Synthesis and evaluation of novel azetidine analogs as potent inhibitors of vesicular [3H]dopamine uptake

Article abstract of DOI:10.1016/j.bmc.2013.08.001

Lobelane analogs that incorporate a central piperidine or pyrrolidine moiety have previously been reported by our group as potent inhibitors of VMAT2 function. Further central ring size reduction of the piperidine moiety in lobelane to a four-membered het

Full text of DOI:10.1016/j.bmc.2013.08.001

Bioorganic & Medicinal Chemistry 21 (2013) 6771–6777
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and evaluation of novel azetidine analogs as potent
3
inhibitors of vesicular [ H]dopamine uptake
a
a
a
b
a
Derong Ding , Justin R. Nickell , Agripina G. Deaciuc , Narsimha Reddy Penthala , Linda P. Dwoskin ,
b,
⇑
Peter A. Crooks
a
Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA
Department of Pharmaceutical Sciences, College of Pharmacy, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Lobelane analogs that incorporate a central piperidine or pyrrolidine moiety have previously been
reported by our group as potent inhibitors of VMAT2 function. Further central ring size reduction of
the piperidine moiety in lobelane to a four-membered heterocyclic ring has been carried out in the cur-
Received 11 June 2013
Revised 24 July 2013
Accepted 1 August 2013
Available online 11 August 2013
rent study to afford novel cis-and trans-azetidine analogs. These azetidine analogs (15a–15c and 22a–
3
2
4
2c) potently inhibited [ H]dopamine (DA) uptake into isolated synaptic vesicles (K
i
6 66 nM). The cis-
= 24 nM), and was twofold more potent that
= 43 nM). The trans-methylenedioxy analog, 15c
= 31 nM), was equipotent with the cis-analog, 22b, in this assay. Thus, cis- and trans-azetidine analogs
2b and 15c represent potential leads in the discovery of new clinical candidates for the treatment of
-methoxy analog 22b was the most potent inhibitor (K
i
Keywords:
Lobelane
either lobelane (2a, K
i
= 45 nM) or norlobelane (2b, K
i
(K
i
Azetidine analogs
Methamphetamine abuse
VMAT2
2
methamphetamine abuse.
[
3H]DA uptake
Ó 2013 Elsevier Ltd. All rights reserved.
⁄
1
. Introduction
Methamphetamine (METH) is an addictive psychostimulant
and a6b2 containing nicotinic acetylcholine receptors (nAChRs) as
well as DAT.
Development of selective, high affinity VMAT2 inhibitors will
allow us to test the hypothesis that selective inhibition of VMAT2
function decreases METH reward. Preliminary structure–activity
relationship (SAR) studies aimed at selectively targeting VMAT2
initially identified N-methyl-2,6-di-(cis-phenylethyl)piperidine
(lobelane) (Fig. 1, 2a), a chemically defunctionalized analog of
lobeline, as a more potent and selective inhibitor of vesicular DA
uptake. Lobelane has negligible affinity for
drug and its abuse has been steadily rising throughout the last dec-
ade to become an escalating worldwide problem. Chronic METH
abuse may cause neural damage, which can result in harmful ef-
fects on cognitive processes, such as attention and memory.
Although there are serious consequences resulting from METH
abuse, there are no FDA-approved pharmacological agents avail-
able to treat METH addiction.
Psychostimulant abuse is primarily thought to result from alter-
ations in the central dopaminergic system. METH interacts with
the vesicular monoamine transporter-2 (VMAT2), promoting vesic-
ular dopamine (DA) release into the cytosol, and also inhibiting
monoamine oxidase, both of which result in an increase in cyto-
solic concentrations of DA. METH also reverses the DA transporter
4
⁄
⁄
3
⁄
a
4b2 ,
a
7 and
a
6b2
containing nAChRs, and has improved affinity for the [ H]dihydro-
tetrabenazine binding site on VMAT2compared to lobeline. Preli-
minary SAR studies with lobelane analogs (Fig. 1) revealed that
(
DAT) to afford elevated synaptic DA concentrations, which is
thought to be associated with its abuse liability. Importantly, re-
search has shown that lobeline (Fig. 1), the principal alkaloid in
3
Lobelia inflata, potently inhibits [ H]DA uptake into synaptic vesi-
cles, inhibits METH-evoked DA release from rat striatal slices,
1
–3
and attenuates METH-induced behavioral effects in rats.
How-
ever, lobeline is a relatively nonselective drug, and has been shown
⁄
⁄
to interact with several different CNS targets, including a4b2 , a7
⇑
Corresponding author. Tel.: +1 501 686 6495; fax: +1 501 686 6057.
E-mail address: pacrooks@uams.edu (P.A. Crooks).
Figure 1. Molecular structures of lobeline (1), lobelane (2a), norlobelane (2b), and
pyrrolidineanalogs of norlobelane (3).
0
968-0896/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2013.08.001

6
772
D. Ding et al. / Bioorg. Med. Chem. 21 (2013) 6771–6777
aromatic substituted analogs exhibited improved affinity for
of Cromwell’s original azetidine synthesis,13 which involved a
⁄
⁄
VMAT2, and also lacked affinity for
a4b2 and
a
7 nicotinic acetyl-
four-step reaction sequence. Initially, bromination of glutaric
0
choline receptors. However, such analogs suffer from poor drug
likeness properties, including poor water-solubility. Other SAR
studies also indicated that removal of the N-methyl substituent
in lobelane (Fig. 1, 2a) to afford norlobelane (Fig. 1, 2b), did not sig-
nificantly affect affinity for VMAT2, suggesting that the presence of
the N-methyl group is not a critical structural requirement for
anhydride afforded
a
,a
-dibromoglutaric anhydride, which was
then converted to its diester with benzyl alcohol in the presence
of oxalyl chloride in DMF. Subsequent reaction of the benzyl dies-
ter with 3 equiv of benzylamine in DMF at 80 °C afforded a mixture
of the trans- and cis-azetidine isomers 4 and 5, in a 2:1 ratio. Sep-
4
aration and treatment of the (±)-trans-isomer 4 with NaBH affor-
5
interaction with VMAT2. Based on these observations, a series of
ded alcohol 6 in 75% yield. When compound 6 was submitted to
Dess–Martin oxidation, a complex mixture was obtained and sig-
nificant decomposition occurred during silica gel column chroma-
tography clean-up; consequently the desired (±)-dialdehyde 7 was
not obtained. Based on these findings, compound 6 was subjected
to Swern oxidation and the resulting (±)-dialdehyde 7 used directly
in a Wittig reaction with benzyl triphenyl phosphonium bromide.
After reaction work-up and product purification, the key interme-
diate 8 was obtained in 42.8% over two steps from 6. We attempted
to effect hydrogenation of the alkene moieties in 8 with simulta-
neous hydrogenolysis of the N-benzyl group.
pyrrolidino analogs of norlobelane (Fig. 1, 3) were synthesized
and evaluated asVMAT2 inhibitors.6 The results indicated that
pyrrolidinonorlobelane analogs were potent inhibitors of vesicular
DA uptake.
–8
In order to broaden the scope of the above SAR study, we now
have synthesized a series of novel cis- and trans-azetidine deriva-
tives (15a–15c and 22a–22c) to assess further the effect of hetero-
cyclic ring size reduction in norlobelane (2b). General synthetic
approaches to azetidine derivatives have been reported previ-
9
–12
ously.
In this communication, we report on a highly efficient
synthetic route for the preparation of a series of novel cis- and
trans-1,4-disubstituted azetidine derivatives that are structurally
related to norlobelane (2b). This synthetic route involves a critical
Wittig reaction and a selective double bond reduction. The azeti-
dine derivatives obtained were evaluated for their ability to inhibit
However, when compound 8 was submitted to hydrogenation
with 20% Pd(OH) , the major product obtained was the ring opened
2
derivative 9, along with other unidentified by-products. As a result
of this failure, we attempted to overcome this difficulty by utilizing
sequential steps for the reduction of the double bonds followed by
the removal the N-benzyl protecting group, respectively, using
3
the uptake of [ H]DA into isolated synaptic vesicles from rat brain.
6
14
NaBH
4
, CuCl
2
/NaBH
4
and then Wilkinson’s reagent.
Unfortu-
nately, none of the desired product, 15a, was obtained utilizing this
strategy.
In view of the above problems, we sought an alternative route
to the target azetidine derivative 15a. Thus, instead of utilizing
an N-benzyl protecting group, the Cbz moiety was used to protect
the N-atom of 10 after removal of the N-benzyl group from 6 with
2
2
. Results and discussion
.1. Chemistry
Our initial strategy for the synthesis of azetidine analogs of nor-
lobelane are summarized in Scheme 1. Isomers 4 and 5 were pre-
pared utilizing literature procedures, that is, Baldwin’s adaptation
2
2 2
0% H /Pd(OH) /MeOH(Scheme 2). Treatment of compound 10
Scheme 1. Reagents and conditions: (a) Ref. 13, four steps; (b) NaBH
THF, ꢀ78 °C ? rt; (e) Pd(OH) , MeOH, H , rt.
4
, THF/EtOH (3:1), rt; (c) oxalyl chloride, DMSO, TEA, CH
2
2
Cl , ꢀ78 °C ? rt; (d) C
6 5 2 6 5 3
H CH P(C H ) Br, n-BuLi,
2
2

D. Ding et al. / Bioorg. Med. Chem. 21 (2013) 6771–6777
6773
Scheme 2. Reagents and conditions: (a) 20% Pd(OH)
tert-BuOK, THF, rt; (e) Wilkinson’s reagent, THF/tert-BuOH (1:1), 55 °C, H ; (f) Pd/C, H , rt.
2 2
2
, MeOH, rt; (b) CbzCl, DIPEA, THF; (c) Swern oxidation,13 ꢀ78 °C ? 0 °C; (d) simple and substituted Ph-CH
2 6 5 3
P(C H ) Br,
Scheme 3. Reagents and conditions: (a) 20% Pd(OH)
tert-BuOK, THF, rt; (e) Wilkinson’s reagent, THF/tert-BuOH (1:1), 55 °C, H ; (f) Pd/C, H , rt.
2 2
2
, MeOH, rt; (b) CbzCl, DIPEA, THF; (c) Swern oxidation,13 ꢀ78 °C ? 0 °C; (d) simple and substituted Ph-CH
2 6 5 3
P(C H ) Br,
with CbzCl/DIPEA in THF afforded the N-CBz protected analogue 11
in 83% yield from 6. Swern oxidation of alcohol 11, followed by
in situ Wittig reaction with the appropriate aromatic substituted
benzyltriphenylphosphonium bromide afforded the unsaturated
esters 13a–13c in 64–69% overall yield from 11. Note, that when
n-BuLi was used as a base in the Wittig reaction of 12 with benzyl-
triphenylphosphonium bromide, the yield of 13a from 11 was only
pounds 15a–15c were obtained through hydrogenation over 10%
Pd/C in 90–93% yield.
When the above procedure was utilized for the synthesis of the
cis-1,4-disubstituted azetidine analogs from cis-intermediate 5, the
target compounds 22a–22c were obtained in comparable yields to
the trans-isomers, 15a–15c.The cis-aromatic substituted azetidine
analogs 22a–22c were prepared as described above utilizing the
appropriate aromatic substituted benzyltriphenylphosphonium
bromide and the cis-N-benzyloxycarbonyl-1,4-diformylazetidine
isomer 19 (Scheme 3).
6
%. When compound 13a was hydrogenated over Pd/C or Pd(OH)
2
an unidentified mixture resulted, and the target compound 15a
was not obtained. Attempts failed at initial removal of the N-Cbz
group of compound 13a with 6 M HCl prior to hydrogenation.
The above azetidine analogs were submitted for pharmacologi-
cal evaluation as their water-soluble hydrochloride salts for their
4
Interestingly, when compound 13a was treated with CuCl/NaBH ,
3
compound 14a was obtained, although the yield was only 40%.
However, when compounds 13a–13c were each treated with Wil-
kinson’s reagent, intermediates 14a–14c were obtained in 95–98%
yield. With the key intermediates 14a–14c in hand, the target com-
ability to inhibit the uptake of [ H]DA into isolated synaptic vesi-
i
cles from rat brain. The inhibition constants (K ) obtained are listed
in Table 1. The water-solubilities of the hydrochloride salts of the
azetidine analogs were generally slightly improved compared to

6
774
D. Ding et al. / Bioorg. Med. Chem. 21 (2013) 6771–6777
Table 1
Inhibition constants (K
synaptic vesicles
sium tartrate solution prior to the final centrifugation (100,000g
for 45 min at 4 °C). Final pellets were resuspended in 2.4 ml of as-
say buffer (25 mM HEPES, 100 mM potassium tartrate, 50
3
i
) for analog-induced inhibition of uptake of [ H]DA into
lM
Compounda
R
VMAT2 [ H]DA uptake,
3
2+
EGTA, 100
.4). Aliquots of the vesicular suspension (100
tubes containing assay buffer, various concentrations of inhibitor
l
M EDTA, 1.7 mM ascorbic acid, 2 mM ATP-Mg , pH
K
i
(nM); mean ± SEMb
7
lL) were added to
METH
—
—
—
H
2460 ± 8.3
45 ± 2.0
43 ± 8.0
48 ± 2.8
66 ± 6.1
31 ± 7.7
62 ± 3.9
24 ± 1.5
55 ± 3.0
2
2
1
1
1
2
2
2
a
b
3
(
5
0.1 nM ꢁ 10 mM) and 0.1
00 L. Nonspecific uptake was determined in the presence of
Ro4-1284 (10 M). Reactions were terminated by filtration, and
lM [ H]DA in a final volume of
l
5a
5b
5c
2a
2b
2c
l
4-OCH
3,4-Methylenedioxy
H
4-OCH
3,4-Methylenedioxy
3
radioactivity retained by the filters was determined by liquid scin-
tillation spectroscopy (Liquid scintillation analyzer; PerkinElmer
Life and Analytical Sciences, Boston, MA).
3
a
All compounds were evaluated as their water-soluble hydrochloride salts.
Each K value represents mean ± SEM from four independent experiments, each
i
4
. Conclusions
b
performed in duplicate. METH: Methamphetamine.
In conclusion, we have developed an efficient synthetic ap-
proach for the preparation of novel azetidine analogs (15a–c and
2a–c) of norlobelane (2b). Conditions were developed for the syn-
thesis of the key intermediate Cbz-protected diphenethylazete-
2
the hydrochloride salts of the corresponding norlobelane analogs,
and were similar to those of the previously reported hydrochloride
salts of the pyrrolidine analogs of norlobelane.8,15 Interestingly, all
dines (14a–c and 21a–c). The cis-analog 22b was the most
3
3
potent inhibitor of [ H]DA uptake (K = 24 nM) and was about two-
of the azetidine analogs exhibited potent inhibition of [ H]DA up-
take into isolated synaptic vesicles (K = 24–66 nM).
i
fold more potent than lobelane (2a, K
i
= 45 nM) and norlobelane
= 43 nM). Interestingly, the trans-methylenedioxy analog
5c was equipotent with 22b. Thus, cis- and trans-azetidine ana-
i
(2b, K
i
Among the compounds tested, it is interesting to note that the
inhibitory activity of the novel cis-azetidine derivative 22b
1
logs 22b and 15c were considered unique structures and potential
leads in the discovery of clinical candidates for the treatment of
methamphetamine abuse.
(
(
K
i
= 24 nM) was about twofold more potent than that of lobelane
2a, K = 45 nM) and norlobelane (2b, K = 43 nM). Comparison of
= 48 nM) and 22a (K = 62 nM), and
= 31 nM) and 22c (K = 55 nM), indicates that the inhib-
i
i
the inhibitory data for 15a (K
for 15c (K
i
i
i
i
itory activities of the trans-type azetidine derivatives are not differ-
ent from those of the cis-type azetidine derivatives. This lack of
difference in inhibition of DA uptake at VMAT2 between the cis-
and trans-azetidine isomers is intriguing, since the cis-scaffold is
structurally quite different from the trans-scaffold with respect to
spatial arrangement of the 2,4-diphenethyl substituents. This indi-
cates that the difference in stereochemistry between these analogs
is not a critical factor that governs their mode of interaction with
their binding site on the cytosolic face of VMAT2, which translo-
cates DA from the cytosol into the vesicle. This phenomenon has
also been observed with the structurally related cis and trans iso-
5. Experimental part
5.1. General chemistry
Unless otherwise noted, moisture sensitive reactions were con-
ducted under dry nitrogen. THF was dried with sodium/benzophe-
none and was freshly distilled before use. Anhydrous DCM was
obtained via distillation over CaH
chromatography plates were obtained from Dynamic Adsorbents
Inc. Flash chromatography silica gel 60, 40–64 m, was obtained
from Silicycle Ultrapure Silica Gels. H and C NMR spectra were
2
. Silica gel 60 F254 thin layer
l
1
13
1
1
mers of 2,5-di-(2-phenethyl)-pyrrolidine, which are also equipo-
recorded in Fourier transform mode at 500( H) or at 300( H)/
tent in their ability to inhibit DA uptake at VMAT2.8
,15
These
13
75( C) MHz in the indicated deuterated solvents. High resolution
azetidine analogs water solubility is same as the lobelanepiperi-
dine and pyrrolidine compounds.
The above data indicate that these novel azetidine derivatives
may represent new ‘lead analogs’ with pharmacological and phys-
iochemical properties favorable for subsequent drug development.
(HRMS) mass spectra were obtained using electron impact (EI) ion-
ization techniques ona magnetic sector instrument at a resolution
greater than 10,000.
5.2. Synthesis
5
.2.1. (± ±)trans)(1)Benzyl azetidine)2,4)diyl±dimethanol (6±
At 0 °C, (±)-trans-benzyl 2,2 -(1-benzylazetidine-2,4-diyl) diace-
3
3
. Pharmacological evaluation
0
3
tate (7.5 g, 18.1 mmol) was dissolved in 200 ml THF/EtOH (3:1).
NaBH (8.2 g, 216 mmol) was added to the stirred solution in por-
.1. [ H]DA uptake inhibition assay
4
3
tions. After completion of the reaction (as indicated by TLC moni-
toring) the resulting mixture was acclimated to room
temperature. The reaction mixture was diluted with EtOAc and
sequentially washed with 1 M citric acid solution and saturated
aqueous NaCl solution. The organic phase was separated and the
aqueous phase was extracted three times with EtOAc. The organic
phases was combined and purified by flash chromatography (DCM/
MeOH 5:1) to give pure compound 6 (2.8 g, 75%) as an amorphous
Inhibition of [ H]DA uptake was conducted using the isolated
synaptic vesicle preparations. Briefly, rat striata were homogenized
0
with 10 strokes of a Teflon pestle homogenizer (clearance ꢁ0.003 )
in 14 ml of 0.32 M sucrose solution. Homogenates were centri-
fuged (2000g for 10 min at 4°C), and the resulting supernatants
were centrifuged again (10,000g for 30 min at 4°C). Pellets were
resuspended in 2 ml of 0.32 M sucrose solution and subjected to
osmotic shock by adding 7 ml of ice-cold water to the preparation,
followed by the immediate restoration of osmolarity by adding
solid, which was crystallized from acetone to give a white solid.
Mp: 127–128 °C. H NMR (300 MHz, CDCl
1
): d 7.21–7.35 (m, 5H),
9
00
trate solution. Samples were centrifuged (20,000g for 20 min at
°C), and the resulting supernatant was centrifuged again
55,000g for 1 h at 4 °C), followed by the addition of 100 L of
L of 1.0 M potas-
lL of 0.25 M HEPES buffer and 900 lL of 1.0 M potassium tar-
3
4
.06 (d, J = 13.5, 1H), 3.72–3.78 (m, 3H), 3.48–3.60 (m, 4H), 3.05
1
3
(
br s, 2H), 2.14 (t, J = 6.9, 2H); C NMR (75 MHz, CDCl
3
): d 138.4,
4
(
1
28.7, 128.5, 128.3, 127.5, 62.8, 62.3, 54.1, 21.5 ppm.
l
1
4
0 mM MgSO , 100 lL of 0.25 M HEPES and 100 l

D. Ding et al. / Bioorg. Med. Chem. 21 (2013) 6771–6777
6775
5
.2.2. (± ±)trans)Benzyl 2,4)bis(hydroxylmethyl±azetidine)1)
5.2.5. (± ±)trans)Benzyl)2,4)bis(2)(benzo[d][1,3]dioxol)5)
yl±vinyl± azetidine)1)carboxylate (13c±
carboxylate (11±
Compound 6 (0.52 g, 2.5 mmol) was dissolved in 10 ml of meth-
anol. To this solution was added 0.13 g 20% Pd(OH)2, and the
The same procedure to that described for the preparation of
compound 13a was utilized. Compound 13c was obtained as an
1
resulting mixture was strongly stirred under 1 atm H
2
at room
oil in 68.8% yield; H NMR (500 MHz, CDCl
3
): d 7.14–7.28 (m,
temperature. After completion of the reaction (determined by
TLC monitoring) the reaction mixture was filtered through a pad
of Celite. The filtrate was evaporated under reduced pressure to af-
ford the crude product, which was used directly in the next step.
The above crude product was dissolved in 15 ml DCM and Cbz-
Cl (0.35 ml, 2.5 mmol) was added to the solution. The reaction mix-
ture was cooled to 0 °C and DIPEA was added slowly. Then the
reaction was acclimated to room temperature. After the reaction
was completed, 30 ml of DCM was added to dilute the mixture,
5H), 6.76–6.91 (m, 6H), 6.42–6.60 (m, 2H), 6.19 (br s, 2H), 5.90–
5.97 (m, 4H), 5.19 (d, J = 12.6, 1H), 4.89–4.95 (m, 3H), 2.33–2.39
(m, 2H) ppm. HRMS (EI) calcd for C29
483.1675.
6
H25NO , 483.1682, found
5.2.6. 5.2.4.(± ±)trans)Benzyl)2,4)diphenethylazetidine)1)
carboxlate (14a±
Compound 13a (0.13 g, 0.32 mmol) was dissolved in 8 ml THF/
tert-BuOH (1/1) and 0.05 g tris(triphenylphosphine)-rhodium chlo-
ride added to the solution. The mixture was hydrogenated at 55 °C
4
and organic solution was washed with saturated aqueous NH Cl
solution. The organic phase was then separated and purified by
flash chromatography (hexane/EtOAc-1:1 to 1:2) and compound
2
under 1 atm H . After the reaction was complete, the mixture was
filtered through a pad of Celite. The filtrate was concentrated and
purified by flash chromatography (hexane/EtOAc 24:1) and com-
1
1 was obtained as an amorphous solid (0.52 g, 82.9%, two steps):
1
pound 14a was obtained as a colorless oil in 97.6% yield; 1
H
H NMR (300 MHz, CDCl
3
): d 7.26–7.37 (m, 5H), 5.05–5.15 (m, 2H),
.28–4.44 (m, 3H), 3.61–3.87 (m, 4H), 2.71 (br s, 1H), 2.14–2.20 (m,
H), 1.95–2.02 (m, 1H); 13C(75 MHz, CDCl
): d 156.9, 136.1, 128.8,
28.5, 128.2, 67.6, 66.6, 63.7, 62.2, 61.6, 21.5 ppm. HRMS (EI) calcd
17NO4, 251.1116, found 251.1113.
4
1
1
NMR (300 MHz, CDCl
4.26 (br s, 2H), 2.24–2.63 (m, 6H), 1.84–2.02 (m, 4H); C NMR
(75 MHz, CDCl ): d 155.2, 141.6, 141.5, 137.0, 128.6, 128.5, 128.4,
128.1, 128.1, 128.0, 126.1, 66.4, 59.5, 58.9, 36.0, 35.6, 31.2, 30.8,
3
): d 7.07–7.39 (m, 15H), 5.02–5.16 (m, 2H),
13
3
3
for C13
H
2
3
8.6 ppm; HRMS (EI) calcd. for C27
99.2191.
2
H29NO , 399.2198, found
5
.2.3. (± ±)trans)Benzyl)2,4)distyrylazetidine)1)carboxylate (13a±
ml of anhydrous DCM was placed in flask and cooled to ꢀ78
°C. Oxalyl chloride (0.3 ml, 3.4 mmol) was added to the flask
5
5
1
.2.7. (± ±)trans)Benzyl)2,4)bis(4)methoxyphenethyl±azetidine)
)carboxylate (14b±
0
and anhydrous DMSO (0.28 ml, 3.9 mmol) was dropped slowly into
the mixture. The solution was stirred for 5 min and compound 11
The same procedure to that described for the preparation of
compound 14a was utilized. Compound 14b was obtained as a col-
(
0.21 g, 0.85 mmol) was dissolved in 1 ml anhydrous DCM and in-
jected into the reaction mixture. After 1 h, 0.89 ml of anhydrous
Et N was added to the reaction flask, which was then acclimated
to room temperature. Saturated aqueous NH Cl solution was used
1
orless oil in 95.3% yield. H (300 MHz, CDCl
3
): d 7.26–7.37 (m, 4H),
.00–7.14 (m, 4H),6.78–6.85 (m, 5H), 5.04–5.19 (m, 2H), 4,27 (br s,
H), 3.73–3.86 (m, 6H),2.46–2.62 (m, 5H), 2.23–2.27 (m, 1H), 1.82–
7
2
2
4
3
4
.07 (m, 4H). HRMS (EI) calcd for C29
59.2407.
4
H33NO , 459.2410, found
to quench the reaction. The mixture was separated and the aque-
ous phase was extracted twice with DCM. The organic phases were
4
combined, dried over anhydrous NaSO , filtered, and the filtrate
evaporated to dryness. The resulting crude product was used
immediately in the next step.
Benzyltriphenylphosphonium bromide (1.1 g, 2.6 mmol) was
suspended in 5 ml anhydrous THF. 2.5 ml of 1 M potassium tert-
butoxide was added slowly to the mixture. The solution was stirred
for 1 h and the above crude aldehyde dissolved in 1 ml DCM was
added to the solution. The reaction was stirred for 2 h at room tem-
5
.2.8. (± ±)trans)Benzyl)2,4)bis(2)(benzo[d][1,3]dioxol)5)
yl±ethyl± azetidine)1)carboxylate (14c±
The same procedure to that described for the preparation of
compound 14a was utilized. Compound 14c was obtained as a col-
orless oil in 96.1% yield. H NMR (500 MHz, CDCl
1
3
): d 7.30–7.36 (m,
5
2
1
4
H), 6.51–6.72 (m, 6H), 5.92 (s, 4H), 5.02–5.16 (m, 2H), 4.23 (br s,
H), 2.43–2.56 (m, 5H), 2.21 (br s, 1H), 1.97 (d, J = 6.5 Hz, 2H),
.82–1.88 (m, 2H) ppm. HRMS (EI) calcd for
87.1995, found 487.1998.
29 6
C H29NO ,
4
perature. Saturated aqueous NH Cl solution was then added to
quench the reaction. 10 ml of EtOAc was added and the mixture
was separated. The aqueous phase was extracted twice with EtOAc.
The organic phases were combined and purified by flash chroma-
5
.2.9. (± ±)trans)2,4)Diphenethylazetidine (15a±
Compound 14a (0.13 g, 0.32 mmol) was dissolved in 10 ml of
MeOH containing 0.019 g 10% Pd/C. The mixture was vigorously
stirred under 1 atm H at room temperature. TLC was used to mon-
itor the reaction. After the reaction was complete, the solution was
filtered through a pad of Celite and the filtrate was concentrated.
tography (hexane/EtOAc 12:1). Compound 13a was obtained as
1
colorless oil in 63.9% yield (two steps). H NMR (300 MHz, CDCl
3
):d
2
6
5
1
1
1
3
3
.99–7.39 (m, 15H), 6.34–6.65 (m, 3H), 5.97–6.04 (m, 1H), 4.94–
.27 (m, 4H), 2.34–2.44 (m, 2H); 13C NMR (75 MHz, CDCl
3
): d
55.4, 136.7, 136.6, 136.5, 133.6, 132.2, 131.4, 130.6, 130.0,
29.4, 129.1, 129.0, 128.8, 128.7, 128.5, 128.4, 128.4, 127.9,
27.8, 127.3, 126.8, 66.6, 61.5, 60.3, 56.6, 32.4, 32.2, 31.7,
The residue was purified by flash chromatography (DCM/CH
3
OH
1
5:1) and compound 15a was afforded as a colorless oil in 93.2%
1
yield; H NMR (500 MHz, CDCl
J = 7.5, 1H), 2.70–2.74 (m, 2H), 2.51–2.61 (m, 3H), 2.36–2.40 (m,
3
):d 7.14–7.26 (m, 10H), 4.27 (t,
0.0 ppm; HRMS (EI) calcd for
95.1873.
27 2
C H25NO 395.1885, found
1
3
2
H), 2.10–2.19 (m, 4H); C NMR (75 MHz, CDCl
3
):d 139.9, 128.8,
1
28.7, 128.6, 128.4, 126.5, 56.9, 36.1, 31.7, 30.3 ppm. HRMS (EI)
5
.2.4. (± ±)trans)Benzyl)2,4)bis(4)methoxystyryl±azetidine)1)
calcd for C19H23N 265.1830, found 265.1823.
carboxylate (13b±
The same procedure to that described for the preparation of
5.2.10. (± ±)trans)2,4)bis(4)methoxyphenethyl±azetidine (15b±
The same procedure to that described for the preparation of
compound 15a was utilized. Compound 15b was obtained as a col-
compound 13a was utilized. Compound 13b was obtained as an
1
oil in 65.1% yield; H NMR (300 MHz, CDCl
3
): d 7.01–7.45 (m,
1
1
4
1H), 6.76–6.93 (m, 4H), 6.44–6.50 (m, 1H), 5.84–5.92 (m, 1H),
.83–5.26 (m, 3H), 3.74–3.87 (m, 7H), 2.32–2.38 (m, 2H) ppm.
, 455.2097, found 455.2099.
orless oil in 91.7% yield. H NMR (300 MHz, CDCl ) d 7.04–7.08 (m,
3
4H), 6.73–6.83 (m, 4H), 4.23–4.31 (m, 2H), 3.67–3.84 (m, 6H),
2.64–2.73 (m, 2H), 2.47–2.60 (m, 2H), 2.31–2.43 (m, 2H), 2.12–
HRMS (EI) calcd for C29H29NO
4

6
776
D. Ding et al. / Bioorg. Med. Chem. 21 (2013) 6771–6777
2
(
.20 (m, 2H), 1.99–2.10 (m, 2H), 1.24–1.31 (m, 1H); 13C NMR
75 MHz, CDCl ): d 158.2, 131.8, 129.6, 129.4, 114.3, 114.1, 56.9,
5.2.17. cis)Benzyl)2,4)diphenethylazetidine)1)carboxlate (21a±
3
The same procedure to that described for the preparation of
5
3
5.5, 36.1, 30.8, 30.1 ppm. HRMS (EI) calcd for
25.2042, found 325.2039.
C
21
H27NO
2
,
compound 14a was utilized. Compound 21a was obtained as an
1
oil in 87.9% yield; H NMR (300 MHz, CDCl
3
): d 7.17–7.38 (m,
1
5H), 5.13 (d, J = 4.5 Hz, 2H), 4.13–4.21 (m, 2H), 2.63–2.68 (m,
5
.2.11. (± ±)trans-2,4)Bis(2)(benzo[d][1,3]dioxol)5)
4H), 2.25–2.47(m, 3H), 1.88–1.97 (m, 2H), 1.52–1.58 (m, 1H);
1
3
yl±ethyl±azetidine (15c±
C(75 MHz, CDCl
128.1, 128.0, 126.0, 66.6, 59.4, 38.6, 31.7, 29.3 ppm. HRMS (EI)
calcd for C27 , 399.2198, found 399.2193.
3
): d 158.0, 141.7, 136.9, 128.6, 128.5, 128.5,
The same procedure to that described for the preparation of
compound 15a was utilized. Compound 15c was obtained as a col-
orless oil in 90.4% yield. H NMR (300 MHz, CDCl
H29NO
2
1
3
): d 6.58–6.73 (m,
6
1
1
H), 5.84–5.97 (m, 4H), 3.68–3.77 (m, 2H), 2.29–2.58 (m, 4H),
5.2.18. cis)Benzyl)2,4)bis(4)methoxyphenethyl±azetidine)1)
carboxa) late (21b±
1
3
.80–2.05 (m, 7H). C NMR (75 MHz, CDCl
35.7, 121.2, 109.0, 108.3, 101.0, 55.4, 40.0, 32.2, 30.1 ppm. HRMS
, 353.1627, found 353.1631.
3
): d 147.7, 145.7,
The same procedure to that described for the preparation of
(
EI) calcd for C21
H23NO
4
compound 14a was utilized. Compound 21b was obtained as an
1
oil in 93.1% yield, H NMR (300 MHz, CDCl
3
): d 7.34–7.38 (m, 4H),
5
.2.12. cis)(1)Benzylazetidine)2,4)diyl± dimethanol (16±
7.08 (d, J = 8.1, 4H), 6.80–6.83 (m, 5H), 5.11 (s, 2H), 4.08–4.15 (m,
2H), 3.72–3.85 (m, 6H), 2.56–2.62 (m, 4H), 2.35–2.44 (m, 1H),
2.19–2.30 (m, 2H), 1.83–1.92 (m, 2H), 1.50–1.56 (m, 1H) ppm.
The same procedure to that described for the preparation of
0
compound 6 was utilized, except that cis-benzyl 2,2 -(1-benzylaz-
etidine-2,4-diyl) diacetate was utilized in place of trans-benzyl
4
HRMS (EI) calcd for C29H33NO 459.2410, found 459.2408.
0
2
,2 -(1-benzylazetidine-2,4-diyl) diacetate. Compound 16 was ob-
1
tained as a white solid, mp 71–73 °C, yield, 65%. H NMR
500 MHz, CDCl ) d 7.24–7.31 (m, 5H), 3.69 (s, 2H), 3.28–3.31 (m,
5.2.19. cis)Benzyl)2,4)bis(2)(benzo[d][1,3]dioxol)5)
yl±ethyl±azetidine)1)carboxylate (21c±
(
6
3
13
H), 2.75 (br s, 2H), 1.99–2.05 (m, 2H); C (75 MHz, CDCl
38.1, 129.3, 128.7, 127.8, 63.6, 63.3, 61.3, 20.5 ppm.
3
): d
The same procedure to that described for the preparation of
1
compound 14a was utilized. Compound 21c was obtained as an
1
oil in 94.7% yield, H NMR(500 MHz, CDCl
3
): d 7.26–7.38 (m, 5H),
5
.2.13. cis)Benzyl)2,4)bis(hydroxylmethyl±azetidine)1)carboxy)
6.60–6.72 (m, 6H), 5.90 (s, 4H), 5.11 (s, 2H), 4.11–4.13 (m, 2H),
2.55–2.58 (m, 4H), 2.37–2.43 (m, 1H), 2.20–2.24 (m, 2H), 1.82–
1.88 (m, 2H), 1.48–1.53 (m, 1H) ppm. HRMS (EI) calcd for
late (18±
The same procedure to that described for the preparation of
compound 11 was utilized. Compound 18 was obtained as a vis-
29 6
C H29NO , 487.1995, found 487.1992.
cous oil, yield, 79.7% (two steps), 1H NMR (500 MHz, CDCl
3
): d
7
2
1
2
2
.27–7.34 (m, 5H), 5.08 (s, 2H), 4.26 (br s, 2H), 3.85 (d, J = 11.5,
5.2.20. cis)2,4)Diphenethylazetidine (22a±
H), 3.60 (br s, 2H), 2.13–2.21 (m, 2H); 13C (75 MHz, CDCl
): d
The same procedure to that described for the preparation of
3
57.8, 136.2, 128.7, 128.5, 128.4, 128.1, 67.3, 64.5, 63.8, 60.7,
compound 15a was utilized. Compound 22a was obtained as a vis-
1
0.1 ppm. HRMS (EI) calcd for
51.1116.
C
13
H17NO
4
251.1116, found,
cous oil, yield 87.3%. HNMR (300 MHz, CDCl
3
): d 7.16–7.29 (m,
10H), 3.59–3.66 (m, 2H), 2.47–2.67 (m, 5H), 2.24–2.32 (m, 1H),
1
3
1
.75–1.92 (m, 4H),1.51–1. 61 (m, 1H); C(75 MHz, CDCl
142.1, 128.5, 128.4, 125.9, 54.2, 40.6, 33.8, 32.1 ppm. HRMS (EI)
calcd for C19 23N, 265.1830, found 265.1822.
3
): d
5
.2.14. cis)Benzyl)2,4)distyrylazetidine)1)carboxlate (20a±
The same procedure to that described for the preparation of
H
compound 13a was utilized. Compound 20a was obtained as an
oil, yield 45.3% (two steps). 1H NMR (300 MHz, CDCl
): d 7.22–
= 6.9, J = 15.9,
H), 5.13 (s, 2H), 4.78–4.85 (m, 2H), 2.75–2.84 (m, 1H), 1.98–
3
5.2.21. cis)2,4)Bis(4)methoxyphenethyl±azetidine (22b±
7
2
2
1
1
.41 (m, 15H), 6.62 (d, J = 15.9, 2H), 6.38 (dd, J
1
2
The same procedure to that described for the preparation of
compound 15a was utilized. Compound 22b was obtained as a vis-
1
3
1
3
.13 (m, 1H); C NMR (75 MHz, CDCl ): d 157.4, 136.7, 136.6,
cous oil, yield 90.5%. H NMR (300 MHz, CDCl
3
): d 6.98–7.26 (m,
33.7, 131.4, 130.6, 130.0, 128.8, 128.7, 128.5, 128.4, 127.9,
27.8, 127.3, 126.8, 66.6, 60.5, 56.6, 32 ppm.
4H), 6.68–6.81 (m, 4H), 4.13–4.21 (m, 2H), 3.67–3.80 (m, 6H),
2.28–2.71 (m, 8H), 1.89–2.11 (m, 3H), 1.23–1.28 (m, 1H); 13
NMR (75 MHz, CDCl ): d 163.9, 158.2, 131.9, 129.6, 129.4, 114.2,
114.1, 56.6, 55.6, 55.5, 36.6, 32.0, 30.7 ppm. HRMS (EI) calcd for
, 325.2042, found 325.2044.
C
3
5
.2.15. cis)Benzyl)2,4)bis(4)methoxylstyryl±azetidine)1)
carboxylate (20b±
21 2
C H27NO
The same procedure to that described for the preparation of
compound 13a was utilized. Compound 20b was obtained as an
5.2.22. cis)2,4)Bis(2)(benzo[d][1,3]dioxol)5)yl±ethyl±azetidine
(22c±
oil in 50.7% yield (two steps). 1H NMR (300 MHz, CDCl
): d 7.17–
.44 (m, 9H), 6.82–6.93 (m, 4H), 6.46–6.58 (m, 2H), 6.19–6.26
m, 1H), 5.86–5.93 (m, 1H), 5.02–5.18 (m, 3H), 4.70–4.78 (m,
3
7
(
The same procedure to that described for the preparation of com-
pound 15a was utilized. Compound 22c was obtained asa viscous oil,
1
1
1
4
H), 3.73–3.88 (m, 6H), 2.73–2.83 (m, 1H), 1.90–1.98 (m,
yield 91.6%. H NMR (300 MHz, CDCl
3 3
): (300 MHz, CDCl ) 6.59–6.72
H) ppm; HRMS (EI) calcd for
55.2089.
C
29
H29NO
4
,
455.2097, found
(m, 6H), 5.89 (s, 4H), 4.16–4.21 (m, 2H), 2.60–2.68 (m, 2H), 2.29–2.52
1
3
(m, 6H), 1.95–2.09 (m, 3H); C NMR (75 MHz, CDCl
35.0, 121.8, 109.5, 108.8, 101.3, 55.5, 36.7, 32.0, 30.9 ppm. HRMS
(EI) calcd for C21 , 353.1627, found 353.1623.
3
):d 147.8, 146.0,
1
5
.2.16. cis)Benzyl)2,4)bis(2)(benzo[d][1,3]dioxol)5)yl±vinyl±
H23NO
4
azetidine)1)carboxylate (20c±
The same procedure to that described for the preparation of
compound 13a was utilized. Compound 20c was obtained as an
Acknowledgments
oil in 53.4% yield (two steps). 1H NMR (300 MHz, CDCl
): d 7.20–
.27 (m, 5H), 6.94 (s, 1H), 6.72–6.83 (m, 5H), 6.42–6.54 (m, 2H),
.15–6.22 (m, 1H), 5.85–5.96 (m, 5H), 5.07–5.17 (m, 3H), 4.71–
.74 (m, 1H), 2.73–2.82 (m, 1H), 1.88–1.97 (m, 1H) ppm. HRMS
This research was supported by NIH grant U01 DA13519. The
University of Kentucky holds patents on lobeline and the analogs
described in the current work. A potential royalty stream to
L.P.D. and P.A.C. may occur consistent with University of Kentucky
policy.
3
7
6
4
(
EI) calcd for C29H25NO6, 483.1682, found 483.1678.

D. Ding et al. / Bioorg. Med. Chem. 21 (2013) 6771–6777
6777
6
7
.
.
Varthak, Ashish P.; Nickell, Justin R.; Jaturaporn, C.; Dwoskin, L. P.; Crooks, P. A.
J. Med. Chem. 2009, 52, 7878.
Zheng, G.; David, B. H.; Reddy, P. N.; Nickell, J. R.; Culver, J. P.; Deaciuc, A. G.;
Dwoskin, L. P.; Crooks, P. A. Med. Chem. Comun. 2013, 4, 564.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmc.2013.08.001.
8. Penthala, N. R.; Ponugoti, P. R.; Deaciuc, A. G.; Nickell, J. R.; Dwoskin, L. P.;
Crooks, P. A. Bioorg. Med. Chem. Lett. 2013, 23, 3342.
9.
Barrett, A. G. M.; Dozzo, P.; White, A. J. P.; Williams, D. J. Tetrahedron 2002, 58,
303.
7
References and notes
10. Evans, G. B.; Furneaux, R. H.; Greatrex, B.; Murkin, A. S.; Vern, L.; Schramm, V.
L.; Tyler, P. C. J. Med. Chem. 2008, 51, 948.
1
2
3
.
.
.
Miller, D. K.; Crooks, P. A.; Teng, L.; Witkin, J. M.; Munzar, P.; Goldberg, S. R.;
Acri, J. B.; Dwoskin, L. P. J. Pharmacol. Exp. Ther. 2001, 296, 1023.
Harrod, S. B.; Dwoskin, L. P.; Crooks, P. A.; Klebaur, J. E.; Bardo, M. T. J.
Pharmacol. Exp. Ther. 2001, 298, 172.
Harrod, S. B.; Dwoskin, L. P.; Green, T. A.; Gehrke, B. J.; Bardo, M. T.
Psychopharmacology (Ber.) 2003, 165, 397.
Wieland, H.; Schopf, C.; Hermsen, W. Liebigs Ann. Chem. 1925, 444, 40.
Zheng, G.; Dwoskin, L. P.; Deaciuc, A. G.; Norrholm, S. D.; Crooks, P. A. J. Med.
Chem. 2005, 48, 5551.
1
1
1. Guanti, G.; Renata, R. Tetrahedron: Asymmetry 2001, 12, 605.
2. Kozikowski, A. P.; Tuckmantel, W.; Reynolds, I. J.; Wroblewski, J. T. J. Med.
Chem. 1990, 33, 1561.
3. Rodenbaugh, R. M.; Cromwell, N. H. J. Heterocycl. Chem. 1968, 5, 309.
4. Baldwin, J. E.; Adlington, R. M.; Jones, R. H.; Schofield, C. J.; Zaracostas, C.;
Greengrass, C. W. Tetrahedron 1986, 42, 4879.
1
1
4
5
.
.
1
5. Beckmann, S. J.; Siripurapu, K. B.; Nickell, J. R.; Horton, D. B.; Denehy, E. D.;
Vartak, A.; Crooks, P. A.; Dwoskin, L. P.; Bardo, M. T. J. Pharmacol. Exp. Ther.
2010, 335, 841.