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In Vitro and In Vivo Evaluation of a F-18-Labeled High Affinity NOTA Conjugated Bombesin Antagonist as a PET Ligand for GRPR-Targeted Tumor Imaging
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preclinical PET Platform.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preclinical PET Platform.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preclinical PET Platform. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Section of Nuclear Medicine and PET.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
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2013 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 12, e81932- p.Article in journal (Refereed) Published
Abstract [en]

Expression of the gastrin-releasing peptide receptor (GRPR) in prostate cancer suggests that this receptor can be used as a potential molecular target to visualize and treat these tumors. We have previously investigated an antagonist analog of bombesin (D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2, RM26) conjugated to 1,4,7-triazacyclononane-N,N',N ''-triacetic acid (NOTA) via a diethylene glycol (PEG(2)) spacer (NOTA-P2-RM26) labeled with Ga-68 and In-111. We found that this conjugate has favorable properties for in vivo imaging of GRPR-expression. The focus of this study was to develop a F-18-labelled PET agent to visualize GRPR. NOTA-P2-RM26 was labeled with F-18 using aluminum-fluoride chelation. Stability, in vitro binding specificity and cellular processing tests were performed. The inhibition efficiency (IC50) of the [F-nat]AlF-NOTA-P2-RM26 was compared to that of the Ga-nat-loaded peptide using I-125-Tyr(4)-BBN as the displacement radioligand. The pharmacokinetics and in vivo binding specificity of the compound were studied. NOTA-P2-RM26 was labeled with F-18 within 1 h (60-65% decay corrected radiochemical yield, 55 GBq/mu mol). The radiopeptide was stable in murine serum and showed high specific binding to PC-3 cells. [F-nat]AlF-NOTA-P2-RM26 showed a low nanomolar inhibition efficiency (IC50=4.4 +/- 0.8 nM). The internalization rate of the tracer was low. Less than 14% of the cell-bound radioactivity was internalized after 4 h. The biodistribution of [F-18]AlF-NOTA-P2-RM26 demonstrated rapid blood clearance, low liver uptake and low kidney retention. The tumor uptake at 3 h p. i. was 5.5 +/- 0.7 % ID/g, and the tumor-to-blood, -muscle and -bone ratios were 87 +/- 42, 159 +/- 47, 38 +/- 16, respectively. The uptake in tumors, pancreas and other GRPR-expressing organs was significantly reduced when excess amount of non-labeled peptide was co-injected. The low uptake in bone suggests a high in vivo stability of the Al-F bond. High contrast PET image was obtained 3 h p. i. The initial biological results suggest that [F-18]AlF-NOTA-P2-RM26 is a promising candidate for PET imaging of GRPR in vivo.

Place, publisher, year, edition, pages
2013. Vol. 8, no 12, e81932- p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-214023DOI: 10.1371/journal.pone.0081932ISI: 000327947800069OAI: oai:DiVA.org:uu-214023DiVA: diva2:684230
Available from: 2014-01-07 Created: 2014-01-07 Last updated: 2017-12-06Bibliographically approved
In thesis
1. Bombesin Antagonists for Targeting Gastrin-Releasing Peptide Receptor-Positive Tumors: Design, Synthesis, Preclinical Evaluation and Optimization of Imaging Agents
Open this publication in new window or tab >>Bombesin Antagonists for Targeting Gastrin-Releasing Peptide Receptor-Positive Tumors: Design, Synthesis, Preclinical Evaluation and Optimization of Imaging Agents
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis is focused on the development, preclinical evaluation, and optimization of radiotracers for the detection of gastrin-releasing peptide receptor (GRPR)-expressing tumors. The work is divided into three distinct parts: (1) the development of bombesin (BN) antagonist (RM26)-based imaging radiotracers for the detection of GRPR-expressing tumors using different positron emission tomography (PET) and single photon emission computed tomography (SPECT) radionuclides (68Ga, 18F and 111In), (2) the establishment of a method to monitor the ligand-G protein-coupled receptor (GPCR) interaction in real time without requiring purification and stabilization of the receptors, and (3) the evaluation of radiopeptide structure-related factors (length of mini-PEG linker and composition of chelator for metal labeling) affecting the in vitro and in vivo characteristics of RM26-based tracers.

We demonstrated the possibility of high-contrast in vivo imaging of GRPR-expressing xenografts despite the physiological expression of GRPR in abdominal organs. Fast radioactivity clearance from the blood and healthy organs, including receptor-positive organs, and long retention in the tumors resulted in high tumor-to-background ratios. A novel real-time assay for measuring the kinetics of the radiotracers targeting GPCR was evaluated. Living cells were used instead of purified receptors in this technology, bringing the developmental work one step closer to the true target environment (imaging in living systems). The comparative study of 68Ga-labeled NOTA-PEGn-RM26 with di-, tri-, tetra- and hexaethylene glycol chains demonstrated that the addition of only a few units of ethylene glycol to the spacer is insufficient to appreciably affect the biodistribution of the radiopeptide. Finally, a comparative study of 68Ga-labeled PEG2-RM26 analogs N-terminally conjugated to NOTA, NODAGA, DOTA or DOTAGA highlighted the influence of the chelator on the targeting properties of the radiopeptide.

The main conclusion that can be drawn from this thesis is that 68Ga-NOTA-PEG2-RM26 has favorable biodistribution properties, such as rapid clearance from blood and tissues with physiological GRPR expression levels and long retention in GRPR-expressing tumors, and that this radiopeptide is potentially suitable for initial clinical investigation.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. 66 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 191
Keyword
Bombesin, Gastrin-releasing peptide receptor (GRPR), Antagonist, Radionuclide molecular imaging
National Category
Pharmaceutical Sciences
Identifiers
urn:nbn:se:uu:diva-232123 (URN)978-91-554-9039-3 (ISBN)
Public defence
2014-10-31, Rudbecksalen, Dag Hammarskjölds väg 20, Uppsala, 09:00 (English)
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Supervisors
Available from: 2014-10-10 Created: 2014-09-12 Last updated: 2015-01-23

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Varasteh, ZohrehÅberg, OlaVelikyan, IrinaLindeberg, GunnarSörensen, JensLarhed, MatsAntoni, GunnarSandström, MattiasTolmachev, VladimirOrlova, Anna

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