Synthesis, biodistribution, and primate imaging of fluorine-18 labeled 2β-carbo-1'-fluoro-2-propoxy-3β-(4-chlorophenyl)tropanes. Ligands for the imaging of dopamine transporters by positron emission tomography

Article abstract of DOI:10.1021/jm9902234

2β-(R)-Carbo-1-fluoro-2-propoxy-3β-(4-chlorophenyl)tropane ((R)-FIPCT, R-6) and 2β-(S)-carbo-1-fluoro-2-propoxy-3β-(4-chlorophenyl)tropane ((S)- FIPCT, S-6) were prepared and evaluated in vitro and in vivo for dopamine transporter (DAT) selectivity and sp

Full text of DOI:10.1021/jm9902234

J . Med. Chem. 2000, 43, 639-648
639
Syn th esis, Biod istr ibu tion , a n d P r im a te Im a gin g of F lu or in e-18 La beled
2â-Ca r bo-1′-flu or o-2-p r op oxy-3â-(4-ch lor op h en yl)tr op a n es. Liga n d s for th e
Im a gin g of Dop a m in e Tr a n sp or ter s by P ositr on Em ission Tom ogr a p h y
Dongxia Xing,†,‡ Ping Chen,†,‡ Robert Keil,†,‡ Clinton D. Kilts,†,§ Bing Shi,†,‡ Vernon M. Camp,†,‡
Gene Malveaux,†,‡ Timothy Ely,§ Michael J . Owens,§ J ohn Votaw,†,‡ Margaret Davis,†,| J ohn M. Hoffman,†,‡
Roy A. E. BaKay, Thygarajan Subramanian,| Ray L. Watts,| and Mark M. Goodman*,†,‡
Emory Center for Positron Emission Tomography and Departments of Radiology, Psychiatry and Behavior Sciences,
Neurology, and Neurosurgery, Emory University, Atlanta, Georgia 30320
Received May 5, 1999
2â-(R)-Carbo-1-fluoro-2-propoxy-3â-(4-chlorophenyl)tropane ((R)-FIPCT, R-6) and 2â-(S)-carbo-
1-fluoro-2-propoxy-3â-(4-chlorophenyl)tropane ((S)-FIPCT, S-6) were prepared and evaluated
in vitro and in vivo for dopamine transporter (DAT) selectivity and specificity. High specific
activity [18F](R)-FIPCT and [18F](S)-FIPCT were synthesized in 5% radiochemical yield (decay-
corrected to end of bombardment (EOB)) by preparation of the precursors 2â-carbo-R-1-
mesyloxy-2-propoxy-3â-(4-chlorophenyl)tropane (R-12) and 2â-carbo-S-1-mesyloxy-2-propoxy-
3â-(4-chlorophenyl)tropane (S-12) followed by treatment with no carrier-added potassium[18F]-
fluoride and kyrptofix K222 in acetonitrile. Competition binding in cells stably expressing the
transfected human DAT and serotonin transporter (SERT) labeled by [3H]WIN 35428 and [3H]-
citalopram, respectively, demonstrated the following order of DAT affinity (Ki in nM): GBR
12909 (0.36) > CIT (0.48) > (S)-FIPCT (0.67) . (R)-FIPCT (3.2). The affinity of (S)-FIPCT and
(R)-FIPCT for SERT was 127- and 20-fold lower, respectively, than for DAT. In vivo
biodistribution studies were performed in male rats and demonstrated that the brain uptake
of [18F](R)-FIPCT and [18F](S)-FIPCT were selective and specific for DAT rich regions (caudate
and putamen). PET brain imaging studies in monkeys demonstrated high [18F](R)-FIPCT and
[18F](S)-FIPCT uptake in the caudate and putamen which resulted in caudate-to-cerebellum
and putamen-to-cerebellum ratios of 2.5-3.5 at 115 min. [18F](R)-FIPCT uptake in the caudate/
putamen achieved transient equilibrium at 75 min. In an imaging experiment with [18F](S)-
FIPCT in a rhesus monkey with its left hemisphere lesioned with MPTP, radioactivity was
reduced to background in the caudate and putamen of the lesioned hemisphere. The high specific
activity one-step radiolabeling preparation and high specificity and selectivity of [18F](R)-FIPCT
and [18F](S)-FIPCT for DAT indicate [18F](R)-FIPCT and [18F](S)-FIPCT are potential radio-
ligands for mapping brain DAT in humans using PET.
In tr od u ction
and euphoric effects of cocaine are initiated by the drug-
induced inhibition of DAT associated with mesolimb-
ocortical DA systems.5-9 These findings have resulted
in the development of a number of positron emitting
radioligands10-16 for quantitating cerebral DAT density
using positron emission tomography (PET). The most
extensively investigated series of carbon-11 and fluorine-
18 labeled radioligands for imaging the density of DAT
are those in which the 3â-(benzoyl) group of (-)-cocaine
was replaced by 3â-(4-substituted-phenyl) groups.17
Several analogues of this series of 2â-carbomethoxy-3â-
(4-substituted-phenyl)tropanes were found to be greater
than 100 times more potent than (-)-cocaine.
The radiolabeling methods utilized for the preparation
of the carbon-11 tropanes involved either N-alkylation
of the 8-position of the corresponding nortropane or
O-alkylation of N-fluoroalkyl nortropane 2â-carboxylic
acids with high specific activity 11CH3I. The carbon-11
labeled DAT imaging agents include [11C]2â-carbo-
methoxy-3â-(4-fluorophenyl)tropane ([11C]-WIN 35,428/
CFT),18-21 [11C]2â-carbomethoxy-3â-(4-iodophenyl)tro-
pane ([11C]CIT/RTI-55),22 [11C]2â-carbomethoxy-3â-(4-
The dopamine transporter (DAT) complex is a protein
that is located on the membranes of presynaptic me-
sotelencephalic dopaminergic neurons which originate
in the substantia nigra and ventral tegmentum located
in the midbrain and converge to the basal ganglia of
the forebrain. The DAT, a member of the family of Na+
Cl--dependent transporters, mediates uptake of dopa-
mine (DA) by an electrogenic sodium and chloride
transport coupled mechanism. Alterations of the density
and function of DAT have been implicated in psycho-
motor disorders such as Parkinson’s disease,1-3 Hun-
tington’s chorea,3 and schizophrenia.4 It is also widely
accepted that the reinforcing, psychomotor stimulant,
* Address correspondence and requests for reprints to Mark M.
Goodman, Ph.D., Emory University, Dept. of Radiology, 1364 Clifton
Road, NE, Atlanta, GA 30322. Phone: (404) 727-9366. Fax: (404) 727-
3488. E-mail: mgoodma@emory.edu.
† Emory Center for Positron Emission Tomography.
‡ Department of Radiology.
§ Department of Psychiatry and Behavior Sciences.
| Department of Neurology.
Department of Neurosurgery.
10.1021/jm9902234 CCC: $19.00 © 2000 American Chemical Society
Published on Web 01/29/2000

640 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4
Xing et al.
Sch em e 1a
chlorophenyl)tropane ([11C]CCT),21 [11C]2â-carbomethoxy-
3â-(3,4-dichlorophenyl)tropane ([11C]CDCT),21 [11C]3â-
(4-iodophenyl)tropane-2â-carboxylic acid isopropyl ester
([11C]RTI-121),23 N-3-fluoropropyl-2â-carbomethoxy-3â-
(4-iodophenyl)nortropane (O-methyl-[11C]FP-CIT) 24 and
N-2-fluoroethyl-2â-carbomethoxy-3â-(4-iodophenyl)nor-
tropane ([11C] â-CIT-FE).25 An undesired property ac-
companying DAT binding by [11C]-WIN 35,428, [11C]-
CCT, [11C]CDCT, and [11C]CIT/RTI-55 in nonhuman
and human primates is the need for a prolonged (>2 h)
period required to reach binding equilibrium with the
DAT. Unlike the parent, [11C]CIT/RTI-55, O-methyl-
[11C]FP-CIT, and [11C]â-CIT-FE DAT binding have been
reported to reach a transient equilibrium.24,25
a
Reagents: (a) LiAlH4, diethyl ether; (b) 1:1 dioxane: H2O; (c)
Fluorine-18 offers an advantage in comparison to the
short 20 min half-life of carbon-11 for ligands which
show delayed binding equilibrium. Fluorine-18’s 110
min half-life allows a longer time (3 × 110 min) for
imaging radioligands which require a longer period of
time to reach a binding equilibrium. Radiosyntheses of
fluorine-18 labeled N-3-fluoropropyl-2â-carbomethoxy-
3â-(4-iodophenyl)nortropane (â-CIT-FP),26 2â-carbo-
2′-fluoroethoxy-3â-(4-chlorophenyl)tropane (FECT),27
2â-carbo-2′-fluoroethoxy-3â-(4-methylphenyl)tropane
(FETT),27 2â-carbomethoxy-3â-(4-chlorophenyl)-8-(3-
fluoropropyl)nortropane (FPCT),28 and 2â-carbometh-
oxy-3â-(4-chlorophenyl)-8-(2-fluoroethyl)nortropane (FE-
CNT)29 in which the methyl group at the 8-position of
the tropane was replaced by a 3-fluoropropyl group and
a 2-fluoroethyl group and the 2â-methyl ester by a 2â-
2′-fluoroethyl ester have recently been reported. Fluorine-
18 labeled FPCT and FECNT were prepared by a two-
step reaction sequence which first involved the prepa-
ration of the radiolabeled precursors 1-[18F]fluoro-3-
iodopropane and 1-[18F]fluoro-2-tosyloxyethane, respec-
tively, followed by alkylation with 2â-carbomethoxy-3â-
(4-chlorophenyl)nortropane to afford [18F]FPCT and
[18F]FECNT in 15-21% overall yield corrected for decay
at EOB. In a similar manner, fluorine-18 labeled FECT
and FETT were prepared from the radiolabeled precur-
sor 1-[18F]fluoro-2-bromopropane followed by alkylation
with 2â-carboxy-3â-(4-chlorophenyl)tropane and 2â-
carboxy-3â-(4-methylphenyl)tropane.27 A successful one-
step radiosynthesis of fluorine-18 labeled FP-CIT has
been reported,26 though in only a 1% radiochemical
yield. The necessity to employ a two-step reaction
sequence to achieve good reproducible radiochemical
yields is a major shortcoming for the use of the N-[18F]-
fluoroalkylnortropanes and O-[18F]fluoroalkyltropanes
for potential widespread clinical use. Fluorine-18 labeled
DAT ligands prepared in high specific activity and good
radiochemical yield by a one-step radiolabeling method
are more desirable for production and distribution in a
hospital based radiopharmacy.
POCl3.
iPr) was found to show low nanomolar potency (IC50
)
1.41 nM vs [3H]CFT) and to be 1000 times more
selective for the DAT than the SERT (IC50 ) 1404 nM
using [3H]paroxetine) and NET (IC50 ) 2154 nM vs [3H]-
mazindol). These results suggested that the replacement
of the 2â-carbomethoxy functionality of CTC-iPr with a
2â-(R,S) 2′-fluoroisopropoxy bioisostere may also lead
to a high affinity and highly selective ligand for the
DAT.
In this paper we report the synthesis of 2â-(R,S)-
carbo-1′-fluoro-2-propoxy-3â-(4-chlorophenyl)tropane
((R,S)-FIPCT), the diastereomeric syntheses of 2â-(R)-
carbo-1′-fluoro-2-propoxy-3â-(4-chlorophenyl)tropane ((R)-
FIPCT) and 2-â-(S)-carbo-1′-fluoro-2-propoxy-3â-(4-
chlorophenyl)tropane ((S)-FIPCT), in vitro and in vivo
evaluation, and a one-step radiochemical synthesis and
PET imaging in nonhuman primates to evaluate the
effect of the 2â-(R)-1′-fluoro-2-propoxy and 2â-(S)-1′-
fluoro-2-propoxy substituents on striatal affinity, up-
take, and retention of the two new tropane analogues
R-FIPCT and S-FIPCT. These new analogues are at-
tractive candidates for labeling with fluorine-18 for
quantitating cerebral DAT density using positron emis-
sion tomography (PET).
Ch em istr y
The 2â-(R,S)-carbo-1′-fluoro-2-propoxy-3â-(4-chloro-
phenyl)tropane (R,S-FIPCT) (6) DAT ligand of this
study was prepared in a four step sequence of reactions
delineated in Scheme 1. In this synthetic approach,
fluoroacetone (1) was treated with LiAlH4 to form an
enantiomeric mixture of (R,S)-1-fluoro-2-propanol (2).
2â-Carboxy-3â-(p-chlorophenyl)tropane (4) was pre-
pared from 2â-carbomethoxy-3â-(p-chlorophenyl)tro-
pane (3) by treatment with a 1:1 mixture of dioxane and
H2O under gentle reflux.29 2â-Carboxy-3â-(p-chloro-
phenyl)tropane (4) was converted to 2â-methanoyl
chloride-3â-(p-chlorophenyl)tropane (5)29 by treatment
with phosphorus oxychloride. Esterification of acid
chloride 5 with (R,S)-1-fluoro-2-propanol (2) gave R,S-
FIPCT (6) as a 3:2 R:S mixture of diastereomers as
determined by high-pressure liquid chromatographic
analysis (HPLC).
In addition to high affinity and specificity, high
selectivity is another important characteristic for a DAT
radioligand developed for in vivo imaging. Recently, it
was reported that replacement of the 2â-carbomethoxy
functionality of 3â-(p-substituted-phenyl)-N-methyl-tro-
pane-2â-carboxylic acid methyl esters with an isopropyl
ester affords analogues exhibiting a significant enhance-
ment in selectivity for the dopamine transporter in
comparison to norepinephrine (NET) and serotonin
(SERT) transporters.29 The 4-chloro derivative (CTC-
The synthetic approach chosen for the preparation of
diastereomerically pure 2â-(R)-carbo-1′-fluoro-2-propoxy-
3â-(4-chlorophenyl)tropane ((R)-FIPCT) (R-6) and 2â-
(S)-carbo-1′-fluoro-2-propoxy-3â-(4-chlorophenyl)tro-
pane ((S)-FIPCT) (S-6) involved the use of the commer-

Ligands for the PET Imaging Of Dopamine Transporters
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4 641
Sch em e 2a
a
Reagents: (a) Ph3CCl, pyridine; (b) LiAlH4, THF; (c) Pd/C, H2, MeOH; (d) MsCl, NEt3, CH2Cl2; (e) 5, NEt3, CH2Cl2; (f) N(C4H9)4F,
THF.
Sch em e 3
cially available optically active C-3 synthons (S)-glycidol
(7) and (S)-1,2-propandiol (S-10), respectively. The use
of these chiral materials facilitated the construction of
either the R- or S- diastereochemical configurations of
6 to enable a study of the structure-activity relation-
ships on DAT binding for the (R)-FIPCT and (S)-FIPCT
diastereomers. In this synthetic approach, 2â-carbo-R-
1′-mesyloxy-2-propoxy-3â-(4-chlorophenyl)tropane (R-
12) and 2â-carbo-S-1′-mesyloxy-2-propoxy-3â-(4-chloro-
phenyl)tropane (S-12) were the key substrates employed
for selectively introducing fluorine onto the 1′-position
of the corresponding 2â-carbo-R-1′-fluoro-2-propoxy (R-
6) and 2â-carbo-S-1′-fluoro-2-propoxy (S-6) esters. The
key substrate R-12 was prepared from a six-step
sequence of reactions outlined in Scheme 2. (S)-Glycidol
(7) was treated with triphenylmethyl chloride to give
(S)-1-triphenylmethylglicydyl ether (8). Reduction of the
aryloxyglycidyl 8 with lithium aluminum hydride af-
forded (R)-1-triphenylmethoxy-2-propanol (9). Hydro-
genolysis of 9 by treatment with Pd/C and H2 gave (R)-
1,2-propandiol (R-10) in quantitative yield. Treatment
of the diol R-10 with methanesulfonyl chloride gave (R)-
1-mesyloxy-2-propandiol (R-11) which was converted to
2â-carbo-(R-1′-mesyloxy-2-propoxy)-3â-(4-chlorophenyl)-
tropane (R-12) by esterification of acid chloride 5. The
2â-carbo-S-1′-mesyloxy-2-propoxy-3â-(4-chlorophenyl)-
tropane (S-12) substrate was prepared in an analogous
manner in two steps from (S)-1,2-propandiol (S-10)
(Scheme 2).
determined by HPLC analysis using a radioactivity
detector. Semipreparative, reverse-phase HPLC puri-
fication of the epimeric mixture gave R,S-FIPCT (6) in
4.6% radiochemical yield, corrected for decay at end of
bombardment (EOB), in a synthesis time of 100 min
following bombardment and with a specific activity of
2 Ci/µmol as determined from the mass measured from
the HPLC UV analysis. The diastereomerically pure
fluorine-18 labeled R-FIPCT ([18F]R-6) and S-FIPCT
([18F]S-6) were prepared in an analogous manner from
their corresponding mesylates.
A shortcoming of the radiosynthesis of fluorine-18
R,S-FIPCT is the significant conversion to its 2R-epimer.
Radiochemical yields of the desired fluorine-18 2â-
epimer ranged from 1.4 to 4.6% at the end of synthesis
and were accompanied by an additional 2.7 to 6.5% of
the undesired 2R-epimer. If the conversion of the 2â- to
the 2R-epimer could be minimized, then the end of
synthesis radiochemical yield of fluorine-18 R,S-FIPCT
may be increased to 7%. It is feasible that the R,S-
FIPCT 2R-epimer resulted from either epimerization,
by K222/K2CO3, of the 2â-mesylate precursor prior to
radiofluorination or epimerization of the final radiola-
beled product. We investigated the preparation of
fluorine-18 labeled R,S-FIPCT by reducing the quantity
of K222/K2CO3. We ran triplicate experiments in which
0.5 equiv of K222/K2CO3 was used. The reduced K222/
K2CO3 preparations produced R,S-FIPCT in 4.9% (n )
3) radiochemical yield at end of bombardment and
resulted in less conversion, 0.17% (n ) 3), of the 2R-
epimer.
The pivotal step in the syntheses of 2â-(R)-carbo-1′-
fluoro-2-propoxy-3â-(4-chlorophenyl)tropane ((R)-FIPCT)
(R-6) and 2â-(S)-carbo-1′-fluoro-2-propoxy-3â-(4-chlo-
rophenyl)tropane ((S)-FIPCT) (S-6) involved introduc-
tion of fluoride into the 2â-carbo-2-propoxy group by
treatment with tetrabutylammonium fluoride. The 2â-
(R)-carbo-1′-fluoro-2-propoxy esters R-6 and S-6 were
purified by flash chromatography followed by crystal-
lization. HPLC analysis (C18) of R-6 and S-6 showed the
presence of only one diastereomer.
Biologica l Resu lts
The in vitro affinities of R-FIPCT and S-FIPCT for
the DAT were determined by competition in DAT
radioligand binding assays using [3H]-WIN 35,428 bind-
ing in homogenates of murine kidney cells stably
expressing the human DAT and are shown in Table 1.
The rank order of affinities was GBR 12909 > CIT >
S-FIPCT > CCT-iPr > R-FIPCT. R-FIPCT had 6 times
Ra d ioch em istr y
Fluorine-18 labeled R,S-FIPCT ([18F]6) was prepared,
shown in Scheme 3, by treatment of the corresponding
mesylate with potassium[18F]fluoride and kryptofix
K222 in acetonitrile at 100 °C to give a mixture of R,S-
FIPCT ([18F]6) and the 2R-epimer in a 4:6 ratio as

642 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4
Xing et al.
Ta ble 1. Inhibition Constants (Ki, nM) of Various Ligands for
Monoamine Transporters
Ta ble 2. Distribution of Radioactivity (Percent Injected Dose
per Gram of Tissue) in Rats after iv Administration of
R,S-[18F]FIPCTa
human
5-HT/DA
rat
time after injection
5-HT/DA
ratio
ligand
DAa 5-HTb
0.48 0.67
0.9 427
3.2 64
0.67 85
ratio
DAc 5-HTc NEc
tissue
blood
2 min
0.06
30 min
60 min
120 minb
0.01
CIT
1.4
474
20
0.01
0.01
CCT-iPr
R-FIPCT
S-FIPCT
(10.05-0.07) (0.01-0.01) (0.01-0.01) (0.01-0.01)
14.7
133 498
6.06 137 690
9
23
heart
lungs
liver
0.76 0.06 0.05 0.06
127
(0.64-0.93) (0.05-0.07) (0.04-0.05) (0.04-0.08)
GBR 12909 0.36
fluvoxamine
2.57
(2.0-3.75)
0.74
0.35
(0.29-0.38) (0.15-0.21) (0.18-0.34)
2.06 2.23 2.09
0.18
0.23
3.08
a
Competitive binding versus [N-methyl-3H]-WIN 35,428 in
(0.69-0.82) (1.82-2.37) (2.13-2.42) (1.64-2.45)
0.74 0.22 0.14 0.14
(0.68-0.85) (0.2-0.26)
1.49 0.18
(1.13-1.94) (0.17-0.18) (0.1-0.11)
0.05 0.05 0.04
murine kidney cells transfected with human dopamine trans-
spleen
kidneys
muscle
testis
bone
b
porter. Competitive binding versus [3H]citalopram in murine
(0.14-0.15) (0.1-0.18)
kidney cells transfected with human serotonin transporter. c Sam-
ples (2-12 nM) of [N-methyl-3H]-WIN 35,428, [3H]citalopram, or
[3H]desmethylipramine were incubated in the presence of 7-11
concentrations of the indicated ligand and membranes prepared
from rat striatum or cortex. The assays were performed by
NOVASCREEN in participation with the NIMH/NOVASCREEN
Psychotherapeutic Drug Discovery Development Program.
0.11
0.12
(0.09-0.16)
0.04
(0.04-0.05) (0.04-0.06)b (0.02-0.05) (0.03-0.05)
0.07
0.09
0.09
0.08
(0.06-0.07) (0.07-0.1)
(0.07-0.1)
0.07
(0.08-0.09)
0.1
0.13
0.08
(0.1-0.15)
(0.07-0.08) (0.06-0.07) (0.01-0.13)
lower affinity than CIT, for the human transfected DAT.
In comparison to S-FIPCT and CCT-iPr, R-FIPCT had
4 and 3 times, respectively, lower affinity for the
transfected human DAT. Similarly, in comparison to
S-FIPCT, R-FIPCT had 2.5 times lower affinity for DAT
binding in rat striatal homogenates. Competition bind-
ing site analyses were also used to evaluate the com-
parative affinity of R-FIPCT and S-FIPCT for the
human SERT and NET labeled by [3H]citalopram and
[3H]desmethylipramine, respectively (Table 1). The af-
finity of R-FIPCT and S-FIPCT for the human SERT
was 20- and 127-fold lower, respectively, than for DAT.
A lower selectivity for R-FIPCT and S-FIPCT for the
DAT vs the SERT, 9 and 23 respectively, was noted in
binding studies using rat cortical homogenates (Table
1). These results indicate that R-FIPCT and S-FIPCT
exhibit low affinity for the rat NET and selective affinity
for human and rat DAT vs SERT and that S-FIPCT
exhibited both higher affinity and selectivity for the
human and rat DAT than R-FIPCT.
a
Mean and range (in parentheses) values for four male Spra-
b
gue-Dawley rats. Mean and range (in parentheses) values for
three male Sprague-Dawley rats.
Ta ble 3. Distribution of Radioactivity (Percent Injected Dose
per Gram of Tissue) in Rat Brain Regions after iv
Administration of R-[18F]FIPCTa
time after injection
region
5 min
3.97
30 min
60 min
120 min
1.20
striatum
4.50
2.28
(3.44-4.80) (3.57-6.08) (2.07-2.83) (1.07-1.22)
2.76 2.04 0.77 0.40
(2.47-3.09) (1.67-2.66) (0.51-1.09) (0.22-0.33)
1.42 0.99 0.38 0.22
(1.26-1.62) (0.75-1.23) (0.32-0.48) (0.19-0.25)
rest of brain 1.36 1.03 0.45 0.27
(1.19-1.53) (0.85-1.21) (0.36-0.52) (0.22-0.33)
2.79 4.55 6.0 5.45
(2.49-3.81) (3.83-4.94) (4.43-6.93) (4.64-6.14)
1.44 2.21 2.96 3.0
(1.32-1.94) (1.67-2.85) (2.59-4.06) (2.61-3.24)
cortex (CX)
cerebellum
S/Cereb
S/CX
a
Mean and range (in parentheses) values for four male
The diastereomeric mixture and the R- and S-isomers
of FIPCT were labeled with fluorine-18, and the distri-
bution of radioactivity expressed as percent dose per
gram in tissues of male rats is shown in Table 2. The
accumulation of radioactivity in the bone was initially
low and exhibited no increase from 2 min (0.13% dose/
g) to 2 h (0.1% dose/g), which demonstrated the expected
stability of the fluoroisopropyl group to significant in
vivo defluorination.
The individual R- and S-diastereomers of FIPCT were
labeled with flourine-18, and the regional distribution
study in the brain of male Sprague Dawley rats was
performed over 2 h. The results shown in Tables 3 and
4 demonstrate that [18F]-R-6 (Table 3) showed high
initial striatal uptake and moderate retention. The
striatum uptake reached a maximum at 30 min (4.50%
dose/g). After 120 min, the striatum uptake decreased
73% when compared to the peak activity at 30 min. The
S/Cereb and S/CX ratios reached a maximum 6.0 and
3.0, respectively, at 60 min. The high uptake of fluorine-
18 in regions of the brain expressing DATs and high
S/Cereb ratio at 60 min and the 73% washout in the
striatum over 120 min demonstrate that [18F]-R-6 has
potential favorable kinetic properties for in vivo quan-
titation and imaging of the DAT by PET.
Sprague-Dawley rats.
Ta ble 4. Distribution of Radioactivity (Percent Injected Dose
per Gram of Tissue) in Rat Brain Regions after iv
Administration of S-[18F]FIPCTa
time after injection
region
5 min
2.47
(1.89-3.73) (2.7-3.03) (2.13-2.89) (1.19-3.39)
cortex (CX) 1.16 1.07 0.45 0.29
(0.92-1.36) (0.92-1.19) (0.36-0.51) (0.21-0.39)
cerebellum 0.77 0.59 0.22 0.14
(0.68-0.89) (0.52-0.73) (0.17-0.27) (0.12-0.17)
rest of brain 0.83 0.69 0.33 0.21
(0.63-0.98) (0.62-0.80) (0.24-0.39) (0.16-0.25)
3.21 4.78 11.82 12.7
(2.12-4.65) (4.2-5.29) (10.37-12.54) (8.04-20.19)
2.13 2.64 5.78 6.14
(1.70-2.74) (2.31-2.93) (4.19-5.82) (4.25-8.69)
30 minb
60 minb
2.6
120 min
striatum
2.82
1.78
S/Cereb
S/CX
a
Mean and range (in parentheses) values for four male
b
Sprague-Dawley rats. Mean and range (in parentheses) values
for three male Sprague-Dawley rats.
The S-diastereomer of FIPCT, [18F]-S-6 (Table 4),
showed higher striatal retention and S/Cereb and S/CX
ratios in comparison to the R-diastereomer. In addition,
in contrast with [18F]-R-6, the [18F]-S-6 striatum uptake

Ligands for the PET Imaging Of Dopamine Transporters
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4 643
F igu r e 1. Trans-axial image for a single plane following 5.69
mCi of [18F](S)-FIPCT (S-6) summed for 105-125 min in a
male rhesus monkey.
F igu r e 3. Time-activity curves for brain regions for a rhesus
monkey after receiving 5.69 mCi of [18F](S)-FIPCT (S-6). Serial
images were acquired for a total time of 125 min (i.e., 18 scans).
F igu r e 4. Trans-axial image for a single plane following 2.95
mCi of [18F](S)-FIPCT (R-6) summed for 105-125 min in a
male rhesus monkey previously given a left intracarotid
infusion of MPTP.
F igu r e 2. Time-activity curves for brain regions for a rhesus
monkey after receiving 3.33 mCi of [18F](R)-FIPCT (R-6). Serial
images were acquired for a total time of 125 min (i.e., 18 scans).
R-6 and [18F]-S-6 in the monkeys (Figure 2 and Figure
3) were consistent with the results found in the rats.
The studies performed in rats demonstrated that [18F]-
R-6 cleared more rapidly than [18F]-S-6 from the stria-
tum. In the PET imaging study with [18F]-R-6 in the
rhesus monkey, the putamen and caudate achieved a
transient equilibrium at 75 min post injection, whereas
the decay-corrected uptake of [18F]-S-6 continued to
increase at 115 min post injection. Following adminis-
tration of [18F]-R-6, the putamen exhibited the highest
uptake in the brain of the monkey and attained maxi-
mum (0.08% dose/g) uptake at 75 min. The uptake
percentage of [18F]-R-6 in the cerebellum (CB) and
cortex (CX) exhibited a maximum value of 0.06% dose/g
at 10. 5 min and 0.03% dose/g at 4.5 min post injection,
respectively. The putamen-to-cerebellum and putamen-
to-cortex exhibited ratios ranged from 3.4 to 3.6 at 115
min. Caudate radioactivity indicated a peak uptake of
0.06% dose/g at 75 min and a caudate-to-cerebellum and
caudate-to-cortex ratio of 2.5-2.6 at 115 min. The [18F]-
S-6 diastereomer, which showed prolonged retention in
the striatum, exhibited 0.06% dose/g and 0.05% dose/g
in putamen and caudate, respectively, at 115 min. The
putamen-to-cerebellum and caudate-to-cerebellum ra-
tios were 2.5-2.6 at 115 min.
decreased only 37% when compared to the peak activity
at 30 min. The S/Cereb and S/CX ratios of [18F]-S-6
exhibited a pronounced increase with time. The S/Cereb
and S/CX ratios at 120 min were 12.7 and 6.14,
respectively. The higher striatal uptake and retention
of the S-diastereomer in comparison to the R-diastere-
omer are consistent with the results of the in vitro
competitive binding data which show that the S-
diastereomer exhibits a 2.5- to 5-fold higher affinity for
the DAT. These results indicate that [18F]-S-6 may also
have very good potential for imaging the DAT in the
striatum and extrastriatal sites by PET.
In Vivo Non h u m a n P r im a te Im a gin g
PET imaging studies were performed with [18F]-R-6
and [18F]-S-6 to compare and determine time-activity
curves for brain regions-of-interest in rhesus monkeys
at 0-115 min post injection. The distribution of fluorine-
18 in the brain following administration of [18F]-R-6 and
[18F]-S-6 was characteristic of binding to DAT sites. The
highest uptake occurred in the caudate and putamen,
with the lowest uptake occurring in the cerebellum and
cortex. (Figure 1).
Time-activity curves of the caudate and putamen
exhibiting the kinetics of the two diastereomers [18F]-

644 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4
Xing et al.
The in vivo binding of [18F]-S-6 to the DAT in the
caudate and putamen of a rhesus monkey was demon-
strated further by the absence of striatal radioactivity
in the left hemisphere of a monkey which previously
received a left intracarotid infusion of MPTP. The
uptake of radioactivity in the caudate and putamen was
only present in the right hemisphere of the rhesus
monkey brain at 115 min following injection of [18F]-S-
6. The putamen-to-cerebellum and caudate-to-cerebel-
lum ratios in the right hemisphere were 3.4-3.5 at 115
min while the putamen-to-cerebellum and caudate-to-
cerebellum ratios in the left hemisphere were 1.2-1.4
at 115 min.
retention within the striatum after 115 min which will
afford higher signal-to-background ratios than [18F]-R-
6 after 115 min.
Exp er im en ta l Section
All chemicals and solvents were analytical grade and were
used without purification. The fluorine-18 fluoride was ob-
tained from 95% enriched oxygen-18 water (Enritiech) by
proton irradiation with a Seimens RDS 112, 11 MeV cyclotron.
The melting points (mp) were determined in capillary tubes
using an Electrothermal apparatus and are uncorrected. The
thin-layer chromatographic analyses (TLC) were performed
using 250 µm thick layers of silica gel G PF-254 coated on
aluminum plates (Whatman). The proton nuclear magnetic
resonance (NMR) spectra were obtained at 300 or 400 MHz
with a Varian instrument. Carbon-13 nuclear magnetic reso-
nance spectra were obtained at 300 or 400 MHz with a Varian
instrument. Elemental analyses were performed by Atlantic
Microlab, Inc. (Norcross, GA) and, unless noted otherwise,
were within (0.4% of the calculated values. High-resolution
voltage scans were performed at 70 EV with reference scans
over narrow mass, •0.0006. Analyses were within 0.002 of
theoretical values. Mass spectra (MS) were determined on a
ZAB-EQ (VG Analytical) hybrid high-resolution, double-focus-
ing mass spectrometer with collision and analyzing quadro-
pole.
Meta bolite An a lysis
Arterial plasma samples from a rhesus monkey fol-
lowing femoral vein injection of S-[18F]FIPCT and
R-[18F]FIPCT were analyzed for nonpolar, potentially
brain permeable metabolites by a solvent extraction and
TLC method. The major radioactive component appear-
ing in the initial arterial plasma samples was ether
extractable, displayed a single peak (>98% pure) on
TLC, and corresponded to unmetabolized authentic
S-[18F]FIPCT. The fraction of plasma radioactivity cor-
responding to unmetabolized S-[18F]FIPCT rapidly de-
creased from 86% at 5 min to 30% at 30 min. The major
radiolabeled metabolite found in arterial plasma was a
polar nonextractable component, which increased to
89% of the plasma activity by 120 min. The identity of
the polar metabolite was not elucidated due to its
improbable brain permeability. The metabolite profile
for R-[18F]FIPCT demonstrated that this diastereomer
metabolized more rapidly. The fraction of plasma ra-
dioactivity corresponding to unmetabolized R-[18F]-
FIPCT rapidly decreased from 84% at 3 min to 25% at
14 min. The major radiolabeled metabolite found in
arterial plasma was a polar nonextractable component,
which increased to 95% of the plasma activity by 120
min. Thus, there was no detectable formation of lipo-
philic radiolabeled metabolites from either diastereomer
that was capable of entering the brain. The absence of
labeled lipophilic metabolites in the arterial plasma that
could reenter the brain permits the metabolism uncor-
rected calculation of the S-[18F]FIPCT and R-[18F]FIPCT
input function and subsequent quantification of binding
sites by tracer kinetic modeling.
All animal experiments were carried out according to
protocols approved by the Institutional University Animal
Care Committee (IUCAC) and Radiation Safety Committees
of Emory University.
Ch em istr y. (R,S)-1-F lu or o-2-p r op a n ol (2). A solution of
fluoroacetone (1.0 g, 13 mmol) in anhydrous ethyl ether (5 mL)
was added dropwise to a stirred suspension of LiAlH4 (131 mg,
3.25 mmol) in 15 mL of anhydrous ethyl ether at -70 °C over
a period of 1 min. The reaction mixture was allowed to warm
to room temperature. The resulting mixture was stirred at 40
°C for 10 min, then cooled to room temperature, and acidified
to pH 3 with 10% H2SO4 solution. The aqueous phase was
separated and extracted with ether (4 × 15 mL). The combined
organic phase was washed with saturated NaHCO3 and brine,
dried over anhydrous MgSO4, and concentrated in vacuo to
give 2 (460 mg, 46%) as a colorless liquid, bp 105 °C. The
product was used without further purification: 1H NMR
(CDCl3) δ 1.18 (dd, J ) 1.5, 3.6 Hz, 3 H), 3.45 (s, 1 H), 3.98-
4.12 (m,1H), 4.24 (ddd, J ) 6.9, 9.3, 48 Hz, 1 H), 4.35 (ddd, J
) 3.0, 9.3, 47 Hz, 1 H).
N-Met h yl-3â(4-ch lor op h en yl)t r op a n e-2â-ca r b oxylic
Acid (4). A solution of 3 (200 mg, 0.68 mmol) in H2O/dioxane
(1:1, 8 mL) was refluxed for 3 days. After the reaction, the
solvents were removed and the residue was purified via flash
column chromatography (eluted employing a gradient with
20% MeOH in CH2Cl2 (0.1% Et3N) to 50% MeOH in CH2Cl2
(0.1% Et3N)) to afford 4 as a colorless microcrystalline solid
(180 mg, 95%): 1H NMR (D2O) δ 1.75-1.85 (m, 1 H), 1.95-
2.13 (m, 2 H), 2.20-2.40 (m, 2 H), 2.56-2.78 (m, 2 H), 2.69 (s,
3 H), 3.27-3.40 (1 H), 3.82-3.95 (m, 2 H), 7.16 (AB, J ) 8.4
Hz, 1 H), 7.26 (AB, J ) 8.4 Hz, 1 H).
Con clu sion s
The two diastereomers of FIPCT (6) have been
synthesized and radiolabeled with fluorine-18 by a one-
step method in a synthesis time of 85 min including
HPLC purification. Competition binding assays in cells
stably expressing the human monoamine transporters
demonstrated that S-6 and R-6 have a high affinity for
the DAT with S-6 showing the highest selectivity, 127-
fold higher for the DAT over the SERT. Biodistribution
studies in rats showed that both [18F]-S-6 and [18F]-R-
6 exhibited high uptake in the striatum, a region rich
in DAT sites. PET imaging studies in rhesus monkeys
demonstrated that [18F]-R-6 achieved a transient equi-
librium in the striatum in less than 90 min and is an
attractive candidate for in vivo quantitation of DAT sites
by PET. In contrast, [18F]-S-6 exhibited potential for
mapping extrastriatal DAT sites due to prolonged
N-Met h yl 2â-ca r b o-(R,S)-1-flu or o-2-isop r op a n oxy-3â-
(4-ch lor op h en yl)t r op a n e (R,S-6). A suspension of 4 (115
mg, 0.41 mmol) in POCl3 (0.8 mL) was stirred under argon at
room temperature for 4 h. Then excess POCl3 was removed
under reduced pressure. Toluene (3 mL) was added and then
removed under reduced pressure to dryness. The freshly
prepared R,S-2 (190 mg, 1.23 mmol) in anhydrous CH2Cl2 (2.5
mL) was added, followed by Et3N (0.2 mL) under argon at 0
°C. The resulting mixture was stirred at 10 °C overnight. H2O
(10 mL) was added, the resulting solution was basified with
NH4OH, extracted with CH2Cl2 (4 × 10 mL), dried (MgSO4),
and filtered, and the filtrate was concentrated in vacuo. The
resulting oil was purified via flash column chromatography
(eluted gradually from Et2O to 1% Et3N in Et2O) to afford R-12
as a colorless microcrystalline solid (141 mg, 83%): mp 109-
1
110 °C; H NMR (CDCl3) δ 1.12 (d, J ) 6.6 Hz, 3 H), 1.55-
1.76 (m, 4 H), 2.02-2.28 (m, 1 H), 2.19 (s, 3 H), 2.51 (dt, J )

Ligands for the PET Imaging Of Dopamine Transporters
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4 645
2.7, 12.3 Hz, 1 H), 2.83 (s, 3 H), 2.86-3.02 (m, 2 H), 3.32-
3.40 (m, 1 H), 3.54-3.60 (m, 1 H), 4.00-4.15 (m, 2 H), 4.98-
5.10 (m, 1 H), 7.16-7.28 (m, 4 H).
NMR (CDCl3) δ 1.12 (d J ) 6.6 Hz, 3 H), 1.55-1.76 (m, 4 H),
2.02-2.28 (m, 1 H), 2.19 (s, 3 H), 2.51 (dt, J ) 2.7, 12.3 Hz, 1
H), 2.83 (s, 3 H), 2.86-3.02 (m, 2 H), 3.32-3.40 (m, 1 H), 3.54-
3.60 (m, 1 H), 4.00-4.15 (m, 2 H), 4.98-5.10 (m, 1 H), 7.16-
7.28 (m, 4 H); HRMS (EI) Calcd for C19H26ClNO5S: 415.1220.
Found: 415.1228.
(S)-1-Tr ip h en ylm eth yl Glycid yl Eth er (8). A solution of
(S)-glycidol (7, 5 g, 64.8 mmol) in pyridine (16 mL) was cooled
to 0 °C under argon. Then triphenylmethyl chloride (18.06 g,
64.8 mmol) was added. The resulting mixture was allowed to
warm to room temperature and stirred overnight. The result-
ing mixture was washed with saturated aqueous CuSO4
solution, extracted with EtOAc, dried (MgSO4), and filtered.
The filtrate was concentrated in vacuo to give an oil that was
purified via flash column chromatography (1% EtOAc in
hexane) to afford a colorless microcrystal solid (8, 6.5 g, 32%):
1H NMR (CDCl3) δ 2.65-2.70 (m, 1 H), 2.79-2.85 (m, 1 H),
3.15-3.25 (m, 2 H), 3.35-3.40 (m, 1 H), 7.25-7.58 (m, 15 H).
(R)-1-Tr ip h en ylm eth oxy-2-p r op a n ol (9). A solution of 8
(3 g, 9.48 mmol) in anhydrous THF (12 mL) was cooled to 0
°C under argon. LiAlH4 (190 mg, 19 mmol, 95%) was added,
and the resulting mixture was stirred continuously overnight.
The reaction was quenched with H2O, followed by 6 N NaOH.
The suspension was filtered, the filtrate was extracted with
CH2Cl2 (4 × 30 mL), dried (MgSO4), and filtered, and the
filtrate was concentrated in vacuo. The resulting oil was
purified via flash column chromatography (1% EtOAc in
hexane) to afford a colorless microcrystalline solid (8, 1.93 g,
64%): 1H NMR (CDCl3) δ 0.98 (d, J ) 6.3 Hz, 3 H), 2.362 (d,
J ) 3 Hz, 1 H for OH), 2.93 (dd, J ) 16.8, 9.0 Hz, 1 H), 3.02
(dd, J ) 9.3, 3.4 Hz, 1 H), 3.80 (m, 1 H), 7.10-7.45 (m, 15 H).
(R)-1,2-P r op a n ed iol (R-10). To a solution of 9 (3.58 g, 21.5
mmol) in methanol (25 mL) was added Pd/C (800 mg) and H2.
The resulting mixture was stirred at room temperature
overnight. The resulting suspension was filtered, and the
residue was washed with CH2Cl2 (2 × 2 mL). The combined
filtrates were concentrated in vacuo, and the product (R-10)
was distilled out under reduced pressure via Kujelrohr to
afford a clear oil (1.64 g, 100%): 1H NMR (CDCl3) δ 1.04 (d,
J ) 6.6 Hz, 3 H), 3.28 (dd, J ) 19.2, 11.2 Hz, 1 H), 3.48 (dd,
J ) 2.7 Hz, 1 H), 3.72-3.85 (m, 1 H), 4.44 (s, 2 H for OHs).
(R)-2-Hyd r oxy-p r op yl Meth a n esu lfa te (R-11). A solution
of R-10 (180 mg, 2.37 mmol) in anhydrous CH2Cl2 (2 mL) and
Et3N (0.24 mL) was cooled to 0 °C under argon, and then
methanesulfonyl chloride (271 mg, 2.37 mmol) was added
dropwise. After the addition, the resulting mixture was allowed
to warm to room temperature over a period of 2 h. Ice water
(5 mL) was added, the product was extracted with CH2Cl2
(4 × 10 mL), dried (MgSO4), and filtered, and the filtrate was
concentrated in vacuo. The resulting oil was purified via flash
column chromatography (5% Et2O in CH2Cl2, 0.03% Et3N) to
afford R-11 as a colorless oil (230 mg, 57%): 1H NMR (CDCl3)
δ 1.11 (d, J ) 6.0 Hz, 3 H), 2.96 (s, 3 H), 3.92-4.12 (m, 3 H).
N -Me t h y l-2â-c a r b o -(R )-1′-flu o r o -2-p r o p o x y -3â-(4-
ch lor op h en yl)tr op a n e (R-6). To a solution of R-12 (18 mg,
0.043 mmol) in 1:1 Et2O/THF (2 mL) was added a solution of
1 M Bu4N+F- in THF (0.05 mL) under argon. The resulting
mixture was refluxed for 7 h. After the reaction, the solvents
were removed, and the residue was purified via flash column
chromatography (eluted gradually from Et2O to 0.5% Et3N in
Et2O) to afford R-6 as a colorless microcrystalline solid (8 mg,
56%): mp 98-99 °C; 1H NMR (CDCl3) δ 1.08 (dd, J ) 0.9, 6.6
Hz, 3 H), 1.48-1.75 (m, 4 H), 2.00-2.25 (m, 1 H), 2.16 (s, 3
H), 2.41 (dt, J ) 3.0, 12.3 Hz, 1 H), 2.75-2.90 (m, 2 H), 3.25-
3.40 (m, 1 H), 3.50-3.60 (m, 1 H), 4.10 (d, J ) 3.9 Hz, 1 H),
4.26 (d, J ) 3.6 Hz, 1 H), 4.80-5.00 (m, 1 H), 7.14-7.25 (m, 4
H); HRMS (EI) Calcd for C18H23ClFNO2: 339.1401. Found:
339.1406.
N -Me t h y l-2â-c a r b o -(S )-1′-m e s y l-2-p r o p o x y -3â-(4-
ch lor op h en yl)tr op a n e (S-12). A suspension of 4 (100 mg,
0.357 mmol) in POCl3 (0.8 mL) were allowed to react as
described for the preparation of R-12. The resulting acid
chloride 5 and mesylate S-11 (180 mg, 1.17 mmol) in anhy-
drous CH2Cl2 (2.5 mL) were reacted and worked up as
described above for the preparation of R-12. The resulting oil
was purified via flash column chromatography (eluted gradu-
ally from Et2O to 1% Et3N in Et2O) to afford S-12 as a colorless
microcrystalline solid (124 mg, 84%): mp 91.0-91.5 °C; 1H
NMR (CDCl3) δ 1.04 (d, J ) 6.6 Hz, 3 H), 1.47-1.72 (m, 4 H),
1.96-2.20 (m, 1 H), 2.15 (s, 3 H), 2.44 (dt, J ) 2.6, 12.5 Hz, 1
H), 2.78-2.98 (m, 2 H), 2.94 (s, 3 H), 3.24-3.34 (m, 1 H), 3.50-
3.56 (m, 1 H), 4.00 (dd, J ) 10.8, 5.4 Hz, 1 H), 4.10 (dd, J )
10.8, 3.6 Hz, 1 H), 4.90-5.00 (m, 1 H), 7.07-7.22 (m, 4 H);
13C NMR (CDCl3) d 16.04 (q), 25.13 (t), 25.83 (t), 33.12 (t), 33.86
(d), 37.61 (q), 41.78 (d), 52.51 (q), 62.10 (d), 65.05 (d), 66.77
(d), 70.59 (t), 128.0 (d), 128.6 (d), 131.5 (s), 141.5 (s), 170.5
(s); HRMS (EI) Calcd for C19H26ClNO5S: 415.1220. Found:
415.1228.
N -Me t h y l-2â-c a r b o -(S )-1′-flu o r o -2-p r o p o x y -3â-(4-
ch lor op h en yl)tr op a n e (S-6). To a solution of S-12 (120 mg,
0.29 mmol) in THF (5 mL) was added a solution of 1 M
Bu4N+F- in THF (0.33 mL) under argon. The resulting mixture
was refluxed for 7 h. After the reaction, the solvent was
removed and the residue was purified via flash column
chromatography (eluted gradually from Et2O to 0.5% Et3N in
Et2O) to afford S-6 as a colorless microcrystalline solid (32 mg,
1
33%): mp 102.5-103.5 °C; H NMR (CDCl3) δ 1.03 (dd, J )
0.9, 6.6 Hz, 3 H), 1.48-1.70 (m, 4 H), 1.95-2.25 (m, 1 H), 2.14
(s, 3 H), 2.48 (dt, J ) 3.0, 12.5 Hz, 1 H), 2.80-2.95 (m, 2 H),
3.26-3.32 (m, 1 H), 3.50-3.57 (m, 1 H), 4.06-4.38 (m, 2 H),
4.81-5.00 (m, 1 H), 7.10-7.20 (m, 4 H); 13C NMR (CDCl3) d
15.05 (q, d, J ) 7.0 Hz), 25.18 (t), 25.71 (t), 33.16 (t), 33.85
(d), 41.73 (d), 52.53 (q), 62.10 (d), 65.23 (d), 67.96 (d, d, J )
19.9 Hz), 84.44 (t, d, J ) 172.6 Hz), 127.9 (d), 128.5 (d), 131.4
(s), 141.6 (s), 170.6 (s); Anal. Calcd for C18H23ClFNO2: C, 63.62;
H, 6.82; N, 4.12. Found: C, 63.55; H, 6.83; N, 4.09.
(S)-2-Hyd r oxy-p r op yl Meth a n esu lfa te (S-11). A solution
of S-10 (180 mg, 2.37 mmol) in anhydrous CH2Cl2 (2 mL) and
Et3N (0.24 mL) was cooled to 0 °C under argon and then
allowed to react with methanesulfonyl chloride (271 mg, 2.37
mmol) as described for R-11. The resulting oil was purified
via flash column chromatography (5% Et2O in CH2Cl2, 0.03%
Et3N) to afford S-11 as a colorless oil (230 mg, 57%): 1H NMR
(CDCl3) δ 1.11 (d, J ) 6.0 Hz, 3 H), 2.96 (s, 3 H), 3.92-4.12
(m, 3 H).
F IP CT Ra d iola belin g. Approximately 280 µL of 95%
enriched [18O]water containing 430 mCi NCA [18F]fluoride
(Seimens RDS 112) was added to an 11 mL glass reaction
vessel containing 1 mL of a solution of 5 mg of Kryptofix (K-
222) and 0.5 mg of potassium carbonate in 0.05 mL of water
and 0.95 mL of CH3CN. The solution was heated at 118 °C for
3.5 min after which three additional portions of 1 mL of CH3-
CN were added and evaporated to dry the fluoride. The vial
was cooled to room temperature, and 3 mg of 2â-carbo-(R,S-
1′-mesyloxy-2-propoxy)-3â-(4-chlorophenyl)tropane (R,S-12) dis-
solved in 1.0 mL of CH3CN was added. The solution was
heated to 100 °C for 10 min, cooled to room temperature,
diluted with 4 mL of ether, and passed through a Waters
classic SiO2 sep-pak into a 10 mL maxi-vial attached to a 50
mL round-bottomed flask. The sep-pak was rinsed with 4 mL
N -Me t h y l-2â-c a r b o -(R )-1′-m e s y l-2-p r o p o x y -3â-(4-
ch lor op h en yl)tr op a n e (R-12). A suspension of 4 (115 mg,
0.41 mmol) in POCl3 (0.8 mL) was stirred at room temperature
for 4 h. The POCl3 was removed under reduced pressure.
Toluene (3 mL) was added and then removed under reduced
pressure to dryness. The freshly prepared R-11 (190 mg, 1.23
mmol) in anhydrous CH2Cl2 (2.5 mL) was added, followed by
Et3N (0.2 mL) under argon at 0 °C. The resulting mixture
stirred at 10 °C overnight. H2O (10 mL) was added, the
resulting solution was basified with NH4OH, extracted with
CH2Cl2 (4 × 10 mL), dried (MgSO4), and filtered, and the
filtrate was concentrated in vacuo. The resulting oil was
purified via flash column chromatography (eluted gradually
from Et2O to 1% Et3N in Et2O) to afford R-12 as a colorless
microcrystalline solid (141 mg, 83%): mp 109-110 °C; 1H

646 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4
Xing et al.
of Et2O, which was added to the maxi-vial, bringing the total
volume to 10 mL. The resulting ethereal solution was evapo-
rated by gentle heating and with a stream of argon gas. The
resulting residue was dissolved in 1 mL of 80/10/0.1% methanol/
water/triethylamine and injected onto a reverse-phase prep
high-pressure liquid chromatography (HPLC) column (Waters,
25 mm × 100 mm, flow rate 6 mL/min). The fraction eluting
at 14 min contained 0.18 mCi (0.07%, end of bombardment
(EOB)) of fluorine-18 labeled 2R-carbo-(R,S-1′-fluoro-2-pro-
poxy)-3â-(4-chlorophenyl)tropane. The fraction eluting at 19
min contained 15.19 mCi (6%, EOB) of the desired product
R,S-FIPCT ([18F]6), in a synthesis/purification time of 85 min.
Thin-layer chromatography (TLC) analysis of R,S-FIPCT ([18F]-
6) using a radioactivity detector (SiO2, 90:10 CH2Cl2:CH3OH,
Rf ) 0.42) and HPLC analysis (Waters C18, 8 mm × 200 mm
Novapak, 75/25/0.1% methanol/water/triethylamine, flow rate
1 mL/min, rt ) 5.38 min) using a UV detector and a radio-
activity detector showed these fractions to have a radiochemi-
cal purity of greater than 99%. Fractions containing the
greatest radioactivity were concentrated in vacuo, dissolved
in sterile saline with 10% ethanol, and filtered through an
Acrodisc 0.2 µm filter for in vivo studies.
An im a l Tissu e Distr ibu tion Exp er im en ts. The distribu-
tion of radioactivity was determined in tissues of male
Sprague-Dawley rats (225-300 g) after intravenous admin-
istration of the radioiodinated tropane. The animals were
allowed food and water ad libitum prior to the course of the
experiment. The radiofluorinated tropanes [18F]R,S-6, [18F]S-
6, and [18F]R-6 (50 µCi) in 0.2 mL of 13% EtOH in 0.9% NaCl
were injected directly into the tail vein of rats under ketamine
anesthesia. The animals were sacrificed at various time points
post injection by cardiac excision under ketamine anesthesia,
and the organs were excised, rinsed, and blotted dry. The
organs were weighed, and the radioactivity of the contents was
determined with a Packard γ automatic counter (Model Cobra).
The percent dose per organ was calculated by a comparison of
the tissue counts to suitably diluted aliquots of injected
material. Total activities of blood and muscle were calculated
under the assumption that they were 7% and 40% of the total
body weight, respectively.
Regional brain distribution was obtained in male Sprague-
Dawley rats (225-300 g) after intravenous administration of
[18F]R,S-6, [18F]S-6, and [18F]R-6. The cortex, striatum, and
cerebellum were dissected and placed in tared test tubes. The
test tubes were weighed, and the radioactivity of the contents
was determined with a γ automatic counter. The percent dose
per gram was calculated by a comparison of the tissue counts
with the counts of the diluted initial injected dose. The uptake
ratio of each brain region was obtained by dividing the percent/
gram of each region by that of the cerebellum.
Tissu e P r ep a r a tion . Male Sprague-Dawley rats (200-
250 g) were decapitated under ether anesthesia, and the brains
were excised and placed in ice. Striatal tissues were dissected,
pooled, and homogenized in 100 volumes (w/v) of ice cold Tris-
HCl buffer (50 mM, pH 7.4). The striatal homogenates were
centrifuged at 20000g for 20 min, and the resultant pellets
were rehomogenized in the same buffer and recentrifuged. The
final pellets were resuspended in a buffer composed of 50 mM
Tris buffer, pH 7.4; 120 mM NaCl; 5 mM KCl; 2 mM CaCl2;
and 1 mM MgCl2 and kept at -20° C for the binding assay
detailed below.
In Vitr o Bin d in g Exp er im en ts. Monoamine transporter
binding assays used cell membranes from either a dog kidney
(MDCK) cell line stably transfected with the human DAT
cDNA (gift of Dr. Gary Rudnick, Yale University) or a human
embryonic kidney cell line (HEK-293) stably transfected with
the human serotonin (hSERTcDNA (gift of Randy Blakely,
Ph.D., Vanderbilt University). Cells were grown to confluency
in DMEM containing 10% fetal bovine serum and Geneticin
sulfate and were harvested using 37° C phosphate-buffered
saline (PBS, pH 7.4) containing ethylenediaminetetracetic acid
(EDTA). Following centrifugation (2000g, 10 min), the super-
natants were decanted and the pellets homogenized with a
Polytron PT3000 (Brinkman, Littau, Switzerland, 11 000 rpm
for 12 s) in 30 volumes of PBS and centrifuged at 43 000g for
10 min. The supernatants were decanted and the resulting
pellets stored at -70° C until assayed.
Binding assays were carried out in 12 × 75 mm polystyrene
tubes in a 1000 µL volume consisting of 700 µL of assay buffer,
100 µL of competing ligand, 100 µL of [3H] ligand, and 100 µL
of membrane suspension. Prior to each assay, the cell lines
were characterized to determine a membrane concentration
that yielded optimal binding. Membrane suspensions were
prepared by resuspending the pellet in 5 mL of assay buffer
and centrifuging (400g, 10 min). The supernatant was de-
canted, and the resulting pellet resuspended in assay buffer
and briefly homogenized using a Polytron PT3000. Competing
ligands were assayed in triplicate at each of nine concentra-
tions (10-13 to 10-6 M). The ligands were first dissolved in
absolute ethanol at 10-3 M then serially diluted in 2.5 mM
HCl. A monoamine transporter-selective ligand of known
binding affinity (DAT, GBR 12909; hSERT, fluvoxamine) was
included as a reference in each assay. For DAT binding, the
assay buffer was 0.03 M phosphate buffer (pH 7.4) containing
0.32 M sucrose, the radioligand was [3H]-WIN 35,428 (Dupont
NEN, Boston, MA, 84.5 Ci/mmol, 2.0 nM final concentration),
and the equilibrium incubation time was 2 h at 4° C. For
hSERT binding, the assay buffer was 50 mM Tris, 120 mM
NaCl, 5 mM KCl (pH 7.9), the radioligand was [3H]citalopram
(Dupont NEN, Boston, MA, 85.7 Ci/mmol, 0.5 nM final
concentration), and the equilibrium incubation time was 1 h
at 22° C. All assays were initiated by the addition of membrane
suspension. Incubations were terminated by the addition of 2
mL of 0.03 M phosphate buffer (pH 7.4, 4° C) and rapid
vacuum filtration through GF/B filters (presoaked in phos-
phate buffer containing 0.3% polyethyleneimine) with four
washes (5 mL) of 0.03 M phosphate buffer (pH 7.4, 4° C). The
filters were then dried and placed in scintillation vials to which
6 mL of scintillation cocktail (Aquasol-2, Packard, Meriden,
CT) was added. The vials were shaken and radioactivity was
determined in a liquid scintillation counter at 66% efficiency.
The data from the competition binding curves were analyzed
and Ki values generated using GraphPad Prism software
(GraphPad Software, San Diego, CA).
Non h u m a n P r im a te Im a gin g. Quantitative brain images
were acquired in a male rhesus monkey weighing 12.1 kg and
a female rhesus monkey weighing 5 kg using a Seimens 951
31 slice PET imaging system. Images were reconstructed with
a Shepp-Logan filter (0.35 × Nyquist frequency) giving a
resolution of 8 mm fwhm. The animal was anesthetized,
initially, with Telazol (3 mg/kg; i.m.) and maintained on a 1%
isofluorane/5% oxygen mixture throughout the imaging pro-
cedure. The animals were intubated with assurance of ad-
equate patency of the airway and were placed on a ventilator
with arterial blood gases monitored throughout the study to
ensure physiologic levels of respiration. An arterial catheter
was placed in a distal leg artery of the animals. The animals
were placed in the tomograph, and the head was immobilized
with a thermoplastic (Tru Scan, MD) face mask. A transmis-
Bin d in g Assa ys. Tissue preparations (50 µL, 40-60 µg
protein) were incubated with appropriate amounts of [3H]
labeled ligand and competitors in a total volume of 0.2 mL of
the assay buffer. The assay mixture was incubated for 60 min
at room temperature with stirring, and the resulting samples
were rapidly filtered through Whatman GF/B glass-fiber filters
pretreated with 0.2% protamine base and washed with 3 × 5
mL of cold (4° C) 50 mM Tris-HCl buffer, pH 7.4. The
nonspecific binding was obtained in the presence of 10 mM
(-)-cocaine. The filters were counted in a γ counter (Beckman
5500) at an efficiency of 70%. Saturation binding, Scatchard,
and competition experiments were analyzed with the iterative
nonlinear least squares curve-fitting program.
sion scan was then obtained with
a gallium source for
attenuation correction of emission data. [18F]FIPCT samples,
5.69 mCi and 3.13 mCi, were injected into an antecubital vein
of the male and female, respectively, in a slow bolus infusion
over 30 s. Arterial blood sampling (0.5 mL samples) was
performed at approximately 10-12 s intervals with 12 samples

Ligands for the PET Imaging Of Dopamine Transporters
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4 647
(11) Kilbourn, M. R.; Haka, M. S. Synthesis of [18F]GBR 13119 A
Presynaptic Dopamine Uptake Antagonist. Appl. Radiat. Isot.
1988, 39, 279-282.
(12) Kilbourn, M. R. In Vivo Binding of [18F]GBR 13119 to the Brain
Dopamine Uptake System. Life Sci. 1988, 42, 1347-1353.
(13) Kilbourn, M. R.; Carey, J . E.; Koeppe, R. A.; Haka, M. S.;
Hutchins, G. A.; Sherman, P. S.; Kuhl, D. E. Biodistribution,
Dosimetry, Metabolism and Monkey PET Studies of [18F]GBR
13119. Imaging the Dopamine Uptake System In Vivo. Nucl.
Med. Biol. 1989, 16, 569-576.
(14) Fowler, J . S.; Volkow, N. D.; Wolf, A. P.; Dewey, S. L.; Schlyer,
D. J .; Macgregor, R. R.; Hitzemann, R.; Logan, J .; Bendriem,
B.; Gatley, S. J . Mapping Cocaine Binding Sites in Human and
Baboon Brain In Vivo. Synapse 1989, 4, 371-377.
(15) Logan, J .; Fowler, J . S.; Volkow, N. D.; Wolf, A. P.; Dewey, S.
L.; Schlyer, D. J .; Macgregor, R. R.; Hitzemann, R.; Bendriem,
B.; Gatley, S. J .; Christman, D. R. Graphical Analysis of
Reversible Radioligand Binding from Tome-Activity Measure-
ments Applied to [N-11C-Methyl]-(-)-Cocaine PET Studies in
Human Subjects. J . Cereb. Blood Flow Metab. 1990, 5, 740-
747.
(16) Ding, Y.; Fowler, J .; Volkow, N.; Logan, J .; Gatley, S.; Sugano,
Y. Carbon-11-d-threo-Methylphenidate Binding to Dopamine
Transporter in Baboon Brain. J . Nucl. Med. 1995, 36, 2298-
2305.
(17) Carroll, F. I.; Lewin, A. H.; Boja, J . W.; Kuhar, M. J .; Cocaine
Receptor: Biochemical Characterization and Structure-Activity
Relationships of Cocaine Analogues at the Dopamine Trans-
porter. J . Med. Chem. 1992, 35, 969-981.
(18) Frost, J . J .; Rosier, A. J .; Reich, S. G.; Smith, J . S.; Ehlers, M.
D.; Snyder, S. H.; Ravert H. T.; Dannals R. F. Positron Emission
Tomographic Imaging of the Dopamine Transporter with 11C-
WIN 35,428 Reveals Marked Declines in Mild Parkinson’s
Disease. Ann. Neurol. 1993, 34, 423-431.
(19) Meltzer, P. C.; Liang, A. Y.; Brownell, A. L.; Elmaleh D. R.;
Madras B. K. Substituted 3-phenyltropane Analogues of Co-
caine: Synthesis, Inhibition of Binding at Cocaine Recognition
Sites, and Positron Emission Tomography Imaging. J . Med.
Chem. 1993, 36, 855-862.
(20) Dannals, R. F.; Neumeyer, J . L.; Milius, R. A.; Ravert, H. T.;
Wilson, A. A.; Wagner, H. N., J r. Synthesis of a Radiotracer for
Studying Dopamine Uptake Site In Vivo Using PET: 2â-
carbomethoxy-3â-(p-fluorophenyl)-[N-11C-methyl]tropane ([11C]-
CFT or (11C-WIN 35,428). J . Lab. Comp. Radiopharm. 1993, 33,
147-152.
(21) Brownell, A. L.; Elmaleh, D. R.; Meltzer, P. C.; Shoup, T. M.;
Brownell, G. L.; Fischman A. J .; Madras B. K. Cocaine Conge-
ners as PET Imaging Probes for Dopamine Terminals. J . Nucl.
Med. 1996, 37, 1186-1192.
(22) Muller, L.; Halldin C.; Farde, L.; Karlson, P.; Hall, H.; Swahn,
C. G.; Neumeyer, J . L.; Wang, S.; Milius, R. A. [11C]â-CIT, a
Cocaine Analogue: Preparation, Autoradiography and Prelimi-
nary PET Investigations. Nucl. Med. Biol. 1993, 20, 249-255.
(23) Brown, D. J .; Luthra, S. K.; Brown, G. D.; Carrol. F. I.; Kuhar,
M. J .; Osman, S.; Waters, S. L.; Brady, F. [11C]RTI-121A
Potential Radioligand For PET Studies of the Dopamine Trans-
porter. J . Lab. Comp. Radiopharm. 1994, 35, 483.
(24) Lundkvist, C.; Halldin C.; Swahn, C. G.; Hall, H.; Karlson, P.;
Nakashima, Y.; Wang, S.; Milius, R. A.; Neumeyer, J . L.; Farde,
L. [O-Methyl-11C]â-CIT-FP, a Potential Radioligand for Quan-
titation of the Dopamine Transporter: Preparation, Autoradi-
ography, Metabolite Studies, and Positron Emission Tomography
Examinations. Nucl. Med. Biol. 1995, 22, 905-913.
obtained over the first 2 min. Arterial samples were also
obtained at 3, 4, 5, 7, 10, 20, 30, 60, 90, and 120 min after
tracer injection. Kinetic emission data were acquired either
using an 18 frame acquisition sequence which included six 30
s scans, four 3 min scans, five 10 min scans, and three 20 min
scans or using a 20 frame acquisition sequence which included
six 30 s scans, four 3 min scans, five 10 min scans, three 20
min scans, and two 30 min scans. Bilateral regions of interest
on the [18F]FIPCT images were drawn manually for the
caudate, putamen, frontal cortex, and cerebellum. The data
were displayed as both % dose/g and nCi/mL and normalized
for the quantity of injected dose and the weight of the monkey.
A left intracarotid infusion of MPTP was performed on the
female rhesus monkey by a previously reported method,30
several weeks prior to the PET study.
[
18F ]F IP CT Meta bolite An a lysis (Mon k ey). Arterial
plasma analysis of S-[18F]FIPCT metabolism was performed
in a rhesus monkey. Metabolite analysis was performed as
described for [18F]FPCT.28 [18F]FIPCT was injected as de-
scribed above, and arterial samples (5.0 mL) were collected
at approximately 2, 5, 15, 30, 60, and 120 min after tracer
injection. Plasma was prepared by centrifugation (3000g for
20 min), and nonpolar, brain permeable 18F labeled metabolites
were extracted with ethyl ether (2 × l mL). The ethyl ether
extracts of each plasma sample were evaporated to dryness
under nitrogen at 35 °C. The resulting residue was dissolved
in 200 µL of 90:10 CH2Cl2:CH3OH and analyzed by TLC (SiO2)
with a radioactivity detector.
Ack n ow led gm en t. This research was sponsored by
the Office of Health and Environmental Research, U.S.
Department of Energy, under Grant No. DE-FG05-
93ER61737 and the National Institute of Mental Health
Drug Screening Program. The authors thank Carolyn
K. Malcolm for assistance in preparing the manuscript,
Delicia Votaw, and Teresa Abak for assistance in the
data collection and analysis.
Refer en ces
(1) Niznik, H. B.; Fogel, E. F.; Fassos, E. F.; Seeman P. The
Dopamine Transporter is Absent in Parkinson’s Putamen and
Reduced in the Caudate Nucleus. J . Neurochem. 1991, 56, 192-
198.
(2) Kaufman, M. J .; Madras, B. K. Severe Depletion of Cocaine
Recognition Sites Associated with the Dopamine Transporter in
Parkinson’s Diseased Striatum. Synapse 1991, 49, 43-49.
(3) Bernheimer, H.; Birkmayer, W.; Hornykiewcz, O. Brain Dopa-
mine and the Syndromes of Parkinson and Huntington: Clinical,
Morphological and Neurochemical Correlations. J . Neurol. Sci.
1973, 20, 415-455.
(4) Haberland, N.; Hetey, L. J . Studies in Postmortem Dopamine
Uptake. II. Synaptosomal Catecholamine Uptake in Postmortem
Brain Regions in Schizophrenia. J . Neural. Transm. 1987, 68,
303-13.
(5) Reith, M. E. A.; Meisler, B. E.; Sershen, H.; Lajtha, A. Structural
Requirements For Cocaine Congeners to Interact with Dopamine
and Serotonin Uptake Sites in Mouse Brain and To Induce
Stereotyped Behavior. Biochem. Pharmacol. 1986, 35, 1123-
1129.
(6) Giros, B.; J aber, M.; J ones, S. R.; Wightman, R. M.; Caron, M.
G. Hyper-locomotion and Indifference to Cocaine and Amphet-
amine in Mice Lacking the Dopamine Transporter. Nature 1996,
379, 606-612.
(7) Ritz, M. C.; Lamb, R. J .; Goldberg, S. R.; Kuhar, M. J . Cocaine
Receptors on Dopamine Transporters are Related to Self-
administration of Cocaine. Science 1987, 237, 1219-1223.
(8) Volkow, N. D.; Wang, G. J .; Fischman, M. W.; Foltin, R. W.;
Fowler, J . S.; Abumrad, N. N.; Vitkun, S.; Logan, J .; Gatley, S.
J .; Pappas, N.; Hitzemann, R.; Shea, C. E. Relationship Between
Subjective Effects of Cocaine and Dopamine Transporter Oc-
cupancy. Nature 1997, 386, 827-830.
(9) Volkow, N. D.; Wang, G. J .; Fowler, J . S.; Logan, J .; Gatley, S.
J .; Hitzemann, R.; Chen, A. D.; Dewey, S. L.; Pappas, N.
Decreased Striatal Dopaminergic Responsiveness in Detoxified
Cocaine-dependent Subjects. Nature 1997, 386, 830-833.
(10) Aquilonius, S. M.; Bergstrom, K.; Eckernas, S. A.; Hartvig, P.;
Leendeers, K. L.; Lundquist, H.; Antoni, G.; Gee, A.; Rimland,
A.; Uhlin, J .; Langstrom, B. In Vivo Evaluation of Striatal
Dopamine Reuptake Sites Using 11C-Nomifensine And Positron
Emission Tomography. Acta Neurol. Scand. 1987, 76, 283-287.
(25) Halldin, C.; Farde, L.; Lundkvist, C.; Ginovart, N.; Nakashima,
Y.; Karlsson, P.; Swahn, C. G. [11C]â-CIT-FE, a Radioligand for
Quantitation of the Dopamine Transporter in the Living Brain
Using Positron Emission Tomography. Synapse 1996, 22, 386-
390.
(26) Chaly, T.; Dhawan, V.; Kazumata, K.; Antonini, A.; Margoulef,
C.; Dahl, R.; Belakhef, A.; Margoulef, D.; Yee, A.; Wang, S.;
Tamagnan, G.; Neumeyer, J . L.; Eidelberg D. Radiosynthesis of
[
18F]N-3-Fluoropropyl-2â-Carbomethoxy-3â-(4-Iodophenyl)Nor-
tropane and the First Human Study with Positron Emission
Tomography. Nucl. Med. Biol. 1996, 23, 999-1004.
(27) Wilson, A. A.; DaSilva, J . N.; Houle, S. In Vivo Evaluation of
[
11C]- and [18F]-Labeled Cocaine Analogues as Potential Dopa-
mine Transporter Ligands for Positron Emission Tomography.
Nucl. Med. Biol. 1996, 23, 141-146.
(28) Goodman, M. M.; Keil, R.; Shoup, T. M.; Eshima, D.; Eshima,
L.; Kilts, C.; Votaw, J .; Camp, V. M.; Votaw, D.; Smith, E.; Kung,
M. P.; Malveaux, E.; Watts, R.; Huerkamp, M.; Garcia, E.;
Hoffman J . M. Fluorine-18 Labeled 2â-Carbomethoxy-3â-(4-
Chlorophenyl)-8-(-3-Fluoropropyl)nortropane (FPCT): A PET
Radiotracer for Imaging Dopamine Transporters. J . Nucl. Med.
1997, 38, 119-126.

648 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 4
Xing et al.
(29) Carroll, F. I.; Abraham, P.; Lewin, A. H.; Parham, K.; Boja, J .
W.; Kuhar, M. J . Isopropyl and Phenyl Esters of 3â-(4-Substi-
tutedphenyl)tropan-2â-carboxylic Acids. Potent and Selective
Compounds for the Dopamine Transporter. J . Med. Chem. 1992,
35, 2497-2500.
PC-12 Cells Demonstrate High Affinity Uptake of Dopamine In
Vitro and [F-18]-DOPA Uptake and Metabolism After Intra-
cerebral Implantation in Nonhuman Primates. Cell Transplant.
1997, 6, 469-477.
(30) Subramanian, T.; Bakay, R. A. E.; Emerich, D. F.; Hoffman, J .
M.; Goodman, M. M.; Miller, G. W.; Levey, A. I.; Hubert, G. W.;
Batchelor, S.; Winn, S. R.; Watts, R. L. Polymer Encapsulated
J M9902234