Synthesis and cytotoxic activity of new azepino[3′,4′:4,5]pyrrolo[2,1-a]isoquinolin-12-ones

Article abstract of DOI:10.1016/j.bmc.2009.01.056

A series of azepino[3′,4′:4,5]pyrrolo[2,1-a]isoquinolin-12-ones (3a-f), that were conformationally restricted analogs of lead compound 2, were designed as potential cytotoxic compounds and synthesized using a radical oxidative aromatic substitution reacti

Full text of DOI:10.1016/j.bmc.2009.01.056

Bioorganic & Medicinal Chemistry 17 (2009) 1849–1856
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and cytotoxic activity of new
azepino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-ones
*
Roberto Martínez , Martha Menes Arzate, Ma. Teresa Ramírez-Apan
Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior, Ciudad Universitaria, Coyoacán 04510, México D.F., Mexico
a r t i c l e i n f o
a b s t r a c t
series of azepino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-ones (3a–f), that were conformationally
Article history:
A
Received 31 October 2008
Revised 21 January 2009
Accepted 24 January 2009
Available online 31 January 2009
restricted analogs of lead compound 2, were designed as potential cytotoxic compounds and synthesized
using a radical oxidative aromatic substitution reaction as the key step. Compounds 3a–f were tested on
five tumor cell lines to determine the conformational requirements for biological activity of compound 2.
The results show that conformational restrictions on compound 2, generating the derivatives 3a–f, do not
appreciably reduce the cytotoxic activity of 2, although compound 3d (R = Br) showed good activity
against U-251 cells. Preliminary structure–activity relationship studies with these compounds revealed
the importance of halogens bonded to the isoquinoline moiety. Additionally, derivatives 3f (R = NO2)
and 3b (R = F) were cytotoxic to PC-3 and K-562 cells. However, none of the azepino[30,40:4,5]pyrrol-
o[2,1-a]isoquinolinones inhibited the enzymatic activity of CDK1/cyclin B, CDK5/p25, or GSK-3.
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
DLP
Radical oxidative
Azepinopyrroloisoquinolinones
Cytotoxic activity
1. Introduction
1.1. Design
Cancer is a leading cause of death worldwide, and the total
number of cancer deaths globally is projected to increase by 45%
from 2007 to 2030 (from 7.9 to 11.5 million), partly because the
world population is increasing and aging. During the same period,
newly diagnosed cases of cancer are also estimated to increase by
37%, from 11.3 million in 2007 to 15.5 million in 2030. In most
developed countries, cancer is the second leading cause of death
after cardiovascular disease, and epidemiological evidence points
to emergence of this trend in the less-developed world.1 Despite
questions on the future of synthetic small molecules as drugs,
and predictions of their downfall, the worldwide drug market is
still dominated by small molecules. At present, more than 80% of
clinical drug candidate development by the top 20 pharmaceutical
firms is based on small molecules,2 and an analysis of developmen-
tal trends through 2007 showed that about half are chemically
synthesized.3 Pharmacophore-based methods have shown promis-
ing results in efforts to find pharmacologically active compounds
relevant to a broad range of therapeutic areas. Drug design begins
with comparisons of active and inactive compounds. This suggests
how structural variations can change the biological activity of a
molecule and permits development of a hypothesis concerning
interactions between the molecule and its receptor. This approach
is known as molecular mimicry and is based on determining the
structural elements necessary for activity of the lead compound.4
As part of our ongoing interest in the development of novel het-
erocyclic compounds inhibiting the growth of cancer cells, we have
employed a structure–activity relationship analysis to generate
new active leads from compound 1.5 Our previous studies sug-
gested that a pyrroloazepine group and two aromatic groups, as
found in compound 2, are required for cytotoxic activity (Fig. 1).6
As the mechanism underlying the antiproliferative activity of com-
pound 2 is still unknown, further exploration of the pyrroloazepi-
none pharmacophore seemed advisable. Introduction of rigidity
into bioactive compounds has proven very useful in studying con-
formational requirements for biological activity. One of the most
efficient strategies for limiting the flexibility of compounds is the
incorporation of conformationally restricted rings.7 We therefore
utilized this approach in the development of new cytotoxic deriv-
atives from compound 2. The conformation represented by struc-
ture 2 could be constrained by elimination of the 1-phenyl
moiety and by attachment of the pyrrole nitrogen to the 2-phenyl
ring of compound 2 through an ethylene linker, yielding the azepi-
nopyrroloisoquinolinones 3. These compounds have part of the
pharmacophore found in the 7,12-dihydroindolo[3,2-d]benzaze-
pin-6(5H)-ones (paullones) (4), which have been shown to restrict
cellular proliferation by competitive inhibition of ATP binding.8
Here, we report the synthesis and cytotoxic activity of the 3-
R-azepino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-one derivatives
3a–f, which lack the 1-phenyl substituent and in which the 2-phe-
nyl group is part of a bicyclic moiety that binds to the pyrroleaze-
pine scaffold. Because compounds 3a-f are closer analogs of
* Corresponding author. Tel.: +52 56224441; fax: +52 56 16 22 03.
E-mail address: robmar@servidor.unam.mx (R. Martínez).
0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.01.056

1850
R. Martínez et al. / Bioorg. Med. Chem. 17 (2009) 1849–1856
1.3. Biological activity
We evaluated the abilities of compounds 3a–f and 9a–f to inhi-
bit the growth of PC-3 prostate cancer, U-251 central nervous sys-
tem cancer, K-562 leukemia, HCT-15 colon cancer, and MFC-7
breast cancer cells (Tables 1 and 2). Compound 3d (R = Br) showed
good inhibition of growth of all five cancer cell lines, with highest
activity against U-251 cells (IC50 = 18.14 1.02 lM). The chloro
derivative 3c showed moderate cytotoxicity against HCT-15 colon
cancer cells, with an IC50 value in the micromolar range
(IC50 = 39.65 1.0
lM). Compound 3b (R = F) was cytotoxic to K-
562 (IC50 = 33.74 3.8
l
M) and MCF-7 (IC50 = 54.51 3.4 M) can-
l
cer cells, suggesting that derivatization of the 3-R-isoquinoline
group with halogens yielded the best anticancer compounds. Inter-
estingly, the N-(4-halogen)-pyrroloazepinone derivatives 2a
(R = Cl) and 2b (R = Br) showed similar cytotoxic tendencies
against the five cancer cell lines.6 In addition, the nitro-derivative
3f (R = NO2) moderately inhibited the growth of three cancer cell
lines, PC-3 (IC50 = 25.79 3.2
lM), HCT-15 (IC50 = 29.50 3.2
lM),
and MCF-7 (IC50 = 43.14 4.4
l
M). By contrast, compounds 3a
(R = H) and 3e (R = OMe) did not inhibit proliferation of any of
the five cancer cell lines tested.
The activities of kenpaullone 4a and alsterpaullone 4b were also
tested against the five cancer cell lines, and the cytotoxic activities
of these compounds were compared with those of compounds 3a–
f. The most active inhibitors of proliferation of PC-3 prostate cells
Figure 1.
were compounds 3d (R = Br; IC50 = 33.56 2.8
lM) and 3f
(R = NO2; IC50 = 25.79 3.2 M). Interestingly, kenpaullone 4a
l
paullones than are the compound 2 derivatives, we hypothesized
that they would exhibit more potent antiproliferative activity.
We also report on the CDK1/cyclin B, CDK5/p25, and GSK-3
inhibitory activities of these compounds. As intermediate com-
pounds 9a–f are chemically connected to the final products, sug-
gesting that they may have pharmacological properties in
common, the cytotoxic activities of the intermediates were also
evaluated.
had no activity against the PC-3 line, whereas alsterpaullone 4b
was over 20-fold more potent than were compounds 3d and 3f.
Similarly, the bromine derivative compound 3d was cytotoxic to
U-251 central nervous system cancer cells and 4a kenpaullone
was inactive, whereas alsterpaullone 4b was over 25-fold more po-
tent than was compound 3d (IC50 = 0.68 0.02
1.02 M). Compounds 3b (IC50 = 33.74 3.8
(IC50 = 36.12 3.6 M) were more active than was compound 4a
lM vs IC50 = 18.14
l
lM) and 3d
l
against the K-562 leukemia cell line, whereas alsterpaullone 4b
was over 20-fold more potent than were either compounds 3b or
1.2. Chemistry
3d (IC50 = 1.58 0.3
pounds 3c, 3d, and 3f against the HCT-15 colon cancer cell line
(IC50 values in the low micromolar range) were similar to that of
lM). The antiproliferative activities of com-
Construction of intermediates 9a–f was the key step in obtain-
ing compounds 3a–f. The bond between the C-2-phenyl group
and the N-ethyliodide moiety can be formed via C-2 radical oxida-
tive aromatic substitution using lauroyl peroxide (DLP) as initiator
and oxidant. The organic peroxides have been shown to be excel-
lent reactants in radical aromatic substitution because they can
act as both initiators and oxidants. Using peroxides avoids the
toxicity of heavy metals present as organotin derivatives, and pre-
vents premature reduction of the intermediate radical.9 We
started with synthesis of the 1,4-dicarbonyl compounds 6a–f, as
described previously10 (Scheme 1). These compounds were con-
densed with 2-aminoethanol to generate the pyrrole derivatives
7a–f. These derivatives were reacted with triphenylphosphine
and iodine to obtain the desired iodo compounds 8a–f in moder-
ate overall yields. Cyclization of these iodides under the condi-
tions described above gave excellent yields of the tetracyclic
compounds 9a–f with the p-fluoro compound 9b being generated
most efficiently.11 Compounds 9a–f were quantitatively oximated
with hydroxylamine hydrochloride in ethanol, in the presence of
sodium carbonate, to yield the oximes 10a–f. Finally, regioselec-
tive Beckmann rearrangement of the syn- and anti-oximes 10a–f
yielded the pyrroloazepinoisoquinolines 3a–f as the sole products.
The group that migrates in the Beckmann rearrangement is
thought to be that anti to the hydroxyl group. However, under
the reaction conditions used, we have verified that the anti oximes
of indolones undergo isomerization to the syn oximes prior to
migration.6,12
compound 4a (IC50 = 23.86
was over 50-fold more potent (IC50 = 0.62 0.05
derivatives compounds 3b (IC50 = 54.51 3.4
43.15 3.6 M), and the nitro derivative compound 3f (IC50
2.9
lM), whereas compound 4b
l
M). The halogen
lM) and 3d (IC50
=
=
l
43.14 4.4), were effective in killing breast cancer cells (MFC-7)
with moderate cytotoxicity (IC50 values in the micromolar range),
and compound 4a was once again inactive, whereas compound
4b was over 35-fold more potent than were compounds 3b, 3d,
and 3f (IC50 = 1.24 0.1
and 3f did not inhibit the protein kinase activities of CDK1/cyclin
B, CDK5/p25, or GSK-3 CDK1 (IC50 > 10
M),13 suggesting that an-
other molecular mechanism is involved in the cytotoxic behavior
of compounds 3a–f.
Of the group of tetrahydroindolo[2,1-a]isoquinolin-11(8H)-
ones 9a–f, the most active were compound 9c (R = Cl) against
K-562 cells (IC50 = 10.21
(R = NO2) against HCT-15 cells (IC50 = 16.69 1.4
these activities are moderate, the potencies of these compounds
were high compared to that of kenpaullone 4a, which had an
IC50 against K-562 cells of over 100
HTC-15 cells of 23.86 2.9 M. However, the potencies of com-
pounds 9c and 9f were lower than that of alsterpaullone 4b. By
contrast, compound 9d (R = Br) showed moderate activity against
all five cancer cell lines, particularly K-562 (IC50 = 23.86
lM). Unfortunately, compounds 3b, 3d,
l
0.1
lM), followed by compound 9f
lM). Although
lM and an IC50 against
l
1.9
lM) and MCF-7 (IC50 = 34.96 2.8 M). Likewise, compound
l

R. Martínez et al. / Bioorg. Med. Chem. 17 (2009) 1849–1856
1851
Scheme 1. Synthesis of pyrroloazepinoisoquinoline derivatives 3a–f. Reagents and conditions: (i) K2CO3, 4-RC6H4COCH2Br, CHCl3, rt, 48 h; (ii) 2-aminoethanol, AcOH, reflux,
12 h; (iii) Ph3P, I2, imidazole, CH2Cl2; (iv) dicumyl peroxide, chlorobenzene, reflux; (v) NH2OHꢀHCl, AcONa, EtOH, reflux, 3 h; (vi) PPA, 80 °C, 3 h.
Table 1
The IC50 values (
l
M) of compounds 3a–f to the five cancer cell linesa
Compd
R
PC-3 (prostate)
U-251 (CNS)
K-562 (leukemia)
HCT-15 (colon)
MCF-7 (breast)
3a
3b
3c
3d
3e
3f
2a
2b
4a
4b
H
F
Cl
Br
OMe
NO2
Cl
Br
Br
>100
>100
>100
33.56 2.8
>100
25.79 3.2
17.44 1.87
21.1 2.1
>100
>100
>100
>100
18.14 1.02
>100
>100
33.74 3.8
>100
36.12 3.6
>100
>100
15.10 3.9
>100
>100
>100
>100
>100
54.51 3.4
>100
43.15 3.6
>100
43.14 4.4
51.66 3.6
>100
39.65 1.0
25.58 2.4
>100
29.50 3.2
20.04 4.7
>100
>100
28.82 3.7
25.6 4.5
>100
23.86 2.9
0.62 0.05
>100
1.24 0.1
NO2
1.12 0.01
0.68 0.02
1.58 0.3
a
Values are means of three experiments, (>100, not active).
Table 2
The IC50 values (
l
M) of compounds 9a–f to the five cancer cell linesa
Compd
R
PC-3 (prostate)
U-251 (CNS)
K-562 (leukemia)
HCT-15 (colon)
MCF-7 (breast)
9a
9b
9c
9d
9e
9f
H
F
Cl
Br
OMe
NO2
Br
>100
>100
>100
50.57 4.3
>100
>100
>100
1.12 0.01
>100
>100
>100
53.33 3.7
>100
>100
>100
0.68 0.02
36.83 2.7
27.22 0.3
10.21 0.1
23.86 1.9
57.50 1.3
73.68 7.8
>100
>100
>100
>100
41.82 0.03
34.96 2.8
>100
>100
>100
1.24 0.1
65.53 0.9
24.81 10
35.99 3.7
>100
16.69 1.4
23.86 2.9
0.62 0.05
4a
4b
NO2
1.58 0.3
a
Values are means of three experiments, (>100, not active).
9a (R = H) was active against K-562 cancer cells (IC50 = 36.83
2.7 M). All of derivatives 9a–f were effective in killing leukemia
1.4. Conclusion
l
cells (K-562) with moderate cytotoxicity in the micromolar
range; compound 4a was inactive and alsterpaullone 4b was
only about 7-fold more potent than was the most active azepi-
The results presented here indicate that conformational restric-
tions on compound 2, generating the derivative compounds 3a–j,
do not appreciably reduce the cytotoxic activity of 2, although
one compound (3d, R = Br) is active mainly against U-251 cells.
In addition, derivatives 3f (R = NO2) and 3b (R = F) were cyto-
nopyrroloisoquinoline 9c (R= Cl) (IC50 = 1.58
= 10.21 2.7 M).
0.1 lM vs. IC50
l

1852
R. Martínez et al. / Bioorg. Med. Chem. 17 (2009) 1849–1856
toxic to PC-3 and K-562 cells. Comparison of results using com-
pounds 3a–f and 9a–f clearly shows that the action mechanisms
of these groups are different, although a preliminary structure–
activity relationship analysis for these compounds also revealed
the importance of halogens in the benzene ring of the isoquinoline
moiety. Likewise, the position of the amide moiety in compounds
3a–g may also play an important role in determining relative activ-
ities. To test this hypothesis, compounds such as compound 11 will
be synthesized and evaluated. Our results indicate that these new
cytotoxic compounds may be useful as lead for the development of
novel anticancer agents. Therefore, future studies will focus on
determining the mechanism by which these new compounds inhi-
bit tumor growth and on testing of combinations of appropriately
positioned substituents.
1321, 1252, 1040, 814; 1H NMR (200 MHz, CDCl3) d 1.05 (s, 6H),
2.32 (s, 4H), 3.99 (s, 2H), 7.46 (dd, J = 1.7, 8.6 Hz, 2H), 8.15 (dd,
J = 1.9, 8.6 Hz, 2H); 13C NMR (75 MHz, CDCl3) d 28.2, 31.2, 31.7,
53.9, 106.3, 129.2, 131.2, 133.4, 141.3, 196.1, 202.3, 203.1; MS
(EI) m/z (rel intensity) 292 (M+, 18), 139 (100).
2.2.1.4. 2-[2-(4-Bromophenyl)-2-oxoethyl]-5,5-dimethylcyclo-
hexane-1,3-dione (6d). Yield 75% as a white solid; mp 154–155 °C;
IR (KBr, cmꢁ1
) mmax 2960, 2925, 2653, 1694, 1581, 1556, 1384,
1234, 1151, 1042, 809; 1H NMR (200 MHz, CDCl3) d 1.05 (s, 6H),
2.32 (s, 4H), 3.98 (s, 2H), 7.60–7.65 (m, 2H), 8.07 (dd, J = 1.9, 8.7
Hz, 2H); 13C NMR (75 MHz, CDCl3) d 28.2, 31.6, 31.7, 48.7, 53.9,
106.3, 129.2, 131.2, 133.4, 141.3, 196.1, 202.3, 203.1; MS (EI) m/z
(rel intensity) 336 (M+, 38), 183 (100).
2.2.1.5.
2-[2-(4-Methoxyphenyl)-2-oxoethyl]-5,5-dimethylcy-
2. Experimental
2.1. Chemistry
clohexane-1,3-dione (6e). Yield 50% as a white solid; mp 151–
153 °C; IR (KBr, cmꢁ1
) mmax 3155, 2961, 2912, 2835, 1677, 1603,
1379, 1251, 1168, 1037, 822; 1H NMR (200 MHz, CDCl3) d 1.04
(s, 6H), 2.31 (s, 4H), 3.88 (s, 3H), 3.97 (s, 2H), 6.95 (dd, J = 2.0, 8.8
Hz, 2H), 8.19 (dd, J = 2.0, 8.8 Hz, 2H); 13C NMR (75 MHz, CDCl3) d
28.2, 31.1, 31.3, 53.9, 55.6, 106.7, 132.5, 132.6, 164.9, 202.1,
203.4; MS (EI) m/z (rel intensity) 288 (M+, 24), 135 (100).
Melting points were determined using a Melt-Temp II melting
point apparatus and are uncorrected. IR spectra were recorded
using a Nicolet FT Magna-IR 750 spectrometer. The 1H and 13C
NMR spectra were recorded using Varian Gemini 200 and UNITY-
300 spectrometers, respectively, in deuterated chloroform solution
containing tetramethylsilane as the internal standard with chemi-
cal shifts (d) expressed downfield from TMS. Mass spectra were ob-
tained using AX505-HA and SX-100 Jeol mass spectrometers.
Reaction mixtures and chromatography fractions were concen-
trated using a rotary evaporator (ca. 20 °C/20 Torr). For column
chromatography, the Merck Silica Gel 60 F254 was employed.
Commercial grade reagents were used without further purification
except where indicated.
2.2.1.6. 5,5-Dimethyl-2-[2-(4-nitrophenyl)-2-oxoethyl]cyclo-
hexane-1,3-dione (6f). Yield 81% as a yellow-orange solid; mp
180–182 °C; IR (KBr, cmꢁ1
)
mmax 3077, 2956, 2907, 2662, 1696,
1565, 1530, 1385, 1350, 1042, 846, 746, 1H NMR (200 MHz,
CDCl3 + DMSO)
d 1.07 (s, 6H), 2.32 (s, 4H), 3.95 (s, 2H),
8.13 (d, J = 8.9 Hz, 2H), 8.28 (d, J = 8.9 Hz, 2H); 13C NMR
(75 MHz, CDCl3) d 27.8, 31.6, 32.9, 46.1, 53.2, 107.6, 123.0,
128.6, 141.6, 149.3, 196.7; MS (EI) m/z (rel intensity) 303
(M+, 94), 151 (100).
2.2.2. General procedure for the synthesis of 1-(2-
hydroxyethyl)-6,6-dimethyl-2-phenyl-1,5,6,7-tetrahydro-4H-
indol-4-ones (7a–f)
2.2. Synthetic procedure
2.2.1. General procedure for the synthesis of 5,5-dimethyl-2-(2-
oxo-2-phenylethyl)cyclohexane-1,3-diones, 6a–f
To a vigorously stirred suspension of triketone 6 (1 g, 3.88
mmol) in acetic acid (5 mL) was added ethanolamine (0.36 mL,
5.82 mmol). The resulting slurry was heated at 60 °C under a
N2 atmosphere until no more starting material was observed
by TLC, and then the reaction mixture was allowed to warm to
room temperature and poured into ice-water (10 mL). The solid
was filtered and washed with cooled water. The crude mixture
was purified by column chromatography on neutral aluminum
oxide using a gradient of 30–50% AcOEt in hexane as eluent.
Evaporation of the collected fractions gave 1-(2-hydroxy-
ethyl)indolones 7a–f.
A slurry of dimedone (0.01 equiv), chloroketone (0.01 equiv),
and anhydrous potassium carbonate (0.01 equiv) in chloroform
was stirred at room temperature for 48 h. The mixture was then fil-
tered; the insoluble salts were dissolved in water and the filtered
solution was made acidic with concentrated HCl. The precipitate
was filtered off, washed with water, and crystallized from aqueous
alcohol.
2.2.1.1. 5,5-Dimethyl-2-(2-oxo-2-phenylethyl)cyclohexane-1,3-
dione (6a). Yield 40% as a white solid; mp 171–173 °C; IR (KBr,
cmꢁ1
) mmax 2961, 2659, 2577, 1685, 1569, 1383, 1252, 1039, 753;
2.2.2.1. 1-(2-Hydroxyethyl)-6,6-dimethyl-2-phenyl-1,5,6,7-tet-
1H NMR (200 MHz, CDCl3) d 1.04 (s, 6H), 2.32 (s, 4H), 4.03 (s,
2H), 7.45–7.55 (m, 3H), 7.55–7.67 (m, 2H); 13C NMR (75 MHz,
CDCl3) d 28.2, 31.4, 31.6, 53.9, 106.6, 128.9, 129.8, 134.6, 135.2,
197.4, 203.4; MS(EI) m/z (rel intensity) 258(M+, 38), 105(100).
rahydro-4H-indol-4-one (7a). Yield 64% as a white solid; mp
172–174 °C; IR (KBr, cmꢁ1
) mmax 3325, 2955, 2883, 1624, 1475,
1061, 779, 703; 1H NMR (300 MHz, CDCl3 + DMSO) d 1.16 (s,
6H), 2.32 (s, 2H), 2.79 (s, 2H), 3.54 (t, J = 6.2 Hz, 2H), 4.02 (t,
J = 6.1 Hz, 3H), 6.42 (s, 1H), 7.41 (m, 5H); 13C NMR (75 MHz,
CDCl3 + DMSO) d 28.2, 34.9, 35.9, 46.1, 51.5, 60.2, 104.6, 118.3,
127.2, 128.2, 128.8, 132.1, 135.4, 144.1, 192.4; MS (EI) m/z (rel
intensity) 283 (M+, 67), 183 (100); HRMS (FAB+): calcd for
C18H22O2N: 284.1651, found: 284.1642.
2.2.1.2. 2-[2-(4-Fluorophenyl)-2-oxoethyl]-5,5-dimethylcyclo-
hexane-1,3-dione (6b). Yield 67% as a white solid; mp 144–146
°C; IR (KBr, cmꢁ1
) mmax 2961, 2908, 2557, 1691, 1598, 1555,
1350, 1256, 1040, 832; 1H NMR (200 MHz, CDCl3) d 1.05 (s, 6H),
2.32 (s, 4H), 3.99 (s, 2H), 7.10–7.20 (m, 2H), 8.22–8.30 (m, 2H);
13C NMR (75 MHz, CDCl3) d 28.2, 31.3, 31.6, 53.9, 106.4, 116.4,
131.0, 132.9, 169.4, 195.9, 202.1, 203.4; MS (EI) m/z (rel intensity)
276 (M+, 36), 123 (100).
2.2.2.2.
2-(4-Fluorophenyl)-1-(2-hydroxyethyl)-6,6-dimethyl-
1,5,6,7-tetrahydro-4H-indol-4-one (7b). Yield 60% as a white so-
lid; mp 161–163 °C; IR (KBr, cmꢁ1
) mmax 3340, 2954, 2879,
1629,1481, 1058, 847; 1H NMR (300 MHz, CDCl3 + DMSO) d 1.16
(s,6H), 2.34 (s, 2H), 2.77 (s, 2H), 3.56 (t, J = 6.3 Hz, 2H), 3.98 (t,
J = 6.3 Hz, 2H), 4.36 (s, 1H), 6.43 (s, 1H), 7.11 (dd, J = 5.4, 8.7 Hz,
2H), 7.41 (dd, J = 5.4, 9.0 Hz, 2H); 13C NMR (75 MHz,
2.2.1.3. 2-[2-(4-Chlorophenyl)-2-oxoethyl]-5,5-dimethylcyclo-
hexane-1,3-dione (6c). Yield 60% as a white solid; mp 163–164;
IR (KBr, cmꢁ1
) mmax 3160, 2958, 2876, 1688, 1599, 1569, 1381,

R. Martínez et al. / Bioorg. Med. Chem. 17 (2009) 1849–1856
1853
CDCl3 + DMSO) d 28.2, 34.8, 35.9, 45.9, 51.4, 60.3, 104.7, 114.7,
115.0, 118.3, 128.1, 130.7, 130.8, 134.3, 143.9, 160.1, 163.3,
192.8; MS (EI) m/z (rel intensity) 301 (M+, 100), 201 (96); HRMS
(FAB+): calcd for C18H21FO2N: 302.1556, found: 302.1554.
2.2.3.1. 1-(2-Iodoethyl)-6,6-dimethyl-2-phenyl-1,5,6,7-tetrahy-
dro-4H-indol-4-one (8a). Yield 85% as a white solid; mp 143–45
°C; IR (KBr, cmꢁ1
) mmax 2952, 2923, 2867, 1656, 1471, 772, 705;
1H NMR (300 MHz, CDCl3) d 1.19 (s, 6H), 2.40 (s, 2H), 2.71 (s,
2H), 3.03 (t, J = 8.1 Hz, 2H), 4.23 (t, J = 8.1 Hz, 2H), 6.56 (s, 1H),
7.34–7.47 (m, 5H); 13C NMR (75 MHz, CDCl3) d 0.7, 28.9, 35.6,
36.4, 46.4, 51.9, 106.2, 119.7, 128.2, 128.9, 129.0, 132.0,135.4,
143.0, 193.6; MS (EI) m/z (rel intensity) 393 (M+, 100); HRMS
(FAB+): calcd for C18H21ONI: 394.0668, found: 394.0666.
2.2.2.3.
1,5,6,7-tetrahydro-4H-indol-4-one (7c). Yield 55% as a white so-
lid; mp 165–66 °C; IR (KBr, cmꢁ1
mmax 3255, 2958, 2870, 1627,
2-(4-Chlorophenyl)-1-(2-hydroxyethyl)-6,6-dimethyl-
)
1477, 1062, 820; 1H NMR (300 MHz, CDCl3) d 1.10 (s, 6H), 2.23
(s, 2H), 2.70 (s, 2H), 2.77 (s,1H), 3.69 (d, J = 6.3 Hz, 2H), 4.02 (t,
J = 6.2 Hz, 2H), 6.50 (s, 1H), 7.38 (m, 4H); 13C NMR (75 MHz, CDCl3)
d 28.7, 35.4, 36.7, 46.5, 51.5, 61.4, 106.1, 119.2, 128.9, 130.7, 130.8,
134.0, 135.1, 144.9, 194.1; MS (EI) m/z (rel intensity) 317 (M+, 98),
217 (100); HRMS (FAB+): calcd for C18H21ClO2N: 318.1261, found:
318.1251.
2.2.3.2. 2-(4-Fluorophenyl)-1-(2-iodoethyl)-6,6-dimethyl-
1,5,6,7-tetrahydro-4H-indol-4-one (8b). yield 79% as a white so-
lid; mp 140–141 °C; IR (KBr, cmꢁ1
) mmax 2952, 2925, 2861, 1658,
1469, 851; 1H NMR (300 MHz, CDCl3) d 1.19 (s, 6H), 2.39 (s, 2H),
2.67 (s, 2H), 3.02 (t, J = 7.9 Hz, 2H), 4.20 (t, J = 7.9 Hz, 2H), 6.53
(s, 1H), 7.14 (dd, J = 8.8, 2.9 Hz, 2H), 7.33 (dd, J = 8.7, 2.7 Hz, 2H);
13C NMR (75 MHz, CDCl3) d 0.5, 28.8, 35.6, 36.4, 46.3, 51.9, 106.4,
115.8, 116.1, 119.6, 130.8, 130.9, 134.2, 142.9, 193.5; MS (EI) m/z
(rel intensity) 411 (M+, 100); HRMS (FAB+): calcd for C18H20ONFI:
412.0582, found: 412.0582.
2.2.2.4. 2-(4-Bromophenyl)-1-(2-hydroxyethyl)-6,6-dimethyl-
1,5,6,7-tetrahydro-4H-indol-4-one (7d). Yield 40% as a white so-
lid; mp 159–160 °C; IR (KBr, cmꢁ1
) mmax 3258, 2953, 2872, 1628,
1475, 1063, 819; 1H NMR (300 MHz, CDCl3) d 1.09 (s, 6H), 2.21
(s, 2H), 2.69 (s, 2H), 2.88 (s,1H), 3.71 (t, J = 5.5 Hz, 2H), 4.02 (t,
J = 5.5 Hz, 2H), 6.49 (s, 1H), 7.32 (dd, J) 8.6, 1.9 Hz, 2H); 7.54 (dd,
J = 8.6, 1.8 Hz, 2H,); 13C NMR (75 MHz, CDCl3) d 28.7, 35.4, 36.6,
46.6, 51.3, 56.9, 61.3, 106.1, 119.0, 122.2, 131.0, 131.1, 131.8,
135.2, 145.4, 194.2; MS (EI) m/z (rel intensity) 361 (M+, 100) 363
(M++2, 99); HRMS (FAB+): calcd for C18H21BrO2N: 362.0756, found:
362.0742.
2.2.3.3.
1,5,6,7-tetrahydro-4H-indol-4-one (8c). Yield 82% as a white so-
lid; mp 125–126 °C; IR (KBr, cmꢁ1
mmax 2957, 2929, 2872, 1651,
2-(4-Chlorophenyl)-1-(2-iodoethyl)-6,6-dimethyl-
)
1472, 803; 1H NMR (300 MHz, CDCl3) d 1.19 (s, 6H), 2.39 (s, 2H),
2.70 (s, 2H), 3.03 (t, J = 7.9 Hz, 2H), 4.22 (t, J = 7.9 Hz, 2H), 6.56
(s, 1H), 7.28 (d, J = 8.7 Hz, 2H), 7.41 (d, J = 8.7 Hz, 2H); 13C NMR
(75 MHz, CDCl3) d 0.5, 28.8, 35.6, 36.4, 46.4, 51.9, 106.6, 119.8,
129.1, 130.1, 130.5, 134.1, 134.3, 143.2, 193.5; MS (EI) m/z (rel
intensity) 427 (M+, 100); HRMS (FAB+): calcd for C18H21ONClI:
428.0278, found: 428.0285.
2.2.2.5.
dimethyl-1,5,6,7-tetrahydro-4H-indol-4-one (7e). Yield 52% as
a white solid; mp 155–56 °C; IR (KBr, cmꢁ1
mmax 3331, 2953,
1-(2-Hydroxyethyl)-2-(4-methoxyphenyl)-6,6-
)
2897, 2835, 1635, 1483, 1250, 1180, 1030, 808; 1H NMR (300
MHz, CDCl3) d 1.12 (s, 6H), 2.28 (s, 2H), 2.62 (br s, 1H), 2.72 (s,
2H), 3.68 (t, J = 6.0 Hz, 2H), 3.85 (s, 3H), 4.01 (t, J = 5.7 Hz, 2H),
6.47 (s, 1H), 6.93 (dd, J = 9.0 Hz, 2H), 7.33 (dd,, J = 9.0 Hz, 2H);
13C NMR (75 MHz, CDCl3) d 28.7, 35.4, 36.7, 46.5, 51.2, 55.3, 61.6,
105.4, 114.0, 118.8, 124.6, 130.9, 136.2, 144.9, 159.4, 194.1; MS
(EI) m/z (rel intensity) 313 (M+, 100); HRMS (FAB+): calcd for
C19H24O3N: 314.1758, found: 314.1756.
2.2.3.4.
1,5,6,7-tetrahydro-4H-indol-4-one (8d). Yield 71% as a white so-
lid; mp 119–120 °C; IR (KBr, cmꢁ1
mmax 2955, 2871, 1654, 1466,
2-(4-Bromophenyl)-1-(2-iodoethyl)-6,6-dimethyl-
)
805; 1H NMR (300 MHz, CDCl3) d 1.19 (s, 6H), 2.39 (s, 2H), 2.70
(s, 2H), 3.03 (t, J = 7.9 Hz, 2H), 4.22 (t, J = 7.9 Hz, 2H), 6.56 (s, 1H),
7.23 (dd, J = 9.0, 1.8 Hz, 2H), 7.57 (dd, J = 8.8, 2.1 Hz, 2H); 13C
NMR (75 MHz, CDCl3) d 0.5, 28.8, 35.6, 36.4, 46.4, 51.9, 106.7,
119.8, 122.5, 130.4, 131.0, 132.1, 134.1, 143.3, 193.5; MS (EI) m/z
(rel intensity) 471 (M+, 06), 473 (M++2, 07), 83 (100); HRMS
(FAB+): calcd for C18H21ONBrI: 471.9773, found: 471.9782.
2.2.2.6.
1,5,6,7-tetrahydro-4H-indol-4-one (7f). Yield 60% as a yellow so-
lid; mp 184–86 °C; IR (KBr, cmꢁ1
mmax 3444, 2957, 2873, 1656,
1-(2-Hydroxyethyl)-6,6-dimethyl-2-(4-nitrophenyl)-
)
1595, 1516, 1475, 1342, 859; 1H NMR (300 MHz, CDCl3 + DMSO)
d 1.13 (s, 6H), 1.95 (s, 1H), 2.30 (s, 2H), 2.74 (s, 2H), 3.75 (t,
J = 5.7 Hz, 2H), 4.12 (t, J = 5.7 Hz, 2H), 6.66 (s, 1H), 7.64 (d, J = 9.0
Hz, 2H), 8.26 (d, J = 8.7 Hz, 2H); 13C NMR (75 MHz, CDCl3) d 28.7,
35.4, 36.8, 46.7, 51.6, 61.5, 108.0, 119.8, 124.0, 129.5, 134.1,
138.9, 146.0, 146.9, 193.9; MS (EI) m/z (rel intensity) 328 (M+,
80); 228 (100); HRMS (FAB+): calcd for C18H21O4N2: 329.1501,
found: 329.1508.
2.2.3.5.
1,5,6,7-tetrahydro-4H-indol-4-one (8e). Yield 72% as a white so-
lid; mp 134–135 °C; IR (KBr, cmꢁ1
max: 2955, 2969, 2840, 1652,
1-(2-Iodoethyl)-2-(4-methoxyphenyl)-6,6-dimethyl-
)
m
1469, 1414, 1248, 1171, 839; 1H NMR (300 MHz, CDCl3) d 1.19
(s, 6H), 2.39 (s, 2H), 2.69 (s, 2H), 3.03 (t, J = 7.4 Hz, 2H), 3.86 (s,
3H), 4.20 (t, J = 7.6 Hz, 2H), 6.50 (s, 1H), 6.96 (dd, J = 8.8, 2.0 Hz,
2H), 7.28 (dd, J = 8.8, 2.1 Hz, 2H); 13C NMR (75 MHz, CDCl3) d 0.8,
28.9, 35.5, 36.4, 46.4, 51.9, 55.3, 105.7, 114.2, 119.5, 124.3, 130.4,
135.2, 142.6, 159.6, 193.6; MS (EI) m/z (rel intensity) 423 (M+,
100), HRMS (FAB+): calcd for C19H23O2NI: 424.0773, found:
424.0774.
2.2.3. General procedure for the preparation of 1-(2-iodoethyl)-
6,6-dimethyl-2-phenyl-1,5,6,7-tetrahydro-4H-indol-4-ones (8a–
f)
2.2.3.6. 1-(2-Iodoethyl)-6,6-dimethyl-2-(4-nitrophenyl)-1,5,6,7-
Triphenylphosphine (1.5 equiv) and imidazole (1.5 equiv) were
dissolved in dry CH2Cl2 (1.5 mL/mmol). The mixture was cooled
in an ice bath, and iodine (1.5 equiv) was added with vigorous
stirring over 10 min. The resulting slurry was warmed to room
temperature, and a solution of hydroxyethyl tetrahydroindolone
7 (1.0 equiv) in CH2Cl2 was added dropwise over 15 min. The
mixture was stirred for 1 h under a N2 atmosphere. The solvent
was removed, and the crude residue was purified by column
chromatography on neutral aluminum oxide using hex/AcOEt as
eluent.
tetrahydro-4H-indol-4-one (8f). Yield 69% as a yellow solid; mp
150–151 °C; IR (KBr, cmꢁ1
) mmax 2954, 2926, 2867, 1656, 1599,
1518, 1470, 1342, 856; 1H NMR (300 MHz, CDCl3) d 1.20 (s, 6H),
2.42 (s, 2H), 2.73 (s, 2H), 3.06 (t, J = 7.5 Hz, 2H), 4.31 (t, J = 7.5
Hz, 2H), 6.72 (s, 1H), 7.53 (d, J = 9.0 Hz, 2H), 8.31 (d, J = 9.0 Hz,
2H); 13C NMR (75 MHz, CDCl3) d 0.5, 28.8, 35.6, 36.5, 46.6, 51.9,
108.6, 120.4, 124.3, 128.9, 133.1, 138.5, 144.6, 147.1, 193.4; MS
(EI) m/z (rel intensity) 438 (M+, 100); HRMS (FAB+): calcd for
C18H20O3N2I: 439.0519, found: 439.0521.

1854
R. Martínez et al. / Bioorg. Med. Chem. 17 (2009) 1849–1856
2.2.4. General procedure for the synthesis of 9,9-dimethyl-
5,8,9,10-tetrahydroindolo[2,1-a]isoquinolin-11(6H)-ones (9a–f)
To a degassed solution of the corresponding iodo derivative (1.0
equiv) in refluxing chlorobenzene (7 mL/mmol) was added dicum-
yl peroxide (1.5 equiv) portionwise (0.3 equiv/1.5 h). The reaction
was carried out under a N2 atmosphere for 7.5 h. The mixture
was then allowed to warm to room temperature and evaporated
to dryness. The crude residue was purified by column chromatog-
raphy on silica gel (AcOEt/hex).
2.2.4.6.
a]isoquinolin-11(6H)-one (9f). Yield 75% as a yellow crystalline
solid; mp 230–231 °C; IR (KBr, cmꢁ1
mmax 2954, 2879, 1653,
9,9-Dimethyl-3-nitro-5,8,9,10-tetrahydroindolo[2,1-
)
1575,1514, 1330, 807; 1H NMR (300 MHz, CDCl3) d 1.17 (s, 6H),
2.41 (s, 2H), 2.69 (s, 2H), 3.21 (t, J = 6.6 Hz, 2H), 4.06 (t, J = 6.6
Hz, 2H), 7.07 (s, 1H), 7.61 (d, J = 8.4 Hz, 1H), 8.10–8.15 (m, 2H);
13C NMR (75 MHz, CDCl3) d 28.8, 28.8, 35.3, 35.7, 40.9, 52.1,
103.9, 120.8, 123.2, 123.6, 129.5, 130.8, 134.8, 143.7, 145.6,
193.2; MS (EI) m/z (rel intensity) 310 (M+, 100); HRMS (FAB+):
calcd for C18H19O3N2: 311.1396, found: 311.1395.
2.2.4.1. 9,9-Dimethyl-5,8,9,10-tetrahydroindolo[2,1-a]isoquino-
lin-11(6H)-one (9a). Yield 82% as a white crystalline solid; mp
2.2.5. General procedure for the synthesis of 9,9-dimethyl-
5,8,9,10-tetrahydroindolo[2,1-a]isoquinolin-11(6H)-one (syn/
anti) oximes (10a–f)
148–150 °C; IR (KBr, cmꢁ1 mmax 2957, 2871, 1647, 1486, 762; 1H
)
NMR (300 MHz, CDCl3) d 1.16 (s, 6H), 2.38 (s, 2H), 2.65 (s, 2H),
3.08 (t, J = 6.6, 2H), 3.97 (t, J = 6.6, 2H), 6.87 (s, 1H), 7.12–7.28 (m,
3H), 7.53 (d, J = 7.5 Hz, 1H); 13C NMR (75 MHz, CDCl3) d 28.8,
35.3, 35.6, 41.0, 52.1, 99.9, 119.8, 122.9, 126.6, 127.5, 127.9,
128.7, 130.3, 131.2, 142.1, 193.3; MS (EI) m/z (rel intensity) 265
(M+, 81), 181 (100); HRMS (FAB+): calcd for C18H20ON: 266.1545,
found: 266.15435.
To a vigorously stirred solution of tetrahydroindolo[2,1-a]iso-
quinoline 9 (0.5 g, 1.88 mmol) in ethanol (10 mL) was added hydrox-
ylamine hydrochloride (0.458 g, 6.59 mmol) and sodium acetate
(0.85 g, 10.36 mmol) dissolved in a minimal quantity of water. The
resulting mixture was refluxed for 3 h, after which the mixture
was allowed to cool to room temperature. The crude product was fil-
tered, washed with cold water and purified by column chromatogra-
phy on silica gel using hexane–ethyl acetate 1:1 as eluent.
2.2.4.2.
a]isoquinolin-11(6H)-one (9b). Yield 91% as a white crystalline
solid; mp 179–180 °C IR (KBr, cmꢁ1
mmax 2955, 2869, 1646,
3-Fluoro-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
)
2.2.5.1. 9,9-Dimethyl-5,8,9,10-tetrahydroindolo[2,1-a]isoquino-
lin-11(6H)-one (syn/anti) oximes (10a). Yield 86% as a white so-
1482, 826; 1H NMR (300 MHz, CDCl3) d 1.16 (s, 6H), 2.39 (s, 2H),
2.65 (s, 2H), 3.08 (t, J = 6.8, 2H), 3.98 (t, J = 6.8, 2H), 6.81 (s, 1H),
6.89–7.01 (m, 2H), 7.49 (dd, J = 8.5, 2.6 Hz, 1H); 13C NMR (75
MHz, CDCl3) d 28.8, 28.9, 35.4, 35.6, 40.7, 52.1, 99.6, 114.7, 119.8,
124.6, 125.0, 130.6, 132.5, 142.1, 159.7, 163.0, 193.5; MS (EI) m/z
(rel intensity) 283 (M+, 100); HRMS (FAB+): calcd for C18H20ONF:
284.1451, found: 284.1452.
lid; mp 218–220 °C; IR (KBr, cmꢁ1
)
mmax 3251, 3155, 1651, 1H
´
NMR (300 MHz, CDCl3 + DMSO) d 1.10–1.20 (H-13,13), 2.30–2.31
(H-10), 2.55–257(H-8), 3.06 (H-5), 3.96 (H-6), 7.05–7.26 (Ar-H),
7.31 (H-12), 7.46–7.59 (Ar-H). 13C NMR (50 MHz, CDCl3 + DMSO)
d 7.6, 28.1, 31.7, 32.1, 34.5, 35.3, 42.4, 97.1, 104.6, 110.9, 121.5,
124.9, 126.4, 127.2, 128.5, 129.3, 132.7, 134.1, 148.2; MS (EI) m/z
(rel intensity) 280(M+, 100).
2.2.4.3.
a]isoquinolin-11(6H)-one (9c). yield 85% as a white crystalline
solid; mp 199–200 °C; IR (KBr, cmꢁ1
mmax 2955, 2868, 1647,
3-Chloro-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
2.2.5.2.
3-Fluoro-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
)
a]isoquinolin-11(6H)-one (syn/anti) oximes (10b). Yield 61% as
1481, 829; 1H NMR (300 MHz, CDCl3) d 1.16 (s, 6H), 2.39 (s, 2H),
2.65 (s, 2H), 3.07 (t, J = 6.7, 2H), 3.97 (t, J = 6.7, 2H), 6.85 (s, 1H),
7.18 (d, J = 1.99 Hz, 1H), 7.22 (dd, J = 2.16, 8.28 Hz, 1H), 7.44 (d,
J = 8.28 Hz, 1H); 13C NMR (75 MHz, CDCl3) d 28.7, 28.8, 28.8,
35.3, 35.6, 40.7, 52.1, 100.4, 119.9, 124.2, 127.2, 127.7, 127.9,
130.3, 131.8, 131.9, 142.4, 193.4; MS (EI) m/z (rel intensity) 299
(M+, 100); 301(M+ + 2, 31); HRMS (FAB+): calcd for C18H20ONCl:
300.1155, found: 300.1149.
a white solid; mp 236–237 °C; IR (KBr, cmꢁ1
)
mmax 3245, 3154,
1
´
1647, H NMR (300 MHz, CDCl3+DMSO) d 1.10 (H-13,13), 2.32–
234 (H-10), 2.54–256 (H-8), 3.06 (H-5), 3.96 (H-6), 6.89–6.95(Ar-
H), 7.27 (H-12), 7.41–7.52 (Ar-H); 13C NMR (75 MHz, CDCl3
+
DMSO) d 27.7, 28.4, 31.9, 32.3, 34.7, 35.5, 42.5, 97.8, 104.5, 111.1,
123.4, 123.5, 125.1, 125.2, 128.1, 131.7, 131.8, 134.3, 148.5; MS
(EI) m/z (rel intensity) 298 (M+, 100).
2.2.5.3.
3-Chloro-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
2.2.4.4. 3-Bromo-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
a]isoquinolin-11(6H)-one (syn/anti) oximes (10c). Yield 83% as
a]isoquinolin-11(6H)-one (9d). Yield 53% as a white crystalline
a white solid; mp 250–254 °C; IR (KBr, cmꢁ1
)
mmax 3242, 3152,
solid; mp 190–191 °C; IR (KBr, cmꢁ1
) mmax 2953, 2871, 1649,
1653, 1H NMR (300 MHz, CDCl3 + DMSO) d 1.08 (H-13,13), 2.24–
´
1470, 825; 1H NMR (300 MHz, CDCl3) d 1.16 (s, 6H), 2.38 (s, 2H),
2.65 (s, 2H), 3.07 (t, J = 6.8 Hz, 2H), 3.97 (t, J = 6.7 Hz, 2H), 6.87
(s, 1H), 7.34 (s, 1H), 7.38 (m, 2H); 13C NMR (75 MHz, CDCl3) d
28.6, 28.8, 35.3, 35.6, 40.7, 52.1, 100.5, 119.8, 120.0, 124.5, 127.7,
130.4, 130.6, 130.8, 132.2, 142.4, 193.4; MS (EI) m/z (rel intensity)
343 (M+, 76), 344 (M+ + 2, 75), 259 (100); HRMS (FAB+): calcd for
C18H20ONBr: 344.0650, found: 344.0661.
2.26 (H-10), 2.55–2.56 (H-8), 3.05 (H-5), 3.96 (H-6), 7.18 (Ar-H),
7.26 (H-12), 7.45 (Ar-H), 10.13 (br s); 13C NMR (75 MHz,CDCl3
+
DMSO) d 27.8, 28.1, 28.4, 32.1, 35.3, 42.7, 97.8, 105.4, 111.5,
123.0, 126.6, 127.4, 127.5, 127.5, 129.7, 131.6, 134.3, 147.6; MS
(EI) m/z (rel intensity) 314 (M+, 100).
2.2.5.4. 3-Bromo-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
a]isoquinolin-11(6H)-one (syn/anti) oximes (10d). Yield 88% as
2.2.4.5. 3-Methoxy-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
a white solid; mp 246–248 °C; IR (KBr, cmꢁ1
)
mmax 3238, 3114, 1649,
a]isoquinolin-11(6H)-one (9e). Yield 53% as a white crystalline
1H NMR (300 MHz, CDCl3 + DMSO) d 1.11–1.12 (H-13,13), 2.37–239
(H-10), 2.57–259 (H-8), 3.06 (H-5), 3.97 (H-6), 7.33–7.39 (Ar-H), 7.56
(H-12); 13C NMR (50 MHz, CDCl3+DMSO) d 27.5, 27.8, 28.2, 32.3,
35.2, 41.9, 97.9, 105.2, 110.7, 118.1, 123.3, 127.5, 128.2, 129.5, 130.1,
131.6, 135.7, 149.0; MS (EI) m/z (rel intensity) 358 (M+, 100).
´
solid; mp 201–203 °C; IR (KBr, cmꢁ1
) mmax: 2951, 2872, 2835,
1654, 1481, 1426, 1161, 1035, 826; 1H NMR (300 MHz, CDCl3) d
1.16 (s, 6H), 2.38 (s, 2H), 2.65 (s, 2H), 3.06 (t, J = 6.4 Hz, 2H), 3.82
(s, 3H), 3.96 (t, J = 6.8 Hz, 2H), 6.75 (s, 1H), 6.83 (dd, J = 2.8, 8.4
Hz, 2H), 7.46 (d, J = 8.4 Hz, 1H); 13C NMR (75 MHz, CDCl3) d 28.9,
29.2, 35.3, 35.6, 40.9, 52.1, 55.3, 98.4, 113.1, 113.3, 119.7, 121.8,
124.4, 131.4, 132.0, 141.8, 158.4, 193.4; MS (EI) m/z (rel intensity)
295 (M+, 85), 211 (100). HRMS (FAB+): calc for C19H22O2N:
296.1657, found: 296.1651.
2.2.5.5. 3-Methoxy-9,9-dimethyl-5,8,9,10-tetrahydroindolo[2,1-
a]isoquinolin-11(6H)-one (syn/anti) oximes (10e). Yield 93% as a
white solid; mp 249–250 °C; IR (KBr, cmꢁ1
) mmax 3246, 3155, 1646,

R. Martínez et al. / Bioorg. Med. Chem. 17 (2009) 1849–1856
1855
1H NMR (300 MHz, CDCl3 + DMSO) d 1.12–1.14 (H-13,13), 2.41–
2.43 (H-10), 2.61–2.63 (H-8), 3.06 (H-5), 3.99 (H-6), 6.77–6.84
(Ar-H), 7.14 (H-12); 7.46 (Ar-H); 13C NMR (75 MHz, CDCl3) d
27.0, 27.8, 32.2, 34.6, 40.8, 102.4, 109.1, 112.1, 120.6, 122.8,
129.3, 131.0, 136.3, 150.3, 157.1; MS (EI) 310.
47% as a white solid, mp 230–232 °C; IR (KBr, cmꢁ1
)
mmax 3402,
´
3274, 3187, 3044, 2957, 1638, 1472, 1314, 1150, 819; 1H NMR
(200 MHz, CDCl3 + DMSO-d6) d 1.13 (s, 6H), 2.71 (s, 2H), 3.08 (t,
J = 6.6 Hz, 2H), 3.13 (d, 2H), 3.95 (t, J = 6.6 Hz, 2H), 6.70 (s, 1H),
7.07 (br s, 1H), 7.34–7.41 (m, 3H); 13C NMR (75 MHz, CDCl3) d
26.3, 28.1, 28.3, 33.1, 40.4, 40.9, 51.8, 105.3, 114.8, 118.9, 122.3,
131.6, 133.7, 168.3; MS (EI) m/z (rel intensity) 358 (M+, 100). Anal.
Calc. for C18H19BrN2O: C, 60.18; H, 5.33; N, 7.80. Found: C, 60.20; H,
5.33; N, 7.82.
2.2.5.6.
9,9-Dimethyl-3-nitro-5,8,9,10-tetrahydroindolo[2,1-
a]isoquinolin-11(6H)-one (syn/anti) oximes (10f). Yield 80% as
a yellow solid, mp 239–240 °C; IR (KBr, cmꢁ1
) mmax 3240, 3152, 1652,
1H NMR (300 MHz, CDCl3 + DMSO) d 1.12–1.14 (H-13,13), 2.43–2.45
´
(H-10)), 2.62–263 (H-8), 3.17–3.24 (H-5), 4.00–4.09 (H-6), 7.55 (H-
2.2.6.5. 3-Methoxy-9,9-dimethyl-5,6,8,9,10,11-hexahydro-12H-
azepino[30,40:4,5]pyrrolo[2,1-a]iso quinolin-12-one (3e). Yield
12), 7.57–7.66 (Ar-H), 8.04–8.13 (Ar-H); 13C NMR (75 MHz, CDCl3
+
DMSO) d 27.4, 27.8, 28.1, 31.8, 32.4, 34.5, 35.2, 35.6, 41.5, 101.5,
108.7, 111.6, 122.8, 127.5, 127.8, 130.0, 134.5, 135.8, 138.0, 143.9,
144.2, 148.9, 150.9; MS (EI) m/z (rel intensity) 325 (M+, 100).
34% as a beige solid, mp 259–261 °C; IR (KBr, cmꢁ1
) mmax 3477,
3272, 3179, 3035, 2960, 1637, 1484, 1392, 1300, 1252, 1166,
816; 1H NMR (300 MHz, CDCl3) d 1.12 (s, 6H), 2.69 (s, 2H), 3.03–
3.10 (m, J = 6.6 Hz, 4H), 3.82 (s, 3H), 3.92 (t, J = 6.6 Hz, 2H), 6.50
(br s, 1H), 6.73 (d, J = 2.4 Hz, 1H), 6.82 (dd, J = 2.7, 8.4 Hz, 1H),
6.91 (s, 1H),7.48 (d, J = 8.4 Hz, 1H); 13C NMR (75 MHz, CDCl3) d
26.9, 29.2, 33.9, 40.4, 40.8, 55.3, 52.7, 103.7, 113.1, 114.9, 121.8,
124.3, 129.9, 131.5, 132.4, 133.2, 158.3, 169.4; MS (EI) m/z (rel
intensity) 310 (M+, 100), Anal. Calc. for C19H22N2O: C, 73.52; H,
7.14; N, 9.03. Found: C, 73.53; H, 7.14; N, 9.04.
2.2.6. General procedure for the synthesis of
azepino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-ones (3a–f)
To a mixture of phosphorus pentoxide (9 g) and phosphoric acid
(5 mL) was added the oxime (0.5 g). The resulting mixture was stir-
red and heated at 80 °C for 3 h. The mixture was allowed to warm
to room temperature and then poured into ice-water, after which it
was neutralized with sodium carbonate and extracted with meth-
ylene chloride (3 ꢂ 10 mL). The organic layer was dried over so-
dium sulfate. The solvent was removed in vacuo, and the crude
residue was purified by column chromatography on silica gel using
a gradient of 50–100% hexane–ethyl acetate as eluent.
2.2.6.6. 9,9-Dimethyl-3-nitro-5,6,8,9,10,11-hexahydro-12H-aze-
pino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-one (3f). Yield 25%
as a pale yellow solid, mp 263–265 °C; IR (KBr, cmꢁ1
) mmax 3405,
3268, 3182, 3041, 2963, 2927, 2878, 1616, 1513, 1470, 1324,
1089, 1025, 800; 1H NMR (300 MHz, CDCl3) d 1.14 (s, 6H), 2.73
(s, 2H), 3.10 (d, 2H), 3.20 (t, J = 6.3 Hz, 2H), 4.02 (t, J = 6.6 Hz,
2H), 6.48 (br s, 1H), 7.23 (s, 1H), 7.60 (d, J = 8.4 Hz, 1H), 8.11 (d,
J = 2.1 Hz, 1H), 8.14 (dd, J = 2.4, 8.4 Hz, 1H); 13C NMR (75 MHz,
CDCl3) d 26.9, 28.8, 34.2, 40.2, 40.6, 52.6, 109.4, 116.9, 123.0,
123.2, 123.5, 128.0, 130.3, 134.8, 135.4, 145.4, 168.7; MS (EI) m/z
(rel intensity) 325(M+, 100), Anal. Calc. for C18H19N3O3: C, 66.45;
H, 5.89; N, 12.9. Found: C, 66.46; H, 5.89; N, 12.93.
2.2.6.1.
no[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-one (3a). Yield 28% as
a white solid; mp 236–237 °C; IR (KBr, cmꢁ1
mmax 3275, 3188,
9,9-Dimethyl-5,6,8,9,10,11-hexahydro-12H-azepi-
)
3049, 2963, 1639, 1472, 1314, 1147, 764; 1H NMR (300 MHz,
CDCl3) d 1.13 (s, 6H), 2.72 (s, 2H), 3.09 (m, 4H), 3.96 (t, J = 6.6,
2H), 6.99 (s, 1H), 7.10–7.29 (m, 3H), 7.49–7.54 (m, 2H); 13C NMR
(75 MHz, CDCl3) d 26.2, 28.2, 33.1, 40.3, 51.8, 104.6, 113.2, 114.6,
122.1, 125.8, 126.8, 127.3, 128.1, 129.1, 129.5, 168.4; MS (EI) m/z
(rel intensity) 280 (M+, 100). Anal. Calc. for C18H20N2O: C, 77.11;
H, 7.19; N, 9.99. Found: C, 77.12; H, 7.19; N, 9.96.
2.3. Cytotoxic activity
Cancer lines were obtained from the National Cancer Institute
of the United States. The lines were those of colon cancer (HTC-
15), breast cancer (MCF-7), leukemia (K-562), central nervous sys-
tem cancer (U-251), and prostate cancer (PC-3). Cytotoxicity data
were determined using the protein-bound-sulforhodamine (SRB)
assay in microculture; the assay measures cell growth as described
by Monks14 method. Cell lines were cultivated in RPMI-1640 med-
2.2.6.2.
3-Fluoro-9,9-dimethyl-5,6,8,9,10,11-hexahydro-12H-
azepino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-one (3b). Yield
49% as a white solid, mp 216–218 °C; IR (KBr, cmꢁ1 mmax 3473,
3275, 3187, 3045, 2906, 2876, 1638, 1478, 1398, 1316, 1157,
818; 1H NMR (300 MHz, CDCl3) d 1.12 (s, 6H), 2.68 (s, 2H), 3.06
(m, J = 6.8, 4H), 3.93 (t, J = 6.8 Hz, 2H), 6.4 (br s, 1H), 6.80–7.02
(m, 2H), 6.97 (s, 1H), 7.5 (dd, J = 5.6, 8.6 Hz, 1H); 13C NMR (75
MHz, CDCl3) d 26.9, 29.0, 34.1, 40.3, 40.6, 52.6, 105.0, 114.3,
114.8, 124.4, 124.6, 133.0, 127.3, 128.1, 129.1, 129.5, 169.2; MS
(EI) m/z (rel intensity) 298 (M+, 100), Anal. Calc. for C18H19FN2O:
C, 72.46; H, 6.42; F, 6.37; N, 9.39. Found: C, 72.48; H, 6.42; N, 9.35.
ium with 10% (v/v) fetal bovine serum, 2
mL penicillin G, 100 mg/mL streptomycin sulfate, and 0.25
l
M
L
-glutamine, 100 IU/
g/mL
l
of amphotericin B (Gibco Grand Island, NY) Incubation was at 37 °C
in a 5%, CO2 atmosphere and 95% relative humidity. After growth,
cells were detached with a 0.1% (w/v) trypsin-EDTA solution,
counted in a hemocytometer, and diluted with complete medium
to 5 ꢂ 104 cells/mL (K-562, MCF-7), 7.5 ꢂ 104 cells/mL (U-251,
PC-3), or 10 ꢂ 104 cells/mL (HTC-15). Microtiter plate wells were
2.2.6.3.
3-Chloro-9,9-dimethyl-5,6,8,9,10,11-hexahydro-12H-
azepino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-one
(3c). Yield
mmax 3491,
65% as a white solid, mp 242–244 °C; IR (KBr, cmꢁ1
)
filled with 100 lL aliquots of cell suspension and incubated. After
3413, 3273, 3183, 3041, 2961, 2901, 1638, 1474, 1314, 1152,
820; 1H NMR (300 MHz, CDCl3) d 1.13 (s, 6H), 2.69 (s, 2H), 3.06
(t, J = 6.6 Hz, 2H), 3.10 (d, J = 5.4 Hz, 2H), 3.94 (t, J = 6.6 Hz, 2H),
6.59 (br s, 1H), 7.01 (s, 1H), 7.18 (d, J = 1.5 Hz, 1H), 7.22 (dd,
J = 2.1, 8.1 Hz, 1H), 7.47 (d, J = 8.1 Hz, 1H); 13C NMR (75 MHz,
CDCl3) d 26.9, 28.8, 33.9, 40.3, 40.7, 52.7, 105.7, 115.2, 124.2,
127.2, 127.7, 127.8, 131.5, 131.6, 133.9, 169.2; MS (EI) m/z (rel
intensity) 314(M+, 100), 316. Anal. Calc. for C18H19ClN2O: C,
68.67; H, 6.08; N, 8.90. Found: C, 68.65; H, 6.08; N, 8.92.
24 h cells were treated with logarithmically-diluted concentrations
of sample compounds. Test samples were dissolved in DMSO to
provide stock solutions (40
solutions of 100, 31, 10, 3.1 and 1
suspensions. After 48 h, cultures were fixed in situ by addition of
50 L 50% (w/v) trichloroacetic acid and incubated for 60 min at
4 °C. Cells were harvested by centrifugation, washed three times,
and dried. Cell pellets were suspended in 100 L 0.4% (w/v) SRB
lM) and were then diluted to yield
lM, for use in treatment of cell
l
l
solution in 1% (v/v) acetic acid for 30 min. Unlinked SRB was re-
moved by washing with 1% (v/v) acetic acid and protein-linked
2.2.6.4.
3-Bromo-9,9-dimethyl-5,6,8,9,10,11-hexahydro-12H-
dye was extracted with 10
solution, and quantitated by spectrometry at 515 nm. The IC50
lM tris(hydroxymethyl)methaneamine
azepino[30,40:4,5]pyrrolo[2,1-a]isoquinolin-12-one (3d). Yield

1856
R. Martínez et al. / Bioorg. Med. Chem. 17 (2009) 1849–1856
values were calculated according to Monks.14 Experiments were
conducted in triplicate, and means and standard errors calculated.
The compounds 9-bromo-7,12-dihydroindolo-[3,2-d][1]benzaze-
pin-6(5H)-one (kenpaullone 4a) and 9-nitro-7,12-dihydroindolo-
[3,2-d][1]benzazepin-6(5H)-one (alsterpaullone 4b) were acquired
from A.G. Scientific, Inc. San Diego,CA.15
6. Martínez, R.; Avila, J. G.; Ramírez, M. T.; Pérez, A.; Martínez, A. Bioorg. Med.
Chem. 2006, 14, 4007.
7. (a) Wermuth, C. G.. The Practice of Medicinal Chemistry. In 2nd Ed; Academic
Press: San Diego, 2003; (b) Ohmori, Y.; Yamashita, A.; Tsujita, R.; Yamamoto, T.;
Taniuchi, T.; Matsuda, A.; Shuto, S. J. Med. Chem. 2003, 46, 5326; (c) Vintonyak,
V. V.; Cal, M.; Lay, F.; Kunze, B.; Sasse, F.; Maier, M. E. Chem. Eur. J. 2008, 14,
3709; (d) Yamaguchi, K.; Kazuta, Y.; Hirano, K.; Yamada, S.; Matsuda, A.; Shuto,
S. Bioorg. Med. Chem. 2008, 16, 8875; (e) Sall, C.; Yapi, A.-D.; Desbois, N.;
Chevalley, S.; Chezal, J. M.; Tan, K.; Teulade, J. C.; Valentin, A.; Blache, Y. Bioorg.
Med. Chem. Lett. 2008, 18, 4666.
8. (a) Zaharevitz, D. W.; Gussio, R.; Leost, M.; Senderowicz, A. M.; Lahusen, T.;
Kunick, C.; Meijer, L.; Sausville, E. A. Cancer Res. 1999, 59, 2566; (b) Meijer, L.;
Thunnissen, A. M.; White, A. W.; Garnier, M.; Nikolic, M.; Tsai, L. H.; Walter, J.;
Cleverley, K. E.; Salinas, P. C.; Wu, Y. Z.; Biernat, J.; Mandelkow, E. M.; Kim, S. H.;
Pettit, G. R. Chem. Biol. 2000, 7, 51; (c) Kunick, C.; Lemcke, Th. Pharm. Ztg 2002,
147, 2428; (d) Kunick, C.; Lauenroth, K.; Wieking, K.; Xie, X.; Schultz, C.; Gussio,
R.; Zaharevitz, D.; Leost, M.; Meijer, L.; Weber, A.; Jørgensen, F. S.; Lemcke, T.
J. Med. Chem. 2004, 47, 22; (e) Wan, Y.; Hur, W.; Cho, C. Y.; Liu, Y.; Adrian, F. J.;
Lozach, O.; Bach, S.; Mayer, T.; Fabbro, D.; Meijer, L.; Gray, N. S. Chem. Biol.
2004, 11, 247.
Enzyme inhibition assays were conducted in the laboratory of
Professor L. Meijer following the protocol described in Ref. 13.
Acknowledgments
We thank Professor L. Meijer and Oliver Lozach, CNRS, Station
Biologique, Roscoff, France, for performing he kinase assays that
were supported by the Ministèrie de la Recherche/INSERM/CNRS
‘Molècules et Cibles Thèrapeutiques’ Programme. Financial support
from the DGAPA, UNAM (Project PAPIIT-IN213407), is gratefully
acknowledged. M.M.A thanks CONACYT for a scholarship and we
also thank R. Patiño, H. Rios, A. Peña, N. Zavala, L. Velasco, and J.
Pérez for technical assistance. Contribution No. 2648 from Instituto
de Química, UNAM.
9. (a) Osornio, Y. M.; Cruz-Almanza, R.; Jiménez-Montaño, V.; Miranda, L. D. Chem.
Commun. 2003, 2316; (b) Zard, S. Z. In Radicals in Organic Synthesis; Renaud, P.,
Sibi, M., Eds.; Wiley VCH: Weinheim, 2001; p 90.
10. Martínez, R.; López, G.; Ávila, J. G. J. Heterocycl. Chem. 1995, 32, 491.
11. Menes-Arzate, M.; Martínez, R.; Cruz-Almanza, R.; Muchowski, J. M.; Osornio,
Y.; Miranda, L. D. J. Org. Chem. 2004, 69, 4001.
12. Martínez, R.; López, G.; Ávila, J. G.; Nava-Salgado, V. O. Heterocycles 2000, 53,
557.
References and notes
13. (a) Leclerk, S.; Garnier, M.; Hoessel, R.; Marko, D.; Bibb, J. A.; Snyder, G. L.;
Greengard, P.; Biernat, J.; Wu, Y.-Z.; Mandelkow, E. M.; Eisenbrand, G.; Meijer,
L. J. Biol. Chem. 2001, 276, 251; (b) Leost, M.; Schultz, C.; Link, A.; Wu, Y.-Z.;
Biernat, J.; Mandelkow, E. M.; Bibb, J. A.; Snyder, G. L.; Greengard, P.;
Zaharevitz, D.; Gussio, R.; Senderovitz, A.; Sausville, E. A.; Kunick, C.; Meijer,
L. Eur. J. Biochem. 2000, 267, 5983.
14. Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.; McMahon, J.; Vistica, D.;
Warren, J. T.; Bokesch, H.; Kenney, S.; Boyd, M. R. J. Natl. Cancer Inst. 1990, 82,
1107.
1. WHO, http://www.who.int/features/qa/15/en/index.html.
2. Mullin, R. Chem. Eng. News 2004, 23.
3. Reichert, J. M.; Wenger, J. B. Drug Discovery Today 2008, 13, 30.
4. (a) Cohen, N. C. In Medicinal Chemistry Today and Tomorrow; Yamazaki, M., Ed.;
Blackwell Science: Oxford, 1997; p 15; (b) Cohen, N. C. In Trends in Medicinal
Chemistry; Van der Goot, H ., Domany, G., Pallos, L., Timmerman, H., Eds.;
Elsevier Science: Amsterdam, 1989; p 13.
15. (a) 9-Bromo-7,12-dihydroindolo-[3,2-d][1]benzazepin-6(5H)-one, kenpaullone,
A.G. Scientific, http://www.agscientific.com/Item/K1050.htm.; (b) 9-Nitro-
5. Martınez, R.; Avila-Zarraga, J. G.; Duran, M. E.; Ramírez, M. T.; Cañas, R. Bioorg.
Med. Chem. Lett. 2002, 12, 1675; National Cancer Institute, Compound
NSC70049, http://dtp.nci.nih.gov/docs/cancer/searches/cancer_open_compounds.
html, personal communication.
7,12-dihydroindolo-[3,2-d][1]benzazepin-6(5H)-one,
alsterpaullone,
A.G.
Scientific, http://www.agscientific.com/Item/A1136.htm.