SAR studies of capsazepinoid bronchodilators 3: The thiourea part (coupling region) and the 2-(4-chlorophenyl)ethyl moiety (C-region)

Article abstract of DOI:10.1016/j.bmc.2007.11.056

Certain derivatives and analogues of capsazepine are potent in vitro inhibitors of bronchoconstriction in human small airways. During an investigation of the dependency of the potency on the structural features of the capsazepinoids in the thiourea moiety (coupling region) and the 2-(4-chlorophenyl)ethyl moiety (C-region), it was revealed that capsazepinoids with a thiourea or an amide link between the B-ring and the C-region in general have a good bronchorelaxing activity, while urea is a less attractive choice. Further, it was shown that 1,2,3,4-tetrahydroisoquinolines with a 2-(phenyl)ethyl derivative as the C-region are considerably more potent than those with an octyl group, while 2,3,4,5-tetrahydro-1H-2-benzazepines were found to be more insensitive to the nature of the C-region.

Full text of DOI:10.1016/j.bmc.2007.11.056

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry 16 (2008) 2529–2540
SAR studies of capsazepinoid bronchodilators 3: The thiourea
part (coupling region) and the 2-(4-chlorophenyl)ethyl
moiety (C-region)
a
a,b
Magnus Berglund, Marıa F. Dalence-Guzman, Staffan Skogvallc and Olov Sternera,
*
´
´
aDivision of Organic Chemistry, Lund University, PO Box 124, SE-221 00 Lund, Sweden
bCentro de Tecnologia Agroindustrial, Facultad de Ciencias y Tecnologia, Universidad Mayor de San Simo´n, Cochabamba, Bolivia
cRespiratorius AB, Magistratsva¨gen 10, 226 43 Lund, Sweden
Received 8 December 2006; revised 14 November 2007; accepted 21 November 2007
Available online 28 November 2007
Abstract—Certain derivatives and analogues of capsazepine are potent in vitro inhibitors of bronchoconstriction in human small
airways. During an investigation of the dependency of the potency on the structural features of the capsazepinoids in the thiourea
moiety (coupling region) and the 2-(4-chlorophenyl)ethyl moiety (C-region), it was revealed that capsazepinoids with a thiourea or
an amide link between the B-ring and the C-region in general have a good bronchorelaxing activity, while urea is a less attractive
choice. Further, it was shown that 1,2,3,4-tetrahydroisoquinolines with a 2-(phenyl)ethyl derivative as the C-region are considerably
more potent than those with an octyl group, while 2,3,4,5-tetrahydro-1H-2-benzazepines were found to be more insensitive to the
nature of the C-region.
ꢀ 2007 Elsevier Ltd. All rights reserved.
1. Introduction
kotriene D4, histamine, acetylcholine, prostaglandin D2)
contraction of human small airway preparations.9 This
inhibitory activity is a class effect and analogues of caps-
azepine, capsazepinoids, also are more or less potent
bronchodilators. One of the most potent class members,
the 1,2,3,4-tetrahydroisoquinoline 4, which is approxi-
mately 10 times as potent compared to capsazepine
(2), was identified after a systematic variation of the
structure of capsazepine (2) (Fig. 2).10
Exposure to capsaicin, the major pungent agent isolated
from chili pepper, is known to cause pain and cough,
but also bronchoconstriction in experimental animals.1
The receptor of capsaicin, TRPV1, was suggested by
Szolcsanyi and Jansco-Gaber in 1975 and cloned for
the first time in 1997.2,3 The bronchoconstricting po-
tency of capsaicin (1) and olvanil (165) (Fig. 1), a capsa-
icin analogue with reduced pungency and analgesic
properties,4 has been compared, and it was found that
olvanil (165) was markedly less bronchoconstricting
than capsaicin (1) in guinea pigs.5 Previous studies have
shown that the constricting effect of capsaicin (1) in gui-
nea pig trachea is inhibited by the TRPV1-antagonist
capsazepine (2),6 but that capsazepine does not inhibit
bronchoconstriction evoked in guinea pig by hista-
mine.7,8 Recently, we have described that capsazepine
(2) is a general in vitro inhibitor of agonist evoked (leu-
The SAR study is described in a series of three parallel
papers dealing with the different regions of the molecule,
O
MeO
N
H
HO
1
O
MeO
N
H
Keywords: Capsazepine; Coupling region; Thiourea; C-region; 2-(Ph-
enyl)ethyl; SAR; Bronchodilator; Small human airways; Asthma;
COPD.
HO
165
*
Corresponding author. Tel.: +46 462228213; fax: +46 462228209;
e-mail: Olov.Sterner@organic.lu.se
Figure 1.
0968-0896/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2007.11.056

2530
M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
O
Cl
O
O
S
S
a
OH
OR
b
HO
HO
N
N
N
H
HO
MeO
167 R=Me
168 R=H
166
2
c
e
Cl
Cl
Cl
OH
d
HO
HO
N
H
RO
169 R=Me
170 R=H
Cl
O
4
e
N
H
Cl
Figure 2.
HO
O
172
N
H
Coupling
HO
171
Cl
S
Scheme 1. Reagents and conditions: (a) MeI, K2CO3, DMF, rt,
overnight, 83%; (b) LiOHÆH2O, THF, reflux, overnight, 79%; (c) 1—
Li, NH3, THF, ꢀ33 ꢁC, 1.5 h; 2—NH4Cl 46%; (d) HBr (48% in H2O),
reflux, 5 h quant.; (e) 2-(4-chlorophenyl)ethylamine, HOBt, EDC,
DMF, rt, 45 h.
HO
HO
N
N
H
A
B
C
Figure 3.
as defined in Fig. 3, which were adopted from a previous
study of capsaicin.11–13 The current paper discusses how
structural variations in the coupling region and the C-re-
gion affect the activity. In the preceding papers, the
importance of a catechol moiety in the A-ring and
how structural differences in the B-ring affect the bron-
chodilating ability of the capsazepinoids12 as well as
the effect of chlorination of the A-ring in phenolic and
catecholic derivatives having different B-rings and in
B-rings substituted with methyl/benzyl groups in various
positions13 is discussed. For the coupling region, the fo-
cus of the study was to determine the importance of the
sulfur and nitrogen atoms as well as the effect of confor-
mational changes. Both derivatives having six-mem-
bered (tetrahydroisoquinoline) and seven-membered
(tetrahydro-2-benzazepine) B-rings were included since
the size of this saturated ring will influence the relative
position of the coupling region and the other functional-
ities present in the molecules. In the part studying the ef-
fect of various C-regions, the A-ring, the B-ring and the
coupling region were retained as in the relatively potent
compounds 2 and 4. To facilitate the comparison of the
results from the complete study, a continuous number-
ing of the compounds has been used. Compounds dis-
cussed in more than one paper consequently have the
same number everywhere. The preparation of com-
pounds having numbers below 165 is described in the
preceding papers.12,13
boxylic acid (170), and coupled with 2-(4-chloro-
phenyl)ethylamine to afford 171. Compound 172 was
prepared by the direct coupling of 166 with 2-(4-
chlorophenyl)ethylamine.
The preparation of amides 173, 174 and 175, described
in Scheme 2, was accomplished by the coupling of 4-
(4-chlorophenyl)butanoic acid15 with the amines 30, 93
and 59 (prepared as reported)12,13 using EDC as cou-
pling reagent and HOBt as additive for the reaction.
The synthesis of the urea derivatives 177, 178 and 179
was carried out by the coupling of the amines 30, 59
Cl
R1
R1
O
HO
HO
HO
HO
NH
n
N
a
n
R2
R2
30 R1=H R2=H n=1
93 R1=Cl R2=Cl n=1
59 R1=H R2=H n=2
173 R1=H R2=H n=1
174 R1=Cl R2=Cl n=1
175 R1=H R2=H n=2
H
N
CCl3
b
O
176
Cl
Cl
R1
R2
O
HO
HO
N
N
H
2. Chemistry
n
The synthesis of the amide 171 is shown in Scheme 1.
6-Hydroxy-2-naphthoic acid (166) was methylated with
methyl iodide to afford 167, followed by hydrolysis
of the methyl ester to 168. Partial reduction of 168
yielded the 1,2,3,4-tetrahydronaphthalene derivative
169,14 which was demethylated with hydrobromic acid
to give 6-hydroxy-1,2,3,4-tetrahydronaphthalene-2-car-
177 R1=H R2=H n=1
178 R1=Cl R2=Cl n=1
179 R1=H R2=H n=2
Scheme 2. Reagents and conditions: (a) 4-(4-chlorophenyl)butanoic
acid, HOBt, EDC, N-methyl-morpholine, DMF, rt, 18 h; (b) DBU,
DMSO, 80 ꢁC, 48 h.

M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
2531
and 93 with 2-(4-chlorophenyl)ethyl isocyanate, gener-
ated in situ by b-elimination of chloroform from 176
in the presence of DBU.16
tive lipophilicity of some of the more hydrophilic caps-
azepinoids was estimated (calculated logD7.4 (ACD/
LogD v8.02 using ACD-I-lab service) values and reten-
tion factors determined by HPLC)19 and compared with
that of capsazepine (2) and 4 (see Table 4).
The derivatives 180–202 were prepared as indicated in
Scheme 3, by the coupling of either of the two amines
59 and 93 with different isothiocyanates, commercially
available or synthesized from different arylalkylamines
by the reaction with 1,10-thiocarbonyldiimidazole.17
Among the nonchlorinated 1,2,3,4-tetrahydroiosquino-
line-6,7-diols, the thiourea 34 is slightly more active
compared to the amide 173, while the urea 177 is only
weakly active. However, in the case of the 5,8-di-
chloro-1,2,3,4-tetrahydroiosquinoline-6,7-diols both the
urea 178 and the amide 174 have the same potency as
the thiourea 4. Although unexpected, this could depend
on limitations in the bioassay to differentiate very potent
compounds from each other. Those compounds were
therefore reassayed at lower concentrations (1 lM)
and the results are shown in Table 2 (vide infra).
3. Results and discussion
The bronchorelaxing effect of the synthesized derivatives
was evaluated in small (0.5–1.5 mm in diameter) human
airway preparations, as previously described.9,18 In
short, the constriction evoked by 10 nM leukotriene
D4 on untreated preparations and on the same prepara-
tions exposed to 10 lM of the test compound for 1.5 h
was compared. The results (Tables 1 and 3) are shown
as the percentage of the remaining contraction com-
pared to the full contraction (100% not active and 0%
maximal inhibition). In addition, the effect of the most
potent compounds (4, 174 and 178) was also assayed
at 1 lM (Table 2), in four consecutive cycles. The con-
striction evoked by 10 nM leukotriene D4 on untreated
preparations and on the same preparations exposed to
leukotriene D4-free physiological solution for one hour
followed by 10 nM leukotriene D4 for 30 min, in the
continuous presence of 1 lM of the test compounds,
was compared. The results are shown in Table 2 as the
percentage of the remaining contraction after each cycle,
that takes 1.5 h, compared to the control contraction.
Amides 175 and 173 show somewhat disparate activities
compared to thioureas 2 and 34, although both are sig-
nificantly more active than the corresponding urea
derivatives. The only difference between urea and amide
derivatives, 179 and 177 compared with 175 and 173,
respectively, is the nitrogen linking the coupling and
the C-region. The conformational restrictions induced
by this second nitrogen in the urea derivatives (the nitro-
gen lone pair lobe should preferably be perpendicular to
the plane of the amide moiety) will not be present in the
more flexible propyl chain of the amide derivatives,
thereby possibly allowing 175 and 173 to adopt more
suitable conformations for binding. The difference in
activity between 175 and 173 compared to each other
was expected, as a similar difference was observed for
the corresponding thiourea derivatives.12
The target of capsazepinoids and its localization are un-
known, but established bronchodilatory principles as
well as TRPV1-antagonism have been excluded as
mechanism of action.9,10 As the target and its localiza-
tion are unknown, it is not possible to assess how phys-
icochemical properties, for example, lipophilicity, will
affect the observed potency of the substances tested.
Consequently the SARs discussed here are only based
on the activity of the compounds in the assay. Varia-
tions in concentration or time of exposure might result
in altered relative potencies. Nevertheless, in order to
get an impression of the solubility properties, the rela-
The evaluation of compounds 48, 171 and 172 addresses
the question of the anchorage of the coupling part in the
B-ring. As demonstrated in the study of the B-ring,12
this should preferably be six-membered. As is discussed
above, a thiourea, urea or amide moiety may be used for
the coupling of the B-ring with the C-region, however,
the necessity of a nitrogen atom in the B-ring needs to
be assesed. The 6-hydroxy-1,2,3,4-tetrahydroiosquino-
line 48 is as potent as capsazepine (2), as the 6-hydroxyl
group appears to be critical for the interaction of tetra-
hydroiosquinolines and tetrahydro-2-benzazepines with
the target.12 However, exchanging the B-ring nitrogen
for a carbon (to compound 171) diminishes the activity,
an effect that cannot be explained by the change of the
thiocarbonyl in 48 for a carbonyl group in 171. The
drop in activity seen for 171 could, however, be due to
other factors: (a) The compound was tested as a racemic
mixture. (b) The rotational barrier of the bond formed
between the nitrogen in the B-ring and the thiocarbonyl
carbon in 48 is bigger than the corresponding bond in
170. (c) The nitrogen atom might be involved in a spe-
cific interaction with the target, exchanging it for a car-
bon will interupt such an interaction. Full planarity of
the B-ring does not enhance activity as the naphthamide
172 is even less active compared to 48 and 170. Although
a constrained B-ring is important for activity,12 the re-
sult obtained with 172 suggests that the angle induced
by the limited flexibility of saturated B-ring between
NH2
R
Cl
Cl
Cl
S
a
HO
HO
HO
HO
R
NH
NH
N
N
H
NCS
R
Cl
93
180—193
b
S
R
N
HO
HO
HO
HO
N
H
59
194—202
Scheme 3. Reagents and conditions: (a) 1,10-thiocarbonyldiimidazole,
triethylamine, DMF, 50 ꢁC, 2 h; (b) triethylamine, DMF, rt.

2532
M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
Table 1.
Cl
Coupling
Type
A-B-ring
Cl
HO
N
HO
HO
HO
HO
N
N
N
HO
HO
HO
HO
A
Cl
B
C
D
F
E
O
S
O
N
H
N
H
I
II
III
Compound
A-B-ring
Coupling type
Remaining contractiona
34b
177
173
A
A
A
I
36 6.5 (5)
69 3.7 (4)
47 6.1 (4)
II
III
4c
178
174
B
B
B
I
14 2.7 (16)
16 1.7 (5)
16 4.3 (4)
II
III
2b
179
175
C
C
C
I
55 3.0 (24)
73 6.6 (3)
54 8.9 (6)
II
III
48b
171
172
D
E
F
I
52 6.8 (4)
78 1.9 (4)
90 2.6 (4)
II
II
a Arithmetic means standard error of the mean of the percentage of remaining contraction in small human airways contracted with LTD4 in the
presence of 10 lM test substance compared to a full contraction evoked by LTD4 in the absence of test compound. Number of times tested in
parentheses.
b Compound reported in Ref. 12.
c Compound reported in Ref. 13.
Table 2. Remaining contraction after treatment with 1 lM of 4, 178
and 174
pounds were therefore reassayed at a lower concentra-
tion (1 lM) for four test cycles, and the results are
shown in Table 2. At this concentration the activity cor-
relates with the results discussed above with 34, 173 and
177, especially after 6 h, showing that the thiourea in
fact is slightly more potent than the amide, while the
urea is significantly less bronchorelaxing.
Compound
Remaining contraction SEMa
3 h 4.5 h 6 h
79 6.8 57 11.0 44 10.2 32 7.2
Nb
1.5 h
4
178
174
5
4
5
91 1.8 81 3.9
80 4.4 65 3.6
78 4.2
55 3.7
71 3.9
45 5.7
See text for details.
a Standard error of the mean.
b Number of experiments performed.
If the substitution of the aromatic A-ring is as that of
4, 174 and 178, the binding of the molecule to the
receptor seems to be optimal and activity increases dra-
matically as seen in studies concerning the A- and B-
rings.12,13 In this case, changes in the coupling region
do not have that dramatical affect on the activity. On
the other hand, when the substitution pattern in the
A-ring is as in 2 or 34, differences in the coupling re-
gion enhance or decrease the activity significantly. Sul-
fur and oxygen atoms in the coupling region are likely
to act as hydrogen bond acceptors or in dipole interac-
tions. In carbonyl groups the oxygen lone pair lobes
are in the R2C@O plane and form angles of about
120ꢁ with the C@O bond. Hydrogen bonding to a car-
bonyl oxygen is at its strongest at this angle, although
the aromatic A-ring and the plane of the coupling region
is critical for activity. A similar hypothesis was sug-
gested by Walpole et al. in their study of constrained
capsaicin derivatives.20
In the standard assay used throughout this investiga-
tion, the 5,8-dichloro-1,2,3,4-tetrahydroiosquinoline-
6,7-diols 4, 178 and 174, differing only in the coupling
type, are equipotent. As all three are very potent and re-
lax the preparations almost completely, this result might
be due to a limitation in the bioassay. All three com-

M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
2533
it also takes place at any other angle between 115ꢁ and
180ꢁ. On the other hand, the acceptor directionality of
the thiocarbonyl group is much more pronounced, with
the lone pair directions forming an angle of only 105ꢁ
with R2C@S.21 Angles between 130ꢁ and 180ꢁ will not
be suitable for hydrogen bonding to a thiocarbonyl.
For 2 and 34, the sulfur atom seems to be placed in
the right position for binding, but once the sulfur atom
is changed to oxygen both the directionality and the
size of the acceptor will change, offering a possible
explanation for the decrease in activity of the urea
derivatives. However, the amides (173, 174 and 175)
are almost as active as the thioureas (2, 4 and 34),
and also significantly more active than the correspond-
ing urea derivatives. The only difference between urea
and amide derivatives (177, 178 and 179) is the nitro-
gen linking the coupling and the C-region. As discussed
above, the conformational restrictions induced by this
second nitrogen in the urea derivatives (the nitrogen
lone pair lobe should preferably be perpendicular to
the plane of the amide moiety) will not be present in
the amide derivatives.
the C-region if they are based on A-B–ring type B or
C (see Tables 1 and 3). For those based on C, that is,
capsazepine (2) analogues, 2 itself with a 2-(4-chloro-
phenyl)ethyl moeity is actually the most potent of all
compounds of this type tested. Only the 2-(2-chloro-
phenyl)ethyl variant (197) has a similar potency. The
octyl derivative 202 is as active as several of the 2-
phenylethyl derivatives. This indicates strongly that the
C-region is less important for the binding of capsazepine
(2) and its closely related analogues, presumably because
the C-region is not positioned in a way to optimize the
interactions with the binding site. As discussed,12 the
B-ring will affect how the A-ring, coupling and C-region
are orientated relative to each other and the B-ring in 4
was shown to be better in this sense.12 In contrast, the
1,2,3,4-tetrahydroisoquinoline derivatives, based on
structure B in Table 3, show a larger dependency on
the C-region. Here it is obvious that an alkyl substituent
(compound 190) is poor compared to the phenylethyl
analogues, even if the size is comparable. This indicates
that some kind of aromatic p-interaction between the
target and the C-region is facilitated for this type of
structure. The length of the link between the thiourea
and the aromatic C-ring (n in Table 3) does not appear
to be critical, as the phenylmethylene 180, the 2-phenyl-
Interestingly, the bronchodilating activity of the caps-
azepine derivatives depends differently on variations in
Table 3.
S
Cl
Cl
S
R
HO
HO
(CH2)n
N
N
H
HO
HO
R
N
N
H
R1
N
N
N
C
I
II
III
B
IV
Compound
A-B-ring
R
n
R1
Remaining contractiona
180
181
194
182
195
183
196
184
197
185
198
4b
B
B
C
B
C
B
C
B
C
B
C
B
C
B
C
B
C
B
B
C
B
C
B
B
B
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
1
H
10 2.8 (5)
19 2.3 (4)
70 8.2 (4)
1
4-Cl
4-Cl
H
1
2
8
4.3 (4)
2
H
71 9.5 (4)
69 7.0 (4)
80 2.5 (4)
10 3.3 (5)
56 8.3 (4)
27 6.4 (4)
76 3.5 (4)
14 2.7 (16)
55 3.0 (24)
2
4-t-Bu
4-t-Bu
2-Cl
2-Cl
3-Cl
3-Cl
4-Cl
4-Cl
4-F
4-F
4-Br
4-Br
4-OH
H
2
2
2
2
2
2
2c
2
186
199
187
200
188
189
201
190
202
191
192
193
2
6
1.4 (4)
2
68 5.4 (4)
17 4.6 (4)
67 3.7 (4)
22 5.3 (4)
11 3.6 (4)
84 2.9 (4)
69 1.6 (4)
69 8.0 (5)
33 5.1 (4)
30 4.5 (4)
26 6.2 (7)
2
2
2
3
3
H
—
n-C8H17
n-C8H17
II
—
—
—
—
—
—
—
III
IV
—
—
a Arithmetic mean the standard error of the mean of the percentage of remaining contraction in small human airways contracted with LTD4 in the
presence of 10 lM test substance compared to a full contraction evoked by LTD4 in the absence of test compound. Number of times tested in
parentheses.
b Compound reported in Ref. 13.
c Compound reported in Ref. 12.

2534
M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
ethylene 182 and the 3-phenylpropylene 189 are approx-
imately equally active, and also comparable with the
2-(4-chlorophenyl)ethylene derivative 4. In addition,
the shorter (4-chlorophenyl)methylene derivative 181
is approximately as active as 4. The different potency
of the three 2-(chlorophenyl)ethylene derivatives 184,
185 and 4 suggests that the volume of the part of the
binding site where the C-region interacts is limited
for a substituent in the meta position, and the same
trend is also seen for the corresponding benzazepines
197, 198 and 2. Steric interactions that hamper binding
are definitely indicated in the case of the 2-(4-tert-
butylphenyl)ethyl derivatives 183 and 196, suggesting
that the binding pocket has a limited size also in the
para direction. In addition it should be noted that
the 4-fluoro derivative 186 actually is better than the
4-chloro, while the 4-bromo (187) is comparable. This
may also be due to steric interactions, although elec-
tronic effects or a combination of both may not be
excluded.
in activity between 4 and 191/192/193/188 is caused by
poorer binding or by other factors. Still, it is interesting
to note that the properties of these compounds can be
modulated in the C-region while retaining the bronchod-
ilating activity. Similarly the amide 174, although being
almost as potent as the thiourea 4, is more hydrophilic,
as judged by the experimentally determined retention
factor.
4. Conclusions
Capsazepinoids, a novel class of bronchodilators, have
been shown to be potent in vitro inhibitors of broncho-
constriction in human small airways. If similar effects
are at hand also in vitro, they constitute a promising no-
vel pharmaceutical principle to prevent and revoke air-
way obstruction caused by asthma or COPD. The
capsazepinoids prepared, assayed and discussed in part
of the study focus on the coupling region and the C-re-
gion (according to Fig. 3), and it is obvious that their
activities depend on their structures.
The dependency of the bronchodilating effect of the
capsazepine derivatives (Table 3) on the C-region dif-
fers compared to the effect of variations of the phen-
ylethyl moiety for the TRPV1 antagonist activity.
Recent studies in 45Ca2+-influx models to assay
TRPV1-antagonism have reported that a tert-butyl
group is the optimum para-substituent in the region
corresponding to the C-region.22 This is in accordance
with the low bronchodilating activity of established
TRPV1-antagonists, for example, iodo-resiniferatoxin,
ruthenium red, JYL1421 and SB366791, previously re-
ported by us,10 and supports the suggestion that the
bronchodilating effect observed in our assay is not
mediated by TRPV1.
The following conclusions could be drawn concerning
the coupling region:
1. The amide derivatives 173, 174 and 175 show similar
bronchorelaxing properties as the corresponding
thioureas, suggesting that the nitrogen atom linking
the coupling unit and the C-region is not of great
importance for the activity. The inverse amides
with no nitrogen in the B-ring (171 and 172) are
considerably less active, especially if the B-ring is
aromatic.
2. The urea derivatives 177, 176 and 178 are less potent
compared to the corresponding thioureas.
Since most of the compounds discussed here are rather
lipophilic and would probably suffer from extensive pro-
tein binding and poor solubility in plasma in vitro,23 the
more hydrophilic pyridine derivatives 191, 192 and 193,
and the 4-hydroxyl derivative 188, were therefore pre-
pared. They all have a similar potency, slightly lower
that of 4, and as can be seen in Table 4, the lipophilicity
of 191, 192, 193 and 188 is considerably lower compared
to compounds 2 and 4. It is at this stage not possible to
draw any conclusions about whether the small difference
Additional conclusions could be drawn concerning the
C-region:
1. The importance of the C-region for the binding
appears to be different for capsazepine and 1,2,3,4-
tetrahydroisoquinoline derivatives, suggesting that
the two compound types present the C-region to the
binding site differently.
2. An aromatic p-interaction between the target and the
C-region is indicated, at least for the 1,2,3,4-tetrahy-
droisoquinoline derivatives.
3. The volume in the binding site where the C-region of
capsazepinoids binds is limited and bigger substitu-
ents in the C-region lower the activity. This also
restricts how long the linker between the thiourea
and the aromatic C-ring can be.
4. Several different C-regions, for example, 2-(2-chloro-
phenyl)ethyl, 2-(4-chlorophenyl)ethyl, 2-(4-fluoro-
phenyl)ethyl, and 2-(phenyl)ethyl are all approximately
equally potent, suggesting that the interaction
between the binding site and the C-region in general
is weak.
Table 4. The lipophilicity of selected capsazepinoids
a
Compound
LogD7:4
Retention factorb
2
4
3.5
4.0
2.7
1.9
2.0
1.9
4.6
8.9
8.6
1.8
0.9
0.9
1.1
5.2
188
191
192
193
174
See text for details.
a The result of ACD/LogD v8.02 was obtained using the ACD/I-lab
service and is given at pH 7.4.
b HPLC, symmetry shield RP8 2.1 · 150 mm, 0.3 ml/min 40% aceto-
nitrile in 20 mM NaH2PO4, pH 7.4.
5. The finding that the 2-(4-tert-butylphenyl)ethyl deriv-
atives are impotent support the suggestion that the
bronchodilating effect of the capsazepinoids is not
mediated by TRPV1.

M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
2535
Work is now in progress to investigate the in vitro activ-
ity of the capsazepinoids as bronchodilators. In this re-
spect, the more hydrophilic derivatives described here,
for example, 174 and 193, might prove useful.
5.2.4. 6-Hydroxy-1,2,3,4-tetrahydronaphthalene-2-car-
boxylic acid (170). Compound 169 was dissolved in
HBr (48% in H2O). The mixture was heated to 105 ꢁC
for 5 h and then concentrated. The residue was sus-
pended in EtOAc and concentrated again affording
1
170 (quantitative). H NMR (CDCl3 300 MHz) d 1.86
5. Experimental
5.1. Materials
(m, 1H), 2.22 (m, 1H), 2.76 (m, 1H), 2.84 (m, 2H),
2.96 (m, 2H), 6.57 (d, J = 2.8 Hz, 1H), 6.63 (dd,
J = 8.2 Hz, J = 2.8 Hz, 1H), 6.54 (d, J = 8.2 Hz, 1H).
Materials were obtained from commercial suppliers and
were used without further purification unless otherwise
noted. DMF were distilled under reduced pressure. All
moisture- and air-sensitive reactions were carried out
under an atmosphere of dry nitrogen using oven-dried
glassware. HRMS (ESI) spectra were recorded with a
Micromass Q-TOF Micro spectrometer. NMR spectra
(in CDCl3 or CD3OD) were recorded with a Bruker
ARX 300 spectrometer at 300 MHz (1H) and at 75
MHz (13C), and with a Bruker DRX 400 spectrometer
at 400 MHz (1H) and at 100 MHz (13C). Chemical shifts
are given in ppm relative to TMS using the residual
CHCl3 peak in CDCl3 or the residual CD2HOD peak
in CD3OD solution as internal standard (7.26 or 3.32
and 77.2 or 49.0 ppm, respectively, relative to TMS).
5.2.5. N-[2-(4-Chlorophenyl)ethyl]-6-hydroxy-1,2,3,4-tet-
rahydronaphthalene-2-carboxamide (171). EDC (1.1 equiv)
and HOBt (1.1 equiv) were dissolved in anhydrous DMF.
Compound 170 (1.0 equiv) and 2-(4-chlorophenyl)ethyla-
mine (1.0 equiv) were added to this solution. The mixture
was stirred for 45 h. Purification was done by flash col-
umn chromatography (silica, Pet. ether/EtOAc/AcOH
33:77:1) affording
7
(65%). 1H NMR (CD3OD,
400 MHz) d 1.75 (m, 1H), 1.92 (m, 1H), 2.47 (m,
1H), 2.75 (m, 4H), 2.81 (t, J = 7.2 Hz, 2H), 3.44 (t,
J = 7.2 Hz, 2H), 6.50 (d, J = 2.3 Hz, 1H), 6.54 (dd,
J = 8.2 Hz, J = 2.3 Hz, 1H), 6.86 (d, J = 8.2 Hz, 1H),
7.21 (d,J = 8.4 Hz, 2H), 7.29 (d,J = 8.4 Hz, 2H). 13C
NMR (CD3OD, 100 MHz) d 27.8, 29.4, 32.7, 35.8, 41.6,
43.3, 114.3, 115.8, 127.1, 129.5, 129.5, 130.8, 131.5, 131.5,
133.2, 137.8, 139.4, 156.3, 178.6. ESI-MS calculated for
C19H21NO2Cl (M+H) 330.1261, found 330.1247.
˚
All flash chromatography was performed on 60 A 35–
70 lm Matrex silica gel (Grace Amicon). TLC analyses
were made on Silica Gel 60 F254 (Merck) plates and visu-
alized with ninhydrin/acetic acid and heating. The purity
5.2.6. N-[2-(4-Chlorophenyl)ethyl]-6-hydroxy-2-naphtha-
mide (172). Compound 171 was prepared from 166,
using the method described for 170. Purification was
done by flash column chromatography (silica, Pet.
1
of the assayed compounds was verified with H NMR
and HPLC, and only used if more than 98% pure.
1
5.2. Synthesis
ether/EtOAc/AcOH 33:77:1) (38%). H NMR (DMSO-
d6, 400 MHz) d 2.87 (t, J = 7.1 Hz, 2H), 3.51 (dd,
J = 5.7 Hz, J = 7.1 Hz, 2H), 7.14 (m, 2H), 7.29 (d,
J = 8.4 Hz, 2H), 7.35 (d, J = 8.4 Hz, 2H), 7.72 (d,
J = 8.6 Hz, 1H), 7.79 (dd, J = 1.7 Hz, J = 8.6 Hz, 1H),
7.85 (d, J = 8.6 Hz, 1H), 8.27 (bs, 1H), 8.58 (t,
J = 5.7 Hz, 1H).13C NMR (DMSO-d6, 100 MHz) d
34.5, 40.7, 108.7, 119.5, 124.4, 126.0, 126.7, 127.4,
128.3, 128.3, 128.7, 130.6, 130.7, 130.7, 130.8, 136.1,
138.7, 156.9, 166.5. ESI-MS calculated for
C19H17NO2Cl (M+H) 326.0948, found 326.0943.
5.2.1. Methyl 6-methoxy-2-naphthoate (167). To a solu-
tion of 6-hydroxy-2-naphthoic acid, 166 (1.0 equiv) and
K2CO3 (2.5 equiv) in anhydrous DMF, MeI (2.2 equiv)
was added. The mixture was stirred overnight, concen-
trated and dissolved in EtOAc. The organic phase was
washed with Na2CO3 (saturated solution), dried (MgSO4)
and concentrated. Recrystallization (heptane/EtOAc 1:1)
of the crude gave 167 (83%). 1H NMR (CDCl3 300 MHz)
d 3.95 (s, 3H), 3.97 (s, 3H), 7.16 (d, J = 2.5 Hz, 1H), 7.20
(dd, J = 8.9 Hz, J = 2.5 Hz, 1H), 7.76 (d, J = 8.7 Hz, 1H),
7.84 (d, J = 8.9 Hz, 1H), 8.03 (dd, J = 8.7 Hz, J = 1.8 Hz,
1H), 8.54 (d, J = 1.8 Hz, 1H).
5.2.7. 2-[4-(4-Chlorophenyl)butanoyl]-1,2,3,4-tetrahydro-
isoquinoline-6,7-diol (173). 4-(4-chlorophenyl)butanoic
acid (1.0 equiv) and 1,2,3,4-tetrahydroisoquinoline-
6,7-diol hydrobromide, 30 (1.0 equiv) were dissolved in
anhydrous DMF, then 1-hydroxy-benzotriazole, HOBt
(1.0 equiv), N0-(3-dimethyl-aminopropyl)-N-ethylcarbo-
diimide hydrochloride, EDC (1.05 equiv) and N-
methyl-morpholine (3.0 equiv) were added. The
reaction mixture was stirred at room temperature for
18 h and then it was concentrated. Purification was done
by flash column chromatography (silica, gradient
elution: 60–100% EtOAc in Pet. ether) affording 173
(rotameric mixture), (48%). 1H NMR (CD3OD 400
MHz) d 1.92 (m, 2H), 2.45 (t, J = 7.1 Hz, 2H), 2.67
(m, 4H), 2.62 (ma) (t, J = 6.0 Hz, 2H), 3.71 (mi) (t,
J = 6.0 Hz, 2H), 4.45 (mi) (s, 2H), 4.50 (ma) (s, 2H),
6.55 (m, 2H), 7.20 (m, 4H). 13C NMR (CD3OD 100
MHz) d 28.0, 28.7 (mi), 29.6 (ma), 33.5, 35.5, 41.7
(mi), 45.0 (ma), 45.0(ma), 48.0 (mi), 113.7 (mi), 113.9
5.2.2. 6-Methoxy-2-naphthoic acid (168). Compound 167
(1.0 equiv) and LiOHÆH2O (10.0 equiv) were suspended
in THF. The slurry was heated to reflux overnight.
The reaction mixture was concentrated and HCl (1%
in H2O) was added. The water phase was extracted with
EtOAc. The combined organic phases were dried
(MgSO4) and concentrated. Recrystallization from
EtOAc afforded 168 (79%). 1H NMR (CDCl3 300
MHz) d 3.96 (s, 3H), 7.17 (d, J = 2.5 Hz, 1H), 7.22
(dd, J = 8.9 Hz, J = 2.5 Hz, 1H), 7.80 (d, J = 8.6 Hz,
1H), 7.88 (d, J = 8.9 Hz, 1H), 8.09 (dd, J = 8.6 Hz,
J = 1.6 Hz, 1H), 8.54 (bs, 1H).
5.2.3. 6-Methoxy-1,2,3,4-tetrahydronaphthalene-2-car-
boxylic acid (169). Compound 169 was prepared from
168. Spectroscopic data as described previously.14

2536
M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
(ma), 115.9 (mi), 116.1 (ma), 124.9, 126.3, 129.4, 129.4,
129.5, 131.1, 131.1, 132.3, 141.9, 145.0, 174.0. ESI-MS
calculated for C19H20ClN2O3 (M+H) 347.1162, found
347.1148.
5.2.11. 5,8-Dichloro-N-[2-(4-chlorophenyl)ethyl]-6,7-dihy-
droxy-3,4-dihydroisoquinoline-2(1H)-carboxamide (178).
Compound 178 was prepared as described for 177
from 5,8-dichloro-1,2,3,4-tetrahydroisoquinoline-6,7-
diol hydrobromide 93. Purification was done by flash
5.2.8. 5,8-Dichloro-2-[4-(4-chlorophenyl)butanoyl]-1,2,3,
4-tetrahydroisoquinoline-6,7-diol (174). Compound 174
was prepared as described for 173 from 5,8-dichloro-
1,2,3,4-tetrahydroisoquinoline-6,7-diol hydrobromide
93. Purification was done by flash column chromatog-
raphy (Pet. ether/EtOAc 1:1) affording 174 (rotameric
column chromatography (silica, gradient elution: 0.5-
2% MeOH in CH2Cl2) (38%). H NMR (CD3OD 400
1
MHz) d 2.68 (t, J = 5.8 Hz, 2H), 2.77 (t, J = 7.3 Hz,
2H), 3.38 (t, J = 7.3 Hz, 2H), 3.55 (t, J = 5.8 Hz, 2H),
4.43 (s, 2H), 7.13 (d, J = 8.3 Hz, 2H), 7.20 (d,
J = 8.3 Hz, 2H). 13C NMR (CD3OD 100 MHz) d 27.1,
36.7, 42.0, 43.2, 45.3, 118.5, 120.5, 124.6, 125.7, 129.4,
129.4, 131.5, 131.5, 133.0, 139.7, 142.5, 142.8, 159.8.
ESI-MS calculated for C18H18N2O3Cl3 (M+H)
415.0383, found 415.0395.
1
mixture) (21%). H NMR (CD3OD 400 MHz) d 1.93
(m, 2H), 2.48 (m, 2H), 2.66 (m, 2H), 2.73 (mi) (t,
J = 6.0 Hz, 2H), 2.79 (ma) (t, J = 6.0 Hz, 2H), 3.69
(ma) (t, J = 6.0 Hz, 2H), 3.78 (mi) (t, J = 6.0 Hz,
2H), 4.51 (mi) (s, 2H), 4.60 (ma) (s, 2H), 7.19 (m,
4H). 13C NMR (CD3OD 100 MHz) d 27.0 (mi),
27.9, 28.1 (ma), 33.1 (ma), 33.6 (mi), 35.4 (mi), 35.5
(ma), 40.4 (mi), 43.6 (ma), 43.9 (ma), 46.9 (mi),
118.7, 124.01, 125.17, 129.4, 129.4, 131.0, 131.1,
131.1, 132.8, 141.6, 141.9, 142.8, 173.9. ESI-MS calcu-
lated for C19H19Cl3NO3 (M+H) 414.0431, found
414.0417.
5.2.12. N-[2-(4-Chlorophenyl)ethyl]-7,8-dihydroxy-1,3,4,5-
tetrahydro-2H-2-benzazepine-2-carboxamide (178). Com-
pound 179 was prepared as described for 177 from
2,3,4,5-tetrahydro-1H-3-benzazepine-7,8-diol
hydro-
bromide 59. Purification was done by flash column
chromatography (silica, CH2Cl2/MeOH 98:2) (29%).
1H NMR (CD3OD 400 MHz) d 1.46 (m, 2H), 2.50
(t, J = 7.3 Hz, 2H), 2.60 (m, 2H), 3.12 (t, J = 7.3 Hz,
2H), 3.40 (m, 2H), 4.11 (s, 2H), 6.43 (s, 1H), 6.54
(s, 1H), 6.83 (d, J = 8.4 Hz, 2H), 6.99 (d, J = 8.4 Hz,
2H) 13C NMR (CD3OD, 100 MHz) d 24.4, 34.3,
35.6, 41.9, 49.9, 51.2, 116.8, 117.1, 128.2, 128.2,
128.4, 130.3, 130.3, 131.7, 133.3, 138.5, 142.5, 143.8,
158.3 ESI-MS calculated for C19H21ClN2O3 (M+H)
360.1241, found 360.1241.
5.2.9. 2-[4-(4-Chlorophenyl)butanoyl]-2,3,4,5-tetrahydro-
1H-2-benzazepine-7,8-diol (175). Compound 175 was
prepared as described for 173 from 2,3,4,5-tetrahydro-
1H-3-benzazepine-7,8-diol hydrobromide 59. Purifica-
tion was done by flash column chromatography (silica,
CH2Cl2/MeOH 98:2) (19%). 1H NMR (CDCl3400
MHz)
d 1.74 (m, 2H), 1.91 (m, 2H), 2.31 (t,
J = 7.4 Hz, 2H), 2.59 (t, J = 7.4 Hz, 2H), 2.90 (m, 2H),
3.69 (bs, 2H), 4.48 (s, 2H), 6.71 (s, 1H), 7.03 (d,
J = 8.3 Hz, 2H), 7.17 (s, 1H), 7.20 (d, J = 8.3 Hz, 2H).
13C NMR (CDCl3 100 MHz) d 26.3, 29.6, 32.2, 34.4,
34.5, 51.0, 52.5, 116.0, 117.0, 128.4, 128.4, 129.1,
129.7, 129.7, 132.5, 132.8, 139.8, 142.0, 143.6, 172.5.
ESI-MS calculated for C20H23ClN2O3 (M+H)
360.1366, found 360.1375.
The isothiocyanates were purchased from commercial
suppliers (1-isothiocyanatooctane and 1-chloro-4-(isoth-
iocyanatomethyl)-benzene) or synthesized from com-
mercially available amines.
Isothiocyanate synthesis, general method: 1,10-thi-
ocarbonyldiimidazole (1.2 equiv) was dissolved in
DMF at 50 ꢁC. To this solution a solution of the
amine (1 equiv) and triethylamine (1 equiv) in DMF
was added dropwise. When the amine was available
as a protonated salt, 3 equivalent of triethylamine
was used. The mixture was stirred at room tempera-
ture for 2 h, diluted with water and extracted with
EtOAc. The combined organic phases were washed
with water, dried (MgSO4) and concentrated. The res-
idue was chromatographed on silica gel (hep-
tane:EtOAc). All isothiocyanates were obtained as
pale yellow liquids.
5.2.10. N-[2-(4-Chlorophenyl)ethyl]-6,7-dihydroxy-3,4-
dihydroisoquinoline-2(1H)-carboxamide (177). 1,2,3,4-
tetrahydroisoquinoline-6,7-diol hydrobromide, 30 (1.0
equiv) was dissolved in anhydrous DMSO, DBU
(1.0 equiv) was added and the solution was stirred for
15 min. Then 176 (1.0 equiv) and DBU (1.0 equiv) were
added. The intermediate 2-(4-chlorophenyl)ethyl isocya-
nate is formed in situ. The reaction mixture was stirred
at 80 ꢁC for 48 h. CH2Cl2 was added to the solution and
the organic phase was washed with HCl (3% in H2O)
and NaHCO3 (saturated solution). The organic phase
was dried (MgSO4) and concentrated. Purification was
done by flash column chromatography (silica, CH2Cl2/
MeOH 98:2) yielding 177 (23%). 1H NMR (CD3OD
1
5.2.13. Isothiocyanatomethylbenzene. H NMR (CDCl3
300 MHz) d 4.70 (s, 2H), 7.33 (m, 5H).
400 MHz)
d
2.64 (t, J = 5.9 Hz, 2H), 2.77 (t,
5.2.14. 1-tert-Butyl-4-(2-isothiocyanatoethyl)-benzene. 1H
NMR (CDCl3 300 MHz) d 1.32 (s, 9H), 2.97 (t,
J = 7.0 Hz, 2H), 3.72 (t, J = 7.0 Hz, 2H), 7.15 (d,
J = 8.2, 2H), 7.37 (d, J = 8.2, 2H).
J = 7.3 Hz, 2H), 3.37 (t, J = 7.3 Hz, 2H), 3.51 (t,
J = 5.9 Hz, 2H), 4.33 (s, 2H), 6.53 (s, 1H), 6.56 (s,
1H), 7.15 (d, J = 8.4 Hz, 2H), 7.22 (d, J = 8.4 Hz, 2H).
13C NMR (CD3OD 100 MHz) d 29.0, 36.8, 42.7, 43.3,
46.2, 113.8, 116.0, 125.6, 127.1, 129.4, 129.4, 131.5,
131.5, 132.9, 139.7, 145.0, 145.2, 160.0. ESI-MS calcu-
lated for C18H20N2O3Cl (M+H) 347.1162, found
347.1148.
5.2.15. 1-Chloro-2-(2-isothiocyanatoethyl)-benzene. 1H
NMR (CDCl3 300 MHz) d 2.96 (t, J = 6.8 Hz, 2H),
3.73 (t, J = 6.8 Hz, 2H), 7.14 (m, 1H), 7.22 (m, 1H),
7.27 (m, 2H).

M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
2537
5.2.16. 1-Chloro-3-(2-isothiocyanatoethyl)-benzene. 1H
NMR (CDCl3 300 MHz) d 3.13 (t, J = 6.8 Hz, 2H),
3.78 (t, J = 6.8 Hz, 2H), 7.26 (m, 3H), 7.39 (m, 1H).
5.2.25. 5,8-Dichloro-N-(4-chlorobenzyl)-6,7-dihydroxy-3,4-
dihydroisoquinoline-2(1H)-carbothioamide (181). Yield:
65%. 1H NMR (CD3OD 400 MHz) d 2.82 (t,
J = 5.8 Hz, 2H), 4.01 (t, J = 5.8 Hz, 2H), 4.88 (s, 2H),
4.91 (s, 2H), 7.25 (d, J = 8.6 Hz, 2H), 7.3 (d,
J = 8.6 Hz, 2H). 13C NMR (CD3OD 100 MHz) d 27.3,
46.1, 49.6, 49.8, 118.4, 120.2, 124.2, 125.9, 129.3,
129.3, 130.1, 130.1, 133.5, 139.5, 142.6, 142.9, 183.6.
HRMS (ESI) calculated for C17H16Cl3N2O2S (M+H)
416.9998, found 417.0004.
5.2.17. 1-Fluoro-4-(2-isothiocyanatoethyl)-benzene. 1H
NMR (CDCl3 300 MHz) d 2.95 (t, J = 6.8 Hz, 2H),
3.70 (t, J = 6.8 Hz, 2H), 7.02 (dd, J = 8.7, JF = 8.7 Hz,
2H), 7.18 (dd, J = 8.7, JF=5.4 Hz, 2H).
5.2.18. 1-Bromo-4-(2-isothiocyanatoethyl)-benzene. 1H
NMR (CDCl3 300 MHz) d 2.94 (t, J = 6.8 Hz, 2H),
3.72 (t, J = 6.8 Hz, 2H), 7.10 (d, J = 8.3 Hz, 2H), 7.47
(d, J = 8.3 Hz, 2H).
5.2.26. 5,8-Dichloro-6,7-dihydroxy-N-(2-phenylethyl)-3,4-
dihydroisoquinoline-2(1H)-carbothioamide (182). Yield:
66%. 1H NMR (CD3OD 400 MHz) d 2.84 (t,
J = 5.9 Hz, 2H), 2.93 (t, J = 7.4 Hz, 2H), 3.83 (t,
J = 7.4 Hz, 2H), 3.94 (t, J = 5.9 Hz, 2H), 4.84 (s, 2H),
7.20 (m, 5H), 13C NMR (CD3OD 100 MHz) d 27.1,
36.3, 45.3, 48.3, 49.3, 118.4, 120.2, 124.2, 125.9, 127.2,
129.4, 129.4, 129.9, 129.9, 140.7, 142.6, 142.9, 182.4.
HRMS (ESI) calculated for C18Cl2H19N2O2S (M+H)
397.0544, found 397.0532.
5.2.19. 4-(2-Isothiocyanatoethyl)-phenol. 1H NMR
(CDCl3 300 MHz) d 2.91 (t, J = 6.9 Hz, 2H), 3.67 (t,
J = 6.9 Hz, 2H), 5.33 (s, 1H), 6.81 (d, J = 8.5 Hz, 2H),
7.08 (d, J = 8.5 Hz, 2H).
5.2.20. (3-Isothiocyanatopropyl)-benzene. 1H NMR
(CDCl3 300 MHz) d 2.02 (m, 2H), 2.76 (t, J = 7.5 Hz,
2H), 3.50 (t, J = 6.5 Hz, 2H), 7.21 (m, 3H), 7.31 (m, 2H).
5.2.27. N-[2-(4-tert-Butylphenyl)ethyl]-5,8-dichloro-6,7-
dihydroxy-3,4-dihydroisoquinoline-2(1H)-carbothioamide
(183). Yield: 25%. 1H NMR (CD3OD 400 MHz) d 1.27,
(s, 9H), 2.77 (t, J = 5.8 Hz, 2H), 2.90 (t, J = 7.4 Hz, 2H),
3.82 (t, J = 7.4 Hz, 2H), 3.96 (t, J = 5.8 Hz, 2H), 4.84 (s,
2H), 7.12 (d, J = 8.4, 2H), 7.27 (d, J = 8.4 Hz, 2H). 13C
NMR (CD3OD 100 MHz) d 27.1, 31.8, 31,8, 31.8, 35.2,
35.7, 45.9, 48.3, 49.3, 118.4, 120.3, 124.3, 125.9, 126.3,
126.3, 129.7, 129.7, 137.7, 142.7, 142.9, 150.1, 182.5.
HRMS (ESI) calculated for C22H27Cl2N2O2S (M+H)
453.1170, found 453.1161.
5.2.21. 4-(2-Isothiocyanatoethyl)-pyridine. 1H NMR
(CDCl3 300 MHz) d 2.95 (t, J = 6.7 Hz, 2H), 3.75 (t,
J = 6.7 Hz, 2H), 7.13 (t, J = 5.9 Hz, 2H), 8.54 (t,
J = 5.9 Hz, 2H).
5.2.22. 3-(2-Isothiocyanatoethyl)-pyridine. 1H NMR
(CDCl3 300 MHz) d 3.00 (t, J = 6.7 Hz, 2H), 3.92 (t,
J = 6.7 Hz, 2H), 7.30 (dd, J = 7.7, 4.8 Hz, 1H), 7.58
(dt, J = 7.7, 1.7 Hz 1H), 8.52 (d, J = 1.7 Hz, 1H), 8.55
(dd, J = 4.8, 1.7 Hz, 1H).
5.2.23. 2-(2-Isothiocyanatoethyl)-pyridine. 1H NMR
(CDCl3 300 MHz) d 3.31 (t, J = 6.7 Hz, 2H), 3.78 (t,
J = 6.7 Hz, 2H), 7.18 (m, 2H), 7.62 (td, J = 7.7, 1.8 Hz
1H), 8.54 (dq, J = 4.8, 0.9 Hz, 1H).
5.2.28. 5,8-Dichloro-N-[2-(2-chlorophenyl)ethyl]-6,7-di-
hydroxy-3,4-dihydroisoquinoline-2(1H)-carbothioamide
(184). Yield: 67%. H NMR (CD3OD 400 MHz) d 2.76
1
(t, J = 5.8 Hz, 2H), 3.10 (t, J = 7.2 Hz, 2H), 3.87 (t,
J = 7.2 Hz, 2H), 3.95 (t, J = 5.8 Hz, 2H), 4.83 (s, 2H),
7.13 (m, 2H), 7.23 (m, 1H), 7.31 (m, 1H). 13C NMR
(CD3OD 100 MHz) d 27.1, 33.8, 45.8, 46.3, 49.3,
118.4, 120.2, 124.2, 125.9, 128.0, 129.0, 130.4, 132.4,
135.1, 138.4, 142.5, 142.8, 182.6. HRMS (ESI) calcu-
lated for C18H18Cl3N2O2S (M+H) 431.0155, found
431.0149.
Coupling method, general method: 5,8-dichloro-1,2,3,4-
tetrahydroisoquinoline-6,7-diol
2,3,4,5-tetrahydro-1H-2-benzazepine-7,8-diol
hydrobromide
or
hydro-
bromide (1 equiv) was dissolved in DMF and trieth-
ylamine (3 equiv) was added. This mixture was
stirred for 15–30 min and then the corresponding iso-
thiocyanate (1.2 equiv) was added. This mixture was
stirred for additional 4 h and then concentrated.
The residue was dissolved in EtOAc and washed
with water. The organic phase was dried (MgSO4)
and concentrated to give the crude product, typically
as a yellow oil. The thiourea was purified by chro-
matography on silica gel (heptane:EtOAc + 1%
AcOH).
5.2.29. 5,8-Dichloro-N-[2-(3-chlorophenyl)ethyl]-6,7-di-
hydroxy-3,4-dihydroisoquinoline-2(1H)-carbothioamide
1
(185). Yield: 65%. H NMR (CD3OD 400 MHz) d 2.79
(t, J = 5.8 Hz, 2H), 2.95 (t, J = 7.3 Hz, 2H), 3.83 (t,
J = 7.3 Hz, 2H), 3.96 (t, J = 5.8 Hz, 2H), 4.85 (s, 2H),
7.18 (m, 4H). 13C NMR (CD3OD 100 MHz) d 27.1,
35.8, 45.8, 47.8, 49.3, 118.5, 120.3, 124.3, 125.9, 127.3,
128.4, 130.0, 130.9, 135.2, 142.7, 142.9, 143.2, 182.7.
HRMS (ESI) calculated for C18H18Cl3N2O2S (M+H)
431.0155, found 431.0152.
5.2.24. N-Benzyl-5,8-dichloro-6,7-dihydroxy-3,4-dihydro-
isoquinoline-2(1H)-carbothioamide (180). Yield: 24%. H
1
NMR (CD3OD 400 MHz) d 2.81 (t, J = 5.8 Hz, 2H),
4.00 (t, J = 5.8 Hz, 2H), 4.90 (s, 2H), 4.94 (s, 2H), 7.18
(m, 1H), 7.28 (m, 4H). 13C NMR (CD3OD 100 MHz)
d 27.3, 46.0, 49.6, 50.2, 118.5, 120.2, 124.3, 125.9,
127.9, 128.4, 128.4, 129.3, 129.3, 140.5, 142.6, 142.9,
183.1. HRMS (ESI) calculated for C17H17Cl2N2O2S
(M+H) 383.0388, found 383.0379.
5.2.30. N-[2-(4-Fluorophenyl)ethyl]-5,8-dichloro-6,7-dihy-
droxy-3,4-dihydroisoquinoline-2(1H)-carbothioamide (186).
Yield: 75%. H NMR (CD3OD 400 MHz) d 2.76 (t,
J = 5.8 Hz, 2H), 2.91 (t, J = 7.4 Hz, 2H), 3.80 (t,
J = 7.4 Hz, 2H), 3.94 (t, J = 5.8 Hz, 2H), 4.84 (s, 2H),
6.93 (m, 2H), 7.19 (m, 2H). 13C NMR (CD3OD 100
1

2538
M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
MHz) d 27.1, 35.4, 45.8, 48.2, 49.3, 115.9 (d, J = 21 Hz),
115.9 (d, J = 21 Hz), 118.4, 120.2, 124.2, 125.8, 131.5 (d,
JF=8 Hz), 131.5 (d, JF=8 Hz), 136.6 (d, JF=3 Hz), 142.5,
142.8, 162.9 (d, JF=241 Hz), 182.4. HRMS (ESI) calcu-
lated for C18H18Cl2FN2O2S (M+H) 415.0450, found
415.0446.
5.2.36. 5,8-Dichloro-6,7-dihydroxy-N-(2-pyridin-3-yleth-
yl)-3,4-dihydroisoquinoline-2(1H)-carbothioamide (192).
Yield: 39%. H NMR (CD3OD 400 MHz) d 2.77 (t,
1
J = 5.8 Hz, 2H), 3.01 (t, J = 7.1 Hz, 2H), 3.87 (t,
J = 7.1 Hz, 2H), 4.08 (t, J = 5.8 Hz, 2H), 4.84 (s, 2H),
7.31 (dd, J = 7.8, 4.9 Hz, 1H), 7.70 (ddd, J = 7.8, 1.8,
1.6 Hz, 1H), 8.35 (dd, J = 4.9, 1.6 Hz, 1H), 8.41 (d,
J = 1.8 Hz, 1H). 13C NMR (CD3OD 100 MHz) d 27.1,
33.3, 45.8, 47.4, 49.3, 118.4, 120.2, 124.2, 125.1, 125.8,
137.4, 138.9, 142.6, 142.9, 147.7, 150.3, 182.7. HRMS
(ESI) calculated for C17H18Cl2N3O2S (M+H)
398.0497, found 398.0511.
5.2.31. N-[2-(4-Bromophenyl)ethyl]-5,8-dichloro-6,7-dihy-
droxy-3,4-dihydroisoquinoline-2(1H)-carbothioamide (187).
1
Yield: 40%. H NMR (CD3OD 400 MHz) d 2.77 (t,
J = 5.8 Hz, 2H), 2.90 (t, J = 7.3 Hz, 2H), 3.81 (t,
J = 7.3 Hz, 2H), 3.94 (t, J = 5.8 Hz, 2H), 4.84 (s, 2H),
7.11 (d, J = 8.4 Hz, 2H), 7.36 (d, J = 8.4 Hz, 2H). 13C
NMR (CD3OD 100 MHz) d 27.1, 35.6, 45.8, 47.8,
49.3, 118.4, 120.3, 120.9, 124.2, 125.8, 131.9, 131.9,
132.4, 132.4, 140.0, 142.6, 142.9, 182.5. HRMS (ESI)
calculated for C18H18Cl2BrN2O2S (M+H) 474.9649,
found 474.9658.
5.2.37. 5,8-Dichloro-6,7-dihydroxy-N-(2-pyridin-2-yleth-
yl)-3,4-dihydroisoquinoline-2(1H)-carbothioamide (193).
1
Yield: 14%. H NMR ((CD3)2SO 400 MHz) d 2.69 (t,
J = 5.7 Hz, 2H), 3.02 (t, J = 7.5 Hz, 2H), 3.83 (m, 2H),
3.91 (t, J = 5.7 Hz, 2H), 4.89 (s, 2H), 7.23 (m, 2H),
7.68 (dt, J= 7.7, 1.8 Hz, 1H), 8.04 (t, J = 5.0 Hz, 1H),
8.49 (d, J = 4.2 Hz, 1H). 13C NMR ((CD3)2SO 100
MHz) d 25.8, 36.8, 43.8, 45.2, 48.2, 117.5, 119.4, 121.5,
123.2, 123.4, 124.4, 136.5, 141.5, 141.8, 149.0, 159.3,
180.1. HRMS (ESI) calculated for C17H18Cl2N3O2S
(M+H) 398.0497, found 398.0478.
5.2.32. 5,8-Dichloro-6,7-dihydroxy-N-[2-(4-hydroxyphenyl)-
ethyl]-3,4-dihydroisoquinoline-2(1H)-carbothioamide (188).
Yield: 20%. 1H NMR (CD3OD 400 MHz) d 2.80 (t,
J = 5.9 Hz, 2H), 2.84 (t, J = 7.3 Hz, 2H), 3.78 (t,
J = 7.3 Hz, 2H), 3.96 (t, J = 5.9 Hz, 2H), 4.87 (s, 2H), 6.69
(dd, J = 6.5, 2.0 Hz, 2H), 7.03 (dd, J = 6.5, 2.0 Hz, 2H).
13C NMR (CD3OD 100 MHz) d 27.2, 35.4, 45.8, 47.9,
49.3, 116.2, 116.2, 118.5, 120.3, 124.3, 125.9, 130.8, 130.8,
131.6, 142.7, 142.9, 156.8, 182.5. HRMS (ESI) calculated
for C18H19Cl2N2O3S (M+H) 413.0493, found 431.0503.
5.2.38. N-(4-Chlorobenzyl)-7,8-dihydroxy-1,3,4,5-tetra-
hydro-2H-2-benzazepine-2-carbothioamide (194). Yield:
63%. 1H NMR (CD3OD 400 MHz) d 1.82 (m, 2H),
2.80 (m, 2H), 4.12 (bs, 2H), 4.73 (s, 2H), 4.80 (s, 2H),
6.61 (s, 1H), 6.81 (s, 1H), 7.11 (d, J = 8.4 Hz, 2H),
7.21 (d, J = 8.4 Hz, 2H). 13C NMR (CD3OD 100
MHz) d 28.8, 34.9, 49.3, 49.8, 55.0, 118.3, 118.5, 128.7,
129.3, 129.3, 129.8, 129.8, 133.4, 134.3, 139.4, 143.7,
5.2.33. 5,8-Dichloro-6,7-dihydroxy-N-(3-phenylpropyl)-
(189).
3,4-dihydroisoquinoline-2(1H)-carbothioamide
1
Yield: 50%. H NMR (CD3OD 400 MHz) d 1.96 (m,
2H), 2.64 (t, J = 5.8 Hz, 2H), 2.79 (t, J = 7.4 Hz, 2H),
3.67 (t, J = 7.4 Hz, 2H), 3.93 (t, J = 5.8 Hz, 2H), 4.83
(s, 2H), 7.10 (m, 1H), 7.21 (m, 4H). 13C NMR (CD3OD
100 MHz) d 27.2, 32.0, 34.4, 45.8, 46.8, 49.3, 118.5,
120.2, 124.3, 125.9, 126.8, 129.3, 129.3, 129.4, 129.4,
142.6, 142.9, 143.2, 182.4. HRMS (ESI) calculated for
C19H21Cl2N2O2S (M+H) 411.0701, found 411.0692.
145.3,
C18H20ClN2O2S (M+H) 363.0934, found 363.0935.
181.9.
HRMS
(ESI)
calculated
for
5.2.39. 7,8-Dihydroxy-N-(2-phenylethyl)-1,3,4,5-tetrahy-
dro-2H-2-benzazepine-2-carbothioamide (195). Yield:
58%. 1H NMR (CD3OD 400 MHz) d 1.76 (m, 2H),
2.77 (m, 2H), 2.87 (t, J = 7.5 Hz, 2H), 3.76 (t,
J = 7.5 Hz, 2H), 4.03 (bs, 2H), 4.67 (s, 2H), 6.59 (s,
1H), 6.78 (s, 1H), 7.15 (m, 3H), 7.24 (m, 2H). 13C
NMR (CD3OD 100 MHz) d 28.8, 34.7, 36.4, 48.2,
54.2, 58.3, 118.2, 118.3, 127.2, 128.8, 129.4, 129.4,
129.9, 129.9, 134.1, 140.7, 143.8, 145.4, 181.2. HRMS
(ESI) calculated for C19H23N2O2S (M+H) 343.1480,
found 343.1493.
5.2.34. 5,8-Dichloro-6,7-dihydroxy-N-octyl-3,4-dihydro-
isoquinoline-2(1H)-carbothioamide (190). Yield: 59%.
1H NMR (CD3OD 400 MHz) d 0.88 (t, J = 7.0 Hz,
3H), 1.30 (m, 10H), 1.62 (bs, 2H), 2.81 (t, J = 5.8 Hz,
2H), 3.61 (t, J = 7.4 Hz, 2H), 3.98 (t, J = 5.8 Hz, 2H),
4.89 (s, 2H). 13C NMR (CD3OD 100 MHz) d 14.4,
23.7, 27.2, 28.0, 30.3, 30.4, 30.5, 33.0, 45.8, 47.1, 49.4,
118.4, 120.2, 124.3, 125.9, 142.6, 142.9, 182.4. HRMS
(ESI) calculated for C18H27Cl2N2O2S (M+H)
405.1170, found 405.1162.
5.2.40. N-[2-(4-tert-Butylphenyl)ethyl]-7,8-dihydroxy-1,3,
4,5-tetrahydro-2H-2-benzazepine-2-carbothioamide (196).
Yield: 72%. 1H NMR (CD3OD 400 MHz) d 1.28 (s, 9H),
1.72 (m, 2H), 2.74 (m, 2H), 2.83 (t, J = 7.5 Hz, 2H), 3.74
(t, J = 7.5 Hz, 2H), 4.00 (bs, 2H), 4.66 (s, 2H), 6.60 (s,
1H), 6.79 (s, 1H), 7.07 (d, J = 8.3 Hz, 2H), 7.28 (d,
J = 8.3 Hz, 2H). 13C NMR (CD3OD 100 MHz) d 28.8,
31.8, 31.8, 31.8, 34.7, 35.2, 35.8, 48.2, 54.5, 55.3, 118.2,
118.4, 126.3, 126.31, 128.5, 129.6, 129.6, 134.1, 137.6,
143.7, 145.3, 150.1, 181.1. HRMS (ESI) calculated for
C23H31N2O2S (M+H) 399.2107 found 399.2108.
5.2.35. 5,8-Dichloro-6,7-dihydroxy-N-(2-pyridin-4-yleth-
yl)-3,4-dihydroisoquinoline-2(1H)-carbothioamide (191).
1
Yield: 13%. H NMR (CD3OD 400 MHz) d 2.79 (t,
J = 5.8 Hz, 2H), 3.03 (t, J = 7.0 Hz, 2H), 3.90 (t,
J = 7.0 Hz, 2H), 3.97 (t, J = 5.8 Hz, 2H), 4.85 (s, 2H),
7.30 (d, J = 6.0 Hz, 2H), 8.38 (d, J = 6.0 Hz, 2H). 13C
NMR (CD3OD 100 MHz) d 27.1, 35.6, 45.9, 46.9,
49.3, 118.5, 120.3, 124.2, 125.9, 126.2, 126.2, 142.7,
143.03, 149.8, 149.8, 151.7, 182.8. HRMS (ESI) calcu-
lated for C17H18Cl2N3O2S (M+H) 498.0497, found
498.0514.
5.2.41. N-[2-(2-Chlorophenyl)ethyl]-7,8-dihydroxy-1,3,4,
5-tetrahydro-2H-2-benzazepine-2-carbothioamide (197).
1
Yield: 22%. H NMR (CD3OD 400 MHz) d 1.75 (m,

M. Berglund et al. / Bioorg. Med. Chem. 16 (2008) 2529–2540
2539
2H), 2.77 (m, 2H), 3.15 (t, J = 7.0 Hz, 2H), 3.80 (t,
5.3. Test procedure
J = 7.0 Hz, 2H), 4.02 (bs, 2H), 4.70 (s, 2H), 6.60 (s,
1H), 6.78 (s, 1H), 7.15 (m, 3H), 7.30 (m, 1H). 13C
NMR (CD3OD 100 MHz) d 28.8, 33.9, 34.7, 46.2,
54.1, 55.2, 118.2, 118.3, 128.1, 129.0, 130.0, 130.3,
132.5, 132.7, 134.1, 138.3, 143.8, 145.3, 181.4. HRMS
(ESI) calculated for C19H22ClN2O2S (M+H) 377.1090,
found 377.1089.
The bronchorelaxing effect of the compounds reported
here was evaluated in human small airways. Human
lung tissue was obtained from patients undergoing
lobectomy due to lung carcinoma in accordance with
procedures approved by Lund Ethical Committee,
and treated as described in Refs. 9,17 In short, lung tis-
sue was placed in a dissection bowl, an airway was
identified and a bronchial preparation from a bronchus
with a diameter of 0.5–1.5 mm was obtained. This was
mounted in the experimental chamber for force mea-
surements. The experimental chamber was kept at
37 ꢁC and was continuously perfused with physiologi-
cal saline solution. The force development of the prep-
aration was registered on a computer. After mounting
in the chamber and a period of adjustment, the prepa-
ration was contracted with LTD4 and stretched repeat-
edly until it exerted a force of 1.2 mN. This was
followed by 2 cycles with control contractions consist-
ing of 60 min relaxation with LTD4-free physiological
saline solution and 30 min contraction with LTD4. If
these two consecutive LTD4 contractions differed by
less than 10%, the preparations were considered to be
stable and the experiments begun. The inhibitory po-
tency of the different substances was evaluated by hav-
ing 10 lM of the test-substance present during one
whole cycle (1 h of LTD4-free PSS followed by
30 min of contraction with 10 nM LTD4). This con-
traction was then compared to the previous one and
the potency is expressed as the arithmetic mean (
standard error of the mean) of the contraction remain-
ing, expressed as percentage, after exposure to test-sub-
stance. Noteworthy, often each test-chamber contained
two preparations from the same patient. The mean of
the observed contractions was then regarded as a single
value. At the end of the experiments, the preparations
were exposed to 0 Ca2+ -solution, to establish the base-
line tension level.
5.2.42. N-[2-(3-Chlorophenyl)ethyl]-7,8-dihydroxy-1,3,4,
5-tetrahydro-2H-2-benzazepine-2-carbothioamide (198).
1
Yield: 66%. H NMR (CD3OD 400 MHz) d 1.76 (m,
2H), 2.76 (m, 2H), 2.87 (t, J = 7.3 Hz, 2H), 3.75 (t,
J = 7.3 Hz, 2H), 4.01 (bs, 2H), 4.68 (s, 2H), 6.59 (s,
1H), 6.79 (s, 1H), 7.05 (dd, J = 7.1, 1.7 Hz, 1H), 7.18
(m, 3H). 13C NMR (CD3OD 100 MHz) d 28.8, 34.7,
36.0, 47.8, 54.3, 55.5, 118.2, 118.3, 127.3, 128.4, 128.6,
129.9, 130.9, 134.1, 135.1, 143.1, 143.7, 145.3, 181.2.
HRMS (ESI) calculated for C19H22ClN2O2S (M+H)
377.1090, found 377.1086.
5.2.43. N-[2-(4-Fluorophenyl)ethyl]-7,8-dihydroxy-1,3,4,
5-tetrahydro-2H-2-benzazepine-2-carbothioamide (199).
Yield: 26.4%. 1H NMR (CD3OD 400 MHz) d 1.75
(m, 2H), 2.77 (m, 2H), 2.85 (t, J = 7.4 Hz, 2H), 3.75
(t, J = 7.4 Hz, 2H), 4.03 (bs, 2H), 4.68 (s, 2H), 6.60
(s, 1H), 6.80 (s, 1H), 6.95 (m, 2H), 7.13 (m, 2H).
13C NMR (CD3OD 100 MHz) d 28.8, 34.8, 35.5,
48.1, 54.3, 55.2, 116.0 (d, JF=21 Hz), 116.0 (d,
JF=21 Hz), 118.2, 118.4, 128.8, 131.5 (d, JF=8 Hz),
131.5 (d, JF=8 Hz), 134.1, 136.6 (d, JF=3 Hz), 143.8,
154.4, 163.0 (d, JF=251 Hz), 181.2. HRMS (ESI) cal-
culated for C19H22FN2O2S (M+H) 361.1386, found
361.1373.
5.2.44. N-[2-(4-Bromophenyl)ethyl]-7,8-dihydroxy-1,3,4,5-
tetrahydro-2H-2-benzazepine-2-carbothioamide (200).
Yield: 32%. 1H NMR (CD3OD 400 MHz) d 1.74
(m, 2H), 2.76 (m, 2H), 2.84 (t, J = 7.3 Hz, 2H),
3.75 (t, J = 7.3 Hz, 2H), 4.02 (bs, 2H), 4.69 (s,
2H), 6.60 (s, 1H), 6.81 (s, 1H), 7.05 (d, J = 8.3 Hz,
2H), 7.38 ( d, J = 8.3 Hz, 2H). 13C NMR (CD3OD
Acknowledgments
100 MHz)
d 28.8, 34.8, 35.8, 47.8, 54.5, 55.6,
118.2, 118.4, 120.9, 128.8, 131.9, 131.9, 132.4, 132.4,
134.1, 140.1, 143.7, 145.3, 181.2. HRMS (ESI) calcu-
lated for C19H20BrN2O2S (M-H) 419.0429, found
419.0413.
Financial support from the Swedish Board for Scientific
Research (VR) and SIDA/Sarec is gratefully
acknowledged.
5.2.45. 7,8-Dihydroxy-N-(3-phenylpropyl)-1,3,4,5-tetra-
hydro-2H-2-benzazepine-2-carbothioamide (201). Yield:
37%. 1H NMR (CD3OD 400 MHz) d 1.79 (m, 2H),
1.88 (dd, J = 7.0 Hz, 7.0 Hz, 2H), 2.55 (t, J = 7.0 Hz,
2H), 2.79 (m, 2H), 3.60 (t, J = 7.0 Hz, 2H), 4.08 (bs,
2H), 4.65 (s, 2H), 6.60 (s, 1H), 6.84 (s, 1H), 7.13 (m,
3H), 7.24 (m, 2H). 13C NMR (CD3OD 100 MHz) d
28.9, 32.3, 34.2, 34.8, 46.6, 54.7, 54.7, 118.3, 118.3,
126.7, 128.8, 129.3, 129.3, 129.4, 129.4, 134.2, 143.3,
143.8, 145.4, 181.1. HRMS (ESI) calculated for
C20H25N2O2S (M+H) 357.1636, found 357.1641.
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