Synthesis and biological evaluation of a small molecule library of 3rd generation multidrug resistance modulators

Article abstract of DOI:10.1016/j.bmc.2009.01.072

The development of new modulators possessing high efficacy, low toxicity and high selectivity is a pivotal approach to overcoming P-glycoprotein (P-gp) mediated multidrug resistance (MDR) in tumour cells. In this study 39 compounds are presented which hav

Full text of DOI:10.1016/j.bmc.2009.01.072

Bioorganic & Medicinal Chemistry 17 (2009) 2524–2535
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and biological evaluation of a small molecule library of 3rd
generation multidrug resistance modulators
Werner Klinkhammer a, , Henrik Müller a, , Christoph Globisch a, Ilza K. Pajeva a,b, Michael Wiese a,
*
a Institute of Pharmacy, University of Bonn, An der Immenburg 4, 53121 Bonn, Germany
b Centre of Biomolecular Engineering, Bulgarian Academy of Science, Sofia, Bulgaria
a r t i c l e i n f o
a b s t r a c t
Article history:
The development of new modulators possessing high efficacy, low toxicity and high selectivity is a pivotal
approach to overcoming P-glycoprotein (P-gp) mediated multidrug resistance (MDR) in tumour cells. In
this study 39 compounds are presented which have been synthesized and pharmacologically investigated
in our laboratory. Similarly to the potent 3rd generation MDR modulator tariquidar (XR9576) the com-
pounds contain a tetrahydroisoquinoline–ethyl-phenylamine substructure that, in contrast to XR9576,
is connected to a smaller hydrophobic part, thus leading to molecules of lower molecular weight. The
connection between the tetrahydroisoquinoline–ethyl-phenylamine substructure and the hydrophobic
part was achieved through four different types of linkers: amide, urea, amide-ether and amide-styryl.
A number of structural modifications in the hydrophobic part were created. The calcein AM assay served
as test system to determine the P-gp transport inhibitory potencies of the compounds. For the amide lin-
ker derivatives a structure–activity relationship analysis was performed outlining which structural mod-
ifications contributed to the inhibitory potency. The compounds containing a bicyclic hydrophobic part
with a particular substituent in a specific orientation were identified as the most potent amide deriva-
tives. Among the urea derivatives the compounds with highest inhibitory potency possessed an ortho-
nitro substituent. The conformational analysis revealed that this position enables the formation of a
hydrogen bond to the urea linker thus stabilizing the conformation. Regarding the amide-styryl deriva-
tives the elongation of the amide linker seemed to be most decisive for the observed increase in activity.
The most promising candidate in the whole library possess an amide-ether linker and an ortho-nitro sub-
stituent in the hydrophobic part. This compound inhibites P-gp slightly less than tariquidar and can serve
as a lead structure for new potent P-gp modulators.
Received 2 December 2008
Revised 20 January 2009
Accepted 23 January 2009
Available online 5 February 2009
Keywords:
Multidrug resistance modulators
P-Glycoprotein
Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction
use the energy of ATP hydrolysis to transport a wide variety of sub-
strates out of cells against the concentration gradient. By actively
The resistance of tumour cells against cytotoxic drugs is a sig-
nificant limitation to successful chemotherapeutic treatment of
cancer. Often the drug efflux by transport proteins is an underlying
mechanism for multidrug resistance (MDR). In MDR tumour cells,
various members of the ATP-binding cassette (ABC) family of
transport proteins can simultaneously be overexpressed: P-glyco-
protein (P-gp, ABCB1), the breast cancer-resistance protein (BCRP,
ABCG2) and the multidrug resistance-associated protein 1 (MRP1,
ABCC family).1 A common feature of these transporters is that they
effluxing substrates their intracellular concentrations are de-
creased thus leading to failure of chemotherapy.
Among the 49 identified human ABC-transporters the most
intensively studied is P-gp. P-gp has the ability to transport a wide
variety of structurally unrelated compounds out of the cells.2 The
protein is associated with poor bioavailability and fast drug elimi-
nation by influencing the drugs’ pharmacokinetics.3
A most striking property of P-gp is the structural diversity of its
substrates, among them many cytotoxic anticancer drugs like
anthracyclines (doxorubicin, daunorubicin), taxanes (paclitaxel,
docetaxel), podophyllotoxin derivatives (etoposid, teniposid), Vin-
ca alkaloids (vinblastine, vincristine) and derivatives from Campt-
Abbreviations: MDR, multidrug resistance; ABC, ATP-binding-cassette; P-gp,
P-glycoprotein; BCRP, breast cancer-resistance protein; MRP, multidrug-resistance
associated protein.
* Corresponding author. Tel.: +49 228 735213; fax: +49 228 737929.
E-mail address: mwiese@uni-bonn.de (M. Wiese).
hoteca acuminata (camptothecin, topotecan, irinotecan).
A
commonly accepted precondition is that P-gp substrates are
amphipathic with a molecular mass between 400 and 1900 Da.
In most cases anionic compounds do not interact with P-gp.4,5
These authors contributed equally.
0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.01.072

W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
2525
Since the first report of MDR overcoming by the calcium chan-
nel blocker verapamil6 a lot of effort has been invested in finding
potent and specific P-gp inhibitors, called also MDR modulators.
The 1st generation modulators (verapamil, cyclosporin A) are
drugs which have been developed for treatment of other diseases.
The use of these substances was limited due to their low efficacy
and the required high doses associated with high toxicity. The sec-
ond generation modulators (dexverapamil, PSC 833) possessed
higher efficacy and lower toxicity, but when applied in patients
serious drug–drug interactions have been observed due to the fact
that both, the cytostatics and 2nd generations modulators are sub-
strates of cytochrome P450 3A. In contrast to the 1st and 2nd gen-
eration modulators the 3rd generation modulators are not
structurally related to or derived from existing drugs. They possess
high efficacy, low toxicity and increased selectivity. Representa-
tives of the 3rd generation modulators which are still under inves-
tigation are: tariquidar (XR9576), elacridar (GF120918),
zosuquidar (LY335979), laniquidar (R1010933).7 In early trials
many of them showed promising activity; nevertheless these mod-
ulators failed later on due to observed toxicities or low survival
advantage.8 Thus to date, the search for new non-toxic, potent
modulators lacking pharmacokinetic interactions is still in process.
XR9576 is one of most potent 3rd generation modulators. It be-
longs to a series of compounds based on the anthranilamide nu-
cleus, called XR compounds, which have been developed and
pharmacologically investigated by Xenova group Ltd.9,10 The mod-
ulator contains a tetrahydroisoquinoline–ethyl-phenylamine sub-
structure which is connected to a highly hydrophobic part via an
amide bond.
an amide group was achieved by reaction of the aniline with isocy-
anato-nitrobenzene. The nitro derivatives (5a, 5b, 7a, 7b, 28, 30a,
30b, 31, 32) were catalytically reduced to the corresponding
amines (6a, 6b, 8a, 8b, 29, 33a, 33b, 34, 35). Two additional linker
variations introduced amide-ether and amide-styryl groups
(Scheme 1).
P-gp modulating activity of the synthesized compounds was
evaluated by the calcein AM assay using P-gp overexpressing, adri-
amycin-resistant A2780adr cells.12,13 As shown in different studies
this cell line is supposed to express high levels of the ABCB1 gene
product P-gp.14–16 This has been confirmed by us applying differ-
ent techniques. Gene expression was confirmed by RT-PCR analy-
17
sis.
The expression of the protein was determined with a
selective FITC-labelled P-gp antibody (17F9 monoclonal P-gp anti-
body) using flow cytometry. Additionally the drug-resistance fac-
tor against the cytotoxic drug doxorubicin was determined,18 and
found to be comparable with resistance factors reported in litera-
ture.15 Further, inhibitory activities of nine structurally unrelated
compounds were measured using MDCK cells transfected with
ABCB1 gene (MDCK-MDR1). The excellent correlation with the cal-
cein AM assay data obtained in A2780adr cells (Fig. 1) clearly
emphasizes that P-gp is the main efflux mechanism influencing
the calcein AM accumulation in these cells.
The inhibitory potencies varied greatly among the synthesized
compounds. To classify the biological activities of the new inhibi-
tors different standard modulators were investigated. We have
chosen verapamil, diltiazem, and cyclosporin A as reference drugs
representing the 1st generation modulators and tariquidar
(XR9576) as a 3rd generation modulator (Table 1). As seen from
the table the activity of the unsubstituted parent compound 4
XR9576 has also been used in our laboratory as a basis for a ra-
tional design of new MDR modulators13. Based on QSAR and
molecular modelling studies of XR compounds important struc-
tural features and pharmacophore elements have been identified11
that have been considered for synthesis of our new P-gp inhibitors.
In contrast to the XR compounds, our compounds have smaller
hydrophobic parts. Thus they are less lipophilic and have lower
molecular weight. This could improve absorption and oral bioavail-
ability. In this paper we describe the synthesis and biological eval-
uation of 39 newly synthesized compounds with MDR modulating
activity. The compounds have been synthesized applying a general
synthesis approach. In the structures the tetrahydroisoquinoline–
ethyl-phenylamine scaffold is combined with the hydrophobic part
using four different types of linkers. The P-gp inhibition activity of
the compounds has been estimated by the functional calcein AM
assay. Structure–activity relationship analysis revealed the struc-
tural elements and modifications that lead to an increased interac-
tion with P-gp. Two promising inhibitors have been identified
which are only three to fourfold less active than tariquidar. These
compounds can serve as lead structures for the future develop-
ment of new potent inhibitors of transport proteins involved in
MDR.
(IC50 = 4.1 lM) is almost equal to that of verapamil (IC50 = 5.2 lM).
Comparing the activity data reported in Table 1 several conclu-
sions about structure–activity relationships in the series can be
drawn.
First, an addition of methoxy groups in positions 6 and 7 of the
tetrahydroisoquinoline substructure (substituent R1) leads to an
increase of the inhibitory potencies compared to the corresponding
unsubstituted derivatives. The positive contribution of the meth-
oxy groups to the biological activities is obvious when comparing
the IC50 values of 5a (IC50 = 5.4
l
l
M) and 5b (IC50 = 14
M), 8a (IC50 = 4.4 M) and 8b
M) Therefore, the tetrahydroisoquinoline ring system
lM), 7a
(IC50 = 1.4
(IC50 = 12
l
l
M) and 7b (IC50 = 5.6
l
supplemented with methoxy groups in position 6 and 7 was cho-
sen as preferred template for the basic substructure; consequently,
this structural element is present in most of the synthesized
substances.
Next, an electron withdrawing nitro-substituent in para posi-
tion of the phenyl ring system (compounds 7a and 7b) influences
the inhibitory potency in a positive manner. Comparing the ni-
tro-substituted derivatives 7a (IC50 = 1.4) and 7b (IC50 = 5.6) with
the corresponding amino-compounds 8a (IC50 = 4.4) 8b (IC50
=
12) a two to three fold decrease in activity is apparent. When alter-
ing the position of the nitro group from para to ortho (substances
7a and 5a), the inhibitory potency decreases considerably, even
below the level of the unsubstituted parent compound 4.
A bromine substitutent at the phenyl ring (compounds 17–19)
has a small positive influence on P-gp inhibitory activity, regard-
less of the position of the bromine varying from ortho over meta
to para. Interestingly, the hydrophilic and electron poor pyridyl
derivative 16 is as active as the phenyl derivative 4.
The inhibitory potency remains unchanged when methoxy
groups in position 3 and 4 of the phenyl ring system (substituent
R2) are present (compare substances 4 and 20). This is in contrast
to anthranilamide MDR modulators where this structural modifi-
cation leads to a twofold increase in activity.9,11
2. Results and discussion
The general strategy used for synthesizing our P-gp related
MDR modulators is shown in Scheme 1. Table 1 reports the struc-
tures and activity data of the 39 compounds.
All compounds possess a tetrahydroisoquinoline–phenylethyl-
amine scaffold (3a or 3b). The latter was obtained in two steps:
(i) alkylation of the corresponding tetrahydroisoquinoline (1a or
1b) with 1-(2-bromoethyl)-4-nitrobenzene leading to the interme-
diate products 2a or 2b followed by catalytic reduction of the nitro
group yielding anilines 3a or 3b. (ii) Reaction of the anilines with
the appropriate carboxylic acid chloride leading to the desired
amide compounds 4–29. Introduction of a urea linker instead of

2526
W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
R1
R1
Br
R1
R1
a
NH
N
NO2
NO2
1a: R1 = OCH3
1b: R1 = H
2a: R1 = OCH3
2b: R1 = H
b
R1
R1
R1
R1
c
N
N
O
NH2
N
R2
3a: R1 = OCH3
3b: R1 = H
4, 5a, 5b, 7a, 7b, 9-28
H
b
6a, 6b, 8a, 8b, 29
d
b
R1
R1
R1
R1
N
N
O
O
NO2
NH2
N
H
N
N
H
N
H
H
30a, 31-32: R1 = OCH3
30b: R1 = H
33a, 34-35: R1 = OCH3
33b: R1 = H
Scheme 1. Synthesis of tetrahydroisoquinoline–phenylethylamine based P-gp inhibitors with different linkers to the hydrophobic substituent R2. Reagents and conditions:
(a) K2CO3, CH3CN, reflux, 24 h, (b) H2, Pd/C, 4 bar, rt, 24 h, (c) R–COCl, THF, Et3N, rt, 48 h, (d) R–NCO, THF, Et3N, reflux, 6 h.
Extending the monocyclic to a bicyclic substituent the inhibi-
tory potency is enhanced. Comparison of the IC50 values of the
sponding amide derivatives 5a and 5b show less inhibitory poten-
cies than the parent compound 4. Again the reduction of the nitro
monocyclic derivatives 4 (IC50 = 4.1
with their corresponding bicyclic analogues 15 (IC50 = 0.63
and 9a (IC50 = 0.57
l
M) and 16 (IC50 = 4.8
l
l
M)
M)
group to an amino group decreases activity (33a: IC50 = 21
lM and
33b: IC50 = 19 M)), and this activity decrease is much more pro-
l
l
M) illustrates that this structural modification
nounced in the urea series and relatively independent of the posi-
yields more effective inhibitors. Interestingly, the orientation of the
bicyclic ring system is important for the biological activity as
the IC50 values of the two different naphthyl derivatives indicate.
tion of the amino group. Also the para-amino compound 35
(IC50 = 31
l
M) is 20 times less active than the nitro-analogue (com-
M).
pound 32, IC50 = 1.5
l
The 2-naphthyl derivative (15) is two fold more active (IC50
=
Extension of the amide linker by a methylenoxy group (27–29)
proved to be an interesting and promising strategy for the design
of new P-gp inhibitors. Among the three different amide-ether
derivatives (27–29) compound 28 showed a superior inhibitory
0.63 M) than the 1-naphthyl derivative (14) (IC50 = 1.4 M). The
l
l
decisive role of the orientation of the bicyclic ring system is even
more obvious when comparing the IC50 values of the quinoline
derivatives 9a and 11.
activity with an IC50 of 0.22 lM, highlighting this substance as
Among the investigated amide derivatives 3-quinolinyl and 2-
quinoxalinyl derivatives are the most active compounds in accor-
dance with the XR-compounds. Thus, the 3-quinolinyl, 2-quinox-
alinyl and 2-naphthyl derivatives are promising lead structures.
They are more than ten times more effective than verapamil and
only four to six times less potent than tariquidar.
the most active compound in our modulator library. Interestingly,
the precondition for an enhanced interaction of substance 28 with
P-gp is due to the nitro substituent in ortho-position. If the nitro
group is absent or reduced to amino, the inhibitory potency consid-
erably decreases. The amino derivative 29 is more than tenfold less
active (IC50 = 3.4
lM) than the most effective modulator 28.
To investigate the role of the linker between the tetrahydroiso-
quinoline–phenylethylamine substructure and the hydrophobic
part for the inhibitory potency the amide linker was replaced by
different linker types.
Eight derivatives with a urea linker have been synthesized.
Among them compound 30a with an ortho-nitro group has the
Comparison of the biological activity of the amide derivative 4
(IC50 = 4.1
(IC50 = 3.1
l
M) with its corresponding amide-ether analogue 27
lM) suggests that the length of the linker seems not to
be decisive for the inhibitory potency. In this context the styryl
derivatives (23–26) differ as elongation of the amide linker is re-
lated to a general increase of activity. The styryl derivative 23
lowest IC50 value (IC50 = 0.33
l
M, Table 1). This substance is among
(IC50 = 1.4
amide counterpart 4 (IC50 = 4.1
in para position (compound 25) leads to a further twofold increase
in activity (IC50 = 0.67 M).
When comparing the influence of an ortho-nitro group on the
activity in the four series substantial difference becomes
apparent. While in case of the amide and styryl linkers activities
l
M) is approximately three times more potent than its
the most active ones found in our library and is about 10-fold more
potent than verapamil. As in the series of amide linked derivatives
the removal of the two methoxy groups at the tetrahydroisoquin-
oline moiety leads to an approximately twofold decrease in activity
(compound 30b with IC50 of 0.77
ortho-nitro group on activity the linker type is decisive. The corre-
lM). Adding a chlorine substituent
l
lM). Regarding the effect of an
a

W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
2527
Table 1
XR9576
Structures of the compounds and their biological activity in the calcein AM assay.
R1
WK-X-34
XR9577
6
5
4
3
N
R1
O
WK-X-50
cyclosporin A
R2
N
H
X
Compound
R1
R2
X
IC50 SD (lM)
4
5a
5b
6a
6b
7a
7b
8a
8b
9°
9b
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30a
OCH3 Phenyl
OCH3 2-Nitrophenyl
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
4.1 1.2
5.4 0.7
verapamil
imatinib
vinblastine
H
2-Nitrophenyl
OCH3 2-Aminophenyl
2-Aminophenyl
OCH3 4-Nitrophenyl
4-Nitrophenyl
OCH3 4-Aminophenyl
4-Aminophenyl
OCH3 3-Quinolinyl
3-Quinolinyl
OCH3 2-Quinolinyl
OCH3 4-Quinolinyl
OCH3 6-Quinolinyl
OCH3 2-Quinoxalinyl
OCH3 1-Naphthyl
14
3
8.5 1.6
9.9 4.7
1.4 0.5
5.6 2.7
4.4 2.8
H
diltiazem
H
4
5
6
7
H
12
3
pIC (A2780adr)
50
0.57 0.18
0.43 0.11
0.85 0.36
4.7 0.7
0.65 0.11
0.47 0.05
1.4 0.4
0.63 0.18
4.8 1.5
3.3 2.3
H
Figure 1. Scatterplot of the pIC50 values of nine structurally different compounds
determined in the calcein AM assay using A2780adr and MDCK-MDR1 cells,
respectively. Data shown are average SD of at least three independent experi-
ments. The squared correlation coefficient is 0.98.
OCH3 2-Naphthyl
OCH3 3-Pyridyl
OCH3 2-Bromophenyl
OCH3 3-Bromophenyl
OCH3 4-Bromophenyl
OCH3 3,4-Dimethoxyphenyl
OCH3 4,5-Dimethoxy-2-nitrophenyl
OCH3 3,4-Methylendioxyphenyl
OCH3 Phenyl
OCH3 2-Nitrophenyl
OCH3 4-Chlorophenyl
OCH3 4,5-Dimethoxy-2-nitrophenyl –CH@CH– 1.5 0.4
OCH3 Phenyl
100
80
60
40
20
0
1.8 0.2
2.4 1.1
4.2 2.1
13
2
2.1 0.5
–CH@CH– 1.4 0.4
–CH@CH– 1.1 0.1
–CH@CH– 0.67 0.09
–CH2–O–
–CH2–O–
–CH2–O–
–NH–
–NH–
–NH–
–NH–
–NH–
–NH–
–NH–
3.1 0.1
0.22 0.03
3.4 1.0
0.33 0.07
0.77 0.15
0.67 0.20
1.5 0.2
OCH3 2-Nitrophenyl
OCH3 2-Aminophenyl
OCH3 2-Nitrophenyl
30b
31
32
33a
33b
34
35
H
2-Nitrophenyl
OCH3 3-Nitrophenyl
OCH3 4-Nitrophenyl
OCH3 2-Aminophenyl
-10
-9
-8
-7
-6
-5
Compound 28, log [M]
21
19
14
3
8
3
H
2-Aminophenyl
Figure 2. Concentration–response curve generated for compound 28 in A2780adr
cells in the calcein AM assay: IC50 = 0.22 0.03 M (pIC50 = 6.65 0.05). Data shown
are average SD from one typical experiment with four replicates out of a series of
OCH3 3-Aminophenyl
OCH3 4-Aminophenyl
l
–NH–
31 17
0.078 0.013
49 20
Tariquidar
Diltiazem
Verapamil
Cyclosporin A
six independent experiments.
5.2 2.0
1.4 0.3
unsubstituted compound 27 (purple) can adopt the same orien-
tation of R2 as the nitro substituted derivative 28. But this con-
formation is less favored among the sampled conformations,
probably because of the missing stabilizing hydrogen bond. The
alignment of 28 with two other active pairs of less active com-
pounds (the amides 4, 5a and the styryl derivatives 23, 24) is
shown in Figure 4. None of those compounds can adopt a planar
conformation similar to the most active compound 28. Thus it
can be speculated that the high activities of compounds 28
and 30a can be explained by the position and planar orientation
of the terminal aromatic substituent. Additionally, the electron
withdrawing properties of substituents like the nitro group could
positively contribute to the effect.
In summary, a number of new molecules have been synthesized
on the basis of selected molecular features and subsequently tested
as P-gp inhibitors. In the synthesized series a number of lead
compounds were found that show P-gp inhibitory potency at
submicromolar concentrations. This is a promising result com-
pared to the activities of the standard inhibitors verapamil and
cyclosporin A determined under the same conditions. The best
compounds are about 10 times more active than verapamil and
four times more active than cyclosporin A.
Values are means with standard deviations from a minimum of three experiments.
of the unsubstituted and ortho-nitro derivatives are comparable,
the nitro derivatives with urea (30a) and ether linker (28) are
the most potent compounds. In order to elucidate the effect
of the nitro group we performed a conformational analysis of
the nitro derivatives and the unsubstituted compounds with
the different linkers. To sample the conformational space 200
cycles of simulated annealing were performed for each com-
pound and the obtained conformations of the molecules were
then sorted by potential energy and compared. Besides the force
field method the described conformers were subsequently opti-
mized by the semiempirical method PM6 with MOPAC2007.
The final conformations yielded 290 only marginal differences
in the geometries. The most active compounds 28 and 30a,
although attached by different linkers in position X, share the
same position of the hydrophobic part R2 (Fig. 3). The amide
group in 28 is flipped allowing the nitro group to form a hydro-
gen bond to the amide hydrogen. Also in case of the urea linker
the planar conformation is stabilized by a hydrogen bond be-
tween the nitro group and the urea nitrogen. The ether attached

2528
W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
Figure 3. Superposition of the lowest energy conformations of the most active compounds 28 (purple) and 30a (grey), showing that both compounds share the same position
of the hydrophobic part in position R2.
Figure 4. Superposition of lowest energy conformations of the most active compound 28 (purple) with the less active compounds 4, 5a and 23, 24; neither of these
compounds can adopt the planar conformation of 28.
Among the studied compounds the amide-ether derivative 28
possesses the strongest inhibitory potency against P-gp. Compar-
ing its structure to that of tariquidar, compound 28 contains an
elongated amide linker between the tetrahydroisoquinoline–phen-
ylethylamine substructure and the hydrophobic part, but lacks the
second amide linker present in the structure of tariquidar. In Figure
2 a typical concentration–response curve for compound 28 ob-
tained with the calcein AM assay is shown (IC50 = 0.22 lM). This
substance inhibits P-gp 20 times stronger than the 1st generation
modulator verapamil and only three times less than the most po-
tent modulator tariquidar (IC50 = 0.078 lM). Therefore, substance
3.2. General procedure for the synthesis of the
nitrophenylethylamines
3.2.1. 6,7-Dimethoxy-2-[2-(4-nitrophenyl)ethyl]-1,2,3,4-tetra-
hydroisoquinoline (2a)
A
mixture of 1-(2-bromoethyl)-4-nitrobenzene (10 mmol,
2.42 g), 1,2,3,4-tetrahydro-6,7-dimethoxyisoquinoline hydrochlo-
ride (10.5 mmol, 2.41 g) and K2CO3 (25 mmol, 3.48 g) in 50 ml ace-
tonitril was heated under reflux for 12 h. The solvent was
evaporated, the residue taken up in 100 ml water and extracted
three times with 75 ml DCM. The solution was dried over MgSO4,
filtered and concentrated. The product was recrystallized from eth-
anol to give yellow crystals: yield 2.6 g (73.7 %), mp 118 °C. 1H
NMR (DMSO-d6): d = 2.68 (m, 4H, 2 Â CH2), 2.72 (t, J = 7.6 Hz, 2H,
CH2), 2.96 (t, J = 7.3 Hz, 2H, CH2), 3.52 (s, 2H, CH2), 3.68 (s, 3H,
OCH3), 3.68 (s, 3H, OCH3), 6.60 (s, 1H, ArH), 6.63 (s, 1H, ArH),
7.54 (d, J = 8.8 Hz, 2H, 2 Â ArH), 8.13 (d, J = 8.8 Hz, 2H, 2 Â ArH).
13C NMR (DMSO-d6): d = 28.4 (CH2), 32.7 (CH2), 50.5 (CH2), 55.1
(CH2), 55.6 (OCH3) 55.7 (OCH3), 58.5 (CH2), 110.2 (Ar-CH), 112.0
(Ar-CH), 123.3 (Ar-CH), 123.4 (2 Â Ar-CH), 126.0 (Ar-C), 126.7
(Ar-C), 130.1 (2 Â Ar-CH), 146.0 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C),
149.4 (Ar-C). Anal. Calcd for C19H22N2O4: C, 66.65; H, 6.48; N,
8.18. Found: C, 66.38; H, 6.46; N, 7.99.
28 could be classified as a promising candidate in our modulator
library and will be used as a lead structure for synthesizing new
effective P-gp modulators in future.
3. Experimental
3.1. General methods
Melting points were measured in open capillary tubes on a Gal-
lenkamp melting point apparatus and are uncorrected. Spectral
data were obtained on the following instruments: IR, Perkin–Elmer
Paragon 1000; mass spectra, Kratos Concept 1-H, A.E.I.; 1H NMR,
Bruker Advance 500 (500 MHz); 13C NMR, Bruker Advance 500
(125.8 MHz); chemical shifts are expressed in d value (ppm) with
using tetramethylsilan as an internal standard; multiplicity of res-
onance peaks is indicated as singlet (s), doublet (d), triplet (t),
quartet (q) and multiplet (m). The marked protons (*) are exchan-
gable with D2O. The 13C signals were assigned with the aid of dis-
tortionless enhancement by polarization transfer (DEPT) and
twodimensional experiments (H–H COSY, C–H COSY), and classifi-
cation of carbon atoms are indicated by CH3 (primary), CH2 (sec-
ondary), CH or Ar-CH (tertiary), or Ar-C (quarternary), the J
values are in Hertz. Elemental analyses were performed on a Vario
EL of Elementar. Found values were all within 0.4% of the theoret-
ical values except when indicated.
3.2.2. 2-[2-(4-Nitrophenyl)ethyl]-1,2,3,4-tetrahydroisoquinoline
(2b)
The compound was prepared as described for the synthesis of
(2a) starting from 1,2,3,4-tetrahydroisoquinoline (24.8 mmol,
3.3 g), 1-(2-bromoethyl)-4-nitrobenzene (20.6 mmol, 5.0 g) and
K2CO3 (50 mmol, 6.96 g): yield 4.55 g (78.3%) of yellow crystals,
mp 103 °C. 1H NMR (DMSO-d6): d = 2.71–2.79 (m, 6H, 3 Â CH2),
2.97 (t, J = 7.3 Hz, 2H, CH2), 3.61 (s, 2H, CH2), 7.01 (m, 1H, ArH),
7.08 (m, 3H, 3 Â ArH), 7.55 (d, J = 8.8 Hz, 2H, 2 Â ArH), 8.13 (d,
J = 8.8 Hz, 2H, 2 Â ArH). 13C NMR (DMSO-d6): d = 28.8 (CH2), 32.6
(CH2), 50.3 (CH2), 55.5 (CH2), 58.5 (CH2), 123.4 (2 Â Ar-CH), 125.6
(Ar-CH), 126.0 (Ar-CH), 126.5 (Ar-CH), 128.5 (Ar-CH), 130.1

W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
2529
(2 Â Ar-CH), 134.3 (Ar-C), 135.0 (Ar-C), 146.0 (Ar-C), 149.4 (Ar-C).
Anal. Calcd for C17H18N2O2: C, 72.32; H, 6.43; N, 9.92. Found: C,
72.21; H, 6.41; N, 9.69.
(78%) as a pale yellow solid, mp 184 °C. 1H NMR (DMSO-d6, 500
MHz): d = 2.67 (m, 6H, 3 Â CH2), 2.80 (t, 2H, CH2), 3.53 (s, 2H,
CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.63 (s, 1H, ArH),
6.65 (s, 1H, ArH), 7.23 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.56 (d, J = 8.5
Hz, 2H, 2 Â ArH), 7.74 (m, 2H, 2 Â ArH), 7.85 (dt, J = 8.4 Hz, 1H,
ArH), 8.12 (dd, J = 8.5 Hz, 1H, ArH), 10.55 (s, 1H*, NH). 13C NMR
(DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2), 55.2
(CH2), 55.6 (OCH3) 55.7 (OCH3), 59.7 (CH2), 110.2 (Ar-CH), 112.0
(Ar-CH), 119.8 (2 Â Ar-CH), 124.3 (Ar-CH), 126.1 (Ar-C), 126.8
(Ar-C), 129.1 (2 Â Ar-CH), 129.4 (Ar-CH), 131.0 (Ar-CH), 132.9
(Ar-C), 134.1 (Ar-CH), 136.3 (Ar-C), 136.9 (Ar-C), 146.7 (Ar-C),
147.1 (Ar-C), 147.3 (Ar-C), 164.0 (CO). Anal. Calcd for
C26H27N3O5Á0.2H2O: C, 67.14; H, 5.94; N, 9.03. Found: C, 67.23;
H, 5.91; N, 8.81.
3.3. General procedure for the hydrogenation
3.3.1. 4-[2-(6,7-Dimethoxy-3,4-dihydro-1H-isoquinolin-2-yl)-
ethyl]phenylamine (3a)
10 mmol (3.42 g) of compound (2a) was dissolved in 200 ml
ethanol and was hydrogenated at room temperature under 4 bar
pressure in a Paar-apparatus in the presence of Pd/C as catalyst.
After hydrogen absorption was completed, the catalyst was filtered
off and the solution was concentrated to give 4-[2-(6,7-dimethoxy-
3,4-dihydro-1H-isoquinoline-2-yl)-ethyl]-phenylamine as a white
solid, yield 2.41 g (87%), mp 129 °C. 1H NMR (DMSO-d6): d = 2.56
(m, 2H, CH2), 2.62 (m, 4H, 2 Â CH2), 2.69 (t, J = 5.7 Hz, 2H, CH2),
3.50 (s, 2H, CH2), 3.68 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 4.78 (s,
2H*, NH2), 6.48 (d, J = 8.5 Hz, 2H, 2 Â ArH), 6.62 (s, 1H, ArH), 6.64
(s, 1H, ArH), 6.87 (d, J = 8.5 Hz, 2H, 2 Â ArH). 13C NMR (DMSO-
d6): d = 28.5 (CH2), 32.4 (CH2), 50.8 (CH2), 55.3 (CH2), 55.6 (OCH3)
55.7 (OCH3), 60.4 (CH2), 110.2 (Ar-CH), 112.0 (Ar-CH), 114.1
(2 Â Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C), 127.5 (Ar-C), 129.1
(2 Â Ar-CH), 146.7 (Ar-C), 147.0 (Ar-C), 147.3 (Ar-C). Anal. Calcd
for C19H24N2O2: C, 73.05; H, 7.74; N, 8.97. Found: C, 72.69; H,
7.68; N, 8.78.
3.4.3. N-{4-[2-(3,4-Dihydro-1H-isoquinolin-2-yl)ethyl]phenyl}-
2-nitrobenzamide (5b)
Following the procedure described for compound 4, starting
from 3b and 2-nitrobenzoyl chloride, compound 5b was obtained
(82%) as a white solid, mp 166 °C.
1H NMR (DMSO-d6, 500 MHz): d = 2.70 (m, 4H, 2 Â CH2), 2.81
(m, 4H, 2 Â CH2), 3.63 (s, 2H, CH2), 7.07 (m, 4H, 4 Â ArH), 7.24 (d,
J = 8.4 Hz, 2H, 2 Â ArH), 7.57 (d, J = 8.4 Hz, 2H, 2 Â ArH), 7.74 (m,
2H, 2 Â ArH), 7.85 (d, J = 7.8 Hz, 1H, ArH), 8.12 (d, J = 7.6 Hz, 1H,
ArH), 10.56 (s, 1H*, NH).
13C NMR (DMSO-d6, 125 MHz): d = 28.8 (CH2), 32.5 (CH2), 50.5
(CH2), 55.5 (CH2), 59.7 (CH2), 119.8 (2 Â Ar-CH), 124.3 (Ar-CH),
125.6 (Ar-CH), 126.0 (Ar-CH), 126.5 (Ar-CH), 128.5 (Ar-CH), 129.1
(2 Â Ar-CH), 129.4 (Ar-CH), 131.0 (Ar-CH), 132.9 (Ar-C), 134.1
(Ar-CH), 134.3 (Ar-C), 135.0 (Ar-C), 136.3 (Ar-C), 136.9 (Ar-C),
146.7 (Ar-C), 164.0 (CO). Anal. Calcd for C24H23N3O3: C, 71.80; H,
5.77; N, 10.47. Found: C, 71.46; H, 5.78; N, 10.40.
3.3.2. 4-[2-(3,4-Dihydro-1H-isoquinolin-2-yl)ethyl]phenylamine
(3b)
Following the procedure described for compound (3a), starting
from (2b) (2.82 g, 10 mmol), compound (3b) (2.32 g, 92%) was ob-
tained as a white solid, mp 90 °C. 1H NMR (DMSO-d6): d = 2.58 (m,
2H, CH2), 2.64 (m, 2H, CH2), 2.68 (t, J = 5.8 Hz, 2H, CH2), 2.78 (t,
J = 5.8 Hz, 2H, CH2), 3.58 (s, 2H, CH2), 4.78 (s, 2H*, NH2), 6.48 (d,
J = 8.4 Hz, 2H, 2 Â ArH), 6.88 (d, J = 8.4 Hz, 2H, 2 Â ArH), 7.02–
7.10 (m, 4H, 4 Â ArH). 13C NMR (DMSO-d6): d = 28.9 (CH2), 32.4
(CH2), 50.6 (CH2), 55.6 (CH2), 60.4 (CH2), 114.1 (2 Â Ar-CH), 125.5
(Ar-CH), 126.0 (Ar-CH), 126.5 (Ar-CH), 127.4 (Ar-C), 128.5 (Ar-
CH), 129.1 (2 Â Ar-CH), 134.3 (Ar-C), 135.1 (Ar-C), 146.7 (Ar-C).
Anal. Calcd for C17H20N2Á0.2H2O: C, 79.77; H, 8.03; N, 10.94. Found:
C, 79.71; H, 8.05; N, 10.83.
3.4.4. 2-Amino-N-{4-[2-(6,7-dimethoxy-3,4-dihydro-1H-
isoquinolin-2-yl)ethyl]phenyl}benzamide (6a)
Following the procedure described for compound 3a, starting
from 5a (1.58 g, 3.4 mmol), compound 6a (89%) was obtained as
a white solid, mp 150 °C.
1H NMR (DMSO-d6, 500 MHz): d = 2.67 (m, 6H, 3 Â CH2), 2.78 (t,
2H, CH2), 3.53 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3),
6.28 (s, 2H*, NH2), 6.57 (d, J = 7.9 Hz, 1H, ArH), 6.62 (s, 1H, ArH),
6.64 (s, 1H, ArH), 6.73 (dd, J = 8.2 Hz, 1H, ArH), 7.18 (m, 3H,
3 Â ArH), 7.59 (m, 3H, 3 Â ArH), 9.89 (s, 1H*, NH).
3.4. General procedure for the synthesis of the amides
13C NMR (DMSO-d6, 125 MHz): d = 28.5 (CH2), 32.6 (CH2), 50.7
(CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.7 (CH2), 110.2 (Ar-
CH), 112.0 (Ar-CH), 114.8 (Ar-CH), 115.6 (Ar-C), 116.5 (Ar-CH),
120.7 (2 Â Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C), 128.8 (Ar-CH),
128.8 (2 Â Ar-CH), 132.1 (Ar-CH), 135.7 (Ar-C), 137.2 (Ar-C),
147.1 (Ar-C), 147.3 (Ar-C), 149.8 (Ar-C), 167.8 (CO). Anal. Calcd
for C26H29N3O3Á0.2H2O: C, 71.77; H, 6.81; N, 9.66. Found: C,
71.81; H, 6.89; N, 9.48.
3.4.1. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)benzamide (4)
5 mmol benzoylchloride was suspended in dry THF and added
dropwise to a stirred solution of 3a (5 mmol) and triethylamine
(6 mmol) in dry THF at room temperature. The solution was stirred
for 12 h. After the reaction was completed the solvent was re-
moved in vacuum and the residue was treated with water and ex-
tracted three times with EtOAc. The organic phase was washed
with dilute NaOH (1 N) and water, dried with MgSO4, and concen-
trated. After recrystallization from DCM/n-hexane the compound
was yielded as a white solid (66%), mp 233 °C. 1H NMR (DMSO-
d6, 500 MHz): d = 2.94–3.19 (m, 8H, 4 Â CH2), 3.72 (s, 3H, OCH3),
3.73 (s, 3H, OCH3), 4.10 (s, 2H, CH2), 6.73 (s, 1H, ArH), 6.76 (s,
1H, ArH), 7.26 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.52 (m, 2H, 2 Â ArH),
7.57 (m, 1H, ArH), 7.74 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.96 (m, 2H,
2 Â ArH), 10.24 (s, 1H*, NH). Anal. Calcd for C26H28N2O3Á0.5H2O:
C, 73.39; H, 6.87; N, 6.58. Found: C, 73.30; H, 6.76; N, 6.30.
3.4.5. 2-Amino-N-{4-[2-(3,4-dihydro-1H-isoquinolin-2-yl)ethyl]
phenyl}benzamide (6b)
Following the procedure described for compound 3a, starting
from 5b (1.05 g, 2.5 mmol), compound 6b (75%) was obtained as
a white solid, mp 104 °C.
1H NMR (DMSO-d6, 500 MHz): d = 2.69 (m, 6H, 3 Â CH2), 2.80 (t,
2H, CH2), 3.62 (s, 2H, CH2), 6.28 (s, 2H*, NH2), 6.57 (m, 1H, ArH),
6.73 (dd, J = 8.4 Hz, 1H, ArH), 7.04 (m, 1H, ArH), 7.09 (m, 3H,
3 Â ArH), 7.18 (m, 3H, 3 Â ArH), 7.60 (m, 3H, 3 Â ArH), 9.89 (s,
1H*, NH).
13C NMR (DMSO-d6, 125 MHz): d = 28.9 (CH2), 32.5 (CH2), 50.6
(CH2), 55.6 (CH2), 59.7 (CH2), 114.8 (Ar-CH), 115.6 (Ar-C), 116.5
(Ar-CH), 120.7 (2 Â Ar-CH), 125.5 (Ar-CH), 126.0 (Ar-CH), 126.5
(Ar-CH), 128.5 (Ar-CH), 128.7 (Ar-CH), 128.8 (2 Â Ar-CH), 132.1
3.4.2. N-{4-[2-(6,7-Dimethoxy-3,4-dihydro-1H-isoquinolin-2-
yl)ethyl]phenyl}-2-nitrobenzamide (5a)
Following the procedure described for compound 4, starting
from 3a and 2-nitrobenzoyl chloride, compound 5a was obtained

2530
W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
(Ar-CH), 134.3 (Ar-C), 135.1 (Ar-C), 135.6 (Ar-C), 137.2 (Ar-C),
149.8 (Ar-C), 167.8 (CO). Anal. Calcd for C24H25N3OÁ1.0C2H5OH: C,
74.79; H, 7.48; N, 10.06. Found: C, 74.41; H, 7.44; N, 9.79.
13C NMR (CDCl3, 125 MHz): d = 29.0 (CH2), 33.3 (CH2), 50.9
(CH2), 56.0 (CH2), 60.2 (CH2), 114.2 (2 Â Ar-CH), 120.3 (2 Â Ar-
CH), 124.2 (Ar-C), 125.6 (Ar-CH), 126.1 (Ar-CH), 126.6 (Ar-CH),
128.6 (Ar-CH), 128.8 (2 Â Ar-CH), 129.2 (2 Â Ar-CH), 134.2 (Ar-C),
134.6 (Ar-C), 136.1 (Ar-C), 136.4 (Ar-C), 149.9 (Ar-C), 165.4 (CO).
Anal. Calcd for C24H25N3OÁ0.8H2O: C, 74.95; H, 6.93; N, 10.93.
Found: C, 74.77; H, 7.06; N, 10.24.
3.4.6. N-{4-[2-(6,7-Dimethoxy-3,4-dihydro-1H-isoquinolin-2-
yl)ethyl]phenyl}-4-nitrobenzamide (7a)
Following the procedure described for compound 4, starting
from 3a and 4-nitrobenzoyl chloride, compound 7a was obtained
(88%) as a yellow solid, mp 134 °C. 1H NMR (CDCl3, 500 MHz):
d = 2.67 (m, 6H, 3 Â CH2), 2.80 (t, 2H, CH2), 3.53 (s, 2H, CH2), 3.69
(s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.62 (s, 1H, ArH), 6.64 (s, 1H,
ArH), 7.25 (d, J = 8.8 Hz, 2H, 2 Â ArH), 7.69 (d, J = 8.6 Hz, 2H,
2 Â ArH), 8.16 (td, J = 8.8 Hz, 2H, 2 Â ArH), 8.35 (td, J = 8.8 Hz,
2H, 2 Â ArH), 10.46 (s, 1H*, NH).
3.4.10. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)quinoline-3-carboxamide (9a)
Following the procedure described for compound 4, starting
from 3a and quinoline-3-carbonyl chloride, compound 9a (75%)
was obtained as a white solid, mp 184 °C. 1H NMR (CDCl3, 500
MHz): d = 2.73–2.79 (m, 4H, 2 Â CH2), 2.83 (t, J = 5.5 Hz, 2H, CH2),
2.89 (q, J = 5.5 Hz, 2H, CH2), 3.63 (s, 2H, CH2), 3.81 (s, 3H, OCH3),
3.81 (s, 3H, OCH3), 6.52 (s, 1H, ArH), 6.58 (s, 1H, ArH), 7.23 (d,
J = 8.5 Hz, 2H, 2 Â ArH), 7.59 (m, 3H, 3 Â ArH), 7.79 (dt, J = 8.2 Hz,
1H, ArH), 7.86 (d, J = 8.2 Hz, 1H, ArH), 8.12 (d, J = 8.5 Hz, 1H,
ArH), 8.19 (s, 1H*, NH), 8.63 (d, J = 2.2 Hz, 1H, ArH), 9.33 (d,
J = 2.2 Hz, 1H, ArH). 13C NMR (CDCl3, 125 MHz): d = 28.7 (CH2),
33.5 (CH2), 51.0 (CH2), 55.7 (CH2), 55.9 (OCH3) 55.9 (OCH3), 60.1
(CH2), 109.5 (Ar-CH), 111.4 (Ar-CH), 120.7 (2 Â Ar-CH), 126.1 (Ar-
C), 126.5 (Ar-CH), 126.7 (Ar-CH), 126.8 (Ar-C), 127.5 (Ar-C), 127.6
(Ar-CH), 129.4 (Ar-CH), 129.4 (2 Â Ar-CH), 131.4 (Ar-CH), 135.6
(Ar-C), 135.7 (Ar-CH), 137.3 (Ar-C), 147.2 (Ar-C), 147.5 (Ar-C),
148.1 (Ar-CH), 149.8 (Ar-C), 163.9 (CO). Anal. Calcd for
C29H29N3O3Á0.2H2O: C, 73.93; H, 6.29; N, 8.92. Found: C, 73.98;
H, 6.21; N, 9.10.
13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7
(CH2), 55.2 (CH2), 55.6 (2 Â OCH3), 59.6 (CH2), 110.2 (Ar-CH),
112.0 (Ar-CH), 120.6 (2 Â Ar-CH), 123.6 (2 Â Ar-CH), 129.0
(2 Â Ar-CH), 129.3 (2 Â Ar-CH), 126.1 (Ar-C), 126.8 (Ar-C), 136.6
(Ar-C), 136.7 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 149.2 (Ar-C),
163.8 (CO). Anal. Calcd for C26H27N3O5 0.5H2O: C, 66.37; H, 6.00;
N, 8.93. Found: C, 66.30; H, 5.94; N, 8.77.
3.4.7. N-{4-[2-(3,4-Dihydro-1H-isoquinolin-2-yl)ethyl]phenyl}-
4-nitrobenzamide (7b)
Following the procedure described for compound 4, starting
from 3b and 4-nitrobenzoyl chloride, compound 7b was obtained
(94%) as a yellow solid, mp 176 °C. 1H NMR (DMSO-d6, 500
MHz): d = 2.69 (m, 4H, 2 Â CH2), 2.80 (m, 4H, 2 Â CH2), 3.62 (s,
2H, CH2), 7.03 (m, 1H, ArH), 7.08 (m, 3H, 3 Â ArH), 7.25 (d, J = 8.7
Hz, 2H, 2 Â ArH), 7.67 (m, 2H, 2 Â ArH), 8.16 (ddd, J = 9 Hz, 2H,
2 Â ArH), 8.35 (ddd, J = 9 Hz, 2H, 2 Â ArH), 10.46 (s, 1H*, NH). 13C
NMR (DMSO-d6, 125 MHz): d = 29.1 (CH2), 32.8 (CH2), 50.9 (CH2),
55.9 (CH2), 60.0 (CH2), 121.2 (2 Â Ar-CH), 124.2 (2 Â Ar-CH),
126.1 (Ar-CH), 126.6 (Ar-CH), 127.0 (Ar-CH), 129.0 (Ar-CH), 129.5
(2 Â Ar-CH), 129.7 (2 Â Ar-CH), 134.7 (Ar-C), 135.3 (Ar-C), 136.8
(Ar-C), 137.2 (Ar-C), 141.2 (Ar-C), 149.7 (Ar-C), 164.6 (CO). Anal.
Calcd for C24H23N3O3Á0.2H2O: C, 71.16; H, 5.82; N, 10.37. Found:
C, 71.14; H, 5.95; N, 10.36.
3.4.11. N-(4-(2-(3,4-Dihydroisoquinolin-2(1H)-yl)ethyl)
phenyl)quinoline-3-carboxamide (9b)
Following the procedure described for compound 4, starting
from 3b and quinoline-3-carbonyl chloride, compound 9b (50%)
was obtained as a white solid, mp 186 °C. 1H NMR (CDCl3, 500
MHz): d = 2.74–2.81 (m, 4H, 2 Â CH2), 2.88–2.93 (m, 4H,
2 Â CH2), 3.71 (s, 2H, CH2), 7.01 (m, 1H, ArH), 7.10 (m, 3H, 3 Â ArH),
7.24 (m, 2H, 2 Â ArH), 7.59 (m, 3H, 3 Â ArH), 7.79 (m, 1H, ArH),
7.87 (d, J = 7.9 Hz, 1H, ArH), 8.14 (d, J = 8.5 Hz, 1H, ArH), 8.16 (s,
1H*, NH), 8.63 (d, J = 2.2 Hz, 1H, ArH), 9.33 (s, 1H, ArH). 13C NMR
(CDCl3, 125 MHz): d = 29.1 (CH2), 33.4 (CH2), 51.0 (CH2), 56.1
(CH2), 60.1 (CH2), 120.7 (2 Â Ar-CH), 125.8 (Ar-CH), 126.1 (Ar-
CH), 126.6 (Ar-CH), 126.8 (Ar-C), 127.5 (Ar-C), 127.7 (Ar-CH),
128.6 (Ar-CH), 128.7 (Ar-CH), 129.4 (Ar-CH), 129.4 (2 Â Ar-CH),
131.4 (Ar-CH), 134.2 (Ar-C), 134.6 (Ar-C), 135.6 (Ar-C), 135.7 (Ar-
CH), 137.3 (Ar-C), 148.0 (Ar-CH), 149.8 (Ar-C), 163.9 (CO). Anal.
Calcd for C27H25N3O: C, 79.58; H, 6.18; N, 10.31. Found: C, 79.37;
H, 6.14; N, 10.25.
3.4.8. 4-Amino-N-(4-(2-(6,7-dimethoxy-3,4-dihydroisoqui-
nolin-2(1H)-yl)ethyl)phenyl)benzamide (8a)
Following the procedure described for compound 3a, starting
from 7a (1.38 g, 3 mmol), compound 8a (89%) was obtained as a
light yellow solid, mp 128 °C. 1H NMR (DMSO-d6, 500 MHz):
d = 2.67 (m, 6H, 3 Â CH2), 2.77 (t, 2H, CH2), 3.53 (s, 2H, CH2), 3.68
(s, 3H, OCH3), 3.69 (s, 3H, OCH3), 5.68 (s, 2H*, NH2), 6.59 (td,
J = 8.5 Hz, 2H, 2 Â ArH), 6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 7.17
(dd, J = 8.5 Hz, 2H, 2 Â ArH), 7.63 (td, J = 8.5 Hz, 2H, 2 Â ArH),
7.69 (td, J = 8.5 Hz, 2H, 2 Â ArH), 9.65 (s, 1H*, NH).
13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7
(CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.8 (CH2), 110.2 (Ar-
CH), 112.0 (Ar-CH), 112.7 (2 Â Ar-CH), 120.3 (2 Â Ar-CH), 126.1
(Ar-C), 126.9 (Ar-C), 128.7 (2 Â Ar-CH), 129.4 (2 Â Ar-CH), 135.1
(Ar-C), 137.8 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 152.2 (Ar-C),
165.3 (CO). Anal. Calcd for C26H29N3O3: C, 72.37; H, 6.77; N, 9.74.
Found: C, 72.14; H, 6.78; N, 9.70.
3.4.12. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)isoquinoline-3-carboxamide (10)
Following the procedure described for compound 4, starting
from 3a and quinoline-2-carbonyl chloride, compound 10 (38%)
was obtained as a white solid, mp 141 °C. 1H NMR (DMSO-d6,
500 MHz): d = 2.67–2.72 (m, 6H, 3 Â CH2), 2.82 (t, 2H, CH2), 3.55
(s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.63 (s, 1H,
ArH), 6.65 (s, 1H, ArH), 7.28 (d, J = 8.2 Hz, 2H, 2 Â ArH), 7.75 (t,
J = 7.4 Hz, 1H, ArH), 7.84 (d, J = 8.2 Hz, 2H, 2 Â ArH), 7.91 (t,
J = 7.6 Hz, 1H, ArH), 8.11 (d, J = 7.9 Hz, 1H, ArH), 8.24 (q, 2H,
2 Â ArH), 8.61 (d, J = 8.5 Hz, 1H, ArH), 10.63 (s, 1H*, NH). 13C
NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2),
55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.6 (CH2), 110.2 (Ar-CH),
112.0 (Ar-CH), 118.8 (Ar-CH), 120.4 (2 Â Ar-CH), 126.1 (Ar-C),
126.8 (Ar-C), 128.2 (Ar-CH), 128.4 (Ar-CH), 129.0 (Ar-C), 129.0
(2 Â Ar-CH), 129.4 (Ar-CH), 130.8 (Ar-CH), 136.3 (Ar-C), 136.4
(Ar-C), 138.3 (Ar-CH), 146.0 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C),
3.4.9. 4-Amino-N-{4-[2-(3,4-dihydro-1H-isoquinolin-2-yl)ethyl]-
phenyl}benzamide (8b)
Following the procedure described for compound 3a, starting
from 7b (0.72 g, 1.8 mmol), compound 8b (83%) was obtained as a
white solid, mp 152 °C.1H NMR (CDCl3, 500 MHz): d = 2.76 (m, 4H,
2 Â CH2), 2.90 (m, 4H, 2 Â CH2), 3.71 (s, 2H, CH2), 4.01 (s, 2H*,
NH2), 6.65 (d, J = 8.7 Hz, 2H, 2 Â ArH), 7.01 (m, 1H, ArH), 7.10 (m,
3H, 3 Â ArH), 7.19 (d, J = 8.4 Hz, 2H, 2 Â ArH), 7.51 (d, J = 8.4 Hz,
2H, 2 Â ArH), 7.67 (d, J = 8.7 Hz, 2H, 2 Â ArH), 7.74 (s, 1H*, NH).

W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
2531
150.3 (Ar-C), 162.6 (CO). Anal. Calcd for C29H29N3O3Á0.5H2O: C,
tained as a white solid, mp 151 °C. 1H NMR (DMSO-d6, 500
MHz): d = 2.70–2.88 (m, 8H, 4 Â CH2), 3.64 (s, 2H, CH2), 3.70 (s,
3H, OCH3), 3.70 (s, 3H, OCH3), 6.65 (s, 1H, ArH), 6.67 (s, 1H, ArH),
7.25 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.58 (m, 3H, 3 Â ArH), 7.72 (d,
J = 7.9 Hz, 3H, 3 Â ArH), 8.01 (m, 1H, ArH), 8.06 (d, J = 8.2 Hz, 1H,
ArH), 8.16 (t, J = 4.9 Hz, 1H, ArH), 10.48 (s, 1H*, NH). 13C NMR
(DMSO-d6, 125 MHz): d = 28.0 (CH2), 32.2 (CH2), 50.8 (CH2), 55.1
(CH2), 55.7 (OCH3) 55.7 (OCH3), 59.3 (CH2), 110.2 (Ar-CH), 112.0
(Ar-CH), 120.0 (2 Â Ar-CH), 125.2 (Ar-CH), 125.3 (Ar-CH), 125.5
(Ar-CH), 125.8 (Ar-C), 126.5 (Ar-CH), 127.1 (Ar-CH), 128.4 (Ar-
CH), 129.0 (2 Â Ar-CH), 129.8 (Ar-C), 130.1 (Ar-CH), 133.3 (Ar-C),
135.0 (Ar-C), 135.6 (Ar-C), 137.5 (Ar-C), 147.2 (Ar-C), 147.5 (Ar-
C), 167.3 (CO). Anal. Calcd for C30H30N2O3Á0.2H2O: C, 76.64; H,
6.52; N, 5.96. Found: C, 76.35; H, 6.65; N, 5.87.
73.09; H, 6.35; N, 8.82. Found: C, 73.00; H, 6.05; N, 9.20.
3.4.13. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)quinoline-4-carboxamide (11)
Following the procedure described for compound 4, starting
from 3a and quinoline-4-carbonyl chloride, compound 11 (55%)
was obtained as a white solid, mp 199 °C. 1H NMR (DMSO-d6,
500 MHz): d = 2.66–2.71 (m, 6H, 3 Â CH2), 2.82 (t, J = 7.5 Hz, 2H,
CH2), 3.54 (s, 2H, CH2), 3.68 (s, 3H, OCH3), 3.69 (s, 3H, OCH3),
6.63 (s, 1H, ArH), 6.65 (s, 1H, ArH), 7.26 (d, J = 8.6 Hz, 2H, 2 Â ArH),
7.67 (m, 4H, 4 Â ArH), 7.82 (dt, J = 5.5 Hz, 1H, ArH), 8.13 (d, J = 8.7
Hz, 2H, 2 Â ArH), 9.02 (d, J = 4.4 Hz, 1H, ArH), 10.66 (s, 1H*, NH). 13
C
NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2),
55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.6 (CH2), 110.2 (Ar-CH),
112.0 (Ar-CH), 119.3 (Ar-CH), 120.1 (2 Â Ar-CH), 124.2 (Ar-C),
125.4 (Ar-CH), 126.1 (Ar-C), 126.8 (Ar-C), 127.7 (Ar-CH), 129.1
(2 Â Ar-CH), 129.6 (Ar-C), 130.0 (Ar-CH), 136.6 (Ar-C), 136.8 (Ar-
C), 142.2 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 148.1 (Ar-C), 150.4
(Ar-CH), 165.2 (CO). Anal. Calcd for C29H29N3O3Á0.8H2O: C, 72.27;
H, 6.40; N, 8.72. Found: C, 72.09; H, 6.29; N, 8.60.
3.4.17. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-2-naphthamide (15)
Following the procedure described for compound 4, starting
from 3a and 2-naphthoyl chloride, compound 15 (73%) was ob-
tained as a white solid, mp 155 °C. 1H NMR (DMSO-d6, 500
MHz): d = 2.64–2.74 (m, 6H, 3 Â CH2), 2.81 (t, J = 7.6 Hz, 2H, CH2),
3.54 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.63 (s,
1H, ArH), 6.65 (s, 1H, ArH), 7.24 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.62
(m, 2H, 2 Â ArH), 7.72 (d, J = 8.6 Hz, 2H, 2 Â ArH), 8.02 (m, 4H,
4 Â ArH), 8.56 (s, 1H, ArH), 10.33 (s, 1H*, NH). 13C NMR (DMSO-
d6, 125 MHz): d = 28.5 (CH2), 32.6 (CH2), 50.7 (CH2), 55.2 (CH2),
55.6 (OCH3) 55.7 (OCH3), 59.7 (CH2), 110.2 (Ar-CH), 112.0 (Ar-
CH), 120.5 (2 Â Ar-CH), 124.6 (Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C),
126.9 (Ar-CH), 127.8 (Ar-CH), 127.9 (Ar-CH), 128.0 (Ar-CH), 128.1
(Ar-CH), 128.9 (2 Â Ar-CH), 129.0 (Ar-CH), 132.2 (Ar-C), 132.5
(Ar-C), 134.4 (Ar-C), 136.0 (Ar-C), 137.3 (Ar-C), 147.1 (Ar-C),
147.3 (Ar-C), 165.5 (CO). Anal. Calcd for C30H30N2O3Á0.2H2O: C,
76.64; H, 6.52; N, 5.96. Found: C, 76.79; H, 6.45; N, 6.01.
3.4.14. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)quinoline-6-carboxamide (12)
Following the procedure described for compound 4, starting
from 3a and quinoline-6-carbonyl chloride, compound 12 (56%)
was obtained as a light yellow solid, mp 177 °C. 1H NMR (DMSO-
d6, 500 MHz): d = 2.65–2.73 (m, 6H, 3 Â CH2), 2.81 (t, J = 7.7 Hz,
2H, CH2), 3.54 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3),
6.63 (s, 1H, ArH), 6.65 (s, 1H, ArH), 7.25 (d, J = 8.5 Hz, 2H, 2 Â ArH),
7.62 (q, J = 4.2 Hz, 1H, ArH), 7.72 (d, J = 8.5 Hz, 2H, 2 Â ArH), 8.12
(d, J = 8.9 Hz, 1H, ArH), 8.25 (d, J = 8.5, 1.9 Hz, 1H, ArH), 8.51 (m,
1H, ArH), 8.62 (d, J = 2.2 Hz, 1H, ArH), 9.00 (q, J = 2.0 Hz, 1H,
ArH), 10.42 (s, 1H*, NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.4
(CH2), 32.6 (CH2), 50.7 (CH2), 55.2 (CH2), 55.6 (OCH3) 55.7
(OCH3), 59.7 (CH2), 110.2 (Ar-CH), 112.0 (Ar-CH), 120.6 (2 Â Ar-
CH), 122.4 (Ar-CH), 126.1 (Ar-C), 126.8 (Ar-C), 127.2 (Ar-CH),
128.2 (Ar-C), 128.5 (Ar-CH), 128.9 (2 Â Ar-CH), 129.2 (Ar-CH),
133.0 (Ar-C), 136.1 (Ar-C), 137.1 (Ar-C), 137.3 (Ar-CH), 147.1 (Ar-
C), 147.3 (Ar-C), 148.9 (Ar-C), 152.3 (Ar-CH), 165.0 (CO). Anal.
Calcd for C29H29N3O3Á0.2H2O: C, 73.93; H, 6.29; N, 8.92. Found: C,
73.76; H, 6.45; N, 8.84.
3.4.18. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)nicotinamide (16)
Following the procedure described for compound 4, starting
from 3a and nicotinoyl chloride, compound 16 (59%) was obtained
as a yellow solid, mp 71 °C. 1H NMR (DMSO-d6, 500 MHz):
d = 2.64–2.71 (m, 6H, 3 Â CH2), 2.80 (t, 2H, CH2), 3.53 (s, 2H,
CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.62 (s, 1H, ArH),
6.64 (s, 1H, ArH), 7.24 (td, J = 8.5 Hz, 2H, 2 Â ArH), 7.55 (m, 1H,
ArH), 7.67 (td, J = 8.5 Hz, 2H, 2 Â ArH), 8.28 (m, 1H, ArH), 8.74 (q,
J = 2.1 Hz, 1H, ArH), 9.09 (q, J = 2.2 Hz, 1H, ArH), 10.36 (s, 1H*,
NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2),
50.7 (CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.7 (CH2), 110.2
(Ar-CH), 112.0 (Ar-CH), 120.5 (2 Â Ar-CH), 123.6 (Ar-CH), 126.1
(Ar-C), 126.8 (Ar-C), 128.9 (2 Â Ar-CH), 130.7 (Ar-C), 135.5 (Ar-
CH), 136.3 (Ar-C), 136.9 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 148.8
(Ar-CH), 152.1 (Ar-CH), 164.0 (CO). Anal. Calcd for
C25H27N3O3Á0.5H2O: C, 70.40; H, 6.62; N, 9.85. Found: C, 70.27;
H, 6.96; N, 9.52.
3.4.15. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)quinoxaline-2-carboxamide (13)
Following the procedure described for compound 4, starting
from 3a and quinoxaline-2-carbonyl chloride, compound 13
(41%) was obtained as a white solid, mp 150 °C. 1H NMR (DMSO-
d6, 500 MHz): d = 2.67–2.72 (m, 6H, 3 Â CH2), 2.82 (t, 2H, CH2),
3.54 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.63 (s,
1H, ArH), 6.65 (s, 1H, ArH), 7.28 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.83
(d, J = 8.9 Hz, 2H, 2 Â ArH), 8.01 (m, 2H, 2 Â ArH), 8.22 (m, 1H,
ArH), 8.29 (m, 1H, ArH), 9.54 (s, 1H, ArH), 10.72 (s, 1H*, NH). 13C
NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2),
55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.6 (CH2), 110.2 (Ar-CH),
112.0 (Ar-CH), 120.7 (2 Â Ar-CH), 126.1 (Ar-C), 126.8 (Ar-C),
129.0 (2 Â Ar-CH), 129.3 (Ar-C), 129.7 (Ar-C), 131.5 (Ar-C), 132.2
(Ar-CH), 136.2 (Ar-C), 136.7 (Ar-C), 139.8 (Ar-C), 143.1 (Ar-C),
144.1 (Ar-CH), 144.9 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 161.9
(CO). Anal. Calcd for C28H28N4O3Á0.2H2O: C, 71.23; H, 6.06; N,
11.87. Found: C, 70.95; H, 5.83; N, 11.84.
3.4.19. 2-Bromo-N-(4-(2-(6,7-dimethoxy-3,4-dihydroiso-
quinolin-2(1H)-yl)ethyl)phenyl)benzamide (17)
Following the procedure described for compound 4, starting
from 3a and 2-bromobenzoyl chloride, compound 17 (77%) was
obtained as a light yellow solid, mp 158 °C. 1H NMR (DMSO-d6,
500 MHz): d = 2.64–2.71 (m, 6H, 3 Â CH2), 2.79 (t, 2H, CH2), 3.53
(s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.62 (s, 1H,
ArH), 6.65 (s, 1H, ArH), 7.21 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.40 (dt,
J = 7.9 Hz, 1H, ArH), 7.49 (m, 2H, 2 Â ArH), 7.61 (d, J = 8.5 Hz, 2H,
2 Â ArH), 7.69 (d, J = 7.9 Hz, 1H, ArH), 10.35 (s, 1H*, NH). 13C
NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2),
55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.7 (CH2), 110.2 (Ar-CH),
112.0 (Ar-CH), 119.1 (Ar-C), 119.7 (2 Â Ar-CH), 126.1 (Ar-C),
3.4.16. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-1-naphthamide (14)
Following the procedure described for compound 4, starting
from 3a and 1-naphthoyl chloride, compound 14 (70%) was ob-

2532
W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
126.8 (Ar-C), 127.8 (Ar-CH), 128.9 (Ar-CH), 129.0 (2 Â Ar-CH),
131.2 (Ar-CH), 132.8 (Ar-CH), 136.1 (Ar-C), 137.0 (Ar-C), 139.4
(Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 165.7 (CO). Anal. Calcd for
C26H27BrN2O3Á0.33 H2O: C, 62.28; H, 5.56; N, 5.59. Found: C,
62.00; H, 5.38; N, 5.99.
ArH), 6.64 (s, 1H, ArH), 7.22 (t, 3H, 3 Â ArH), 7.56 (d, J = 8.5 Hz,
2H, 2 Â ArH), 7.68 (s, 1H, ArH), 10.38 (s, 1H*, NH). 13C NMR
(DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2), 55.2
(CH2), 55.6 (OCH3), 55.7 (OCH3), 56.5 (OCH3), 56.8 (OCH3), 59.7
(CH2), 107.4 (Ar-CH), 110.2 (Ar-CH), 111.2 (Ar-CH), 112.0 (Ar-
CH), 119.7 (2 Â Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C), 127.6 (Ar-CH),
129.0 (2 Â Ar-CH), 136.1 (Ar-C), 137.1 (Ar-C), 138.9 (Ar-C), 147.1
(Ar-C), 147.3 (Ar-C), 149.1 (Ar-C), 153.2 (Ar-C), 164.0 (CO). Anal.
Calcd for C28H31N3O7Á0.33H2O: C, 63.75; H, 6.05; N, 7.96. Found:
C, 63.66; H, 5.86; N, 7.98.
3.4.20. 3-Bromo-N-(4-(2-(6,7-dimethoxy-3,4-dihydroiso-
quinolin-2(1H)-yl)ethyl)phenyl)benzamide (18)
Following the procedure described for compound 4, starting
from 3a and 3-bromobenzoyl chloride, compound 18 (69%) was
obtained as a pale yellow solid, mp 230 °C. 1H NMR (DMSO-d6,
500 MHz): 1H NMR (DMSO-d6, 500 MHz): d = 2.93–3.07 (m, 4H,
2 Â CH2), 3.15–3.28 (m, 4H, 2 Â CH2), 3.72 (s, 3H, OCH3), 3.73 (s,
3H, OCH3), 4.11 (s, 2H, CH2), 6.73 (s, 1H, ArH), 6.76 (s, 1H, ArH),
7.27 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.49 (t, J = 7.9 Hz, 1H, ArH), 7.72
(d, J = 8.5 Hz, 2H, 2 Â ArH), 7.78 (m, 1H, ArH), 7.96 (dt, J = 8.2 Hz,
1H, ArH), 8.14 (t, J = 1.9 Hz, 1H, ArH), 10.35 (s, 1H*, NH). Anal. Calcd
for C26H27BrN2O3Á0.2H2O: C, 62.58; H, 5.53; N, 5.61. Found: C,
62.54; H, 5.59; N, 5.55.
3.4.24. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)benzo[d][1,3]-dioxole-5-carboxamide (22)
Followingthe procedure described for compound 4, starting from
3a and benzo[d][1,3]dioxole-5-carbonyl chloride, compound 22
(80%) was obtained as a white solid, mp 180 °C. 1H NMR (DMSO-
d6, 500 MHz): d = 2.66–2.71 (m, 6H, 3 Â CH2), 2.78 (t, J = 7.5 Hz, 2H,
CH2), 3.56 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 6.11
(s, 2H, CH2), 6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 7.03 (d, J = 8.2 Hz,
1H, ArH), 7.20 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.50 (d, J = 1.5 Hz, 1H,
ArH), 7.56 (dd, J = 8.2 Hz, 1H, ArH), 7.64 (d, J = 8.5 Hz, 2H, 2 Â ArH),
9.96 (s, 1H*, NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.3 (CH2),
32.5 (CH2), 50.7 (CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.6
(CH2), 101.9 (CH2), 107.8 (Ar-CH), 108.0 (Ar-CH), 110.2 (Ar-CH),
112.0 (Ar-CH), 120.5 (2 Â Ar-CH), 122.9 (Ar-CH), 126.0 (Ar-C),
126.6 (Ar-C), 128.8 (2 Â Ar-CH), 129.0 (Ar-C), 135.7 (Ar-C), 137.3
(Ar-C), 147.1 (Ar-C), 147.5 (Ar-C), 147.7 (Ar-C), 150. 1 (Ar-C), 164.4
(CO). Anal. Calcd for C27H28N2O5Á0.2H2O: C, 69.87; H, 6.17; N, 6.04.
Found: C, 69.93; H, 6.22; N, 6.02.
3.4.21. 4-Bromo-N-(4-(2-(6,7-dimethoxy-3,4-dihydro-
isoquinolin-2(1H)-yl)ethyl)phenyl)benzamide (19)
Following the procedure described for compound 4, starting
from 3a and 4-bromobenzoyl chloride, compound 19 (74%) was
obtained as a white solid, mp 183 °C. 1H NMR (CDCl3, 500 MHz):
d = 2.76 (m, 4H, 2 Â CH2), 2.83 (t, 2H, CH2), 2.88 (q, 2H, CH2),
3.63 (s, 2H, CH2), 3.81 (s, 3H, OCH3), 3.82 (s, 3H, OCH3), 6.52 (s,
1H, ArH), 6.58 (s, 1H, ArH), 7.21 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.52
(d, J = 8.2 Hz, 2H, 2 Â ArH), 7.57 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.70
(d, J = 8.5 Hz, 2H, 2 Â ArH), 7.85 (s, 1H*, NH). 13C NMR (CDCl3,
125 MHz): d = 28.6 (CH2), 33.4 (CH2), 51.0 (CH2), 55.7 (CH2), 55.9
(OCH3) 55.9 (OCH3), 60.1 (CH2), 109.5 (Ar-CH), 111.4 (Ar-CH),
120.5 (2 Â Ar-CH), 126.1 (Ar-C), 126.4 (Ar-C), 126.5 (Ar-C), 128.6
(2 Â Ar-CH), 129.3 (2 Â Ar-CH), 132.0 (2 Â Ar-CH), 133.8 (Ar-C),
135.7 (Ar-C), 137.0 (Ar-C), 147.2 (Ar-C), 147.5 (Ar-C), 164.7 (CO).
Anal. Calcd for C26H27BrN2O3Á0.5H2O: C, 61.91; H, 5.60; N, 5.55.
Found: C, 61.58; H, 5.43; N, 5.59.
3.4.25. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)cinnamamide (23)
Following the procedure described for compound 4, starting
from 3a and cinnamoyl chloride, compound 23 (38%) was obtained
as a pale yellow solid, mp 150 °C. 1H NMR (DMSO-d6, 500 MHz):
d = 2.69–2.83 (m, 8H, 4 Â CH2), 3.61 (s, 2H, CH2), 3.69 (s, 3H,
OCH3), 3.70 (s, 3H, OCH3), 6.64 (s, 1H, ArH), 6.66 (s, 1H, ArH),
6.83 (d, J = 15.5 Hz, 1H, CO–CH@CH), 7.21 (d, J = 8.6 Hz, 2H,
2 Â ArH), 7.41 (m, 3H, 3 Â ArH), 7.56 (d, J = 15.8 Hz, 1H, Ar-
CH@CH), 7.61 (m, 4H, 4 Â ArH), 10.14 (s, 1H*, NH). 13C NMR
(DMSO-d6, 125 MHz): d = 28.1 (CH2), 32.2 (CH2), 50.6 (CH2), 54.9
(CH2), 55.6 (OCH3) 55.7 (OCH3), 59.3 (CH2), 110.2 (Ar-CH), 112.0
(Ar-CH), 119.4 (2 Â Ar-CH), 122.6 (CO-CH=CH), 125.8 (Ar-C),
126.1 (Ar-C), 127.8 (2 Â Ar-CH), 129.1 (2 Â Ar-CH), 129.1 (2 Â Ar-
CH), 129.8 (Ar-CH), 134.9 (Ar-C), 135.3 (Ar-C), 137.4 (Ar-C), 140.0
(Ar-CH=CH), 147.1 (Ar-C), 147.4 (Ar-C), 163.5 (CO). Anal. Calcd
for C28H30N2O3Á0.4H2O: C, 74.77; H, 6.90; N, 6.23. Found: C,
74.48; H, 7.20; N, 6.61.
3.4.22. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-3,4-dimethoxybenzamide (20)
Following the procedure described for compound 4, starting
from 3a and 3,4-dimethoxybenzoyl chloride, compound 20 (55%)
was obtained as a white solid, mp 181 °C. 1H NMR (DMSO-d6,
500 MHz): d = 2.66–2.82 (m, 8H, 4 Â CH2), 3.57 (s, 2H, CH2), 3.69
(s, 3H, OCH3), 3.69 (s, 3H, OCH3), 3.83 (s, 3H, OCH3), 3.83 (s, 3H,
OCH3), 6.63 (s, 1H, ArH), 6.65 (s, 1H, ArH), 7.06 (d, J = 8.5 Hz, 1H,
ArH), 7.21 (d, J = 8.2 Hz, 2H, 2 Â ArH), 7.52 (d, J = 1.9 Hz, 1H,
ArH), 7.60 (dd, J = 8.2 Hz, J = 1.9 Hz, 1H, ArH), 7.65 (d, J = 8.5 Hz,
2H, 2 Â ArH), 9.97 (s, 1H*, NH). 13C NMR (DMSO-d6, 125 MHz):
d = 28.3 (CH2), 32.4 (CH2), 50.9 (CH2), 55.1 (CH2), 55.6 (OCH3),
55.7 (OCH3), 55.8 (OCH3), 55.8 (OCH3), 59.5 (CH2), 110.2 (Ar-CH),
111.1 (Ar-CH), 111.3 (Ar-CH), 112.0 (Ar-CH), 120.7 (2 Â Ar-CH),
121.1 (Ar-CH), 126.0 (Ar-C), 126.6 (Ar-C), 127.2 (Ar-C), 128.8
(2 Â Ar-CH), 135.5 (Ar-C), 137.3 (Ar-C), 147.1 (Ar-C), 147.4 (Ar-C),
148.5 (Ar-C), 151.7 (Ar-C), 164.9 (CO). Anal. Calcd for
C28H32N2O5Á0.67H2O: C, 68.83; H, 6.88; N, 5.73. Found: C, 68.78;
H, 6.94; N, 6.07.
3.4.26. (E)-N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-3-(2-nitrophenyl)acrylamide (24)
Following the procedure described for compound 4, starting
from 3a and (E)-3-(2-nitrophenyl)acryloyl chloride, compound 24
(72%) was obtained as a pale yellow solid, mp 169 °C. 1H NMR
(DMSO-d6, 500 MHz): d = 2.64–2.72 (m, 6H, 3 Â CH2), 2.79 (m,
J = 8.6 Hz, 2H, CH2), 3.53 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s,
3H, OCH3), 6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 6.81 (d, J = 15.8
Hz, 1H, CO-CH=CH), 7.22 (d, J = 8.6 Hz, 2H, 2 Â ArH), 7.60 (d,
J = 8.6 Hz, 2H, 2 Â ArH), 7.65 (m, 1H, ArH), 7.81 (m, 2H, 2 Â ArH),
7.83 (d, J = 15.8 Hz, 1H, Ar-CH=CH), 8.06 (d, J = 7.9 Hz, 1H, ArH),
10.25 (s, 1H*, NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2),
32.6 (CH2), 50.7 (CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.6
(CH2), 110.2 (Ar-CH), 112.0 (Ar-CH), 119.5 (2 Â Ar-CH), 124.8
(CO-CH=CH), 126.1 (Ar-C), 126.8 (Ar-C), 127.3 (Ar-CH), 128.9 (Ar-
CH), 129.1 (2 Â Ar-CH), 130.1 (Ar-C), 130.5 (Ar-CH), 134.0 (Ar-
CH), 135.0 (Ar-CH@CH), 136.0 (Ar-C), 137.0 (Ar-C), 147.1 (Ar-C),
3.4.23. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-4,5-dimethoxy-2-nitrobenzamide (21)
Following the procedure described for compound 4, starting
from 3a and 4,5-dimethoxy-2-nitrobenzoyl chloride, compound
21 (66%) was obtained as a pale yellow solid, mp 152 °C. 1H
NMR (DMSO-d6, 500 MHz): d = 2.64–2.70 (m, 6H, 3 Â CH2), 2.79
(t, J = 7.5 Hz, 2H, CH2), 3.62 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69
(s, 3H, OCH3), 3.91 (s, 3H, OCH3), 3.93 (s, 3H, OCH3), 6.62 (s, 1H,

W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
2533
147.3 (Ar-C), 148.4 (Ar-C), 162.7 (CO). Anal. Calcd for
C28H29N3O5Á0.5H2O: C, 67.73; H, 6.09; N, 8.46. Found: C, 67.69;
H, 5.86; N, 8.49.
(81%) was obtained as a white solid, mp 131 °C. MS m/z 492.2
(MH+, 60), 307.1 (18), 289.1 (10), 206.2 (100), 165.1 (11). 1H
NMR (DMSO-d6, 500 MHz): d = 2.62–2.69 (m, 6H, 3 Â CH2), 2.77
(t, J = 7.7 Hz, 2H, CH2), 3.51 (s, 2H, CH2), 3.68 (s, 3H, OCH3), 3.69
(s, 3H, OCH3), 4.90 (s, 2H, OCH2), 6.61 (s, 1H, ArH), 6.64 (s, 1H,
ArH), 7.15 (dt, J = 8.2 Hz, 1H, ArH), 7.20 (d, J = 8.5 Hz, 2H, 2 Â
ArH), 7.28 (d, J = 8 Hz, 1H, ArH), 7.48 (d, J = 8.5 Hz, 2H, 2 Â ArH),
7.64 (m, 1H, ArH), 7.91 (dd, J = 8.2 Hz, 1H, ArH), 9.95 (s, 1H*,
NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.5 (CH2),
50.7 (CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.7 (CH2), 68.0
(OCH2), 110.2 (Ar-CH), 112.0 (Ar-CH), 115.6 (Ar-CH), 119.5
(2 Â Ar-CH), 121.4 (Ar-CH), 125.4 (Ar-CH), 126.1 (Ar-C), 126.9
(Ar-C), 129.1 (2 Â Ar-CH), 134.7 (Ar-CH), 136.2 (Ar-C), 136.2 (Ar-
C), 139.6 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 151.0 (Ar-C), 165.4
(CO). Anal. Calcd for C27H29N3O6Á0.2H2O: C, 65.49; H, 5.98; N,
8.49. Found: C, 65.55; H, 6.07; N, 8.47.
3.4.27. (E)-3-(4-Chlorophenyl)-N-(4-(2-(6,7-dimethoxy-3,4-
dihydroisoquinolin-2(1H)-yl)ethyl)phenyl)acrylamide (25)
Following the procedure described for compound 4, starting
from 3a and (E)-3-(4-chlorophenyl)acryloyl chloride, compound
25 (28%) was obtained as a pale grey solid, mp 198 °C. 1H NMR
(DMSO-d6, 500 MHz): d = 2.64–2.70 (m, 6H, 3 Â CH2), 2.78 (t, 2H,
CH2), 3.53 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69 (s, 3H, OCH3),
6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 6.78 (d, J = 15.5 Hz, 1H, CO-
CH=CH), 7.20 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.48–7.64 (m, 7H,
6 Â ArH, 1 Â Ar-CH=CH), 10.11 (s, 1H*, NH). 13C NMR (DMSO-d6,
125 MHz): d = 28.4 (CH2), 32.5 (CH2), 50.7 (CH2), 55.2 (CH2), 55.6
(OCH3) 55.7 (OCH3), 59.6 (CH2), 110.2 (Ar-CH), 112.0 (Ar-CH),
119.4 (2 Â Ar-CH), 123.4 (CO-CH=CH), 126.1 (Ar-C), 126.8 (Ar-C),
129.1 (2 Â Ar-CH), 129.2 (2 Â Ar-CH), 129.5 (2 Â Ar-CH), 133.9
(Ar-C), 134.2 (Ar-C), 135.7 (Ar-C), 137.2 (Ar-C), 138.6 (Ar-CH@CH),
147.1 (Ar-C), 147.3 (Ar-C), 163.2 (CO). Anal. Calcd for
C28H29ClN2O3Á0.5H2O: C, 69.20; H, 6.22, Cl, 7.29; N, 5.76. Found:
C, 69.16; H, 6.38; N, 5.70.
3.4.31. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-2-(2-aminophenoxy)acetamide (29)
Following the procedure described for compound 3a, starting
from 28 (1.03 g, 2.1 mmol), compound 29 (89%) was obtained as
a
white solid, mp 163 °C. 1H NMR (DMSO-d6, 500 MHz):
d = 2.62–2.70 (m, 6H, 3 Â CH2), 2.78 (t, J = 7.7 Hz, 2H, CH2), 3.52
(s, 2H, CH2), 3.68 (s, 3H, OCH3), 3.69 (s, 3H, OCH3), 4.59 (s, 2H,
OCH2), 5.03 (s, 2H*, NH2), 6.49 (m, 1H, ArH), 6.62 (s, 1H, ArH),
6.64 (s, 1H, ArH), 6.66 (dd, J = 6 Hz, 1H, ArH), 6.72 (dt, J = 7.2 Hz,
1H, ArH), 6.81 (dd, J = 8.2 Hz, 1H, ArH), 7.20 (d, J = 8.5 Hz, 2H, 2 Â
ArH), 7.53 (d, J = 8.5 Hz, 2H, 2 Â ArH), 9.86 (s, 1H*, NH). 13C NMR
(DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2), 55.2
(CH2), 55.6 (OCH3) 55.7 (OCH3), 59.7 (CH2), 68.1 (OCH2), 110.2
(Ar-CH), 112.0 (Ar-CH), 112.9 (Ar-CH), 114.6 (Ar-CH), 116.2 (Ar-
CH), 120.6 (2 Â Ar-CH), 122.2 (Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C),
128.9 (2 Â Ar-CH), 136.1 (Ar-C), 136.3 (Ar-C), 138.4 (Ar-C), 145.0
(Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 166.8 (CO). Anal. Calcd for
C27H31N3O4: C, 70.26; H, 6.77; N, 9.10. Found: C, 70.03; H, 6.93;
N, 9.03.
3.4.28. (E)-3-(4,5-Dimethoxy-2-nitrophenyl)-N-(4-(2-(6,7-
dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)phenyl)-
acrylamide (26)
Following the procedure described for compound 4, starting
from 3a and (E)-3-(4,5-dimethoxy-2-nitrophenyl)acryloyl chloride,
compound 26 (84%) was obtained as a yellow solid, mp 184 °C. 1H
NMR (DMSO-d6, 500 MHz): d = 2.64–2.70 (m, 6H, 3 Â CH2), 2.78 (t,
J = 7.5 Hz, 2H, CH2), 3.53 (s, 2H, CH2), 3.68 (s, 3H, OCH3), 3.69 (s, 3H,
OCH3), 3.89 (s, 3H, OCH3), 3.95 (s, 3H, OCH3), 6.62 (s, 1H, ArH), 6.64
(s, 1H, ArH), 6.78 (d, J = 15.5 Hz, 1H, ArH), 7.20 (d, J = 8.5 Hz, 2H,
2 Â ArH), 7.26 (s, 1H, ArH), 7.60 (d, J = 8.9 Hz, 2H, 2 Â ArH), 7.65
(s, 1H, ArH), 7.90 (d, J = 15.5 Hz, 1H, ArH), 10.21 (s, 1H*, NH). 13C
NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.6 (CH2), 50.7 (CH2),
55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 56.3 (OCH3) 56.4 (OCH3),
59.6 (CH2), 108.1 (Ar-CH), 110.1 (Ar-CH), 110.2 (Ar-CH), 112.0
(Ar-CH), 119.4 (2 Â Ar-CH), 124.7 (Ar-C), 126.1 (CH), 126.1 (Ar-C),
126.8 (Ar-C), 129.1 (2 Â Ar-CH), 135.4 (CH), 135.8 (Ar-C), 137.1
(Ar-C), 141.2 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 149.4 (Ar-C),
153.0 (Ar-C), 162.9 (CO). Anal. Calcd for C30H33N3O7Á0.75H2O: C,
64.22; H, 6.20; N, 7.49. Found: C, 64.01; H, 5.97; N, 7.44.
3.4.32. 1-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-3-(2-nitrophenyl)urea (30a)
3 mmol of 1-isocyanato-2-nitrobenzene was suspended in dry
THF and added dropwise to a stirred solution of 3a (3 mmol) in
50 ml of dry THF at room temperature. After a few minutes cata-
lytic amounts of triethylamine was added and the solution was re-
fluxed for 6 h. After the reaction was completed the solvent was
removed in vacuo. To the residue 50 ml of water was added and ex-
tracted three times with EtOAc. The organic phase was dried with
MgSO4, and concentrated. Recrystallization from DCM/n-hexane
the compound 30a was yielded as a yellow solid (71%), mp 167
°C. 1H NMR (DMSO-d6, 500 MHz): d = 2.62–2.70 (m, 6H, 3 Â CH2),
2.76 (t, J = 7.5 Hz, 2H, CH2), 3.52 (s, 2H, CH2), 3.68 (s, 3H, OCH3),
3.69 (s, 3H, OCH3), 6.61 (s, 1H, ArH), 6.64 (s, 1H, ArH), 7.18 (m,
3H, 3 Â ArH), 7.38 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.67 (dt, J = 8.5 Hz,
1H, ArH), 8.07 (dd, J = 8.2 Hz, 1H, ArH), 8.29 (dd, J = 8.5 Hz, 1H,
ArH), 9.56 (s, 1H*, NH), 9.73 (s, 1H*, NH). 13C NMR (DMSO-d6,
125 MHz): d = 28.4 (CH2), 32.5 (CH2), 50.7 (CH2), 55.2 (CH2), 55.6
(OCH3) 55.7 (OCH3), 59.7 (CH2), 110.2 (Ar-CH), 112.0 (Ar-CH),
118.9 (2 Â Ar-CH), 122.2 (Ar-CH), 122.6 (Ar-CH), 125.5 (Ar-CH),
126.1 (Ar-C), 126.8 (Ar-C), 129.1 (2 Â Ar-CH), 134.8 (Ar-C), 135.1
(Ar-CH), 135.2 (Ar-C), 137.1 (Ar-C), 137.7 (Ar-C), 147.0 (Ar-C),
147.3 (Ar-C), 151.9 (CO). Anal. Calcd for C26H28N4O5: C, 65.53; H,
5.92; N, 11.76. Found: C, 65.32; H, 6.06; N, 11.61.
3.4.29. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-2-phenoxyacetamide (27)
Following the procedure described for compound 4, starting
from 3a and 2-phenoxyacetyl chloride, compound 27 (44%) was
obtained as a pale white solid, mp 122 °C. 1H NMR (DMSO-d6,
500 MHz): d = 2.66–2.80 (m, 8H, 4 Â CH2), 3.57 (s, 2H, CH2), 3.69
(s, 3H, OCH3), 3.69 (s, 3H, OCH3), 4.66 (s, 2H, OCH2), 6.63 (s, 1H,
ArH), 6.65 (s, 1H, ArH), 6.98 (m, 3H, 3 Â ArH), 7.19 (d, J = 8.5 Hz,
2H, 2 Â ArH), 7.30 (m, 2H, 2 Â ArH), 7.54 (d, J = 8.5 Hz, 2H,
2 Â ArH), 9.95 (s, 1H*, NH). 13C NMR (DMSO-d6, 125 MHz):
d = 28.2 (CH2), 32.3 (CH2), 50.6 (CH2), 55.0 (CH2), 55.6 (OCH3)
55.7 (OCH3), 59.4 (CH2), 67.3 (CH2), 110.2 (Ar-CH), 112.0 (Ar-CH),
114.8 (2 Â Ar-CH), 119.9 (2 Â Ar-CH), 121.3 (Ar-CH), 125.9 (Ar-C),
126.4 (Ar-C), 129.0 (2 Â Ar-CH), 129.6 (2 Â Ar-CH), 135.8 (Ar-C),
136.5 (Ar-C), 147.1 (Ar-C), 147.4 (Ar-C), 158.0 (Ar-C), 166.5 (CO).
Anal. Calcd for C27H30N2O4Á0.75H2O: C, 70.49; H, 6.90; N, 6.09.
Found: C, 70.35; H, 7.06; N, 5.88.
3.4.30. N-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-2-(2-nitrophenoxy)acetamide (28)
3.4.33. 1-(4-(2-(3,4-Dihydroisoquinolin-2(1H)-yl)ethyl)phenyl)-
3-(2-nitrophenyl)urea (30b)
Following the procedure described for compound 4, starting
from 3a and 2-(2-nitrophenoxy)acetyl chloride, compound 28
Following the procedure described for compound 30a, starting
from 3b and 1-isocyanato-2-nitrobenzene, compound 30b (85%)

2534
W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
was obtained as a yellow solid, mp 157 °C. 1H NMR (CDCl3, 500
MHz): d = 2.75 (m, 2H, CH2), 2.80 (t, 2H, CH2), 2.87–2.93 (m, 4H,
2 Â CH2), 3.70 (s, 2H, CH2), 7.00–7.11 (m, 5H, 5 Â ArH), 7.21 (d,
J = 8.5 Hz, 2H, 2 Â ArH), 7.32 (m, 3H, 2 Â ArH, NH*), 7.56 (m, 1H,
ArH), 8.13 (d, J = 8.5 Hz, 1H, ArH), 8.64 (d, J = 8.5 Hz, 1H, ArH),
9.91 (s, 1H*, NH). 13C NMR (CDCl3, 125 MHz): d = 29.0 (CH2), 33.3
(CH2), 51.0 (CH2), 56.0 (CH2), 60.1 (CH2), 121.8 (Ar-CH), 121.8
(2 Â Ar-CH), 122.2 (Ar-CH), 125.6 (Ar-CH), 125.7 (Ar-CH), 126.1
(Ar-CH), 126.6 (Ar-CH), 128.7 (Ar-CH), 129.6 (2 Â Ar-CH) 134.2
(Ar-C), 134.6 (Ar-C), 135.1 (Ar-C), 135.8 (Ar-CH), 135.9 (Ar-C),
136.4 (Ar-C), 137.4 (Ar-C), 152.5 (CO). Anal. Calcd for
C24H24N4O3Á0.5H2O: C, 67.75; H, 5.92; N, 13.17. Found: C, 67.96;
H, 5.90; N, 13.34.
(2 Â Ar-CH), 123.7 (Ar-CH), 124.4 (Ar-CH), 125.0 (Ar-C), 126.1
(Ar-C), 126.8 (Ar-C), 129.0 (2 Â Ar-CH), 133.6 (Ar-C), 138.1 (Ar-C),
140.9 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 153.3 (CO). Anal. Calcd
for C26H30N4O3Á0.2H2O: C, 69.37; H, 6.81; N, 12.45. Found: C,
69.27; H, 6.90; N, 12.34.
3.4.37. 1-(2-Aminophenyl)-3-(4-(2-(3,4-dihydroisoquinolin-
2(1H)-yl)ethyl)phenyl)urea (33b)
Following the procedure described for compound 3a, starting
from 30b (0.84 g, 2 mmol), compound 33b (58%) was obtained as
white solid, mp 176 °C. 1H NMR (DMSO-d6, 500 MHz): d = 2.66
(m, 2H, CH2), 2.71 (d, J = 5.8 Hz, 2H, CH2), 2.75–2.81 (m, 4H,
2 Â CH2), 3.58 (s, 2H, CH2), 4.73 (s, 2H*, NH2), 6.56 (dt, J = 7.6 Hz,
1H, ArH), 6.72 (dd, J = 7.6 Hz, 1H, ArH), 6.82 (dt, J = 7.6 Hz, 1H,
ArH), 7.03 (m, 1H, ArH), 7.09 (m, 3H, 3 Â ArH), 7.14 (d, J = 8.5 Hz,
2H, 2 Â ArH), 7.33 (m, 3H, 3 Â ArH), 7.67 (s, 1H*, NH), 8.62 (s,
1H*, NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.6 (CH2), 32.4
(CH2), 50.5 (CH2), 55.6 (CH2), 59.8 (CH2), 116.0 (Ar-CH), 117.0
(Ar-CH), 118.2 (2 Â Ar-CH), 123.7 (Ar-CH), 124.4 (Ar-CH), 125.0
(Ar-C), 125.5 (Ar-CH), 126.0 (Ar-CH), 126.5 (Ar-CH), 128.5 (Ar-
CH), 129.0 (2 Â Ar-CH) 133.6 (Ar-CH), 134.3 (Ar-C), 135.1 (Ar-C),
138.1 (Ar-C), 140.9 (Ar-C), 153.3 (CO). Anal. Calcd for C24H26N4O:
C, 74.58; H, 6.78; N, 14.50. Found: C, 74.25; H, 6.62; N, 14.87.
3.4.34. 1-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-3-(3-nitrophenyl)urea (31)
Following the procedure described for compound 30a, starting
from 3a and 1-isocyanato-3-nitrobenzene, compound 31 (93%)
was obtained as a yellow solid, mp 166 °C. 1H NMR (DMSO-d6,
500 MHz): d = 2.62–2.70 (m, 6H, 3 Â CH2), 2.76 (t, J = 7.6 Hz, 2H,
CH2), 3.52 (s, 2H, CH2), 3.68 (s, 3H, OCH3), 3.69 (s, 3H, OCH3),
6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 7.13 (d, J = 8.5 Hz, 2H, 2 Â ArH),
7.37 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.54 (t, J = 8.2 Hz, 1H, ArH), 7.69
(dd, J = 8.2 Hz, 1H, ArH), 7.79 (dd, J = 8.2 Hz, 1H, ArH), 8.54 (t,
J = 2.2 Hz, 1H, ArH), 8.75 (s, 1H*, NH), 9.19 (s, 1H*, NH). 13C NMR
(DMSO-d6, 125 MHz): d = 28.5 (CH2), 32.5 (CH2), 50.7 (CH2), 55.2
(CH2), 55.6 (OCH3) 55.7 (OCH3), 59.6 (CH2), 110.2 (Ar-CH), 112.0
(Ar-CH), 112.2 (Ar-CH), 116.4 (Ar-CH), 118.9 (2 Â Ar-CH), 124.4
(Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C), 129.1 (2 Â Ar-CH), 130.1 (Ar-
CH), 134.6 (Ar-C), 137.2 (Ar-C), 141.3 (Ar-C), 147.0 (Ar-C), 147.3
3.4.38. 1-(3-Aminophenyl)-3-(4-(2-(6,7-dimethoxy-3,4-dihydro-
isoquinolin-2(1H)-yl)ethyl)-phenyl)urea (34)
Following the procedure described for compound 3a, starting
from 31 (0.95 g, 2 mmol), compound 34 (81%) was obtained as a
white solid, mp 172 °C. 1H NMR (DMSO-d6, 500 MHz): d = 2.61–
2.75 (m, 8H, 4 Â CH2), 3.52 (s, 2H, CH2), 3.69 (s, 3H, OCH3), 3.69
(s, 3H, OCH3), 4.97 (s, 2H*, NH2), 6.17 (d, J = 7.9 Hz, 1H, ArH),
6.53 (d, J = 8.2 Hz, 1H, ArH), 6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH),
6.75 (s, 1H, ArH), 6.87 (t, 1H, J = 7.9 Hz, ArH), 7.13 (d, J = 8.5 Hz,
2H, 2 Â ArH), 7.32 (d, J = 8.5 Hz, 2H, 2 Â ArH), 8.28 (s, 1H*, NH),
8.43 (s, 1H*, NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2),
32.5 (CH2), 50.7 (CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.8
(CH2), 103.9 (Ar-CH), 106.2 (Ar-CH), 108.2 (Ar-CH), 110.2 (Ar-
CH), 112.0 (Ar-CH), 118.2 (2 Â Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C),
129.0 (2 Â Ar-CH), 129.1 (Ar-CH), 133.8 (Ar-C), 137.9 (Ar-C),
140.5 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C), 149.2 (Ar-C), 152.6 (CO).
Anal. Calcd for C26H30N4O3Á?H2O: C, 69.00; H, 6.83; N, 12.38.
Found: C, 68.75; H, 6.76; N, 12.38.
(Ar-C),
148.3
(Ar-C),
152.6
(CO).
Anal.
Calcd
for
C26H28N4O5Á0.67H2O: C, 63.92; H, 6.05; N, 11.47. Found: C, 63.79;
H, 6.26; N, 11.25.
3.4.35. 1-(4-(2-(6,7-Dimethoxy-3,4-dihydroisoquinolin-2(1H)-
yl)ethyl)phenyl)-3-(4-nitrophenyl)urea (32)
Following the procedure described for compound 30a, starting
from 3a and 1-isocyanato-4-nitrobenzene, compound 32 (90%)
was obtained as a yellow solid, mp 138 °C. 1H NMR (DMSO-d6,
500 MHz): d = 2.62–2.70 (m, 6H, 3 Â CH2), 2.76(t, J = 7.5, 2H,
CH2), 3.52 (s, 2H, CH2), 3.68 (s, 3H, OCH3), 3.69 (s, 3H, OCH3),
6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 7.17 (d, J = 8.5 Hz, 2H, 2 Â ArH),
7.37 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.67 (d, J = 9.5 Hz, 2H, 2 Â ArH),
8.16 (d, J = 9.5 Hz, 2H, 2 Â ArH), 8.84 (s, 1H*, NH), 9.41 (s, 1H*,
NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.5 (CH2),
50.7 (CH2), 55.2 (CH2), 55.6 (OCH3), 55.7 (OCH3), 59.7 (CH2),
110.2 (Ar-CH), 112.0 (Ar-CH), 117.5 (2 Â Ar-CH), 118.9 (2 Â Ar-
CH), 125.3 (2 Â Ar-CH), 126.1 (Ar-C), 126.9 (Ar-C), 129.1 (2 Â Ar-
CH), 134.8 (Ar-C), 137.0 (Ar-C), 141.1 (Ar-C), 146.6 (Ar-C), 147.1
3.4.39. 1-(4-Aminophenyl)-3-(4-(2-(6,7-dimethoxy-3,4-dihydro-
isoquinolin-2(1H)-yl)ethyl)-phenyl)urea (35)
Following the procedure described for compound 3a, starting
from 32 (0.95 g, 2 mmol), compound 35 (76%) was obtained as a
light yellow solid, mp 187 °C. 1H NMR (DMSO-d6, 500 MHz):
d = 2.61–2.75 (m, 8H, 4 Â CH2), 3.52 (s, 2H, CH2), 3.68 (s, 3H,
OCH3), 3.69 (s, 3H, OCH3), 4.72 (s, 2H*, NH2), 6.50 (d, J = 8.8 Hz,
2H, 2 Â ArH), 6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 7.05 (d, J = 8.8
Hz, 2H, 2 Â ArH), 7.11 (d, J = 8.5 Hz, 2H, 2 Â ArH), 7.31 (d, J = 8.5
Hz, 2H, 2 Â ArH), 8.07 (s, 1H*, NH), 8.35 (s, 1H*, NH). 13C NMR
(DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.5 (CH2), 50.7 (CH2), 55.2
(CH2), 55.6 (OCH3), 55.7 (OCH3), 59.9 (CH2), 110.2 (Ar-CH), 112.0
(Ar-CH), 114.3 (2 Â Ar-CH), 118.2 (2 Â Ar-CH), 120.8 (2 Â Ar-CH),
126.1 (Ar-C), 126.9 (Ar-C), 128.8 (Ar-C), 129.0 (2 Â Ar-CH), 133.5
(Ar-C), 138.2 (Ar-C), 144.1 (Ar-C), 147.1 (Ar-C), 147.3 (Ar-C),
153.1 (CO). Anal. Calcd for C26H30N4O3Á0.4H2O: C, 68.82; H, 6.84;
N, 12.35. Found: C, 68.76; H, 6.69; N, 12.23.
(Ar-C),
147.3
(Ar-C),
152.1
(CO).
Anal.
Calcd
for
C26H28N4O5Á1.0H2O: C, 63.15; H, 6.11; N, 11.33. Found: C, 63.15;
H, 6.23; N, 11.03.
3.4.36. 1-(2-Aminophenyl)-3-(4-(2-(6,7-dimethoxy-3,4-dihydro-
isoquinolin-2(1H)-yl)ethyl)-phenyl)urea (33a)
Following the procedure described for compound 3a, starting
from 30a (0.95 g, 2 mmol), compound 31a (88%) was obtained as
a light yellow solid, mp 171 °C. 1H NMR (DMSO-d6, 500 MHz):
d = 2.63–2.76 (m, 8H, 4 Â CH2), 3.53 (s, 2H, CH2), 3.68 (s, 3H,
OCH3), 3.69 (s, 3H, OCH3), 4.74 (s, 2H*, NH2), 6.56 (dt, J = 7.9 Hz,
1H, ArH), 6.62 (s, 1H, ArH), 6.64 (s, 1H, ArH), 6.72 (dd, J = 7.9 Hz,
1H, ArH), 6.82 (dt, J = 7.9 Hz, 1H, ArH), 7.13 (d, J = 8.5 Hz, 2H,
2 Â ArH), 7.33 (m, 3H, 3 Â ArH), 7.70 (s, 1H*, NH), 8.65 (s, 1H*,
NH). 13C NMR (DMSO-d6, 125 MHz): d = 28.4 (CH2), 32.4 (CH2),
50.7 (CH2), 55.2 (CH2), 55.6 (OCH3) 55.7 (OCH3), 59.8 (CH2), 110.2
(Ar-CH), 112.0 (Ar-CH), 116.0 (Ar-CH), 117.0 (Ar-CH), 118.2
3.5. Calcein AM assay
Cells were grown under standard conditions in T75- or T175-
flasks. After reaching confluence of approximately 80%, cells were
harvested by short trypsination (0.05 % trypsin/0.02 % EDTA). Pel-

W. Klinkhammer et al. / Bioorg. Med. Chem. 17 (2009) 2524–2535
2535
leted cells were resuspended in fresh culture medium and counted
with a Casy I Modell TT cell counter (Schaerfe System GmbH, Reut-
lingen, Germany). Cells were washed three times with Krebs-
HEPES buffer, and then seeded into colourless 96 well plates (Gre-
iner, Frickenhausen, Germany) at a density of approximately
Germany). This work was partly funded by a grant from Deutsche
Forschungsgemeinschaft (Grant No. WI 874/4-1).
References and notes
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30,000 cells in a volume of 90
compounds were added, resulting in a final volume of 100
well. The prepared 96 well plates were preincubated for 30 min.
After the preincubation period, 33 l of a 1.25 M Calcein AM solu-
ll per well. Then, 10
ll of the test
l
l per
l
l
tion which was protected from light were added to each well.
The fluorescence was measured immediately in constant time
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485 nm and an emission wavelength of 520 nm with a BMG
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3.6. Conformational analysis
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As no X-ray data on the compounds were available the energy
minimum conformers of the compounds were generated from
their sketched and minimized structures (MOE MMFF94x,
0.05 kcal molÀ1 ÅÀ1 gradient, MMFF94x charges) using simulated
annealing (Sybyl MMFF94 force field). Simulated annealing was
performed for 200 cycles, 1000 K initial temperature for heating
for 2000 fs equilibration, 0 K target temperature for 10,000 fs
annealing time, and exponential annealing function. The 200 local
minima obtained were then optimized by the MMFF94x force field
in MOE.
Acknowledgments
The authors thank Dr. Armin Buschauer for providing tariquidar
(Institute of Pharmaceutical Chemistry, University of Regensburg,