Discovery of orally active 3-pyridinyl-tropane as a potent nociceptin receptor agonist for the management of cough

Article abstract of DOI:10.1021/jm9008218

A series of 3-pyridinyl-tropane analogues based on previously reported compound 1 have been synthesized and shown to bind to the nociceptin receptor with high affinity. From the SAR study and our lead optimization efforts, compound 10 was found to possess

Full text of DOI:10.1021/jm9008218

J. Med. Chem. 2009, 52, 5323–5329 5323
DOI: 10.1021/jm9008218
Discovery of Orally Active 3-Pyridinyl-tropane As a Potent Nociceptin Receptor Agonist for the
Management of Cough
Shu-Wei Yang,*,† Ginny Ho,† Deen Tulshian,† William J. Greenlee,† John Anthes,‡ Xiomara Fernandez,‡ Robbie L. McLeod,‡
John A. Hey,‡ and Xiaoying Xu§
†Department of Chemical Research;CV & CNS and ‡Neurobiology Department of Biological Research and §Department of Drug Metabolism,
Schering-Plough Research Institute, 2015 Galloping Hill Road, Kenilworth, New Jersey 07033
Received June 5, 2009
A series of 3-pyridinyl-tropane analogues based on previously reported compound 1 have been
synthesized and shown to bind to the nociceptin receptor with high affinity. From the SAR study
and our lead optimization efforts, compound 10 was found to possess potent oral antitussive activity in
the capsaicin-induced guinea pig model. The rationale for compound selection and the biological profile
of the optimized lead (10) are disclosed.
Introduction
Genetic and pharmacological studies indicate that NOP
modulates an anxiety response in rodents.6,7 It was found that
local (ICV) administration of N/OFQ or IP administration of
NOP agonist Ro64-6198 led to anxiolytic-like activity in rats.7,8
Recently, NOP agonists, SCH 221510 and its derivatives, were
unveiled to possess the oral anxiolytic-like activity in the rat
conditioned lick suppression (CLS) model and/or the separa-
tion-induced guinea pig pup vocalization (GPPV) model.9,10
The reported data prompted us to investigate the selectivity
profile of antitussive vs anxiolytic activities for the tropane
analogues.10b,c
In previous communications, we have reported SAR devel-
opment on the C-3 substituted tropane scaffold, an advance
lead moiety derived from the piperidine series.5 Compound
1 was identified previously as a NOP agonist possessing
potent oral dual antitussive and anxiolytic-like activities in
the guinea pig models from a series of 3-carboxylamide-
tropane analogues.10 However, compound 1 and its series of
compounds (carboxylamide) did not show high selectivity
over MOP receptor. The major goal of our further efforts was
focused on identification of a compound with improved NOP
selectivityoverMOPand exhibiting potentantitussive activity
and less CNS effect, measured using the GPPV and CLS
assays to characterize its therapeutic window. Synthesis and
SAR of a novel 3-monosubstituted-tropane series in contrast
to the previous 3-disubstituted tropane series10 are disclosed
along with the discovery of a potential antitussive agent.
Many of G-protein coupled receptors (GPCRsa) were
cloned after the human genome was sequenced. Among those
GPCRs, the nociceptin/orphanin FQ receptor (NOP, NOP1,
ORL-1, or N/OFQR) was first cloned in 1994 from a search of
opioid subtype receptors. It displays low binding affinity for
the classical opioid ligands (ex. naloxone), albeit its sequence
has ∼50% homology to those of the opioid receptor family μ,
κ, and δ (or MOP, KOP, and DOP, respectively).1 NOP is
widely distributed throughout the brain and spinal cord and
thus is expected to participate in a wide range of physiological
events. NOP was deorphanized in 1995. Its endogenous
heptadecapeptide ligand, nociceptin/orphanin FQ(N/OFQ),2
does not interact with the other opioid receptors and has been
reported to mediate various physiological responses through
peripheral or central nervous system (CNS).3 Following the
discovery of NOP and its ligands, there has been remarkable
advance toward understanding its pharmacological signifi-
cance. The pathological and physiological roles of NOP have
recently been reviewed.3d,e Therefore selective NOP agonists or
antagonists could have broad therapeutic potential for the
treatment of related diseases, for instance, pain, cough, anxiety,
stroke, heart failure, cognition dosorder, sleep disorder, sub-
stance abuse, immune disorder, and neurogenic bladder.3d,e
N/OFQ hasbeenshowntodisplay antitussive activityin the
guinea pig and cat capsaicin-induced cough models after
peripheral (IV) or central (ICV) administration,4a-d and to
reduce ex vivo airway contractility in the human isolated
bronchi.4e,f Thus, selective NOP agonists provide a novel
therapeutic approach for the treatment of cough.5
*To whom correspondence should be addressed. Phone: (908) 740-
7291. Fax: (908) 740-7164. E-mail: shu-wei.yang@spcorp.com.
a Abbreviations: CDP, chlordiazepoxide; CHO cell, Chinese hamster
ovary cell; CLS, conditioned lick suppression; CNS, central nervous
system; DOP, δ receptor; GDP, guanosine diphosphate; GPCRs,
G-protein coupled receptors; GPPV, guinea pig pup vocalization; GTPγS,
guanosine 50-O-[γ-thio]triphosphate; hERG, human ether-a-go-go re-
lated gene; hPXR, human pregnane X receptor gene; ICV, intracereb-
roventricular; IP, intraperitoneal; KHMDS, potassium bis(trimethyl-
silyl)amide; KOP, κ receptor; MOP, μ receptor; NaHMDS, sodium bis-
(trimethylsilyl)amide; NOE, nuclear Overhauser effect; NOEL, no effect
level; N/OFQ, nociceptin/orphanin FQ; NOP, nociceptin/orphanin FQ
receptor; Rb, rubidium; RIF, rifampicin.
Chemistry
The synthetic route to the C-3-R-substituted analogues is
summarized in Scheme 1. The commercially available tropi-
none was transformed to ketone 2 and then nitrile 3 using
r
2009 American Chemical Society
Published on Web 08/13/2009
pubs.acs.org/jmc

5324 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17
Yang et al.
Scheme 1a
Scheme 3a
a Reagents and conditions: (a) PtO2, H2, 1 atm, DCM, rt; (b) 37%
formaldehyde, formic acid, 70 °C; (c) Ac2O, pyridine, 0 °C; (d) Br-R,
K2CO3, DMF, 60 °C.
a Reagents and conditions: (a) R-chloroethyl chloroformate, DCE,
reflux; then MeOH, reflux; (b) K2CO3, CH3CN, reflux; (c) KO-tBu,
tosylmethyl isocyanide, DME, -40 °C to rt; (d) NaHMDS, RX, THF, -
78 °C to rt (for the pyridinyl analogues); (e) RPhF, KHMDS, neat,
microwave, 100 °C (for the phenyl analogues); (f) KOH, ethylene glycol,
190 °C; (g) LiAlH4, THF, rt or 60 °C.
Scheme 4a
Scheme 2a
a Reagents and conditions: (a) DIBAL in toluene, 0 °C; (b) tosyl-
methylisocyanide, K2CO3, MeOH, reflux.
Target compounds were tested for their affinity at the cloned
human NOP receptor expressed in CHO cell membranes by
measuring their ability to compete with [125I][Tyr14]N/OFQ.
The opioid receptor binding assays were performed with CHO
cell membranes expressing the human opioid receptors using
[3H]-diprenorphine as the radioligand.4a The functional activ-
ities of selected compounds were evaluated for their ability
to enhance the binding of [35S]GTPγS in the presence of GDP,
using membranes isolated from CHO cells transfected with the
human NOP gene. Because most of the tropane analogues
identified previously showed good selectivity over DOP and
KOP,10 only selectivity over MOP are presented for the most of
the analogues. Some analogues were further evaluated for in
vitro safety assays including hERG rubidium (Rb) efflux and/
or human pregnane X receptor gene (hPXR) assays.11 The
hPXR gene is an important factor regulating P450 CYP3A4,
an important hepatic enzyme related to drug metabolism.
Activation of hPXR, a part of CYP3A4 gene, could result in
induction of CYP3A4 level and subsequently leads to unde-
sired drug-drug interaction.
a Reagents and conditions: (a) NBS, (PhCO2)2, CCl4, reflux; (b) piper-
idine, K2CO3, DMF, rt; (c) 3, NaHMDS, THF, -78 °C to rt; (d) LAH,
THF, rt.
previously established procedure.5,10 Subsequent nucleophilic
addition of a phenyl or pyridinyl group to the less hindered R
face was achieved using the corresponding halides under the
NaHMDS or KHMDS condition to give 4.10 The C-3 stere-
ochemistry was established with NOE experiment as de-
scribed previously.10 The coupling constant of axial H-3
(e.g., tt, 12.0, 5.5 Hz) of 5 further confirmed the stereochem-
istry at the C-3 position.
The cyano group of the nitriles (4) was hydrolyzed to the
carboxylic acid, followed by decarboxylation using KOH in
ethylene glycol at ∼190 °C to give 5. Alternatively, decyana-
tion could be carried out under LAH reductive conditions.
This method was especially useful for the C-3 phenyl analo-
gues because decarboxylation was difficult to achieve under
the KOH conditions in the phenyl series.
To prepare the 3-piperidinylmethyl pyridinyl analogue (9), a
four-step synthesis was designed as shown in Scheme 2. Bro-
mination of 2-bromo-3-methyl-pyridine was conducted using
N-bromosuccinimide to afford 6. Displacement of the bromine
was performed using piperidine in the presence of K2CO3 to
give 7. The same procedure described in Scheme 1 was utilized
to prepare the nitrile (8) and the C-3-β-H analogue (9).
The C-3 piperidinyl analogues were synthesized as de-
scribed in Scheme 3. Reduction of the pyridinyl analogue
(10) with H2 and PtO2 catalyst in DCM gave 11. Reductive
methylation and acetylation of 11 afforded 12 and 13, respec-
tively. Alkylation of 11 further afforded analogues 14 and 15.
The C-3 oxazole analogue (17) was obtained through the
aldehyde intermediate (16) using tosylmethylisocyanide as
shown in Scheme 4.
Results and Discussion
The unsubstituted 2-pyridinyl analogue (10) showed potent
NOP binding affinity with a Ki of 5 nM and ∼215-fold
selectivity over MOP receptor. On the basis of this promising
result, SAR concentrating on pyridinyl substitution was
quickly developed using various commercially available
pyridinyl halides. Introduction of a simple substituent such
as a fluoro, bromo, methyl, hydroxyl, or methylthio group
(18-22) at the 5 position of the pyridine ring led to ∼3-7-fold
reduction of NOP binding affinity (Ki between 14 and
34 nM, Table 1) with selectivity less than 47-fold over MOP
receptor. The two 3-substituted analogues 23 and 24 did not
show desired potency and selectivity. On the basis of the
previous SAR,5b a piperidinylmethyl group was introduced
at the 3-position to regain potency and selectivity. Analogue
9 displayed potent and selective affinity for NOP receptor but
showed high induction in the hPXR assay: 1.76-fold of

Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5325
Table 3. SAR of the C-3 2-Piperidinyl Analogues
Table 1. SAR of the C-3 2-Pyridinyl Analogues
compd
R
NOP Ki (nM)
MOP Ki (nM)
compd
R
NOP Ki (nM)
MOP Ki (nM)
10
18
19
20
21
22
23
24
9
H
5-F
5
34
22
14
24
23
29
467
7
1073
1136
894
11
12
13
14
15
H
2
412
596
nda
309
325
Me
Ac
1
5-Br
5-Me
5-OH
5-SMe
3-Me
3-Br
71
3
661
234
-CH2CN
-(CH2)2OH
2
584
a nd: not determined.
1684
8382
918
affinity and poor selectivity over MOP (11-fold). The C-3
oxazole (17) substitution was not tolerated for NOP affinity
(272 nM). However, the pyrimidinyl analogue (32) exhibited
potent and selective affinity for NOP receptor, comparable to
the lead compound 10. However, 32 displayed higher inhibi-
tion in the hERG Rb eflux assay (15% at 5 μM) compared to
10 (9%) and was not further evaluated. Thus the above
modification did not improve the overall profile of 10.
3-piperidinylmethyl
4-Me
6-F
25
26
27
28
18
25
10
9
891
5420
276
6-Me
6-OMe
421
Table 2. SAR of the Additional C-3 Heterocyclic Analoguesa
To investigate the SAR of the basicity of the C-3 ring verses
NOP affinity, the more basic piperidinyl analogue (11) was
prepared. Compound 11 was identified as a highly potent and
selective NOP ligand with a Ki of 2 nM and >200-fold
selectivity over MOP. However, 11 showed an unacceptable
inhibition of the hERG channel in the hERG Rb efflux assay
(73% at 5 μg/mL). Substitution on the piperidinyl nitrogen
was attempted to address this hERG issue. Hence compounds
12-15 were prepared and evaluated. All of the compounds
except 13 showed potent and selective NOP binding affinity
(Table 3) and were identified as full agonist in the NOP
functional assay (EC50 20-60 nM). Compound 12 displayed
the best NOP affinity (Ki 1 nM) and selectivity over MOP
(∼600-fold). Unfortuneately, these analogues (12, 14, and 15)
still displayed significant inhibition of the hERG channel
(46%, 36%, and 30%, respectively) although the inhibitory
level was reduced, compared to that of 11. In general, potent
and selective NOP ligands were identified in the piperidinyl
series but the hERG issue could not be completely addressed.
The hERG and hPXR data for some analogues are listed
in Table 4. In the hPXR assay, the induction level at 1 μM was
compared to the positive control RIF, and the ratio of 0.4 was
set to be the screening cutoff to minimize the possibility of
false-positive results. From the previous study, it was found
that substitution on the 5-pyridyl position reduced the hPXR
liability compared to that of the unsubstituted pyridyl analo-
gue.10b This was also found true for 5-bromo analogue
19 (0.10, ratio to RIF) compared to that of the unsubstituted
10 (0.38). In this study, 3- or 4-methyl and 3-bromo analogues
(23, 24, and 25, respectively) also displayed reduced hPXR
induction (<0.22) compared to that of 10. All tested pyridyl
analogues displayed acceptable hERG channel inhibition
(<20%). Previously, we found that the terminal acetamide
and methyl carbamate groups were strong inducers for
CYP3A4 through the hPXR reporter gene pathway in the
C-3 pyridyl series.10b Thesame phenomenon was observedfor
13, possessing a N-acetyl moiety, which led to the significant
hPXR liability (1.6, ratio to RIF).
a nd: not determined.
induction relative to the standard rifampicin (RIF) at 1 μM.
Therefore, 9 was not further pursued. The 4-methyl analogue
(25) showed a similar profile to the 5-methyl analogue (20).
Compound 26, having 6-fluoro substitution, displayed excel-
lent selectivity over MOP (∼217-fold) but showed some
induction in the hPXR assay (0.66-fold relative to RIF, above
our cutoff value of 0.4). The 6-methyl (27) or 6-methoxy (28)
substitution on the pyridine ring was tolerated for NOP
binding with inferior selectivity (28- and 47-fold, respectively)
compared to that of 10. Overall substitution on the pyridine
ring in thisstudydid not result ina better NOP-binding profile
compared to unsubstituted 10.
To compare aryl or other heteroaryl substitution at the
C-3 position of the tropane with 2-pyridinyl analogues (10),
a series of compounds were prepared, including phenyl,
3-pyridinyl, 4-pyridinyl, oxazole, and pyrimidinyl analogues.
The SAR data of these analogues are listed in Table 2. The
phenyl analogue (29) displayed potent NOP binding affinity
(Ki 9 nM) with ∼37-fold selectivity over MOP receptor. The
3-pyridinyl analogue (30) retained NOP binding affinity but
showed reduced selectivity over MOP (57-fold), while the
4-pyridinyl analogue (31) showed ∼14-fold decrease of NOP
In summary, removal of C-3 carboxylamide and fluoro of
1 led to the discovery of 10 with improved selectivity over
MOP affinity. Although substitution on the pyridine ring

5326 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17
Table 4. hERG and hPXR Data of Some Tropane Analogues
Yang et al.
Table 5. Selected Pharmacokinetic Data of 10 in Rats and Dogs
hERG inhibition at 5 μM
(Rb efflux assay), %
hPXR, ratio to
RIF at 1 μM
rat (3 mg/kg, PO)
dog (1 mg/kg, PO)
compd
AUC (0-inf), μM h
Cmax, nM
T1/2, h
0.4
60
0.4
180
0.9
24
3
1
4
0.29
0.38
0.10
0.22
0.14
1.76
0.19
0.66
0.23
nd
10
19
23
24
9
9
4.5
23
-1
20
12
11
nda
-3
nd
10
-7
19
15
65
74
46
19
36
29
F (oral), %
CL, mL/min/kg
Vd, L/kg
74
12.6
24
1.5
25
26
27
28
29
30
32
17
11
12
13
14
15
Table 6. In Vivo Activities of Compound 10
antitussive activitya
anxiolytic-like activity
capsaicin-induced
cough model,
ED90, mg/kg
nd
GPPV (NOEL),
mg/kg
rat CLS,
ED50
0.16
nd
compd
0.16
0.10
0.11
1.60
0.11
0.18
10
0.04 (2 h)
0.73 (6 h)
6.8 (2 h)b
0.3 (2 h)
>3 (2 h)
codeine
ndc
nd
a Activity was determined 2 or 6 h after dosing. b ED50 is presented
here. c nd: not determined.
a nd: not determined.
compared to that of the placebo. Further study deter-
mined the no effect level (NOEL) of 10 as 0.3 mg/kg com-
pared to that of the reference compound chlordiazep-
oxide (CDP) (ED50 3.2 mg/kg). In the rat CLS assay,9 10
did not show significant activity at 3 mg/kg compared to
that of the reference compound CDP (ED50 8.8 mg/kg).
Compound 10 exhibited potent oral antitussive activity in
the capsaicin-inducedguineapig coughmodelwithanED90 of
0.04 at 2 h and an ED90 of 0.73 mg/kg at 6 h compared to that
of the reference compound codeine (ED50 6.8 mg/kg, 2 h).4a
The potent antitussive activity observed at 6 h suggested
that 10 could show a long-lasting pharmacological effect in
higher animals or humans. Compound 10 displayed signifi-
cant higher efficacy in the antitussive assay (ED90 0.04 mg/kg)
than that in the anxiolytic-like GPPV assay (NOEL 0.3 mg/
kg). The antitussive/anxiolytic-like efficacy ratio was greater
than 10. Compared to the previous lead compounds 1 and
33,10b,c compound 10 was more selective over MOP. Com-
pound 10 also exhibited better antitussive efficacy and less
CNS effect than those of 1 and 33. In the rat pharmacokinetic
studies, 10 and 33 showed similar profiles. However, 10
displayed significant longer half-life (4.5 h) compared to that
of 33 (1.8 h).
resulted in reduced hPXR induction, the NOP affinity and
selectivity over MOP deteriorated. The saturation of the
pyridine ring led to the identification of potent NOP ligands
(11-15), whereas they suffered from the hERG-inhibition or
hPXR-induction liability. Further replacement of 2-pyridyl
ring with other aromatic rings (e.g., 17, 29-32) did not lead to
compounds with better profile.
From these in vitro studies, 10 was found to possess the
best overall profile and was subjected to various profiling
assays. Compound 10 showed potent NOP agonist activity in
the GTPγS functional assay (EC50: 28 nM) and ∼78-fold
selectivityover MOP(EC50: 2193 nM). Itdisplayed ∼100-and
>2000-fold selectivity over KOP and DOP, respectively. It
showed lowinhibition (<10%at5 μg/mL) in hERG Rb assay
and an acceptable ratio of 0.38 relative to RIF in the hPXR
reporter gene assay. Compound 10 displayed no inhibition of
CYP P450 enzymes (2D6, 2C9, and 3A4) up to 30 μM and no
positive response in the Ames test. In the pharmacokinetic
assay, 10 showed a reasonable oral pharmacokinetic profile
in rats and dogs. These data including AUC, maximum
concentration, half-life, oral bioavailability, clearance, and
volume distribution are summarized in Table 5.
Conclusion
A novel series of potent NOP receptor ligands based on
C-3-monosubstituted tropane scaffold were discovered. In
general, these compounds showed better selectivity over
MOP compared to that of the previously reported C-3
carboxylamide series.10 Among the potent NOP ligands iden-
tified, compound10demonstratedthe best overallprofilewith
potent antitussive activity and less CNS effect compared to
that of 1. It showed antitussive activity at 6 h in the guinea pig
model, indicating that it could be a potential long-acting
antitussive agent. With these superior properties, compound
10 was advanced into preclinical toxicology evaluation.
In Vivo Studies
Compound 10 was further evaluated for anxiolytic-like9
and antitussive activities4a by means of oral administration
(Table 6). In the anxiolytic-like GPPV assay,9 10 displayed
a significant reduction of number of guinea pig vocalizations
under stress condition induced by separation at 3 mg/kg

Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5327
Experimental Section
tography with EtOAc-hexane as mobile phase, and pure
product 10 (633 mg, 17% yield) was obtained.
General Information. NMR spectra of the presented com-
pounds were recorded in CDCl3 on a Varian Unity XL-400
(400 MHz for 1H; 100 MHz for 13C) using standard Varian pulse
sequence programs. All chemicals purchased from commercial
source (e.g., Acros or Aldrich) were used without further pur-
ification. All compounds tested for in vitro and in vivo assays
were hydrochloride salts, which were prepared by mixing the
free base with one or multiple eqivalent of 1 M hydrochloric
acid in ether followed by evaporation of ether or filtration.
Final compounds were analyzed for their purity from LC-ESIMS
instruments coupled with UV detector. Most of the presented
compounds including key compounds (e.g., 10, 11, 12, 27, 28, 29,
30, and 32) showed purity >95%. The compounds with purity
less than 95% are 9 (92%), 13 (90%), 14 (92%), 15 (91%), 17
(91%), and 18 (90%). The purity of the compound was deter-
mined by LC/MS analysis using an Applied Biosystems API-
150Ex mass spectrometer and Shimadzu SCL-10A LC column
(Altech platinum C18, 33 mm ꢀ 7 mm). A gradient flow was used
as follows: 0 min, 10% MeCN; 5 min, 95% MeCN).
exo-8-[Bis(2-chlorophenyl)methyl]-3-(5-fluoro-2-pyridinyl)-
8-azabicyclo[3.2.1]octane (18) and exo-6-[8-[Bis(2-chlorophenyl)-
methyl]-8-azabicyclo[3.2.1]oct-3-yl]-3-pyridinol (21). Compounds
18 and 21 were prepared according to general methods 1 and
2 using 2-bromo-5-fluoropyridine. The corresponding nitrile:
ESI-MS: m/z 466 (C26H22Cl2FN3 Hþ). A mixture of the nitrile
3
(67 mg, 0.144 mmol), NaOH (400 mg), and ethylene glycol (4 mL)
was stirred and heated to 150 °C overnight. After cooled to rt, the
mixture was distributed toH2OandEtOAc. The organiclayerwas
washed with brine, dried over Na2SO4, filtered, and evaporated to
dryness. Purification of the residue by SiO2 chromatography
(0-100% EtOAc/hexane) gave 18 (∼2 mg, 90% purity, 3% yield).
1H NMR (CDCl3) δ 1.70 (m, 4H), 1.92 (t, J = 12.4 Hz, 2H), 2.22
(m, 2H), 3.03 (tt, J = 11.8, 5.2 Hz, 1H), 3.11 (brs, 2H), 5.47 (s,
1H), 7.00-7.3 (m, 8H), 7.85 (d, J = 6.7 Hz, 2H), 8.22 (s, 1H). LC/
ESI-MS: m/z 441 (C25H23Cl2FN2 Hþ) and 21 (5 mg, 8% yield).
3
1H NMR(CDCl3) δ 1.60 (m, 2H), 1.68 (dd, J = 14.3, 6.4 Hz, 2H),
1.90 (t, J = 12.4 Hz, 2H), 2.20 (m, 2H), 3.08 (m, 3H), 5.45 (s, 1H),
7.06 (td, J = 7.6, 1.6 Hz, 2H), 7.15 (t, J = 7.3 Hz, 2H), 7.25 (m,
4H), 7.81 (d, J=7.7 Hz, 2H), 8.12 (d, J=2.6 Hz, 1H). LC/ESI-
General Method 1. endo-8-[Bis(2-chlorophenyl)methyl]-3-(2-
pyridinyl)-8-azabicyclo[3.2.1]octane-3-carbonitrile (4a). To a
stirred solution of the nitrile (3) (20 g, 53.9 mmol) in 100 mL
THF was added NaHMDS/THF (40.4 mL, 2M) dropwise
at -78 °C under N2 atmosphere. The solution was stirred
at -78 °C for ∼1 h, and then 2-bromopyridine (17 g, 108 mmol)
in THF (20 mL) was added dropwise. After stirring for another
hour at this temperature, the reaction flask was moved to an
acetonitrile/dry ice bath. The reaction mixture was slowly
warmed to room temperature and stirred at room temperature
overnight. It was quenched with sat. aq NH4Cl at -78 °C and
extracted with EtOAc at room temperature,. The combined
organic layer was dried over Na2SO4, filtered, and evaporated to
dryness. The residue was washed with ether several times to give
the desired compound (19.1 g, ∼79% yield), which was used
in the next reaction without further purification. Nitrile 4a
(2-pyridinyl analogue): 1H NMR (CDCl3) δ 2.11 (brd, J =
12.1 Hz, 2H), 2.35 (brs, 4H), 2.54 (dd, J = 13.8, 2.9 Hz, 2H), 3.22
(brs, 2H), 5.54 (s, 1H), 7.09 (ddd, J = 9.3, 7.6, 1.7 Hz, 2H), 7.21
(m, 3H), 7.26 (dd, J = 7.9, 1.5 Hz, 2H), 7.58 (ddd, J = 8.1, 1.7,
0.9 Hz, 1H), 7.68 (ddd, J = 9.6, 7.8, 1.8 Hz, 1H), 7.80 (dd, J=
7.9, 1.7 Hz, 2H), 8.62 (ddd, J = 2.7, 1.8, 0.9 Hz, 1H). LC/ESI-
MS: m/z 439 (C25H24Cl2N2O Hþ)
3
exo-8-[Bis(2-chlorophenyl)methyl]-3-phenyl-8-azabicyclo[3.2.1]-
octane (29). To a mixture of 3 (580 mg, 1.56 mmol) and fluo-
robenzene (∼1.5 mL) was added potassium bis(trimethyl-
silyl)amide (580 mg) in fluorobenzene (∼2.5 mL) under N2
atmosphere. The mixture was prestirred for 10 min and then
microwaved at 100 °C for 18 min. After cooled to rt, the mixture
was quenched with saturated aq NH4Cl and partitioned with
EtOAc. The organic layer was dried over Na2SO4, filtered, and
concentrated. The residue was washed with ether to give the
corresponding nitrile (∼450 mg, 65% yield). LC/ESI-MS:
m/z 447 (C27H24Cl2N2 Hþ). To a stirred solution of the nitrile
3
(310 mg, 0.693 mmol) in THF was added 1 M LiALH4/THF
solution (0.693 mL, 0.693 mmol) dropwise at rt under N2 atmo-
sphere. The mixture was warmed to 60 °C and stirred overnight.
The following solutions were added sequentially: 50 μL of water,
150 μL of 15% NaOH, and 50 μL of water. The mixture was
stirred, filtered, and evaporated to dryness. Purification of the
residue by SiO2 chromatography (0-50% EtOAc/hexane) gave
29 (∼80 mg, 27% yield). 1H NMR (CDCl3) δ 1.58 (dt, J = 11.8,
3.3 Hz, 2H), 1.69 (dd, J = 14.1, 6.7 Hz, 2H), 1.86 (t, J = 12.5 Hz,
2H), 2.22 (m, 2H), 2.88 (tt, J = 12.5, 5.9 Hz, 1H), 3.09 (brs, 2H),
5.50 (s, 1H), 7.09 (t, J = 7.3 Hz, 2H), 7.14 (m, 1H), 7.21 (t, J = 8.0
Hz, 2H), 7.26 (m, 6H), 7.84 (d, J = 8.3 Hz, 2H). 13C NMR
(CDCl3) 27.5, 35.4, 40.2, 58.2, 60.0, 126.0, 126.8, 127.2, 127.9,
128.3, 129.4, 130.5, 134.5, 140.1, 146.2. LC/ESI-MS: m/z 422
MS: m/z 448 (C26H23Cl2N3 Hþ). HRMS (ESI) calcd for
3
C26H24Cl2N3, 448.1347 (M þ Hþ); found, 448.1360.
General Method 2. exo-8-[Bis(2-chlorophenyl)methyl]-3-(2-
pyridinyl)-8-azabicyclo[3.2.1]octane (10). A mixture of nitrile
4a (5 g, 11.2 mmol), NaOH (20 g, 500 mmol), and ethylene
glycol (40 mL) in a round-bottom flask was stirred and heated to
190-200 °C under N2 atmosphere for two to three days. After
reaction completed, the mixture was cooled to rt and dissolved
in 1N HCl solution. The suspension was partitioned with
dichloromethane under basic condition. The organic layer was
dried over Na2SO4, filtered, and evaporated to dryness. Purifi-
cation of the residue by SiO2 chromatography (EtOAc/hexane)
gave compound 10 (4.3 g, ∼91% yield, 99.5% purity). LC/ESI-
MS: m/z 423 (C25H24Cl2N2.Hþ). Compound 10: 1H NMR
(CDCl3) δ 1.67 (m, 2H), 1.72 (m, 2H), 1.97 (td, J = 12.5, 2.5
Hz, 2H), 2.24 (dd, J = 7.0, 2.5 Hz, 2H), 3.07 (tt, 12.0, 5.5 Hz, H-
3, 1H), 3.13 (brs, H-1, 2H), 5.48 (s, 1H), 7.07 (m, 3H), 7.22 (m,
5H), 7.57 (td, J = 7.7, 1.8 Hz, 1H), 7.86 (dd, J = 8.0, 1.5 Hz 2H),
8.50 (ddd, J = 4.9, 1.8, 0.9 Hz, 1H).
(C26H25Cl2N Hþ).
3
2-Bromo-3-bromomethyl-pyridine (6). A mixture of 2-bromo-3-
methylpyridine (1.114 mL, 10 mmol), NBS (1780 mg, 10 mmol),
and benzoyl peroxide (45 mg) in CCl4 (30 mL) was refluxed for 3
h. After cooling to rt, the suspension was filtered. Purification of
the residue by SiO2 chromatography (EtOAc/hexane) gave the
desired compound 6 (600 mg, 24% yield). LC-ESIMS: m/z 250,
252, and 254 (C6H5Br2N Hþ)
3
2-Bromo-3-(1-piperidinylmethyl)pyridine (7). To a solution of
2-bromo-3-bromomethyl-pyridine (6, 595 mg, 2.36 mmol) in
DMF (10 mL) was added piperidine (205 mg, 2.4 mmol) and
K2CO3 (979 mg, 7.08 mmol), sequentially. The mixture was
stirred at rt overnight, quenched with ice-water, and then
partitioned with ether. The organic layer was dried over
Na2SO4, filtered, and concentrated. Purification of the residue
by SiO2 chromatography (0-50% EtOAc/hexane) gave the
desired compound 7 (∼450 mg, 75% yield). C11H15BrN2, LC-
General Method 3. To a stirred solution of pyridyl-nitrile 4a
(4 g, 8.93 mmol) in THF (40 mL) was added 1 M LiALH4/THF
solution (9.8 mL, 9.8 mmol) dropwise at rt under N2 atmo-
sphere. The mixture was stirred at 50 °C for one hour. After
cooled to rt, the following solutions were added sequentially:
380 μL of water, 1440 μL of 15% NaOH, and 380 μL of water.
The mixture was stirred, filtered, and evaporated to dryness.
The crude material was purified through Si gel column chroma-
ESIMS: m/z 255 and 257 (C11H15BrN2 Hþ)
3
exo-8-[Bis(2-chlorophenyl)methyl]-3-[3-(1-piperidinylmethyl)-
2-pyridinyl]-8-azabicyclo[3.2.1]octane (9). Compounds 8 and
9 were prepared according to the general methods 1 and 3
using 2-bromo-3-(1-piperidinylmethyl)pyridine (7). The nitrile

5328 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17
Yang et al.
intermediate 8 (72% yield). 1H NMR (CDCl3) δ 1.43 (m, 2H),
1.52 (m, 4H), 2.40 (m, 10H), 2.65 (dd, J = 14.0, 2.8 Hz, 2H), 3.24
(brs, 2H), 3.78 (s, 2H, -CH2N-), 5.52 (s, 1H), 7.08 (td, J=7.8,
1.7 Hz, 2H), 7.20 (m, 3H), 7.26 (dd, J = 7.9, 1.3 Hz, 2H), 7.79 (d,
J = 7.7 Hz, 2H), 8.08 (dd, J = 7.8, 1.3 Hz, 1H), 8.44 (dd, J=4.6,
extracted with ether. The organic solution was dried over Na2SO4,
filtered, and concentrated. Purification of the residue by SiO2
chromatography gave 15 (9.4 mg, 91% purity, 21% yield). H
1
NMR (CDCl3) δ 1.1-1.7 (m, 13H), 2.0-2.3 (m, 3H), 2.38 (brd,
J = 14.1 Hz, 1H), 2.67 (m, 1H), 2.76 (m, 1H), 2.86 (td, 1H), 3.00
(brs, 2H), 3.46 (t, J=5.3 Hz, 2H), 5.39 (s, 1H), 7.07 (t, J = 7.5 Hz,
2H), 7.19 (m, 2H), 7.23 (m, 2H), 7.75 (dd, J = 7.2, 2H). LC/ESI-
1.7 Hz, 1H). 13C NMR (CDCl3)
δ 24.5, 26.2, 26.5,
38.5, 40.7, 54.5, 57.8, 59.8 (2C), 122.8, 125.0, 126.8, 128.0,
129.5, 130.1, 133.7, 134.4, 139.0, 139.4, 146.3, and 156.0 ppm.
MS: m/z 473 (C27H34Cl2N2O Hþ)
3
LC/ESI-MS: m/z 545 (C32H34Cl2N4 Hþ); compound 9 (purity
exo-8-[Bis(2-chlorophenyl)methyl]-3-(5-oxazolyl)-8-azabicyclo-
[3.2.1]octane (17). To the solution of nitrile 3 (1484 mg, 4 mmol)
in toluene (20 mL) was added 1.5 M DIBAL solution in toluene
(5.87mL, 8.8mmol) at0°CunderN2 atmosphere. After stirred for
three hours at this temperature, reaction was brought to -78 °C
and quenched with 1.8 mL of MeOH and 2.7 mL saturated aq
NH4Cl solution. The mixture was brought to room temperature
and stirred. EtOAc and 1N NaOH were added to the mixture for
partition. The organic layer was combined and dried over Na2SO4,
filtered, and evaporated to get crude material. This material was
further mixed with 1N HCl and then stirred at 60 °C overnight.
The mixture was adjusted to a basic solution with 1N NaOH
solution, and then extracted with ether (3ꢀ). Organic layer was
combined and dried over Na2SO4, filtered, and evaporated to get
product 16 (1 g, 67% yield), which was used directly in the next
3
1
92%, 20% yield): H NMR (CDCl3) δ 1.46 (m, 8H), 1.72 (dd,
J = 14.2, 6.3 Hz, 2H), 2.10 (t, J = 12 Hz, 2H), 2.27 (m, 6H),
3.13 (m, 2H), 3.35 (s, 2H), 3.57 (tt, J = 12.2, 5.0 Hz, 1H), 5.43
(s, 1H), 6.97 (dd, J=7.6, 4.7 Hz, 1H), 7.06 (m, 2H), 7.18 (m, 2H),
7.23 (m, 2H), 7.39 (d, J = 7.2 Hz, 1H), 7.95 (d, J = 8.2 Hz, 2H),
8.50 (dd, J = 5, 2 Hz, 1H). LC/ESI-MS: m/z 520 (C31H35-
Cl2N3 Hþ)
3
exo-8-[Bis(2-chlorophenyl)methyl]-3-(2-piperidinyl)-8-azabi-
cyclo[3.2.1]octane (11). To a solution of 10 (200 mg, 0.474 mmol)
in dichloromethane (10 mL) was added PtO2 (40 mg). The
mixture was stirred at room temperature under 1 atm H2
environment through a balloon for ∼24 h, filtered, and con-
centrated. Purification of the residue by SiO2 column chroma-
1
tography gave compound 11 (∼190 mg, 94% yield). H NMR
(CDCl3) δ 1.20-2.00 (12H), 2.08 (m, 2H), 2.20 (m, 1H), 2.71 (m,
3H), 3.04 (m, 2H), 3.41 (d, J = 12.3 Hz, 1H), 5.38 (brs, 1H), 7.09
(m, 2H), 7.19 (m, 2H), 7.25 (m, 2H), 7.71 (m, 2H). LC/ESI-MS:
reaction. ESI-MS: m/z 374 (C21H21Cl2NO Hþ). To a solution of
3
16 (75 mg, 0.2 mmol) in MeOH (3 mL) was added K2CO3 (83 mg,
0.6 mmol) and tosylmethyl isocyanide (39.1 mg, 0.2 mmol). The
mixture was refluxed under nitrogen for 2 h, cooled to rt, and
extracted with EtOAc. The organic solution was dried over
Na2SO4 and concentrated. Purification of the residue by SiO2
m/z 429 (C25H30Cl2N2 Hþ).
3
exo-8-[Bis(2-chlorophenyl)methyl]-3-(1-methyl-2-piperidinyl)-
8-azabicyclo[3.2.1]octane (12). A suspension of 11 (30 mg,
0.07 mmol) in formic acid (100 μL) and 37% aq formaldehyde
(200 μL) was stirred and heated at 70 °C for ∼7 h. The mixture
was evaporated to dryness and then distributed to EtOAc and
1 N NaOH solution. The organic layer was dried over Na2SO4,
filtered, and concentrated. Purification of the residue by SiO2
1
chromatography gave 17 (11.5 mg, 91% purity, 14% yield). H
NMR (CDCl3) δ 1.64 (m, 2H), 1.68 (m, 2H), 1.82 (t, J=12.5 Hz,
2H), 2.24 (m, 2H), 3.02 (tt, J= 11.8, 5.9 Hz, 1H), 3.09 (brs, 2H),
5.45(s, 1H), 6.71(s, 1H), 7.09(t, J=7.1 Hz, 2H), 7.22 (m, 4H), 7.72
(s, 1H), 7.79 (d, J = 7.4 Hz, 2H). LC/ESI-MS: m/z 413
(C23H22Cl2N2O Hþ)
1
chromatography gave 12 (17.8 mg, 58%). H NMR (CDCl3)
3
δ 1.10-1.65 (∼12H), 1.72 (t, J = 13.2 Hz, 2H), 2.10 (m, 3H),
2.19 (s, 3H), 2.83 (d, J = 11.8 Hz, 1H), 3.01 (brs, 2H), 5.42 (s,
1H), 7.07 (t, J = 7.6 Hz, 2H), 7.17 (t, J = 6.8 Hz, 2H), 7.23 (d,
J = 7.7 Hz, 2H), 7.77 (dd, J = 6.7 Hz, 2H). LC/ESI-MS: m/z
Acknowledgment. We acknowledge Dr. Geoffrey Varty
and his group for providing anxiolytic-like activity data, Dr.
Jianshe Kong, Dr. Jesse Wong, and Meng Tao for prepara-
tion of the intermediates, Dr. Tze-Ming Chan and his group
for structure confirmation of some analogues, Dr. Xiaoming
Cui and the DMPK group for acquiring hPXR and pharma-
cokinetic data, and Drs. Steve Sorota and Tony Priestley for
providing hERG data.
443 (C26H32Cl2N2 Hþ).
3
1-Acetyl-2-[exo-8-[bis(2-chlorophenyl)methyl]-8-azabicyclo[3.2.1]-
oct-3-yl]piperidine (13). Amixtureof11 (34 mg, 0.0784 mmol), Ac2O
(0.2 mL), and pyridine (0.2 mL) was stirred at 0 °C for 12 h. The
solvent was removed in vacuo. Purification of the residue by SiO2
chromatography gave 13 (13 mg, 90% purity, 35% yield). 1H NMR
(CDCl3) (rotamers observed) δ 1.10-1.80 (12H), 2.01 and 2.09 (s,
Me, 3H), 2.16 (m, 3H), 3.01 (m, 3H), 3.49 (m, 1H), 4.47 (m, 1H),
5.35 and 5.44 (s, 1H), 7.08 (m, 2H), 7.22 (m, 4H), 7.75 (m, 2H). LC/
Supporting Information Available: Information for charac-
terization of compounds 19, 20, 22-28, and 30-32 and refer-
ences for biological evaluation. This material is available free of
charge via the Internet at http://pubs.acs.org.
ESI-MS: m/z 471 (C27H32Cl2N2O Hþ).
3
2-[exo-8-[Bis(2-chlorophenyl)methyl]-8-azabicyclo[3.2.1]oct-
3-yl]-1-piperidineacetonitrile (14). A mixture of 11 (55 mg,
0.128 mmol) and potassium carbonate (53 mg, 0.385 mmol)
in DMF (1.5 mL) was stirred at room temperature for 20 min,
and then bromoacetonitrile (17.8 μL, 0.256 mmol) was added.
The mixture was stirred at rt for additional 30 min and then
heated at 60 °C overnight. The mixture was cooled to rt,
quenched with water extracted with ether, dried over MgSO4,
filtered, and concentrated. Recrystallization of the residue in
ether gave 14 (16 mg, 92% purity, 27% yield). 1H NMR (CDCl3)
δ 1.08-1.68 (m, 10H), 1.75 (t, J = 14.7 Hz, 2H), 2.00 (m, 1H),
2.14 (m, 3H), 2.46 (td, J = 11.5, 2.8 Hz, 1H), 2.73 (d, J = 10.8
Hz, 1H), 3.03 (brs, 2H), 3.32 (d, J = 16.9 Hz, 1H), 3.76 (d, J=
16.9 Hz, 1H), 5.43 (s, 1H), 7.08 (td, J = 7.7, 1.6 Hz, 2H), 7.19 (t,
J = 7.7 Hz, 2H), 7.23 (d, J = 8.0 Hz, 2H), 7.76 (dd, J = 7.3, 2H).
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