Imaging of α7 nicotinic acetylcholinereceptors (nAChRs): Automated synthesis and biological evaluation of [¹⁸F]NS14490 in mice and pig

Aim
There is evidence that α7nAChRs play an important role in cancer and neurodegenerative diseases. Therefore, quantitative imaging of α7nAChRs using PET represents a new approach for investigation of those diseases.
Due to its high target affinity and selectivity [¹⁸F]NS14490, an oxadiazolyl-diazabicyclononane derivative, is a promising radiotracer for α7nAChRs imaging. Manual radiosynthesis of [¹⁸F]NS14490 and its in vivo evaluation in mice were already reported [1]. The subsequent assessment of its imaging potential by dynamic PET studies in piglets and PET/CT studies in tumor-bearing mice required the transfer of the manual synthesis to an automated synthesis module.

Materials and Methods
For dynamic PET studies, the manual radiosynthesis of [¹⁸F]NS14490 [1] was transferred to an automatic synthesis module (Tracerlab FX-N) including azeotropic drying of [¹⁸F]F- with Kryptofix K222/K₂CO₃ in acetonitrile, direct nucleophilic substitution at the precursor, semipreparative HPLC and solid phase extraction. After evaporation the radiotracer was formulated in phosphate buffered saline containing 5% ethanol.
Dynamic PET studies in piglets (female, 15-18 kg) were performed under control and blocking conditions (n=2 each) with a highly selective α7nAChRs ligand (NS6740; bolus: 3 mg kg-¹ h^-1; infusion: 1 mg kg^-1 h^-1) for 4 hours. The metabolism of [¹⁸F]NS14490 in piglets was analyzed by chromatography of plasma samples.
PET/CT studies were performed in tumor-bearing nude mice under control and blocking conditions with NS6740.

Results and Conclusion
During transfer of radiosynthesis into an automated synthesis module, solid phase extraction was challenging. For desorption of the radiotracer from divinyl/polystyrene cartridge, the elution solvent ethanol/acetic acid used in manual synthesis was replaced by acetonitrile/formic acid resulting in more efficient elution, formerly not accomplished. The radiotracer was achieved within 1.25 hours and radiochemical yields (32%), radiochemical purity (> 90%) and specific activity (> 150 GBq μmol_-1) were comparable to the manual synthesis [1]. Brain uptake of [¹⁸F]NS14490 peaked in piglets at 3 min p.i. and is with a value of SUV_max = 0.504 about tenfold higher than in mice. Blocking by NS6740 decreased the specific uptake in brain by 28% (SUV 240 min p.i. 0.174 vs. 0.125 under baseline and blocking conditions, respectively).The metabolic stability of [¹⁸F]NS14490 in piglets is lower than in mice. At 60 min p.i. intact radiotracer represented 30% of plasma activity in piglets compared to 55% in mice.
Preliminary animal PET/CT studies in tumor-bearing mice provide evidence that [¹⁸F]NS14490 may also be suitable for tumor imaging.
[1] Rötering, S. et al. Bioorg. Med. Chem. 2013