkers has been reported (29). Other studies indicated that prostate tumor uptake of 64 Cu-labeled DOTA-bombesin peptide with a GGG-linker was higher than that with a GSG-linker, although kidney uptake was higher (30). Future work will focus on evaluating and optimizing the pharmacokinetic profile of 64 Cu-NO2A-TFpep using more than 1 linker group based on hydrocarbons, PEG, or amino acids.…”

“…MDA-MB-435 and 184A1 cells grown in culture flasks were trypsinized, released, and washed once in cell-binding medium (RPMI 1640 with 25 mM N-(2-hydroxyethyl)piperazine-N9-(2-ethanesulfonic acid), pH 7.4, 0.2% bovine serum albumin [BSA], and 3 mM 1,10-phenanthroline). Cells (2 · 10 5 cells per tube) were transferred to microcentrifuge tubes containing 0.3 mL of cell-binding medium and were incubated at 37°C for different times (10,20,30,40, and 60 min) with 1 · 10 5 counts per min (cpm) of radiolabeled and scrambled peptide. After incubation, medium was removed and the cells were rinsed with ice-cold 0.01 M phosphate-buffered saline (PBS), pH 7.4, and 0.2% BSA, and then centrifuged.…”