Radiosynthesis and evaluation of [11C]EMPA as a potential PET tracer for orexin 2 receptors

doi:10.1016/j.bmcl.2013.03.079

Bioorganic & Medicinal Chemistry Letters

Volume 23, Issue 11, 1 June 2013, Pages 3389-3392

https://doi.org/10.1016/j.bmcl.2013.03.079 Get rights and content

Abstract

EMPA is a selective antagonist of orexin 2 (OX₂) receptors. Previous literature with [³H]-EMPA suggest that it may be used as an imaging agent for OX₂ receptors; however, brain penetration is known to be modest. To evaluate the potential of EMPA as a PET radiotracer in non-human primate (as a step to imaging in man), we radiolabeled EMPA with carbon-11. Radiosynthesis of [¹¹C]N-ethyl-2-(N-(6-methoxypyridin-3-yl)-2-methylphenylsulfonamido)-N-(pyridin-3-ylmethyl)acetamide ([¹¹C]EMPA), and evaluation as a potential PET tracer for OX₂ receptors is described. Synthesis of an appropriate non-radioactive O-desmethyl precursor was achieved from EMPA with sodium iodide and chlorotrimethylsilane. Selective O-methylation using [¹¹C]CH₃I in the presence of cesium carbonate in DMSO at room temp afforded [¹¹C]EMPA in 1.5–2.5% yield (non-decay corrected relative to trapped [¹¹C]CH₃I at EOS) with ⩾95% chemical and radiochemical purities. The total synthesis time was 34–36 min from EOB. Studies in rodent suggested that uptake in tissue was dominated by nonspecific binding. However, [¹¹C]EMPA also showed poor uptake in both rats and baboon as measured with PET imaging.

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Acknowledgments

This research was carried out at the Athinoula A. Martinos Center for Biomedical Imaging at the Massachusetts General Hospital, using resources provided by the Center for Functional Neuroimaging Technologies, P41EB015896, a P41 Regional Resource supported by the National Institute of Biomedical Imaging and Bioengineering (NIBIB), National Institutes of Health. This work also involved the use of instrumentation supported by the NIH Shared Instrumentation Grant Program and/or High-End

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Although the orexin 1 receptor (OX₁R) in the brain is considered to regulate reward and feeding, the in vivo function of OX₁R has not been fully elucidated. In vivo imaging of OX₁R with positron emission tomography (PET) may be useful to further understand the molecular details of OX₁R. In this study, we newly designed and synthesized a phenylbenzofuran-2-carboxamide (PBC) derivative ([¹⁸F]PBC-1) and evaluated its utility as a PET probe targeting OX₁R in the brain. The results of cell binding assays suggested that [¹⁸F]PBC-1 has affinity for OX₁R. In an in vitro competitive inhibition assay, PBC-1 showed selective binding affinity for OX₁R (IC₅₀ = 19.5 nM) over orexin 2 receptor (IC₅₀ = 456.7 nM). Furthermore, [¹⁸F]PBC-1 displayed sufficient brain uptake for in vivo imaging with PET in a biodistribution study using normal mice, but in vivo instability was observed. These results suggest that further modifications for improvement of the pharmacokinetics are needed, but the PBC scaffold has potential for the development of useful PET probes targeting OX₁R in the brain.
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Thus, the orexin-2 receptor ligand 176 has been labeled in good formulated yield through O-methylation of an α-hydroxy-pyridinyl precursor 175 with [11C]methyl iodide in DMF with aqueous potassium carbonate as base (Scheme 57) (2013JMC6371). Other base-solvent pairs have been successful for similar 11C-O-methylation reactions, including solid potassium hydroxide-acetonitrile (2005NMB631) and cesium carbonate-DMF (2013BMCL3389). An effective radioligand for studying β-amyloid in AD, [11C]AZD4694 ([11C]3), has been synthesized in high formulated yield through N-methylation of nor-precursor 177 with [11C]methyl iodide and solid potassium hydroxide in DMF followed by acid removal of the ethoxymethyl (EOM) protecting group (Scheme 58) (2014MIB173).
Over the last half-century, the molecular imaging technique of positron emission tomography (PET) has emerged as an important tool for biomedical research, drug development and medical diagnosis. The power of PET derives from its utilization of radiotracers that can report specific information on pharmacology, biochemistry, and physiology. Many of these radiotracers are heterocycles that have been labeled with a short-lived cyclotron-produced radioisotope, either carbon-11 (t_1/2 = 20.4 min) or fluorine-18 (t_1/2 = 109.8 min). This review addresses the chemical challenges to be met in producing PET radiotracers with these radioisotope labels at heterocyclic moieties and the methods now available for doing so.
Molecular modeling technology studies of novel pyrazoylethylbenzamide derivatives as selective orexin receptor 1 antagonists
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EMPA, JNJ-10397049, MK-3697, and JNJ-42847922 are 2-SORAs. Some of the 2-SORAs are inefficient, for example, EMPA showed poor uptake in brain in rats and baboon [18]. At present, research on OX1R antagonists that play an important role in sleep-wake effect is still in its infancy.
Orexin neuropeptide signal is the core promoter of arousal, and orexin signaling is activated by orexin receptor 1 (OX₁R), which makes OX₁R an attractive target for the treatment of insomnia. In this study, OX₁R quantitative structure-activity relationship (QSAR) models (CoMFA: q² ₌0.788, r² ₌0.988; CoMSIA: q² = 0.783, r² = 0.953) were constructed based on the pIC₅₀ values of 36 pyrazoylethylbenzamide derivatives. Results indicated that the models have good reliability and predictability. Furthermore, 3D-QSAR contour maps reveal that the bioactivity of antagonists is most affected by hydrogen bond receptors. Molecular docking showed significant hydrogen bonds between key residues (ASN318 and SER103) and OX₁R pocket were determined. Seven new more selective OX₁R antagonists (DS01–DS07) were designed based on the contour maps and molecular docking results. The results of ADME prediction illustrated that the designed compounds had potential druggability and that the selectivity index value was higher than that of template molecule 33 (SI = 265). The molecular dynamics simulation results of compound 33 and DS01 indicated that their stability can be attributed to the hydrogen bonds between the nitrogen atom in the R₁ region, the fluorine atom in the R₄ region and the protein pockets. We constructed the orexin receptor 2 (OX₂R) QSAR models (CoMFA: q² = 0.852, r² = 0.979; CoMSIA: q² = 0.823, r² = 0.965) and compared them with models 1. Four potential new skeletal antagonists (Z1–Z4) were screened from the ZINC12 database containing 1,020,679 compounds based on the pharmacophore model. The molecular docking indicated that the screened compounds formed additional hydrogen bond interaction with residues GLN176 and ARG322 of OX₁R. The present study could offer theoretical guidance for future structural optimization, design, and synthesis of more selective OX₁R antagonists. Designed and screened compounds might also be as candidate compounds for related research groups.
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Orexin 1 receptor (OX₁R) is thought to be involved in various body functions, including arousal maintenance and emotional control, but the full details of its function remain unknown. OX₁R imaging with positron emission tomography (PET) would be useful in elucidating the orexin system including OX₁R, but no PET probes targeting OX₁R have been reported. We, therefore, designed and synthesized tetrahydroisoquinoline (THIQ) derivatives as novel PET probes targeting OX₁R, and evaluated their utility. In an in vitro competitive binding assay, THIQ-1 and THIQ-2 showed significantly higher binding to OX₁R (IC₅₀ = 30 and 31 nM, respectively) than OX₂R (IC₅₀ = 160 and 332 nM, respectively). These features were also observed in a cell binding assay using [¹⁸F]THIQ-1 and [¹⁸F]THIQ-2, demonstrating their OX₁R-specific binding property in vitro. In a biodistribution study using normal mice, the brain uptake of [¹⁸F]THIQ-1 was higher than that of [¹⁸F]THIQ-2, but further improvement is required for in vivo imaging with PET. Taken together, [¹⁸F]THIQ-1 and [¹⁸F]THIQ-2 have the potential to become useful imaging probes for PET targeting the OX₁R, but require additional structural changes to improve their brain uptake.
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Orexin receptors are subject to active drug discovery. Currently this is strongly directed toward insomnia, but uses will surely also be found in other indications and for other types of drugs than antagonists. This ligand discovery has also been of great value for the investigations or orexin physiology, though some of the conclusions are based on poor pharmacological thinking. The current review presents some key orexin receptor ligands, but mainly focuses on their potential uses in research and therapy, and the potential complexities—both complications and promises—of this.

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Radiosynthesis and evaluation of [11C]EMPA as a potential PET tracer for orexin 2 receptors

Abstract

Graphical abstract

Section snippets

Acknowledgments

Peptides

Proc. Natl. Acad. Sci. U.S.A.

Bioorg. Med. Chem. Lett.

Bioorg. Med. Chem. Lett.

Br. J. Pharmacol.

Br. J. Pharmacol.

Annu. Rev. Psychol.

J. Headache Pain

Headache

Nat. Med.

Handb. Exp. Pharmacol.

Physiol. Behav.

Radiosynthesis and evaluation of [¹¹C]EMPA as a potential PET tracer for orexin 2 receptors