Cholecystokinin-2/gastrin receptor (CCK2R) is a metabotropic receptor. CCK2R is overexpressed on the surface of cells in certain types of tumors, allowing for its use in nuclear medicine in the field of theranostics. Several radiolabeled agonists of CCK2R are in clinical trials, however, they can be problematic due to agonism-related side effects and pharmacokinetic properties. In the somatostatin receptor (SSTR) system and its ligands, it has been found that SSTR antagonists are potentially more effective candidates for theranostic use, as they recognize a greater number of binding sites, ensure higher uptake in tumors, and thus improve diagnostic value, especially in cases of low receptor expression density. Therefore, our research group is one of the first to explore CCK2R antagonists as potential radiopharmaceuticals in the theranostics of selected cancers.
Within the group, we have developed several radiolabeled CCK2R antagonists, such as the compounds DOTA-CRL1 and DOTAGA-CRL1, which were evaluated in in vitro and in vivo preclinical tests and found to be good candidates for further improvements and eventual clinical translation. Based on the DOTA-conjugated compound, we proceeded with structure based design and in silico modifications of the tripeptide linker to improve the pharmacokinetic profile of the compound. As part of one of the master's theses, we synthesized a series of compounds with modified tripeptide linkers.
In this master's thesis, we evaluated six selected compounds using a set of in vitro biological cellular assays. A series of compounds radiolabeled with gallium-68 (⁶⁸Ga) and/or indium-111 (¹¹¹In) were evaluated for uptake and retention in cells expressing CCK2R, as well as their binding properties (determination of Kd and Bmax values), using the transfected A431-CCK2R+ cell line. To assess the extent of nonspecific internalization and binding, the A431-mock cell line (transfected with an empty vector) was used. We also examined the effects of the compounds on the metabolic activity of the selected cell lines. Radioactivity measurements were performed using a gamma counter and the results were statistically processed. We determined the logD7.4 and plasma protein binding of the compounds, as well as calculated TPSA and cLogP using software tools. Based on the results obtained, we concluded that modifications to the tripeptide linker did not represent a step forward in the translation process, as none of the compounds showed improved properties compared to our antagonist DOTAGA-CRL1, primarily due to the high nonspecific binding of the new series of compounds.
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