In this communication, we report the synthesis and characterization of a library of small molecule antagonists of the human gonadotropin releasing hormone receptor based upon the 2-(4-tert-butylphenyl)-4-piperazinyl-benzimidazole scaffold via Cu-catalysed azide alkyne cycloaddition.
Herein, we report a straightforward sequential acylation-Finkelstein approach to achieve iodination of amine containing bioactives. The utility was demonstrated by successful radiolabelling with 123 I in high radiochemical yield. Moreover, microwave-assisted Finkelstein reaction can be employed to enhance conversion and reaction rates to obtain the desired iodides. The method is of interest for radioiodination of amine-containing bioactives. The mechanistic details of the iodination process were studied by kinetics and density functional theory calculations, which revealed the mechanistic complexity of the reaction involving amide group anchimeric assistance. We disclose a number of fundamental aspects of amide group anchimeric assistance in substitution reactions.
The increased expression of gonadotropin releasing hormone receptor (GnRH-R) in brain has been strongly linked to Alzheimer disease. Therefore, the development of radiolabeled imaging agents for GnRH-R is relevant for early diagnosis of Alzheimer disease. We have recently disclosed the discovery of two promising compounds displaying nanomolar-range affinity for the GnRH-R. In the present study, a preclinical evaluation of the compound properties was performed to evaluate their potential as single photon emission computed tomography (SPECT) radiotracers for imaging the GnRH-receptor. The compounds were assessed in vitro by performing serum stability analysis by human and rat serum, metabolic profiling by human liver microsomes, and exploratory rat brain autoradiography. The investigated compounds displayed satisfactory stability against human, rat serum, and liver microsomal metabolism, which favors their potential as SPECT-imaging agents. Additionally, we identified and quantified the formation rate of the metabolites by fragmentation of up to LIST OF ABBREVIATIONS: GNRH, gonadotropin releasing hormone; GNRH-R, gonadotropin releasing hormone receptor; HPG, hypothalamic pituitary gland; HPLC-MS/MS, high performance liquid chromatography mass spectrometry; IHC, immunohistochemistry; LC-MS, liquid chromatography mass spectrometry; LHRH, luteinizing hormone releasing hormone; NADP, nicotinamide adenine dinucleotide phosphate; NADPH, dihydronicotinamide adenine dinucleotide phosphate; SPECT, single photon emission computed tomography.
72J Label Compd Radiopharm. 2020;63:72-84. wileyonlinelibrary.com/journal/jlcr five mass spectrometric stages. The GnRH-R rat brain specificity of these compounds was tested in competition with a known ligand for the receptor and the in vitro autoradiography confirmed that compounds 3 and 4 binds to rat GnRH-R in different rat brain regions.
K E Y W O R D Sautoradiography, gonadotropin, in vitro, metabolic profiling, SPECT
We report the synthesis, radiosynthesis and biological characterisation of two gonadotropin‐releasing hormone receptor (GnRH−R) antagonists with nanomolar binding affinity. A small library of GnRH−R antagonists was synthesised in 20–67 % overall yield with the aim of identifying a high‐affinity antagonist capable of crossing the blood–brain barrier. Binding affinity to rat GnRH−R was determined by autoradiography in competitive‐binding studies against [125I]buserelin, and inhibition constants were calculated by using the Cheng–Prusoff equation. The radioligands were obtained in 46–79 % radiochemical yield and >95 % purity and with a molar activity of 19–38 MBq/nmol by direct nucleophilic radiofluorination. Positron emission tomography imaging in rat under baseline conditions in comparison to pretreatment with a receptor‐saturating dose of GnRH antagonist revealed saturable uptake (0.1 %ID/mL) into the brain.
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