Pyrrolidine analogues of lobelane: Relationship of affinity for the dihydrotetrabenazine binding site with function of the vesicular monoamine transporter 2 (VMAT2)

Article abstract of DOI:10.1021/jm900770h

Ring size reduction of the central piperidine ring of lobelane yielded pyrrolidine analogues that showed marked inconsistencies in their ability to bind to the dihydrotetrabenazine (DTBZ) binding site on the vesicular monoamine transporter-2 (VMAT2) and t

Full text of DOI:10.1021/jm900770h

7878 J. Med. Chem. 2009, 52, 7878–7882
DOI: 10.1021/jm900770h
Pyrrolidine Analogues of Lobelane: Relationship of Affinity for the Dihydrotetrabenazine Binding Site
with Function of the Vesicular Monoamine Transporter 2 (VMAT2)†
Ashish P. Vartak, Justin R. Nickell, Jaturaporn Chagkutip, Linda P. Dwoskin, and Peter A. Crooks*
Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 725 Rose Street, Lexington, Kentucky 40536
Received May 31, 2009
Ring size reduction of the central piperidine ring of lobelane yielded pyrrolidine analogues that showed marked
inconsistencies in their ability to bind to the dihydrotetrabenazine (DTBZ) binding site on the vesicular mono-
amine transporter-2 (VMAT2) and their ability to inhibit VMAT2 function. The structure-activity relation-
ships indicate that structural modification within the pyrrolidine series resulted in analogues that interact with two
different sites, i.e., the DTBZ binding site and an alternative site on VMAT2 to inhibit transporter function.
Introduction
possessing little affinity for R4β2* nicotinic acetylcholine receptors
(nAChRs), to which 1 binds with high potency.8 The cis-2,6-
diethylarylpiperidine structural motif in 2 was therefore regarded
as an initial pharmacophore on which to develop subsequent
generations of VMAT2 inhibitors. Initial structure-activity stu-
dies on 2 were focused on the incorporation of substituents into
the aromatic rings, variation in the number of methylenes between
the central piperidine ring and the phenyl rings, and the position-
ing of the two phenethyl moieties at different sites around the
central piperidine ring. Modest increases in affinity for the DTBZ
binding site on VMAT2 were obtained through substitution of
anisyl and naphthyl moieties for phenyl moieties in the phenethyl
side chains,7 while departure from the 2,6-diphenethyl substitution
pattern around the piperidine ring was found to be generally
deleterious.9 A minimum of two methylene units in the alkane
linker moiety was found to be essential for retention of binding
affinity exhibited by 2at the DTBZ binding site on VMAT2. Last,
the trans-configured enantiomeric analogues of 2 (trans-2s, 4, and
5) exhibited diminished affinity for the DTBZ binding site, as did
N-demethylated analogues, including nor-2 (3) (Figure 1).
Design Rationale. Available SAR suggests that the intra-
molecular distance between the two aromatic rings in 2, and the
relative orientation of the phenethyl moieties, are determinants for
affinity at the DTBZ binding site on VMAT2. To improve
affinity, we investigated the effect of restricting the flexibility of
the central piperidine ring to afford more conformationally
defined analogues. In this respect, the current study investigates
the effect of replacing the central piperidine ring in 2 with a more
conformationally restricted pyrrolidine ring. A series of six pyrroli-
dine analogues 6-11 were chosen as the initial target molecules
(Figure 2). Compound 7 is a pyrrolidino analogue of 2, and the
trans-configured analogues 9 and 11 represent the pyrrolidino-
analogues of trans-2s 4, and 5. Analogues 6, 8, and 10 were also
included as analogues of nor-2 (3) to determine the importance of
the N-methyl group in the SAR of these pyrrolidine analogues.
The vesicular monoamine transporter-2 (VMAT2a) transports
monoamines from the neuronal cytosol into the synaptic vesicle,
a process that prevents metabolism by monoamine oxidase
(MAO) and stores monoamines, such as dopamine (DA), for
eventual release by exocytosis.1 Psychostimulants, such as
methamphetamine (METH), inhibit the function of VMAT2,
thereby increasing cytosolic levels of DA.2 METH also inhibits
MAO and reverses the DA transporter (DAT), which together
with the increased cytosolic levels of DA results in a substantial
efflux of neurotransmitter into the synaptic cleft. Thus, the
interaction of METH with VMAT2 is the first step in a series
of events that leads ultimately to its pharmacological effects,
including the psychological high that results from METH admin-
istration. The design and development of VMAT2- selective
inhibitors is therefore a worthwhile pursuit toward the broader
goal of developing efficacious therapeutics as a means to clinically
intervene in the abuse of psychostimulants such as METH.3
The piperidine alkaloid and principal constituent of Lobelia
inflata, lobeline (1), was found to inhibit the uptake of DA into
synaptic vesicles.4 Because lobeline and its analogues competi-
tively inhibit the binding of [3H]dihydrotetrabenazine (DTBZ),
a known VMAT2 inhibitor, this binding site was identified as a
target for the development of potential therapeutic agents for
the treatment METH abuse. Thus, the DTBZ binding site on
VMAT2 was utilized as a probe in preliminary screening of
lobeline analogues for potency and selectivity for VMAT2
interaction. Large increases in potency were achieved through
complete deoxygenation and reduction of the lobeline mole-
cule, which resulted in the identification of a lead compound,
cis-N-methyl-2,6-diphenethylpiperidine (2, lobelane).5-7
Compound 2, which is also a minor alkaloid of Lobelia inflata,
was found to be more selective than 1 as a VMAT2 ligand,
† We express profound gratitude to the ACS Division of Medicinal
Chemistry for its relentless efforts toward fostering research and training
in the discipline of Medicinal Chemistry over the past 100 years.
*To whom correspondence should be addressed. Phone: 859-257-
1718. Fax: 859-257-7585. E-mail: pcrooks@email.uky.edu.
a Abbreviations: VMAT2, vesicular monoamine transporter 2;
MAO, monoamine oxidase; DA, dopamine; METH, methampheta-
mine; DTBZ, dihydrotetrabenazine; TBZ, tetrabenazine; nAChRs,
nicotinic acetylcholine receptors.
Results
Scheme 1 shows the synthesis of the initial 2,5-diphenethyl-
pyrrolidine analogues. The absolute and relative stereochemis-
tries were achieved utilizing Katritzky’s elegant benzotriazole-
oxazolidine approach.10 Briefly, L- and D-phenylglycinol were
r
pubs.acs.org/jmc
Published on Web 08/19/2009
2009 American Chemical Society

Brief Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23 7879
both obtained through NaBH4 reduction of the corresponding
phenylglycine methyl esters. A three-component condensation
of L-phenylglycinol with benzotriazole and succinaldehyde
afforded the key synthon 12 in useful quantities. Addition of
a 6-fold excess of phenethyl magnesium bromide to synthon 12
at room temperature led to a slightly exothermic reaction,
affording a 2:1 mixture of the cis- and trans-configured
diastereomers of 13 within a few hours; these diastereomers
were separated by silica gel chromatography. The 2R,5S- and
2R,5R-diastereomers, 14 and 15, were hydrogenolyzed sepa-
rately by catalytic-transfer hydrogenation with palladium
hydroxide-over carbon, employing ammonium formate as
the hydrogen source in refluxing methanol. These conditions
afforded quantitative conversion to the respective products, 6
and 8, within 20 min, as opposed to 12 h utilizing alternate
conditions (i.e., H2/Pd-C).5 The resulting meso- and (þ)-
trans-2R,5R pyrrolidines 6 and 8 were then each N-methylated
with paraformaldehyde and sodium triacetoxyborohydride/
acetic acid in acetonitrile followed by treatment with HCl in
diethyl ether to afford the meso isomer 7 and the (þ)-trans-
2R,5R isomer 9, respectively. Utilization of D-phenylglycinol
in place of L-phenylglycinol in the above synthetic scheme
provided synthon 16, which could be converted into 18 and 19.
Hydrogenolysis of 19 followed by hydrochloride salt forma-
tion afforded 10 ((-)-trans-2S,5S), which in turn was
N-methylated to 11 ((-)-trans-2S,5S). Note that compound
18 also forms the cis-meso isomer 6 upon hydrogenolysis,
making further elaboration of this compound redundant.
The pyrrolidine analogues in Scheme 1 were evaluated for
affinity at the DTBZ binding site on VMAT2 (Table 1). When
the 2,5-diphenethyl substituents are in the meso cis-configura-
tion, overall binding affinity clearly decreases upon replacement
of the piperidine ring with the pyrrolidine ring, because 7is about
10 times less potent than 2 (2). When the phenethyl substituents
are in the 2,5-trans configuration (analogues 9 and 11), the
binding affinity does not appreciably change with azacyclic ring-
size reduction when compared to the piperidine (2) counterparts
4 and 5, respectively. Interestingly, the (S,S)-trans-nor-N-methyl
analogue 10 is about 10-fold more potent than its N-methyl
derivative 11, whereas replacement of the N-methyl group in
both the cis- and trans-analogues with an N-phenethanolyl
substituent afforded no significant change in binding affinity,
with analogues 14, 15, 18, and 19 possessing affinities compar-
able to the nor- and N-methyl pyrrolidine analogues.
The ultimate goal of this SAR study was to identify chemical
entities that would inhibit VMAT2 function. It is known
that there are multiple binding sites on VMAT2 that may
influence transporter function, i.e., a ketanserine binding site near
the N-terminus, a tetrabenazine (TBZ) binding site near the
C-terminus, and two reserpine binding sites (a high affinity site
and a low affinity site).11 In this respect, the affinity of a ligand
toward the DTBZ binding site on VMAT2 may not necessarily be
a sufficient indicator of the activity of the ligand in altering function
of VMAT2. In light of these data and the possibility that other, as
yet unidentified, binding sites on VMAT2 may exist, the ability
of the current analogues to inhibit the uptake of [3H]-DA into
isolated synaptic vesicles was also evaluated (Table 1).
Figure 1. Structures of 1, 2, and synthetic analogues.
While the affinities of the pyrrolidine ligands for the DTBZ
binding site did not vary much with respect to structural
change (Ki values were in the range of 0.3-14 μM), their
ability to inhibit the uptake of [3H]-DA into synaptic vesicles
varied greatly with subtle structural changes, with Ki values
ranging from 9 nM to 1.94 μM. Ring-size reduction of the
piperidine moiety in 2 afforded a 10-fold decrease in the
Figure 2. Structures of initial pyrrolidine analogues of 2 and nor-2.
Scheme 1. Synthesis of 2,5-Diphenethylpyrrolidines

7880 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23
Table 1. Interaction of 2 and Pyrrolidine Analogues with VMAT2
Vartak et al.
Scheme 2. Synthesis of Second-Generation Pyrrolidine Analogues
[3H]DTBZ;
Ki ( SEM
(μM)
VMAT2 [3H]-DA
uptake
ratio [3H]DTBZ
binding/[3H]-
DA uptake
analogue
Ki ( SEM (μM)
TBZ
1
0.013 ( 0.001
2.76 ( 0.64
0.97 ( 0.19
2.31 ( 0.21
5.32 ( 0.45
6.46 ( 1.70
3.06 ( 0.17
8.80 ( 2.30
3.40 ( 0.030
3.97 ( 0.64
1.33 ( 0.19
14.45 ( 2.13
4.34 ( 0.71
5.05 ( 1.05
2.75 ( 0.63
5.61 ( 0.91
1.64 ( 0.12
2.46 ( 0.42
0.27 ( 0.0067
2.78 ( 0.63
0.054 ( 0.015
0.470 ( 0.045
0.033 ( 0.041
0.023 ( 0.004
4.05 ( 0.612
0.24
5.87
29.4
100
2
3
4
1.31
3.06
106
5
2.11 ( 0.50
6
0.029 ( 0.007
0.270 ( 0.035
0.054 ( 0.017
0.120 ( 0.015
0.035 ( 0.006
0.170 ( 0.030
1.290 ( 0.240
1.940 ( 0.470
0.140 ( 0.049
0.270 ( 0.053
0.0093 ( 0.00075
0.019 ( 0.00091
0.014 ( 0.0011
0.12 ( 0.011
to bind to the DTBZ binding site on VMAT2 (Ki 2.75 and
5.61 μM, respectively) and their ability to inhibit [3H]-DA
uptake into vesicles (Ki 140 and 270 nM, respectively). Pyrro-
lidines 18 and 19 are more potent as [3H]-DA uptake inhibitors,
compared to their corresponding 20S-configured analogues
14 (Ki (binding)=4.34 μM, Ki (uptake)=1.29 μM)) and 15
(Ki (binding)=5.05 μM, Ki (uptake)=1.94 μM).
7
32.6
63.0
33.1
38.0
85.0
3.36
2.60
19.6
20.7
176
8
9
10
11
14
15
18
19
22
23
25
26
cis-2,5-Diphenethylpyrrolidine (6) was identified as the
most potent pyrrolidine analogue for the inhibition of
VMAT2 function, and structural optimization of this lead
molecule was further investigated. Thus, compound 22, in
which the methylene linker moieties have been truncated to a
single methylene unit, as well as compound 25, in which the
methylene units have been increased from two to three, were
synthesized. The synthesis of pyrrolidine 22 was approached
by utilizing the method illustrated in Scheme 2. Treatment of
12 with benzyl magnesium chloride led to an inseparable ∼3:1
mixture of diastereomers with the 2,5-cis-configured diaster-
eomer 21 being the predominant isomer. Because diastereo-
mer 21 was required for further elaboration, conditions were
identified to increase the proportion of this diastereomer in
the reaction product. During the synthesis of the 2,5-diphe-
nethyl pyrrolidines, it was noted that the presence of MgBr2 in
the reaction mixture (originating from the activation of Mg
turnings by dibromoethane) resulted in diastereomeric ratios
that favored the 2,5-cis diastereomers (14 and 18) over the
2,5-trans diastereomers (15 and 19). A solution of 12 was
treated with progressively increasing equivalents of MgBr2
(generated in situ from Mg and dibromoethane) prior to the
addition of benzylmagnesium chloride. When 10 equiv of
MgBr2 had been utilized, the product mixture consisted solely
of the free-base of21 (the trans-configured diastereomer could
130
19.3
23.2
potency to inhibit [3H]-DA uptake into vesicles when com-
pared to 2 (meso cis-analogues 7 and 2) However, a similar
ring-size reduction in nor-2 had no effect on inhibition of
[3H]-DA uptake because 3 and 6 were equipotent. The most
striking difference in [3H]-DA uptake inhibition was observed
between the trans-configured piperidine- and pyrrolidine-
analogues. Thus, (S,S)-trans-2 (4) (Ki = 4.05 μM) and
(R,R)-trans-2 (Ki=2.11 μM) (5) are 33- and 12-fold, respec-
tively, less potent than their corresponding pyrrolidine analo-
gues 9 (Ki = 120 nM) and 11 (Ki = 170 nM), respectively.
Within the pyrrolidine series, the presence of the N-methyl
group is clearly deleterious toward potency of inhibition of
[3H]-DA uptake because the N-methyl pyrrolidines 7, 9, and
11areall less potent than theirnor-N-methyl counterparts6, 8,
and 10, with the effect of N-methyl substitution being most
prominent in the cis-configured analogues 6 and 7. Also,
within the N-phenethanolyl pyrrolidine series of analogues,
the configuration of the pendant phenethanolyl stereocenter
appears to be a factor that contributes to the potency for [3H]-
DA uptake inhibition because the R-configured analogues 18
and 19 are about 10-fold more potent than the S-configured
analogues 14 and 15.
A marked disparity exists in the abilities of the above series
of analogues to bind to the DTBZ binding site on VMAT2
and to inhibit [3H]-DA uptake into vesicles, as can be seen in
the ratios of the Ki values, as well as in the rank order of
potency of the compounds in the two assays; this is evident for
both the piperidine and pyrrolidine analogues (Table 1, right
column). Compound 3 and its pyrrolidine analogue 6 are both
about 100-fold more potent in inhibiting [3H]-DA uptake into
isolated synaptic vesicles than in binding to the DTBZ-bind-
ing site, while their N-methyl counterparts 2 and 7 are both
about 30 times more potent in this regard. While the N-methyl
trans-configured pyrrolidines 9 and 11 are both significantly
more potent in inhibiting [3H]-DA uptake than in binding to
the DTBZ binding site, their piperidine counterparts (i.e., the
trans-2 enantiomers 4 and 5) are relatively weak [3H]-DA
uptake inhibitors (Ki>1 μM) and show relatively low po-
tency in the DTBZ binding assay (Kis 5.32 and 6.46 μM,
respectively). In the N-phenethanolyl series, the 20R-analo-
gues 18 and 19 exhibit significant differences in their ability
1
not be detected by H NMR or by gas chromatography),
however, this favorable exclusive formation of the desired
diastereomer was achieved at the expense of a significantly
lower yield (42%), as compared to the 80% yield obtained in
the reaction without MgBr2. Hydrogenolysis of 21 led to 22,
which was N-methylated and treated with HCl in diethylether
to afford 23. The diastereomerically pure propyl homologue
25 and its N-methylated derivative 26 were prepared in a
similar manner by utilizing the aforementioned procedure
employing phenpropyl magnesium bromide.
The methylene truncated analogues 22 and 23 bound to the
DTBZ-binding site of VMAT2 with affinities comparable to
that of the parent pyrrolidines 6 and 7, whereas the methylene
elongated nor-analogue 25, bound with high affinity (Ki =
270 nM), making it one of the most potent 2-derived ligands
known for this binding site. N-Methylation of 25 afforded 26,
which was 10 times less potent than 25.
Pyrrolidine 22 was the most potent [3H]-DA uptake inhibitor
in this series of analogues, with a Ki of 9.3 nM. Its N-methylated
derivative 23 was also a good uptake inhibitor (Ki=19 nM).
Pyrrolidine 25 was also a potent uptake inhibitor (Ki=14 nM);
its N-methylated analogue 26 was about 10 times less potent.

Brief Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23 7881
As seen in Table 1, the disparity between [3H]-DTBZ binding
and [3H]-DA uptake inhibition is more prominent for the
methylene truncated pyrrolidines 22 and 23, which inhibit
[3H]-DA uptake 176- and 129-fold more potently than they
inhibit [3H]-DTBZ binding to VMAT2. The methylene elon-
gated pyrrolidines 25 and 26 inhibit [3H]-DA uptake about
20 times more potently than they inhibit DTBZ binding,
irrespective of their relative potencies in the individual assays.
Figure 3. Pyrrolidine 22 may be regarded structurally a “dimer” of
methamphetamine.
yellow oil. The oil was dissolved in 20 mL of dry diethyl ether
through which HCl gas had been bubbled with stirring for 10 min.
The supernatant was decanted, and the resulting gummy sediment
was triturated with diethyl ether (5 ꢀ 50 mL). Aqueous NaOH
solution (2N, 20 mL) was then added, and the suspension extracted
with diethyl ether (2ꢀ100 mL). The organic layers were washed with
brine, dried, and the oily residue was applied to a silica gel column
(10 g). The column was eluted with 15:1 (hexanes/diethyl ether) to
afford pure fractions of compounds 14 (240 mg, 60%) and 15
(60 mg, 15%) as colorless oils. 14 Rf = 0.4 (8:1, hexanes/EtOAc).
1H NMR (300 MHz, CDCl3) δ ppm 7.45-7.03 (m, 15H, Ar),
4.02-3.83 (m, 2H), 3.78-3.60 (m, 1H), 3.35-3.20 (br, s, 1H),
Discussion
In summary, ring size reduction of the central piperidine
ring of 2 yielded pyrrolidine analogues that showed marked
disparities in their ability to bind to the [3H]-DTBZ binding
site on VMAT2 and their ability to inhibit VMAT2 function.
Unlike TBZ, the compounds in this series of pyrrolidine
analogues of 2were more potent in inhibiting VMAT2 function
than they were in inhibiting the binding of [3H]-DTBZ to
VMAT2. The SAR indicate that structural modification within
the pyrrolidine series results in analogues that likely interact
with two different sites on VMAT2, i.e., the DTBZ binding site,
and perhaps an alternative site on VMAT2, to inhibit trans-
porter function. While the affinity of the pyrrolidine analogues
toward the DTBZ binding site is not significantly sensitive to
structural changes, the potency for [3H]-DA uptake inhibition
is sensitive to subtle structural changes, such as alteration in
methylene linker length, N-substitution, and stereochemistry in
the case of the N-phenethanolyl analogues. Such sensitivity
suggests that the putative binding site on VMAT2 whose
interaction with the pyrrolidine analogues leads to functional
inhibition, is topographically distinct from the DTBZ ligand
binding site. It should be noted that compound 22, which has
the greatest potency to inhibit [3H]-DA uptake into synaptic
vesicles, can be considered a structural dimer of methamphe-
tamine (Figure 3), which is itself known to inhibit VMAT2
function.12 In this respect, 2 (2) could also be considered to be a
“dimeric” methamphetamine analogue; however, this com-
pound does not itself support self-administration but rather
decreases methamphetamine self-administration in animal
models.13
The results of this study are expected to have a significant
impact on the future design of lobelane-derived VMAT2 inhibi-
tors. Due caution needs to be exercised in utilizing the SAR from
[3H]-DTBZ-binding assays for the design of lobelane-derived
VMAT2 inhibitors because such data may not be functionally
relevant. The pyrrolidine ring has been identified as a valid
replacement for the piperidine ring in 2, and nor-analogues (i.e.,
N-demethylated homologues of 2) appear to be optimal for both,
binding to the DTBZ binding site on VMAT2 and inhibition of
VMAT2 function. A potent VMAT2 ligand, 25, and a potent
water-soluble inhibitor of VMAT2 function (22) were also
identified during this study, which appear to be suitable for
further development and advancement into behavioral studies.
3.15-2.95 (m, 2H), 2.80-2.45 (m, 4H), 1.95-1.20 (m, 10H). 13
C
NMR (75 MHz, CDCl3) δ ppm 143.7, 137.2, 129.1, 128.5, 128.5,
127.9, 125.9, 64.8, 63.6, 61.9, 57.6, 38.1, 36.4, 29.8, 29.2, 28.9.
20
[R]D -13.2° (c 1.0, CH2Cl2). Anal. C28 H33NO (C,H,N) 15
Rf = 0.2 (8:1, hexanes/EtOAc). H NMR (300 MHz, CDCl3)
1
δ ppm 7.50-6.98 (m, 15H), 4.30-3.92 (m, 2H), 3.84-3.79 (m, 1H),
3.32-3.16 (br, s, 1H), 3.17-2.92 (m, 2H), 2.73-2.56 (m, 4H),
2.03-1.20 (m, 10H). 13C NMR (75 MHz, CDCl3) δ ppm 146.3,
138.2, 125.4, 127.9, 128.8, 126.1, 125.4, 122.1, 66.2, 61.6, 57.6, 38.1,
29.2, 28.9. [R]D20 -2.0° (c 1.0, CH2Cl2). Anal. C28 H33NO (C,H,N).
(20R)-2-(2S,5R)-2,5-Di-(2-phenethyl)-tetrahydro-1H-1-pyrro-
lyl-2-phenylethan-1-ol (18). Compound 18 was synthesized from
1
16 and was the major diastereomer (55% yield). H and 13C
23
NMR spectra were identical to 3. [R]D þ12.1° (c 1.0, CH2Cl2).
Anal. C28 H33NO (C,H,N).
(20R)-2-(2S,5S)-2,5-Di(2-phenethyl)-tetrahydro-1H-1-pyrrolyl-
2-phenylethan-1-ol (19). Compound 19 was synthesized from 16 as
the minor diastereomer (20% yield). 1H and 13C NMR spectra were
20
identical to 4.[R]D þ2.2° (c1.0, CH2Cl2).Anal.C28 H33NO (C,H,N).
cis-2,5-Di-(2-phenethyl)-pyrrolidine Hydrochloride (6). To a
solution of 14 (100 mg, 0.25 mmol) in MeOH (5 mL) was added
Pd(OH)2-C (10 mg, 10 wt %) and ammonium formate (100 mg,
100 wt %). The mixture was refluxed for 20 min before being
diluted with CH2Cl2 (30 mL) and filtered through celite. The
filtrate was treated with 1 mL conc HCl and evaporated to a
crusty white solid under reduced pressure. The solid was tritu-
rated with diethyl ether and precipitated from a mixture of
CH2Cl2 and hexanes by addition of hexanes, to yield 6 (60 mg,
77%) as a white solid, mp=155 °C. 1H NMR (300 MHz, CDCl3)
δ ppm 10.22 (br, s, 1H), 9.03 (br, s, 1H), 7.32-7.08 (m, 10H),
3.62-3.43 (apparent s, br, 2H), 2.63-2.47 (m, 4H), 1.83-1.62
(m, 8H). 13C NMR (75 MHz, CDCl3) δ ppm 141.7, 128.6, 128.6,
126.2, 60.4, 41.1, 35.8, 29.0. Anal. C20H26ClN (C,H,N).
(2R,5R)-Di-(2-phenethyl)-pyrrolidine Hydrochloride (8). Com-
pound 15 (100 mg, 0.25 mmol) was prepared as described for the
preparation of 6, to yield 38 mg (48%) of 8 as a white solid, mp =
1
95 °C (deliquescent). H NMR (300 MHz, CDCl3) δ ppm 9.80
(br, s, 1H), 9.72 (br, s, 1H), 7.40-7.00 (m, 10H), 3.52-3.30 (s, br,
2H), 2.40-1.99 (m, 4H), 1.70-1.53 (m, 8H). 13C NMR (75 MHz,
CDCl3) δ ppm 140.0, 129.6, 128.5, 126.1, 58.3, 43.8, 33.0, 29.4.
[R]D20 = þ7.5° (c = 1.0, CH2Cl2). Anal. C20H26ClN (C,H,N).
(2S,5S)-di-(2-phenethyl)-pyrrolidine Hydrochloride (10). Com-
pound 10 was synthesized from 19 in a manner similar to that described
for 8 (73% yield). Spectral characteristics were identical to those of 8.
[R]D20=-8.1° (c=1.0, CH2Cl2). Anal. C20H26ClN (C,H,N).
1-Methyl-cis-2,5-di-(2-phenethyl)-pyrrolidine Hydrochloride (7).
Toasolutionof 6(50mg, 0.14 mmol) inCH3CN (5mL) andacetic
acid (1 mL) was added NaBH4 (50 mg, excess) and paraformal-
dehyde (50 mg, excess). The initial suspension afforded a clear
solution upon stirring for 12 h. The solution was then treated with
Experimental Section
(20S)-2-[(2R,5S)-2,5-Di-(2-phenethyl)-tetrahydro-1H-1-pyrrolyl]-
2-phenylethan-1-ol (14) and (20S)-2-[(2R,5R)-2,5-Di-(2-phenethyl)-
tetrahydro-1H-1-pyrrolyl]-2-phenylethan-1-ol (15). To 306 mg
(1 mmol) of 12 suspended in 20 mL of THF was added a solution
of phenethylmagnesium bromide in THF (6 equiv) over a period of
10 min at 0 °C. The solution was warmed to ambient temperature
over 8 h, 10 mL of 2N NaOH were added, and the biphasic mixture
was extracted with diethyl ether (3ꢀ100 mL). The etherial layers
were combined, diluted with 50 mL of CH2Cl2, dried with anhy-
drous Na2SO4 and evaporated under reduced pressure to afford a

7882 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23
Vartak et al.
conc HCl (0.5 mL), stirred for 2 h, poured into 50% aqueous
NaOH (10 mL), and extracted with CH2Cl2 (6 ꢀ 20 mL). The
CH2Cl2 extracts were combined, dried (anhydrous MgSO4),
filtered, and the solvent evaporated to yield an oil that was treated
with conc HCl (1 mL) and azeotroped twice with toluene.
Trituration of the residue with diethyl ether afforded a white solid
7 (43 mg, 83%), mp = 94 °C (deliquescent). 1H NMR (300 MHz,
CDCl3) δppm 9.48(br,s,1H), 7.25-7.02 (m, 10H), 3.73-3.48(m,
2H), 2.63-2.47 (m, 4H), 2.34 (s, 3H), 1.74-1.38 (m, 8H). 13C
NMR (75 MHz, CDCl3) δ ppm 144.8, 127.2, 127.1, 126.5, 65.2,
(1 mL) and stirred for 2 h, during which time all of the cyanobor-
ohydride and hydrogen cyanide was assumed to be decomposed.
The mixture was then poured into 20% aq NaOH (3 mL) and
extracted with CH2Cl2 (10 mL). Drying and evaporation of the
organic layer yielded an oil that was treated with conc HCl (1 mL)
and coevaporated with toluene. Trituration of the residue afforded a
white solid (18 mg, 87%), mp=78 °C. 1H NMR (300 MHz, CDCl3)
δ ppm 9.48 (s, 1H), 7.45-7.03 (m, 10H), 3.47-3.18 (m, 2H), 2.58-
2.47 (m, 4H), 2.19 (s, 3H), 1.91-1.36 (m, 6H). 13C NMR (75 MHz,
CDCl3) δ ppm 140.6, 128.2, 128.13, 128.02, 64.1, 37.6, 36.4, 29.5,
48.3, 38.3, 32.2, 30.1. Anal. C21H28ClN 0.5H2O (C,H,N).
29.2, 28.3. Anal. C23H32ClN H2O (C,H,N).
3
3
1-Methyl-(2R,5R)-di-(2-phenethyl)-pyrrolidine Hydrochloride
(9). Compound 8 was subjected to identical conditions to those
described for the preparation of 7toyield30mg(58%) of9as a white
solid, mp=113 °C. 1H NMR (300 MHz, CDCl3) δ ppm 9.90 (br, s,
1H), 7.30-6.99 (m, 10H), 3.84-3.65 (m, 2H), 2.50-2.23 (m, 4H),
Acknowledgment. We thank Agripina G. Deaciuc for
technical assistance and the National Institutes of Health,
NIDA grant DA013519, for financial support.
2.10 (s, 3H), 1.66-1.20 (m, 8H). 13CNMR(75 MHz, CDCl3) δppm
Supporting Information Available: Combustion analysis data
for all compounds, preparation and characterization data for
21 and 24 and procedures for the determination of [3H]-DTBZ
binding and [3H]-DA uptake inhibition. This material is avail-
able free of charge via the Internet at http://pubs.acs.org.
20
144.3, 127.06, 127.00, 126.9, 63.1, 45.3, 37.2, 31.6, 31.1. [R]D
=
þ46.2° (c=1.5, MeOH). Anal. C21H28ClN H2O (C,H,N).
3
1-Methyl-(2S,5S)-di-(2-phenethyl)-pyrrolidine Hydrochloride
(11). Compound 10 was subjected to conditions similar to those
described for the preparation of 9. Spectral characteristics were
identical to those of 7. [R]D20 = -48.7° (c = 1.0, MeOH). Anal.
C21H28ClN (C,H,N).
References
cis-2,5-Di(2-benzyl)-pyrrolidine Hydrochloride (22). To a so-
lution of 21 (200 mg, 0.50 mmol) in MeOH (5 mL) was added
Pd(OH)2-C (10 mg, 10 wt %) and ammonium formate (100 mg,
50 wt %). The mixture was refluxed for 20 min, diluted with
CH2Cl2 (60 mL), and filtered through celite. The filtrate was
treated with 5 mL of conc HCl and evaporated to a crusty white
solid under reduced pressure. The solid was triturated with
diethyl ether and precipitated from a mixture of CH2Cl2 and
hexanes by addition of hexanes, to yield 22 (108 mg, 74%) as a
white solid, mp = 173 °C. 1H NMR (300 MHz, CDCl3) δ ppm
9.4 (s, 1H), 9.2 (br, s, 1H), 7.45-7.00 (m, 10H), 4.00-3.64 (s, br,
2H), 2.48-2.14 (m, 4H), 1.70-1.52 (m, 6H). 13C NMR (75 MHz,
CDCl3) δ ppm 139.0, 126.3, 124.6, 120.8, 58.4, 37.1, 34.3, 33.1.
Anal. C18H22ClN (C,H,N).
1-Methyl-cis-2,5-di(2-benzyl)-pyrrolidine Hydrochloride (23).
To a solution of 22 (100 mg, 0.32 mmol) in MeOH (3 mL)
was added NaCNBH3 (50 mg, excess) and paraformaldehyde
(50 mg, excess). The initial suspension became a clear solution
upon stirring for 6 h. The solution was diluted with conc HCl
(10 mL) and stirred for 2 h, during which time all of the
cyanoborohydride and hydrogen cyanide was assumed to be
decomposed. The mixture was poured into 20% aq NaOH
(10 mL) and extracted with CH2Cl2 (100 mL). Drying and
evaporation of the organic layer yielded an oil that was treated
with conc HCl (5 mL) and coevaporated with toluene. Tritura-
tion of the residue afforded a white solid 22 (80 mg, 81%). mp =
126 °C (deliquescent). 1H NMR (300 MHz, CDCl3) δ ppm 9.98
(br, s, 1H), 7.33-7.00 (m, 10H), 3.63-3.53 (m, 2H), 2.46-2.23
(m, 4H), 2.42 (s, 3H), 1.51-1.22 (m, 6H). 13C NMR (75 MHz,
CDCl3) 139.1, 128.9, 127.4, 125.1, 65.2, 48.3, 39.0, 30.1. Anal.
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C19H24ClN H2O (C,H,N).
3
cis-2,5-Di(3-phenylpropyl)-pyrrolidine Hydrochloride (25). To
compound 24 (100 mg, 0.21 mmol) was added Pd(OH)2-on-
carbon (10 wt %), MeOH (10 mL) and ammonium formate
(500 mg). After refluxing for 2 h, the solution was poured into
CH2Cl2 (50 mL), filtered, and evaporated to obtain a white solid
(70 mg, 94%). 1H NMR (300 MHz, CDCl3) δ ppm 10.31 (s, 1H),
9.03 (s, 1H), 7.27-7.13 (m, 15H, Ar), 3.50 (s, 1H), 2.62-2.14 (m,
4H), 2.12-1.68 (m, 14H). 13C NMR (75 MHz, CDCl3) δ ppm
140.2, 129.7, 127.2, 126.2, 65.66, 37.38, 29.5, 28.2, 27.85. Anal.
C22H30ClN (C,H,N).
1-Methyl-cis-2,5-di(3-phenylpropyl)-pyrrolidine Hydrochloride
(26). To a solution of 25 (20 mg, 0.08 mmol) in MeOH (3 mL) was
added NaCNBH3 (20 mg, excess) and paraformaldehyde (20 mg,
excess). The initial suspension became a clear solution upon
stirring for 12 h. The solution was diluted with conc HCl