Enhanced Specific Activity by Multichelation of Exendin-3 Leads To Improved Image Quality and In Vivo Beta Cell Imaging

Abstract

Glucagon-like peptide-1 receptor (GLP-1R) targeting using radiolabeled exendin is a promising approach to noninvasively visualize and determine beta cell mass (BCM), which could help to unravel the pathophysiology of diabetes. However, saturation of the GLP-1R on beta cells occurs at low peptide doses, since the number of receptors expressed under physiological conditions is low. Therefore, tracers with high specific activities are required to sensitively image small variations in BCM. Here, we describe a novel exendin-3-based radiotracer with multiple chelators and determine its potential for in vivo beta cell imaging. Exendin-3 was modified by adding six lysine residues C-terminally conjugated with one, two, or six DTPA moieties. All compounds were labeled with 111In and their GLP-1R affinity was determined in vitro using GLP-1R expressing cells. The in vivo behavior of the 111In-labeled tracers was examined in BALB/c nude mice with a subcutaneous GLP-1R expressing tumor (INS-1). Brown Norway rats were used for SPECT visualization of the pancreatic BCM. Addition of six lysine and six DTPA residues (hexendin(40–45)) resulted in a 7-fold increase in specific activity (from 0.73 GBq/nmol to 5.54 GBq/nmol). IC50 values varied between 5.2 and 69.5 nM. All compounds with two or six lysine and DTPA residues had a significantly lower receptor affinity than [Lys40(DTPA)]exendin-3 (4.4 nM, p < 0.05). The biodistribution in mice revealed no significant decrease in pancreatic uptake after addition of six lysine and DTPA molecules. Hexendin(40–45) showed a 6-fold increase in absolute 111In uptake in the pancreas of Brown Norway rats compared to [Lys40(DTPA)]exendin-3 (182.7 ± 42.3 kBq vs 28.8 ± 6.0 kBq, p < 0.001). Visualization of the pancreas on SPECT was improved using hexendin(40–45), due to the higher count rate, achieved at the same peptide dose. In conclusion, hexendin(40–45) showed an improved visualization of the pancreas with SPECT. This tracer holds promise to sensitively and specifically detect small variations in BCM.

Multivalent γ‐PGA‐Exendin‐4 Conjugates to Target Pancreatic β‐Cells

“…The derivatization site on lysine is a crucial aspect to be considered, since observations of crystal structure reveal that Ex-4 interacts with GLP-1R with multiple hydrophilic and hydrophobic residues. [16,71] Nevertheless, literature reveals how different modifications on more positions have been successfully carried out and evaluated, including positions 1 (histidine), [72] 12 (lysine, Lys), [73,74] 14 (methionine), [75] 27 (Lys), [7,76] 39 (serine), [77] and 40 (Lys). [78][79][80][81][82][83] Lysins are the most exploited amino acid in Ex-4 for bioconjugates synthesis, displaying a primary amine suitable for chemical derivatization.…”

Advances in Development of Radiometal Labeled Amino Acid-Based Compounds for Cancer Imaging and Diagnostics

“…Studies covering 111 In-labeled biomolecules are aimed at somatostatin receptor imaging [ 66 ], glucagon-like peptide receptor [ 67 , 68 ], gastrin-releasing peptide receptor [ 69 ], or integrins [ 70 ].…”

Advances in GLP-1 receptor targeting radiolabeled agent development and prospective of theranostics

“…In order to enhance SRA and enable detection and quantification of small changes in beta cell mass responsible for diabetic pathophysiology progression, a number of exendin-3 analogues carrying one, two, or six DTPA moieties were developed 118. The analogues were labeled with 111 In and the one comprising six chelator moieties demonstrated 7-fold increase in SRA.…”