Synthesis and biological evaluation of 2-(arylethynyl)quinoline derivatives as mGluR5 antagonists for the treatment of neuropathic pain

Article abstract of DOI:10.1016/j.bmcl.2012.12.056

We described here the synthesis and biological evaluation of mGluR5 antagonists containing a quinoline ring structure. Using intracellular calcium mobilization assay (FDSS assay), we identified compound 5n, showing high inhibitory activity against mGluR5. In addition, it was found that compound 5n has excellent stability profile. Finally, this compound exhibited favorable analgesic effects in spinal nerve ligation model of neuropathic pain, which is comparable to gabapentin.

Full text of DOI:10.1016/j.bmcl.2012.12.056

Bioorganic & Medicinal Chemistry Letters 23 (2013) 1472–1476
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of 2-(arylethynyl)quinoline
derivatives as mGluR5 antagonists for the treatment of neuropathic
pain
Myung-Hee Son a,b, Ji Young Kim a,c, Eun Jeong Lim a, Du-Jong Baek b, Kihang Choi c, Jae Kyun Lee a,
Ae Nim Pae a,d, Sun-Joon Min a,d, , Yong Seo Cho a,d,
⇑
⇑
a Center for Neuro-Medicine, Brain Science Institute, Korea Institute of Science and Technology (KIST), Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791, Republic of Korea
b Department of Chemistry, College of Natural Sciences, Sangmyung University, Hong-ji moon 2-gil 20, Jongno-gu, Seoul 110-743, Republic of Korea
c Department of Chemistry, Korea University, Seoul 136-701, Republic of Korea
d Biomolecular Science, University of Science and Technology (UST), 176 Gajung-dong, 217 Gajungro, Yuseong-gu, Daejeon 305-333, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
We described here the synthesis and biological evaluation of mGluR5 antagonists containing a quinoline
ring structure. Using intracellular calcium mobilization assay (FDSS assay), we identified compound 5n,
showing high inhibitory activity against mGluR5. In addition, it was found that compound 5n has excel-
lent stability profile. Finally, this compound exhibited favorable analgesic effects in spinal nerve ligation
model of neuropathic pain, which is comparable to gabapentin.
Received 9 November 2012
Revised 13 December 2012
Accepted 15 December 2012
Available online 22 December 2012
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
Metabotropic glutamate receptor
Antagonist
Quinoline
Sonogashira reaction
Neuropathic pain
Glutamate, the major excitatory neurotransmitter in the mam-
malian central nervous system, is involved in a wide range of phys-
iological functions acting through various glutamatergic
transmission.1 At the synaptic level, its transmission of neuronal
signals is mediated by two main families of glutamate receptors:
ionotropic receptors (iGluRs) and metabotropic glutamate recep-
tors (mGluRs). The iGluRs, known as ligand-gated ion channels,
are classified into three different receptors on the basis of the
mGluR8) are located in presynaptic regions and inhibit activated
adenylate cyclase activity by coupling to a Gi protein.3 mGluR5 is
highly expressed in brain regions and its modulation represents a
potential therapeutic approach for treatment of a wide range of
CNS-related disorders including pain,4 anxiety,5 Parkinson’s dis-
ease6 and drug dependency.7
There has been a huge study to indentify selective mGluR5
modulators suitable for clinical use (Fig. 1). For example, MPEP,
2-methyl-6-(phenylethynyl)-pyridine8 was first reported as a po-
tent, selective mGluR5 NAM (negative allosteric modulator) show-
ing high efficacies in various preclinical disease models, such as
pain, anxiety, and depression although it did not proceed to clinical
trials due to low pharmacokinetic profile. In addition, different
kinds of mGluR5 NAMs such as fenobam,9 ADX10059,10 and
AFQ05611 have been discovered and known to be effective for Frag-
ile X syndrome, gastro-esophageal reflux disease (GERD), and mi-
graine. Many of mGluR5 antagonists contain an alkyne subunit as
a key structural component. Much effort has focused on synthesis
of compounds bearing acetylene itself or its isostere.12 Despite the
substantial activity of the reported acetylenic analogues, however,
the identification of selective and clinically efficacious mGluR5
antagonists still remains a challenge.
interaction with their specific ligands, that is, N-methyl-D-aspartic
acid (NMDA), kainate, and (S)-2-amino-3-(3-hydroxy-5-methyl-4-
isoxazolyl)propionic acid (AMPA). In general, they are responsible
for fast excitatory synaptic transmission and plasticity. On the
other hand, the mGluRs are members of the G-protein-coupled
receptors (GPCRs) and divided into three groups according to their
sequence homology, receptor pharmacology, and signal transduc-
tion pathways.2 Currently, eight distinct mGluR subtypes are dis-
covered. Group I consists of mGluR1 and mGluR5, which are
mainly distributed in postsynaptic regions and positively coupled
to phospholipase C via a Gq protein, whereas group II (mGluR2
and mGluR3) and group III (mGluR4, mGluR6, mGluR7, and
In this context, we report the synthesis and biological evalua-
tion of novel acetylenic quinoline derivatives as selective mGluR5
⇑
Corresponding author.
E-mail addresses: sjmin@kist.re.kr (S.-J. Min), ys4049@kist.re.kr (Y.S. Cho).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.12.056

M.-H. Son et al. / Bioorg. Med. Chem. Lett. 23 (2013) 1472–1476
1473
Me
N
H2N
Me
N
H
N
Cl
N
N
Me
HO
Me
F
O
HN
O
MPEP
Fenobam
ADX10059
O
Me
Me
H
O
N
N
Me
H
OMe
AFQ-056
MRZ-8676
Figure 1. Representative mGluR5 antagonists.
antagonists including in vivo study of a lead compound in animal
models of neuropathic pain.
reaction of 5-aminoquinoline 3,14 2-chloroquinoline derivatives 4
having different substituents at the 5-postion were synthesized
via N-oxidation followed by chlorination. Installation of arylethynyl
groups was conducted in three different ways. Sonogashira reaction
of 4 with arylethyne15 directly afforded the 5-substituted 2-(aryle-
thynyl)quinolines 5 (method A).16 Alternatively, the 2-arylethynyl
components were introduced by a stepwise procedure. Thus, palla-
dium-catalyzed coupling reaction of 2-chloroquinolines 4 with
2-methylbut-3-yn-2-ol or ethynyltrimethylsilane gave the C-termi-
nal protected ethynyl derivatives 7 and 8. Compound 7 was sub-
jected to another Sonogashira reaction in the presence of
potassium hydroxide to give its corresponding 2-ethynylquinolines
5 or 6 through in situ deprotection of 2-propanol group (method
B).17 Removal of trimethylsilyl group of 8 and subsequent Sono-
gashira reaction produced the desired 2-(arylethynyl)quinolines 6
containing pyridine or pyrimidine rings (method C).
Considering the previously reported mGluR5 NAMs, we first de-
signed 2-(arylethynyl)quinoline as a potential starting scaffold. We
reasoned that a quinoline group can act as a potential isostere of
heteoaromatic groups existing in most acetylene-based mGluR5
NAMs such as MPEP, MTEP, and AFQ056. It has been reported that
MRZ-867613 having a hydroquinolinone scaffold (Fig. 1) showed a
significant negative modulation against mGluR5 with an IC50 value
of 20 nM, which also supports our idea that the quinoline structure
may be a suitable scaffold for an alternative pharmacophore.
Moreover, we thought that introduction of various substituents
to the quinoline ring system provides a good strategy to lead
optimization through potency improvement or physicochemical
modulation.
The synthesis of 2-(arylethynyl)quinoline derivatives is depicted
in Scheme 1. Starting from 5-substituted quinolines 2, prepared
from alkylation/acylation of 5-hydroxyquinoline 1 or Sandmeyer
In vitro activities of our synthesized compounds against
mGluR5 were determined by the ability of the compound to inhibit
R1
OH
i
h
N
OH
N
method B
1
2
Me Me
7
a or b
R1
R1
R1
g
e,f
N
N
method A
N
N
Cl
Ar
4
5a-r
6a-q
X
Y
c or d
Ar:
R1
X
R2
NH2
j
N
l
method C
N
N
N
R
k
N
3
8
9
(R = TMS)
(R = H)
R1 = H, OH, MeO, EtO, tBuCO2, Cl, Br
X, Y = CH or N
R
2 = F, Me, CN, CF3, MeO
Scheme 1. Reagents and conditions: (a) TMSCHN2, MeOH, rt, 42% or K2CO3, EtI, DMF, 89%; (b) PvCl, pyridine, CH2Cl2, rt, 84%; (c) CuCl, aq HCl, NaNO2, 0 °C to rt, 68%; (d) CuBr,
NaNO2, 48% HBr, 0 °C to rt, 35%; (e) mCPBA, DCM, 0 °C to rt; (f) POCl3, CH2Cl2, reflux, 32–47% (2 steps); (g) arylacetylene, PdCl2(PPh3)2, CuI, Et3N, 80 °C, 9–100%; (h) 2-
methylbut-3-yn-2-ol, PdCl2(PPh3)2, CuI, Et3N, 80 °C, 52–100%; (i) aryl bromide, KOH, PdCl2(PPh3)2, CuI, iPr2NH, 95 °C, 8–35%; (j) TMS–acetylene, PdCl2(PPh3)2, CuI, Et3N, 60 °C,
93%; (k) K2CO3, CH2Cl2/MeOH, rt, 85%; (l) 2-bromopyrimidine or 5-bromopyrimidine, PdCl2(PPh3)2, CuI, Et3N, 80 °C, 11–32%

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M.-H. Son et al. / Bioorg. Med. Chem. Lett. 23 (2013) 1472–1476
Table 2
calcium mobilization caused by a high concentration of glutamate
in human mGluR5/HEK293 cells using FDSS6000 system.18 First,
we investigated the effect of substituents at the 5-position of quin-
olines (Table 1). 2-Phenylethynyl or 2-(pyridine-3-ylethynyl)quin-
olines having hydrogen and alkoxy groups at the 5-position (5a, b,
g–i, n) gave good inhibitory activities with higher than 60% and
In vitro inhibitory activity of the synthesized compounds against mGluR5
N
N
X
N
R2
N
30% inhibition values at the concentration of 10 and 1 lM,
6a-p
6q
% Inhibition (mGluR5)a
10
N
respectively. On the other hand, most 2-(pyridine-2-yl)quinoline
derivatives maintain the high negative modulation of mGluR5
except R = OH. This result indicated that hydrogen bonding accep-
tors at the 5-postion of quinoline moiety as well as pyridine-2-yl
group on the opposite side are crucial to interact with the mGluR5
receptor.
Based on the SAR of the first series, we then examined a second
set of compounds, which derived from compound 5n, to study the
influence of substituted pyridine-2-yl groups on the potency (Table
2). In general, inhibitory activities of most compounds in this series
against the mGluR5 were not superior or comparable to that of 5n
at 10 lM. Best results were obtained with small substituents such
as nitrile, fluorine, and methyl group at the 4- and 5-positions (6b,
c, e, f, h, i). Although we found that the compound 6e was the most
potent derivative of this set at 10 lM, its potency at 1 lM was two-
fold lower than that of 5n. In addition, replacement of the pyridin-
2-yl group with pyrimidin-2-yl or 5-yl group (6n–q) exhibited low
potency regardless of a substituent character, which also supports
that the position of nitrogen atom in the pyridine fragment of this
series is closely associated with the binding site of the mGluR5
receptor.
Compounds
R2
X
Methodb
l
M
1 lM
6a
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
6m
6n
6o
6p
6q
3-CF3
3-F
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
N
16.43
55.24
40.09
58.04
76.09
66.48
50.66
63.29
66.27
28.66
64.82
46.90
48.62
28.63
15.87
15.59
22.45
9.49
C
B
B
C
C
B
C
C
B
C
C
B
C
C
C
C
C
31.31
18.85
23.26
35.22
21.89
12.99
33.87
29.00
15.77
32.96
29.59
16.60
17.08
17.34
15.96
13.60
3-Me
4-CF3
4-CN
4-Me
5-CF3
5-F
5-Me
6-CF3
6-CN
6-F
6-MeO
H
4-CF3
5-F
N
N
a
Ca2+ flux assay using glutamate as agonist.
See Scheme 1 for synthetic methods.
b
Next, we selected several compounds with over 60% (10
lM)
and 50% (1 M) inhibition in the FDSS assay and examined their
l
whereas only the compound 5n displayed excellent
pharmacological properties in terms of microsomal stability
and CYP inhibition.
IC50 values, hERG inhibition, microsomal stability, and CYP
inhibition. The results are summarized in Table 3. In fact,
most of the compounds showed good IC50 values against
the mGluR5 and low activity against the hERG channel blockade
The pharmacokinetic parameters for compound 5n following
intravenous and oral administration in rats are presented in
Table 4. Despite of high stability in human liver microsomes
and CYP enzymes, compound 5n showed high clearance and
low bioavailability. The considerably high value of the mean
volume of distribution suggests that it tends to bind to tissue
components or plasma proteins. The brain to plasma ratios in
intravenous and oral administration were low and moderate,
which might be due to nonspecific binding in brain or high
clearance.
Table 1
In vitro inhibitory activity of the synthesized compounds against mGluR5
R1
Given the good in vitro potency and stability along with moder-
ate pharmacokinetic profile in rats, the in vivo efficacy of com-
pound 5n was evaluated in the SNL (spinal nerve ligation)
neuropathic pain model (Fig. 2).19 In this model, a neuropathic pain
state was induced by tight ligation of the L5 spinal nerve at a site
distal to the DRG. Two behavioral tests (mechanical allodynia
and cold allodynia) were performed after 14 days of surgical
manipulation.20 The rats were treated orally with 100 mg/kg of
compound 5n or gabapentin (a positive control). While 5n exhib-
ited high suppression effect on mechanical allodynia at 5 h, high
paw withdrawal response was observed in cold allodynia at 3 h.
This result showed that the efficacy of compound 5n is comparable
to that of Gabapentin in this behavior test. Therefore, it should be
further investigated as a viable mGluR5 antagonist for treatment of
neuropathic pain.
In conclusion, we have synthesized and evaluated 2-(arylethy-
nyl)quinolines 5 and 6 as potential mGluR5 antagonists. The SAR
study of acetylenic quinoline derivatives on the mGluR5 receptor
led to the identification of 5n, a compound showing high inhibitory
activity against mGluR5 and excellent stability profile. Despite rel-
atively low oral bioavailability, 5n was evaluated in animal model
of neuropathic pain and it significantly reduced both mechanical
allodynia and cold allodynia by oral administration, which is com-
parable to gabapentin. Based on the in vitro and in vivo data
N
X
5
Y
Compounds
R1
X
Y
% Inhibition (mGluR5)a
Methodb
10
l
M
1 lM
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
5q
5r
H
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
N
CH
CH
CH
CH
CH
CH
N
N
N
N
N
67.71
68.03
23.77
24.32
8.02
20.67
65.78
67.72
66.63
28.5
17.58
28.35
40.07
76.66
62.44
22.15
64.31
77.52
35.88
63.58
24.03
18.01
11.72
21.09
43.93
67.68
47.16
10.56
25.22
11.16
22.02
58.81
17.12
17.36
31.02
56.73
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
B
B
MeO
tBuCO2
OH
Cl
Br
H
MeO
EtO
tBuCO2
OH
Cl
Br
H
tBuCO2
OH
Cl
N
N
CH
CH
CH
CH
CH
N
N
N
N
Br
Ca2+ flux assay using glutamate as agonist.
See Scheme 1 for synthetic methods.
a
b

M.-H. Son et al. / Bioorg. Med. Chem. Lett. 23 (2013) 1472–1476
1475
Table 3
The results of IC50 (mGluR5 and hERG channel), microsomal stability, and CYP inhibition
a
a
Compounds
mGluR5 IC50
(lM)
hERG IC50
(l
M)
HLM% remaining @ 1
lM after 30 min
CYP (% remaining @ 10
lM)
CYP2D6
CYP2C9
CYP3A4
5b
5h
5n
5r
0.41 0.07
0.43 0.02
0.94 0.25
0.75 0.17
61.70 11.20
65.30 17.70
20.30 6.11
22.70 7.04
<1
<1
88
98
>99
86
98
55
50
29
77
56
>99
>99
52
78
a
IC50 value( SD) was obtained from a dose–response curve.
Table 4
Acknowledgements
Mean pharmacokinetic parameters in rat plasma following intravenous (n = 4) and
oral (n = 3) administration (10 mg/kg) of 5n
This research was supported by the Korea Institute of Science
and Technology (KIST) and by a grant of the Korea Health technol-
ogy R&D Project, the Ministry of Health and Welfare, Republic of
Korea (A111035).
Intravenous
Oral
Cmax
Tmax (min)
T1/2 (min)
CL (ml/min/kg)
Vdss (mL/kg)
B/P ratio at 2 h
F (%)
(
l
g/mL)
—
—
0.43( 0.18)
10 (5–30)a
48.53 ( 8.06)
58.24 ( 62.41)
84.93 8.98
1269.02 ( 158.55)
0.08
Supplementary data
—
0.35
15.9%
—
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2012.12.
056.
Values are presented as mean (standard deviation in parentheses). Cmax, peak
plasma concentration; Tmax, time to reach Cmax; Vdss, apparent volume of distribu-
tion at steady state; F, bioavailability.
a
Median (range) for Tmax
.
References and notes
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B. Mechanical allodynia (%MPE)
100
A. Mechanical allodynia
5n 100mg/kg (n=5)
15
Gabapentin 100mg/kg (n=4)
80
60
40
20
0
*
12
*
9
*
6
3
0
Base
1h
3h
5h
Pre N14D 1h
3h
5h
Experimental time (hour)
Experimental time (day or hour)
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C. Cold allodynia
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Gabapentin 100mg/kg (n=4)
210
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*
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*
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Experimental time (hour)
Experimental time (day or hour)
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neuropathic pain-induced rats. Experimental time expressed as D for days after neuropathic injury (N) and h for hours after gabapentin or 5n administration, ⁄P <0.05
(gabapentin), #P <0.05 (5n) versus pre-administration value (paired t-test), |P <0.05 gabapentin versus 5n (unpaired t-test).

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(dd, J = 7.8, 7.2 Hz, 1H), 7.70–7.79 (m, 4H), 7.84 (d, J = 8.4 Hz, 1H), 8.17 (t,
J = 8.7 Hz, 2H), 8.69 (d, J = 4.8 Hz, 1H); 13C NMR (CDCl3, 75 MHz) d 88.3, 88.4,
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18. Experimental procedure for in vitro assay. Human embryonic kidney cells which
stably express mGluR5 were obtained from Yonsei University. Cells were
grown in DMEM medium supplemented with 10% (v/v) fetal bovine serum,
penicillin (100 U/mL), streptomycin (100 lg/mL), and puromycin (10 lg/mL) at
37 °C in a humid atmosphere of 5% CO2 and 95% air. For calcium assay, cells
were harvested and dispensed into 96-well black wall clear bottom plates at a
density of 40,000 cells per a well. After 18 h of incubation, cells were treated
with Calcium-5 assay reagent, which is prepared by manufacture’s instruction
(Molecular Devices Corporation, California). During fluorescence-based
FDSS6000 assay, mGluR5 was activated using
a high concentration of L-
glutamate (10 M) in HBSS, and various concentrations of synthesized
l
9. Porter, R. H.; Jaeschke, G.; Spooren, W.; Ballard, T. M.; Büttelmann, B.;
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20. Experimental procedure for in vivo assay. Two behavioral tests (mechanical
allodynia and cold allodynia) were conducted for rats at 1 day prior to surgery
and 14 days after surgery. After the postoperative behavioral test, the animals
were treated orally with 50 mg/kg compound 5n or gabapentin. The tests were
re-evaluated at 1 h, 3 h, and 5 h after administration. Mechanical allodynia:
Testing was performed according to methods described in the literature
(Chaplan, S. R.; Bach, F. W.; Pogrel, J. W.; Chung, J. M.; Yaksh, T. L. J. Neurosci.
Methods 1994, 53, 55). Rats were acclimated in a transparent plastic boxes
permitting freedom of movement with a wire mesh floor to allow access to the
planter surface of the hind paws. A von Frey filament (Stoelting, Wood Dale, IL)
was applied 5 times (once every 3–4 s) to hind paw. Von Frey filaments were
used to assess the 50% mechanical threshold for paw withdrawal. The 50%
withdrawal threshold was determined by using the up-down method and
formula given by Dixon (Dixon, W. J. Ann. Rev. Pharmacol. Toxicol. 1980, 20,
441): 50% threshold = 10 (X + kd)/104, where X is the value of the final von Frey
hair used (in log units), k is the tabular value for the pattern of positive/
negative responses modified from the same literature, and d is the mean
difference between stimuli in log units (0.17). Cold allodynia: To quantify cold
sensitivity of the paw, brisk paw withdrawal in response to acetone application
was measured as reported previously (Choi, Y.; Yoon, Y. W.; Na, H. S.; Kim, S.
H.; Chung, J. M. Pain 1994, 59, 369). The rat was placed under a transparent
plastic box on a metal mesh floor and acetone was applied to the plantar
surface of the hind paw. To do this, an acetone bubble was formed at the end of
a small piece of polyethylene tubing which was connected to a syringe. The
bubble was then gently touched to the heel. The acetone quickly spread over
the proximal half of the plantar surface of the hind paw. The acetone was
11. Levenga, J.; Hayashi, S.; De Vrij, F. M.; Koekkoek, S. K.; Van Der Linde, H. C.;
Nieuwenhuizen, I.; Song, C.; Buijsen, R. A.; Pop, A. S.; Gomezmancilla, B.;
Nelson, D. L.; Willemsen, R.; Gasparini, F.; Oostra, B. A. Neurobiol. Dis. 2011, 42,
311.
12. For recent mGluR5 antagonists or NAMs, see: (a) Pilla, M.; Andreoli, M.; Tessari,
T.; Delle-Fratte, S.; Roth, A.; Butler, S.; Brown, F.; Shah, P.; Bettini, E.; Cavallini,
P.; Benedetti, R.; Minick, D.; Smith, P.; Tehan, B.; D’Alessandro, P.; Lorthioir, O.;
Ball, C.; Garzya, V.; Goodacre, C.; Watson, S. Bioorg. Med. Chem. Lett. 2010, 20,
7521; (b) Spanka, C.; Glatthar, R.; Desrayaud, D.; Fendt, F.; Orain, D.; Troxler, T.;
Vranesic, I. Bioorg. Med. Chem. Lett. 2010, 20, 184; (c) Weiss, J. M.; Jimenez, H.
N.; Li, G.; April, M.; Uberti, M. A.; Bacolod, M. D.; Brodbeck, R. B.; Doller, D.
Bioorg. Med. Chem. Lett. 2011, 21, 4891; (d) Gilbert, A. M.; Bursavich, M. G.;
Lombardi, S.; Adedoyin, A.; Dwyer, J. M.; Hughes, Z.; Kern, J. C.; Khawaja, X.;
Rosenzweig-Lipson, S.; Moore, W. J.; Neal, S. J.; Olsen, M.; Rizzo, S. J. S.;
Springer, D. Bioorg. Med. Chem. Lett. 2011, 21, 195; (e) Alagille, D.; DaCosta, H.;
Chen, Y.; Hemstapat, K.; Rodriguez, A.; Baldwin, R. M.; Conn, J. P.; Tamagnan, G.
D. Bioorg. Med. Chem. Lett. 2011, 21, 3243.
13. Dekundy, A.; Gravius, A.; Hechenberger, M.; Pietraszek, M.; Nagel, J.; Tober, C.;
van der Elst, M.; Mela, F.; Parsons, C. G.; Danysz, W. J. Neural. Transm. 2011, 118,
1703.
14. Karramkam, M.; Dolle, F.; Valette, H.; Besret, L.; Bramoulle, Y.; Hinnen, H.;
Vaufrey, F.; Franklin, C.; Bourg, S.; Coulon, C.; Ottaviani, M.; Delaforge, M.;
Loc´h, C.; Bottlaendera, B.; Crouzela, C. Bioorg. Med. Chem. Lett. 2002, 10, 2611.
15. Zou, H.; Zhou, L.; Li, Y.; Cui, Y.; Zhong, H.; Pan, Z.; Yang, Z.; Quan, J. J. Med. Chem.
2010, 53, 994.
16. Experimental procedure for synthesis of compound 5n. To
a
solution of 2-
applied 5 times to hind paw at 2 min interval. The frequency of paw
chloroquinoline (871 mg, 5.26 mmol) in THF (1.5 mL) was added PdCl2(PPh3)2
(183 mg, 0.26 mmol), CuI (110 mg, 0.58 mmol). The reaction mixture was
stirred for 5 min and triethylamine (14.4 mL) and 2-ethynylpyridine (1.05 mL,
10.4 mmol) were added. After the resulting mixture was stirred at 100 °C for
48 h, it was allowed to cool to room temperature and filtered through a pad of
Celite by the aid of EtOAc. The filtrate was treated with water and extracted
with EtOAc (3 ꢀ 100 mL). The organic layer was washed with water and brine,
dried over anhydrous MgSO4, and concentrated under reduced pressure. The
crude oil was purified by column chromatography on silica gel (EtOAc/
withdrawal was expressed as a percentage [(no. of trials accompanied by
brisk foot withdrawal/total no. of trials) ꢀ l00]. Data analysis: The results of
behavioral tests are expressed as a %MPE. For example, paw withdrawal
thresholds were converted to %MPE by the following formula, by using a cutoff
of 15 g (the threshold for normal rats) to define maximum possible effect:
(post
drug
threshold ꢁ baseline
threshold)/(cutoff ꢁ baseline
threshold) ꢀ 100.%MPE values near 100 indicate normal mechanical
thresholds (i.e., at or near 15 g), whereas values near 0 indicate allodynia.
The result of cold allodynia was also expressed as %MPE.