Abstract: Objectives: The a7-nAChR properties are impaired in schizophrenia, brain trauma and neurodegenerative diseases. Furthermore, a7-nAChRs are overexpressed in numerous tumor cells and represent a potential target for therapy. The outstanding diversity of cellular properties mediated by neuronal and non-neuronal a7-nAChRs points to the diagnostic potential of quantitative molecular imaging of a7-nAChRs. We have developed [18F]NS10743 (KD = 7.7 nM), the first 18F-labeled radioligand for PET imaging of a7-nAChRs, and assessed the radiotracer selectivity in mice and pigs. Methods: [18F]NS10743 was synthesised by nucleophilic substitution of the nitro precursor with specific activity > 150 GBq/µmol and radiochemical purity > 99%. Its biodistribution was determined in mice at 5 min, 20 min and 60 min p.i. To determine the specificity, separate animals were pre-treated with the a7 agonist SSR180711 (10 mg/kg). Ex vivo autoradiography was performed on mice brain at 60 min after injection of 250 MBq [18F]NS10743. In vitro autoradiography was performed on porcine brain using [125I]a-bungarotoxin as a7 ligand. Furthermore, anesthetized piglets were intravenously injected with 250-500 MBq of [18F]NS10743. Selected animals additionally received 3 mg/kg SSR180711 at 10 min prior to radiotracer application followed by a continuous infusion (2 mg/kg/2h). Acquisition of dynamic PET scans (2 h) started with injection of [18F]NS10743. About 50 plasma samples were obtained, and plasma metabolites were analyzed with HPLC. Regions of interest (brain and eye) were drawn on summed PET images and distribution volumes were estimated. Results: The brain uptake of [18F]NS10743 in mice was 4.8 ± 0.4 %ID/g at 5 min, 4.0 ± 0.4 %ID/g at 20 min, and 1.6 ± 0.3 %ID/g at 60 min p.i. High initial radioactivity uptake was also observed in adrenal glands (11.3 %ID/g), spleen (7.7 %ID/g), pancreas (6.0 %ID/g), and thymus (4.4 %ID/g). Organ distribution of mice pre-treated with SSR180711 shows a significant reduction of radioactivity uptake at 60 min p.i. in adrenals, brain, thymus, and pancreas, all organs which express a7-nAChRs. Ex vivo autoradiography revealed a pattern of [18F]NS10743 binding in anatomically defined brain structures such as hippocampus and cortex which closely matches regions with high a7-nAChRs expression. Summed PET images obtained in piglets show a distribution pattern similar to in vitro autoradiographs of [125I]a-bungarotoxin with high radiotracer binding in cortex, moderate binding in cerebellum and low binding in white matter. The brain uptake is comparable to that of the a4b2 nAChR-ligand 2-[18F]F-A-85380. Application of SSR180711 resulted in more than two-fold increase of brain uptake. However, the distribution volume was decreased by about 20%, indicating inhibition of specific radiotracer binding. Furthermore, the flow-independent uptake of [18F]NS10743 in eye, which is known to express a7-nAChRs, was reduced by about 30%. Metabolite studies revealed the presence of a single major metabolite in mice and piglets, which is probably the result of an enzymatic oxidation of the nitrogen at position 1 in the diaza-bicyclononane moiety of NS10743. Conclusions: [18F]NS10743 is a high affinity ligand for a7-nAChRs with good brain uptake, specific receptor binding in vivo and good metabolic stability, which makes it suitable for neuroimaging with PET.

Peter Brust¹, Winnie Deuther-Conrad¹, Steffen Fischer¹, Achim Hiller¹, Peter-Georg Hoffmeister¹, Cornelius Donat¹, Reinhard Bauer², Elsbeth Ostergaard-Nielsen³, Daniel Brunicardi Timmermann³, Osama Sabri⁴, Joerg Steinbach¹, Dan Peters³

¹Institute of Interdisciplinary Isotope Research Leipzig, Germany, ²Institute of Molecular Cell Biology Jena, Germany, ³NeuroSearch A/S, Copenhagen, Denmark, ⁴University of Leipzig, Germany

Keywords: positron emission tomography , nicotine , neurodegeneration