Synthesis and in vitro evaluation of fluorinated diphenyloxide derivatives and sulfur analogs as serotonin transporter ligands

doi:10.1016/j.bmc.2009.10.062

Bioorganic & Medicinal Chemistry

Volume 18, Issue 1, 1 January 2010, Pages 236-241

https://doi.org/10.1016/j.bmc.2009.10.062 Get rights and content

Abstract

As the serotonin transporter (SERT) is involved in several neurodegenerative and psychiatric disorders; radiopharmaceuticals to image the SERT by PET would be valuable in studying these diseases. To this end we synthesized diphenyloxide derivatives and sulfide analogs, as new tracers, incorporating a fluorine or oxyalkyl fluorinated group on 4′ or 5′-position on phenyl ring B. Three of these exhibited good to high in vitro affinity (7 < K_i < 8 nM) and selectivity for the SERT over the other monoamine transporters.

Graphical abstract

Introduction

The serotonin transporter (SERT) plays a pivotal role in the regulation of serotoninergic neurotransmission by clearance of serotonin from the synaptic cleft via reuptake into the presynaptic neuron.1, 2 Alteration of serotoninergic neurotransmission is associated with psychiatric disorders3, 4, 5 such as depression and several neurodegenerative6, 7 such as Alzheimer’s and Parkinson’s diseases. In the brain the SERT is widely distributed in the thalamus and the raphe. In humans, SERT in vivo imaging to assess the transporter density or functionality by positron emission tomography (PET) would assist in the early diagnosis and follow-up of treatment of these diseases.8, 9 Different classes of compounds have been screened for their SERT affinity (for review see Ref. 10) such as the 3-amino-4-[2-(dimethylaminomethyl)phenylsulphanyl] benzonitrile (DASB)11, 12 or N,N-dimethyl-2-(2-amino-4-methylphenylthio)benzylamine (MADAM).¹³ These derivatives which are labeled with carbon-11 have to be used at the site of C-11 production and have to display an appropriate in vivo kinetics commensurate with the short half life of C-11 (T_1/2 = 20.4 min). Today, fluorine-18 (half-life: 109.8 min) represents one of the most attractive positron-emitting radioisotope. Its half time is long enough to do kinetic studies or plasma metabolite analysis. The radiolabeling could be performed by multi-step synthetic pathways, by aliphatic nucleophilic fluorination or aromatic electrophilic fluorination. For these reasons, several fluorinated diphenylsulfide analogs have been synthesized as PET tracers for imaging the SERT. These include: [¹⁸F]AFM,¹⁴ [¹⁸F]AFA,14, 15 [¹⁸F]AFC¹⁶ or 5-[¹⁸F]F-ADAM,¹⁷ [¹⁸F]AFE and [¹⁸F]AFP¹⁸ (Fig. 1).

Our contribution to this field has been the synthesis of several of these derivatives expanding the structure–activity profile of these classes.13, 19 As a results of these studies, we proposed that a N,N-dimethylaminomethyl group in position 1 on phenyl ring A and an amino function at position 2′ of ring B of the diphenyl sulfide structure yield compounds such as MADAM¹³ (Fig. 1) with high in vitro and in vivo affinity in monkeys²⁰ and the good selectivity for the SERT that could be attributed to the methyl in 5′-position on ring B.

Several studies have been performed evaluating the impact of substituent on the 2′-, 4′- or 5′-position on ring B (Fig. 1) but very few studies have been achieved on ring A. Recently 5-position has been studied,21, 22, 23 with a diphenylsulfide core, but to our knowledge, nothing has been performed at the 4-position.

In parallel, the substitution of the sulfur atom of IDAM by an oxygen (ODAM,²⁴ Fig. 1) results in a little effect on the in vitro SERT affinity (K_iSERT = 0.171 nM), increased brain uptake, slower kinetics and peripheral metabolism.24, 25 Vercouillie et al. have also observed this result in an in vitro study.19, 23 Furthermore, it has been shown in our previous study that for one derivative of these compounds, the sulfone analog (which could be an in vivo MADAM metabolite) does not possess any affinity for the SERT.¹⁹ For these two reasons, it could be interestingly to investigate the diphenyloxide skeleton.

So, we hypothesized that a series of fluoro (on A ring) diphenyl oxide or sulfide derivatives built on the MADAM structure should provide insight into SERT binding properties. In this paper, we report the synthesis and in vitro evaluation of eight new fluoro derivatives.

Section snippets

Chemistry

All of the compounds tested for their in vitro binding properties (7a–c and 14–16) were synthesized as shown in Scheme 1, Scheme 2. 2-Amino-4-fluoro-benzoic acid 1a was converted to a diazonium, displaced by sulfur. The intermediate disulfide was reduced with zinc in acetic acid to give the thiol 2 in 62% yield. All the carboxylic acids (2, 1b,c and 8a,b) were converted to the amides 3, 4a,b, and 9a,b, respectively, by amidification with P₂O₅ in DMF, for 2 days under reflux (36%, 52%, 85%, 38%

In vitro evaluation

In vitro affinities of derivatives 7 and 14–16 were evaluated by competition studies using tritiated ligands of SERT ([³H]citalopram), DAT ([³H]GBR12935), and NET ([³H]nisoxetine) (see Table 1). For each compound and each transporter, K_i values were only determined when 100 nM of a target compound inhibited at least 50% of tritiated ligand binding (IC₅₀ <100 nM) in accordance with the published procedure.²⁶

Results and discussion

Regarding the excellent affinity for the SERT and their good selectivity toward the two other monoamines transporters (DAT and NET), many diphenylsulfide analogs radiolabeled with carbon-11 have been described; however no suitable fluorine-18 ligand has been developed to date. Several chemical modulations have been reported on the 4- and 5-position on B ring, but A ring is not fully explored, especially the 5-position. As previously mentioned by Vercouillie et al.²³ the 5-position on A ring

Conclusion

Eight novel fluoro diphenyloxide and sulfur analog were synthesized and tested as serotonin transporter ligands for potential use as PET imaging agents. Fluorophenyl derivatives on 4- or 5-position showed good in vitro affinity and selectivity for the SERT. The position of the substitution by an O-alkyl chain influenced the affinity for the SERT. Only for the 5-position, a 3-fluoropropoxy or 4-fluoro-butoxy lead to compounds with high affinities for the SERT.

Thus, the corresponding tosylate

Chemistry

NMR spectra were recorded on a Brüker DPX Avance 200 spectrometer (200 MHz for ¹H, 50.3 MHz for ¹³C). CDCl₃ or DMSO-d₆ was used as solvent; chemical shifts are expressed in ppm relative to TMS as an internal standard. Mass spectra were obtained on a CG–MS Hewlett Packard 5989A spectrometer (electronic impact at 70 eV). The thin-layer chromatographic (TLC) analyses were performed using Merck 60-F₂₅₄ silica gel plates. Flash chromatography was used for routine purification of reaction products using

Acknowledgments

This study was funded in part by the EC-FP6-project DiMI, LSHB-CT-2005-512146, and by INSERM, Programme National de Recherche – Imagerie 2007 (A07124NS). We thank the ‘Département d’analyses Chimiques et S.R.M. biologique et médicale’ (Tours, France) for chemical analyses. K_i determinations were generously provided by the National Institute of Mental Health’s Psychoactive Drug Screening Program, Contract # NO1MH32004 (NIMH PDSP). The NIMH PDSP is directed by Bryan L. Roth MD, PhD at the

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Intramolecular interception of the Newman-Kwart rearrangement by carboxylic acids
2020, Tetrahedron Letters
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As the mass loss in DSC-TGA is only consistent with the loss of carbonylsulfide, we rationalized the formation of 2-hydroxyterephthalic acid, which requires further mass loss, from acidolysis of some product during the melt reaction utilizing the free carboxylic acid functionality in (10). Indeed, quantitative crude recoveries were obtained when the reaction was repeated on 2-((dimethylcarbamothioyl)oxy)benzoic acid and 2-((dimethylcarbamothioyl)oxy)-5-methoxybenzoic acid giving 2-hydroxy-N,N-dimethylbenzamide [20] and 2-hydroxy-5-methoxy-N,N-dimethylbenzamide [21] respectively (Figs. S28–S33, ESI). Similarly, heating (6) saw the rapid loss of 33% mass upon melting at ~200 °C in DSC-TGA, matching that expected for the loss of two molar equivalents of carbonylsulfide (32%) (Figs. S34 and S35, ESI) and pure material could be recovered in near quantitative yield after trituration and filtration.
[18F]Diphenyl sulfide derivatives for imaging serotonin transporter (SERT) in the brain
2018, Nuclear Medicine and Biology
Citation Excerpt :
Positron emission tomography (PET) imaging with suitably 18F-labeled SERT inhibitors or 123I-labeled diphenyl sulfides for single photon emission tomography (SPECT) may be useful as a method for probing pathophysiological and therapeutic mechanisms in various psychological diseases [3]. A number of SERT ligands for in vivo imaging (Fig. 1) have been developed [4–14]. [ 11C]McN5652 was the first SERT PET imaging tracer used in humans.
Serotonin transporters (SERT) play an important role in controlling serotonin concentration in the synaptic cleft and in managing postsynaptic signal transduction. Inhibitors of SERT binding are well known as selective serotonin reuptake inhibitors (SSRIs) such as fluoxetine, sertraline, paroxetine, and escitalopram, that are commonly prescribed antidepressants. Positron emission tomography (PET) and single photon emission tomography (SPECT) imaging agents targeting SERT may be useful for studying its function and providing a tool for monitoring drug treatment.
A series of novel ¹⁸F-labeled diphenyl sulfide derivatives were prepared and tested for their binding affinity. Among them, 2-((2-((dimethylamino)-methyl)-4-(2-(2-fluoroethoxy)ethoxy)phenyl)thio)aniline, 1, which showed excellent binding toward serotonin transporter (SERT) in the brain (K_i = 0.09 nM), was selected for further evaluation. An active OTs intermediate, 7, was treated with [¹⁸F]F⁻/K₂₂₂ to provide [¹⁸F]1 in one step and in high radiochemical yields. This new SERT targeting agent was evaluated in rats by biodistribution studies and animal PET imaging studies.
The radiolabeling reaction led to the desired [¹⁸F]1. After HPLC purification no-carrier-added [¹⁸F]1 was obtained (radiochemical yield, 23–47% (n = 10,); radiochemical purity >99%; molar activity, 15–28 GBq/μmol). Biodistribution studies with [¹⁸F]1 showed good brain uptake (1.04% dose/g at 2 min post-injection), high uptake into the hypothalamus (1.55% dose/g at 30 min), and a high target-to-non-target (hypothalamus to cerebellum) ratio of 6.1 at 120 min post-injection. A PET imaging study in normal rats showed excellent uptake in the midbrain and thalamus regions known to be rich in SERT binding sites at 60 min after iv injection. Chasing experiment with escitalopram (iv, 2 mg/kg) in a rat at 60 min after iv injection caused a noticeable reduction in the regional radioactivity and the target-to-non-target ratio, suggesting binding by [¹⁸F]1 was highly specific and reversible for SERT binding sites in the brain.
A novel diphenyl sulfide derivative, [¹⁸F]1 for SERT imaging was successfully prepared and evaluated. Results suggest that this new chemical entity is targeting SERT binding sites in the brain, and it is a suitable candidate for future commercial development.
One-step preparation of [18F]FPBM for PET imaging of serotonin transporter (SERT) in the brain
2016, Nuclear Medicine and Biology
Citation Excerpt :
In vivo positron emission tomography (PET) imaging of SERT binding in the brain may be useful as a method for probing pathophysiological and therapeutic mechanisms in various psychological diseases [8–10] such as OCD [11], eating disorders, and addiction [3,4,12]. A number of SERT ligands for in vivo imaging (Fig. 1) have been developed [13–25]. [ 11C]McN5652 was the first SERT PET imaging tracer used in humans.
Serotonin transporters (SERT) in the brain play an important role in normal brain function. Selective serotonin reuptake inhibitors such as fluoxetine, sertraline, paroxetine, escitalopram, etc., specifically target SERT binding in the brain. Development of SERT imaging agents may be useful for studying the function of SERT by in vivo imaging. A one-step preparation of [¹⁸F]FPBM, 2-(2′-(dimethylamino)methyl)-4′-(3-([¹⁸F]fluoropropoxy)phenylthio)benzenamine, for positron emission tomography (PET) imaging of SERT binding in the brain was achieved. An active OTs intermediate, 9, was reacted with [¹⁸F]F⁻/K₂₂₂ to produce [¹⁸F]FPBM in one step and in high radiochemical yield. This labeling reaction was evaluated and optimized under different temperatures, bases, solvents, and varying amounts of precursor 9. The radiolabeling reaction led to the desired [¹⁸F]FPBM in one step and the crude product was purified by HPLC purification to give no-carrier-added [¹⁸F]FPBM (radiochemical yield, 24–33%, decay corrected; radiochemical purity > 99%). PET imaging studies in normal monkeys (n = 4) showed fast, pronounced uptakes in the midbrain and thalamus, regions known to be rich in SERT binding sites. A displacement experiment with escitalopram (5 mg/kg iv injection at 30 min after [¹⁸F]FPBM injection) showed a rapid and complete reversal of SERT binding, suggesting that binding by [¹⁸F]FPBM was highly specific and reversible. A one-step radiolabeling method coupled with HPLC purification for preparation of [¹⁸F]FPBM was developed. Imaging studies suggest that it is feasible to use this method to prepare [¹⁸F]FPBM for in vivo PET imaging of SERT binding in the brain.
Mapping the distribution of serotonin transporter in the human brainstem with high-resolution PET: Validation using postmortem autoradiography data
2016, NeuroImage
Citation Excerpt :
For each region of interest, specific binding was calculated by subtracting nonspecific binding, defined in the presence of 10 μM fluoxetine, from the total [3H]MADAM binding. Finally, for comparison purposes, binding density values were converted into the in vitro analog of BPND (Innis et al., 2007) using the in vitro Bmax value (density of the protein in vitro) as an estimate of Bavail (density of the available protein to bind the radioligand in vivo) and in vitro estimates of KD (dissociation kostant) and fND (free fraction in the non displaceable compartment) obtained from the literature (Mavel et al., 2010; Fridén et al., 2014). Pearson correlation analysis was used to assess the correlation between estimates of regional BPND and VOI volumes obtained with the template-based and individual-based methods of regional BPND.
The human brainstem is a complex structure with several small nuclei and neural pathways of interest in the pathophysiology of central nervous system (CNS) disorders. In common with other monoaminergic systems, serotoninergic neurons originate from a group of nuclei located in the brainstem. The present study was designed to validate a user-independent approach for a detailed in vivo quantification of serotonin transporter (5-HTT) availability in the human brainstem using a template-based approach that consisted of three steps. First, 3 T-MR images and parametric binding potential (BP_ND) [¹¹C]MADAM images of ten healthy subjects were used to generate a PET template of 5-HTT availability. In the second step, volumes of interest (VOIs) for different brainstem nuclei were obtained using a method in which VOIs are initially delineated on MRI images using anatomical landmarks and then are finally tailored on the distribution of 5-HTT binding using a thresholding approach applied to the 5-HTT template. In the final step, the VOIs were transformed and applied individually to BP_ND images of 16 healthy subjects (14 M/2F, 20–64 years). The in vivo distribution of BP_ND values obtained with the template-based method were in good agreement with an individual-based approach taken as gold standard. Results were also in agreement with 5-HTT quantification using in vitro binding data obtained with autoradiography (ARG) studies using [³H]MADAM. The proposed template-based method can be applied to PET data acquired in several CNS disorders in which serotonin neurons in the brainstem might be affected.
An improved preparation of [18F]FPBM: A potential serotonin transporter (SERT) imaging agent
2013, Nuclear Medicine and Biology
Citation Excerpt :
The imaging study may also be employed for examining neurodegenerative diseases (such as Alzheimer's disease and Parkinson's disease) that involve the serotonergic system [14–18]. A number of SERT ligands for in vivo imaging have been developed [19–34]. Ligands with a core structure of diarylsulfide showed the most promising results as in vivo imaging agents (Fig. 1).
In vivo positron emission tomography (PET) imaging of the serotonin transporter (SERT) is a valuable tool in drug development and in monitoring brain diseases with altered serotonergic function. We have developed a two-step labeling reaction for the preparation of the high serotonin affinity ligand [¹⁸F]FPBM ([¹⁸F]2-(2′-((dimethylamino)methyl)-4′-(3-fluoropropoxy)phenylthio)benzenamine, 1).
To improve and automate the radiolabeling of [¹⁸F]FPBM, 1, an intermediate, [¹⁸F]3-fluoropropyltosylate, [¹⁸F]4, was prepared first, and then it was reacted with the phenol precursor (4-(2-aminophenylthio)-3-((dimethylamino)methyl)phenol, 3) to afford [¹⁸F]FPBM, 1. To optimize the labeling, this O-alkylation reaction was evaluated under different temperatures, using different bases and varying amounts of precursor 3. The desired product was obtained after a solid phase extraction (SPE) purification.
This two-step radiolabeling reaction successfully produced the desired [¹⁸F]FPBM, 1, with an excellent radiochemical purity (> 95%, n = 8). Radiochemical yields were between 31% and 39% (decay corrected, total time of labeling: 70 min, n = 8). The SPE purification cannot completely remove pseudo-carriers in the final dose of [¹⁸F]FPBM, 1. The concentrations of major pseudo-carriers were measured by UV-HPLC (476–676, 68–95 and 50–71 μg for precursor 3, O-hydroxypropyl and O-allyloxy derivatives, 5 and 6, respectively). To investigate the potential inhibition of SERT binding of these pseudo-carriers, we performed in vitro competition experiments evaluated by autoradiography. Known amounts of ‘standard’ FPBM, 1, of the pseudo-carriers, 5 and 6, were added to the HPLC-purified [¹⁸F]1 dose. The inhibition of ‘standard’ FPBM, 1, binding to the SERT binding sites, using monkey brain sections, were measured (EC₅₀ = 13, 46, 7.1 and 8.3 nM, respectively for 1, precursor 3, O-hydroxypropyl and O-allyloxy derivative of 3).
An improved radiolabeling method by a SPE purification for preparation of [¹⁸F]FPBM, 1, was developed. The results suggest that it is feasible to use this labeling method to prepare [¹⁸F]FPBM, 1, without affecting in vivo SERT binding.
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Bioorganic & Medicinal Chemistry

Abstract

Graphical abstract

Introduction

Section snippets

Chemistry

In vitro evaluation

Results and discussion

Conclusion

Chemistry

Acknowledgments

References and notes (26)

Biol. Psychiatry

Lancet

Neuroscience

Eur. Neuropsychopharmacol.

Neurosci. Biobehav. Rev.

Appl. Radiat. Isot.

Bioorg. Med. Chem. Lett.

Bioorg. Med. Chem. Lett.

Nucl. Med. Biol.

Pharmacol. Rev.

Pharmacol. Rev.

Clin. Chem.

Neuroreport

Cited by (9)

Intramolecular interception of the Newman-Kwart rearrangement by carboxylic acids

[<sup>18</sup>F]Diphenyl sulfide derivatives for imaging serotonin transporter (SERT) in the brain

One-step preparation of [<sup>18</sup>F]FPBM for PET imaging of serotonin transporter (SERT) in the brain

Mapping the distribution of serotonin transporter in the human brainstem with high-resolution PET: Validation using postmortem autoradiography data

An improved preparation of [<sup>18</sup>F]FPBM: A potential serotonin transporter (SERT) imaging agent

Novel Diarylthioether Compounds as Agents for the Treatment of Chagas Disease