Synthesis and pharmacology of site-specific cocaine abuse treatment agents: 2-Substituted-6-amino-5-phenylbicyclo[2.2.2]octanes

Article abstract of DOI:10.1021/jm990306k

A series of 2-substituted-6-amino-5-phenylbicyclo[2.2.2]octanes was synthesized and tested for inhibitor potency in [³H]WIN 35,428 (WIN) binding at the dopamine (DA) transporter and [³H]DA uptake assays. To demonstrate transporter se

Full text of DOI:10.1021/jm990306k

4836
J . Med. Chem. 1999, 42, 4836-4843
Syn th esis a n d P h a r m a cology of Site-Sp ecific Coca in e Abu se Tr ea tm en t Agen ts:
2-Su bstitu ted -6-a m in o-5-p h en ylbicyclo[2.2.2]octa n es
Sahar J avanmard, Howard M. Deutsch,* David M. Collard, Michael J . Kuhar,‡ and Margaret M. Schweri†
School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, Georgia 30332-0400, Yerkes Regional Primate
Center, Emory University, Atlanta, Georgia 30329, and Mercer University School of Medicine, Macon, Georgia 31201-1554
Received J une 14, 1999
A series of 2-substituted-6-amino-5-phenylbicyclo[2.2.2]octanes was synthesized and tested for
inhibitor potency in [3H]WIN 35,428 (WIN) binding at the dopamine (DA) transporter and
[3H]DA uptake assays. To demonstrate transporter selectivity for the compounds, inhibitor
potency was also tested using [3H]nisoxetine and [3H]paroxetine binding assays for the
norepinephrine (NE) and serotonin (5-HT) transporters, respectively. Synthesis was ac-
complished by bisannulation of the enamine derived from phenylacetaldehyde and dimethyl-
amine with 2-cyclohexenone to give a mixture of endo- and exo-trans-6-amino-5-phenylbicyclo[2.2.2]-
octan-2-ones. The separated ketones were reduced to the four diastereomeric alcohols which
were converted to acetyl and benzoyl esters. The ketones, alcohols, and acetyl esters generally
have low affinity for the three transporters and do not effectively inhibit the uptake of [3H]DA.
In all cases, the benzoates show significantly greater inhibition of WIN binding compared to
the corresponding ketones, alcohols, or acetate esters. One compound, (1R/S,4R/S)-6R/S-(N,N-
dimethylamino)-5R/S-phenylbicyclo[2.2.2]oct-2S/ R-yl benzoate, is almost as potent as cocaine
in binding to the DA transporter (IC50 ) 270 nM versus 159 nM for cocaine). The C-2 epimer,
(1R/S,4R/S)-6R/S-(N,N-dimethylamino)-5R/S-phenylbicyclo[2.2.2]oct-2R/S-yl benzoate, was se-
lective and potent in binding to the 5-HT transporter (IC50 ) 53 nM versus 1050 nM for cocaine)
as compared to the DA transporter (IC50 ) 358 nM). A preliminary molecular modeling study
of the benzoyl esters indicates that their relative potencies in the WIN binding assay are not
correlated to the nitrogen to benzoate phenyl (centroid) distance or to the deviation of the
nitrogen from the plane defined by the benzoate ring.
In tr od u ction
provide guidance in the design of potential treatment
agents. For example, WIN 35,428 (2, WIN) and GBR
12783 (4) are phenylpropylamines, whereas methylpheni-
A natural component of coca leaves (Erythroxylum
coca), (-)-cocaine, 1, is a psychostimulant and powerful
reinforcer.1,2 It has become one of the most prevalent
and problematic drugs of abuse, and considerable effort
has been expended in the study of its mechanism of
action. It is now widely accepted that (-)-cocaine binds
to specific recognition sites associated with monoamine
transporters3,4 in the mammalian brain. Its primary
mechanism of action has been ascribed to its ability to
inhibit the dopamine (DA) transporter for the reuptake
of DA5-7 into the presynaptic neuron, thus increasing
the concentration of DA in the synapse.
Several approaches to the development of effective
treatment agents for cocaine abuse have been proposed.
One approach is to identify agonists (or partial agonists)
which can serve as cocaine substitutes, the pharmaco-
logical equivalent of methadone in the treatment of
opiate abuse.8 Such a substance would elicit some of the
same effects in the user as cocaine itself but, perhaps,
not cause the same degree of euphoria. The most
promising drug of this type might be a long-acting, slow-
onset agonist. An alternative approach would be to
make use of an antagonist, a substance which blocks
the binding of cocaine yet still allows DA reuptake.
However, no drugs of this type are currently known. The
structures of known potent DA reuptake inhibitors
date (3) is a phenylethylamine. To develop further leads
to agonists, partial agonists, and antagonists of cocaine,
we have undertaken a program of study of rigid bicyclic,
non-tropane amines upon which a myriad of substitu-
ents can be placed in various regio- and stereochemical
arrangements. These are used to probe structure-
activity relationships for DA binding and reuptake,
along with binding at the serotonin (5-HT) and norepi-
nephrine (NE) transporters. An approach9 that ap-
peared attractive was based on a series of 2-aminomethyl-
3-phenylbicyclo[2.2.2]octane analogues which were
‡ Emory University.
† Mercer University School of Medicine.
10.1021/jm990306k CCC: $18.00 © 1999 American Chemical Society
Published on Web 10/29/1999

Site-Specific Cocaine Abuse Treatment Agents
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 23 4837
initially synthesized10 and tested for biological
activity10-13 in the late 1970s. The most potent of these
compounds, LR 5182 (5), was shown to be selective for
the DA transporter, with Ki values of 3, 58, and 1700
nM for inhibition of [3H]DA, [3H]NE, and [3H]5-HT,
respectively.11 Despite a lack of the tropane core, LR
5182 has a reasonable degree of structural analogy with
cocaine, including the three-dimensional spatial ar-
rangement of the basic nitrogen and phenyl ring.
In contrast to our previous work9 on variants of 5,
the study described here involves synthesis of rigid
bicyclic amines 6-19 which: (i) lack the exocyclic
methylene group present in 5 and (ii) incorporate an
oxygen functionality (ketone, alcohol, or ester) on the
bicyclic framework. Incorporation of the oxygen func-
tionality provides the opportunity to enhance the water
solubility of agents by increasing the hydrophilicity of
the hydrocarbon face of 5 and provides for additional
structural diversity. Bicyclic amines 6-19 were studied
for their effect on the DA transporter by measuring
inhibition of [3H]WIN binding and [3H]DA uptake. To
demonstrate transporter selectivity of the compounds,
data for the NE and 5-HT transporters are also reported
by measuring the displacement of [3H]nisoxetine and
[3H]paroxetine binding, respectively.
and endo isomers, respectively. Apparently the initial
Michael addition leads to equal amounts of the R/S (S/
R) and S/S (R/R) diastereomers, since we observe equal
amounts of the exo and endo isomers.
Reduction of the endo amino ketone 6 with NaBH4
in absolute EtOH gave exclusively 8 with the hydroxyl
group in the endo position presumably due to shielding
of one face of the carbonyl carbon of 6 by the N,N-
dimethylamino group. The structure of 8 was confirmed
by single-crystal X-ray crystallography. Treatment of 6
with borane-THF at -78 °C led exclusively to the
diastereomeric exo alcohol 7 in which hydride has been
introduced to the most sterically hindered face of the
ketone. This observation is consistent with the com-
plexation of the electron-rich amine and electron-
deficient borane prior to hydride transfer to the carbo-
1
nyl. H NMR spectroscopy clearly differentiates 7 and
8 by observation of chemical shifts for the carbinol
methine at 4.4 and 4.0 ppm, respectively.
Reduction of the exo amino ketone 13 with NaBH4
gave a mixture of diastereomeric alcohols, 14 and 15
(1:1 ratio), which were separated by triturating the
mixture with EtOAc. Single-crystal X-ray crystallogra-
phy showed that the EtOAc-insoluble material is alcohol
15, which confirms that it is an exo amine and shows
that the hydroxyl substituent is in the endo position.
The EtOAc-soluble fraction contained the diastereomer
with the hydroxyl in the exo position (i.e., 14). The
1
carbinol proton of alcohols 14 and 15 coincide in the H
NMR spectrum (δ 4.1). Alcohols 7, 8, 14, and 15 were
converted to acetyl and benzoyl esters, 9-12 and 16-
19, by treatment with the appropriate acid chlorides in
benzene containing triethylamine.
P h a r m a cology. The IC50 values for the inhibition of
ligand binding to the DA,16 5-HT, and NE17 transporters
and the inhibition of [3H]DA uptake18 into synaptosomes
were determined according to established protocols and
are shown in Table 1. Binding to the DA transporter
was measured using [3H]WIN 35,428 (WIN), to the 5-HT
transporter using [3H]paroxetine (PAR), and to the NE
transporter using [3H]nisoxetine (NIS).
In h ibition of WIN Bin d in g. In all cases, the endo
amines are more potent than the corresponding exo
amines by 3-8-fold. In addition, in all cases, the
benzoate esters are more potent than the corresponding
alcohols (3-8-fold). The alcohols have little difference
in potency as compared to the corresponding ketones,
which are somewhat more potent than the correspond-
ing acetates (1.5-4-fold). The most potent compound is
endo amine endo benzoyl ester 12 (IC50 ) 0.270 µM)
which is comparable to (-)-cocaine (IC50 ) 0.159 µM).
For all of the synthesized compounds, the stereochem-
istry at the 2-position has little effect on the potency.
In h ibition of [3H]DA Up ta k e. Similar conclusions
to those discussed about WIN binding can be reached
when considering the inhibition of DA uptake.
In h ibition of P AR Bin d in g. Generally, but not in
all cases, the endo amines are more potent than the
corresponding exo amines (2-14-fold more potent in four
cases, 6-8-fold less potent in two cases). Somewhat
similar trends in potency are observed for the 5-HT
transporter as for the DA transporter for the different
derivatives. In all cases, the benzoate esters are more
potent than the corresponding alcohols (2-, 16-, 50-, and
Resu lts a n d Discu ssion
Ch em istr y. Enamine 20, which was prepared by
condensation of phenylacetaldehyde with dimethyl-
amine,14 underwent reaction with cyclohexenone15 to
give a mixture of substituted bicyclo[2.2.2]octan-2-ones,
6 and 13, as shown in Scheme 1. The isomeric ketones
were formed in approximately a 1:1 ratio and were
separated by flash chromatography on silica gel. It has
been previously shown15 that similar bisannulation
procedures yield mixtures of endo and exo isomers in
some cases and only the endo isomer in other cases.
Single-crystal X-ray crystallography showed that the
less polar (thin-layer chromatography: silica gel/EtOAc,
Rf ) 0.38) ketone was 6, which has the N,N-dimethy-
lamino substituent in the endo position (i.e., the amine
is syn to the carbonyl) and the trans relationship
between the phenyl and amino substituents.
We speculate that the mechanism of this reaction
would involve an initial Michael addition of the enamine
20 to 2-cyclohexenone to form enolate A, as shown in
Figure 1. This enolate would be in equilibrium with the
regioisomeric enolate B which could ring close from the
conformations shown in which the phenyl and iminium
groups are approximately anti to each other. This would
lead to only trans isomers, and in the case illustrated
here, the S/S and R/S diastereomers would form the exo

4838 J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 23
J avanmard et al.
Sch em e 1a
a
Reagents and conditions: (a) (CH3)2NH, Et2O, CaCl2; (b) cyclohexenone, benzene, ∆; (c) BH3-THF, THF, -78 °C; (d) NaBH4, EtOH,
∆; (e) RC(O)Cl, Et3N, benzene.
F igu r e 1. Possible mechanism of the bisannulation reaction.
160-fold). The alcohols are generally slightly more
potent than the corresponding ketones, which are not
uniformly more potent than the corresponding acetates
(in one case 6-fold more potent, in three cases 3-13-
fold less potent). For PAR binding at the 5-HT trans-
porter, the stereochemistry of the 2-position has a large
influence on activity. In all cases the exo isomers are
more potent than the corresponding endo isomers (3-
210-fold) with the endo amine exo benzoate 11 being
210-fold more potent than the endo benzoate isomer 12.
However, in the exo amine series, there is only a 2.5-
fold decrease in potency when the benzoate function is
moved from the exo to the endo position (18 versus 19).
As in the case of the benzoate esters in the endo amine
series, there is also a significant effect of the stereo-
chemistry at C-2 for the acetate esters in the endo series.
Changing the stereochemistry of the acetyl group from
exo (9) to endo (10) reduces the potency 71-fold. Overall,
the most effective compound at inhibiting PAR binding
at the 5-HT transporter is 11 (IC50 ) 0.053 µM).
In h ibition of NIS Bin d in g. In all cases, the endo
amines are more potent than or equipotent to the
corresponding exo amines. The compound most potent
for the NE transporter is the endo amine exo benzoate
11 (IC50 ) 0.225 µM). The order of potency of the
compounds in inhibiting the binding of NIS at the NE
transporter generally, but not always, follows the pat-
tern benzoyl > ketone > alcohol > acetate. Somewhat
surprisingly, the endo amine exo acetate 9 was the
second most potent compound (IC50 ) 0.535 µM). As
with the 5-HT transporter, the stereochemistry of the
2-position has a much greater effect on activity in the

Site-Specific Cocaine Abuse Treatment Agents
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 23 4839
Ta ble 1. Binding and Uptake Data for 2-Substituted-6-amino-5-phenylbicyclo[2.2.2]octanes and Reference Compounds
IC50 (µM)
IC50
NE/DA
IC50
5-HT/DA
compda
DA [3H]WINb
NE [3H]NISc
5-HT [3H]PARc
[3H]DA uptaked
(-)-cocaine
0.160 ( 0.015
0.0204 ( 0.0017
0.083 ( 0.0079
0.0142 ( 0.0016
4.01 ( 0.40
3.30 ( 0.29
0.835 ( 0.045
0.989 ( 0.054
0.0960 ( 0.003
4.12 ( 0.29
1.51 ( 0.21
7.23 ( 0.71
0.535 ( 0.025
55.4 ( 11.9
0.225 ( 0.044
1.47 ( 0.017
10.6 ( 1.6
1.05 ( 0.089
0.809 ( 0.059
224 ( 19
0.404 ( 0.026
0.0513 ( 0.00030
0.024 ( 0.019
0.0293 ( 0.0017
9.14 ( 0.63
4.45 ( 0.12
2.81 ( 0.270
4.93 ( 0.63
25.8 ( 3.00
0.372 ( 0.061
0.687 ( 0.015
24.7 ( 0.45
10.9 ( 2.3
21
41
12
6.6
40
2700
18
7.8
4.7
11
0.46
14
WIN 35,428
methylphenidate
LR 51829
0.261 ( 0.016
31.4 ( 7.6
6.8
1.0
0.85
3.5
0.10
4.5
0.65
5.4
1.0
1.1
3.5
0.76
1.0
3.4
1.8
6
7
8
9
1.77 ( 0.13
8.38 ( 1.1
2.04 ( 0.24
21.8 ( 2.75
2.45 ( 0.18
176 ( 5.5
5.33 ( 0.36
10
11
12
13
14
15
16
17
18
19
12.2 ( 0.69
0.358 ( 0.067
0.270 ( 0.029
10.3 ( 0.40
0.053 ( 0.012
11.1 ( 1.39
61.2 ( 17
12.3 ( 2.5
>100
4.77 ( 0.12
22.8 ( 0.51
0.760 ( 0.081
1.88 ( 0.039
0.15
41
5.9
7.04 ( 0.29
7.73 ( 0.80
34.4 ( 4.7
1.7
9.92 ( 0.94
23.0 ( 1.6
>10
0.35
0.50
0.35
0.83
13.6 ( 1.43
10.4 ( 3.6
46.8 ( 2.6
7.38 ( 0.66
4.19 ( 0.74
16.3 ( 0.16
45.6 ( 6.24
∼75e
2.20 ( 0.31
4.84 ( 0.064
8.24 ( 0.33
2.27 ( 0.066
a
b
Compounds tested as the HCl salts. All values are the mean of two experiments performed in triplicate. c All values are the mean
of three experiments performed in triplicate. All values are the mean of two experiments performed in duplicate. e 55.3 ( 0.7% inhibition
at 75 µM.
d
endo amine series than in the exo series. For example,
changing the stereochemistry of the acetyl and benzoyl
substituents from exo (9 and 11) to endo (10 and 12)
decreases the potency by 100- and 7-fold, respectively.
Similar comparisons in the endo series show much
smaller (or no) decreases in potency: exo (16 and 18) to
endo (17 and 19) result in 4.5- and 0.6-fold potency
reductions, respectively.
Tr a n sp or ter Selectivity. Selectivity of compounds
6-19 for the three transporters was calculated by
comparing the ratio of IC50 values, NIS/WIN for NE/
DA selectivity and PAR/WIN for 5-HT/DA selectivity,
as shown in Table 1. The compound that shows the
highest selectivity for the DA transporter was endo
amine endo benzoate 12, which is 41 times more potent
for the DA transporter than for the 5-HT transporter
and 5 times more potent for the DA transporter than
for the NE transporter. Interestingly, endo amine exo
benzoate 11 is most active and selective for the 5-HT
transporter, being 7-fold selective compared to the DA
transporter and 4-fold selective compared to the NE
transporter.
Ta ble 2. Interatomic Distances and Relative Distribution of
Atoms in Space
distances (Å)
a
b
c
N-Phc
N-Phfa
N-Php
WIN
PAR
11
12
18
5.57
6.91
7.68
5.43
8.64
7.19
8.30
10.81
10.18
8.15
11.01
9.61
0.07
1.4
0.43
0.74
1.1
19
0.35
a
b
Phc ) centroid of benzoate phenyl ring. Phfa ) furthest atom
on benzoate phenyl ring. c Php ) benzoate phenyl plane.
F igu r e 2. Definitions of distances used in the description of
models
Gen er a l Con sid er a tion s. Since the benzoate esters
were generally the most potent derivatives, it was
hypothesized that the benzoate ester phenyl ring was
important in the binding interactions of these com-
pounds. To test if the distance and orientation of the
benzoate phenyl ring from the nitrogen atom are
important parameters in the pharmacophore of these
compounds, energy-minimized (MM2)19 conformational
searches were computed to obtain the global minima.
Different conformations about the two C-Ph and two
C-O bonds were manually searched. The distances
between the nitrogen and the centroid of the benzoate
phenyl ring (N-PhC) and between the nitrogen and the
plane of the phenyl ring20 (N-PhP) were calculated and
are shown in Table 2 (also see Figure 2). In its energy-
minimized (MM2)19 conformation, the highly potent DA
uptake inhibitor WIN, 2, has a N-PhC distance of 5.57
Å with the nitrogen only slightly out of the plane defined
by the phenyl ring (0.07 Å). Examination of the interac-
tion of the test compounds with the DA transporter
revealed that the most active of the benzoyl esters (12,
IC50 ) 0.270 µM) has a N-PhC distance of 5.43 Å with
the nitrogen further out of the plane of the phenyl ring
(0.74 Å). One of the less active of the benzoate esters
(18, IC50 ) 2.20 µM) has a larger N-PhC distance (8.64
Å) and a considerably larger deviation of the nitrogen
from the plane of the phenyl ring (1.1 Å). However, the
other less active benzoate ester (19, IC50 ) 2.27 µM) has
a much shorter N-PhC distance (7.19 Å) and smaller
out of plane distance (0.35 Å). This is comparible to the
more active 11 (IC50 ) 0.358 µM, N-PhC ) 7.68 Å,
N-Php ) 0.43 Å). These data alone do not show how
either the orientation of the benzoate phenyl ring with
respect to the nitrogen or the distance between the two
is of significance in determining the potency of these
agents in inhibiting binding to the DA transporter.
In contrast, the 5-HT transporter seems to be able to
accommodate a larger N-PhC distance. Although the
N-Phc distance has been correlated to potency be-
fore,21,22 such an analysis did not take the phenyl
substituents into account. Thus, we also tabulate the
distance between the nitrogen and the furthest atom

4840 J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 23
J avanmard et al.
of potent and selective inhibitors of the monoamine
transporters.
Exp er im en ta l Section
Gen er a l Meth od s. Melting points were determined on a
Mel-Temp apparatus and are uncorrected. 1H and 13C NMR
spectra were obtained on a Varian Gemini spectrometer at 300
and 75 MHz, respectively. IR spectra were obtained using KBr
pellets on a Nicolet 520FT spectrometer. All starting materials
were used as received from Aldrich Chemical Co. Tetrahydro-
furan (THF) was dried over sodium benzophenone ketyl prior
to distillation under nitrogen. Benzene was dried over 4 Å
molecular sieves. Free bases were dissolved in minimum
amount of MeOH and were converted to HCl salts by the
addition of 1.5 equiv of concentrated HCl. MeOH was added
and the HCl was removed under reduced pressure. The
resulting solid was recrystallized from MeOH/EtOAc. Elemen-
tal analyses were obtained from Atlantic Microlabs, Atlanta,
GA.
F igu r e 3. A. Overlay of PAR (shaded) and 6R enantiomer 11
(white). B. Overlay of WIN (shaded) and 6R enantiomer of 12
(white). C. Overlay of WIN (shaded) and 6S enantiomer of 12
(white).
E-2-(N,N-Dim eth yla m in o)-1-p h en yleth en e (20). A mix-
ture of dimethylamine (2.76 mL, 41.7 mmol), Et2O (34 mL),
phenylacetaldehyde (1.95 mL, 6.60 mmol), and CaCl2 pellets
(3.75 g) was stirred at room temperature for 64 h. The mixture
was filtered and concentrated under reduced pressure to give
20 (2.25 g, 91.8%) as a dark yellow viscous oil. Distillation gave
1.65 g (73.3%) of 20 as a yellow liquid: bp 165-168 °C at 0.5
(N-Phfa) on the ring in Table 2 (also see Figure 2). The
compound most effective at inhibiting PAR binding at
the 5-HT transporter, 11 (IC50 ) 0.052 µM), has an
N-Phfa distance (10.18 Å) comparable to that of PAR
(10.81 Å). In contrast the least active benzoate, 12 (IC50
) 11.1 µM), has a much shorter N-Phfa distance (8.15
Å). Figure 3A shows an overlay of PAR and the 6R
enantiomer of benzoate ester 11 and illustrates the good
overlap of the phenyl groups and nitrogen atoms.
Figure 3 also shows an overlay of WIN and the 6R
enantiomer of benzoate ester 12 (B) and the 6S enan-
tiomer of 12 (C). Substituents on the two-carbon bridge
often dramatically decrease the potency of of tropane-
based compounds.23 Comparison of these two overlays
demonstrates that the enantiomer shown in Figure 3C
has atoms in the region of the two-carbon bridge of the
tropane whereas that shown in Figure 3B does not; this
might provide future direction in targeting the develop-
ment of potent DA uptake inhibitors.
1
mmHg (lit.14 bp 100-102 °C/0.5 mmHg); H NMR (CDCl3) δ
2.82 (s, 6H, CH3), 5.22 (d, J ) 13 Hz, 1H, CHdCHN), 6.84
(dd, J ) 13 Hz, J ) 2.1 Hz, 1H, CHdCHN), 7.00-7.25 (m,
5H, Ar).
(1R/S,4R/S)-6-(N,N-Dim et h yla m in o)-5-p h en ylb icyclo-
[2.2.2]octa n -2-on es 6 a n d 13. A solution of 20 (1.5 g, 10
mmol) and 2-cyclohexenone (1.54 mL, 15.9 mmol) in benzene
was heated at reflux for 48 h. The solvent was removed under
reduced pressure. The residue was dissolved in CH2Cl2 (10 mL)
and the solution was extracted with 6 M aqueous HCl (3 × 10
mL). The aqueous extracts were combined, made basic with
NaOH pellets, and extracted with CH2Cl2 (3 × 10 mL). The
combined organic extracts were dried over MgSO4 and filtered
and the solvent was removed under reduced pressure to afford
a viscous brown oil. Purification by flash column chromatog-
raphy (silica gel; eluted sequentially with 2:1 hexane:EtOAc,
1:1 hexane:EtOAc, and EtOAc) gave 6 as an off-white solid
(0.19 g, 25%) and further elution (1:1 EtOAc:MeOH then
MeOH) gave 13 as a viscous brown oil (0.15 g, 20%).
(1R/S,4R/S)-6R/S-(N,N-Dim et h yla m in o)-5R/S-p h en yl-
bicyclo[2.2.2]octa n -2-on e (6): 1H NMR (CDCl3) δ 1.04-1.28
(m, 1H), 1.46-62 (m, 1H), 1.74-1.79 (m, 2H), 1.99-2.01 (m,
7H, N(CH3)2), 2.16 (dd, J ) 19 Hz, J ) 3, 1H), 2.37 (dd, J )
19 Hz, J ) 3 Hz, 1H), 2.64-2.69 (m, 2H, C-5), 2.88 (br s, 1H,
C-6), 7.12-7.30 (m, 5H, aromatic); 13C NMR (CDCl3) δ 12.7,
16.3, 30.9, 37.7, 39.7, 40.2, 44.9, 62.8 (aliphatic); 120.9, 122.4,
123.1, 138.5 (aryl); 209.7 (CdO).
6-HCl: 97% yield; mp 203-204 °C; 1H NMR (D2O) δ 1.25-
1.34 (m, 1H), 1.52-1.62 (m, 1H), 1.81-1.92 (m, 1H), 1.95-
2.05 (m, 1H), 2.10 (q, J ) 3.0 Hz, 1H), 2.45 (d, J ) 2.7 Hz,
1H), 2.49 (t, J ) 2.7 Hz, 1H), 2.56 (s, 6H, N(CH3)2), 2.84 (q, J
) 2.7 Hz, 1H, C-5), 3.12 (d, J ) 7.2 Hz, 1H, C-6), 4.07 (dt, J )
7.2 Hz, J ) 1.8 Hz, 1H, C-2), 7.18-7.29 (m, 5H, Ar); IR 2657-
2591, 1729 cm-1. Anal. (C16H22ONCl‚1.2H2O) C, H, N, Cl.
(1R/S,4R/S)-6S/R-(N,N-Dim et h yla m in o)-5S/R-p h en yl-
bicyclo[2.2.2]octa n -2-on e (13): 1H NMR (CDCl3) δ 1.45-
1.60 (m, 2H), 1.75 (s, 1H), 1.82-1.88 (m, 1H), 1.92-1.97 (m,
7H, N(CH3)2), 2.04-2.18 (m, 2H), 2.48 (br s, 1H), 2.61 (br s,
1H, C-5), 2.78 (br s, 1H, C-6), 6.98-7.21 (m, 5H, Ar); 13C NMR
(CDCl3) δ 10.1, 20.6, 31.2, 33.1, 38.7, 41.6, 45.3, 59.6 (aliphatic);
120.9, 122.5, 123.2, 139.0 (aryl); 210.8 (CdO).
Due to the many possible low-energy conformations
of NIS, it is difficult to comment on the distances that
are optimal for effective binding.
Con clu sion s
A series of trans-6-(N,N-dimethylamino)-5-phenyl-
bicyclo[2.2.2]octanes was prepared by a bisannulation
procedure with a variety of substituents at the 2-posi-
tion. Interestingly, the ketone 6 could be stereospecifi-
cally reduced to either alcohol 7 or 8 with diborane or
sodium borohydride, respectively. The ketones (6, 13),
alcohols (7, 8, 14, 15), acetates (9, 10, 16, 17), and
benzoates (18 and 19) usually have moderate to low
affinity for the DA, 5-HT, and NE transporters and do
not effectively inhibit the uptake of [3H]DA, relative to
cocaine. The benzoate ester 12 is the most potent
derivative for inhibiting radioligand binding to the DA
transporter, while benzoate 11 is quite potent and
somewhat selective for inhibiting the 5-HT transporter.
In these benzoate esters (11 and 12) the ester phenyl
ring (and not the phenyl at C-5) might be binding to
the “aromatic ring binding region” in the various
transporters. Future work will test the importance of
the benzoate ester phenyl ring by removal of the phenyl
at C-5. In addition, the enantiomers of 12 will be
prepared to determine which is most active; this will
provide further input into a model for the development
13-HCl: 74% yield; mp 184-186 °C; 1H NMR (D2O) δ 1.64-
1.77 (m, 2H), 1.88-1.91 (m, 2H), 2.10-2.25 (m, 2H), 2.50-
2.77 (m, 6H, N(CH3)2), 2.95 (br s, 1H), 3.11-3.15 (m, 1H), 3.17
(dd, J ) 4.8 Hz, J ) 3.3 Hz, 1H, C-5) 3.88-3.98 (m, 1H, C-6),
7.15-7.33 (m, 5H, Ar); IR 2664-2460, 1729, 1130 cm-1. Anal.
(C16H22ONCl‚1.52H2O) C, H, N, Cl.

Site-Specific Cocaine Abuse Treatment Agents
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 23 4841
(1R/S,4R/S)-6R/S-(N,N-Dim et h yla m in o)-5R/S-p h en yl-
bicyclo[2.2.2]octa n -2R/S-ol (7). BH3-THF (8 mL) was slowly
added to a solution of 6 (2.26 g, 9.92 mmol) dissolved in THF
(20 mL) at -78 °C and stirred under nitrogen for 20 h. Aqueous
6 M HCl (10 mL) was added to the mixture and THF was
removed under reduced pressure. The resulting solution was
made basic with NaOH pellets and extracted with CH2Cl2 (4
× 10 mL). The organic extracts were combined, dried over
MgSO4, and filtered, and the solvent was removed under
reduced pressure to afford 7 as a white solid (2.27 g, 100%
yield): 1H NMR (CDCl3) δ 1.25-1.35 (m, 1H), 1.40-1.50 (m,
3H), 1.61 (d, J ) 2.7 Hz, 1H), 2.12-2.21 (m, 8H, N(CH3)2),
2.34-2.43 (m, 1H), 2.62 (br s, 1H, C-5), 2.69 (br s, 1H, C-6),
4.42 (d, J ) 9.9 Hz, 1H, C-2), 7.10-7.35 (m, 5H, Ar); 13C NMR
(CDCl3) δ 12.5, 13.5, 29.6, 29.6, 33.5, 38.8, 45.2, 58.5, 62.8
(aliphatic); 120.6, 122.6, 123.0, 139.8 (aryl).
15-HCl: 78% yield; mp 225-226 °C; 1H NMR (D2O) δ 1.20-
1.25 (m, 1H), 1.43-1.61 (m, 6H), 2.25 (br s, 1H, C-1), 2.62 (br
s, 6H, N(CH3)2), 2.82 (d, J ) 6.0 Hz, 1H, C-5), 3.89 (d, J ) 6.0
Hz, 1H, C-6), 4.06-4.20 (m, 1H, C-2), 7.21 (m, 5H, Ar); IR
3473-3361, 2710, 1110, 1045 cm-1. Anal. (C16H24ONCl‚0.20H2O)
C, H, N, Cl.
6-(N,N-Dim eth yla m in o)-5-p h en ylbicyclo[2.2.2]octa n -2-
yl Aceta tes a n d Ben zoa tes 9-12 a n d 16-19. The conver-
sion of 14 to 16 is illustrated. Acetyl chloride (40.0 µL, 0.567
mmol) and Et3N (91.0 µL, 0.654 mmol) were added to a solution
of 14 (0.107 g, 0.436 mmol) in benzene (6 mL). The solution
was stirred under nitrogen at room temperature for 24 h. The
mixture was washed with water (1 × 10 mL) and 5% aqueous
Na2CO3 (3 × 10 mL). The organic layer was dried over MgSO4
and filtered and the solvent was removed under reduced
pressure to afford 16 as a white solid (0.120 g, 96% yield).
(1R/S,4R/S)-6S/R-(N,N-Dim et h yla m in o)-5S/R-p h en yl-
bicyclo[2.2.2]oct-2R/S-yl a ceta te (16): 73% yield; 1H NMR
(CDCl3) δ 1.13 (dd, J ) 14.7 Hz, J ) 1.2 Hz, 1H), 1.58-1.69
(m, 5H), 1.96-2.02 (m, 1H), 2.05 (s, 3H, CH3), 2.10 (s, 6H,
N(CH3)2), 2.22-2.26 (m, 1H), 2.42-2.44 (m, 1H, C-5), 2.75 (d,
J ) 5.1 Hz, 1H, C-6), 4.98 (dt, J ) 9.9 Hz, J ) 3.3 Hz, 1H,
C-2), 7.10-7.20 (m, 5H, Ar); 13C NMR (CDCl3) δ 7.4, 15.8, 21.3,
23.2, 26.7, 28.5, 38.6, 45.3, 61.1, 66.4 (aliphatic); 120.7, 122.5,
123.1, 139.8 (aryl); 165.5 (CdO).
16-HCl: 71% yield; mp 250-251 °C; 1H NMR (D2O) δ 1.10-
1.14 (m, 1H), 1.30-1.45 (m, 1H), 1.50-1.68 (m, 3H), 1.79-
1.87 (m, 2H), 1.93 (br s, 3H, CH3), 2.39 (br s, 1H, C-1), 2.59
(br s, 6H, N(CH3)2), 2.80-2.84 (m, 1H, C-5), 3.60-3.68 (m, 1H,
C-6), 4.83-4.92 (m, 1H, C-2), 7.15-7.28 (m, 5H, Ar); IR 2670-
2591, 1729, 1262, 1038 cm-1. Anal. (C18H26O2NCl‚0.75H2O) C,
H, N, Cl.
(1R/S,4R/S)-6R/S-(N,N-Dim et h yla m in o)-5R/S-p h en yl-
bicyclo[2.2.2]oct-2R/S-yl a ceta te (9): 100% yield; 1H NMR
(CDCl3) δ 1.22-1.49 (m, 3H), 1.54 (dt, J ) 14.4 Hz, J ) 3.0
Hz, 1H), 1.63 (dd, J ) 3.0 Hz, J ) 2.7 Hz, 1H), 1.91-2.03 (m,
1H), 2.05 (s, 3H, CH3), 2,13 (m, 6H, N(CH3)2), 2.32 (m, 2H,
C-1 and C-4), 2.44 (dd, J ) 4.8 Hz, J ) 2.7 Hz, 1H, C-5), 2.70
(br s, 1H, C-6), 5.27 (dt, J ) 10.2 Hz, J ) 3.0 Hz, C-2), 7.15-
7.39 (m, 5H, Ar); 13C NMR (CDCl3) δ 13.0, 13.2, 15.9, 26.2,
28.9, 30.2, 38.9, 45.8, 62.3, 63.3 (aliphatic); 120.6, 122.6, 122.9,
139.9 (aryl); 165.7 (CdO).
9-HCl: 74% yield; mp 197-199 °C; 1H NMR (D2O) δ 1.21-
1.65 (m, 7H), 1.90 (br s, 1H), 1.97 (s, 3H, CH3), 2.18-2.26 (m,
1H), 2.46-2.85 (m, 6H, N(CH3)2), 3.74 (dd, J ) 15 Hz, J ) 3.9
Hz, 1H, C-6), 4.95 (m, 1H, C-2), 7.18-7.40 (m, 5H, Ar); IR
2670-2591, 1729, 1262, 1038 cm-1. Anal. (C18H26O2NCl‚
0.30H2O) C, H, N, Cl.
(1R/S,4R/S)-6R/S-(N,N-Dim et h yla m in o)-5R/S-p h en yl-
bicyclo[2.2.2]oct-2S/R-yl a ceta te (10): 74% yield; 1H NMR
(CDCl3) δ 1.10-1.14 (m, 1H), 1.39-1.42 (m, 1H), 1.58-1.68
(m, 3H), 1.79 (dt, J ) 13.8 Hz, J ) 4.8 Hz, J ) 2.7 Hz, 1H),
2.07 (s, 3H, CH3), 2.1 (s, 6H, N(CH3)2), 2.17-2.28 (m, 1H),
2.41-2.45 (m, 1H), 2.51 (d, J ) 4.8 Hz, 1H, C-5), 3.02 (br s,
1H, C-6), 4.94-4.97 (m, 1H, C-2), 7.10-7.35 (m, 5H, Ar); 13C
NMR (CDCl3) δ 12.3, 15.9, 18.6, 25.6, 28.3, 30.6, 39.0, 46.2,
63.0, 66.3 (aliphatic); 120.7, 122.4, 123.4, 123.0, 139.8 (aryl);
166.0 (CdO).
10-HCl: 95% yield; mp 252-253 °C; 1H NMR (D2O) δ 0.80-
1.10 (m, 1H), 1.08-1.38 (m, 1H), 1.56-1.67 (m, 4H), 2.00 (s,
3H, CH3), 2.17-2.25 (m, 1H, C-4), 2.58-2.62 (m, 7H, N(CH3)2
and C-1), 3.01 (d, J ) 6.0 Hz, 1H, C-5), 3.67 (d, J ) 6.0 Hz,
1H, C-6), 4.94 (br d, J ) 10 Hz, 1H, C-2), 7.19-7.40 (m, 5H,
Ar); IR 1749, 1249, 1038 cm-1. Anal. (C18H26O2NCl) C, H, N,
Cl.
(1R/S,4R/S)-6S/R-(N,N-Dim et h yla m in o)-5S/R-p h en yl-
bicyclo[2.2.2]oct-2S/R-yl a ceta te (17): 92% yield; 1H NMR
(CDCl3) δ 1.32-1.49 (m, 3H), 1.59 (br s, 1H), 1.68-1.78 (m,
2H), 1.90-1.99 (m, 1H), 2.04 (m, 6H, N(CH3)2), 2.12 (s, 3H,
CH3), 2.20 (d, J ) 2.7 Hz, 1, C-1), 2.66 (br s, J ) 5.6 Hz, 1H,
C-5), 2.73 (br d, J ) 5.7 Hz, 1H, C-6), 4.95 (dt, J ) 10.2 Hz, J
) 3.6 Hz, C-2), 7.15-7.28 (m, 5H, Ar); 13C (CDCl3) δ 11.4, 15.9,
20.5, 22.7, 26.7, 29.4, 38.8, 45.9, 56.2, 66.7 (aliphatic); 120.5,
122.9, 122.9, 140.1 (aryl); 165.3 (CdO).
7-HCl: 84% yield; mp 296-297 °C; 1H NMR (D2O) δ 1.10-
1.37 (m, 4H), 1.51-1.56 (m, 1H), 1.91-1.95 (m, 1H), 2.12-
2.22 (m, 2H), 2.56 (br s, 6H, N(CH3)2), 2.73-2.76 (m, 1H, C-5),
3.67 (dd, J ) 15.5 Hz, J ) 6.6 Hz, C-6), 4.02 (br s, 1H, C-2),
7.09-7.35 (m, 5H, Ar); IR 3381, 2703, 2479, 1077 cm-1. Anal.
(C16H24ONCl) C, H, N, Cl.
(1R/S,4R/S)-6R/S-(N,N-Dim et h yla m in o)-5R/S-p h en yl-
bicyclo[2.2.2]octa n -2S/R-ol (8). To a solution of 6 (1.49 g,
6.12 mmol) in absolute EtOH (50 mL) was added slowly NaBH4
(0.464 g, 12.3 mmol) and the mixture was heated at reflux
under nitrogen for 3 h. The resulting solution was poured onto
ice, EtOH was removed under reduced pressure, and the
aqueous solution was extracted with CH2Cl2 (3 × 10 mL). The
organic extracts were combined, dried over MgSO4, and filtered
and the solvent was removed under reduced pressure to afford
8 as a white solid (1.50 g, 100% yield): 1H NMR (CDCl3) δ
0.91-0.95 (m, 1H), 1.31-1.56 (m, 5H), 1.75 (dd, J ) 14.4 Hz,
J
) 1.6 Hz, 1H), 1.91-2.00 (m, 1H), 2.05-2.07 (m, 6H,
N(CH3)2), 2.43 (d, J ) 1.2 Hz, 1H), 2.65 (dd, J ) 6.5 Hz, J )
1.6 Hz, 1H, C-5), 2.99 (d, J ) 6.5 Hz, 1H, C-6), 3.92-3.96 (m,
1H, C-2), 7.05-7.25 (m, 5H, Ar); 13C NMR (CDCl3) δ 12.3, 16.6,
25.7, 28.9, 35.7, 38.4, 44.1, 62.4, 64.0 (aliphatic); 120.6, 122.6,
123.0, 140.1 (aryl).
8-HCl: 87% yield; mp 245-247 °C; 1H NMR (D2O) δ 0.98-
1.02 (m, 1H), 1.26-1.28 (m, 1H), 1.49-1.60 (m, 1H), 1.63 (s,
3H), 1.97-2.06 (m, 1H), 2.40 (br s, 4H), 2.50 (br s, 1H), 2.83
(s, 2H), 3.10 (d, J ) 6.9 Hz, 1H, C-5), 3.65 (d, J ) 6.9, 1H,
C-6), 4.07 (d, J ) 9.3 Hz, 1H, C-2), 7.21-7.35 (m, 5H, Ar); IR
3335-3295, 1150, 1031 cm-1. Anal. (C16H24ONCl‚0.07H2O) C,
H, N, Cl.
6-(N,N-Dim eth yla m in o)-5-p h en ylbicyclo[2.2.2]octa n -2-
ols 14 a n d 15. Treatment of 13 according to the procedure
for the synthesis of 8 afforded a mixture of 14 and 15 which
was triturated with EtOAc (10 mL) and filtered to give 14 as
a white solid (1.26 g, 57%). The solvent was removed from the
filtrate to afford 15 as a white solid (0.95 g, 43%).
(1R/S,4R/S)-6S/R-(N,N-Dim eth yla m in o)-5S/R-p h en ylbi-
cyclo[2.2.2]-octa n -2R/S-ol (14): 1H NMR (CDCl3) δ 1.09 (d,
J ) 14.1 Hz, 1H), 1.56 (d, J ) 2.1 Hz, 1H), 1.64-1.95 (m, 5H),
2.10 (m, 7H, N(CH3)2), 2.31 (d, J ) 3.6 Hz, 1H, C-5), 2.70 (d,
J ) 4.8 Hz, 1H, C-6), 3.6 (br s, 1H, OH), 4.01 (dd, J ) 3.3 Hz,
J ) 6.6 Hz, 1H, C-2), 7.10-7.30 (m, 5H, Ar); 13C NMR (CDCl3)
δ 6.9, 21.7, 26.0, 28.9, 30.2, 39.1, 45.5, 62.1, 63.3 (aliphatic);
120.5, 122.5, 123.0, 140.3 (aryl).
14-HCl: 86% yield; mp 250-251 °C; 1H NMR (D2O) δ 0.93
(br d, J ) 14.7 Hz, 1H), 1.32-1.37 (m, 1H), 1.55-1.60 (m, 3H),
1.73-1.76 (m, 3H), 2.19 (br s, 1H), 2.57-2.79 (m, 6H, N(CH3)2),
3.51 (d, J ) 4.5 Hz, 1H, C-6), 4.00 (d, J ) 9.9 Hz, 1H, C-2),
7.01-7.25 (m, 5H, Ar); IR 3368-3328, 2683, 1170, 1097 cm-1
Anal. (C16H24ONCl) C, H, N, Cl.
.
(1R/S,4R/S)-6S/R-(N,N-Dim et h yla m in o)-5S/R-p h en yl-
bicyclo[2.2.2]octa n -2S/R-ol (15): 1H NMR (CDCl3) δ 1.32-
1.41 (m, 3H), 1.45-1.52 (m, 3H), 1.60-1.80 (m, 3H), 1.95-
2.02 (m, 2H), 2.11 (m, 7H, N(CH3)2), 2.67 (d, J ) 5.9 Hz, 1H,
C-5), 2.92 (d, J ) 5.9 Hz, 1H, C-6), 4.06-4.08 (m, 1H, C-2),
7.18-7.30 (m, 5H, Ar); 13C NMR (CDCl3) δ 11.9, 20.7, 25.3,
29.8, 30.1, 38.9, 46.4, 55.8, 63.8 (aliphatic); 120.3, 122.9, 123.2,
140.5 (aryl).

4842 J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 23
J avanmard et al.
17-HCl: 80% yield; mp 118-120 °C; 1H NMR (D2O) δ 1.26-
1.32 (m, 1H), 1.48-1.77 (m, 6H), 2.04 (s, 3H, CH3), 2.41 (br s,
1H, C-5), 2.60 (br s, 6H, N(CH3)2), 2.85 (d, J ) 6 Hz, 1H, C-6),
4.90 (dt, J ) 9.9 Hz, J ) 3.9 Hz, 1H, C-2), 7.15-7.28 (m, 5H,
Ar); IR 1755, 1262, 1038 cm-1. Anal. (C18H26O2NCl‚2.5H2O)
C, H, N, Cl.
(1R/S,4R/S)-6R/S-(N,N-Dim et h yla m in o)-5R/S-p h en yl-
bicyclo[2.2.2]oct-2R/S-yl ben zoa te (11): 90% yield; 1H NMR
(CDCl3) δ 1.33-1.56 (m, 3H), 1.72 (br s, 1H), 1.77-1.78 (m,
1H), 2.06-2.17 (m, 1H), 2.21-2.22 (m, 6H, N(CH3)2), 2.42-
2.46 (m, 1H), 2.51-2.53 (m, 2H, C-5 and C-1), 2.79 (br s, 1H,
C-6), 5.54 (br d, J ) 9.9 Hz, 1H, C-2), 7.19-8.22 (m, 10H, Ar);
13C (CDCl3) δ 13.2, 13.3, 26.4, 28.9, 30.4, 38.9, 45.8, 62.4, 64.0
(aliphatic); 120.6, 122.7, 123.0, 123.6, 124.1, 125.2, 125.5,
127.4, 129.2, 139.8 (aryl); 161.1 (CdO).
11-HCl: 82% yield; mp 261-262 °C; 1H NMR (D2O) δ 1.29-
1.41 (m, 2H), 1.45-1.61 (m, 1H), 1.73 (br s, 1H), 1.75-1.77
(m, 1H), 2.04-2.18 (m, 1H), 2.28-2.38 (m, 1H), 2.45-2.58 (m,
2H), 2.62 (br s, 1H), 2.83-2.95 (m, 4H), 3.18 (br s, 1H, C-5),
3.78-3.85 (m, 1H, C-6), 5.19-5.24 (m, 1H, C-2), 7.19-8.01 (m,
10H, Ar); IR 2677-2473, 1722, 1275, 1117 cm-1. Anal.
(C23H28O2NCl‚0.33H2) C, H, N, Cl.
(1R/S,4R/S)-6R/S-(N,N-Dim et h yla m in o)-5R/S-p h en yl-
bicyclo[2.2.2]oct-2S/R-yl ben zoa te (12): 94% yield; 1H NMR
(CDCl3) δ 1.15 (br s, 1H), 1.38-1.57 (m, 1H), 1.60-1.79 (m,
3H), 1.90 (br s, 1H, C-4), 2.10 (br s, 6H, N(CH3)2), 2.22-2.41
(m, 1H), 2.50 (br s, 2H, C-5 and C-1), 3.09 (br s, 1H, C-6), 5.20
(br s, 1H, C-2), 7.10-7.58 (m, 8H, Ar), 8.1 (s, 2H, Ar); 13C NMR
(CDCl3) δ 12.4, 18.6, 25.9, 28.5, 31.3, 39.4, 46.6, 63.0, 67.0;
120.6, 122.6, 122.9, 123.0, 124.5, 125.7, 127.3, 140.1 (aryl);
161.6 (CdO).
ference in the KD or Bmax of [3H]WIN binding in unanesthetized
rats versus anesthetized rats; data not shown.) Their brains
were quickly removed and placed in ice-cold 0.32 M sucrose.
The striatal tissue was removed and homogenized in 20
volumes of 0.32 M sucrose, using 10 up/down strokes of a
motorized Potter-Elvejhm homogenizer. The supernatant ob-
tained after centrifugation for 10 min at 0 °C (S1 fraction) was
removed and centrifuged for 20 min at 20000g and 0 °C to
obtain the P2 fraction, which was then resuspended in 50
volumes (original wet weight) of ice-cold 25 mM sodium
phosphate buffer (pH 7.7) using a Tekmar tissuemizer. Samples
containing 750 µL phosphate buffer, 150 µL of the P2 suspen-
sion, 50 µL of the test compound, 25 µL of water or amfonelic
acid (to define nonspecific binding; final concentration, 10 µM)
and 25 µL of [3H]WIN (final concentration, 2 nM) were
incubated for 2 h at 0 °C. The incubation was terminated by
vacuum filtration through Whatman GF/B filters presoaked
with 0.05% (w/v) poly(ethylenimine), mounted in Millipore
filtration manifolds. A 5-mL aliquot of assay buffer was added
to the sample immediately before filtering it, and a second
5-mL aliquot of assay buffer was used to wash the filter. The
filters were transferred to scintillation vials, shaken vigorously
in the presence of 8 mL of Beckman Ready-Safe scintillation
fluid for 30 min, and counted in a liquid scintillation counter.
IC50 values (that concentration of test compound required
to inhibit 50% of the control specific binding of [3H]WIN) were
determined from dose-response curves usually containing a
range of seven concentrations of the test drug, with triplicate
determinations made at each concentration. The IC50 for WIN
was 22.2 ( 4.7 nM (average ( SEM) under these conditions.
[3H]DA Up ta k e. Accumulation of [3H]DA was determined
as previously described.18 Briefly, 250 µL of an S1 fraction
prepared as described above from striatal tissue of CO2-
anesthetized rats was diluted 4-fold with a modified Krebs-
phosphate buffer (120 mM NaCl, 4.9 mM KCl, 1.2 mM MgSO4,
11 mM glucose, 0.16 mM Na2EDTA, 1.1 mM ascorbic acid, 0.01
mM pargyline, and 15.5 mM Na2PO4, equilibrated with 95%
O2-5% CO2, and adjusted to pH 7.4 with NaOH) and prein-
cubated with 1100 µL of Krebs-phosphate buffer and 100 µL
of vehicle or drug for 10 min at 37 °C. A solution of [3H]DA
hydrochloride (50 µL; DuPont/NEN, Boston, MA, or Amersham
Corp., Arlington Heights, IL) which had been previously
diluted with sufficient unlabeled DA-HCl to bring the specific
activity to approximately 5 Ci/mmol was added to give a final
concentration of DA of about 30 nM. The samples were exposed
to the [3H]DA for exactly 2.0 min. Nonspecific dopamine
transport was determined by following the same protocol at 0
°C. Accumulation of [3H]DA was terminated by rapid addition
of 5 mL of the chilled Krebs-phosphate buffer to each sample,
followed by filtration through a Whatman GF/C filter under
vacuum, after which the filter was washed with an additional
5-mL aliquot of buffer. The filters were extracted, and the
trapped radioactivity was quantified as described for the
[3H]WIN binding assay.
12-HCl: 75% yield; mp 260-261 °C; 1H NMR (D2O) δ 1.05-
1.20 (m, 1H), 1.21-1.39 (m, 1H), 1.72 (br s, 4H), 2.35-2.44
(m, 1H), 2.49 (s, 3H, NCH3), 2.73 (br s, 1H, C-1), 2.97 (s, 3H,
NCH3), 3.10 (br s, 1H, C-5), 3.79 (br d, 1H, J ) 5.4 Hz, C-6),
5.08-5.20 (m, 1H, C-2), 7.19-7.81 (m, 10H, Ar); IR 1722, 1277,
1113 cm-1. Anal. (C23
H28O2NCl‚0.66H2O) C, H, N, Cl.
(1R/S,4R/S)-6S/R-(N,N-Dim et h yla m in o)-5S/R-p h en yl-
bicyclo[2.2.2]oct-2R/S-yl ben zoa te (18): 98% yield; 1H NMR
(CDCl3) δ 1.25-1.39 (m, 1H), 1.67 (br s, 1H), 1.75-1.87 (m,
4H), 2.16-2.21 (m, 7H, N(CH3)2 and C-4), 2.41 (br s, 1H, C-1),
2.50 (dd, J ) 5.4 Hz, J ) 3.3 Hz, 1H, C-5), 2.80 (br s, 1H,
C-6), 5.27 (dt, J ) 9.9 Hz, J ) 3.3 Hz, 1H, C-2), 7.19-8.20 (m,
10H, Ar); 13C NMR (CDCl3) δ 7.7, 21.5, 23.4, 27.0, 28.6, 39.0,
45.5, 61.0, 67.1 (aliphatic); 120.7, 122.6, 123.0, 123.1, 123.5,
124.2, 125.2, 127.6, 129.2, 140.0 (aryl); 160.9.
18-HCl: 85% yield; mp 273-275 °C; 1H NMR (D2O) δ 1.28-
1.33 (m, 1H), 1.48-1.55 (m, 1H), 1.69-1.73 (m, 3H), 1.91-
2.01 (m, 2H), 2.55-2.59 (m, 4H), 2.79 (s, 3H), 2.92 (d, J ) 5.7
Hz, 1H, C-5), 3.73 (d, J ) 5.7 Hz, 1H, C-6), 5.18 (m, 1H, C-2),
7.19-7.98 (m, 10H, Ar); IR 1722, 1275, 1123 cm-1. Anal.
(C23H28O2NCl‚0.33H2O) C, H, N, Cl.
(1R/S,4R/S)-6S/R-(N,N-Dim et h yla m in o)-5S/R-p h en yl-
bicyclo[2.2.2]oct-2S/R-yl ben zoa te (19): 91% yield; 1H NMR
(CDCl3) δ 1.54-1.62 (m, 2H), 1.66-1.72 (m, 2H), 1.80-1.94
(m, 2H), 2.02-2.14 (m, 7H, N(CH3)2 and C-1), 2.43 (t, J ) 2.4
Hz, 1H), 2.79 (d, J ) 6.15 Hz, 1H, C-5), 2.94 (d, J ) 6.15 Hz,
The IC50 values were determined from dose-response curves
usually containing a range of five concentrations of test drug,
with [3H]DA uptake measured in duplicate samples at each
concentration.
1H, C-6), 5.22-5.29 (m, 1H, C-2), 7.25-8.19 (m, 10H, Ar); 13
C
[3H]P AR17 a n d [3H]NIS24,25 Bin d in g. Brains from male
Sprague-Dawley rats weighing 200-250 g (Harlan Labs,
Indianapolis, IN) were removed, dissected, and rapidly frozen.
Ligand binding experiments for the serotonin transporter were
conducted in assay tubes containing 4 mL of buffer (50 mM
Tris, 120 mM NaCl, 0.5 mM KCl, pH 7.4 at 25 °C) for 90 min
at room temperature. Each assay tube contained 0.2 nM
[3H]PAR and 1.5 mg of hindbrain tissue (original wet weight).
Nonspecific binding of [3H]PAR was defined by 1 µM citalo-
pram. Ligand binding experiments for the norepinephrine
transporter were conducted in Tris buffer (50 mM Tris, 120
mM NaCl, 0.5 mM KCl, pH 7.4 at 4 °C) at a total volume of
0.5 mL for 60 min. Each assay tube contained 0.5 nM [3H]NIS
and 8 mg of frontal cerebral cortex. Nonspecific binding of
[3H]NIS was defined using 1 µM desiprimine. Incubations were
terminated by filtration with two 5-mL washes of ice-cold
NMR (CDCl3) δ 11.5, 20.6, 22.9, 27.0, 29.6, 38.9, 45.8, 56.7,
67.5 (aliphatic); 120.6, 122.9, 123.0, 123.2, 124.2, 125.4, 127.7,
140.1 (aryl); 160.8 (CdO).
19-HCl: 91% yield; mp 163-165 °C; 1H NMR (D2O) δ 1.36-
1.42 (m, 1H), 1.53-1.70 (m, 5H), 1.77-1.90 (m, 1H), 2.50 (br
s, 3H, NCH3), 2.76 (br s, 3H, NCH3), 2.91 (br s, 1H, C-5), 3.91-
4.00 (m, 1H, C-6), 5.05-5.18 (m, 1H, C-2), 7.15-7.97 (m, 10H,
Ar); IR 1729, 1281, 1117 cm-1. Anal. (C23H28O2NCl‚0.66H2O)
C, H, N, Cl.
P h a r m a cology. [3H]]WIN Bin d in g. The synthesized com-
pounds were screened for activity in a striatal tissue prepara-
tion using a modification of the [3H]WIN binding assay
described by Reith and Selmeci.16 Male Sprague-Dawley rats
(Harlan Sprague-Dawley, Indianapolis, IN) weighing 150-
300 g were anesthetized using CO2 gas and sacrificed by
decapitation. (Preliminary experiments demonstrated no dif-

Site-Specific Cocaine Abuse Treatment Agents
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Ack n ow led gm en t. This research was supported in
part by the National Institute on Drug Abuse, Grant
DA06305 (H.M.D. and M.M.S.); Grants DA00418 and
RR00165 (M.J .K.); and Predoctoral Fellowship DA05869
(S.J .). The technical assistance of Ms. Monica Stafford
and Mr. Richard Hunter is much appreciated.
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