Radiopharmaceuticals with antagonistic activity on CCK2R




01. September 2020 - 31. August 2023


1,13 FTE


Marko Anderluh


1-09 Natural sciences and Mathematics - Pharmacy

Research Organisation:




Citations for bibliographic records:





Nuclear medicine investigations have an integral role in timely identification and confirmation of cancer, staging, follow-up of treatment efficiency and identification of the disease relapse. Certain tumour types and entities can be visualized by nuclear medicine imaging techniques because of the (over)expression of different receptors for certain regulatory peptide hormones on tumour cell surface. One of these regulatory peptides is gastrin, which binds and acts through CCK2/gastrin receptors (CCK2R), specifically. The CCK2R is (over)expressed in several tumour types, including neuroendocrine neoplasms (NEN), such as medullary thyroid cancer (MTC). By the use of corresponding radiopharmaceutical - radiolabelled vector for targeting of CCK2R - and depending on the radionuclide used, these molecules can be utilized for diagnostic purposes or peptide receptor radionuclide therapy (PRRT). 
Currently, there are radiolabelled minigastrin analogues targetting CCK2R suitable for diagnostic purposes of NENs, however candidates for PRRT still need to be found. The main limiting factors of these agonistic acting compounds are either too low tumour uptake, possible nephrotoxicity and/or pentagastrin test-like adverse effect. It was generally considered, that the internalization of agonists is prerequisite for efficient molecular imaging and therapy. Consequently, non-internalizing antagonist-based radiopharmaceuticals were not of a great interest until recently, when it was proven that SSTR radiolabelled antagonists provide superior SSTR visualization than currently used SSTR agonists. In this project proposal, we suggest the same switch from agonists to antagonists for the gastrin/CCK2R system.The purpose of the project is to synthesize a series of bifunctional chelator-conjugated CCK2R antagonists using structure-based drug design methodology, biochemically evaluate these series of novel and optimized CCK2R antagonists, and compare them in in vivo pre-clinical studies to standard agonistic CCK2R analogue. The latter would finally elucidate whether the radiolabelled CCK2R antagonists outperform CCK2R agonists.
The management of MTC mainly relies on surgical resection. However, recurrent disease commonly develops, and long-term responses by radiotherapy or systemic therapy are uncommon. Although in clinical practice an evaluation of patients with MTC engages various available diagnostic options from anatomic to nuclear medicine imaging methods, there are still many patients with undetected distant metastases. Therefore, MTC remains one of the most challenging cancers since there is no well-established diagnostic imaging method or effective treatment available.
To pursue the objectives as defined above the predefined work programme will be followed. In short, novel CCK2R ligands/antagonist will be designed using structure-based in silico drug design methodology followed by synthesis of designed potential antagonists. To assure the integrity of the radiolabel, macrocyclic DOTA bifunctional chelator will be used to provide high complex stability and enable radiolabelling with variety of medicinal radionuclides. Antagonistic properties of synthesized conjugates will be tested using cell-based IP-1 functional assay to enable proper screening and SAR-based optimization. Next, the optimal DOTA-conjugated antagonist for further radiolabelling and subsequent biochemical evaluation will be selected. Biochemical evaluation of series of 111In-labelled CCK2R antagonists in comparison to standard 111In-labelled minigastrin analogue will serve as a basis of selection of best candidates for further animal studies. 
Biodistribution studies together with micro-imaging of radiolabelled antagonists in BALB/c animal model in comparison to a standard agonist will give a final indication whether CCK2R antagonist outperform agonists in the gastrin/CCK2R system.


Work package 1: Drug design & molecular modelling studies (WP1 leader: prof. dr. Marko Anderluh)
Task 1.1: Homology modelling. 
Task 1.2: Docking studies. 
 Currently undergoing.

Work package 2: Synthesis of CCK2R ligands and DOTA-conjugates (WP2 leader: prof. dr. Marko Anderluh)
Task 2.1: Synthesis of the conjugates of CCK2R binding moiety with the linker. 
 Currently undergoing.
Task 2.2: Synthesis of DOTA-conjugates. 
 Currently undergoing.

Work package 3: Radiolabelling studies (WP3 leader: dr. Petra Kolenc Peitl)
Task 3.1: Radiolabelling. 
Task 3.2: Cold labelling. 
Work package 4: Screening and biochemical evaluation (WP4 leader: dr. Petra Kolenc Peitl and prof. dr. Marko Anderluh)
Task 4.1: Cell line. 
Task 4.2: Cell membrane preparation. 
Task 4.3: Screening and SAR-based optimization. 
Task 4.4: In vitro receptor binding studies. 
Task 4.5: In vitro cell internalization studies. 
Task 4.6: In vitro enzymatic stability study and serum half-life. 
Task 4.7: Data analysis and statistics. 

Work package 5: In vivo studies (WP5 leader: dr. Petra Kolenc Peitl and prof. Dr. Melpomeni Fani)
Task 5.1: Animals. 
Task 5.2: Animal studies. 


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