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Facile synthesis of carbon-11-labeled arylpiperazinylthioalkyl derivatives as new PET radioligands for imaging of 5-HT1AR
Applied Radiation and Isotopes 70 (2012) 498–504
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Facile synthesis of carbon-11-labeled arylpiperazinylthioalkyl derivatives as new PET radioligands for imaging of 5-HT1AR Mingzhang Gao, Min Wang, Qi-Huang Zheng n Department of Radiology and Imaging Sciences, Indiana University School of Medicine, 1345 West 16th Street, L3-208, Indianapolis, IN 46202-2111, USA
a r t i c l e i n f o
a b s t r a c t
Article history: Received 12 August 2011 Received in revised form 26 October 2011 Accepted 14 November 2011 Available online 25 November 2011
Carbon-11-labeled arylpiperazinylthioalkyl derivatives, 2-((4-(4-(2-[11C]methoxyphenyl)piperazin1-yl)butyl)thio)benzo[d]oxazole ([11C]5a), 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)butyl)thio)5,7-dimethylbenzo[d]oxazole ([11C]5c), 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)butyl)thio) benzo[d]thiazole ([11C]5e), 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]oxazole 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7-dimethylbenzo[d]oxazole ([11C]5g), ([11C]5i), and 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]thiazole ([11C]5k), were prepared from their corresponding phenol precursors with [11C]CH3OTf through O-[11C]methylation and isolated by a simplified solid-phase extraction (SPE) method using a Sep-Pak Plus C18 cartridge in 50–60% (n ¼5) radiochemical yields based on [11C]CO2 and decay corrected to end of bombardment (EOB). The overall synthesis time from EOB was 23 min, the radiochemical purity was 499%, and the specific activity at end of synthesis (EOS) was 277.57 92.5 GBq/mmol (n ¼5). & 2011 Elsevier Ltd. All rights reserved.
Keywords: Carbon-11-labeled arylpiperazinylthioalkyl derivatives Positron emission tomography (PET) Serotonin-1A receptor (5-HT1AR) Radioligands Imaging
1. Introduction Serotonin (5-hydroxytryptamine, 5-HT) is an important neurotransmitter that is involved in various physiological and pathological processes in peripheral and central nervous system (Siracusa et al., 2008; Yoshida et al., 2005). To date at least 14 different serotonin receptors (5-HTRs) have been identified, and these receptors can be divided into distinct families—denoted by numbers 1, 2, 3, 4, 5, 6, and 7, with subtypes in each family denoted by letters like a, b, and c (Savitz et al., 2009). Among 5-HTRs, the 5-HT1AR was one of the first serotonin receptor subtypes pharmacologically characterized and is the best studied (Caliendo et al., 2005; Lacivita et al., 2008). 5-HT1AR is generally accepted to be implicated in the pathophysiology of major neuropsychiatric disorders, including depression, suicidal behavior, panic disorder, epilepsy, bulimia, schizophrenia, Parkinson’s disease, and Alzheimer’s disease (Akimova et al., 2009; Drevets et al., 2007; Kumar and Mann, 2007; la Fougere et al., 2009). Therefore, 5-HT1AR is an important target for drug therapy, and numerous new therapeutic agents 5-HT1AR ligands belonging to different chemical classes have been developed in recent years (Lacivita et al., 2008). 5-HT1AR ligands can be grouped into two classes: antagonistic and agonistic ligands. 5-HT1AR is overexpressed in the brain. Because of the higher concentration and
n
Corresponding author. Tel.: þ317 278 4671; fax: þ317 278 9711. E-mail address: [email protected] (Q.-H. Zheng).
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heterogeneous distribution of this receptor in living brain, 5-HT1AR is one of the most attractive targets for noninvasive in vivo imaging of the related brain diseases using biomedical imaging technique positron emission tomography (PET), and various PET radioligands employed for quantification of brain 5-HT1AR have been developed accordingly (Kumar and Mann, 2007). Likewise, 5-HT1AR radioligands can be grouped into two classes: antagonistic and agonistic radioligands. Most of the 5-HT1AR radioligands belong to two structural families: (1) compounds structurally similar to the 5-HT1A antagonist WAY100635 (N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclohexane carboxamide), and (2) derivatives of the 5-HT1A agonist 8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)tetralin). Among these radioligands, [carbonyl-11C]WAY100635 (KD ¼ 0.2 nM for 5-HT1AR) is the most commonly used 5-HT1AR radioligand for in vivo human studies (Brooks and Piccini, 2006; Hall et al., 1997; Kumar and Mann, 2007; Turner et al., 2005). Despite recognition of the potential value of 5-HT1AR radioligands, there is a requirement for continued in vivo investigation using superior agents to establish the clinical utility of this strategy, before these radiotracers are clinically approved (Moghbel et al., 2011). Long-chain arylpiperazine derivatives are one of the most important classes of 5-HT1AR ligands. Recently, a new series of arylpiperazinylthioalkyl derivatives has been found to be potent and selective 5-HT1AR ligands (Siracusa et al., 2008). These ligands possess 2-methoxyphenyl amenable to labeling with the positron emitting radionuclide carbon-11 as 5-HT1AR radioligands. Arylpiperazinylthioalkyl derivatives labeled with carbon-11 as radiotracers using
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
PET may enable noninvasive monitoring of the density of 5-HT1AR and its response to 5-HT1AR antagonist and agonist treatment. Here we report the design, synthesis, and radiosynthesis of novel 5-HT1AR radioligands carbon-11-labeled arylpiperazinylthioalkyl derivatives, 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d]oxazole ([11C]5a), 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl) butyl)thio)-5,7-dimethylbenzo[d]oxazole ([11C]5c), 2-((4-(4-(2-[11C] methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d]thiazole ([11C]5e), 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]o xazole ([11C]5g), 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7-dimethylbenzo[d]oxazole ([11C]5i), and 2-((6-(4-(2-[11C] methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]thiazole ([11C]5k).
2. Results and discussion 2.1. Chemistry Six title compounds, 2-((4-(4-(2-methoxyphenyl)piperazin1-yl)butyl)thio)benzo[d]oxazole (5a), 2-((4-(4-(2-methoxyphenyl)piperazin-1-yl)butyl)thio)-5,7-dimethylbenzo[d]oxazole (5c), 2-((4-(4-(2-methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d] thiazole (5e) 2-((6-(4-(2-methoxyphenyl)piperazin-1-yl)hexyl) thio)benzo[d]oxazole (5g), 2-((6-(4-(2-methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7-dimethylbenzo[d]oxazole (5i), and 2((6-(4-(2-methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]thiazole (5k), served as the reference standards and selected for radiolabeling. Compounds 5a, 5e, 5g, and 5k have been reported previously (Siracusa et al., 2008). These arylpiperazinylthioalkyl derivatives are potent and selective 5-HT1A ligands with low nanomolar Ki values, and their reported binding data with 5-HT1A, a1-adrenergic, D1, D2, and 5-HT2A receptors and calculated selectivity are summarized in Tables 1 and 2 (Siracusa et al., 2008). In particular, representative compound 5a displayed very high 5-HT1AR affinity (Ki 0.094 nM) and selectivity over all other investigated receptors including a1-adrenergic (Ki ratio 830), D1 (Ki ratio 17021), D2 (Ki ratio 397), and 5-HT2A (Ki ratio 3170) receptors. Compounds 5c and 5i are new compounds. Previous study on the structure–activity relationship (SAR) of 2(4-methylpiperazinyl)benzoxazole derivatives, 5-HT3 ligands indicated that introduction of small lipophilic substituents such as CH3 at the 5- and 7-position of the benzoxazole ring greatly increased the affinity for the 5-HT3 receptor (Gao et al., 2008a; Sato et al., 1998; Yoshida et al., 2005). Thus, we hypothesize that 5,7-dimethylbenzoxazole derivatives 5c and 5i will exhibit higher affinity for the 5-HT1AR than known benzoxazole derivatives 5a, 5e, 5g, and 5k without bearing any lipophilic substituent in the benzoxazole ring, and new compounds 5c and 5i were designed and synthesized in this regard. Further SAR study of the substituent effect is currently under way to confirm this hypothesis. Arylpiperazinylalkylthio benzoxazole or benzothiazole derivative reference standards and precursors were obtained by two-step condensation as outlined in Scheme 1, according to the published
Table 1 5-HT1A and a1-adrenergic receptors binding data of arylpiperazinylthioalkyl derivative standards 5a, 5e, 5g, and 5k (Siracusa et al., 2008). Compound
5a 5e 5g 5k
Ki (nM)
Ki ratio
5-HT1A
a1
a1/5-HT1A
0.094 70.008 0.27 70.01 0.52 70.01 1.30 70.004
78.07 23.2 16.67 1.4 52.57 7.4 46.07 7.3
830 61 101 35
499
Table 2 D1, D2, and 5-HT2A receptors binding data of arylpiperazinylthioalkyl derivative standards 5a, 5e, and 5g (Siracusa et al., 2008). Compound Ki (nM) D1
5a 5e 5g
Ki ratio D2
5-HT2A
D1/5HT1A
16007 82 37.3 7 8.3 298 7 44 17021 20057 388 58.3 7 12.2 106 7 23 7426 922 7 85 46.6 7 7.0 207 7 19 1773
D1/5HT1A
5-HT1A/5HT1A
397 216 90
3170 393 398
procedures (Siracusa et al., 2008) with slight modifications. The simple old method was used for preparation of a series of new compounds. The commercially available starting materials benzo [d]oxazole-2-thiol (1a), 5,7-diemthyl benzo[d]oxazole-2-thiol (1b), or benzo[d]thiazole-2-thiol (1c) were reacted with appropriate 1-bromo-4-chlorobutane or 1-bromo-6-chlorohexane (2) in acetonitrile at reflux for 1.5 h in the presence of potassium carbonate to give compounds 3a–f with high chemical yields (65–82% purified yield). The intermediates 3a–f were then reacted with 1-(2-methoxyphenyl)piperazine in acetonitrile at reflux for 30 h in the presence of potassium carbonate and potassium iodide to obtain the desired final standard products 5a, 5c, 5e, 5g, 5i, and 5k. The precursor compounds 5b, 5d, 5f, 5h, 5j, and 5l were designed and prepared by the same procedure as their corresponding standard compounds, using just 2-(piperazin-1-yl)phenol instead of 1-(2-methoxyphenyl)piperazine. The chemical yields of compounds 5a–l varied from 22% to 61%. In comparison with the literature methods, the slight modifications included the change of stoichiometric ratio of the starting materials, and the change of reaction conditions and work-up procedures, such as the use of different base, the extension of the reflux reaction time, and the use of the column chromatography for further purification. These modifications improved the chemical yield and chemical purity of the intermediates, standards, and precursors we prepared. 2.2. Radiochemistry Synthesis of carbon-11-labeled arylpiperazinylalkylthio benzoxazole or benzothiazole derivatives, [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k, is indicated in Scheme 2. Phenolic hydroxyl precursors 5b, 5d, 5f, 5h, 5j, and 5l were labeled by a reactive [11C]methylating agent, [11C]methyl triflate ([11C] CH3OTf) (Jewett, 1992; Mock et al., 1999) prepared from [11C]CO2, under basic conditions (2N NaOH) in acetonitrile through the O-[11C11C]methylation and isolated by a simplified solid-phase extraction (SPE) method (Gao et al., 2008b, 2009) to provide target tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k in 50–60% (n¼5) radiochemical yields, decay corrected to end of bombardment (EOB), based on [11C]CO2. The large polarity difference between the phenolic hydroxyl precursor and the corresponding labeled O-methylated ether product permitted the use of SPE technique for purification of the labeled product from the radiolabeling reaction mixture. A C-18 Plus Sep-Pak cartridge was used in SPE purification technique. The crude reaction mixture was treated with aqueous NaHCO3 and loaded onto the cartridge by gas pressure. The pH of freshly prepared 0.1 M NaHCO3 might be too low to effectively deprotonate a phenol. However, an excess of 2N NaOH used in the reaction provided a final pH after addition of 0.1 M NaHCO3 high enough to deprotonate all phenol. Any non-reacted phenolic hydroxyl precursor was actually converted to the corresponding sodium salt, and any non-reacted [11C]CH3OTf was actually hydrolyzed to [11C]CH3OH, which would not be trapped to the C-18 Sep-Pak. The cartridge was washed with water to remove non-reacted [11C]CH3OTf, remaining phenolic hydroxyl precursor,
500
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
R1
N
Cl
SH
X
n
Br 2
R1
X
i
R1
N X
Cl
HN
n
S
R2
N
n
S
O
ii
3a, n = 2, R1 = H, X = O 3b, n = 2, R1 = Me, X = O 3c, n = 2, R1 = H, X = S 3d, n = 4, R1 = H, X = O 3e, n = 4, R1 = Me, X = O 3f, n = 4, R1 = H, X = S
N
N 4
R1
1a, R1 = H, X = O 1b, R1 = Me, X = O 1c, R1 = H, X = S
R1
N
O
R2 R1 5a, n = 2, R1 = H, X = O, R2 = Me 5b, n = 2, R1 = H, X = O, R2 = H 5c, n = 2, R1 = Me, X = O, R2 = Me 5d, n = 2, R1 = Me, X = O, R2 = H 5e, n = 2, R1 = H, X = S, R2 = Me 5f, n = 2, R1 = H, X = S, R2 = H 5g, n = 4, R1 = H, X = O, R2 = Me 5h, n = 4, R1 = H, X = O, R2 = H 5i, n = 4, R1 = Me, X = O, R2 = Me 5j, n = 4, R1 = Me, X = O, R2 = H 5k, n = 4, R1 = H, X = S, R2 = Me 5l, n = 4, R1 = H, X = S, R2 = H Scheme 1. Synthesis of arylpiperazinylthioalkyl derivatives 5a–l. Reagents, conditions, and yields: (i) K2CO3/CH3CN, reflux; 65–82% yield. (ii) KI, K2CO3/CH3CN, reflux; 22–61% yield.
R1
N X
N S
n
N
i
R1
N
HO
R1 5b, n = 2, R1 = H, X = O 5d, n = 2, R1 = Me, X = O 5f, n = 2, R1 = H, X = S 5h, n = 4, R1 = H, X = O 5j, n = 4, R1 = Me, X = O 5l, n = 4, R1 = H, X = S
X
N S
n
N H311CO
1
R
[11C]5a, n = 2, R1 = H, X = O [11C]5c, n = 2, R1 = Me, X = O [11C]5e, n = 2, R1 = H, X = S [11C]5g, n = 4, R1 = H, X = O [11C]5i, n = 4, R1 = Me, X = O [11C]5k, n = 4, R1 = H, X = S
Scheme 2. Synthesis of carbon-11-labeled arylpiperazinylthioalkyl derivatives [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k. Reagents, conditions, and yields: (i) [11C]CH3OTf, CH3CN, 2N NaOH, 80 oC; 50–60% decay corrected radiochemical yield.
and reaction solvent, and total 6 mL (2 3 mL) volume of water was enough to wash off all phenol. The final labeled product was eluted with ethanol (2 2 mL), concentrated by rotary evaporation, and reformulated in saline (10 mL). In our fully automated radiosynthesis module (Mock et al., 2005b), it is difficult to directly elute the labeled product from a C-18 Sep-Pak to a vial using either 1 1 mL ethanol or 2 0.5 mL ethanol, due to the back pressure in the C-18 Sep-Pak and dead volume in the transfer tubing. In order to elute most of the labeled product from the C-18 Sep-Pak, we need to increase the volume of the eluent ethanol. For the radiotracers produced for animal study, we used 2 2 mL ethanol for elution, and evaporation was required before reformulation. For the radiotracer produced for human study, we used 2 1 mL ethanol, no evaporation was required, and a C-18 Sep-Pak was used for direct reformulation (Gao et al., 2010; Wang et al., 2011). Overall synthesis time was 23 min from EOB, including approximately 11 min for [11C]CH3OTf production, 5 min for O-[11C]methylation reaction, and 7 min for SPE purification, evaporation, and reformulation.
SPE technique is fast, efficient, and convenient and works very well for the O-methylated ether tracer purification using the phenolic hydroxyl precursor for radiolabeling (Gao et al., 2008b). The radiosynthesis was performed in an automated selfdesigned multi-purpose 11C-radiosynthesis module, allowing measurement of specific activity at EOB during synthesis (Mock et al., 2005a,b). Online determination of specific activity at EOB is accurate when high performance liquid chromatography (HPLC) is used as purification method. However, the on-the-fly technique to determine specific activity at EOB is not applied when SPE is used as the purification method. In general, the specific activity for all 11C-tracers produced in our PET radiochemistry facility is basically same, because the specific activity of a carrier-free radionuclide sample (11C) is only related to its half-life (20.4 min). In particular, the specific activity for all 11C-tracers produced in our facility is dependent on two parts: (1) carrier from the 11C gas target, and (2) carrier from the 11C radiolabeled precursor [11C]CH3OTf production system. The 11C gas target we used is the Siemens RDS-111 Eclipse cyclotron 11C gas target.
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
501
mV
600.00 400.00 200.00 0.00 1.00
2.00
3.00
4.00
5.00 6.00 Minutes
7.00
8.00
9.00
10.00
1.00
2.00
3.00
4.00
5.00 6.00 Minutes
7.00
8.00
9.00
10.00
AU
0.006 0.004 0.002 0.000
Fig. 1. A representative analytical HPLC chromatographic profile for the tracer [11C]5a produced by SPE purification: analytical radioactive (A) and UV (B) traces, tR [11C]5a ¼ 5.32 and 5.22 min, respectively.
The [11C]CH3OTf production system we used is an Eckert & Ziegler Modular Lab C-11 Methyl Iodide/Triflate module (Mock et al., 1999). The specific activity for our 11C-tracers usually ranges from 370 to 925 GBq/mmol at EOB according to our previous works. Actually the specific activity of all our 11C-tracers is essentially the same, because in the same radiochemistry facility, the same targetry conditions, C-11 Methyl Iodide/Triflate module, and 11Cradiosynthesis module are employed. The specific activity of carbon-11-labeled arylpiperazinylthioalkyl derivatives was estimated in the range of 185–370 GBq/mmol at the end of synthesis (EOS) based on other compounds produced in our facility using the same targetry conditions, which have been measured by the on-the-fly technique or the same SPE purification method (Mock et al., 2005b; Zheng et al., 2007). The actual measurement of specific activity at EOS was performed by analytical HPLC (Zheng and Mock, 2005) and calculated. The exact values of the specific activity for the tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k were 277.5 792.5 GBq/mmol (n¼5), which are in agreement with the estimated values and the ‘‘online’’ determined values. Chemical purity and radiochemical purity were determined by analytical HPLC (Zheng and Mock, 2005). The chemical purity of the precursors and reference standards was 496%. The radiochemical purity of the target tracers was 499% determined by radio-HPLC through g-ray (PIN diode) flow detector, and the chemical purity of the target tracers was 495% determined by reversed-phase HPLC through UV flow detector. Representative analytical HPLC chromatograms for the target tracer [11C]5a produced by SPE purification are shown in Fig. 1.
3. Experimental 3.1. General All commercial reagents and solvents were purchased from Sigma-Aldrich and Fisher Scientific and used without further purification. [11C]CH3OTf was prepared according to a literature procedure (Mock et al., 1999). Melting points were determined on a MEL-TEMP II capillary tube apparatus and were uncorrected. 1H NMR and 13C NMR spectra were recorded at 500 and 125 MHz, respectively, on Bruker Avance II 500 NMR spectrometers using tetramethylsilane (TMS) as an internal standard. Chemical shift data for the proton resonances were reported in parts per million (ppm, d scale) relative to internal standard TMS (d: 0.0), and coupling constants (J) were reported in hertz (Hz). Liquid chromatography-mass spectra (LC-MS) analysis was performed on an
Agilent system, consisting of an 1100 series HPLC connected to a diode array detector and a 1946D mass spectrometer configured for positive-ion/negative-ion electrospray ionization. The high resolution mass spectra (HRMS) were obtained using a Waters/ Micromass LCT Classic spectrometer. Chromatographic solvent proportions are indicated in a volume:volume ratio. Thin-layer chromatography (TLC) was run using Analtech silica gel GF uniplates (5 10 cm2). Plates were visualized under UV light. Normal phase flash column chromatography was carried out on EM Science silica gel 60 (230–400 mesh) with a forced flow of the indicated solvent system in the proportions described below. All moisture- and/or air-sensitive reactions were performed under a positive pressure of nitrogen maintained by a direct line from a nitrogen source. Analytical HPLC was performed using a Prodigy (Phenomenex) 5 mm C-18 column, 4.6 250 mm; 3:1:1 CH3CN:MeOH:20 mM phosphate buffer solution (pH¼6.7) mobile phase; 1.5 mL/min flow rate; UV (254 nm) and g-ray (PIN diode) flow detectors. C-18 Plus Sep-Pak cartridges (55–105 mm; part number WAT036575) were obtained from Waters Corporation, Milford, MA. Sterile Millex-GS 0.22 mm vented filter unit was obtained from Millipore Corporation, Bedford, MA. 3.2. General procedure for the preparation of compounds 3a–f A solution of compounds 1 (20 mmol), 1-bromo-4-chlorobutane or 1-bromo-6-chlorohexane (50 mmol), and potassium carbonate (4.15g, 30 mmol) in acetonitrile (120 mL) was refluxed for 1.5 h. The reaction mixture was cooled down to room temperature (RT), filtered to eliminate inorganic material, and washed with additional acetonitrile (30 mL). The organic solution was concentrated, and the residue was purified by column chromatography on silica gel with eluent (10% EtOAc/hexanes) to give pure compound 3 as a light-yellow oil in 65–82% yield. Rf ¼0.72–0.80 (1:3 EtOAc/hexanes). 2-((4-Chlorobutyl)thio)benzo[d]oxazole (3a). 1H NMR (CDCl3) d: 1.95–2.05 (m, 4H, CH2CH2), 3.34 (t, J ¼7.0 Hz, 2H, CH2), 3.59 (t, J¼6.5 Hz, 2H, CH2), 7.23 (dt, J ¼1.5, 7.5 Hz, 1H, Ph-H), 7.28 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 7.42 (dd, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.59 (dd, J ¼1.0, 7.5 Hz, 1H, Ph-H). MS (ESI): 242 ([MþH] þ , 100%). 2-((4-Chlorobutyl)thio)-5,7-dimethylbenzo[d]oxazole (3b). 1H NMR (CDCl3) d: 1.95–2.02 (m, 4H, CH2CH2), 2.39 (s, 3H, CH3), 2.43 (s, 3H, CH3), 3.33 (t, J ¼7.0 Hz, 2H, CH2), 3.58 (t, J¼6.0 Hz, 2H, CH2), 6.85 (s, 1H, Ph-H), 7.20 (s, 1H, Ph-H). MS (ESI): 270 ([MþH] þ , 100%). 2-((4-Chlorobutyl)thio)benzo[d]thiazole (3c). 1H NMR (CDCl3) d: 1.94–2.03 (m, 4H, CH2CH2), 3.38 (t, J ¼6.5 Hz, 2H, CH2), 3.58
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(t, J¼6.0 Hz, 2H, CH2), 7.28 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.41 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.73 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). MS (ESI): 258 ([MþH] þ , 100%). 2-((6-Chlorohexyl)thio)benzo[d]oxazole (3d). 1H NMR (CDCl3) d: 1.49–1.54 (m, 4H, 2CH2), 1.76–1.81 (m, 2H, CH2), 1.82–1.88 (m, 2H, CH2), 3.31 (t, J¼7.0 Hz, 2H, CH2), 3.53 (t, J¼6.5 Hz, 2H, CH2), 7.23 (dt, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.27 (dt, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.42 (d, J¼8.0 Hz, 1H, Ph-H), 7.58 (dd, J¼1.0, 8.0 Hz, 1H, Ph-H). MS (ESI): 270 ([MþH] þ , 100%). 2-((6-Chlorohexyl)thio)-5,7-dimethylbenzo[d]oxazole (3e). 1H NMR (CDCl3) d: 1.49–1.51 (m, 4H, 2CH2), 1.77–1.80 (m, 2H, CH2), 1.81–1.86 (m, 2H, CH2), 2.39 (s, 3H, CH3), 2.42 (s, 3H, CH3), 3.29 (t, J ¼7.0 Hz, 2H, CH2), 3.53 (t, J¼6.5 Hz, 2H, CH2), 6.84 (s, 1H, PhH), 7.20 (s, 1H, Ph-H). MS (ESI): 298 ([MþH] þ , 100%). 2-((6-Chlorohexyl)thio)benzo[d]thiazole (3f). 1H NMR (CDCl3) d: 1.48–1.53 (m, 4H, 2CH2), 1.76–1.80 (m, 2H, CH2), 1.81–1.86 (m, 2H, CH2), 3.34 (t, J¼7.5 Hz, 2H, CH2), 3.53 (t, J¼ 6.5 Hz, 2H, CH2), 7.28 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.40 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.73 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). MS (ESI): 286 ([MþH] þ , 100%).
3.3. General procedure for the preparation of compounds 5a-l A suspension of 2-(chloroalkylthio)benzoxazole or 2-(chloroalkylthio)benzothiazole (3, 2.0 mmol), 1-(2-substitutephenyl) piperazine (4, 2.5 mmol), potassium carbonate (0.346 g, 2.5 mmol), and potassium iodide (0.332 g, 2.0 mmol) in acetonitrile (60 mL) was refluxed for 30 h. The reaction mixture was concentrated, and the residue was diluted with water and extracted with CH2Cl2 (3 80 mL). The combined organic layers were dried over Na2SO4 and concentrated by evaporation in vacuo. The residue was purified by column chromatography on silica gel with eluent (1% MeOH/ CH2Cl2) to provide compound 5 in 22–61% yield. Rf ¼ 0.62–0.70 (10% MeOH/CH2Cl2). 2-((4-(4-(2-Methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d]oxazole (5a). Yellowish oil, 31% yield. 1H NMR (CDCl3) d: 1.72– 1.75 (m, 2H, CH2), 1.87–1.93 (m, 2H, CH2), 2.46 (t, J¼7.5 Hz, 2H, NCH2), 2.65 (br m, 4H, piperazine-H), 3.09 (br m, 4H, piperazineH), 3.35 (t, J ¼7.0 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (d, J¼7.5 Hz, 1H, Ph-H), 6.90–6.94 (m, 2H, Ph-H), 6.97–7.00 (m, 1H, Ph-H), 7.21–7.28 (m, 2H, Ph-H), 7.41 (d, J¼ 7.5 Hz, 1H, Ph-H), 7.58 (d, J¼7.5 Hz, 1H, Ph-H). MS (ESI): 398 ([MþH] þ , 100%). 2-(4-(4-(Benzo[d]oxazol-2-ylthio)butyl)piperazin-1-yl)phenol (5b). Yellow oil, 26% yield. 1H NMR (CDCl3) d: 1.70–1.76 (m, 2H, CH2), 1.83–1.94 (m, 2H, CH2), 2.48 (t, J¼7.5 Hz, 2H, NCH2), 2.62 (br m, 4H, piperazine-H), 2.83–2.91 (m, 4H, piperazine-H), 3.36 (t, J¼ 7.0 Hz, 2H, SCH2), 6.83 (dt, J¼1.5, 8.0 Hz, 1H, Ph-H), 6.93 (dd, J ¼1.0, 8.0 Hz, 1H, Ph-H), 7.06 (dt, J¼ 1.0, 7.5 Hz, 1H, Ph-H), 7.14 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.23 (dt, J¼1.0, 7.5 Hz, 1H, PhH), 7.28 (dt, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.42 (dd, J¼0.5, 7.5 Hz, 1H, Ph-H), 7.59 (dd, J¼ 0.5, 7.5 Hz, 1H, Ph-H). MS (ESI): 384 ([MþH] þ , 100%). HRMS (ESI): calcd for C21H26N3O2S 384.1746 ([MþH] þ ), found 384.1740. 2-((4-(4-(2-Methoxyphenyl)piperazin-1-yl)butyl)thio)-5.7-dimethylbenzo[d]oxazole (5c). Yellow oil, 30% yield. 1H NMR (CDCl3) d: 1.69–1.75 (m, 2H, CH2), 1.85–1.91 (m, 2H, CH2), 2.39 (s, 3H, CH3), 2.42 (s, 3H, CH3), 2.45 (t, J¼7.5 Hz, 2H, NCH2), 2.64 (br m, 4H, piperazine-H), 3.08 (br m, 4H, piperazine-H), 3.34 (t, J¼7.0 Hz, 2H, CH2), 3.85 (s, 3 H, OCH3), 6.84–6.85 (m, 2H, Ph-H), 6.89–6.94 (m 2H, Ph-H), 6.97–7.00 (m, 1H, Ph-H), 7.20 (s, 1H, Ph-H). 13C NMR (CDCl3) d: 15.14, 21.52, 25.99, 27.49, 32.34, 50.77, 53.57, 55.45, 58.10, 111.24, 115.85, 118.30, 119.76, 121.08, 122.98, 126.15, 133.15, 141.48, 141.82, 149.38, 152.38, 164.59. HRMS (ESI): calcd for C24H32N3O2S 426.2215 ([MþH] þ ), found 426.2197.
2-(4-(4-((5,7-Dimethylbenzo[d]oxazol-2-yl)thio)butyl)piperazin-1-yl)phenol (5d). Yellow oil, 24% yield. 1H NMR (CDCl3) d: 1.69–1.75 (m, 2H, CH2), 1.87–1.93 (m, 2H, CH2), 2.39 (s, 3H, CH3), 2.43 (s, 3H, CH3), 2.46 (t, J¼7.5 Hz, 2H, NCH2), 2.62 (br m, 4H, piperazine-H), 2.89–2.90 (m, 4H, piperazine-H), 3.34 (t, J ¼7.0 Hz, 2H, CH2), 6.83 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 6.85 (s, 1H, Ph-H), 6.93 (dd, J¼1.0, 8.0, 1H, Ph-H), 7.06 (dt, J¼ 1.5, 8.0 Hz, 1H, Ph-H), 7.13 (dd, J ¼1.5, 5.5 Hz, 1H, Ph-H), 7.20 (s, 1H, Ph-H). HRMS (ESI): calcd for C23H30N3O2S 412.2059 ([MþH] þ ), found 412.2045. 2-((4-(4-(2-Methoxyphenyl)piperazin-1-yl)butyl)thio)benzo [d]thiazole (5e). Yellowish oil, 27% yield. 1H NMR (CDCl3) d: 1.69– 1.75 (m, 2H, CH2), 1.85–1.91 (m, 2H, CH2), 2.45 (t, J¼ 7.5 Hz, 2H, NCH2), 2.64 (br m, 4H, piperazine-H), 3.08 (br m, 4H, piperazineH), 3.38 (t, J¼7.5 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (d, J¼7.5 Hz, 1H, Ph-H), 6.90–6.92 (m, 2H, Ph-H), 6.97–7.00 (m, 1H, Ph-H), 7.25–7.29 (m, 1H, Ph-H), 7.38 (dt, J¼1.0, 7.0 Hz, 1H, Ph-H), 7.73 (d, J ¼7.5 Hz, 1H, Ph-H), 7.85 (d, J ¼7.5 Hz, 1H, Ph-H). MS (ESI): 414 ([MþH] þ , 100%). 2-(4-(4-(Benzo[d]thiazol-2-ylthio)butyl)piperazin-1-yl)phenol (5f). Yellowish semisolid, 22% yield. 1H NMR (CDCl3) d: 1.70–1.74 (m, 2H, CH2), 1.86–1.92 (m, 2H, CH2), 2.46 (t, J¼7.5 Hz, 2H, NCH2), 2.60 (br m, 4H, piperazine-H), 2.88–2.89 (m, 4H, piperazine-H), 3.40 (t, J ¼7.0 Hz, 2H, SCH2), 6.84 (t, J¼7.5 Hz, 1H, Ph-H), 6.93 (d, J¼8.0 Hz, 1H, Ph-H), 7.04 (t, J¼8.0 Hz, 1H, Ph-H), 7.12 (d, J¼8.0 Hz, 1H, Ph-H), 7.28 (dd, J ¼7.5, 8.0 Hz, 1H, Ph-H), 7.40 (dd, J¼7.5, 8.0 Hz, 1H, Ph-H), 7.74 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). HRMS (ESI): calcd for C21H26N3OS2 400.1517 ([MþH] þ ), found 400.1525. 2-((6-(4-(2-Methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]oxazole (5g). White semisolid, 61% yield. 1H NMR (CDCl3) d: 1.38–1.43 (m, 2H, CH2), 1.50–1.56 (m, 2H, CH2), 1.58–1.64 (m, 2H, CH2), 1.82–1.88 (m, 2H, CH2), 2.47 (t, J¼7.5 Hz, 2H, NCH2), 2.72 (br m, 4H, piperazine-H), 3.15 (br m, 4H, piperazine-H), 3.31 (t, J¼7.5 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (dd, J¼1.0, 8.0 Hz, 1H, Ph-H), 6.89–6.95 (m, 2H, Ph-H), 6.98–7.01 (m, 1H, Ph-H), 7.22 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.25–7.27 (m, 1H, Ph-H), 7.42 (dd, J¼0.5, 7.5 Hz, 1H, Ph-H), 7.59 (d, J¼0.5, 7.5 Hz, 1H, Ph-H). MS (ESI): 426 ([MþH] þ , 100%). 2-(4-(6-(Benzo[d]oxazol-2-ylthio)hexyl)piperazin-1-yl)phenol (5h). White solid, mp 95–97 1C, 44% yield. 1H NMR (CDCl3) d: 1.38–1.43 (m, 2H, CH2), 1.50–1.58 (m, 4H, CH2CH2), 1.83–1.89 (m, 2H, CH2), 2.40 (t, J¼7.5 Hz, 2H, NCH2), 2.60 (br m, 4H, piperazineH), 2.89–2.91 (m, 4H, piperazine-H), 3.32 (t, J¼7.0 Hz, 2H, SCH2), 6.85 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 6.93 (dd, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.15 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.23–7.28 (m, 2H, Ph-H), 7.42 (dd, J¼0.5, 7.5 Hz, 1H, Ph-H), 7.59 (d, J¼ 0.5, 7.5 Hz, 1H, Ph-H). HRMS (ESI): calcd for C23H30N3O2S 412.2059 ([MþH] þ ), found 412.2070. 2-((6-(4-(2-Methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7dimethylbenzo[d]oxazole (5i). White solid, mp 68–70 1C, 55% yield. 1H NMR (CDCl3) d: 1.38–1.42 (m, 2H, CH2), 1.49–1.55 (m, 2H, CH2), 1.57–1.61 (m, 2H, CH2), 1.81–1.87 (m, 2H, CH2), 2.17 (s, 3H, CH3), 2.39 (s, 1H, CH3), 2.44 (t, J¼7.0 Hz, 2H, NCH2), 2.69 (br m, 4H, piperazine-H), 3.13 (br m, 4H, piperazine-H), 3.30 (t, J¼ 7.0 Hz, 2H, SCH2), 3.86 (s, 3H, OCH3), 6.84 (s, 1H, Ph-H), 6.86 (d, J ¼9.0 Hz, 1H, Ph-H), 6.90–6.95 (m, 2H, Ph-H), 6.99 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 7.20 (s, 1H, Ph-H). 13C NMR (CDCl3) d: 15.13, 21.50, 26.44, 27.08, 28.58, 29.27, 32.26, 50.33, 53.46, 55.45, 58.65, 111.25, 115.84, 118.40, 119.75, 121.11, 123.16, 126.13, 133.96, 141.16, 141.82, 149.36, 152.34, 164.66. HRMS (ESI): calcd for C26H36N3O2S 454.2528 ([Mþ H] þ ), found 454.2508. 2-(4-(6-((5,7-Dimethylbenzo[d]oxazol-2-yl)thio)hexyl)piperazin-1-yl)phenol (5j). White solid, mp 74–75 1C, 42% yield. 1H NMR (CDCl3) d: 1.36–1.42 (m, 2H, CH2), 1.49–1.57 (m, 4H, CH2CH2), 1.81–1.87 (m, 2H, CH2), 2.38–2.42 (m, 5H, CH3, and NCH2), 2.43 (s, 1H, CH3), 2.60 (br m, 4H, piperazine-H), 2.89–2.95 (m, 4H,
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
piperazine-H), 3.30 (t, J¼7.5 Hz, 2H, SCH2), 6.84 (s, 1H, Ph-H), 6.83–6.86 (m, 2H, Ph-H), 6.93 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.05 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 7.16 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.20 (s, 1H, Ph-H). HRMS (ESI): calcd for C25H24N3O2S 440.2372 ([MþH] þ ), found 440.2371. 2-((6-(4-(2-Methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo [d]thiazole (5k). Light-yellow semisolid, 52% yield. 1H NMR (CDCl3) d: 1.38–1.42 (m, 2H, CH2), 1.49–1.55 (m, 2H, CH2), 1.59–1.66 (m, 2H, CH2), 1.81–1.87 (m, 2H, CH2), 2.50 (t, J¼8.0 Hz, 2H, NCH2), 2.75 (br m, 4H, piperazine-H), 3.17 (br m, 4H, piperazine-H), 3.34 (t, J¼7.5 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (d, J¼8.0 Hz, 1H, Ph-H), 6.89–6.95 (m, 2H, Ph-H), 6.98–7.02 (m, 1H, Ph-H), 7.28 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.40 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.73 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). MS (ESI): 442 ([MþH] þ , 100%). 2-(4-(6-(Benzo[d]thiazol-2-ylthio)hexyl)piperazin-1-yl)phenol (5l). Yellowish solid, mp 77–79 1C, 41% yield. 1H NMR (CDCl3) d: 1.38–1.43 (m, 2H, CH2), 1.51–1.56 (m, 4H, CH2CH2), 1.83–1.86 (m, 2H, CH2), 2.40 (t, J¼ 7.5 Hz, 2H, NCH2), 2.60 (br m, 4H, piperazineH), 2.89–2.91 (m, 4H, piperazine-H), 3.36 (t, J¼7.5 Hz, 2H, SCH2), 6.85 (dt, J ¼1.0, 7.5 Hz, 1H, Ph-H), 6.93 (dd, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.05 (dt, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.15 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.28 (dt, J¼1.5, 8.0 Hz, 1H), 7.40 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.74 (d, J ¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). HRMS (ESI): calcd for C23H30N3OS2 428.1830 ([MþH] þ ), found 428.1839. 3.4. General procedure for the preparation of the target tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k [11C]CO2 was produced by the 14N(p,a)11C nuclear reaction in the small volume (9.5 cm3) aluminum gas target provided with the Siemens RDS-111 Eclipse cyclotron. The target gas consisted of 1% oxygen in nitrogen purchased as a specialty gas from Praxair, Indianapolis, IN. Typical irradiation used for the development was 50 mA beam current for 15 min on target. The production run produced approximately 25.9 GBq of [11C]CO2 at EOB. The phenolic hydroxyl precursor (0.1–0.3 mg) was dissolved in CH3CN (300 mL). To this solution was added 2N NaOH (2 mL). The mixture was transferred to a small reaction vial. No-carrier-added (high specific activity) [11C]CH3OTf (13.9 GBq), that was produced by the gas-phase production method (Mock et al., 1999) within 11 min from [11C]CO2 (25.9 GBq) through [11C]CH4 (21.8 GBq) and [11C]CH3Br (13.9 GBq) with silver triflate (AgOTf) column, was passed into the reaction vial at RT until radioactivity reached a maximum (2 min), and then the reaction vial was isolated and heated at 80 1C for 3 min. The contents of the reaction vial were diluted with NaHCO3 (0.1 M, 1 mL). The reaction vial was connected to a C-18 Plus Sep-Pak cartridge. The labeled product mixture solution was passed onto the cartridge for SPE purification by gas pressure. The cartridge was washed with H2O (2 3 mL), and the aqueous washing was discarded. The product was eluted from the cartridge with EtOH (2 2 mL), and then passed onto a rotatory evaporator. The solvent was removed by evaporation (3 min) under vacuum. The final volume of ethanol after evaporation was 1 mL. The labeled product was reformulated with saline (10 mL), sterile-filtered through a sterile vented Millex-GS 0.22 mm cellulose acetate membrane, and collected into a sterile vial. Total radioactivity (5.9–7.1 GBq) was assayed and the total volume (10–11 mL) was noted for tracer dose dispensing. The overall synthesis time including SPE purification and reformulation was 23 min. The radiochemical yields decay corrected to end of bombardment (EOB), from [11C]CO2, were 50–60% (n¼5). The same procedure was used to prepare the target tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k from their corresponding precursors 5b, 5d, 5f,
503
Table 3 Analytical HPLC retention times (tR) of arylpiperazinylthioalkyl derivatives. tR (min) Precursor 5b 5d 5f 5h 5j 5l
Standard 2.12 2.28 2.39 2.27 2.41 2.57
5a 5c 5e 5g 5i 5k
Tracer 5.32 5.45 5.66 5.34 5.76 5.92
[11C]5a [11C]5c [11C]5e [11C]5g [11C]5i [11C]5k
5.32 5.45 5.66 5.34 5.76 5.92
5h, 5j, and 5l. Retention times of arylpiperazinylthioalkyl derivatives including the precursors, reference standards, and target tracers in the analytical HPLC system are listed in Table 3.
4. Conclusion An efficient and convenient synthesis of new 5-HT1AR radioligands, carbon-11-labeled arylpiperazinylthioalkyl derivatives, has been developed. The synthetic methodology employed simple twostep condensation to prepare a series of new arylpiperazinylthioalkyl derivatives, phenolic hydroxyl precursors, and their corresponding ether standard compounds. The target tracers were easily prepared by O-[11C]methylation of their corresponding precursors using a reactive [11C]methylating agent, [11C]CH3OTf, and isolated by a simplified SPE purification procedure in high radiochemical yields, short overall synthesis time, and high specific radioactivities. These chemistry results combined with the reported in vitro biological data (Siracusa et al., 2008) encourage further in vivo biological evaluation of new carbon-11-labeled arylpiperazinylthioalkyl derivatives as candidate PET agents for imaging of 5-HT1AR.
Acknowledgments 1
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Savitz, J., Lucki, I., Drevets, W.C., 2009. 5-HT1A receptor function in major depressive disorder. Prog. Neurobiol. 88, 17–31. Siracusa, M.A., Salerno, L., Modica, M.N., Pittala, V., Romeo, G., Amato, M.E., Nowak, M., Bojarski, A.J., Mereghetti, I., Cagnotto, A., Mennini, T., 2008. Synthesis of new arylpiperazinylalkylthiobenzimidazole, benzothiazole, or benzoxazole derivatives as potent and selective 5-HT1A serotonin receptor ligands. J. Med. Chem. 51, 4529–4538. Turner, M.R., Rabiner, E.A., Hammers, A., Al-Chalabi, A., Grasby, P.M., Shaw, C.E., Brooks, D.J., Leigh, P.N., 2005. [11C]-WAY100635 PET demonstrates marked 5-HT1A receptor changes in sporadic ALS. Brain 128, 896–905. Wang, M., Gao, M., Miller, K.D., Zheng, Q.-H., 2011. Synthesis of [11C]PBR06 and [18F]PBR06 as agents for positron emission tomographic (PET) imaging of the translocator protein (TSPO). Steroids 76, 1331–1340. Yoshida, S., Shiokawa, S., Kawano, K.-i., Ito, T., Murakami, H., Suzuki, H., Sato, Y., 2005. Orally active benzoxazole derivatives as 5-HT3 receptor partial agonist for treatment of diarrhea-predominant irritable bowel syndrome. J. Med. Chem. 48, 7075–7079. Zheng, Q.-H., Mock, B.H., 2005. Purification of carbon-11 PET radiotracers from unlabeled precursors by preparative HPLC and SPE. Biomed. Chromatogr. 19, 671–676. Zheng, Q.-H., Gao, M., Mock, B.H., Wang, S., Hara, T., Nazih, R., Miller, M.A., Receveur, T.J., Lopshire, J.C., Groh, W.J., Zipes, Z.P., Hutchins, G.D., DeGrado, T.R., 2007. Synthesis and biodistribution of new radiolabeled high-affinity choline transporter inhibitors [11C]hemicholinium-3 and [18F]hemicholinium3. Bioorg. Med. Chem. Lett. 17, 2220–2224.
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Applied Radiation and Isotopes journal homepage: www.elsevier.com/locate/apradiso
Facile synthesis of carbon-11-labeled arylpiperazinylthioalkyl derivatives as new PET radioligands for imaging of 5-HT1AR Mingzhang Gao, Min Wang, Qi-Huang Zheng n Department of Radiology and Imaging Sciences, Indiana University School of Medicine, 1345 West 16th Street, L3-208, Indianapolis, IN 46202-2111, USA
a r t i c l e i n f o
a b s t r a c t
Article history: Received 12 August 2011 Received in revised form 26 October 2011 Accepted 14 November 2011 Available online 25 November 2011
Carbon-11-labeled arylpiperazinylthioalkyl derivatives, 2-((4-(4-(2-[11C]methoxyphenyl)piperazin1-yl)butyl)thio)benzo[d]oxazole ([11C]5a), 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)butyl)thio)5,7-dimethylbenzo[d]oxazole ([11C]5c), 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)butyl)thio) benzo[d]thiazole ([11C]5e), 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]oxazole 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7-dimethylbenzo[d]oxazole ([11C]5g), ([11C]5i), and 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]thiazole ([11C]5k), were prepared from their corresponding phenol precursors with [11C]CH3OTf through O-[11C]methylation and isolated by a simplified solid-phase extraction (SPE) method using a Sep-Pak Plus C18 cartridge in 50–60% (n ¼5) radiochemical yields based on [11C]CO2 and decay corrected to end of bombardment (EOB). The overall synthesis time from EOB was 23 min, the radiochemical purity was 499%, and the specific activity at end of synthesis (EOS) was 277.57 92.5 GBq/mmol (n ¼5). & 2011 Elsevier Ltd. All rights reserved.
Keywords: Carbon-11-labeled arylpiperazinylthioalkyl derivatives Positron emission tomography (PET) Serotonin-1A receptor (5-HT1AR) Radioligands Imaging
1. Introduction Serotonin (5-hydroxytryptamine, 5-HT) is an important neurotransmitter that is involved in various physiological and pathological processes in peripheral and central nervous system (Siracusa et al., 2008; Yoshida et al., 2005). To date at least 14 different serotonin receptors (5-HTRs) have been identified, and these receptors can be divided into distinct families—denoted by numbers 1, 2, 3, 4, 5, 6, and 7, with subtypes in each family denoted by letters like a, b, and c (Savitz et al., 2009). Among 5-HTRs, the 5-HT1AR was one of the first serotonin receptor subtypes pharmacologically characterized and is the best studied (Caliendo et al., 2005; Lacivita et al., 2008). 5-HT1AR is generally accepted to be implicated in the pathophysiology of major neuropsychiatric disorders, including depression, suicidal behavior, panic disorder, epilepsy, bulimia, schizophrenia, Parkinson’s disease, and Alzheimer’s disease (Akimova et al., 2009; Drevets et al., 2007; Kumar and Mann, 2007; la Fougere et al., 2009). Therefore, 5-HT1AR is an important target for drug therapy, and numerous new therapeutic agents 5-HT1AR ligands belonging to different chemical classes have been developed in recent years (Lacivita et al., 2008). 5-HT1AR ligands can be grouped into two classes: antagonistic and agonistic ligands. 5-HT1AR is overexpressed in the brain. Because of the higher concentration and
n
Corresponding author. Tel.: þ317 278 4671; fax: þ317 278 9711. E-mail address: [email protected] (Q.-H. Zheng).
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heterogeneous distribution of this receptor in living brain, 5-HT1AR is one of the most attractive targets for noninvasive in vivo imaging of the related brain diseases using biomedical imaging technique positron emission tomography (PET), and various PET radioligands employed for quantification of brain 5-HT1AR have been developed accordingly (Kumar and Mann, 2007). Likewise, 5-HT1AR radioligands can be grouped into two classes: antagonistic and agonistic radioligands. Most of the 5-HT1AR radioligands belong to two structural families: (1) compounds structurally similar to the 5-HT1A antagonist WAY100635 (N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclohexane carboxamide), and (2) derivatives of the 5-HT1A agonist 8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)tetralin). Among these radioligands, [carbonyl-11C]WAY100635 (KD ¼ 0.2 nM for 5-HT1AR) is the most commonly used 5-HT1AR radioligand for in vivo human studies (Brooks and Piccini, 2006; Hall et al., 1997; Kumar and Mann, 2007; Turner et al., 2005). Despite recognition of the potential value of 5-HT1AR radioligands, there is a requirement for continued in vivo investigation using superior agents to establish the clinical utility of this strategy, before these radiotracers are clinically approved (Moghbel et al., 2011). Long-chain arylpiperazine derivatives are one of the most important classes of 5-HT1AR ligands. Recently, a new series of arylpiperazinylthioalkyl derivatives has been found to be potent and selective 5-HT1AR ligands (Siracusa et al., 2008). These ligands possess 2-methoxyphenyl amenable to labeling with the positron emitting radionuclide carbon-11 as 5-HT1AR radioligands. Arylpiperazinylthioalkyl derivatives labeled with carbon-11 as radiotracers using
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
PET may enable noninvasive monitoring of the density of 5-HT1AR and its response to 5-HT1AR antagonist and agonist treatment. Here we report the design, synthesis, and radiosynthesis of novel 5-HT1AR radioligands carbon-11-labeled arylpiperazinylthioalkyl derivatives, 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d]oxazole ([11C]5a), 2-((4-(4-(2-[11C]methoxyphenyl)piperazin-1-yl) butyl)thio)-5,7-dimethylbenzo[d]oxazole ([11C]5c), 2-((4-(4-(2-[11C] methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d]thiazole ([11C]5e), 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]o xazole ([11C]5g), 2-((6-(4-(2-[11C]methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7-dimethylbenzo[d]oxazole ([11C]5i), and 2-((6-(4-(2-[11C] methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]thiazole ([11C]5k).
2. Results and discussion 2.1. Chemistry Six title compounds, 2-((4-(4-(2-methoxyphenyl)piperazin1-yl)butyl)thio)benzo[d]oxazole (5a), 2-((4-(4-(2-methoxyphenyl)piperazin-1-yl)butyl)thio)-5,7-dimethylbenzo[d]oxazole (5c), 2-((4-(4-(2-methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d] thiazole (5e) 2-((6-(4-(2-methoxyphenyl)piperazin-1-yl)hexyl) thio)benzo[d]oxazole (5g), 2-((6-(4-(2-methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7-dimethylbenzo[d]oxazole (5i), and 2((6-(4-(2-methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]thiazole (5k), served as the reference standards and selected for radiolabeling. Compounds 5a, 5e, 5g, and 5k have been reported previously (Siracusa et al., 2008). These arylpiperazinylthioalkyl derivatives are potent and selective 5-HT1A ligands with low nanomolar Ki values, and their reported binding data with 5-HT1A, a1-adrenergic, D1, D2, and 5-HT2A receptors and calculated selectivity are summarized in Tables 1 and 2 (Siracusa et al., 2008). In particular, representative compound 5a displayed very high 5-HT1AR affinity (Ki 0.094 nM) and selectivity over all other investigated receptors including a1-adrenergic (Ki ratio 830), D1 (Ki ratio 17021), D2 (Ki ratio 397), and 5-HT2A (Ki ratio 3170) receptors. Compounds 5c and 5i are new compounds. Previous study on the structure–activity relationship (SAR) of 2(4-methylpiperazinyl)benzoxazole derivatives, 5-HT3 ligands indicated that introduction of small lipophilic substituents such as CH3 at the 5- and 7-position of the benzoxazole ring greatly increased the affinity for the 5-HT3 receptor (Gao et al., 2008a; Sato et al., 1998; Yoshida et al., 2005). Thus, we hypothesize that 5,7-dimethylbenzoxazole derivatives 5c and 5i will exhibit higher affinity for the 5-HT1AR than known benzoxazole derivatives 5a, 5e, 5g, and 5k without bearing any lipophilic substituent in the benzoxazole ring, and new compounds 5c and 5i were designed and synthesized in this regard. Further SAR study of the substituent effect is currently under way to confirm this hypothesis. Arylpiperazinylalkylthio benzoxazole or benzothiazole derivative reference standards and precursors were obtained by two-step condensation as outlined in Scheme 1, according to the published
Table 1 5-HT1A and a1-adrenergic receptors binding data of arylpiperazinylthioalkyl derivative standards 5a, 5e, 5g, and 5k (Siracusa et al., 2008). Compound
5a 5e 5g 5k
Ki (nM)
Ki ratio
5-HT1A
a1
a1/5-HT1A
0.094 70.008 0.27 70.01 0.52 70.01 1.30 70.004
78.07 23.2 16.67 1.4 52.57 7.4 46.07 7.3
830 61 101 35
499
Table 2 D1, D2, and 5-HT2A receptors binding data of arylpiperazinylthioalkyl derivative standards 5a, 5e, and 5g (Siracusa et al., 2008). Compound Ki (nM) D1
5a 5e 5g
Ki ratio D2
5-HT2A
D1/5HT1A
16007 82 37.3 7 8.3 298 7 44 17021 20057 388 58.3 7 12.2 106 7 23 7426 922 7 85 46.6 7 7.0 207 7 19 1773
D1/5HT1A
5-HT1A/5HT1A
397 216 90
3170 393 398
procedures (Siracusa et al., 2008) with slight modifications. The simple old method was used for preparation of a series of new compounds. The commercially available starting materials benzo [d]oxazole-2-thiol (1a), 5,7-diemthyl benzo[d]oxazole-2-thiol (1b), or benzo[d]thiazole-2-thiol (1c) were reacted with appropriate 1-bromo-4-chlorobutane or 1-bromo-6-chlorohexane (2) in acetonitrile at reflux for 1.5 h in the presence of potassium carbonate to give compounds 3a–f with high chemical yields (65–82% purified yield). The intermediates 3a–f were then reacted with 1-(2-methoxyphenyl)piperazine in acetonitrile at reflux for 30 h in the presence of potassium carbonate and potassium iodide to obtain the desired final standard products 5a, 5c, 5e, 5g, 5i, and 5k. The precursor compounds 5b, 5d, 5f, 5h, 5j, and 5l were designed and prepared by the same procedure as their corresponding standard compounds, using just 2-(piperazin-1-yl)phenol instead of 1-(2-methoxyphenyl)piperazine. The chemical yields of compounds 5a–l varied from 22% to 61%. In comparison with the literature methods, the slight modifications included the change of stoichiometric ratio of the starting materials, and the change of reaction conditions and work-up procedures, such as the use of different base, the extension of the reflux reaction time, and the use of the column chromatography for further purification. These modifications improved the chemical yield and chemical purity of the intermediates, standards, and precursors we prepared. 2.2. Radiochemistry Synthesis of carbon-11-labeled arylpiperazinylalkylthio benzoxazole or benzothiazole derivatives, [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k, is indicated in Scheme 2. Phenolic hydroxyl precursors 5b, 5d, 5f, 5h, 5j, and 5l were labeled by a reactive [11C]methylating agent, [11C]methyl triflate ([11C] CH3OTf) (Jewett, 1992; Mock et al., 1999) prepared from [11C]CO2, under basic conditions (2N NaOH) in acetonitrile through the O-[11C11C]methylation and isolated by a simplified solid-phase extraction (SPE) method (Gao et al., 2008b, 2009) to provide target tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k in 50–60% (n¼5) radiochemical yields, decay corrected to end of bombardment (EOB), based on [11C]CO2. The large polarity difference between the phenolic hydroxyl precursor and the corresponding labeled O-methylated ether product permitted the use of SPE technique for purification of the labeled product from the radiolabeling reaction mixture. A C-18 Plus Sep-Pak cartridge was used in SPE purification technique. The crude reaction mixture was treated with aqueous NaHCO3 and loaded onto the cartridge by gas pressure. The pH of freshly prepared 0.1 M NaHCO3 might be too low to effectively deprotonate a phenol. However, an excess of 2N NaOH used in the reaction provided a final pH after addition of 0.1 M NaHCO3 high enough to deprotonate all phenol. Any non-reacted phenolic hydroxyl precursor was actually converted to the corresponding sodium salt, and any non-reacted [11C]CH3OTf was actually hydrolyzed to [11C]CH3OH, which would not be trapped to the C-18 Sep-Pak. The cartridge was washed with water to remove non-reacted [11C]CH3OTf, remaining phenolic hydroxyl precursor,
500
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
R1
N
Cl
SH
X
n
Br 2
R1
X
i
R1
N X
Cl
HN
n
S
R2
N
n
S
O
ii
3a, n = 2, R1 = H, X = O 3b, n = 2, R1 = Me, X = O 3c, n = 2, R1 = H, X = S 3d, n = 4, R1 = H, X = O 3e, n = 4, R1 = Me, X = O 3f, n = 4, R1 = H, X = S
N
N 4
R1
1a, R1 = H, X = O 1b, R1 = Me, X = O 1c, R1 = H, X = S
R1
N
O
R2 R1 5a, n = 2, R1 = H, X = O, R2 = Me 5b, n = 2, R1 = H, X = O, R2 = H 5c, n = 2, R1 = Me, X = O, R2 = Me 5d, n = 2, R1 = Me, X = O, R2 = H 5e, n = 2, R1 = H, X = S, R2 = Me 5f, n = 2, R1 = H, X = S, R2 = H 5g, n = 4, R1 = H, X = O, R2 = Me 5h, n = 4, R1 = H, X = O, R2 = H 5i, n = 4, R1 = Me, X = O, R2 = Me 5j, n = 4, R1 = Me, X = O, R2 = H 5k, n = 4, R1 = H, X = S, R2 = Me 5l, n = 4, R1 = H, X = S, R2 = H Scheme 1. Synthesis of arylpiperazinylthioalkyl derivatives 5a–l. Reagents, conditions, and yields: (i) K2CO3/CH3CN, reflux; 65–82% yield. (ii) KI, K2CO3/CH3CN, reflux; 22–61% yield.
R1
N X
N S
n
N
i
R1
N
HO
R1 5b, n = 2, R1 = H, X = O 5d, n = 2, R1 = Me, X = O 5f, n = 2, R1 = H, X = S 5h, n = 4, R1 = H, X = O 5j, n = 4, R1 = Me, X = O 5l, n = 4, R1 = H, X = S
X
N S
n
N H311CO
1
R
[11C]5a, n = 2, R1 = H, X = O [11C]5c, n = 2, R1 = Me, X = O [11C]5e, n = 2, R1 = H, X = S [11C]5g, n = 4, R1 = H, X = O [11C]5i, n = 4, R1 = Me, X = O [11C]5k, n = 4, R1 = H, X = S
Scheme 2. Synthesis of carbon-11-labeled arylpiperazinylthioalkyl derivatives [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k. Reagents, conditions, and yields: (i) [11C]CH3OTf, CH3CN, 2N NaOH, 80 oC; 50–60% decay corrected radiochemical yield.
and reaction solvent, and total 6 mL (2 3 mL) volume of water was enough to wash off all phenol. The final labeled product was eluted with ethanol (2 2 mL), concentrated by rotary evaporation, and reformulated in saline (10 mL). In our fully automated radiosynthesis module (Mock et al., 2005b), it is difficult to directly elute the labeled product from a C-18 Sep-Pak to a vial using either 1 1 mL ethanol or 2 0.5 mL ethanol, due to the back pressure in the C-18 Sep-Pak and dead volume in the transfer tubing. In order to elute most of the labeled product from the C-18 Sep-Pak, we need to increase the volume of the eluent ethanol. For the radiotracers produced for animal study, we used 2 2 mL ethanol for elution, and evaporation was required before reformulation. For the radiotracer produced for human study, we used 2 1 mL ethanol, no evaporation was required, and a C-18 Sep-Pak was used for direct reformulation (Gao et al., 2010; Wang et al., 2011). Overall synthesis time was 23 min from EOB, including approximately 11 min for [11C]CH3OTf production, 5 min for O-[11C]methylation reaction, and 7 min for SPE purification, evaporation, and reformulation.
SPE technique is fast, efficient, and convenient and works very well for the O-methylated ether tracer purification using the phenolic hydroxyl precursor for radiolabeling (Gao et al., 2008b). The radiosynthesis was performed in an automated selfdesigned multi-purpose 11C-radiosynthesis module, allowing measurement of specific activity at EOB during synthesis (Mock et al., 2005a,b). Online determination of specific activity at EOB is accurate when high performance liquid chromatography (HPLC) is used as purification method. However, the on-the-fly technique to determine specific activity at EOB is not applied when SPE is used as the purification method. In general, the specific activity for all 11C-tracers produced in our PET radiochemistry facility is basically same, because the specific activity of a carrier-free radionuclide sample (11C) is only related to its half-life (20.4 min). In particular, the specific activity for all 11C-tracers produced in our facility is dependent on two parts: (1) carrier from the 11C gas target, and (2) carrier from the 11C radiolabeled precursor [11C]CH3OTf production system. The 11C gas target we used is the Siemens RDS-111 Eclipse cyclotron 11C gas target.
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
501
mV
600.00 400.00 200.00 0.00 1.00
2.00
3.00
4.00
5.00 6.00 Minutes
7.00
8.00
9.00
10.00
1.00
2.00
3.00
4.00
5.00 6.00 Minutes
7.00
8.00
9.00
10.00
AU
0.006 0.004 0.002 0.000
Fig. 1. A representative analytical HPLC chromatographic profile for the tracer [11C]5a produced by SPE purification: analytical radioactive (A) and UV (B) traces, tR [11C]5a ¼ 5.32 and 5.22 min, respectively.
The [11C]CH3OTf production system we used is an Eckert & Ziegler Modular Lab C-11 Methyl Iodide/Triflate module (Mock et al., 1999). The specific activity for our 11C-tracers usually ranges from 370 to 925 GBq/mmol at EOB according to our previous works. Actually the specific activity of all our 11C-tracers is essentially the same, because in the same radiochemistry facility, the same targetry conditions, C-11 Methyl Iodide/Triflate module, and 11Cradiosynthesis module are employed. The specific activity of carbon-11-labeled arylpiperazinylthioalkyl derivatives was estimated in the range of 185–370 GBq/mmol at the end of synthesis (EOS) based on other compounds produced in our facility using the same targetry conditions, which have been measured by the on-the-fly technique or the same SPE purification method (Mock et al., 2005b; Zheng et al., 2007). The actual measurement of specific activity at EOS was performed by analytical HPLC (Zheng and Mock, 2005) and calculated. The exact values of the specific activity for the tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k were 277.5 792.5 GBq/mmol (n¼5), which are in agreement with the estimated values and the ‘‘online’’ determined values. Chemical purity and radiochemical purity were determined by analytical HPLC (Zheng and Mock, 2005). The chemical purity of the precursors and reference standards was 496%. The radiochemical purity of the target tracers was 499% determined by radio-HPLC through g-ray (PIN diode) flow detector, and the chemical purity of the target tracers was 495% determined by reversed-phase HPLC through UV flow detector. Representative analytical HPLC chromatograms for the target tracer [11C]5a produced by SPE purification are shown in Fig. 1.
3. Experimental 3.1. General All commercial reagents and solvents were purchased from Sigma-Aldrich and Fisher Scientific and used without further purification. [11C]CH3OTf was prepared according to a literature procedure (Mock et al., 1999). Melting points were determined on a MEL-TEMP II capillary tube apparatus and were uncorrected. 1H NMR and 13C NMR spectra were recorded at 500 and 125 MHz, respectively, on Bruker Avance II 500 NMR spectrometers using tetramethylsilane (TMS) as an internal standard. Chemical shift data for the proton resonances were reported in parts per million (ppm, d scale) relative to internal standard TMS (d: 0.0), and coupling constants (J) were reported in hertz (Hz). Liquid chromatography-mass spectra (LC-MS) analysis was performed on an
Agilent system, consisting of an 1100 series HPLC connected to a diode array detector and a 1946D mass spectrometer configured for positive-ion/negative-ion electrospray ionization. The high resolution mass spectra (HRMS) were obtained using a Waters/ Micromass LCT Classic spectrometer. Chromatographic solvent proportions are indicated in a volume:volume ratio. Thin-layer chromatography (TLC) was run using Analtech silica gel GF uniplates (5 10 cm2). Plates were visualized under UV light. Normal phase flash column chromatography was carried out on EM Science silica gel 60 (230–400 mesh) with a forced flow of the indicated solvent system in the proportions described below. All moisture- and/or air-sensitive reactions were performed under a positive pressure of nitrogen maintained by a direct line from a nitrogen source. Analytical HPLC was performed using a Prodigy (Phenomenex) 5 mm C-18 column, 4.6 250 mm; 3:1:1 CH3CN:MeOH:20 mM phosphate buffer solution (pH¼6.7) mobile phase; 1.5 mL/min flow rate; UV (254 nm) and g-ray (PIN diode) flow detectors. C-18 Plus Sep-Pak cartridges (55–105 mm; part number WAT036575) were obtained from Waters Corporation, Milford, MA. Sterile Millex-GS 0.22 mm vented filter unit was obtained from Millipore Corporation, Bedford, MA. 3.2. General procedure for the preparation of compounds 3a–f A solution of compounds 1 (20 mmol), 1-bromo-4-chlorobutane or 1-bromo-6-chlorohexane (50 mmol), and potassium carbonate (4.15g, 30 mmol) in acetonitrile (120 mL) was refluxed for 1.5 h. The reaction mixture was cooled down to room temperature (RT), filtered to eliminate inorganic material, and washed with additional acetonitrile (30 mL). The organic solution was concentrated, and the residue was purified by column chromatography on silica gel with eluent (10% EtOAc/hexanes) to give pure compound 3 as a light-yellow oil in 65–82% yield. Rf ¼0.72–0.80 (1:3 EtOAc/hexanes). 2-((4-Chlorobutyl)thio)benzo[d]oxazole (3a). 1H NMR (CDCl3) d: 1.95–2.05 (m, 4H, CH2CH2), 3.34 (t, J ¼7.0 Hz, 2H, CH2), 3.59 (t, J¼6.5 Hz, 2H, CH2), 7.23 (dt, J ¼1.5, 7.5 Hz, 1H, Ph-H), 7.28 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 7.42 (dd, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.59 (dd, J ¼1.0, 7.5 Hz, 1H, Ph-H). MS (ESI): 242 ([MþH] þ , 100%). 2-((4-Chlorobutyl)thio)-5,7-dimethylbenzo[d]oxazole (3b). 1H NMR (CDCl3) d: 1.95–2.02 (m, 4H, CH2CH2), 2.39 (s, 3H, CH3), 2.43 (s, 3H, CH3), 3.33 (t, J ¼7.0 Hz, 2H, CH2), 3.58 (t, J¼6.0 Hz, 2H, CH2), 6.85 (s, 1H, Ph-H), 7.20 (s, 1H, Ph-H). MS (ESI): 270 ([MþH] þ , 100%). 2-((4-Chlorobutyl)thio)benzo[d]thiazole (3c). 1H NMR (CDCl3) d: 1.94–2.03 (m, 4H, CH2CH2), 3.38 (t, J ¼6.5 Hz, 2H, CH2), 3.58
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(t, J¼6.0 Hz, 2H, CH2), 7.28 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.41 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.73 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). MS (ESI): 258 ([MþH] þ , 100%). 2-((6-Chlorohexyl)thio)benzo[d]oxazole (3d). 1H NMR (CDCl3) d: 1.49–1.54 (m, 4H, 2CH2), 1.76–1.81 (m, 2H, CH2), 1.82–1.88 (m, 2H, CH2), 3.31 (t, J¼7.0 Hz, 2H, CH2), 3.53 (t, J¼6.5 Hz, 2H, CH2), 7.23 (dt, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.27 (dt, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.42 (d, J¼8.0 Hz, 1H, Ph-H), 7.58 (dd, J¼1.0, 8.0 Hz, 1H, Ph-H). MS (ESI): 270 ([MþH] þ , 100%). 2-((6-Chlorohexyl)thio)-5,7-dimethylbenzo[d]oxazole (3e). 1H NMR (CDCl3) d: 1.49–1.51 (m, 4H, 2CH2), 1.77–1.80 (m, 2H, CH2), 1.81–1.86 (m, 2H, CH2), 2.39 (s, 3H, CH3), 2.42 (s, 3H, CH3), 3.29 (t, J ¼7.0 Hz, 2H, CH2), 3.53 (t, J¼6.5 Hz, 2H, CH2), 6.84 (s, 1H, PhH), 7.20 (s, 1H, Ph-H). MS (ESI): 298 ([MþH] þ , 100%). 2-((6-Chlorohexyl)thio)benzo[d]thiazole (3f). 1H NMR (CDCl3) d: 1.48–1.53 (m, 4H, 2CH2), 1.76–1.80 (m, 2H, CH2), 1.81–1.86 (m, 2H, CH2), 3.34 (t, J¼7.5 Hz, 2H, CH2), 3.53 (t, J¼ 6.5 Hz, 2H, CH2), 7.28 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.40 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.73 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). MS (ESI): 286 ([MþH] þ , 100%).
3.3. General procedure for the preparation of compounds 5a-l A suspension of 2-(chloroalkylthio)benzoxazole or 2-(chloroalkylthio)benzothiazole (3, 2.0 mmol), 1-(2-substitutephenyl) piperazine (4, 2.5 mmol), potassium carbonate (0.346 g, 2.5 mmol), and potassium iodide (0.332 g, 2.0 mmol) in acetonitrile (60 mL) was refluxed for 30 h. The reaction mixture was concentrated, and the residue was diluted with water and extracted with CH2Cl2 (3 80 mL). The combined organic layers were dried over Na2SO4 and concentrated by evaporation in vacuo. The residue was purified by column chromatography on silica gel with eluent (1% MeOH/ CH2Cl2) to provide compound 5 in 22–61% yield. Rf ¼ 0.62–0.70 (10% MeOH/CH2Cl2). 2-((4-(4-(2-Methoxyphenyl)piperazin-1-yl)butyl)thio)benzo[d]oxazole (5a). Yellowish oil, 31% yield. 1H NMR (CDCl3) d: 1.72– 1.75 (m, 2H, CH2), 1.87–1.93 (m, 2H, CH2), 2.46 (t, J¼7.5 Hz, 2H, NCH2), 2.65 (br m, 4H, piperazine-H), 3.09 (br m, 4H, piperazineH), 3.35 (t, J ¼7.0 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (d, J¼7.5 Hz, 1H, Ph-H), 6.90–6.94 (m, 2H, Ph-H), 6.97–7.00 (m, 1H, Ph-H), 7.21–7.28 (m, 2H, Ph-H), 7.41 (d, J¼ 7.5 Hz, 1H, Ph-H), 7.58 (d, J¼7.5 Hz, 1H, Ph-H). MS (ESI): 398 ([MþH] þ , 100%). 2-(4-(4-(Benzo[d]oxazol-2-ylthio)butyl)piperazin-1-yl)phenol (5b). Yellow oil, 26% yield. 1H NMR (CDCl3) d: 1.70–1.76 (m, 2H, CH2), 1.83–1.94 (m, 2H, CH2), 2.48 (t, J¼7.5 Hz, 2H, NCH2), 2.62 (br m, 4H, piperazine-H), 2.83–2.91 (m, 4H, piperazine-H), 3.36 (t, J¼ 7.0 Hz, 2H, SCH2), 6.83 (dt, J¼1.5, 8.0 Hz, 1H, Ph-H), 6.93 (dd, J ¼1.0, 8.0 Hz, 1H, Ph-H), 7.06 (dt, J¼ 1.0, 7.5 Hz, 1H, Ph-H), 7.14 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.23 (dt, J¼1.0, 7.5 Hz, 1H, PhH), 7.28 (dt, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.42 (dd, J¼0.5, 7.5 Hz, 1H, Ph-H), 7.59 (dd, J¼ 0.5, 7.5 Hz, 1H, Ph-H). MS (ESI): 384 ([MþH] þ , 100%). HRMS (ESI): calcd for C21H26N3O2S 384.1746 ([MþH] þ ), found 384.1740. 2-((4-(4-(2-Methoxyphenyl)piperazin-1-yl)butyl)thio)-5.7-dimethylbenzo[d]oxazole (5c). Yellow oil, 30% yield. 1H NMR (CDCl3) d: 1.69–1.75 (m, 2H, CH2), 1.85–1.91 (m, 2H, CH2), 2.39 (s, 3H, CH3), 2.42 (s, 3H, CH3), 2.45 (t, J¼7.5 Hz, 2H, NCH2), 2.64 (br m, 4H, piperazine-H), 3.08 (br m, 4H, piperazine-H), 3.34 (t, J¼7.0 Hz, 2H, CH2), 3.85 (s, 3 H, OCH3), 6.84–6.85 (m, 2H, Ph-H), 6.89–6.94 (m 2H, Ph-H), 6.97–7.00 (m, 1H, Ph-H), 7.20 (s, 1H, Ph-H). 13C NMR (CDCl3) d: 15.14, 21.52, 25.99, 27.49, 32.34, 50.77, 53.57, 55.45, 58.10, 111.24, 115.85, 118.30, 119.76, 121.08, 122.98, 126.15, 133.15, 141.48, 141.82, 149.38, 152.38, 164.59. HRMS (ESI): calcd for C24H32N3O2S 426.2215 ([MþH] þ ), found 426.2197.
2-(4-(4-((5,7-Dimethylbenzo[d]oxazol-2-yl)thio)butyl)piperazin-1-yl)phenol (5d). Yellow oil, 24% yield. 1H NMR (CDCl3) d: 1.69–1.75 (m, 2H, CH2), 1.87–1.93 (m, 2H, CH2), 2.39 (s, 3H, CH3), 2.43 (s, 3H, CH3), 2.46 (t, J¼7.5 Hz, 2H, NCH2), 2.62 (br m, 4H, piperazine-H), 2.89–2.90 (m, 4H, piperazine-H), 3.34 (t, J ¼7.0 Hz, 2H, CH2), 6.83 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 6.85 (s, 1H, Ph-H), 6.93 (dd, J¼1.0, 8.0, 1H, Ph-H), 7.06 (dt, J¼ 1.5, 8.0 Hz, 1H, Ph-H), 7.13 (dd, J ¼1.5, 5.5 Hz, 1H, Ph-H), 7.20 (s, 1H, Ph-H). HRMS (ESI): calcd for C23H30N3O2S 412.2059 ([MþH] þ ), found 412.2045. 2-((4-(4-(2-Methoxyphenyl)piperazin-1-yl)butyl)thio)benzo [d]thiazole (5e). Yellowish oil, 27% yield. 1H NMR (CDCl3) d: 1.69– 1.75 (m, 2H, CH2), 1.85–1.91 (m, 2H, CH2), 2.45 (t, J¼ 7.5 Hz, 2H, NCH2), 2.64 (br m, 4H, piperazine-H), 3.08 (br m, 4H, piperazineH), 3.38 (t, J¼7.5 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (d, J¼7.5 Hz, 1H, Ph-H), 6.90–6.92 (m, 2H, Ph-H), 6.97–7.00 (m, 1H, Ph-H), 7.25–7.29 (m, 1H, Ph-H), 7.38 (dt, J¼1.0, 7.0 Hz, 1H, Ph-H), 7.73 (d, J ¼7.5 Hz, 1H, Ph-H), 7.85 (d, J ¼7.5 Hz, 1H, Ph-H). MS (ESI): 414 ([MþH] þ , 100%). 2-(4-(4-(Benzo[d]thiazol-2-ylthio)butyl)piperazin-1-yl)phenol (5f). Yellowish semisolid, 22% yield. 1H NMR (CDCl3) d: 1.70–1.74 (m, 2H, CH2), 1.86–1.92 (m, 2H, CH2), 2.46 (t, J¼7.5 Hz, 2H, NCH2), 2.60 (br m, 4H, piperazine-H), 2.88–2.89 (m, 4H, piperazine-H), 3.40 (t, J ¼7.0 Hz, 2H, SCH2), 6.84 (t, J¼7.5 Hz, 1H, Ph-H), 6.93 (d, J¼8.0 Hz, 1H, Ph-H), 7.04 (t, J¼8.0 Hz, 1H, Ph-H), 7.12 (d, J¼8.0 Hz, 1H, Ph-H), 7.28 (dd, J ¼7.5, 8.0 Hz, 1H, Ph-H), 7.40 (dd, J¼7.5, 8.0 Hz, 1H, Ph-H), 7.74 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). HRMS (ESI): calcd for C21H26N3OS2 400.1517 ([MþH] þ ), found 400.1525. 2-((6-(4-(2-Methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo[d]oxazole (5g). White semisolid, 61% yield. 1H NMR (CDCl3) d: 1.38–1.43 (m, 2H, CH2), 1.50–1.56 (m, 2H, CH2), 1.58–1.64 (m, 2H, CH2), 1.82–1.88 (m, 2H, CH2), 2.47 (t, J¼7.5 Hz, 2H, NCH2), 2.72 (br m, 4H, piperazine-H), 3.15 (br m, 4H, piperazine-H), 3.31 (t, J¼7.5 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (dd, J¼1.0, 8.0 Hz, 1H, Ph-H), 6.89–6.95 (m, 2H, Ph-H), 6.98–7.01 (m, 1H, Ph-H), 7.22 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.25–7.27 (m, 1H, Ph-H), 7.42 (dd, J¼0.5, 7.5 Hz, 1H, Ph-H), 7.59 (d, J¼0.5, 7.5 Hz, 1H, Ph-H). MS (ESI): 426 ([MþH] þ , 100%). 2-(4-(6-(Benzo[d]oxazol-2-ylthio)hexyl)piperazin-1-yl)phenol (5h). White solid, mp 95–97 1C, 44% yield. 1H NMR (CDCl3) d: 1.38–1.43 (m, 2H, CH2), 1.50–1.58 (m, 4H, CH2CH2), 1.83–1.89 (m, 2H, CH2), 2.40 (t, J¼7.5 Hz, 2H, NCH2), 2.60 (br m, 4H, piperazineH), 2.89–2.91 (m, 4H, piperazine-H), 3.32 (t, J¼7.0 Hz, 2H, SCH2), 6.85 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 6.93 (dd, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.15 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.23–7.28 (m, 2H, Ph-H), 7.42 (dd, J¼0.5, 7.5 Hz, 1H, Ph-H), 7.59 (d, J¼ 0.5, 7.5 Hz, 1H, Ph-H). HRMS (ESI): calcd for C23H30N3O2S 412.2059 ([MþH] þ ), found 412.2070. 2-((6-(4-(2-Methoxyphenyl)piperazin-1-yl)hexyl)thio)-5,7dimethylbenzo[d]oxazole (5i). White solid, mp 68–70 1C, 55% yield. 1H NMR (CDCl3) d: 1.38–1.42 (m, 2H, CH2), 1.49–1.55 (m, 2H, CH2), 1.57–1.61 (m, 2H, CH2), 1.81–1.87 (m, 2H, CH2), 2.17 (s, 3H, CH3), 2.39 (s, 1H, CH3), 2.44 (t, J¼7.0 Hz, 2H, NCH2), 2.69 (br m, 4H, piperazine-H), 3.13 (br m, 4H, piperazine-H), 3.30 (t, J¼ 7.0 Hz, 2H, SCH2), 3.86 (s, 3H, OCH3), 6.84 (s, 1H, Ph-H), 6.86 (d, J ¼9.0 Hz, 1H, Ph-H), 6.90–6.95 (m, 2H, Ph-H), 6.99 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 7.20 (s, 1H, Ph-H). 13C NMR (CDCl3) d: 15.13, 21.50, 26.44, 27.08, 28.58, 29.27, 32.26, 50.33, 53.46, 55.45, 58.65, 111.25, 115.84, 118.40, 119.75, 121.11, 123.16, 126.13, 133.96, 141.16, 141.82, 149.36, 152.34, 164.66. HRMS (ESI): calcd for C26H36N3O2S 454.2528 ([Mþ H] þ ), found 454.2508. 2-(4-(6-((5,7-Dimethylbenzo[d]oxazol-2-yl)thio)hexyl)piperazin-1-yl)phenol (5j). White solid, mp 74–75 1C, 42% yield. 1H NMR (CDCl3) d: 1.36–1.42 (m, 2H, CH2), 1.49–1.57 (m, 4H, CH2CH2), 1.81–1.87 (m, 2H, CH2), 2.38–2.42 (m, 5H, CH3, and NCH2), 2.43 (s, 1H, CH3), 2.60 (br m, 4H, piperazine-H), 2.89–2.95 (m, 4H,
M. Gao et al. / Applied Radiation and Isotopes 70 (2012) 498–504
piperazine-H), 3.30 (t, J¼7.5 Hz, 2H, SCH2), 6.84 (s, 1H, Ph-H), 6.83–6.86 (m, 2H, Ph-H), 6.93 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.05 (dt, J¼1.5, 7.5 Hz, 1H, Ph-H), 7.16 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.20 (s, 1H, Ph-H). HRMS (ESI): calcd for C25H24N3O2S 440.2372 ([MþH] þ ), found 440.2371. 2-((6-(4-(2-Methoxyphenyl)piperazin-1-yl)hexyl)thio)benzo [d]thiazole (5k). Light-yellow semisolid, 52% yield. 1H NMR (CDCl3) d: 1.38–1.42 (m, 2H, CH2), 1.49–1.55 (m, 2H, CH2), 1.59–1.66 (m, 2H, CH2), 1.81–1.87 (m, 2H, CH2), 2.50 (t, J¼8.0 Hz, 2H, NCH2), 2.75 (br m, 4H, piperazine-H), 3.17 (br m, 4H, piperazine-H), 3.34 (t, J¼7.5 Hz, 2H, SCH2), 3.85 (s, 3H, OCH3), 6.84 (d, J¼8.0 Hz, 1H, Ph-H), 6.89–6.95 (m, 2H, Ph-H), 6.98–7.02 (m, 1H, Ph-H), 7.28 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.40 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.73 (d, J¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). MS (ESI): 442 ([MþH] þ , 100%). 2-(4-(6-(Benzo[d]thiazol-2-ylthio)hexyl)piperazin-1-yl)phenol (5l). Yellowish solid, mp 77–79 1C, 41% yield. 1H NMR (CDCl3) d: 1.38–1.43 (m, 2H, CH2), 1.51–1.56 (m, 4H, CH2CH2), 1.83–1.86 (m, 2H, CH2), 2.40 (t, J¼ 7.5 Hz, 2H, NCH2), 2.60 (br m, 4H, piperazineH), 2.89–2.91 (m, 4H, piperazine-H), 3.36 (t, J¼7.5 Hz, 2H, SCH2), 6.85 (dt, J ¼1.0, 7.5 Hz, 1H, Ph-H), 6.93 (dd, J¼1.0, 7.5 Hz, 1H, Ph-H), 7.05 (dt, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.15 (dd, J¼1.5, 8.0 Hz, 1H, Ph-H), 7.28 (dt, J¼1.5, 8.0 Hz, 1H), 7.40 (dt, J¼1.0, 8.0 Hz, 1H, Ph-H), 7.74 (d, J ¼8.0 Hz, 1H, Ph-H), 7.85 (d, J¼8.0 Hz, 1H, Ph-H). HRMS (ESI): calcd for C23H30N3OS2 428.1830 ([MþH] þ ), found 428.1839. 3.4. General procedure for the preparation of the target tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k [11C]CO2 was produced by the 14N(p,a)11C nuclear reaction in the small volume (9.5 cm3) aluminum gas target provided with the Siemens RDS-111 Eclipse cyclotron. The target gas consisted of 1% oxygen in nitrogen purchased as a specialty gas from Praxair, Indianapolis, IN. Typical irradiation used for the development was 50 mA beam current for 15 min on target. The production run produced approximately 25.9 GBq of [11C]CO2 at EOB. The phenolic hydroxyl precursor (0.1–0.3 mg) was dissolved in CH3CN (300 mL). To this solution was added 2N NaOH (2 mL). The mixture was transferred to a small reaction vial. No-carrier-added (high specific activity) [11C]CH3OTf (13.9 GBq), that was produced by the gas-phase production method (Mock et al., 1999) within 11 min from [11C]CO2 (25.9 GBq) through [11C]CH4 (21.8 GBq) and [11C]CH3Br (13.9 GBq) with silver triflate (AgOTf) column, was passed into the reaction vial at RT until radioactivity reached a maximum (2 min), and then the reaction vial was isolated and heated at 80 1C for 3 min. The contents of the reaction vial were diluted with NaHCO3 (0.1 M, 1 mL). The reaction vial was connected to a C-18 Plus Sep-Pak cartridge. The labeled product mixture solution was passed onto the cartridge for SPE purification by gas pressure. The cartridge was washed with H2O (2 3 mL), and the aqueous washing was discarded. The product was eluted from the cartridge with EtOH (2 2 mL), and then passed onto a rotatory evaporator. The solvent was removed by evaporation (3 min) under vacuum. The final volume of ethanol after evaporation was 1 mL. The labeled product was reformulated with saline (10 mL), sterile-filtered through a sterile vented Millex-GS 0.22 mm cellulose acetate membrane, and collected into a sterile vial. Total radioactivity (5.9–7.1 GBq) was assayed and the total volume (10–11 mL) was noted for tracer dose dispensing. The overall synthesis time including SPE purification and reformulation was 23 min. The radiochemical yields decay corrected to end of bombardment (EOB), from [11C]CO2, were 50–60% (n¼5). The same procedure was used to prepare the target tracers [11C]5a, [11C]5c, [11C]5e, [11C]5g, [11C]5i, and [11C]5k from their corresponding precursors 5b, 5d, 5f,
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Table 3 Analytical HPLC retention times (tR) of arylpiperazinylthioalkyl derivatives. tR (min) Precursor 5b 5d 5f 5h 5j 5l
Standard 2.12 2.28 2.39 2.27 2.41 2.57
5a 5c 5e 5g 5i 5k
Tracer 5.32 5.45 5.66 5.34 5.76 5.92
[11C]5a [11C]5c [11C]5e [11C]5g [11C]5i [11C]5k
5.32 5.45 5.66 5.34 5.76 5.92
5h, 5j, and 5l. Retention times of arylpiperazinylthioalkyl derivatives including the precursors, reference standards, and target tracers in the analytical HPLC system are listed in Table 3.
4. Conclusion An efficient and convenient synthesis of new 5-HT1AR radioligands, carbon-11-labeled arylpiperazinylthioalkyl derivatives, has been developed. The synthetic methodology employed simple twostep condensation to prepare a series of new arylpiperazinylthioalkyl derivatives, phenolic hydroxyl precursors, and their corresponding ether standard compounds. The target tracers were easily prepared by O-[11C]methylation of their corresponding precursors using a reactive [11C]methylating agent, [11C]CH3OTf, and isolated by a simplified SPE purification procedure in high radiochemical yields, short overall synthesis time, and high specific radioactivities. These chemistry results combined with the reported in vitro biological data (Siracusa et al., 2008) encourage further in vivo biological evaluation of new carbon-11-labeled arylpiperazinylthioalkyl derivatives as candidate PET agents for imaging of 5-HT1AR.
Acknowledgments 1
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