Initial In Vitro and In Vivo Evaluation of a Novel CCK2R Targeting Peptide Analog Labeled with Lutetium-177
Anton Amadeus Hörmann1, Maximilian Klingler1, Maliheh Rezaeianpour1,2
1Department of Nuclear Medicine, Medical University of Innsbruck, 6020 Innsbruck, Austria.
Abstract:
Targeting of cholecystokinin-2 receptor (CCK2R) expressing tumors using radiolabeled minigastrin (MG) analogs is hampered by rapid digestion of the linear peptide in vivo. In this study, a new MG analog stabilized against enzymatic degradation was investigated in preclinical studies to characterize the metabolites formed in vivo. The new MG analog DOTA-DGlu-Pro-Tyr-Gly-Trp-(N-Me)Nle-Asp-1Nal-NH2 comprising site-specific amino acid substitutions in position 2, 6 and 8 and different possible metabolites thereof were synthesized. The receptor interaction of the peptide and selected metabolites was evaluated in a CCK2R-expressing cell line. The enzymatic stability of the 177Lu-labeled peptide analog was evaluated in vitro in different media as well as in BALB/c mice up to 1 h after injection and the metabolites were identified based on radio-HPLC analysis. The new radiopeptide showed a highly increased stability in vivo with >56% intact radiopeptide in the blood of BALB/c mice 1 h after injection. High CCK2R affinity and cell uptake was confirmed only for the intact peptide, whereas enzymatic cleavage within the receptor specific C-terminal amino acid sequence resulted in complete loss of affinity and cell uptake. A favorable biodistribution profile was observed in BALB/c mice with low background activity, preferential renal excretion and prolonged uptake in CCK2R-expressing tissues. The novel stabilized MG analog shows high potential for diagnostic and therapeutic use. The radiometabolites characterized give new insights into the enzymatic degradation in vivo.
Insights
A new stabilized minigastrin analog shows increased stability in vivo for targeting cholecystokinin-2 receptor (CCK2R) expressing tumors. This radiopeptide demonstrates potential for improved diagnostic and therapeutic applications in oncology.
Area of Science:
- Biomedical research
- Radiopharmaceutical chemistry
- Oncology
Background:
- Targeting cholecystokinin-2 receptor (CCK2R) expressing tumors with radiolabeled minigastrin (MG) analogs is limited by in vivo peptide degradation.
- Development of stabilized analogs is crucial for effective in vivo delivery and therapeutic efficacy.
Purpose of the Study:
- To synthesize and characterize a novel, stabilized minigastrin analog for CCK2R targeting.
- To evaluate the in vivo enzymatic stability, receptor binding, and biodistribution of the new analog and its metabolites.
Main Methods:
- Synthesis of a novel MG analog with site-specific amino acid substitutions (DOTA-DGlu-Pro-Tyr-Gly-Trp-(NMe)Nle-Asp-1Nal-NH2).
- In vitro evaluation of receptor interaction in a CCK2R-expressing cell line.
- In vitro and in vivo (BALB/c mice) enzymatic stability assessment of the 177Lu-labeled analog.
- Radio-HPLC analysis for metabolite identification.
- Biodistribution studies in BALB/c mice.
Main Results:
- The novel radiopeptide exhibited significantly increased in vivo stability, with >56% intact peptide remaining in blood after 1 hour in BALB/c mice.
- High CCK2R affinity and cell uptake were observed only for the intact peptide; enzymatic cleavage abolished these properties.
- Favorable biodistribution was noted, characterized by low background activity, renal excretion, and prolonged uptake in CCK2R-expressing tissues.
Conclusions:
- The stabilized minigastrin analog demonstrates high potential for diagnostic and therapeutic applications in CCK2R-expressing tumors.
- Characterization of radiometabolites provides valuable insights into the in vivo enzymatic degradation pathways.
- This stabilized analog offers an improved approach for targeted radionuclide therapy and imaging.
