Synthesis and radioligand binding studies of methoxylated 1,2,3,4-tetrahydroisoquinolinium derivatives as ligands of the apamin-sensitive Ca2+-activated K+ channels

Article abstract of DOI:10.1021/jm0607395

Several methoxylated 1,2,3,4-tetrahydroisoquinoliniums derived from N-methyl-laudanosine and N-methyl-noscapine were synthesized and evaluated for their affinity for apamin-sensitive binding sites. The quaternary ammonium derivatives have a higher affinit

Full text of DOI:10.1021/jm0607395

7208
J. Med. Chem. 2006, 49, 7208-7214
Synthesis and Radioligand Binding Studies of Methoxylated 1,2,3,4-Tetrahydroisoquinolinium
Derivatives as Ligands of the Apamin-Sensitive Ca2+-Activated K+ Channels
Amaury Graulich,*,† Jacqueline Scuve´e-Moreau,‡ Livia Alleva,‡ Ce´dric Lamy,‡ Olivier Waroux,‡ Vincent Seutin,‡ and
Jean-Franc¸ois Lie´geois†
Drug Research Center, Laboratory of Medicinal Chemistry, UniVersity of Lie`ge, aVenue de l’Hoˆpital, 1 (B36), B-4000 Lie`ge 1, Belgium, and
Research Center for Cellular and Molecular Neurobiology, Laboratory of Pharmacology, UniVersity of Lie`ge, aVenue de l’Hoˆpital, 1 (B36),
B-4000 Lie`ge 1, Belgium
ReceiVed June 20, 2006
Several methoxylated 1,2,3,4-tetrahydroisoquinoliniums derived from N-methyl-laudanosine and N-methyl-
noscapine were synthesized and evaluated for their affinity for apamin-sensitive binding sites. The quaternary
ammonium derivatives have a higher affinity with regard to the tertiary amines. 6,7-Dimethoxy analogues
possess a higher affinity than the 6,8- and 7,8-dimethoxy isomers. A 3,4-dimethoxybenzyl or a
2-naphthylmethyl moiety in C-1 position are more favorable than a 3,4-dimethoxyphenethyl group. Smaller
groups such as propyl or isobutyl are unfavorable. In 6,7-dimethoxy analogues, increasing the size and
lipophilicity with a naphthyl group in the C-1 position leads to a slight increase of affinity, while the same
group in the 6,7,8-trimethoxy series is less favorable. The 6,7,8-trimethoxy derivative 3f is the first tertiary
amine in the series to possess an affinity close to that of N-methyl-laudanosine and N-methyl-noscapine.
Moreover, electrophysiological studies show that the most effective compound 4f blocks the apamin-sensitive
afterhyperpolarization in rat dopaminergic neurons.
Introduction
octadecapeptide structure isolated from honey bee (Apis mel-
lifera) venom, which possesses an affinity in the picomolar
range.17 Leiurotoxin I, another peptide isolated from the scorpion
Androctonus mauretanicus mauretanicus, blocks human SK2
and SK3, but not SK1 channels.18 Recently isolated from the
venom of the scorpion Mesobuthus tamulus, tamapin represents
a novel, promising pharmacological tool, as it blocks SK2
channels ∼1750- and ∼70-fold more potently than SK1 and
SK3 channels, respectively, making it the most potent and
selective SK2 channel natural toxin characterized so far.19
Besides these toxins, a small number of nonpeptidic blockers
have been discovered. Dequalinium has an affinity of 245 nM
for rat cortical apamin-sensitive sites.20 Extensive structure-
activity relationship studies on dequalinium (Figure 1) led to a
family of dimeric compounds, including UCL1684 and UCL1848
(Figure 1), with a low nanomolar affinity.21 On the other hand,
N-methyl-bicuculline (Figure 2) was shown to potentiate burst
firing in dopaminergic neurons by blocking the apamin-sensitive
Ca2+-activated K+ current.22,23 Unfortunately, this compound
also possesses an unwanted GABAA receptor antagonist effect.
Ion channels have been shown to play a role in many
functions, including neuronal communication and behavioral
plasticity, secretion, and cell proliferation. Therefore, ion
channels represent promising targets for the development of new
CNS drugs.1 In addition to their role in regulating cell
excitability, the channels themselves can be modulated in a cell-
specific manner through second messengers, hormones, and
neurotransmitters. Calcium-activated potassium channels are
separated in three families: BK, IK, and SK, according to their
large, intermediate, and small conductance, respectively.2,3 Small
conductance Ca2+-activated K+ (SK) channels are voltage-
insensitive and are activated by an increase in the intracellular
calcium concentration. They play an important role in modulat-
ing the firing rate and the firing pattern of several types of
neurons, because their activation induces a prolonged postspike
afterhyperpolarization (AHP).4-6 Three SK channel subunits
have been identified by DNA cloning, namely, SK1, SK2, and
SK3.4 The distribution of the SK channel subunits has been
investigated in the rat by using in situ hybridization and
immunohistochemistry. These experiments revealed that SK1
and SK2 subunits are mostly expressed in the cortex and
hippocampus,7 while SK3 channel expression is higher in
subcortical areas, especially in the monoamine cell regions.
According to the literature, SK channel modulation could have
therapeutic effects in various diseases such as cognitive
dysfunction,8-12 neuronal hyperexcitability,13 dopamine related
disorders,14-16 and depression.11
Nature is frequently the source of potent ionotropic agents.
Indeed, venoms of arthropods or insects contain natural peptides,
which are high affinity blockers of SK channels. So far, the
most potent SK channel blocker is apamin, a toxin with an
* To whom correspondence should be addressed: Tel.: +32 (0) 43 66
43 77. Fax: +32 (0)43 66 43 62. E-mail: a.graulich@student.ulg.ac.be.
† Drug Research Center.
‡ Research Center for Cellular and Molecular Neurobiology.
Figure 1. Chemical structure of non-peptidic SK channel blockers.
10.1021/jm0607395 CCC: $33.50 © 2006 American Chemical Society
Published on Web 10/28/2006

Methoxylated 1,2,3,4-Tetrahydroisoquinoliniums
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 24 7209
Scheme 1. Synthesis of 1-Substituted Di- and Trimethoxylated
2,2-Dimethyl-1,2,3,4-tetrahydroisoquinolinium Derivatives by
the Bischler-Napieralski Procedurea
a Key: (i) R4COCl, NEt3, MeCOOEt, rt; (ii) P2O5, ArMe, reflux; (iii)
MeI, MeCN, reflux; (iv) NaBH4, MeOH, rt; (v) MeI, MeCN, reflux. R1,
R2, R3 ) H or OMe. R4 ) n-propyl (a), isobutyl (b), 2-(3,4-dimethoxyphe-
nyl)-ethyl (c), 3,4-dimethoxybenzyl (d,f), 2-naphthylmethyl (e).
Figure 2. Chemical structure of benzyltetrahydroisoquinoline deriva-
tives interacting with SK channels.
Scheme 2. Synthesis of 1-Substituted Di- and Trimethoxylated
2,2-Dimethyl-1,2,3,4-tetrahydroisoquinolinium Derivatives from
Reissert Compoundsa
Therefore, different molecules structurally close to N-methyl-
bicuculline were evaluated as potential specific SK channel
blockers. A first series of studies started with N-methyl-
laudanosine (NML;24-26 Figure 2) and more recently with
N-methyl-noscapine (NMN;26,27 Figure 2). These two molecules
block the AHP with a medium potency and are quickly
reversible.27 Different modifications of these compounds have
been carried out, such as the size and the substitution of the
side chain in the C-1 position,25 the absence of substituents in
the C-6 and C-7 positions,25 the nature of the fourth group on
the nitrogen,25 and also the presence of a halogen, an alkyl, or
a methoxy group in the C-8 position26 and the presence of a
halogen in the C-5 position.26 Because NML is chiral, both
enantiomers have been separated and tested.25 As no difference
was found between these two stereoisomers, all compounds were
tested as racemate.
In the present paper, we focus on the synthesis and biological
evaluation of new NML and NMN derivatives obtained by
changing the position of the methoxy groups on the tetrahy-
droisoquinoline ring and the nature of the substituent in the C-1
position.
a Key : (i) (Me)3SiCN, AlCl3, BzCl; (ii) NaH, DMF, -10 °C; (iii) R4X,
DMF, -10 °C; (iv) 50% NaOH, EtOH-H2O, reflux; (v) MeI, MeCN,
reflux; (vi) NaBH4, MeOH, rt; (vii) MeI, MeCN, reflux. R1, R2, R3 ) H or
OMe. R4X ) 3,4-dimethoxybenzyl chloride (b), 2-naphthylmethyl bromide
(a,c,d,e).
Chemistry
Two procedures are used depending on the benzene substitu-
tion of the tetrahydroisoquinoline ring. Indeed, some compounds
are prepared from a Bischler-Napieralski procedure (Scheme
1). The appropriate acid anhydride or acid chloride, the latter
being commercially available or directly prepared by reaction
of the corresponding carboxylic acid with thionyl chloride and
used without further purification, reacts with the adequate amine
to obtain the amide (1a-f). In the next step, the amide is
cyclodehydrated by heating to reflux with phosphorus pentoxide
in toluene to afford the 3,4-dihydroisoquinoline derivative (2a-
f).28 Subsequently, the 3,4-dihydroisoquinoline is methylated
by methyl iodide in refluxing MeCN. The resulting 2-methyl-
3,4-dihydroisoquinolinium analogue is immediately reduced by
sodium borohydride in MeOH to give the tertiary amine
(3a-f). By methylation of the 1,2,3,4-tetrahydroisoquinoline
derivative (3a-f) with methyl iodide, the 2,2-dimethyl-1,2,3,4-
tetrahydroisoquinolinium derivative is obtained (4a-f).
The other procedure follows Reissert compound pathway
(Scheme 2) for the alkylation of the isoquinoline nucleus29 in
the C-1 position. These intermediates are usually synthesized
from the appropriate nitrogen heterocycles and acyl chlorides
in the presence of a cyanide source.30 Indeed, the isoquinolines
react with benzoyl chloride, and the resulting acyliminium
undergoes a nucleophilic addition with cyanide to afford the
Reissert compounds (5a-d).31 This reaction is carried out with

7210 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 24
Graulich et al.
Table 1. Tertiary Analogues (3a-f, 7a-e): Physical Data and Preliminary Screening Results for Binding on Rat Cortical Apamin-Sensitive Sites
b
No
R1
R2
R3
R4
yield (%)
mp (°C)
formulaa
C15H23NO2‚HCl
%
3a
3b
3c
3d
3e
3f
7a
7b
7c
7d
7e
OMe
OMe
OMe
OMe
OMe
OMe
OMe
H
OMe
OMe
OMe
H
H
H
H
OMe
OMe
OMe
OMe
OMe
OMe
H
n-C3H7
i-C4H9
90
94
85
65
81
75
27
59
30
47
53
183-184
148-150
74-76
0
0
C16H25NO2‚HCl‚0.5H2O
C22H29NO4
3,4-(MeO)2C6H3CH2CH2
3,4-(MeO)2C6H3CH2
2-(C10H7)CH2
3,4-(MeO)2C6H3CH2
2-(C10H7)CH2
3,4-(MeO)2C6H3CH2
2-(C10H7)CH2
2-(C10H7)CH2
11
0
99-101
137-138
94-95
C21H27NO4‚C2H2O4‚0.75H2O
C23H25NO2
C22H29NO5‚C4H4O4‚H2O
C24H27NO3
H
0
OMe
OMe
OMe
OMe
OMe
H
27
0
86-88
159-161
108-110
212-214
149-151
C21H27NO4‚HCl
3
0
6
0
H
OMe
H
C23H25NO2‚C2H2O4
C23H25NO2‚HCl
H
2-(C10H7)CH2
C21H21N‚HCl‚0.75H2O
a According to CHN analysis. b The % of 125I-apamin displaced at 10 µM.
Table 2. Quaternary Analogues (4a-f, 8a-e): Physical Data and Preliminary Screening Results for Binding on Rat Cortical Apamin-Sensitive Sites
b
No
R1
R2
R3
R4
yield (%)
mp (°C)
formulaa
C16H26NO2I
%
4a
4b
4c
4d
4e
4f
8a
8b
8c
8d
8e
OMe
OMe
OMe
OMe
OMe
OMe
OMe
H
OMe
OMe
OMe
H
H
H
H
OMe
OMe
OMe
OMe
OMe
OMe
H
n-C3H7
i-C4H9
85
93
78
77
83
88
57
67
58
75
75
228-229
189-191
165-166
151-152
140-142
103-104
133-135
204-206
121-122
212-214
219-221
6
0
C17H28NO2I
3,4-(MeO)2C6H3CH2CH2
3,4-(MeO)2C6H3CH2
2-(C10H7)CH2
3,4-(MeO)2C6H3CH2
2-(C10H7)CH2
3,4-(MeO)2C6H3CH2
2-(C10H7)CH2
2-(C10H7)CH2
C23H32NO4I‚0.5H2O
C22H30NO4I‚0.5H2O
C24H28NO2I‚0.25H2O
C23H32NO5I‚0.25H2O
C25H30NO3I‚0.5H2O
C22H30NO4I
C24H28NO2I‚0.5H2O
C24H28NO2I‚0.5H2O
C22H24NI
32
20
2
72
53
24
18
70
1
H
OMe
OMe
OMe
OMe
OMe
H
H
OMe
H
H
2-(C10H7)CH2
a According to CHN analysis. b The % of 125I-apamin displaced at 10 µM.
trimethylsilyl cyanide in anhydrous CH2Cl2 in good yields. All
Reissert compounds (5a-d) are then deprotonated by sodium
hydride in DMF. The resulting Reissert anions are alkylated
by using 3,4-dimethoxybenzyl chloride or 2-(bromomethyl)-
naphthalene. Then, the alkylated Reissert compounds are
hydrolyzed to 1-(3,4-dimethoxybenzyl)-isoquinolines and 1-(2-
naphthylmethyl)-isoquinolines (6a-e). Compounds 6a-e are
methylated by methyl iodide in refluxing MeCN. Due to their
chemical lability, the resulting N-methylisoquinoliniums are
directly reduced to N-methyl-1,2,3,4-tetrahydroisoquinolines
(7a-e) by using an excess of sodium borohydride in MeOH.
A further methylation by using methyl iodide in refluxing MeCN
gives the quaternary ammoniums (8a-e).
respectively. The affinities (Ki) for the apamin-sensitive binding
sites of the rat cortex preparation are 5.4, 0.73, 2.2, and 0.91
µM for 4c, 4f, 8a, and 8d, respectively.
Compounds 3a-e, 4a, 4b, 4d, 4e, 7a-e, 8b, 8c, and 8e have
no significant affinity, because a negligible fraction of radio-
ligand is displaced.
In electrophysiological experiments, compound 4f blocks the
apamin-sensitive AHP in dopaminergic neurons with an IC50
of 9.0 ( 0.7 µM (n ) 3), while compound 3f blocks 51.9 (
1.2% of the AHP at 100 µM (n ) 3).
Discussion
The first benzylisoquinoline alkaloid reported to be active
on apamin-sensitive binding sites was the N-methyl derivative
of bicuculline (Figure 2). Unfortunately, this drug is not an
optimal tool because it presents a GABAA receptor antagonist
activity and a poor stability due to its lactone function in
physiological conditions.23 Then NML, a compound structurally
related to the bicuculline template, was prepared and tested to
determine its potency to block the apamin-sensitive AHP. This
compound revealed an interesting binding profile24 and a very
quickly reversible blocking effect on the apamin-sensitive AHP,
with no effect on GABAA transmission.27 However, after
extensive biological evaluation, a non-SK mediated effect on
the serotonergic neurons of the dorsal raphe has been detected.27
Structurally close to NML and N-methyl-bicuculline, NMN was
also prepared and evaluated in in vitro binding and electro-
physiological experiments. It was found that this drug has a
Results
Biological data from binding experiments on rat cortical
apamin-sensitive sites are summarized in Tables 1-3. A
preliminary screening is performed at a concentration of 10 µM
(Tables 1 and 2), then drugs that displace at least 25% of 125I-
apamin are further tested to determine their affinity (Table 3).
In our experimental conditions, 125I-apamin has a Kd of 2.02
( 0.31 pM, while the unlabeled apamin has an affinity of 3.8
( 1.1 pM.
The tertiary compound 3f displaces 27% of radioligand at
10 µM and has a Ki of 5.8 µM for the apamin-sensitive bind-
ing sites.
The quaternary ammonium compounds 4c, 4f, 8a, and 8d
displace 32, 72, 53, and 70% of the radioligand at 10 µM,

Methoxylated 1,2,3,4-Tetrahydroisoquinoliniums
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 24 7211
Table 3. Affinity of N-Methyl-laudanosine (NML), N-Methyl-noscapine (NMN), and Methoxylated Analogues for Binding on Rat Cortical
Apamin-Sensitive Sites
a
compd
R1
R2
R3
R4
%
Ki ( SDb (µM)
NML
NMN
3f
4c
4f
8a
8d
apamin
OMe
OMe
H
3,4-(MeO)2C6H3CH2
57
27
27
32
72
53
70
1.3 ( 0.02
3.6 ( 1.3
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
H
OMe
OMe
H
3,4-(MeO)2C6H3CH2
3,4-(MeO)2C6H3CH2CH2
3,4-(MeO)2C6H3CH2
2-(C10H7)CH2
5.8 ( 0.4
5.4 ( 0.6
0.73 ( 0.11
2.2 ( 0.4
0.91 ( 0.11
3.8 ( 1.1 pM
2-(C10H7)CH2
a The % of 125I-apamin displaced at 10 µM. b Only compounds displacing more than 25% of 125I-apamin at 10 µM were further evaluated; n ) 3 except
for NML (n ) 8).
quickly reversible effect on SK channels without affecting
serotonergic neurons but possesses a lower affinity than NML.27
Like NML, NMN is devoid of GABAA receptor antagonist
effects.27 From the structural point of view, the substitution of
the isoquinoline nucleus of NMN differs from that of NML by
a methoxy group in the C-8 position and by an unstable lactone
in the side chain in the C-1 position (Figure 2). Following these
observations, different compounds structurally close to the NML
and NMN templates were synthesized and evaluated by radio-
ligand binding studies to extend our medicinal chemistry
program.24-26
(Table 3). In this case, lipophilicity of the naphthyl group does
not appear favorable with such a substitution on the main ring
unless steric hindrance by the presence of the group in the C-8
position would be critical. This can be illustrated by the fact
that 6,8- (4d,e) and 7,8-dimethoxy (8b,c) substitution are
unfavorable (Table 2). Nevertheless, other parameters could be
involved since we previously reported that the compound
possessing only a methoxy group in the C-8 position does not
interact with the SK channel.26 This observation is interesting
and challenging because the 8-isopropyl derivative previously
described has an affinity similar to that of NML.26 The presence
of an electronically rich atom could be responsible for this
absence of affinity as we also reported that a chlorine or a
bromine in the C-8 position is not favorable.26
In our binding conditions, binding parameters of 125I-apamin
and apamin are in the same range than those previously
reported.20,32
To demonstrate the blocking potential of the drugs on SK
channels, the most effective compound of this series, compound
4f, was tested in electrophysiological experiments performed
in rat brain slices. In these experiments, compound 4f blocks
the AHP recorded in midbrain dopaminergic neurons with an
IC50 of 9.0 ( 0.7 µM, meaning that this compound is twice
more active than NML (IC50 ) 15 µM, n ) 3).24 These data
are in good agreement with in vitro binding data in which the
affinity of compound 4f (IC50 ) 0.73 ( 0.11 µM, n ) 3) is
also almost twice superior to that of NML (IC50 ) 1.3 ( 0.02
µM, n ) 8; Table 3). The lower activity of both drugs when
evaluated in brain slices may be due in part to a more difficult
access to the targets in such a preparation. Compound 3f, having
an affinity ∼5 times weaker than NML, induces a 50%
inhibition of the AHP at 100 µM. As a result of the low potency
of this compound further evaluations on selectivity were not
performed in this study.
In vitro binding results show that the quaternary compounds
(4a-f, 8a-e; Table 2) have a higher affinity than the tertiary
derivatives (3a-f, 7a-e; Table 1) for apamin-sensitive binding
sites. However, the tertiary amine 3f has an affinity of 5.8 µM
(Table 3). This is the first basic compound in this series with a
significant affinity for examined sites. Presently, all known SK
channel blockers have at least one ionized function. Indeed,
animal toxins like apamin, leiurotoxin, or tityus κ possess one
or more arginine residues.18,33-35 Synthetic blockers such as
dequalinium and UCL compounds (Figure 1) possess two
charged centers, namely, two quinolinium nuclei.20,21,33
When comparing the affinities of C-1-substituted compounds,
it is clear that the presence of a small alkyl group in this position
(4a,b) is deleterious for the affinity for rat brain apamin-sensitive
sites (Table 2). These data are in accordance with the inef-
ficiency of C-1-unsubstituted, methylated, and unsubstituted
benzylated derivatives previously reported.25 Otherwise, the
homologous derivative of NML 4c, possessing a 3,4-dimethox-
yphenethyl group, shows a lower affinity (Ki ) 5.4 ( 0.6 µM,
n ) 3) than NML (Ki ) 1.3 ( 0.02 µM, n ) 8; Table 3). Thus,
the presence of a 3,4-dimethoxybenzyl or a 2-naphthylmethyl
group in the C-1 position represents so far the most effective
substitution in these series. Moreover, these side chains have a
different impact according to the nature of the 1,2,3,4-tetrahy-
droisoquinoline nucleus. Indeed, in the 6,7-dimethoxy series,
8d with a naphthylmethyl group has an affinity inferior to 1
µM. Thus, increasing lipophilicity of this group would appear
favorable in this series. 6,7,8-Trimethoxy analogue 4f with a
3,4-dimethoxybenzyl moiety in the C-1 position is two times
more effective than NML with an affinity of 0.73 µM, while
the 2-naphthylmethyl analogue 8a has an affinity of 2.2 µM
Thus, among the chemical modifications performed on NML
and NMN structures, it appears that some limited changes
increase the interaction with the apamin-sensitive sites. The
presence of a 6,7,8-trimethoxy substitution on the 1,2,3,4-
tetrahydroisoquinoline nucleus is the most effective, followed
by 6,7-dimethoxy (NML) and 8-alkyl, while 6,8- and 7,8-
dimethoxy, 8-methoxy, 5-halogeno, and 8-halogeno substitution
are less interesting.25,26 Concerning the group in the C-1 position
of the 1,2,3,4-tetrahydroisoquinoline ring, a 3,4-dimethoxyben-
zyl (NML, 4f) or a 2-naphthylmethyl (8a, 8d) moiety appears
to be the most efficient to increase the affinity in these series
of compounds, while a small alkyl group must be avoided. The
most interesting combinations found are, on the one hand, a
6,7,8-trimethoxy-1,2,3,4-tetrahydroisoquinoline with a 3,4-

7212 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 24
Graulich et al.
aqueous HCl (100 mL). The acidic layer was washed with Et2O (3
× 20 mL) and then basified with NH4OH. The suspension was
extracted with CH2Cl2 (3 × 30 mL). The organic layers were
collected, dried over anhydrous MgSO4, and evaporated under
reduced pressure to afford colorless oil, which was isolated as
hydrochloride salt. Further recrystallization from MeCN/Et2O gave
compound 3a as a white solid (0.74 g); yield, 90%; mp, 183-184
°C. Anal. (C15H23NO2‚HCl) C, H, N.
General Preparation of Quaternary Ammonium Derivatives
(4a-f, 8a-e): 6,7-Dimethoxy-2,2-dimethyl-1-propyl-1,2,3,4-
tetrahydroisoquinolinium Iodide (4a). A solution of compound
3a (0.25 g; 1.0 mmol) in MeCN (10 mL) was refluxed with an
excess of methyl iodide (0.5 mL; 8 mmol). After 4 h, the solvent
was removed under reduced pressure. The white residue (0.33 g)
recrystallized from MeCN gave 4a as a white solid; yield, 85%;
mp, 228-229 °C. Anal. (C16H26NO2I) C, H, N.
dimethoxybenzyl in the C-1 position (4f), and, on the other hand,
a 6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline with a 2-naph-
thylmethyl in the C-1 position (8d). In the first case, it is
interesting to mention that such a combination leads for the first
time to a tertiary amine (3f) with a significant affinity for
apamin-sensitive binding sites. Although its affinity remains in
the micromolar range, compound 3f with its nonpermanent
ionized function may be useful for the development of more
potent lipophilic blockers, able to cross the blood brain barrier.
Indeed, a critical step in the evaluation of the consequences of
SK channel modulation in the CNS is the development of drugs
that can be administrated systemically. When such a compound
will be developed, its putative selectivity for SK channel
subtypes and other targets will be investigated.
General Procedure of Reissert Compounds (5a-c): 2-Ben-
zoyl-1-cyano-6,7,8-trimethoxy-1,2-dihydroisoquinoline (5a). An-
hydrous aluminum chloride (10 mg) was added to a stirred solution
of 6,7,8-trimethoxyisoquinoline (3.44 g; 15.7 mmol) and trimeth-
ylsilyl cyanide (3.9 mL; 31.4 mmol) in anhydrous CH2Cl2 (50 mL)
at room temperature. Then benzoyl chloride (3.6 mL; 31.4 mmol)
was dropwise added to the stirred solution over a course of 5 min.
The mixture was warmed to 30 °C if no exotherm had begun after
the addition of benzoyl chloride. After stirring for an additional 3
h period, water (50 mL) was added and stirring was continued for
30 min. The organic layer was collected and washed successively
with 1 N aqueous HCl (2 × 50 mL), water (50 mL), 1 N aqueous
NaOH (2 × 50 mL), and finally water (50 mL). The organic
solution was dried over anhydrous MgSO4 and evapored under
reduced pressure to give an oil that was triturated with Et2O (20
mL), resulting in crystallization. The solid was collected, washed
with small volumes of Et2O, and dried (3.3 g) to give 5a as a white
crystal; yield, 60%; mp, 155-157 °C. Anal. (C20H18N2O4) C, H,
N.
Experimental Section
Chemistry. Melting points were determined on a Bu¨chi-Tottoli
capillary melting point apparatus in open capillary and are uncor-
rected. NMR spectra were recorded on a Bruker Avance 500
spectrometer at 500 MHz. IR spectra were performed on a Perkin-
Elmer FTIR-1750 spectrometer. IR spectra were measured using
KBr discs. Only significant bands from IR are reported. Elemental
analyses were determined using a Carlo-Erba elemental analyzer
CHNS-O model EA1108, and the results are within 0.4% of the
theoretical values. All starting materials and reagents were obtained
from Aldrich Chemical Co. or from Acros Organics Co. and were
used without further purification. 6,7- and 7,8-dimethoxy-isoquino-
line and 6,7,8,-trimethoxy-isoquinoline are prepared according to
the procedure previously described.29 Concentration and evaporation
refer to removal of volatile materials under reduced pressure (10-
15 mmHg at 30-50 °C) on a Buchi rotavapor.
General Preparation of Amides (1a-f): N-[2-(3,4-Dimethox-
yphenyl)-ethyl]-butanamide (1a). Butyric anhydride (3.6 mL; 23.5
mmol) was added dropwise to a solution of 2-(3,4-dimethoxyphe-
nyl)-ethylamine (4.0 mL; 23.5 mmol) and of an excess of
triethylamine (5 mL) in MeCOOEt (30 mL) at room temperature.
After 1 h, the organic medium was washed with 5% aqueous Na2-
CO3 (2 × 50 mL) and then with 1 N aqueous HCl until the washing
phase became acidic. The organic layer was dried over anhydrous
MgSO4 and evaporated under reduced pressure. The crude product
was recrystallized from petroleum ether 100-140 °C to afford 1a
as a white solid (4.7 g); yield, 79%; mp 47-48 °C. Anal. (C14H21-
NO3) C, H, N.
General Procedure for Preparing 3,4-Dihydroisoquinoline
Derivatives (2a-f): 6,7-Dimethoxy-1-propyl-3,4-dihydroiso-
quinoline Hydrochloride (2a). A solution of compound 1a (1.5
g; 6 mmol) in refluxing ArMe (50 mL) was treated with phosphorus
pentoxide (5 g) added during 15 min. After the mixture had refluxed
30 min, ArMe was decanted and the sticky residue was dissolved
in water and washed with Et2O (2 × 30 mL). The aqueous solution
was made alkaline with NH4OH and extracted with CH2Cl2 (3 ×
20 mL). The organic layer was dried over anhydrous MgSO4 and
evaporated under reduced pressure. The crude oil was treated with
a saturated etherous HCl solution to isolate 2a as a hydrochloride
salt. Recrystallization from MeCN/Et2O gave 2a as a white solid;
yield, 81%; mp, 175-176 °C. Anal. (C14H19NO2. HCl) C, H, N.
General Procedure for Preparing 1,2,3,4-Tetrahydroiso-
quinoline Analogues (3a-f, 7a-e): 6,7-Dimethoxy-2-methyl-
1-propyl-1,2,3,4-tetrahydroisoquinoline Hydrochloride (3a). A
solution of 6,7-dimethoxy-1-propyl-3,4-dihydroisoquinoline (0.69
g; 2.9 mmol) in MeCN (10 mL) was refluxed with an excess of
methyl iodide (1.0 mL; 16 mmol). After 2 h, Et2O was added,
resulting in a rapid crystallization of yellow solid. The precipitate
was filtered off, washed with Et2O (2 × 10 mL), dried, and used
without further purification. Under an inert atmosphere, NaBH4
(0.82 g; 21.6 mmol) was added to a solution of the corresponding
isoquinolinium compound (1 g; 2.7 mmol) in MeOH (50 mL) at
room temperature. After 15 min, MeOH was removed under
reduced pressure, and the crude residue was dissolved in a 1 N
2-Benzoyl-1-cyano-1,2-dihydroisoquinoline (5d) is prepared ac-
cording to a previous report.36
General Procedure for Preparing 1-Substituted Isoquinolines
(6a-e): 6,7,8-Trimethoxy-1-(2-naphthylmethyl)-isoquinoline (6a).
A solution of 2-benzoyl-1-cyano-6,7,8-trimethoxy-1,2-dihydroiso-
quinoline 5a (2.41 g; 6.89 mmol) and of 2-(bromomethyl)-
naphthalene (1.3 g; 6.89 mmol) in DMF (15 mL) was dropwise
added to a stirred suspension of sodium hydride (0.2 g; 8.33 mmol)
in DMF (30 mL) at -10 °C. The medium was stirred for 4 h and
poured into ice-cold water (200 mL). The creamy solid was filtered
off. After drying, the solid was hydrolyzed by treatment with 50%
NaOH in a 1:1 EtOH-water solution at reflux. After removal of
EtOH, the crude residue was dissolved in ArMe (50 mL) and water
(50 mL). The organic layer was collected, washed with water (50
mL), and then extracted with 1 N aqueous HCl (2 × 50 mL). The
acidic layers were basified with concentrated NH4OH and finally
extracted with CH2Cl2 (3 × 30 mL). The organic layers were dried
over anhydrous MgSO4 and evaporated under reduced pressure to
afford a solid that was purified by flash chromatography (Me2-
CO). Finally the compound was recrystallized from petroleum ether
100-140 °C to give 6a as a cream solid; yield, 90%; mp, 184-
185 °C dec. Anal. (C23H21NO3) C, H, N.
Radioligand Binding Studies and Data Analysis. Synapto-
somes Preparation. Rats (male Wistar, (250 g) were killed by
decapitation, and the brains were quickly removed and kept on ice
during dissection. Crude cortex was dispersed in 0.32 M sucrose
by using a Potter homogenizer. After a first centrifugation at 1500
× g for 10 min, the supernatant was centrifuged at 25 000 × g for
10 min. The resulting pellet was dispersed in 5 mL of 0.32 M
sucrose to be aliquoted. Protein concentration was determined by
the method of Hartree with bovine serum albumin as a standard.37
Binding Experiments. The incubation buffer consisted of a 10
mM Tris-HCl (pH 7.5) solution containing 5.4 mM KCl and 0.1%
bovine serum albumin. The radioligand was 125I-apamin (Perkin-
Elmer, specific activity 81.4 TBq mmol-1). Glass fiber filters
(Whatman GF/C) used in these experiments were coated for 1 h in

Methoxylated 1,2,3,4-Tetrahydroisoquinoliniums
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 24 7213
0.5% polyethylenimine and then washed with 2.5 mL of the ice-
cold buffer just before use. Binding experiments were always
terminated as follows. Aliquots were filtered under reduced pressure
through Whatman filters. Filters were rapidly washed twice with
2.5 mL of buffer. The radioactivity remaining on the filter was
evaluated with a Packard Tri-Carb 1600TR liquid scintillation
analyzer with an efficacy of 69%. 125I-apamin binding to the filters
was also estimated in the absence of synaptosomes. This binding
was also subtracted from the total binding. Curve fitting was carried
out using GraphPad Prism.
Supporting Information Available: Routine experimental,
spectroscopic, and elemental analysis data. This material is available
free of charge via the Internet at http://pubs.acs.org.
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Acknowledgment. The technical assistance of Y. Abrassart,
S. Counerotte, L. Massotte, and J.-C. Van Heugen is gratefully
acknowledged. A.G. is a Research Fellow of the Fonds pour la
formation a` la Recherche Industrielle et Agricole (F.R.I.A.) and
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was financially supported in part by F.N.R.S. Grant Nos.
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pour la Recherche 2002 and 2003 of the University of Lie`ge
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