⁶⁴Cu Labeled Sarcophagine Exendin-4 for MicroPET Imaging of Glucagon like Peptide-1 Receptor Expression

Abstract

The Glucagon-like peptide 1 receptor (GLP-1R) has become an important target for imaging due to its elevated expression profile in pancreatic islets, insulinoma, and the cardiovascular system. Because native GLP-1 is degraded rapidly by dipeptidyl peptidase-IV (DPP-IV), several studies have conjugated different chelators to a more stable analog of GLP-1 (such as exendin-4) as PET or SPECT imaging agents with various advantages and disadvantages. Based on the recently developed Sarcophagin chelator, here, we describe the construction of GLP-1R targeted PET probes containing monomeric and dimeric exendin-4 subunit. The in vitro binding affinity of BarMalSar-exendin-4 and Mal2Sar-(exendin-4)2 was evaluated in INS-1 cells, which over-express GLP-1R. Mal2Sar-(exendin-4)2 demonstrated around 3 times higher binding affinity compared with BaMalSar-exendin-4. After 64Cu labeling, microPET imaging of 64Cu-BaMalSar-exendin-4 and 64Cu-Mal2Sar-(exendin-4)2 were performed on subcutaneous INS-1 tumors, which were clearly visualized with both probes. The tumor uptake of 64Cu-Mal2Sar-(exendin-4)2 was significantly higher than that of 64Cu-BaMaSarl-exendin-4, which could be caused by polyvalency effect. The receptor specificity of these probes was confirmed by effective blocking of the uptake in both tumor and normal positive organs with 20-fold excess of unlabeled exendin-4. In conclusion, sarcophagine cage conjugated exendin-4 demonstrated persistent and specific uptake in INS-1 insulinoma model. Dimerization of exendin-4 could successfully lead to increased tumor uptake in vivo. Both 64Cu-BaMalSar-exendin-4 and 64Cu-Mal2Sar-(exendin-4)2 hold a great potential for GLP-1R targeted imaging.

Radioiodinated Exendin-4 Is Superior to the Radiometal-Labelled Glucagon-Like Peptide-1 Receptor Probes Overcoming Their High Kidney Uptake

“…This prompted an intensive research into developing more sensitive and non-invasive imaging modalities to localize benign insulinoma in the pancreas. A variety of GLP-1R targeting peptide probes labelled with 111 In [7,8], 99m Tc [8], 123 I [9] for SPECT and with 68 Ga [3,8,10–13], 64 Cu [3,14], 18 F [5,15,16], and 89 Zr [13] for PET imaging showed promising results in preclinical studies. Some of the tracers have been successfully translated into the clinic [17–22] and showed highest sensitivity among the available techniques in detecting ‘‘hidden” insulinomas in patients.…”

Recent advances in biological applications of cage metal complexes

“…The latter may also be synthesized with no need for the isolation of an intermediate copper(II) sarcophaginate 29 [44]. Similarly, 64 Cu 2+ cage complexes of exendin-4functionalized sarcophagines 48 and 49, having high kidney radioactivity levels [53], showed a persistent and specific uptake in a given insulinoma model; an increased tumor uptake in vivo has been also observed for 49. Mono-and bifunctionalized copper(II) sarcophaginates 34 -36 (Scheme 10) with di-or trimethylene linkers and with terminal reactive carboxylic groups [47] have been obtained from octaamine precursors; their further functionalization with various peptides has allowed obtaining 64 Cu 2+ sarcophaginates that were stable for a long time under the conditions of in vivo experiments [48].…”

“…In this case, the intermediate sarcophagine to be further functionalized has been obtained from a corresponding cobalt(III) cage complex by either its demetallation followed by base hydrolysis of the free ligand or by its base hydrolysis followed by demetallation (Scheme 8). The receptor of this peptide with elevated expression profile in pancreatic islets, insulinoma and cardiovascular system [53] and its accurate visualization and quantification of β-cell mass are critical for improved understanding, diagnosis and treatment of type 1 diabetes and insulinoma. Another sarcophaginate-based bioconjugate 38 (Scheme 11) with the encapsulated 64 Cu 2+ ion, which performed well in both in vitro and in vivo experiments [49], has been synthesized via 1,3-dipolar cycloaddition of a dibenzocyclooctyne-functionalized 64 A 64 Cu-labelled complex of a dipeptide sarcophagine 47 (Scheme 13), synthesized from a diazide-containing sarcophagine and a corresponding alkyne-containing peptide, has been used [52] for microPET imaging of the receptor CD13 (a tumor vasculature biomarker) expressed in vitro and in vivo in a living mice; it displayed a good binding affinity and CD13 specificity with given tumor cells and an excellent tumor uptake.…”

Advancement in treatment and diagnosis of pancreatic cancer with radiopharmaceuticals

“…In the past few years, exendin-4 analogs have been labeled with PET radionuclides for preclinical insulinomas imaging. Exendin-4 labeled with radio metals ( 68 Ga, 64 Cu) showed significant uptake in INS-1 insulinoma xenografts [78,79] . However, the substantial kidney uptake may limit their use in clinical practice due to high radiation exposure to the organs.…”