Synthesis, in vitro characterization, and radiolabeling of N,N-dimethyl-2-(2′-amino-4′-substituted-phenylthio)benzylamines: Potential candidates as selective serotonin transporter radioligands

Article abstract of DOI:10.1021/jm050079o

A series of N,N-dimethylated and N-monomethylated analogues of N,N-dimethyl-2-(2′-amino-4′-iodophenylthio)benzylamine substituted at the 4′-phenyl position have been prepared and evaluated in vitro for serotonin transporter (SERT) selectivity. Several der

Full text of DOI:10.1021/jm050079o

4254
J. Med. Chem. 2005, 48, 4254-4265
Synthesis, in Vitro Characterization, and Radiolabeling of
N,N-Dimethyl-2-(2′-amino-4′-substituted-phenylthio)benzylamines: Potential
Candidates as Selective Serotonin Transporter Radioligands
Nachwa Jarkas,† Jonathan McConathy,† Ronald J. Voll,† and Mark M. Goodman*,†,‡
Division of Radiological Sciences, Department of Radiology, and Department of Psychiatry and Behavior Sciences,
Emory University, Atlanta, Georgia 30322
Received January 28, 2005
A series of N,N-dimethylated and N-monomethylated analogues of N,N-dimethyl-2-(2′-amino-
4′-iodophenylthio)benzylamine substituted at the 4′-phenyl position have been prepared and
evaluated in vitro for serotonin transporter (SERT) selectivity. Several derivatives were
prepared where the 4′-position was either unsubstituted 13 and 33a or substituted with methyl
14a and 33b, ethenyl 14b and 34, ethyl 16 and 35, hydroxymethyl 20 and 41, hydroxyethyl
22, fluoroethyl 23, hydroxypropyl 27, and fluoropropyl 28. Competition binding in cells stably
expressing the transfected human SERT, dopamine transporter (DAT), and norepinephrine
transporter (NET) using [3H]citalopram, [3H]WIN 35,428 or [125I]RTI-55, and [3H]nisoxetine,
respectively, demonstrated the following order of SERT affinity (Ki (nM)): 14a (0.25) > 16
(0.49) > 20 (0.57) > 14b (1.12) > 13 (1.59) > 33b (1.94) ) 35 (2.04) . 23 (8.50) ) 28 (8.55) >
41 (15.11) . 22 (51) > 33a (83.43) > 27 (92). The Ki values revealed that most of these
derivatives displayed a high affinity for the SERT and a high selectivity over the DAT and
NET. Moreover, substitution at the 4′-position of the dimethylated and monomethylated
benzylamines differently influenced SERT binding: (i) the dimethylated benzylamines exhibited
higher SERT affinity than the monomethylated ones, (ii) alkyl, alkenyl, or hydroxymethyl
functions at the 4′-position afford compounds with high SERT affinity, and (iii) ω-hydroxy and
fluoro-substituted ethyl and propyl groups at the 4′-position decrease the SERT affinity. From
this series, the dimethylated derivatives 13, 14a, 14b, 16, and 20 were radiolabeled with carbon-
11 and their log P7.4 was calculated as a measure of their potential brain penetrance as positron
emission tomography SERT imaging agents.
Introduction
density.8-11 However, the precise role of the SERT
system in these neuropsychiatric disorders remains to
be clarified. For this reason, in vivo mapping of the
SERT in the living human brain would provide a useful
tool to better understand how alterations of this system
are related to depressive illness and other neurodegen-
erative disorders as well as to assess the quantification
of SERT occupancy in relation to the antidepressant
drug treatment.
There has been a considerable interest in the develop-
ment of suitable radiotracers for imaging the SERT
either by positron emission tomography (PET) or by
single photon emission computed tomography (SPECT).
Many classes of compounds such as pyrroloisoquino-
line,12 quipazine,13 and phenyl nortropane14 derivatives
have been screened for their affinities for the SERT. The
(+) enantiomer of trans-1,2,3,5,6,10â-hexahydro-6-[4-
(methylthio)phenyl]pyrrolo-[2,1-a]-isoquinoline ([11C]-
(+)McN5652),15 from the pyrroloisoquinoline series, was
the first successful ligand for imaging the SERT in
humans by PET. Unfortunately, this radiotracer has
several shortcomings: (i) the time required to reach
quasi-equilibrium, a condition when the ratio of region
of interest to reference region stays constant, is slow,
requiring at least 120 min of data acquisition; (ii) the
nonspecific binding is relatively high, which precludes
the measurements of lower SERT density regions such
as the frontal cortex and cingulate; (iii) the plasma free
Serotonin is an essential neurotransmitter in both the
central and peripheral nervous system. The widespread
distribution of serotonin neurons and fibers provides the
anatomical basis for the involvement of serotonin in the
modulation of cortical functions.1,2 Serotonin neurons
originate in the raphe nuclei, from where they project
into numerous brain regions including the olfactory
bulb, cerebral cortex, hippocampus, and basal ganglia.
The serotonin transporter (SERT), located on the cell
bodies of the raphe nuclei and the terminals of the
presynaptic serotonergic neurons in the brain, is neces-
sary for the regulation of synaptic serotonin levels.3-5
SERT is the target of several antidepressant drugs
(selective serotonin reuptake inhibitors), such as fluox-
etine, paroxetine, and sertraline which are designed to
preferentially increase serotonin transmission by in-
hibiting binding of serotonin to SERT.6,7 Recent reviews
have suggested that dysfunctions of serotonin neu-
rotransmission could be associated with neurological
and psychiatric disorders. Postmortem brain sections of
patients with depression and Alzheimer’s and Parkin-
son’s diseases have shown a decrease in the SERT
* To whom correspondence should be addressed: Mark M. Goodman,
Ph. D. Tel: (404) 727-9366. Fax: (404) 727-3488.
† Department of Radiology, Emory University.
‡ Department of Psychiatry and Behavior Sciences, Emory Univer-
sity.
10.1021/jm050079o CCC: $30.25 © 2005 American Chemical Society
Published on Web 06/04/2005

Serotonin Transporter Radioligands
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13 4255
Figure 1. Diphenyl sulfide derivatives.
fraction is very low (<1%), and it interferes with the
kinetic modeling using this tracer.16 In the quipazine
derivatives, only 5-iodo-6-nitroquipazine showed prom-
ising properties as an in vivo tracer for mapping the
SERT sites in a monkey’s brain. However, human study
of this agent has not been reported. In the phenyl
nortropane series, such as 3â-(4-ethyl-3-iodophenyl)-
nortropane-2-beta-carboxylic acid methyl ester,17 3â-(4′-
n-propylphenyl)nortropane-2â-carboxylic acid methyl
ester,14 3â-(4′-isopropylphenyl)nortropane-2â-carboxylic
acid methyl ester,14 2â-carbomethoxy-3â-(4′-(2-fluoro-
isopropenyl)phenyl)nortropane,18 2â-carbomethoxy-3â-
(4′-(2-fluoroethyl)-3′-iodophenyl)nortropane,19 2â-carbo-
methoxy-3â-(4′-(2-fluoroethyl)-3′-bromophenyl)nortro-
pane,19and2â-carbomethoxy-3â-(4′-(2-fluoroethyl)-3′-chloro-
phenyl)nortropane,19 none of these compounds except
2â-carbomethoxy-3â-(4′-((Z-2-iodoethenyl)phenyl)nor-
tropane20 and2â-carbomethoxy-3â-(4′-((Z-2-iodoethenyl)-
phenyl)tropane21 have physiochemical and kinetic prop-
erties desirable for optimal imaging of the SERT in the
human brain: the most common limitation has been a
relatively poor signal-to-noise ratio, limiting their use
in vivo for the quantification of the SERT. In contrast,
2â-carbomethoxy-3â-(4′-((Z-2-iodoethenyl)phenyl)tro-
pane and 2â-carbomethoxy-3â-(4′-((Z-2-iodoethenyl)-
phenyl)nortropane displayed a high SERT specific bind-
ing with a low nonspecific accumulation and have been
proposed as potential radioligands for PET and SPECT
imaging of the SERT, respectively.
Recently, it has been reported that a novel antide-
pressant, N,N-dimethyl-2-(4′-chloro-2′-hydroxymeth-
ylphenylthio)benzylamine (1) (Figure 1), was a high
affinity and selective SERT ligand.22 On the basis of the
substituted phenylthiophenyl motif of this compound,
a number of analogues were prepared and were reported
as potent and selective candidate SERT radioligands
(Figure 1). The iodo analogues N,N-dimethyl-2-(2′-
hydroxymethyl-4′-iodophenylthio)benzylamine (2) and
N,N-dimethyl-2-(2′-amino-4′-iodophenylthio)benzyl-
amine (3) (Figure 1) were labeled with iodine-123 and
showed promise as new SPECT imaging agents for the
SERT.22-28 [11C]-3 was prepared and evaluated as a PET
SERT radioligand. However, the slow binding kinetics
displayed by [11C]-3 were found to be inappropriate for
the short-lived carbon-11 labeled PET radioligand.29 In
attempts to develop a SERT imaging agent with kinetic
properties compatible with the 20 min half-life of
carbon-11, several alternative substituted benzylamines
have been synthesized, characterized, and evaluated by
PET imaging in nonhuman primates and humans. PET
imaging with carbon-11 labeled N,N-dimethyl-2-(2′-
amino-4′-methoxyphenylthio)benzylamine ([11C]-4)30 and
N,N-dimethyl-2-(2′-amino-4′-cyanophenylthio)benzyl-
amine ([11C]-5)30 in humans and [11C]-5,16 N,N-dimethyl-
2-(2′-amino-4′-methylphenylthio)benzylamine ([11C]-
6),31-33 N,N-dimethyl-2-(2′-amino-4′-fluoromethylphenyl-
thio)benzylamine ([11C]-7),16,34 and N,N-dimethyl-2-(2′-
amino-4′-bromophenylthio)benzylamine ([11C]-8)16,35 (Fig-
ure 1) in nonhuman primates has been reported. These
carbon-11 compounds displayed good specific-to-non-
specific (cerebellum) ratios in SERT-rich thalamus from
2:1 to 3.5:1. PET imaging studies in baboons with
carbon-11 analogues [11C]-3, [11C]-8, [11C]-5, and [11C]-
716 demonstrated that only [11C]-5 reached a state of
quasi-equilibrium in the thalamus within 40 min postin-
jection as determined from fitted time activity curves
between thalamus and cerebellum and supported its
candidacy as the preferred radioligand for imaging
SERT sites by PET. Thalamus-to-cerebellum ratios of
[11C]-3, [11C]-8, and [11C]-7 increased continuously up
to 90 min postinjection. [11C]-7 displayed the highest
specific-to-nonspecific ratio in the thalamus (3.5) but did
not achieve a state of quasi-equilibrium in the thalamus
by 90 min postinjection. These findings prompted us to
incorporate alternative alkyl-containing substituents at
the 4′-position of the diphenyl sulfide. These new N,N-
dimethyl-2-(2′-amino-4′-iodophenylthio)benzylamine 3
analogues may provide better candidates for developing
PET ligands for imaging the SERT in the living human
brain. Reported herein are the synthesis and in vitro
evaluation of several derivatives of N,N-dimethyl-2-(2′-
amino-4′-iodophenylthio)benzylamine where the 4′-posi-
tion of the aniline ring was either unsubstituted 13 and
33a or substituted with methyl 14a31-33 and 33b,
ethenyl 14b33 and 34, ethyl 1633,36 and 35, hydroxymeth-
yl 2037 and 41, hydroxyethyl 22, fluoroethyl 23,33,34,38

4256 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13
Jarkas et al.
Scheme 1
were commercially available. Compounds 9a, 9b, and
10c were prepared from thiosalicylic acid and 10b,
respectively, using previously described procedures.39,40,41
The 4-bromo-3-nitro-styrene (10d) was obtained from
the 4-bromo-3-nitro-benzaldehyde (10c)41 under Wittig
reaction conditions. Copper-catalyzed coupling42 of com-
pounds 10a, 10b, and 10d with either 2-thio-N,N-
dimethylbenzamide (9a)39 or ethylthiosalicylate (9b)40
gave the aryl thioethers 11a-c and 29a-c, respectively,
in good yields (Schemes 1 and 3). In the case of the
dimethyl derivatives (Schemes 1 and 2), only when
compound 9a was freshly prepared was a good coupling
yield achieved. Palladium-catalyzed reduction of the
nitro group of compound 11a under a hydrogen atmo-
sphere provided 12 which, after reduction of the amide
group with the diborane-tetrahydrofuran (1 M) com-
plex, was transformed to 13 in good yield. Reduction of
the nitro as well as the amide groups of compounds
11b-c was achieved using the lithium aluminum hy-
dride-tetrahydrofuran (1 M) complex to give N,N-
dimethyl-2-(2′-amino-4′-methylphenylthio)benzyla-
mine (14a) and N,N-dimethyl-2-(2′-amino-4′-ethenyl-
phenylthio)benzylamine (14b). N,N-Dimethyl-2-(2′-
amino-4′-ethylphenylthio)benzylamine (16) was ob-
tained from 11c following the same conditions used to
prepare 13. Radical bromination with N-bromosuccin-
imide of 11b proceeded smoothly in carbon tetrachlo-
ride43 yielding 17 which was transformed to 18 after a
nucleophilic substitution with potassium acetate. N,N-
Dimethyl-2-(2′-amino-4′-hydroxymethylphenylthio)ben-
zylamine (20) was obtained in good yield from 18, after
reduction with the diborane-tetrahydrofuran (1 M)
complex followed by a palladium-catalyzed reduction
(Scheme 2).
hydroxypropyl 27, and fluoropropyl 28.33 The in vitro
biological properties of the 3 analogues were character-
ized. Compounds 13, 14a, 14b, 16, and 20 showed both
good affinity and selectivity for the SERT and were
radiolabeled with carbon-11, and their lipophilicities
were determined: [11C]-13, [11C]-14a, [11C]-14b, [11C]-
16, and [11C]-20 were prepared by a one-step reaction
involving the alkylation of the precursors (the mono-
methylbenzylamines) 33a, 33b, 34, 35, and 41, respec-
tively, with the carbon-11 iodomethane.
Results and Discussion
Chemistry. The synthesis of the new target ligands
13, 14a, 14b, 16, 20, 22, 23, 27, and 28, as well as the
normethyl derivatives 33a, 33b, 34, 35, and 41, required
as precursors for radiolabeling with [11C]iodomethane,
is presented in Schemes 1-4. Compounds 10a and 10b
The syntheses of the ω-hydroxyalkyl and the ω-fluo-
roalkyl derivatives 22 and 27 and 23 and 28, respec-
Scheme 2

Serotonin Transporter Radioligands
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13 4257
Scheme 3
tively, are outlined in Scheme 2. The 4′-(2-hydroxyethyl)-
dimethylamine (21), was prepared by hydroboration of
11c, followed by hydrogenation in the presence of a
stoichiometric amount of palladium on charcoal to afford
N,N-dimethyl-2-(2′-amino-4′-(ethan-2-ol)phenylthio)ben-
zylamine (22). Compound 22 was promptly converted
to N,N-dimethyl-2-(2′-amino-4′-(2-fluoroethyl)phenyl-
thio)benzylamine (23) with diethylaminosulfur trifluo-
ride (DAST)44 at low temperature (Scheme 2). The
synthesis of fluoropropyl aniline 28 was initiated by the
copper-catalyzed coupling of 24, prepared by the Wittig
reaction of 10c using ethyl (triphenylphosphoranylidine)
acetate,45 with 9a to afford 25 in satisfactory yield
(Scheme 2). Hydrogenolysis of 25 over palladium hy-
droxide in absolute ethanol gave 26 which, after reduc-
tion with the diborane-tetrahydrofuran (1 M) complex,
was converted to N,N-dimethyl-2-(2′-amino-4′-(propan-
3-ol)phenylthio)benzylamine (27). Conversion of 27 to
N,N-dimethyl-2-(2′-amino-4′-(3-fluoropropyl) phenyl-
thio)benzylamine (28) was accomplished using DAST in
dichloromethane (DCM) (Scheme 2).The syntheses of
monomethylaminoanilines 33a, 33b, 34, 35, and 41
were analogous to the synthetic approaches reported in
Scheme 3 and Scheme 4. Saponification of the previ-
ously prepared esters 29a, 29b, and 29c afforded the
acids 30a, 30b, and 30c in very good yields. Treatment
of these acids with thionyl chloride in refluxing DCM
with a catalytic amount of dimethylformamide gave
crude acid chlorides which reacted, without further
purification, with methylamine hydrochloride in dichlo-
romethane to yield the desired benzamides 31a, 31b,
and 31c (Scheme 3). Monomethylaminoanilines 33a and
33b (Scheme 3) were prepared starting from amides 31a
and 31b using the same reaction conditions applied to
obtain 13. Lithium aluminum hydride reduction of nitro
amide 31c yielded monomethylaminoaniline 34 which,
after chromatographic purification, was converted to
amine 35 under the same hydrogenolysis conditions
used to reduce compound 25 (Scheme 3). Under the
same conditions used to prepare 18, 10b was trans-
formed to 37. Condensation of 37 with thiosalicylic acid
proceeded smoothly in dimethylformamide yielding 38
which, under the appropriate conditions, was trans-
formed to 39. Compound 39 gave 41 in good yield
(Scheme 4) when the same procedure was used to
prepare 13.
Radiochemistry. The radiotracers [11C]-13, [11C]-
14a, [11C]-14b, [11C]-16, and [11C]-20 were all synthe-
sized in a similar fashion as depicted in Scheme 5.
Cyclotron-produced [11C]iodomethane was trapped in
solutions of the monomethyl precursors 33a-35 and 41
at -20 °C in N,N-dimethylformamide and heated to 90
°C for 10 min. Purification of the resultant reaction
mixtures was accomplished by reverse-phase high-
performance liquid chromatography (HPLC) followed by
a solid-phase extraction purification46 (see Experimental
Section) and formulation in saline (containing 10%
ethanol) to give [11C]-13, 14a, 14b, 16, and 20 in 24 (
3% (n ) 2), 30 ( 9% (n ) 6), 4 ( 1% (n ) 3), 24 ( 1%
(n ) 6), and 21 ( 6% (n ) 6), respectively, radiochemical
yield EOS (decay corrected, from [11C]iodomethane
production). Quality control tests showed that the final
products were sterile, pyrogen free, and radiochemically
pure (>98%) and contained no precursors. The total
synthesis time was 65 min from the end of the bom-
bardment and the specific activity was 500-1200 mCi/
µmol at the end of the synthesis. A possible side reaction
in these syntheses is the methylation of the aromatic

4258 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13
Jarkas et al.
Scheme 4
Scheme 5
5000) except for the 4′-unsubstituted derivative 13 with
Ki ) 279 nM and Ki ) 4.95 nM for the DAT and NET,
respectively. In comparison with the N,N-dimethyl
derivatives, the hNET affinities of the corresponding
N-methyl derivatives were an order of magnitude lower.
Thus, the substitution at the 4′-position of the aniline
ring of the phenylthiophenyl core structure and demeth-
ylation of the N,N-dimethylbenzylamine substituent are
responsible for the decrease of the DAT and NET
affinities.
In our series, derivatives with hydrogen (13), methyl
(14a), ethenyl (14b), ethyl (16), and hydroxymethyl (20)
at the 4′-position showed nM or higher affinity, Ki )
1.59, 0.25, 1.12, 0.49, and 0.57 nM, respectively, for the
SERT. These five compounds differ from the other
derivatives by introduction of a short chain alkyl,
alkenyl, and ω-hydroxylalkyl group onto the 4′-position
of the aniline ring. Compared to the ethyl derivative (16)
(Ki ) 0.49 nM), the ω-hydroxyethyl (22) and ω-fluoro-
ethyl (23) derivatives displayed lower affinities Ki ) 51
and 8.50 nM, respectively, for the SERT. Lengthening
the ω-hydroxyethyl (22) and ω-fluoroethyl (23) deriva-
tives by one carbon afforded ω-propyl derivatives 27 and
28, respectively, that possessed approximately the same
SERT affinity as their ethyl homologues, Ki ) 92 and
8.55 nM, respectively. Thus, a bulk steric effect at that
4′-position of the aniline ring and the introduction of
polar atoms on the alkyl groups diminished the binding
potency of such derivatives.
The derivatives with the highest SERT binding, 13,
14a, 14b, 16, and 20, were radiolabeled with carbon-
11 to determine their log P at pH 7.4. The log P values
of compounds 13, 14a, 14b, 16, and 20 are presented
in Table 1. The lipophilicity (octanol/water partition),
log P7.4, is an important parameter for correlating
structure with brain penetrance and ligand-transporter
kinetic behavior. Compounds 13, 14a, 14b, 16, and 20
had log P7.4 values of 2.28, 2.46, 2.47, 2.60, and 1.6,
respectively. Comparison between the parent 4′-unsub-
stituted compound 13 and the 4′-substituted series
amino group, a site that should be less nucleophilic than
the targeted aliphatic amino group: no radioactive
byproducts were observed upon HPLC analysis of the
final products.
Biological Results: in Vitro Competition Assays.
The affinities of N,N-dimethyl derivatives 13, 14a, 14b,
16, 20, 22, 23, 27, and 28 and N-methyl derivatives 33a,
33b, 35, and 41 for the human SERT (hSERT), human
DAT (hDAT), and human NET (hNET) were determined
through in vitro competition assays. These data are
shown in Table 1 along with the previously determined
Ki values for 3.
[3H]Citalopram (SERT ligand), [3H]WIN 35,428 or
[
125I]RTI-55 (DAT ligand), and [3H]nisoxetine (NET
ligand) were used as radiotracers during the in vitro
displacement experiments. The N,N-dimethyl deriva-
tives 13, 14a, 14b, 16, 20, 22, 23, 27, and 28 displayed
high affinity and selectivity for the SERT over the DAT
and NET. The rank of order of hSERT affinity (Ki in
nM) for the N,N-dimethyl derivatives was 326 (0.013) >
14a (0.25) > 16 (0.49) > 20 (0.57) > 14b (1.12) > 13
(1.59) . 23 (8.50) ) 28 (8.55) . 22 (51) > 27 (92). In
comparison with the N,N-dimethyl derivatives, the
hSERT affinities of the corresponding N-methyl deriva-
tives were an order of magnitude or lower.
The N,N-dimethyl derivatives exhibited very low
affinities for the DAT (∼600 e Ki e 5000) as well as
moderate to very low affinities for the NET (37 e Ki e

Serotonin Transporter Radioligands
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13 4259
Table 1. Relative Transporters Affinities Ki (nM) of Candidate SERT Ligands in Transfected Cell Lines and log P
compd
326
13
X
R
SERTa (n)d
DATb (n)
NETc (n)
log P7.4
2.52
2.28 ( 0.02
2.46 ( 0.09
2.47 ( 0.09
2.60 ( 0.02
1.60 ( 0.02
I
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
H
0.013 ( 0.003
1.59 ( 1.23 (3)
0.25 ( 0.12 (4)
1.12 ( 1.40 (3)
0.49 ( 0.38 (5)
0.57 ( 0.05 (3)
50.58 (1)
8.50 ( 3.53 (1)
92.26 (1)
8.55 ( 3.46 (1)
83.43 (1)
840 ( 100
699 ( 80
H
279 ( 51.10 (3)
532 ( 106.00 (3)
962 ( 116.60 (3)
4.95 ( 0.71 (3)
61 ( 14.85 (2)
269 ( 113.85 (2)
370 ( 181.70 (3)
303 ( 225.57 (2)
194 (1)
14a
14b
16
CH3
CHdCH2
C2H5
>2000 (3)
20
CH2OH
(CH2)2OH
(CH2)2F
(CH2)3OH
(CH2)3F
H
>1000 (3)
>1000 (1)
>5000 (1)
>5000 (1)
>5000 (1)
22
23
>5000 (1)
27
108 (1)
28
219 (1)
31a
31b
35
969 ( 27.03 (1)
698 ( 250 (2)
>1000 (2)
>1000 (2)
37.52 ( 2.18 (2)
191 ( 14.14 (2)
445.36 ( 134.30 (2)
483.55 ( 207.11 (2)
CH3
H
1.94 ( 0.31 (3)
2.04 ( 0.29 (3)
15.11 ( 2.25 (2)
C2H5
CH2OH
H
41
H
a Competitive binding vs [3H]citalopram in human kidney cells transfected with human serotonin transporters. b Competitive binding
vs[3H]WIN 35,428 or [125I]RTI-55 in canine kidney cells transfected with human dopamine transporters. c Competitive binding vs
[3H]nisoxetine in human kidney cells transfected with human norepinephrine transporters. d Number of assays. Values were obtained
from the mean ( standard deviation of at least two independent assays, each in duplicates, except for compounds 22, 23, 27, 28, and 31a.
demonstrated that a methyl, vinyl, and ethyl group
increased log P7.4 by 0.18-0.32 and that the hydroxyl-
methyl group decreased log P7.4 by 0.68. These effects
can be explained by an increased polarity of 20, result-
ing in an increased solubility in the buffer caused by
the hydroxyl group forming intermolecular hydrogen
bonds with the aqueous buffer. Introduction of methyl,
vinyl, and ethyl groups caused the opposite effect
because the nonpolar alkyl and vinyl groups will orient
themselves toward and be attracted to the nonpolar
octanol. The initial brain uptake and specific and
nonspecific binding ratios in rats of a series carbon-11
analogues with CN (2.71, 5), CF3 (3.77), Cl (3.55), and
OCH3 (2.83) substituted at the 4′-position of the aniline
findings reported with [11C]2â-carbomethoxy-3â-(4′-((Z-
2-iodoethenyl)phenyl)tropane and [11C]2â-carbomethoxy-
3â-(4′-((Z-2-bromoethenyl)phenyl)tropane. In addition,
20 binding at the SERT exhibited the fastest kinetics
and achieved a quasi-equilibrium within 55 min.
Conclusion
Chemical modifications of a series of diphenyl sulfides
at the 2- and 4′-positions were performed. Evaluation
of the in vitro affinities to the human monoamine
transporters, SERT, DAT, and NET and log P7.4 values
showed that (i) the dimethylamino group at the 2-posi-
tion increases the SERT affinity; (ii) the substituent at
the 4′-position is critical for SERT binding; (iii) the
introduction of an alkyl substituent at the 4′-position
increases SERT affinity; (iv) the introduction of OH and
F at the ω-position of an alkyl group markedly decreases
the SERT affinity; (v) the introduction of an OH and F
at the ω-position of increasing larger alkyl groups does
not increase SERT affinity; (vi) the substituent at the
4′-position significantly influenced log P7.4 values with
short chain alkyl and alkenyl groups yielding log P7.4
values ) ∼2.5 and the introduction of OH, 20, affording
a significantly lower log P7.4 ) 1.6. Preliminary micro-
PET screening of carbon-11 labeled 13, 14a, 16, and 20
in anesthetized macaques demonstrated that 20 had the
highest peak midbrain-to-cerebellum ratios (∼ 4) and
the most favorable SERT binding kinetics reaching
quasi-equilibrium by 55 min postinjection. The full in
vivo characterization of 20 for mapping the SERT in the
macaque and human brain is currently under investiga-
tion.
ring have been reported to be optimum when log P7.4
)
2.71 (5).47-49 The analogue with the lowest log P7.4 value
of 2.71, 5, appeared to provide the best combination of
initial brain uptake (1.22% in dose/g) and specific to
nonspecific binding ratios, 7.9 to 1, for SERT-rich
midbrain regions to cerebellum, respectively, at 60 min
postinjection. The octanol/water partition method at pH
7.4 used for this study was identical to the method
employed for 5.47-49 Compounds 13, 14a, 14b, and 16
exhibiting log P7.4 values 2.28-2.60 were lower than 5.
The measured log P7.4 value for 20 of 1.6 was signifi-
cantly lower than 5. Although the log P7.4 value for 20
was 1.6, the recently reported tropanes [11C]2â-car-
bomethoxy-3â-(4′-((Z-2-iodoethenyl)phenyl)tropane 21 and
[11C]2â-carbomethoxy-3â-(4′-((Z-2-bromoethenyl)phen-
21
yl)tropane
with log P7.4 values 1.5 and 1.3, respec-
tively, showed good brain penetrance and good specific
(midbrain) to nonspecific binding (cerebellum) ratios
2.1:1 and 1.9:1, respectively, at 85 min postinjection in
nonhuman primates. A preliminary microPET screening
of carbon-11 labeled 13, 14a, 16, and 20 in anesthetized
macaques demonstrated that the cooperative properties
of Ki ) 0.57 and log P7.4 ) 1.6 for 20 resulted in the
highest specific accumulation in brain regions rich in
SERT, with a midbrain-to-cerebellum uptake ratio of
4.0. These results with 20 correlated well with the
Experimental Section
Melting points were determined with a Laboratory Device
MEL-TEMP II and uncorrected. Proton NMR spectra were run
on a Varian Unity 400 at 400 MHz (1H) in CDCl3 with
tetramethylsilane as an internal standard. Elemental analyses
were performed by Atlantic Microlabs, Inc. (Atlanta, GA) and
were within (0.4% of the theoretical values. Starting materials

4260 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13
Jarkas et al.
9b40 and 10c41 were prepared and purified by methods
previously reported.40,41 Compound 9a was freshly prepared
and used without further purification. All of the other chemi-
cals were purchased from Aldrich Chemical Co. and were used
without further purification. Purification and analyses of
radioactive compounds by HPLC were performed with in-line
UV (254 nm) detectors in series with a radioactivity detector.
The HPLC columns used were either a Waters C18 reverse
phase (25 mm × 100 mm) (column A) for purification or a
Waters C18 reverse phase (3.9 mm × 150 mm) (column B) for
quality control.
Celite, and the Celite filter cake was washed with several
portions of EtOH. The combined filtrate was concentrated
under vacuo. Purification by silica gel flash column chroma-
tography (DCM/MeOH, 9:1) afforded 12 as an orange oil (0.87
g, 82%). 1H NMR (CDCl3, δ): 2.90 (s, 3H, NCH3), 3.15 (s, 3H,
NCH3), 4.45 (s, 2H, NH2), 6.72 (m, 2H, ArH), 6.98 (m, 1H,
ArH), 7.19 (m, 4H, ArH), 7.46 (dd, 1H, J ) 7.6, 1.6 Hz, ArH).
Anal. (C15H16N2OS) C, H, N.
N,N-Dimethyl-2-(2′-aminophenylthio)benzylamine (13).
BH3-THF complex (1 M solution in THF) (15 mL, 15 mmol)
was added dropwise at 5 °C to a solution of compound 12 (0.83
g, 3.67 mmol) in dry THF (15 mL). The mixture was heated to
reflux for 2 h and left at room temperature overnight. The
reaction was quenched by the addition of concentrated HCl
at 0 °C, and the solvent was removed under vacuo. To the
residue, water (15 mL) was added, and the mixture was
refluxed for 30 min with vigorous stirring. After the mixture
was cooled to room temperature, a saturated solution of
NaHCO3 was added to neutralize the solution to pH values of
∼7-8. The resulting aqueous solution was extracted with
Et2O. Combined organic layers were dried over Na2SO4, and
the solvent was removed under vacuo. Silica gel flash column
chromatography purification (DCM/MeOH, 9:1) afforded 13 as
4-Bromo-3-nitrostyrene (10d). To a suspension of meth-
yltriphenylphosphonium bromide (1.98 g, 5.6 mmol) in dry
tetrahydrofuran (THF) (15 mL), was added butyllithium (1.6
M in hexane, 3.48 mL, 5.6 mmol) at room temperature and
stirred for 20 min. The mixture was then cooled to -78 °C
and 10c (0.64 g, 2.8 mmol) in dry THF (10 mL) was added
dropwise. After complete addition, the reaction mixture was
allowed to warm to room temperature and stirred for 30 min.
The solvent was concentrated under vacuo, and the residue
was extracted with dichloromethane (DCM). Organic layers
were washed with saturated aqueous Na2CO3 solution and
water, dried over Na2SO4, and concentrated under vacuo to
give the crude product. Purification by silica gel flash column
chromatography (DCM) afforded the product 10d as a yellow
1
an orange oil (0.48 g, 50%). H NMR (CDCl3, δ): 2.49 (s, 6H,
NCH3), 3.76 (s, 2H, CH2), 4.67 (s, 2H, NH2), 6.91 (m, 2H, ArH),
7.06 (m, 1H, ArH), 7.25 (m, 2H, ArH), 7.40 (m, 2H, ArH), 7.66
(dd, 1H, J ) 8.0, 1.6 Hz, ArH). Anal. (C15H18N2S) C, H, N.
1
oil (400 mg, 63%). H NMR (CDCl3, δ): 5.45 (d, 1H, J ) 10.8
Hz, sCHdCH2), 5.85 (d, 1H, J ) 18 Hz, sCHdCH2), 6.66 (dd,
1H, J ) 17.2, 11.2 Hz, sCHdCH2), 7.42 (dd, 1H, J ) 8.4, 2.0
Hz, ArH), 7.66 (d, 1H, J ) 8.8 Hz, ArH), 7.83 (d, 1H, J ) 1.6
Hz, ArH). Anal. (C8H6BrNO2) C, H, N.
N,N-Dimethyl-2-(2′-amino-4′-methylphenylthio)ben-
zylamine31-33 (14a). To a 1 M lithium aluminum hydride
solution in THF (9.48 mL, 9.48 mmol) was added a solution of
amide 11b (0.60 g, 1.89 mmol) in dry THF (3 mL) at room
temperature under argon. The resulting brown solution was
refluxed overnight. After the mixture was cooled to room
temperature, it was quenched with a saturated NH4Cl solution
and extracted with ether. Combined organic layers were
washed with 1 N HCl solution. Aqueous layers were basified
with saturated Na2CO3 solution and extracted with Et2O.
Concentration of the solvent gave the crude product. Silica gel
flash column chromatography purification (DCM/MeOH, 9:1)
afforded 14a as an orange oil (0.20 g, 39%). 1H (CDCl3, δ): 2.29
(s, 3H, CH3), 2.31 (s, 6H, NCH3), 3.57 (s, 2H, CH2N), 4.30 (s,
2H, NH2), 6.59 (m, 2H, ArH), 6.86 (m, 1H, ArH), 7.07 (m, 2H,
ArH), 7.23 (m, 1H, ArH), 7.37 (dd, 1H, J ) 7.2, 0.8 Hz, ArH).
Anal. (C16H20N2S) C, H, N.
N,N-Dimethyl-2-(2′-amino-4′-ethenylphenylthio)ben-
zylamine (14b). The alkene 14b was prepared by the method
described for 14a, by reduction of amide 11c (0.10 g, 0.30
mmol) with 1 M LAH/THF (1.52 mL, 1.52 mmol). Silica gel
flash column chromatography purification (DCM/MeOH, 9:1)
afforded 14b as a colorless oil (30 mg, 35%). 1H NMR (CDCl3,
δ): 2.55 (s, 6H, NCH3), 3.94 (s, 2H, CH2N), 5.30 (d, 1H, J )
11.2 Hz, sCHdCH2), 5.78 (d, 1H, J ) 16.8 Hz, -CHdCH2),
6.64 (dd, 1H, J ) 10.4, 7.2 Hz, sCHdCH2), 6.83 (m, 2H, ArH),
6.90 (m, 1H, ArH), 7.15 (m, 2H, ArH), 7.35 (d, 1H, J ) 7.6 Hz,
ArH), 7.45 (m, 1H, ArH). Anal. (C17H20N2S) C, H, N.
General Procedure for the Copper-Catalyzed Cou-
pling. To a solution of differently substituted bromo-nitro
compounds 10a, 10b, 10d, or 24 (1 equiv) in toluene was added
Cs2CO3 (3 equiv) followed by CuI (0.1 equiv) as a catalyst. The
reaction mixture was stirred at 100 °C for 15 min. 9a (2 equiv)
or 9b (2 equiv) was added, and the mixture was refluxed
overnight. After the mixture was cooled to room temperature,
the solvent was concentrated under vacuo and the residue was
extracted with DCM. Organic layers were washed with satu-
rated aqueous Na2CO3 solution and water, dried over Na2SO4,
and concentrated under vacuo to give the crude product.
Purification by silica gel flash column chromatography (hex-
ane/EtOAc, 9:1) afforded the desired compound in good yield.
N,N-Dimethyl-2-(2′-nitrophenylthio)benzamide (11a).
When the general coupling procedure was followed, condensa-
tion of 10a (1.11 g, 5.14 mmol) with 9a (1.86 g, 10.28 mmol)
afforded 11a as a light brown oil (1.18 g, 76%). 1H NMR
(CDCl3, δ): 2.85 (s, 3H, NCH3), 3.03 (s, 3H, NCH3), 7.21 (m,
1H, ArH), 7.43 (m, 4H, ArH), 7.57 (m, 2H, ArH), 8.18 (dd, 1H,
J ) 10.8, 2.0 Hz, ArH). Anal. (C15H14N2O3S) C, H, N.
N,N-Dimethyl-2-(4′-methyl-2′-nitrophenylthio)benza-
mide (11b). When the general coupling procedure was fol-
lowed, condensation of 10b (3.29 g, 15.23 mmol) with 9a (5.52
g, 30.5 mmol) afforded 11b as an orange solid (2.13 g, 44%):
1
mp ) 102 °C. H NMR (CDCl3, δ): 2.33 (s, 3H, CH3), 2.84 (s,
3H, NCH3), 3.03 (s, 3H, NCH3), 6.84 (d, 1H, J ) 8.0 Hz, ArH),
7.18 (dd, 1H, J ) 8.0, 1.6 Hz, ArH), 7.44 (m, 2H, ArH), 7.55
(m, 2H, ArH), 7.98 (d, 1H, J ) 1.2 Hz, ArH). Anal. (C16H16N2-
O3S) C, H, N.
N,N-Dimethyl-2-(4′-ethenyl-2′-nitrophenylthio)benza-
mide (11c). When the general coupling procedure was fol-
lowed, condensation of 10d (3.02 g, 13.24 mmol) with 9a (4.80
g, 26.49 mmol) afforded 11c as a light brown oil (0.93 g, 21%).
1H NMR (CDCl3, δ): 2.85 (s, 3H, NCH3), 3.04 (s, 3H, NCH3),
5.34 (d, 1H, J ) 10.8 Hz, sCHdCH2), 5.76 (d, 1H, J ) 17.6
Hz, sCHdCH2), 6.63 (dd, 1H, J ) 17.2, 11.1 Hz, sCHdCH2),
6.89 (d, 1H, J ) 8.6 Hz, ArH), 7.45 (m, 3H, ArH), 7.55 (m, 2H,
ArH), 8.17 (d, 1H, J ) 1.6 Hz, ArH). Anal. (C17H16N2O3S) C,
H, N.
N,N-Dimethyl-2-(2′-amino-4′-ethylphenylthio)benza-
mide (15). Compound 15 was prepared by the method
described for amine 12, through the reduction of compound
11c (0.12 g, 0.36 mmol) with 10% Pd/C (0.12 g). Silica gel flash
column chromatography purification (DCM/EtOAc, 8:2) af-
forded 15 as a yellow solid (90 mg, 82%): mp ) 105 °C. 1H
NMR (CDCl3, δ): 1.32 (t, 3Η, J ) 7.6 Ηz, CΗ3), 2.65 (q, 2Η,
J ) 8.0 Hz, CH2), 2.99 (s, 3H, NCH3), 3.24 (s, 3H, NCH3), 4.51
(s, 2H, NH2), 6.67 (m, 2H, ArH), 7.05 (d, 1H, J ) 8.0 Hz, ArH),
7.25 (m, 3H, ArH), 7.44 (d, 1H, J ) 8.0 Hz, ArH). Anal.
(C17H20N2OS) C, H, N.
N,N-Dimethyl-2-(2′-amino-4′-ethylphenylthio)benzyl-
amine (16). Compound 16 was prepared by the method
described for amine 13, through the reduction of amide 15
(0.20 g, 0.67 mmol) with BH3-THF (1 M solution in THF) (3.35
mL, 3.35 mmol). Silica gel flash column chromatography
purification (DCM/MeOH, 9:1) afforded 16 as a yellow oil (80
mg, 42%). 1H NMR (CDCl3, δ): 1.23 (t, 3H, J ) 10.0 Hz, CH3),
2.31 (s, 6H, NCH3), 2.58 (q, 2H, J ) 10.0 Hz, CH2), 3.57 (s,
N,N-Dimethyl-2-(2′-aminophenylthio)benzamide (12).
To a solution of the compound 11a (1.18 g, 3.90 mmol)
dissolved in absolute EtOH (25 mL) was added 10% Pd/C (1.18
g). The resulting black slurry was stirred under a hydrogen
atmosphere for 24 h. The slurry was then filtered through

Serotonin Transporter Radioligands
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13 4261
2H, CH2N), 4.35 (s, 2H, NH2), 6.60 (m, 2H, ArH), 6.87 (m, 1H,
ArH), 7.05 (m, 2H, ArH), 7.28 (m, 1H, ArH), 7.38 (dd, 1H, J )
6.8, 4.0 Hz, ArH). Anal. (C17H22N2S) C, H, N.
a colorless oil (8.40 mg, 19%). 1H NMR (CDCl3, δ): 2.65 (s,
6H, NCH3), 2.82 (t, 2H, J ) 6.4 Hz, PhCH2), 3.89 (t, 2H, J )
6.4 Hz, CH2OH), 4.19 (s, 2H, NH2), 4.29 (s, 2H, CH2N), 6.65
(dd, 1H, J ) 8.0, 1.6 Hz, ArH), 6.69 (d, 1H, J ) 1.6 Hz, ArH),
6.89 (dd, 1H, J ) 7.6, 1.6 Hz, ArH), 7.18 (m, 2H, ArH), 7.29
(d, 1H, J ) 8.0 Hz, ArH), 7.37 (dd, 1H, J ) 7.2, 1.6 Hz, ArH).
Anal. (C17H22N2OS) C, H, N.
N,N-Dimethyl-2-(2′-amino-4′-(2-fluoroethyl)phenyl-
thio)benzylamine (23). A solution of DAST (0.13 mL, 0.96
mmol) in DCM (1 mL) was slowly added to a solution of
compound 22 (0.15 g, 0.49 mmol) in DCM (2 mL) cooled to
-70 °C. The reaction mixture was warmed to room tempera-
ture and mixed with cold water. The organic phase was washed
with saturated aqueous Na2CO3 followed by water, and the
resulting organic layers were combined, dried over Na2SO4,
and concentrated under vacuo. After silica gel flash column
chromatography purification (DCM/MeOH, 9:1), compound 23
was obtained as a yellow oil (16.8 mg, 11%). 1H NMR (CDCl3,
δ): 2.67 (s, 6H, NCH3), 2.85 (dt, 2H, J ) 24.3, 6.1 Hz, PhCH2),
4.59 (s, 2H, CH2N), 4.69 (d t, 2H, J ) 47.2, 6.1 Hz, CH2F),
6.68 (dd, 1H, J ) 8.0, 1.6 Hz, ArH), 6.80 (m, 2H, ArH), 7.20
(m, 2H, ArH), 7.32 (m, 1H, ArH), 7.40 (d, 1H, J ) 8.1 Hz, ArH).
Anal. (C17H21FN2S) C, H, N.
N,N-Dimethyl-2-(4′-bromomethyl-2′-nitrophenylthio)-
benzamide (17). A mixture of 11b (0.50 g, 1.6 mmol),
N-bromosuccinimide (0.42 g, 2.4 mmol), and azobisisobutyro-
nitrile (AIBN) (25 mg) was refluxed for 16 h in CCl4 (15 mL).
The solvent was removed under vacuo, and the residue was
purified by silica gel flash column chromatography using
(DCM/EtOAc, 8:2) as the eluent. Compound 17 was obtained
as a yellow oil (0.30 g, 47%). 1H NMR (CDCl3, δ): 2.85 (s, 3H,
NCH3), 3.03 (s, 3H, NCH3), 4.42 (s, 2H, CH2Br), 7.38 (dd, 1H,
J ) 8.4, 2.0 Hz, ArH), 7.51 (m, 1H, ArH), 7.56 (m, 2H, ArH),
7.60 (m, 2H, ArH), 8.21 (d, 1H, J ) 2.0 Hz, ArH). Anal. (C16H15-
BrN2O3S) C, H, N.
N,N-Dimethyl-2-(4′-acetoxymethyl-2′-nitrophenylthio)-
benzamide (18). A mixture of compound 17 (69.0 mg, 0.17
mmol) and CH3CO2K (17.17 mg, 0.17 mmol) was stirred at
100 °C for 15 h in DMF (2 mL). The mixture was cooled to
room temperature, poured into cold acidic water, and extracted
several times with DCM. The combined organic layers were
washed thoroughly with water, dried over Na2SO4, and
concentrated under vacuo. Silica gel flash column chromatog-
raphy purification (DCM/EtOAc, 8:2) afforded 18 as a yellow
oil (31.8 mg, 49%). 1H NMR (CDCl3, δ): 2.09 (s, 3H, OCOCH3),
2.85 (s, 3H, NCH3), 3.04 (s, 3H, NCH3), 5.06 (s, 2H, CH2O),
6.93 (d, 1H, J ) 8.4 Hz, ArH), 7.34 (dd, 1H, J ) 8.4, 2.4 Hz,
ArH), 7.47 (m, 2H, ArH), 7.57 (m, 2H, ArH), 8.19 (d, 1H, J )
2.0 Hz, ArH). Anal. (C16H16N2O3S) C, H, N.
N,N-Dimethyl-2-(4′-hydroxymethyl-2′-nitrophenylthio)-
benzylamine (19). Compound 19 was prepared by the method
described for amine 13, through the reduction of amide 18
(31.8 mg, 0.085 mmol) with BH3-THF (1 M solution in THF)
(0.42 mL, 0.42 mmol). Silica gel flash column chromatography
purification (DCM/MeOH, 9:1) afforded 19 as an orange oil
(11.6 mg, 43%). 1H NMR (CDCl3, δ): 2.21 (s, 6H, NCH3), 3.57
(s, 2H, CH2N), 4.71 (s, 2H, CH2OH), 6.68 (d, 1H, J ) 8.4 Hz,
ArH), 7.33 (m, 2H, ArH), 7.52 (m, 2H, ArH), 7.70 (d, 1H, J )
7.6 Hz, ArH), 8.24 (m, 1H, ArH). Anal. (C16H18N2O3S) C, H,
N.
N,N-Dimethyl-2-(2′-amino-4′-hydroxymethylphenyl-
thio)benzylamine (20). Compound 20 was prepared by the
method described for amine 12, through the reduction of 19
(35.6 mg, 0.11 mmol) with 10% Pd/C (36 mg). Silica gel flash
column chromatography purification (DCM/MeOH/Et3N, 9:1:
0.1) afforded 20 as a yellow oil (29 mg, 90%). 1H NMR (CDCl3,
δ): 2.34 (s, 6H, NCH3), 3.64 (s, 2H, CH2N), 4.59 (s, 2H, CH2-
OH), 6.69 (dd, 1H, J ) 7.6, 1.6 Hz, ArH), 6.83 (m, 1H, ArH),
7.06 (m, 2H, ArH), 7.09 (m, 1H, ArH), 7.27 (m, 1H, ArH), 7.40
(d, 1H, J ) 8.0 Hz, ArH). Anal. (C16H20N2OS) C, H, N.
trans-Ethyl-3-(5-(2-bromonitrophenyl))-2-propeno-
ate (24). A solution of ethyl triphenylphosphoranylidine
acetate (0.38 g, 1.08 mmol) dissolved in CH3CN (3 mL) was
added while stirring, to a CH3CN solution (2 mL) containing
10c (0.25 g, 1.08 mmol). The reaction mixture was refluxed
overnight. After the mixture was cooled, the solvent was
removed under vacuo to afford a crude solid that was purified
by silica gel flash column chromatography using (hexane/
EtOAc, 9:1) as the eluent. Compound 24 was obtained as a
1
white solid (0.26 g, 80%): mp ) 130 °C. H NMR (CDCl3, δ):
1.34 (t, 3H, J ) 6.8 Hz, CH3), 4.29 (q, 2H, J ) 7.2 Hz, CH2),
6.53 (d, 1H, J ) 16.4 Hz, -CHCO2C2H5), 7.55 (dd, 1H, J )
8.8, 2.6 Hz, ArH), 7.63 (d, 1H, J ) 16.4 Hz, sCHdCHCO2C2H5),
7.77 (d, 1H, J ) 8.6 Hz, ArH), 7.97 (d, 1H, J ) 2.6 Hz, ArH).
Anal. (C11H10BrNO4) C, H, N.
trans-1-Ethyl-3-(4′-(2-(N,N-dimethylamido)phenylthio)-
2′-nitro-phenyl)-2-propenoate (25). When the general cou-
pling procedure was followed, condensation of 24 (0.23 g, 0.76
mmol) with 9a (0.28 g, 1.53 mmol) afforded, after silica gel
flash column chromatography purification (DCM/EtOAc, 8:2),
1
25 as an orange solid (0.21 g, 68%): mp ) 105 °C. H NMR
(CDCl3, δ): 1.32 (t, 3H, J ) 7.6 Hz, CH3), 2.85 (s, 3H, NCH3),
3.04 (s, 3H, NCH3), 4.25 (q, 2H, J ) 7.2 Hz, CH2), 6.40 (d, 1H,
J ) 16.4 Hz, -CHCO2C2H5), 6.92 (d, 1H, J ) 8.8 Hz, ArH),
7.50 (m, 3H, ArH), 7.57 (d, 1H, J ) 16.4 Hz, sCHd
CHCO2C2H5), 7.60 (m, 2H, ArH), 8.32 (d, 1H, J ) 2.8 Hz, ArH).
Anal. (C20H20N2O5S) C, H, N.
1-Ethyl-3-(4′-(2-(N,N-dimethylamido)phenylthio)-2′-
phenylamine)propanoate (26). To a suspension of pal-
ladium hydroxide (1.00 g) in absolute EtOH (10 mL) was slowly
added the amide 25 (1.50 g, 3.74 mmol) in EtOH (40 mL). The
mixture was stirred under a hydrogen atmosphere for 20 h.
Filtration through Celite and concentration gave a green solid.
Purification by silica gel flash column chromatography (DCM/
EtOAc, 8:2) afforded 26 as a yellow oil (0.28 g, 20%). 1H NMR
(CDCl3, δ): 1.23 (t, 3H, J ) 7.6 Hz, CH3), 2.59 (t, 2H, J ) 7.6
Hz, CH2), 2.86 (t, 2H, J ) 8.0 Hz, CH2), 2.89 (s, 3H, NCH3),
3.15 (s, 3H, NCH3), 4.14 (q, 2H, J ) 7.6 Hz, CH2), 6.96 (m,
1H, ArH), 7.18 (m, 2H, ArH), 7.37 (d, 1H, J ) 8.0 Hz, ArH),
7.45 (m, 2H, ArH), 7.67 (d, 1H, J ) 8.4 Hz, ArH). Anal.
(C20H24N2O3S) C, H, N.
N,N-Dimethyl-2-(2′-amino-4′-(propan-3-ol)phenylthio)-
benzylamine (27). Compound 27 was prepared by the method
described for amine 13, through the reduction of compound
26 (0.50 g, 1.34 mmol) with BH3-THF (1 M solution in THF)
(6.71 mL, 6.71 mmol). Silica gel flash column chromatography
purification (DCM/MeOH, 9:1) afforded 27 as a white solid
(0.11 g, 26%): mp ) 94 °C. 1H NMR (CDCl3, δ): 1.89 (m, 2H,
CH2), 2.31 (s, 6H, NCH3), 2.64 (t, 2H, J ) 7.2 Hz, PhCH2),
3.58 (s, 2H, CH2N), 3.68 (t, 2H, J ) 6.4 Hz, CH2OH), 4.41 (s,
2H, NH2), 6.60 (dd, 1H, J ) 12.0, 1.6 Hz, ArH), 6.86 (m, 2H,
N,N-Dimethyl-2-(4′-(ethan-2-ol)-2′-nitrophenylthio)-
benzylamine (21). BH3-THF complex (1 M solution in THF)
(3.04 mL, 3.04 mmol) was added dropwise at 5 °C to a solution
of compound 11c (0.20 g, 0.61 mmol) in dry THF (4 mL). The
mixture was heated to reflux for 15 h. A few drops of 3 N
NaOH solution followed by a few drops of 30-35% H2O2 were
added at 0 °C, and the reaction mixture was left at room
temperature for 1 h. The solvent was removed under vacuo,
and the resulting aqueous solution was extracted with DCM.
Combined organic layers were dried over Na2SO4 and concen-
trated under vacuo. Silica gel flash column chromatography
purification (DCM/EtOAc, 8:2) afforded 21 as an orange oil
(24 mg, 12%).1H NMR (CDCl3, δ): 2.60 (s, 6H, NCH3), 2.87 (t,
2H, J ) 6.4 Hz, PhCH2), 3.88 (t, 2H, J ) 6.0 Hz, CH2OH),
4.20 (s, 2H, CH2N), 6.43 (d, 1H, J ) 8.4 Hz, ArH), 7.23 (dd,
1H, J ) 6.8, 2.0 Hz, ArH), 7.49 (m, 1H, ArH), 7.56 (m, 1H,
ArH), 7.64 (d, 1H, J ) 7.6 Hz, ArH), 7.75 (d, 1H, J ) 6.4 Hz,
ArH), 8.15 (d, 1H, J ) 6.4 Hz, ArH). Anal. (C17H20N2O3S) C,
H, N.
N,N-Dimethyl-2-(2′-amino-4′-(ethan-2-ol)phenylthio)-
benzylamine (22). Compound 22 was prepared by the method
described for amine 12, through the reduction of 21 (48 mg,
0.14 mmol) with 10% Pd/C (48 mg). Silica gel flash column
chromatography purification (DCM/EtOAc, 8:2) afforded 22 as

4262 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13
Jarkas et al.
ArH), 7.07 (m, 2H, ArH), 7.24 (m, 1H, ArH), 7.39 (d, 1H, J )
8.4 Hz, ArH). Anal. (C18H24N2OS) C, H, N.
and a few drops of DMF. The reaction mixture was refluxed
for 90 min under argon. The solvent was evaporated, and the
solid was used without further purification.
N,N-Dimethyl-2-(2′-amino-4′-(3-fluoropropyl)phenylthio)-
benzylamine (28). The fluoropropylamine 28 was prepared
by the method described for the fluoroethylamine 23, by the
reaction of 27 (0.20 g, 0.63 mmol) with DAST (0.18 mL, 1.3
mmol). After silica gel flash column chromatography purifica-
tion (DCM/MeOH, 9:1), compound 28 was obtained as a yellow
oil (20 mg, 10%). 1H NMR (CDCl3, δ): 2.01 (dtt, 2H, J ) 25.2,
7.2, 6.0 Hz, CH2), 2.33 (s, 6H, NCH3), 2.67 (t, 2H, J ) 8.0 Hz,
PhCH2), 3.61(s, 2H, CH2N), 4.48 (d t, 2H, J ) 47.2, 6.0 Hz,
CH2F), 6.58 (m, 2H, ArH), 6.86 (m, 1H, ArH), 7.08 (m, 2H,
ArH), 7.24 (m, 1H, ArH), 7.38 (d, 1H, J ) 8.0 Hz, ArH). Anal.
(C18H23FN2S) C, H, N.
Ethyl 2-(2′-Nitrophenylthio)benzoate (29a). When the
general coupling procedure was followed, condensation of 10a
(0.33 g, 1.63 mmol) with 9b (0.6 g, 3.27 mmol) afforded 29a
as a yellow oil (0.41 g, 83%). 1H NMR (CDCl3, δ): 0.96 (t, 3H,
J ) 7.2 Hz, CH3), 4.03 (q, 2H, J ) 6.8 Hz, CH2), 6.78 (dd, 1H,
J ) 8.4, 1.2 Hz, ArH), 7.04 (m, 1H, ArH), 7.14 (m, 1H, ArH),
7.25 (m, 3H, ArH), 7.67 (m, 1H, ArH), 7.93 (dd, 1H, J ) 8.5,
1.6 Hz, ArH). Anal. (C15H13NO4S) C, H, N.
A solution of triethylamine (1.20 g, 11.91 mmol) and
methylamine hydrochloride (0.40 g, 5.96 mmol) in anhydrous
DCM (20 mL) was cooled to -70 °C. The acid chloride (freshly
prepared) in DCM was added dropwise under argon. The
reaction mixture was allowed to warm to room temperature,
stirred for 30 min, and mixed with cold water. The organic
phase was washed with 1 N HCl solution followed by water,
and the resulting organic layers were combined, dried over
Na2SO4, and concentrated under vacuo. The product was
purified by silica gel flash column chromatography (DCM/
EtOAc, 8:2) to yield the desired 31a as an orange oil (0.37 g,
86%). 1H NMR (CDCl3, δ): 2.55 (d, 3H, NCH3), 6.09 (s, 1H,
NH), 6.63 (d, 1H, J ) 8.0 Hz, ArH), 6.96 (m, 1H, ArH), 7.08
(m, 1H, ArH), 7.24 (m, 3H, ArH), 7.48 (dd, 1H, J ) 7.6, 1.6
Hz, ArH), 7.91 (dd, 1H, J ) 8.4, 0.8 Hz, ArH). Anal.
(C14H12N2O3S) C, H, N.
N-Methyl-2-(4′-methyl-2′-nitrophenylthio)benzamide
(31b). Acid 30b (0.89 g, 3.08 mmol) was treated with thionyl
chloride (0.91 g, 7.69 mmol) as described for 31a. The crude
acid chloride was treated with methylamine hydrochloride
(0.83 g, 12.30 mmol) in the presence of triethylamine (2.49 g,
24.61 mmol). The monomethylamide derivative 31b was
purified by silica gel flash column chromatography (DCM/
EtOAc, 8:2) to give a yellow solid (0.67 g, 72%): mp ) 140 °C.
1H NMR (CDCl3, δ): 2.35 (s, 3H, CH3), 2.84 (d, 3H, J ) 4.8
Hz, NCH3), 6.41 (s, 1H, NH), 6.81 (d, 1H, J ) 8.0 Hz, ArH),
7.22 (dd, 1H, J ) 8.0, 1.2 Hz, ArH), 7.52 (m, 3H, ArH), 7.77
(dd, 1H, J ) 7.6, 1.6 Hz, ArH), 7.98 (m, 1H, ArH). Anal.
(C15H14N2O3S) C, H, N.
N-Methyl-2-(4′-ethenyl-2′-nitrophenylthio)benza-
mide (31c). Acid 30c (0.21 g, 0.70 mmol) was treated with
thionyl chloride (0.21 g, 1.74 mmol) as described for 31a. The
crude acid chloride was treated with methylamine hydrochlo-
ride (0.19 g, 2.79 mmol) in the presence of triethylamine (0.56
g, 5.57 mmol). The monomethylamide derivative 31c was
purified by silica gel flash column chromatography (DCM/
EtOAc, 8:2) to give an orange oil (0.19 g, 87%). 1H NMR
(CDCl3, δ): 2.84 (d, 3H, J ) 4.4 Hz, NCH3), 5.36 (d, 1H, J )
10.8 Hz, sCHdCH2), 5.77 (d, 1H, J ) 17.6 Hz, sCHdCH2),
6.32 (s, 1H, NH), 6.65 (dd, 1H, J ) 18.0, 10.8 Hz, sCHdCH2),
6.80 (d, 1H, J ) 8.4 Hz, ArH), 7.41 (dd, 1H, J ) 8.4, 1.6 Hz,
ArH), 7.55 (m, 3H, ArH), 7.75 (dd, 1H, J ) 7.6, 1.6 Hz, ArH),
8.18 (d, 1H, J ) 1.6 Hz, ArH). Anal. (C16H14N2O3S) C, H, N.
N-Methyl-2-(2′-aminophenylthio)benzamide (32a). Amide
31a (0.37 g, 1.28 mmol) was treated with 10% Pd/C (0.37 g)
as described for 12. The desired compound 32a was obtained
as an orange oil (0.27 g, 82%). 32a was spectroscopically pure
and used in the next step without further purification. 1H NMR
(CDCl3, δ): 3.04 (d, 3H, NCH3), 4.38 (s, 2H, NH2), 6.11 (s, 1H,
NH), 6.76 (m, 1H, ArH), 6.86 (dd, 1H, J ) 8.0, 0.8 Hz, ArH),
7.18 (m, 4H, ArH), 7.44 (m, 2H, ArH).
Ethyl 2-(4′-Methyl-2′-nitrophenylthio)benzoate (29b).
When the general coupling procedure was followed, condensa-
tion of 10b (2.00 g, 9.26 mmol) with 9b (3.37 g, 18.52 mmol)
1
afforded 29b as a yellow solid (1.69 g, 58%): mp ) 80 °C. H
NMR (CDCl3, δ): 1.25 (t, 3H, J ) 7.2 Hz, CH3), 2.39 (s, 3H,
CH3), 4.29 (q, 2H, J ) 6.8 Hz, CH2), 7.00 (d, 1H, J ) 8.4 Hz,
ArH), 7.23 (dd, 1H, J ) 8.4, 1.2 Hz, ArH), 7.35 (m, 1H, ArH),
7.44 (m, 2H, ArH), 7.92 (m, 2H, ArH). Anal. (C16H15NO4S) C,
H, N.
Ethyl 2-(4′-Ethenyl-2′-nitrophenylthio)benzoate (29c).
When the general coupling procedure was followed, condensa-
tion of 10d (0.31 g, 1.36 mmol) with 9b (0.50 g, 2.72 mmol)
afforded 29c as a yellow oil (0.30 g, 67%). 1H NMR (CDCl3, δ):
1.22 (t, 3H, J ) 6.8 Hz, CH3), 4.27 (q, 2H, J ) 7.2 Hz, CH2),
5.39 (d, 1H, J ) 10.8 Hz, sCHdCH2), 5.82 (d, 1H, J ) 17.6
Hz, sCHdCH2), 6.67 (dd, 1H, J ) 18.0, 10.8 Hz, sCHdCH2),
6.96 (d, 1H, J ) 17.6 Hz, ArH), 7.42 (dd, 1H, J ) 8.0, 2.4 Hz,
ArH), 7.48 (m, 3H, ArH), 7.92 (m, 1H, ArH), 8.15 (d, 1H, J )
2.0 Hz, ArH). Anal. (C17H15NO4S) C, H, N.
2-(2′-Nitrophenylthio)benzoic Acid (30a). Compound
29a (0.51 g, 1.68 mmol) was heated in 3 N NaOH (16 mL)
solution at 120 °C for 20 min. After the solution was cooled to
room temperature, the reaction mixture was acidified with
concentrated HCl. Filtration gave the desired compound 30a
as an orange solid (0.41 g, 89%): mp >250 °C. 30a was
spectroscopically pure and used in the next step without
further purification. 1H NMR (CDCl3, δ): 7.11 (dd, 1H, J )
8.0, 1.2 Hz, ArH), 7.34 (m, 1H, ArH) 7.41 (dd, 1H, J ) 7.6, 1.6
Hz, ArH), 7.49 (m, 3H, ArH), 8.11 (m, 2H, ArH). Anal. (C13H9-
NO4S) C, H, N.
2-(4′-Methyl-2′-nitrophenylthio)benzoic Acid (30b). Acid
30b was prepared from ester 29b (1.64 g, 5.17 mmol) as
described for the preparation of 30a. Hydrolysis of 29b
afforded 30b as an orange solid (1.46 g, 98%): mp ) 245 °C.
30b was spectroscopically pure and used in the next step
without further purification. 1H NMR (CDCl3, δ): 2.42 (s, 3H,
CH3), 7.12 (d, 1H, J ) 8.4 Hz, ArH), 7.27 (m, 2H, ArH), 7.40
(m, 1H, ArH), 7.47 (m, 1H, ArH), 7.89 (m, 1H, ArH), 8.10 (dd,
1H, J ) 8.0, 1.6 Hz, ArH). Anal. (C14H11NO4S) C, H, N.
2-(4′-Ethenyl-2′-nitrophenylthio)benzoic Acid (30c).
Acid 30c was prepared from ester 29c (0.33 g, 1.00 mmol) as
described for the preparation of 30a. Hydrolysis of 29c afforded
30c as a yellow solid (0.21 g, 70%): mp ) 172 °C. 30c was
spectroscopically pure and used in the next step without
further purification. 1H NMR (CDCl3, δ): 5.41 (d, 1H, J ) 10.8
Hz, sCHdCH2), 5.83 (d, 1H, J ) 17.6 Hz, sCHdCH2), 6.69
(dd, 1H, J ) 17.6, 10.8 Hz, sCHdCH2), 7.06 (d, 1H, J ) 8.4
Hz, ArH), 7.45 (m, 4H, ArH), 8.10 (m, 2H, ArH). Anal. (C17H11-
NO4S) C, H, N.
N-Methyl-2-(4′-methyl-2′-aminophenylthio)benza-
mide (32b). Amide 31b (0.67 g, 2.22 mmol) was treated with
10% Pd/C (0.63 g) as described for 12. The desired compound
32b was obtained following purification by silica gel flash
column chromatography (DCM/EtOAc, 8:2) as a yellow solid
(0.52 g, 86%): mp ) 130 °C. 1H NMR (CDCl3, δ): 2.29 (s, 3H,
CH3), 3.05 (d, 3H, J ) 4.8 Hz, NCH3), 4.29 (s, 2H, NH2), 6.08
(s, 1H, NH), 6.59 (m, 2H, ArH), 6.85 (dd, 1H, J ) 8.0, 1.2 Hz,
ArH), 7.11 (m, 1H, ArH), 7.19 (m, 1H, ArH), 7.32 (d, 1H, J )
8.0 Hz, ArH), 7.46 (dd, 1H, J ) 7.6, 1.6 Hz, ArH). Anal.
(C15H16N2OS) C, H, N.
N-Methyl-2-(2′-aminophenylthio)benzylamine (33a).
Amide 32a (0.27 g, 1.04 mmol) was treated with BH3-THF (1
M solution in THF) (4.18 mL, 4.18 mmol) as described for 13.
The desired compound 33a was obtained following purification
by silica gel flash column chromatography (DCM/MeOH, 9:1)
1
as a colorless oil (84 mg, 33%). H NMR (CDCl3, δ): 2.55 (s,
N-Methyl-2-(2′-nitrophenylthio)benzamide (31a). To a
suspension of the acid 30a (0.41 g, 1.49 mmol) in anhydrous
DCM (16 mL) were added thionyl chloride (0.27 g, 2.23 mmol)
3H, NCH3), 3.97 (s, 2H, CH2N), 6.79 (m, 2H, ArH), 6.86 (m,
1H, ArH), 7.13 (m, 2H, ArH), 7.28 (m, 1H, ArH), 7.35 (dd, 1H,

Serotonin Transporter Radioligands
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13 4263
J ) 7.2, 1.2 Hz, ArH), 7.44 (dd, 1H, J ) 7.6, 1.6 Hz, ArH).
Anal. (C14H16N2S) C, H, N.
δ): 2.09 (s, 3H, OCOCH3), 2.86 (d, 3H, NCH3), 5.06 (s, 2H,
CH2O), 6.31 (s, 1H, NH), 6.88 (d, 1H, J ) 8.4 Hz, ArH), 7.35
(dd, 1H, J ) 8.4, 2.0 Hz, ArH), 7.53 (m, 3H, ArH), 7.75 (dd,
1H, J ) 7.6, 1.6 Hz, ArH), 8.19 (d, 1H, J ) 2.0 Hz, ArH). Anal.
(C17H16N2O5S) C, H, N.
N-Methyl-2-(4′-acetoxymethyl-2′-aminophenylthio)-
benzamide (40). Amide 39 (0.35 g, 0.97 mmol) was treated
with 10% Pd/C (0.35 g) as described for 12. The desired
compound 40 was obtained as an orange oil and used without
further purification.
N-Methyl-2-(2′-amino-4′-methylphenylthio)benzyla-
mine (33b). Amide 32b (0.12 g, 0.44 mmol) was treated with
BH3-THF (1 M solution in THF) (1.76 mL, 1.76 mmol) as
described for 13. The desired compound 33b was obtained
following purification by silica gel flash column chromatogra-
phy (DCM/MeOH/Et3N, 95:5:0.1) as yellow crystals (75 mg,
66%): mp ) 76 °C. 1H NMR (CDCl3, δ): 2.30 (s, 3H, CH3),
2.51 (s, 3H, NCH3), 3.92 (s, 2H, CH2N), 6.57 (dd, 1H, J ) 7.6,
1.6 Hz, ArH), 6.61 (m, 1H, ArH), 6.80 (m, 1H, ArH), 7.08 (m,
2H, ArH), 7.29 (m, 2H, ArH). Anal. (C15H18N2S) C, H, N.
N-Methyl-2-(2′-amino-4′-ethenylphenylthio)benzyla-
mine (34). Amide 31c (0.10 g, 0.32 mmol) was treated with 1
M LAH/THF (1.60 mL, 1.60 mmol) as described for 14a. The
desired compound 34 was obtained following purification by
silica gel flash column chromatography (DCM/MeOH, 9:1) as
N-Methyl-2-(2′-amino-4′-hydroxymethylphenylthio)-
benzylamine (41). Amide 40 (0.22 g, 0.66 mmol) was treated
with BH3-THF (1 M solution in THF) (3.0 mL, 3.0 mmol) as
described for 13. The desired compound 41 was obtained
following purification by silica gel flash column chromatogra-
1
phy (DCM/MeOH, 9:1) as a yellow oil (59 mg, 32%). H NMR
(CDCl3, δ): 2.93 (s, 3H, NCH3), 4.35 (s, 2H, CH2N), 5.05 (s,
2H, CH2OH), 7.15 (dd, 1H, J ) 8.0, 1.6 Hz, ArH), 7.22 (d, 1H,
J ) 1.6 Hz, ArH), 7.24 (dd, 1H, J ) 7.2, 2.0 Hz, ArH), 7.52 (m,
2H, ArH), 7.74 (dd, 1H, J ) 7.2, 2.0 Hz, ArH), 7.81 (d, 1H,
J ) 8.0 Hz, ArH). Anal. (C15H18N2OS) C, H, N.
1
an orange oil (37 mg, 43%). H NMR (CDCl3, δ): 2.59 (s, 3H,
NCH3), 4.16 (s, 2H, CH2N), 5.30 (d, 1H, J ) 10.8 Hz, sCHd
CH2), 5.78 (d, 1H, J ) 17.6 Hz, sCHdCH2), 6.63 (dd, 1H, J )
17.6, 10.8 Hz, sCHdCH2), 6.81 (m, 3H, ArH), 7.13 (m, 2H,
ArH), 7.30 (d, 1H, J ) 8.0 Hz, ArH), 7.51 (dd, 1H, J ) 6.8, 1.6
Hz, ArH). Anal. (C16H18N2S) C, H, N.
Radiochemistry. (1) Radiosynthesis of [11C]-(13). [11C]-
Iodomethane, produced from 11CO2 as described previously,21
was swept by a flow of argon gas (25 mL/min) into a solution
of 33a (1.0 mg) in DMF (0.2 mL) at -20 °C. When the
radioactivity had peaked, the reaction mixture was heated to
90 °C for 10 min and quenched with HPLC buffer (0.5 mL).
The solution was purified by semipreparative HPLC on column
A: 50% CH3CN, 50% H2O, 6.3 g of CH3CO2NH4 (0.082 M), 9
mL/min (retention time (rt) of [11C]-13 ) 14.86 min, rt of
33a ) 7.85 min). The appropriate fractions were collected and
condensed via a solid-phase extraction procedure based upon
Lemaire’s method.46 An aliquot (0.1 mL) of the formulated
solution was used to establish the chemical and radiochemical
purity and specific activity of the final solution by analytical
HPLC on column B: 70% MeOH, 30% H2O, 0.1% Et3N, 1 mL/
min (rt of [11C]-13 ) 5.08 min). The radiochemical yield for
[11C]-13 was 24 ( 3% (n ) 2). The chemical and radiochemical
purity was >99%. [11C]-14a, [11C]-14b, [11C]-16, and [11C]-20
were prepared in a similar manner with minor adjustments
to the HPLC conditions. The radiochemical yields for [11C]-
14a, [11C]-14b, [11C]-16, and [11C]-20 were 30 ( 9% (n ) 6),
4 ( 1% (n ) 3), 24 ( 1% (n ) 6), and 21 ( 6% (n ) 6),
respectively. Their chemical and radiochemical purities were
>99%. Further evidence for the identity of the radiolabeled
products was achieved by co-injection with authentic “cold”
material on column B.
(2) Solid-Phase Extraction. The 4.5 mL fractions of [11C]-
13 collected from the semipreparative HPLC column were
diluted with sodium carbonate solution (6 mL, 12 mg/mL) and
combined in one flask. The homogeneous solution was trans-
ferred with vacuum, through a Cook line, to a C18 Sep Pak
(previously activated with ethanol (10 mL) and water (10 mL))
after stirring; the eluate was directed to the waste. After being
washed five successive times, four with saline (0.9% NaCl, 10
mL) and one with ethanol (0.5 mL), all being directed to the
waste, the [11C]-13 was eluted with ethanol (1.5 mL) and
driven to a sterile empty vial containing 0.9% NaCl solution
(3.5 mL). The residue was pushed with argon, through a
Millipore filter (pore size 1.0 µm) followed by a smaller one
(pore size 0.2 µm), to a 30 mL sterile, empty vial containing
0.9% NaCl solution (10 mL) for log P determinations.
N-Methyl-2-(2′-amino-4′-ethylphenylthio)benzyla-
mine (35). Amine 34 (80 mg, 0.29 mmol) was treated with
Pd(OH)2 (100 mg) as described for 26. The desired compound
35 was obtained following purification by silica gel flash
column chromatography (DCM/MeOH, 9:1) as a yellow solid
1
(22 mg, 27%): mp ) 56 °C. H NMR (CDCl3, δ): 1.24 (t, 3H,
J ) 7.6 Hz, CH3), 2.50 (s, 3H, NCH3), 2.60 (q, 2H, J ) 7.6 Hz,
CH2), 3.92 (s, 2H, CH2N), 6.60 (m, 2H, ArH), 6.80 (m, 1H, ArH),
7.07 (m, 2H, ArH), 7.28 (m, 1H, ArH), 7.33 (d, 1H, J ) 7.6 Hz,
ArH). Anal. (C16H20N2S) C, H, N.
2-Bromo-5-bromomethyl-nitrobenzene (36). Compound
10b (1.0 g, 4.63 mmol) reacted with N-bromosuccinimide (1.23
g, 6.94 mmol) and a catalytic amount of AIBN (72 mg) in CCl4
(40 mL) as described for 17. In this case, the reaction mixture
was refluxed for 3 days. After silica gel flash column chroma-
tography was used (hexane/EtOAc, 9:1), 36 was obtained as a
yellow oil (1.03 g, 75%). 1H NMR (CDCl3, δ): 4.45 (s, 2H, CH2-
Br), 7.47 (dd, 1H, J ) 8.4, 2.4 Hz, ArH), 7.72 (d, 1H, J ) 8.4
Hz, ArH), 7.87 (d, 1H, J ) 2.0 Hz, ArH). Anal. (C7H5Br2NO2)
C, H, N.
2-Bromo-5-acetoxymethyl-nitrobenzene (37). Com-
pound 36 (0.20 g, 0.68 mmol) was treated with CH3CO2K (0.68
mmol, 66 mg) in DMF (7 mL) as described for 18. The desired
compound 37 was obtained following purification by silica gel
flash column chromatography (hexane/EtOAc, 9:1) as a yellow
1
oil (0.15 g, 81%). H NMR (CDCl3, δ): 2.13 (s, 3H, OCOCH3),
5.11 (s, 2H, CH2O), 7.43 (dd, 1H, J ) 8.0, 2.0 Hz, ArH), 7.74
(d, 1H, J ) 8.4 Hz, ArH), 7.84 (d, 1H, J ) 2.0 Hz, ArH). Anal.
(C9H8BrNO4) C, H, N.
2-(4′-Acetoxymethyl-2′-nitrophenylthio)benzoic acid
(38). Thiosalicylic acid (0.14 g, 0.91 mmol) followed by K2CO3
(0.38 g, 2.73 mmol) was added to a solution of 37 (0.25 g, 0.91
mmol) in DMF (3 mL). The reaction mixture was stirred at
130 °C for 18 h. The brown solution was poured into cold acidic
water after it was cooled to room temperature, and the residue
extracted with EtOAc. Organic layers were washed with water,
dried over Na2SO4, and concentrated under vacuo to give the
crude product. Purification by silica gel flash column chroma-
tography (DCM/MeOH, 9:1) afforded the desired compound 38
as a yellow oil (55 mg, 17%). 1H NMR (CDCl3, δ): 2.10 (s, 3H,
OCOCH3), 5.06 (s, 2H, CH2O), 7.03 (d, 1H, J ) 8.4 Hz, ArH),
7.36 (m, 4H, ArH), 7.98 (d, 1H, J ) 7.2 Hz, ArH), 8.05 (d, 1H,
J ) 0.8 Hz, ArH). Anal. (C16H13NO6S) C, H, N.
N-Methyl-2-(4′-acetoxymethyl-2′-nitrophenylthio)benz-
amide (39). Acid 38 (0.32 g, 0.92 mmol) was treated with
thionyl chloride (0.27 g, 2.30 mmol) as described for 31a. The
crude acid chloride was treated with methylamine hydrochlo-
ride (0.25 g, 3.71 mmol) in the presence of triethylamine (0.75
g, 7.43 mmol). The monomethylamide derivative 39 was
purified by silica gel flash column chromatography (DCM/
MeOH, 9:1) to give a yellow oil (0.27 g, 81%). 1H NMR (CDCl3,
(3) log P Measurements. The determination of partition
coefficients of radioactive compounds [11C]-13, [11C]-14a, [11C]-
14b, [11C]-16, and [11C]-20 between 1-octanol and 0.02 M
phosphate buffer at pH 7.4 was performed as reported
previously.47-49
In Vitro Competition Assays. Competition assays were
performed based on methods reported previously.21,50 Cell
membranes from HEK-293 cells stably expressing hSERT or
hNET (a gift from Dr. Randy Blakely, Ph.D., Vanderbilt
University) and Madin Darby canine kidney cells stably
expressing hDAT (a gift from Dr. Gary Rudnick, Yale Univer-
sity) were used in these assays. Cells were grown to confluency

4264 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 13
Jarkas et al.
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11 000 rpm for 12 s in 30 vol of PBS. The resulting cell
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polystyrene tubes in a 2.0 mL final volume consisting of 1.7
mL of assay buffer, 100 µL of competing ligand in assay buffer,
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and then serially diluted in 5 mM HCl.
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For SERT assays, the assay buffer consisted of 53 mM Tris
buffer, 126 mM NaCl, and 5.3 mM KCl (pH 7.9 at room
temperature) and the equilibrium incubation time was 2 h at
room temperature. The radioligand used for SERT assays was
[3H]citalopram obtained from Dupont NEN (Boston, MA, 3100
GBq/mmol). For NET assays, the assay buffer consisted of 53
mM Tris buffer, 320 mM NaCl, and 5.3 mM KCl (pH 7.4 at 4
°C) and the equilibrium incubation time was 4 h at 4 °C. The
radioligand used for NET assays was [3H]nisoxetine obtained
from Dupont NEN (3000 GBq/mmol). For the DAT assay, the
assay buffer consisted of 42 mM sodium phosphate buffer and
320 mM sucrose (pH 7.4 at room temperature) and the
equilibrium incubation time was 1 h at room temperature. The
radioligand used for DAT assays was [3H]WIN 35,428 obtained
from Dupont NEN (Boston, MA, 84.5 Ci/mmol, 2.0 nM final
concentration) or [125I]RTI-55 obtained from Dupont NEN
(Boston, MA, 2200 Ci/mmol, 2.0 nM final concentration). All
of the assays were initiated by the addition of the cell
membrane suspension.
At the end of the incubation, the assays were terminated
by the addition of ∼5 mL of assay buffer at 4 °C followed by
rapid vacuum filtration with 3 × 5 mL washes with assay
buffer at 4 °C through GF/B filters (Whatman, Inc., Clifton,
NJ) presoaked in assay buffer containing 0.3% polyethylene-
imine. The data from the competition curves were analyzed,
and Ki values were calculated using GraphPad Prism software
(GraphPad software, San Diego, CA). The reported Ki values
are the average of at least two separate assays, each in
duplicates, for each compound.
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examination of three potent cocaine derivatives as specific
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H. F.; Laruelle, M. Comparative evaluation in nonhuman
primates of five PET radiotracers for imaging the serotonin
transposters:
[ [ [ [
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Supporting Information Available: Theoretical and
experimental elemental analyses for compounds 10d-41. This
material is available free of charge via the Internet at http://
pubs.acs.org.
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