Synthesis and biological evaluation of new penta- and heptacyclic indolo- and quinolinocarbazole ring systems obtained via Pd0 catalysed reductive N-heteroannulation

Article abstract of DOI:10.1039/c0ob00149j

A short route, involving a tetramolecular condensation reaction and a Pd/C catalyst-H₂-mediated reductive N-heteroannulation as the key-steps, has been found for the synthesis of some new penta- and heptacyclic indolo- (12), quinolino- (13) and indoloquinolinocarbazole (11) derivatives. HF-DFT (B3LYP) energy profiles and NMR calculations were carried out to help in the understanding of the experimental results. N-Alkylated indoloquinolinocarbazoles (16b, 17a, 17b and 18) were prepared and screened essentially toward some cancer-(G-quadruplex, DNA, topoisomerase I) and CNS-related (kinases) targets. Biological results evidenced 13 as a potent CDK-5 and GSK-3β kinases inhibitor, while di- or triaminopropyl-substituted indoloquinolinocarbazoles 17b or 18 targeted rather DNA-duplex or telomeric G-quadruplex structures, respectively.

Full text of DOI:10.1039/c0ob00149j

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PAPER
www.rsc.org/obc | Organic & Biomolecular Chemistry
Synthesis and biological evaluation of new penta- and heptacyclic indolo- and
quinolinocarbazole ring systems obtained via Pd0 catalysed reductive
N-heteroannulation†
Marie Laronze-Cochard,a Fabien Cochard,a Etienne Daras,a Ame´lie Lansiaux,b Bertrand Brassart,c
Enguerran Vanquelef,a,d Elise Prost,e, f Jean-Marc Nuzillard,e Brigitte Baldeyrou,b Jean-Franc¸ois Goosens,g
Olivier Lozach,h Laurent Meijer,h Jean-Franc¸ois Riou,i Eric Henon*j and Janos Sapi*a
Received 19th May 2010, Accepted 14th July 2010
DOI: 10.1039/c0ob00149j
A short route, involving a tetramolecular condensation reaction and a Pd/C catalyst–H2-mediated
reductive N-heteroannulation as the key-steps, has been found for the synthesis of some new penta- and
heptacyclic indolo- (12), quinolino- (13) and indoloquinolinocarbazole (11) derivatives. HF-DFT
(B3LYP) energy profiles and NMR calculations were carried out to help in the understanding of the
experimental results. N-Alkylated indoloquinolinocarbazoles (16b, 17a, 17b and 18) were prepared and
screened essentially toward some cancer-(G-quadruplex, DNA, topoisomerase I) and CNS-related
(kinases) targets. Biological results evidenced 13 as a potent CDK-5 and GSK-3b kinases inhibitor,
while di- or triaminopropyl-substituted indoloquinolinocarbazoles 17b or 18 targeted rather
DNA-duplex or telomeric G-quadruplex structures, respectively.
simple functionalised carbazole alkaloids2 display antibacterial,
antifungal, antiviral, anti-inflammatory and antitumor properties.
Among such structures, some have proved to be active against
metabolic3 and neurodegenerative4 disorders. Synthetic analogs
such as 3-aryl, 3,6-diaryl, or 4-alkoxycarbazole derivatives have
been developed for their antitumoral or antihypertensive activities,
respectively.5
Introduction
The carbazole or benzo[b]indole ring system is widely represented
in natural or synthetic biologically active substances.1 Many of the
aUniversite´ de Reims-Champagne-Ardenne, Institut de Chimie Mole´culaire
de Reims, CNRS UMR 6229, Groupe BSMA, UFR de Pharmacie, 51
rue Cognacq-Jay, F-51096 Reims, Cedex, France. E-mail: janos.sapi@
univ-reims.fr; Fax: +33(0)326918029
Heterocycle, especially aza-heterocycle-annulated natural car-
bazole alkaloids have been reported as potential anticancer
agents interacting with various targets.6 Ellipticine,7 a pyrido[4,3-
b]carbazole-type alkaloid has been identified as a strong DNA
interfering substance with topoisomerase II inhibitory properties.
Indolo[2,3-a]pyrrolo[3,4-c]carbazole rebeccamycin8 was found to
be a selective topoisomerase I inhibitor. Staurosporine was
identified as a potent but non-selective kinase inhibitor (protein
bLaboratoire de Pharmacologie Antitumorale, INSERM U-837, Centre
Oscar Lambret, Universite´ Nord de France, Institut de Recherches sur le
Cancer de Lille, IMPRT, 1 Place de Verdun, F-59045 Lille, Cedex, France
cUniversite´ de Reims-Champagne-Ardenne, CNRS UMR 6237, IFR 53
Biomole´cules, UFR des Sciences Exactes et Naturelles, BP 1039, F-51687
Reims, Cedex 2, France
dUniversite´ de Picardie Jules Verne, Laboratoire des Glucides, CNRS UMR
6219, Equipe THERA, Faculte´ de Pharmacie, 1 rue des Louvels, F-80037
Amiens, Cedex 1, France
10
kinase C9 (PKC), glycogen synthase kinase-3b (GSK-3b)) and
eUniversite´ de Reims-Champagne-Ardenne, Institut de Chimie Mole´culaire
de Reims, CNRS UMR 6229, Groupe IS, UFR des Sciences Exactes et
Naturelles, Bat. 18., BP 1039, F-51687 Reims, Cedex 2, France
also implicated in cell signaling process, while its hydroxy analog
UCN-0111 or granulatimide12 proved to be more selective cell-cycle
controlling agents inhibiting cyclin dependant kinases (CDKs)
and/or check-point kinase 1 (Chk1) (Fig. 1). Such polycyclic
natural products have also been selected as leads for structure-
based drug discovery programs (cancer, type II diabetes and CNS
diseases).13
Thus, simple aryl-substituted pyrrolo[3,4-c]carbazole analogs
(I) have been reported as inhibitors of checkpoint kinase Wee1,14
while aryl or heteroaryl fused counterparts (II) were found
to be potent against other kinases (CDK-1,15 CDK-5,15 GSK-
3b,15 Cyclin D1/CDK-4,16 Chk-113c,17). Closely related aryl-
indolylmaleimides18 (III) were developed as potent inhibitors of
GSK-3b, an enzyme involved with CDK-5 in Alzheimer disease.19
Isomer pyrrolo[2,3-a]carbazole derivatives (IV) have been iden-
tified as potential CDK-120 or Pim-kinase21 inhibitors. Substi-
tuted indolo[3,2-c]quinoline-type derivatives have been reported
as antineoplastic agents inducing G2/M cell cycle arrest and
apoptosis. Pentacyclic indolo[3,2-c]quinoline derivative (V)22 and
f
Universite´ Paris Descartes, Synthe`se et Structure de Mole´cules d’Inte´reˆt
Pharmacologique, CNRS UMR 8638, Faculte´ des Sciences Pharmaceu-
tiques et Biologiques, 4 avenue de l¢Observatoire, F-75270 Paris, Cedex 06,
France
gUniversite´ de Lille 2, EA 4034, Laboratoire de Chimie Analytique, Faculte´
des Sciences Pharmaceutiques et Biologiques, 3 rue du Professeur Laguesse,
BP 83, F-59006 Lille, Cedex, France
hCNRS, Protein Phosphorylation & Human Disease Group, USR3151,
Station Biologique, BP 74, F-29682 Roscoff, France
i
Re´gulation et Dynamique des Ge´nomes, INSERM U565, CNRS UMR
5153, USM 503, Muse´um National d’Histoire Naturelle, 43 rue Cuvier,
CP26, F-75231 Paris, Cedex 5, France
j
Universite´ de Reims-Champagne-Ardenne, Institut de Chimie Mole´culaire
de Reims, CNRS UMR 6229, Groupe BSMA, UFR des Sciences Exactes
et Naturelles, Bat. 18., BP 1039, F-51687 Reims, Cedex 2, France. E-mail:
eric.henon@univ-reims.fr; Fax: +33(0)326918497
† Electronic supplementary information (ESI) available: Computational
study of the key intermediate 8a; Synthesis and analytical data of 6b, 6c,
8b, 8c, 17a; Biological evaluation of 11, 13, 14, 16b, 17a–b, 18; References;
Copies of NMR Spectra of 6b–c, 8a–c, 10, 11, 12, 13, 14, 16b, 17a–b, 18;
Computational data of 8a. See DOI: 10.1039/c0ob00149j
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Fig. 1 Selected examples of natural and synthetic carbazoles displaying anticancer properties.
5-N-methylated quinoline (indolo[3,2-b]quinolinium) derivatives
(VI)23) have displayed potent telomeric G-quadruplex binding
affinities and in vitro telomerase inhibitory activities.
In addition to medicinal chemistry applications carbazole
derivatives have emerged as promising chemically stable mate-
rials for electronic devices.24 Substituted indolo[3,2-b]carbazole,
poly(indolo[3,2-b]carbazole) or poly(2,7-carbazole) derivatives
have been described as organic semiconductive materials, organic
thin-film transistors or components for photovoltaic cells.25
The prevalence of the carbazole core in medicinal chem-
istry and in organic materials has triggered considerable syn-
thetic efforts during the past years towards the prepara-
tion of this important structural unit. Apart from the well-
documented26 cyclisation of ortho-nitrogen-substituted diphenyl
derivatives via nitrene, recent development of organometallic
chemistry has provided new opportunities for metal-catalysed
synthesis of functionalised carbazoles. Thus, carbazoles have
been prepared by Pd-catalysed intramolecular arylation of N-(o-
halogenophenyl)aniline derivatives obtained by cross-coupling or
Buchwald–Hartwig amination method.27 Nozaki and colleagues
have developed a double N-arylation approach of 2,2¢-dihalogeno-
or 2,2¢-bis(trifluoromethansulfonyloxy) biphenyls28 while another
strategy consists of Pd-catalysed oxidative aryl–aryl coupling of
N-arylaniline derivatives.29
In this context development of simple synthetic approaches for
the preparation of new heterocycle fused carbazole ring systems
continues to be a challenge in organic chemistry.
Scheme 1 Proposed synthetic pathway to “angular” (4) and “linear” (5)
Some years ago we found that tetramolecular condensation
between 2-substituted indoles 1, Meldrum’s acid 2 and two equiv-
alents of aromatic aldehydes smoothly afforded the corresponding
1,3-diaryl substituted tetrahydrocarbazoles bearing a spiro Mel-
drum’s acid appendage on the C-3 atom.30 In continuation of
our work aiming at DNA-interfering carbazoles31 tetramolecular
condensation product 3a obtained with 2-(indol-2-yl)ethyl acetate
and o-nitrobenzaldehyde seemed to be a versatile intermediate
for the preparation of polycyclic aza-heterocycles (4, 5) by
simple functional group transformations such as intramolecular
cyclisation(s)/decarboxylation/aromatisation (Scheme 1).
heterocycle annulated carbazoles.
Herein we disclose our results leading to the preparation
of some new penta- and heptacyclic indolo- and quinolinocar-
bazoles by simple Pd–C/H2-mediated reductive cyclisation of
conformationally restricted o-nitrobiaryl derivatives, as the key
step. A DFT computational approach was used to confirm the
observed atropoisomers for the key intermediate 8a. N-Alkylation
reactions of indoloquinolinocarbazoles 11 and 14 and preliminary
biological in vitro evaluation of carbazoles (16b–18) were carried
out.
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We have never obtained the corresponding acid 6a probably due
to the highly congested and hydrophobic environment around the
C-3 carbon atom. Since thermolysis of Meldrum’s acid derivatives
is known to afford ketenes,34 tetramolecular condensation product
3a adsorbed on silicagel was heat◦ed under microwave irradiation35
Results and discussion
(i) Synthesis of 2,4-diarylcarbazole derivatives 8a–c
Firstly, diastereomerically homogenous tetrahydrocarbazole 3a30
was subjected to catalytic hydrogenation (10% Pd–C, H2, 4 bar)
prospecting for an intramolecular cyclisation between the in situ
formed aromatic amine function(s) and the Meldrum’s acid core,
followed by spontaneous decarboxylation.32 Such kind of domino
transformations have recently allowed the preparation of chi-
ral pyrano-, and pyrrolidino-fused tryptamines.33 Unfortunately,
catalytic hydrogenation under varying conditions led to a very
complex and inseparable mixture. After this unsuccessful reaction
we envisaged the transformation of the Meldrum’s acid moiety
to the corresponding monoacid function (6a) by hydrolysis and
decarboxylation (Scheme 2).34 With such an acid in hand two
different pentacyclic derivatives (4 and 5) could be envisaged by
aryl nitro group reduction and subsequent cyclisation.
R
(CEM Discoverꢀ (300 W), 200 C) aiming at the corresponding
carboxylic acid 6a, accessible via the postulated ketene 7a by
hydratation (Scheme 2). Surprisingly, instead of monoacid 6a 2,4-
diaryl substituted carbazole 8a was isolated (41%) as a result
of a thermally-induced decarboxylation–aromatisation process.
The 1H NMR spectrum of the chromatographically homogenous
derivative 8a featured great complexity. Thus, three separable
peaks have been found for the indole NH proton in 50, 29 and 21%
ratio at d = 10.60, 10.40 and 10.15 ppm, respectively, while only two
distinct signals were present for both ester methyl and methylene
protons. A careful analysis of the 1H NMR spectrum emphasised
the presence of three isomers. Methylene protons displayed two
groups of quadruplex at d = 4.76 and 4.08 ppm with an integrated
ratio of 1 to 4. Comparing the integration of NH protons to that of
the deshielded quadruplex (d = 4.76 ppm) we evidenced that this
latter could unambiguously be attributed to the minor isomer (a3 -
21%) while methylene protons of the two major isomers (a1 - 50%,
a2 - 29%) appeared at d = 4.08 ppm. Moreover, the corresponding
relative intensities have not been altered by prolonged heating
(180 ◦C, overnight). Altogether, these data give support to the
coexistence of several long lifetime isomers. A theoretical approach
was proposed to analyse these observed NMR results.
Neither lower microwave performance, nor use of alumina as
solid support allowed the isolation of monoacid 6a in the nitro
series. On the contrary, irradiation of tetramolecular adducts
3b,c adsorbed on silicagel afforded the corresponding acids 6b,c
(mixture of diastereomers) in 71–86% yield depending on the
microwave oven type. Higher irradiation temperature and longer
reaction time permitted a direct transformation of 3b,c to aromatic
carbazoles 8b,c in moderate yield. It is important to note that the
above mentioned decarboxylation–aromatisation could be carried
out without solvents and heavy metal catalysis36 meeting the
requirements of “green chemistry procedures”.
(ii) Computational study37,38,39 of the key intermediate 8a
The NMR spectra of 8a have provided evidence for the existence
of several stable isomers, even at high temperature (180 ◦C).
Complementary to the experimental work, computations were
performed to help in the understanding of these results. The
details of the quantum mechanical calculations are reported in
the Electronic Supplementary Information.†
Geometry variations of 8a were considered by examining the
internal rotation involving the nitrophenyl groups and the ester
group. Four conformers were found (Fig. 2) with the ethyl group
lying in the plane of symmetry of the carbazole tricyclic structure.
Due to steric hindrance the carbazole and nitrophenyl groups are
non-planar with rotation dihedral angles varying between 67 and
89◦. The H ◊ ◊ ◊ O distance between the carbazole NH group and an
Scheme 2 Preparation of 2,4-diaryl substituted carbazoles (8a–c).
An “angular” type heptacyclic system 4 could be generated
by a double cyclisation (paths 1 and 3), while a “linear” type
pentacycle 5 may result from a monocyclisation according to path
2 (Scheme 1). To this end, Meldrum’s acid containing derivative
3a was refluxed with sodium ethylate in ethanol, in aqueous
hydrochloric acid or in 30% aqueous sodium hydroxide, in vain.
˚
ester oxygen atom was found to be short (about 2 A) in the four
isomers, showing a significant interaction. The S and A notations
have been reserved for the structures with the two nitro functions
on the same side (syn) or on opposite sides (anti) of the carbazole
structure, respectively. The * notation refers to geometries with the
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a competition between simple reduction and reductive insertion
should be considered.
In fact, when 2,4-bis(o-nitrophenyl) substituted carbazole 8a
dissolved in a mixture of ethanol and toluene was submitted to
medium pressure hydrogenation conditions (10% Pd–C, H2 10
bar, room temperature, 25 h) a mixture of polycyclic compounds
was formed. Purification by column chromatography allowed
the isolation of two new heptacyclic and two new pentacyclic
derivatives (Scheme 3).
In the series of the so-called “angular” compounds (type 4)
reductive N-heteroannulation was accompanied by lactamisation
between the amino and ester groups affording a new indolo-
quinolinocarbazole 11 in 55% yield. It is interesting to note, that
in the first phase of the reaction lactim ether 10 was formed,
perhaps via an initial insertion of the nitrene moiety into the
carbonyl group, and 10 progressively converted into lactam11. The
expected indolo[3,2-b]carbazole derivative 12 was obtained with
reproducible yield (17%) resulting from reductive insertion on the
C-3 carbon atom. Less than 1% of pentacyclic quinolinocarbazole
13 was generally observed.
The structures of the new polycyclic systems were unam-
biguously ascertained by exhaustive NMR studies (HSQC,
HMBC, COSY, ROESY) and MS analyses. Thus, “angular-type”
molecules 10 and 11 were characterised by three deshielded
protons H-10, H-9, H-4 (observed between d = 8.80 and 9.30 ppm
in DMSO-d6) and by a strong ROE effect between H-10 and H-9
in ROESY experiment, while “linear” molecule 12 shows only one
downfielded proton (H-7: d = 8.75 ppm in DMSO-d6).
Analysis of the experimental results evidenced that in our
conditions, exhaustive reduction of nitro groups was avoided and
completely reduced derivative 13 was isolated in very low yield
(1%). Our DFT theoretical results suggested that the ratio of
11 and 12 could be related to the different atropoisomers of the
starting structure 8a. Actually, the geometrical requirement for the
nitrene insertion should be the proximity of the reacting nitrogen
atom with the targeted C–H or C O bonds. Even though the
detailed mechanism leading from 8a to 11 and 12 is unknown,
the analysis of the N–C distances in 8a shed light on the possible
conformer dependence of nitrogen insertion.
Fig. 2 Optimised 8a structures. A and S stand for anti and syn; the *
notation refers to the spatial orientation of the ester; geometries were fully
optimised within the HF-DFT(B3LYP/6-31G**) toluene PCM model.
ester carbonyl group directed in the opposite direction to the NH
function.
To achieve a deeper interpretation of these NMR data, chemical
shifts were subsequently computed for all stable conformers
relative to the absolute shielding constants of TMS (see ESI†).
The theoretical study combined with the experimental data
evidenced that the relative spatial orientation of the ester function
with respect to the neighbouring nitrophenyl group was respon-
sible for the peak separation observed in the NMR spectrum of
8a. It is interesting to note that the most stable atropoisomer (A)
gave signals for CH2 (or CH3, or NH) that presumably correspond
1
to the major conformer in the experimental H NMR spectrum.
Furthermore, the 50, 29, 21% ratio (from NMR data) fits well with
the relative stability determined for A, S and A* (within the DFT
precision). This is of special importance in view of understanding
the yields observed for products 11 and 12 (see below).
(iii) Synthesis of new polycyclic indolo- and quinolinocarbazole
derivatives (10, 11, 12, 13, 14, 15)
As can be seen from Table 1, conformers A and A* would favour
the formation of 11 rather than 12 because the reacting nitrogen
Respecting our objectives 2,4-bis(o-nitrophenyl) substituted car-
bazole 8a appeared to be a proper intermediate for the preparation
of both “angular” (4) and “linear” type (5) indole-heterocycles.
First of all, the well-documented trivalent phosphorous-mediated
(PPh3, (EtO)3P) reductive cyclisation was attempted.40 In accor-
dance with some reported observations,41 in our case deoxygena-
tion of 8a and subsequent nitrene insertion occurred with low
yields affording a multitude of side products. As an alternative
to Cadogan-reaction transition metal catalysed methods using
carbon monoxide as reductant have recently been developed for
the synthesis of indoles42 or carbazoles43 from o-nitrostyrenes or o-
nitrobiphenyls, respectively. However, in some cases formation of
side-products or harsh reaction conditions (high temperature and
high pressure of CO) have been reported, limiting the synthetic
application of this method.
˚
atom Na is significantly too far from the C-3 carbon atom (ꢀ 4 A)
˚
with respect to the ester carbon atom (ꢀ 3.6 A) in these species.
In contrast, conformer S may only favour the formation of 12
bringing the nitrogen atom Na closer to the C-3 carbon atom.
From this, 8a would give predominantly 11 as found experimen-
tally. These results provide additional support that several isomers
of 8a coexist in thermal equilibrium. This qualitative analysis
should be of course completed by a full study of the reaction
mechanism for a final clarification of these hypotheses. From a
mechanistic point of view, a palladium-bound nitrene intermediate
˚
Table 1 Selected distances (in A) in the B3LYP/6-31G** geometry of
molecule 8a
Distance
A
S
A*
The above mentioned inconveniences were circumvented by
Alajar´ın and colleagues reporting a Pd/C–hydrogen-mediated
reductive cyclisation of o-nitrophenylaryl derivatives.44 This simple
approach retained our attention even if under such conditions
O2Na ◊ ◊ ◊ C Oester
O2Na ◊ ◊ ◊ C–3
O2Nb ◊ ◊ ◊ C–3
3.65
3.97
3.36
4.09
3.70
3.98
3.62
3.93
3.42
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Scheme 3 Pd/C–H2-mediated reductive N-heteroannulation toward new indoloquinolino- (10,11), indolo-(12), and quinolinocarbazoles (13).
(9) can be involved similar to So¨derberg et al.’s proposition.45
However, a competitive pathway via partially deoxygenated in-
termediates may also be considered even if their insertion into
aromatic rings requires higher activation energy.46
(iv) Synthesis of N-alkylated indolo[2,3-
c]quinolino[4,3-a]carbazoles (16b) and (17a,b) and of
N-substituted 15-oxo-5H,14H-15,16-dihydroindolo[2,3-
c]quinolino[4,3-a]carbazole (18)
Fully aromatic heptacyclic derivative 14 was obtained by the
partial reduction of the lactam carbonyl group of 11 (Scheme 4).
As classical methods (POCl3, P4S10, Lawesson’s reagent, BH3–
Me2S) failed,◦we turned to LiAlH4 used in large excess. Lactam 11
heated at 60 C in DME was transformed to the corresponding
heptacycle 14 (71%) sometimes isolated together with a small
quantity of dihydropyridine derivative (15). Due to its instability
this latter could only be detected by MS analysis of the crude
mixture (peak (ESI) at m/z 358 [M+H]+). Nevertheless, the
mixture of 14 + 15 could be smoothly converted into 14 by heating
in the presence of p-chloranil.
In order to enhance the solubility and provide a better interaction
with DNA minor groove’s phosphates aminoalkyl side-chains have
been introduced according to the Scheme 5.
Two different side-chains were selected with tertiary amine
functions attached by two or three carbons to the aromatic ring-
system.
Treatment of heptacyclic derivative 14 with 6 eq. of sodium
hydride and 3-chloropropyldimethylamine hydrochloride led to
monoalkylated (16b) and dialkylated derivatives (17b) in 12 and
55% yield, respectively.
When 14 was reacted with sodium hydride and 2-
chloroethyldimethylamine hydrochloride under the same condi-
tions disubstituted compound 17a was isolated in 41% yield.
In the “lactam-series” 11 was submitted to alkylation using a
larger excess (9 eq.) of 3-chloropropyldimethylamine hydrochlo-
ride and sodium hydride to afford trialkylated derivative 18 (53%)
and an inseparable mixture of mono- and dialkylated derivatives
(approx. 17%).
(v) Biological multitarget screening of compounds (10–14 and
16b–18)
With these new polycyclic indole-heterocycles in hand we were
interested in conducting a preliminary biological screening toward
some CNS or cancer targets (kinases, telomeric G-quadruplex,
DNA-binding, cytotoxicity tests).
Scheme 4 Preparation of 5H,14H-indolo[2,3-c]quinolino[4,3-a]carbazole
(14).
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Table 2 Telomeric G-quadruplex stabilisations measured by FRET
FRET DTm/◦C
20 mM
Nos
10 mM
5 mM
16b
2.6
4.7
3.7
10.7
—
—
1.7
—
8.2
—
—
0.7
—
4.0
27.7
17a
17b
18
BRACO-19
—: Not Tested.
Table 3 Binding capacity to DNA and cytotoxicity measurements
a
b
Compounds
DTm
Kapp 105 M-1
IC50 (HL-60)/mMc
11
1.7
11.1
1.8
6.7
15.2
0.93 0.07
7.51 0.58
0.74 0.06
4.22 0.62
7.79 0.66
ND
10.5 1.98
ND
7.3 0.64
6.0 0.21
0.01 0.003
18
14
17a
17b
Camptothecin
a DTm = Tm of drug–DNA complex – Tm of CT-DNA alone in ◦C. Tm
measurements were performed in BPE buffer, in 1 cm quartz cuvettes at
◦
260 nm with a heating rate of 1 C min-1. The Tm values were obtained
Scheme 5 Synthesis of N-alkylated compounds 16b, 17a, 17b and 18.
from first-derivative plots (ratio 0.5). b Apparent affinity constant (Kapp
)
calculated from intrinsic fluorescence of compounds (1 mM in BPE;
lexc = 380 nm and lem = 510 nm) and CT-DNA titration n = 3. c drug
concentration (mM) that inhibits HL-60 cell growth by 50% after 72 h of
incubation ND: Not Determined.
Kinases inhibition. Firstly we tested the main penta- (12, 13)
and heptacyclic derivatives (10, 11, 14) and their four aminoalkyl-
substituted counterparts (16b, 17a, 17b and 18) as potential
inhibitors of DYRK1A, cyclin-dependent kinase-5 (CDK-5)10a,58
and glycogen synthase kinase-3b (GSK-3b)59 enzymes, implicated
in diverse CNS disorders. Among the screened products only 13
showed significant inhibitory activity for CDK-5 and GSK-3b
kinases with IC50 = 0.93 mM and IC50 = 1.2 mM, respectively (see
ESI†).
Telomeric G-quadruplex binding affinity measurement. The
telomeric G-quadruplex binding capacity of the four aminoalkyl-
substituted compounds (16b, 17a, 17b and 18) was firstly evalu-
ated by FRET (fluorescence resonance energy transfer) melting
experiments60 onto telomeric sequence (F21D) at a 20 mM ligand
concentration. At this concentration, mono- and disubstituted
compounds 16b, 17a, 17b showed modest stabilisation properties
toward G-quadruplex DNA (DTm value <5 ◦C), whereas the
◦
trisubstituted one 18 exhibited a significant DTm value (>10 C)
with a dose-response effect (Table 2). In the case of 18, the presence
of a third aminoalkylated branched chain might be responsible for
a specific position of the molecule allowing thus more interactions
with guanine tetrad and/or with potassium channel. However,
even if compound 18 showed the best affinity of the series
toward G-quadruplex DNA, it exhibited lower stabilisation ability
than the reference tri-substituted acridine derivative BRACO-196i
(DTm = 27.7 ◦C at 5 mM).
Fig. 3 Affinity measurements of di- and trisubstituted heptacyclic
carbazoles (17a, 17b and 18) toward DNA.
At the pH of serum polycyclic derivatives without protonable
functions (10, 11) or with aromatic amine groups (12, 13, 14)
showed no interaction with DNA. Introduction of dimethy-
laminoalkyl side-chain at indole or lactam nitrogen stabilizes the
DNA duplex against heat denaturation. Thus, simple 11 displayed
no significant effect on DNA stability, while its trisubstituted
analog 18 gave an important growth of Tm values with both
CT-DNA and poly(dATdT)2 and a ten fold Kapp value (Fig. 3,
Table 3).
DNA duplex binding affinity measurements. To evaluate the
extent of DNA binding, the variations of melting temperature of
DNA (calf thymus DNA or CT-DNA and poly(dAdT)2) were
measured and depicted in Fig. 3 and Table 3. Moreover, calf
thymus DNA binding affinities (Kapp) were measured from the
intrinsic fluorescence of compounds 11, 18, 14, 17a and 17b, and
DNA titration (Table 3).
4630 | Org. Biomol. Chem., 2010, 8, 4625–4636
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Similarly, disubstituted heptacyclic analogs (17a, 17b) proved to
be more efficient DNA duplex stabilizing agents. Comparison of
DTm data and apparent binding data of 17a and 17b evidenced the
importance of side chain length on the DNA stabilizing capacity,
with the dimethylaminopropyl moiety favored. Experiments car-
ried out at different poly(dAdT)2 ratios confirmed the affinity of
17a and 17b to DNA duplex.
on a Perkin-Elmer Spectrum BX FTIR instrument. NMR spectra
of compounds 6b–c, 8a–c, 10, 12, 13, 14, 16b, 17a,b and 18 were
recorded on a Bruker AC 300 spectrometer. All 1H NMR and 13
C
NMR spectra are reported in d units (ppm) using CDCl3 (diluted
with 10% of CD3OD for compound 16b (n = 2)) or DMSO-
d6 as solvent. Couplings expressed as s, br s, d, br d, ddd, t,
m correspond to singlet, broad-singlet, doublet, broad-doublet,
doublet of doublets of doublets, triplet and multiplet, respectively.
For compounds 8b and 8c, the CPD experiment was used to record
the 13C NMR spectra; in all other cases (compounds 6b–c, 8a, 10,
11, 12, 13, 14, 16b, 17a,b and 18), the J-Modulated one was chosen.
Partial or complete chemical shift assignment of compounds 6b–c,
8a–c, 10, 12, 13, 14, 16b, 17a,b and 18 was achieved using various
two dimensional NMR experiments (COSY, HSQC, HMBC).
For compound 11, mono and two dimensional exhaustive NMR
studies (COSY, HSQC, HMBC, ROESY) were realized using
a Bruker DR X 500 spectrometer and DMSO-d6 as solvent.
Mass spectra were recorded either on an MSQ ThermoFinnigan
apparatus using electronspray (ESI) in positive ion mode or on
a GCT Waters apparatus using electronimpact (EI, HRMS).
Microwave activated reactions were carried out in a Normalab
Inhibition of the topoisomerase I activity. In parallel, we
have investigated the interaction with human topoisomerase I.61
None of these derivatives induced topoisomerase I dependent
DNA cleavable complex stabilisation (see ESI†). Consequently,
topoisomerase I is not a real target of these molecules even if
some of them (17a, 17b and 18) showed interesting DNA binding
properties.
Cytotoxicity. Cytotoxicity measurements to determine the
drug concentration (in mM) required to inhibit HL-60 cell growth
by 50% have also been carried out with dimethylaminoalkyl-
substituted heptacyclic derivatives (17a, 17b and 18). From these
results it appeared that derivatives with strong affinity to DNA
showed cytotoxicity against HL-60 cell line at the micromolar
range (Table 3).
R
R
Analis Normatron 112ꢀ oven or in a CEM Discoverꢀ (300 W)
apparatus.
Conclusions
General procedure for the preparation of carbazoles (8a–c) by
microwave irradiation
In summary, we have found a short and efficient synthesis, based
on a Pd/C–H2-mediated reductive N-heteroannulation as the key
step, for the preparation of new penta- and heptacyclic indolo-,
quinolino- or indoloquinolinocarbazoles. From DFT molecular
orbital computations the signal separation observed for 8a could
be attributed to the specific spatial orientation of the ethyl group
relatively to the neighbouring nitrophenyl group. In addition, this
study might explain the observed conformer-dependent nitrene
insertion (8a → 11 or 12).
To a solution of the diastereomerically pure tetramolecular
condensation product 3a, 3b or 3c in CH2Cl2 was added silicagel.
After concentration under reduced pressure the residue was heated
under microwave irradiation in an open reaction vessel. Procedure
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A: Normalab Normatron 112ꢀ apparatus (1000 W); Procedure
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B: CEM Discoverꢀ apparatus. Purification of the crude solid by
column chromatography on silica gel afforded the corresponding
carbazoles 8a, 8b or 8c.
Preliminary screening led to the identification of new poly-
cyclic carbazole-type lead compounds displaying some intriguing
pharmacological properties. Thus, the pentacyclic compound 13
proved to be an efficient inhibitor of CDK-5 and GSK-3b kinases
implicated in Alzheimer disease.
Moreover, aminoalkyl-substituted heptacyclic carbazoles 17a,
17b and 18 interacted with DNA. Tris(aminopropyl)-substituted
analog 18 also interfered with telomeric G-quadruplex structure,
a promising new target in anticancer research. Pharmacomodula-
tions of the new skeletons identified are under investigation aiming
at structure–activity relationship studies and optimisation of the
inhibitory potency and/or selectivity.
2,4-Bis(2¢-nitrophenyl)-1-ethoxycarbonyl-9H-carbazole
mixture of three inseparable atropoisomers (a1/a2/a3).
Procedure B: starting from 3a (100 mg, 0.13 mmol) mixed
(8a)
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with silicagel (250 mg); Irradiation: CEM Discoverꢀ (200
◦C for 15 min); Purification (eluent: cyclohexane/CH2Cl2:
40/60→CH2Cl2) gave the orange foam 8a (32 mg, 41%), as a
mixture of three inseparable atropoisomers (1H NMR ratio of
a1/a2/a3: 50/29/21). nmax(film)/cm-1 3418, 1683, 1674, 1520,
1344, 1304, 1274, 1243, 1168 and 1141; dH(300 MHz; CDCl3;
Me4Si) 0.88 (6 H, 2 ¥ t, Ja1 and Ja2 7.1, 2 ¥ CO2CH2CH3)a1,a2
,
1.65 (3 H, t, J 7.1 Hz, CO2CH2CH3)a3, 4.08 (4 H, 2 ¥ quartet,
Ja1 and Ja2 7.1, 2 ¥ CO2CH2CH3)a1,a2, 4.76 (2 H, quartet, J 7.1,
CO2CH2CH3)a3, 6.78 (1 Ha3, d, J 8.0), 6.82–7.03 (4 H, m, 2 ¥
Ha2 and 2 ¥ Ha3), 7.19–7.89 (26 H, m, 10 ¥ Ha1, 8 ¥ Ha2 and 8
¥ Ha3), 8.09 (1 Ha2, d, J 8.0), 8.14–8.26 (2 H, m, 1 ¥ Ha2 and
1 ¥ Ha1), 8.30 (1 Ha3, d, J 7.9), 8.81–8.94 (3 H, m, 1 ¥ Ha2
and 2 ¥ Ha1), 10.15 (1 H, s, NH)a3, 10.40 (1 H, s, NH)a2 and
10.60 (1 H, s, NH)a1; dC(75 MHz; CDCl3; Me4Si) 13.3 (2 ¥
CO2CH2CH3)a1,a2, 14.6 (CO2CH2CH3)a3, 60.8 (CO2CH2CH3)a1,
60.9 (CO2CH2CH3)a2, 61.4 (CO2CH2CH3)a3, 107.7, 109.6, 110.5,
110.8, 111.0a1, 111.2a1, 111.3, 117.5a1, 118.9, 119.1, 119.2, 119.4a1,
119.7a1, 120.0, 120.8a1, 121.0, 121.1, 121.2, 121.3a1, 121.5, 121.8a1,
122.4a1, 122.5, 123.3a1, 123.7, 124.5 (2 ¥ CH)a1, 124.6, 124.8, 125.2,
125.8a1, 126.0, 126.5, 126.7, 126.8, 127.0a1, 127.9, 128.8a1, 129.4,
Experimental
Chemistry
General methods. All solvents were of reagent grade and,
when necessary, were purified and dried by standard meth-
ods. Reactions and products were routinely monitored by thin-
layer chromatography (TLC) on silica gel (Kieselgel 60 F254,
Merck). Column chromatography purifications were performed
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on CHROMAGELꢀ Silice 60 ACC 70–200 mm silica gel.
Melting points were determined on a Reichert Thermovar hot-
stage apparatus and are uncorrected. IR spectra were measured
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130.2, 130.8, 131.8, 131.9, 132.1a1, 132.3, 132.5, 132.7, 132.8,
133.0a1, 133.2a1, 133.3, 133.7, 134.1a1, 134.2, 134.6, 136.2a1, 136.3,
137.4, 137.8, 138.2, 138.3, 139.5a1, 140.2, 140.8, 141.1a1, 141.5,
148.4 (2 ¥ C–NO2), 149.3 (2 ¥ C–NO2), 149.4 (2 ¥ C–NO2)a1,
167.6 (2 ¥ CO2CH2CH3)a1,a2 and 168.3 (CO2CH2CH3)a3; m/z (EI)
481 (M+ 100%); HRMS calcd. for C27H19N3O6 481.1274, found
481.1248.
15-Oxo-5H,14H-15,16-dihydroindolo[2,3-c]quinolino[4,3-
a]carbazole (11)
◦
Yellow powder; mp >350 C; nmax (KBr)/cm-1 3447, 3433, 3381,
3026, 1661, 1613, 1591 and 1320; dH(500 MHz; DMSO-d6) 7.42 (1
H, t, J 7.5, 11-H), 7.46 (1 H, t, J 7.5, 8-H), 7.51 (1 H, ddd, J 1.9,
6.7 and 8.2, 3-H), 7.53 (1 H, t, J 7.5, 12-H), 7.58–7.67 (3 H, m,
1-H, 2-H and 7-H), 7.95 (1 H, d, J 8.1, 6-H), 8.04 (1 H, d, J 8.1,
13-H), 8.87 (1 H, d, J 8.2, 10-H), 8.96 (1 H, d, J 8.2, 9-H), 8.99 (1
H, d, J 8.2, 4-H), 11.89 (1 H, s, 5-NH), 12.09 (1 H, s, 16-NH) and
12.22 (1 H, s, 14-NH); dC(125 MHz; DMSO-d6) 108.3 (14b-C),
112.8 (13-CH), 112.9 (6-CH), 115.2 (9c-C), 116.3 (1-CH), 117.5
(4a-C), 119.3 (11-CH), 119.5 (4b-C), 119.9 (8-CH), 120.7 (9d-C),
121.4 (9b-C), 121.5 (9a-C), 122.7 (3-CH), 122.7 (10-CH), 123.3 (9-
CH), 125.2 (12-CH), 126.0 (4-CH), 126.4 (7-CH), 128.8 (2-CH),
129.6 (4c-C), 133.5 (14a-C), 136.5 (16a-C), 139.8 (13a-C), 141.9
(5a-C) and 162.4 (CO); m/z (EI) 373 (M+ 100%); HRMS calcd.
for C25H15N3O 373.1215, found 373.1229.
In some cases purification by column chromatography of the
crude mixture of three atropoisomers of 8a allowed the isolation
of a small quantity of the pure atropoisomer 8a1 as a pale orange
foam.
n
max(film)/cm-1 3426, 1680, 1522, 1349, 1307, 1277, 1246,
and 1171; dH(300 MHz; CDCl3; Me4Si) 0.90 (3 H, t, J 7.1,
CO2CH2CH3), 4.11 (2 H, quartet, J 7.1, CO2CH2CH3), 7.39–7.82
(10 H, m), 8.25 (1 H, dd, J 1.3 and 8.0), 8.88 (1 H, d, J 7.9), 8.91
(1 H, d, J 7.8) and 10.61 (1 H, s, NH); dC(75 MHz; CDCl3; Me4Si)
13.4 (CO2CH2CH3), 60.8 (CO2CH2CH3), 111.0, 111.2, 117.6 (4a-
C), 119.4, 119.8, 121.0, 121.4 (4b-C), 121.9, 122.5, 123.3, 124.5,
124.6, 125.8, 127.0, 128.9, 132.2, 133.1, 133.2, 134.2, 136.2, 139.6,
141.1, 149.4 (2 ¥ C–NO2) and 167.6 (CO2CH2CH3); m/z (EI)
481 (M+ 100%); HRMS calcd. for C27H19N3O6 481.1274, found
481.1267.
6-Ethoxycarbonyl-12-(2¢-aminophenyl)-5H,11H-indolo[3,2-
b]carbazole (12)
◦
Green solid; mp 190–192 C; nmax (KBr)/cm-1 3436, 3048, 1661,
1613, 1318, 1265, 1243, 1164 and 1146; dH(300 MHz; DMSO-d6)
1.55 (3 H, t, J 6.7, OCH2CH3), 4.65 (2 H, s, NH2), 4.75 (2 H, q, J
6.7, OCH2CH3), 6.88 (1 H, t, J 7.9, 5¢-H), 6.92 (1 H, t, J 8.0, 8-H),
7.02 (d, J 7.9, 1 H, 3¢-H), 7.10 (1 H, d, J 7.9, 1-H), 7.15 (1 H, t, J
7.9, 2-H), 7.20 (1 H, d, J 7.9, 6¢-H), 7.35 (1 H, t, J 7.9, 3-H), 7.38
(1 H, t, J 7.9, 4¢-H), 7.42 (1 H, t, J 8.0, 9-H), 7.60 (1 H, d, J 7.9,
4-H), 7.70 (1 H, d, J 8.0, 10-H), 8.75 (1 H, d, J 8.0, 7-H), 10.75
(1 H, s, 11-NH) and 11.10 (1 H, s, 5-NH); dC(75 MHz; DMSO-
d6) 14.7 (CO2CH2CH3), 61.0 (CO2CH2CH3), 104.5 (6-C), 111.4
(10-CH), 111.6 (4-CH), 115.4 (3¢-CH), 117.0 (5¢-CH), 117.9 (2-
CH), 118.5 (8-CH), 119.7 (1¢-C), 120.3 (6a-C), 120.8 (12-C), 121.4
(12a-C), 121.6 (1-CH), 121.6 (12b-C), 121.9 (6b-C), 124.9 (7-CH),
126.0 (3-CH), 126.1 (9-CH), 129.7 (4¢-CH), 130.4 (6¢-CH), 134.1
(11a-C), 136.2 (5a-C), 141.4 (4a-C), 142.1 (10a-C), 146.4 (2¢-C)
and 167.6 (CO2CH2CH3); m/z (EI) 419 (M+ 100%) and 373 (37);
HRMS calcd. for C27H21N3O2 419.1634, found: 419.1629.
Preparation of polycyclic derivatives (10), (11), (12), (13) and (14)
To a solution of the three inseparable atropoisomers 8a(a1,a2,a3)
(836 mg, 1.74 mmol) in a mixture of toluene–ethanol (20 mL/50
mL) was added Pd/C 10% (124 mg). The suspension was stirred
in an autoclave under hydrogen atmosphere (10 bar). After 25 h,
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the green suspension was filtered off on a Celiteꢀ pad and
the catalyst was successively washed with toluene, CH2Cl2 and
methanol affording a first filtrate I. The remaining solid was
finally washed twice with distilled THF to give the second filtrate
II. After concentration of the filtrate I, the dark-yellow residue
(352 mg) was purified by column chromatography on silica gel to
obtain successively 10 (5 mg, 0.7%), as a yellow powder (eluent:
petroleum ether/CH2Cl2: 70/30), 12 (122 mg, 17%), as a green
powder (eluent: petroleum ether/CH2Cl2: 50/50) and 13 (7 mg,
1%), as a pale yellow powder (eluent: CH2Cl2–MeOH: 99/1).
Concentration of the filtrate II in vacuo at room temperature
afforded 11 (356 mg, 55%), as a yellow powder.
6-(2¢-Aminophenyl)-12-oxo-11H-12,13-dihydroquinolino[4,3-
a]carbazole (13)
15-Ethoxy-5H,14H-indolo[2,3-c]quinolino[4,3-a]carbazole (10)
Pale yellow solid; mp 175–177 ◦C; nmax (KBr)/cm-1 3401, 3216,
3048, 1652, 1647, 1613, 1335 and 1318; dH(300 MHz; DMSO-d6)
4.65 (2 H, s, NH2), 6.80 (1 H, t, J 7.6, 5¢-H), 6.92 (1 H, d, J 7.6,
3¢-H), 7.03 (1 H, t, J 8.1, 8-H), 7.22 (1 H, d, J 7.6, 6¢-H), 7.25 (1 H,
d, J 8.1, 7-H), 7.29 (1 H, t, J 7.4, 3-H), 7.30 (1 H, t, J 7.6, 4¢-H),
7.38 (1 H, t, J 8.1, 9-H), 7.50 (1 H, d, J 7.4, 1-H), 7.55 (1 H, t, J
7.4, 2-H), 7.90 (1 H, d, J 8.1, 10-H), 8.05 (1 H, s, 5-H), 8.50 (1 H,
d, J 7.4, 4-H), 11.90 (1 H, s, 13-NH) and 12.07 (1 H, s, 11-NH);
dH(300 MHz; DMSO-d6) 108.9 (11b-C), 112.6 (10-CH), 114.2 (5-
CH), 114.9 (3¢-CH), 116.4 (1-CH), 116.5 (5¢-CH), 118.5 (4a-C),
119.5 (8-CH), 120.1 (6a-C), 121.3 (7-CH), 121.4 (6b-C), 122.6 (3-
CH), 124.1 (4-CH), 124.6 (1¢-C), 125.7 (9-CH), 129.3 (4¢-CH),
129.3 (2-CH), 129.9 (6¢-CH), 133.1 (4b-C), 136.7 (13a-C), 138.7
(11a-C), 139.3 (6-C), 140.4 (10a-C), 145.8 (2¢-C) and 162.1 (CO);
m/z (EI) 375 (M+ 100%); HRMS calcd. for C25H17N3O 375.1372,
found: 375.1373.
Yellow powder; mp 184–186 ◦C; [Found: C, 81.01; H, 4.68; N,
10.69. C27H19N3O requires C, 80.77; H, 4.77; N, 10.47%];.nmax
(KBr)/cm-1 3463, 1604, 1591, 1458, 1419, 1392, 1326, 1309, 1230,
1216, 1141 and 1036; dH(300 MHz; DMSO-d6) 1.70 (3 H, t, J
7.0, OCH2CH3), 5.07 (2 H, q, J 7.0, OCH2CH3), 7.42–7.53 (2 H,
m, 8-H and 11-H), 7.56–7.67 (2 H, m, 7-H and 12-H), 7.74–7.85
(2 H, m, 2-H and 3-H), 8.03 (2 H, dl, J 8.5), 8.08 (1 H, d, J
8.1), 8.94 (1 H, d, J 8.1, 9-H or 10-H), 9.0 (1 H, d, J 8.1, 9-H
or 10-H), 9.23 (1 H, d, J 7.6, 4-H), 11.60 (1 H, s, 5-NH) and
12.03 (1 H, s, 14-NH); dC(75 MHz; DMSO-d6) 15.0 (OCH2CH3),
62.2 (OCH2CH3), 104.9, 112.7, 113.1, 115.4, 119.4, 119.8, 119.9,
121.0, 121.2, 121.5, 121.8, 122.7 (9-CH or 10-CH), 123.2 (9-CH or
10-CH), 124.9 (3-CH), 125.1, 125.6 (4-CH), 126.1, 127.7 (1-CH),
128.4 (2-CH), 129.8, 130.1, 139.4, 141.6, 143.1 (16a-C) and 158.6
(15-C). m/z (ESI) 402 (M + H)+.
4632 | Org. Biomol. Chem., 2010, 8, 4625–4636
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5H,14H-Indolo[2,3-c]quinolino[4,3-a]carbazole (14)
chloropropyldimethylamine hydrochloride (series b) (146 mg, 0.92
mmol), Na2CO3 (340 mg, 2.46 mmol), TBAI (6 mg, 0.016 mmol)
in DMF (12 mL); Heating: 85 ◦C for 13 h; Extraction: sat.
aq. K2CO3 (10 mL) and ethyl acetate (10 mL ¥ 3); Purification
(eluent: CH2Cl2–CH3OH: 98/2→ CH2Cl2–CH3OH: 60/40) gave
the orange foam 16b (8 mg, 12%) and 17b (45 mg, 55%) as an
amorphous orange solid.
To a suspension of LiAlH4 (2.10 g, 55.4 mmol) in dry DME (70
mL) was added at 0 ◦C under nitrogen atmosphere a solution
of 11 (153 mg, 0.41 mmol) in dry DME (50 mL). The orange
suspension was stirred at room temperature and then heated at
60 ◦C (oil bath) for 11 h under nitrogen atmosphere. To the
orange suspension water (17 mL) was added dropwise at 0 ◦C
under stirring until precipitation occurred. The precipitate was
washed with DME, ethyl acetate, the filtrate was dried (MgSO4),
filtered and concentrated in vacuo (180 mg). Purification by
chromatography on silica gel (eluent: CH2Cl2–CH3OH: 98/2) gave
14 (104 mg, 71%), as a yellow powder.
5-[3¢-(N,N-Dimethylamino)propyl]-14H-indolo[2,3-
c]quinolino[4,3-a]carbazole (16b)
Orange foam; nmax(film)/cm-1 2919, 2844, 1595, 1581, 1455, 1378,
1361 and 1328; dH(300 MHz; CDCl3; CD3OD; Me4Si) 2.18–2.32
(2 H, m, 2¢-CH2), 2.32 (6 H, s, N(Me)2), 2.53 (2 H, t, J 7.4, 3¢-
CH2), 4.72 (2 H, t, J 7.4, 1¢-CH2), 7.43 (2 H, t, J 7.7, 11-H and
8-H), 7.59 (2 H, t, J 8.3, 12-H and 7-H), 7.69 (1 H, d, J 8.3, 6-H),
7.73–7.88 (3 H, m, 13-H, 2-H and 3-H), 8.21 (1 H, d, J 8.7, 1-H),
8.82 (1 H, d, J 8.3, 10-H), 8.86 (1 H, d, J 8.3, 9-H), 9.00 (1 H, d,
J 7.7, 4-H) and 9.83 (1 H, s, 15-H); dC(75 MHz; CDCl3; CD3OD;
Me4Si) 26.5 (2¢-CH2), 44.6 (N(Me)2), 44.9 (1¢-CH2), 55.9 (3¢-CH2),
109.5 (6-CH), 111.9 (13-CH), 112.8 (4b-C or 14b-C), 117.3 (9c-
C), 119.0 (4b-C or 14b-C), 119.6 (8-CH), 119.8 (11-CH), 120.3
(9b-C), 122.0 (9a-C), 122.5 (9d-C), 122.9 (10-CH), 123.5 (9-CH),
123.6 (4a-C), 124.6 (4-CH), 125.3 (12-CH), 126.1 (7-CH), 127.3
(3-CH), 127.9 (2-CH), 128.9 (1-CH), 130.2 (14a-C), 131.3 (4c-
C), 141.3 (5a-C or 13a-C), 141.4 (5a-C or 13a-C), 143.0 (16a-C)
and 146.8 (15-CH); m/z (EI) 443 (M+ + 1, 4%), 442 (M+, 13%),
368 (31), 129 (100); HRMS calcd. for C30H26N4 442.2157, found
442.2147.
In some cases, especially in longer reactions (15–16 h at 60 ◦C) a
mixture of 14 and its dihydro derivative (15) was obtained. In that
case, after destruction of the excess of LiAlH4 the concentrated
filtrate was dissolved in a mixture of toluene (20 mL) and DME (10
mL), p-chloranil (376 mg, 1.53 mmol) was added to the solution
and the reaction mixture was heated at 50 ◦C for 6 h under nitrogen
atmosphere until the complete conversion of 15 into 14 (TLC
monitoring). After concentration in vacuo the brown residue was
washed with CH2Cl2, filtered and concentrated to give 14 (386 mg,
61%), as a yellow powder; mp >350 ◦C; nmax (KBr)/cm-1 3410,
1
3233, 2916, 1613, 1577, 1458, 1379, 1309 and 1243; H NMR
(300 MHz; DMSO-d6) 7.41–7.55 (2 H, m, 8-H and 11-H), 7.62 (1
H, t, J 8.1, 12-H), 7.69 (1 H, t, J 8.1, 7-H), 7.82 (1 H, d, J 8.1,
13-H), 7.98 (1 H, d, J 8.1, 6-H), 8.03–8.18 (2 H, m, 2-H and 3-H),
8.46 (1 H, d, J 7.5, 1-H), 8.85 (1 H, d, J 8.1, 10-H), 8.92 (1 H, d,
J 8.1, 9-H), 9.41 (1 H, d, J 7.5, 4-H), 10.56 (1 H, s, 15-H), 12.28
(1 H, s, 5-NH) and 13.21 (1 H, s, 14-NH); dC(75 MHz; DMSO-
d6) 110.4, 112.1 (13-CH), 113.3 (6-CH), 115.4 (9c-C), 118.8, 120.1
(11-CH), 120.5 (8-CH), 121.2 (9a-C), 121.4 (9d-C), 123.0 (9b-
C), 123.3 (10-CH), 123.4, 123.8 (9-CH), 126.0 (2 ¥ CH, 12-CH
and 1-CH), 126.5 (4-CH), 127.8 (7-CH), 128.7 (4c-C), 129.3 (2-
CH), 129.9 (3-CH), 133.0 (14a-C), 139.0 (16a-C), 139.8 (13a-C),
142.7 (5a-C) and 146.0 (15-CH); m/z (ESI +) 358 (M + H)+; m/z
(EI) 357 (M+ 100); HRMS calcd. for C25H15N3 357.1266, found:
357.1259.
5-[3¢-(N,N-Dimethylamino)propyl]-14-[3¢¢-(N,N-dimethylamino)-
propyl]-indolo[2,3-c]quinolino[4,3-a]carbazole (17b)
Amorphous orange solid; nmax (KBr)/cm-1 3047, 2929, 2754, 1598,
1579, 1456, 1446, 1373 and 1321; dH(300 MHz; CDCl3; Me4Si)
0.92–1.10 (2 H, m, 2¢-CH2), 1.41 (2 H, t, J 7.0, 3¢-CH2), 1.68
(6 H, s, N(Me)2), 2.31 (6 H, s, N(Me)2), 2.32–2.40 (2 H, m, 2¢¢-
CH2), 2.52 (2 H, t, J 6.9, 3¢¢-CH2), 4.68 (2 H, t, J 7.0, 1¢-CH2),
4.99 (2 H, t, J 6.9, 1¢¢-CH2), 7.46–7.56 (2 H, m, 11-H and 3-H),
7.57–7.72 (3 H, m, 12-H, 7-H and 8-H), 7.73–7.75 (1 H, m, 2-
H), 7.75–7.84 (2 H, m, 13-H and 1-H), 8.26 (1 H, dd, J 1.1 and
8.0, 6-H), 8.80 (1 H, dd, J 1.1 and 8.1, 4-H), 8.97 (1 H, d, J
8.0, 10-H), 9.04 (1 H, d, J 8.0, 9-H) and 10.20 (1 H, s, 15-H);
dC(75 MHz; CDCl3; Me4Si) 25.0 (2¢-CH2), 27.7 (2¢¢-CH2), 44.7
(N(Me)2), 45.0 (1¢¢-CH2), 45.5 (N(Me)2), 46.7 (1¢-CH2), 55.9 (3¢-
CH2), 56.5 (3¢¢-CH2), 109.7 (13-CH), 112.9 (1-CH), 113.3 (4b-C),
116.8 (9c-C), 119.6 (11-CH), 120.6 (3-CH), 122.0 (9d-C), 122.8
(9a-C), 123.1 (10-CH), 123.4 (9b-C), 123.8 (14b-C), 124.1 (9-CH),
125.3 (12-CH), 125.4 (4a-C), 125.8 (8-CH), 126.2 (7-CH), 127.4
(4-CH), 127.9 (2-CH), 128.3 (6-CH), 132.3 (14a-C), 132.8 (4c-C),
141.6 (13a-C), 143.5 (16a-C), 145.8 (5a-C) and 147.5 (15-CH); m/z
(ESI+) 528 (M + 1)+; m/z (EI) 528 (M+ + 1, 26%), 527 (M+ 100%)
469 (67), 455 (41); HRMS calcd. for C35H37N5 527.3049, found
527.3042.
General procedure for the preparation of N-substituted
indolo[2,3-c]quinolino[4,3-a]carbazoles (16b) and (17a,b) and of
N-substituted 15-oxo-15,16-dihydroindolo[2,3-
c]quinolino[4,3-a]carbazole (18)
Sodium hydride 60% was carefully introduced at -50 ◦C under
nitrogen atmosphere to a stirred solution of 11 or 14 in dry
DMF. The suspension of the halide (series a (n = 1) or series
b (n = 2)), Na2CO3 (powder) and tetrabutylammonium iodide
(TBAI) in dry DMF was dropwise added to the latter under
nitrogen atmosphere. The mixture was heated, diluted with ethyl
acetate and extracted by a saturated aqueous potassium carbonate
solution. The combined organic layers were dried over MgSO4,
filtered and concentrated. Purification of the crude mixture by
column chromatography on silica gel afforded the corresponding
N-substituted carbazoles.
Preparation of N-substituted 15-oxo-15,16-dihydroindolo[2,3-
c]quinolino[4,3-a]carbazole (18)
Preparation of carbazoles 16b and 17b
According to General Procedure: Starting from 14 (55 mg,
According to General Procedure: Starting from 11 (78 mg,
0.15 mmol) reacted with NaH 60% (37 mg, 0.92 mmol), 3-
0.21 mmol) reacted with NaH 60% (76 mg, 1.89 mmol),
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3-chloropropyldimethylamine hydrochloride (series b) (199 mg,
1.26 mmol), Na2CO3 (356 mg, 3.36 mmol), TBAI (8 mg, 0.021
mmol) in DMF (12 mL); Heating: 85 ◦C for 20 h; Extraction: sat.
aq. K2CO3 (10 mL) and ethyl acetate (10 mL ¥ 3); Purification
(eluent: CH2Cl2–CH3OH: 95/5→ CH3OH/NH4OH: 90/10) gave
the inseparable mixture of mono- and dialkylated compounds
(11 mg; approx. 17%) as an amorphous orange solid, and 18
(70 mg, 53%) as an amorphous orange solid.
Acknowledgements
Authors thank Pr. J.-Y. Laronze (Universite´ de Reims) for helpful
discussions. Financial support of CNRS, Re´gion Champagne-
Ardenne, Fond Europe´en pour le De´veloppement Re´gional (FEDER)
and University of Reims is gratefully acknowledged. F.C. thanks
the French Ministry of Education and Research for a fellowship.
The C.R.I.H.A.N computing centre and the computational centre
of the Universite´ de Reims Champagne-Ardenne (ROMEO)
are acknowledged for the CPU time donated. Spectroscopic
measurements by C. Petermann (NMR), D. Patigny (MS) and
P. Sigaut (MS) are gratefully acknowledged. J.-F.R. and B.B. are
supported by the Ligue Nationale contre le Cancer, e´quipe labellise´e
2010.
5,14,16-Tris[3-(N,N-dimethylamino)propyl]-15-oxo-15,16-
dihydroindolo[2,3-c]quinolino[4,3-a]carbazole (18)
Amorphous orange solid; nmax (KBr)/cm-1 2943, 2706, 1635, 1598,
1458, 1370 and 1115; dH(300 MHz; CDCl3, Me4Si) 1.15–1.29 (2 H,
m, 2¢¢¢-CH2), 1.44–1.58 (2 H, m, 3¢¢¢-CH2), 1.72 (6 H, s, N(Me)2),
1.73–1.83 (2 H, m, 2¢-CH2 or 2¢¢-CH2), 2.09–2.19 (2 H, m, 2¢-CH2
or 2¢¢-CH2), 2.31 (6 H, s, N(Me)2), 2.33 (6 H, s, N(Me)2), 2.50
(2 H, t, J 7.3, 3¢-CH2 or 3¢¢-CH2), 2.53–2.67 (2 H, m, 3¢-CH2 or
3¢¢-CH2), 3.48 (2 H, t, J 6.4, 1¢-CH2 or 1¢¢-CH2), 4.51 (2 H, t, J
7.5, 1¢-CH2 or 1¢¢-CH2), 4.58–4.75 (2 H, m, 1¢¢¢-CH2), 7.30 (1 H,
t, J 7.9, 3-H), 7.41–7.54 (3 H, m, 2-H, 8-H and 11-H), 7.57–7.68
(3 H, m, 1-H, 7-H, 12-H), 7.73 (2 H, d, J 8.2, 6-H and 13-H), 8.27
(1 H, dd, J 1.1 and 7.9, 4-H), 8.94 (1 H, d, J 8.0, 9-H or 10-H),
9.02 (1 H, d, J 8.0, 9-H or 10-H); dC(125 MHz; CDCl3; Me4Si)
24.8 (2¢¢¢-CH2), 25.9 (2¢-CH2 or 2¢¢-CH2), 27.3 (2¢-CH2 or 2¢¢-CH2),
41.3 (1¢-CH2 or 1¢¢-CH2), 44.9 (N(Me)2), 45.0 (N(Me)2), 45.4 (1¢“-
CH2), 45.5 (N(Me)2), 55.9 (3¢¢¢-CH2), 56.4 (3¢-CH2 or 3¢¢-CH2),
57.0 (3¢-CH2 or 3¢¢-CH2), 68.7 (1¢-CH2 or 1¢¢-CH2), 110.6, 111.1
(6-CH or 13-CH), 112.4 (6-CH or 13-CH), 114.3 (1-CH), 119.4,
119.5 (8-CH or 11-CH), 119.9 (4a-C), 120.3 (8-CH or 11-CH),
121.7 (3-CH), 123.0 (9-CH or 10-CH), 124.3 (9-CH or 10-CH),
124.9 (9a-C or 9d-C), 125.5 (7-CH or 12-CH), 126.5 (7-CH or
12-CH), 128.2 (2-CH), 128.3 (4-CH), 132.4, 135.5 (16a-C), 136.5,
143.4 (5a-C or 13a-C), 145.7 (5a-C or 13a-C) and 160.6 (CO);
m/z (EI) 629 (M+ + 1, 13%), 628 (M+, 51%), 543 (100), 472 (39),
458 (67), 357 (42); HRMS calcd. for C40H48N6O 628.3890, found
628.3906.
Notes and references
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4634 | Org. Biomol. Chem., 2010, 8, 4625–4636
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