Synthesis of chromenoindole derivatives from Robinia pseudoacacia

Article abstract of DOI:10.1039/c3md00213f

An unusual tetrahydrochromeno[4,3-b]indole type analogue of medicarpin has been isolated from the roots of the Black Locust and subsequently has been synthesized together with an aromatic derivative which shows considerable biological activity against a range of significant targets.

Full text of DOI:10.1039/c3md00213f

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Synthesis of chromenoindole derivatives from Robinia
pseudoacacia†
Cite this: Med. Chem. Commun., 2013,
4, 1580
a
b
b
b
a
*
Lisa Dejon, Hamdoon Mohammed, Peng Du, Claus Jacob and Andreas Speicher
Received 24th July 2013
An unusual tetrahydrochromeno[4,3-b]indole type analogue of medicarpin has been isolated from the
roots of the Black Locust and subsequently has been synthesized together with an aromatic derivative
which shows considerable biological activity against a range of significant targets.
Accepted 5th October 2013
DOI: 10.1039/c3md00213f
www.rsc.org/medchemcomm
the roots of the Black Locust (Robinia pseudoacacia), a tree
known to be rich in (biologically active) avonoids (for details
Introduction
The phytoalexin medicarpin (3) is found in a wide range of
legumes, such as barrel medic (Medicago truncatula, a Medi-
terranean form of clover) and alfalfa, where it is formed upon
fungal infection of the plant.1 During the last couple of decades,
several studies have been conducted to investigate the various
biological activities with this plant protective/defensive pter-
ocarpan (for a review see ref. 1). Not surprisingly, medicarpin
exhibits a good fungicidal activity against a range of pathogenic
fungi,2 including Cladosporium cladosporioides.3 The compound
is also effective against other microbes, including certain
bacteria,4 and has shown the ability to induce apoptosis in
human lung broblasts and peripheral lymphocytes.5 In the
context of human health, medicarpin and its analogue cou-
mestrol form a group of tetracyclic isoavone-derivatives with
phyto-estrogenic properties and have a potential impact on
bone formation.6 Interestingly, medicarpin is also present in
the dried root of Taverniera abyssinica, and forms part of
an analgesic and antipyretic Ethiopian drug called “Dinge-
tegna” which, like medicarpin itself, shows strong nematicidal
activity, for instance against Caenorhabditis elegans.7
see ESI†). This compound is rather unusual for a range of
reasons. First of all, medicarpin (3) is synthesized from its
(open) isoavone derivative formononetin by formal ring
closure involving B and the keto-function in ring A. In sharp
contrast, the biosynthesis of 1, which is hitherto unknown,
would have to proceed via an imine, which would be fairly
unusual. Secondly, whilst the overall three-dimensional struc-
ture of this compound may be similar to the one of medicarpin,
the presence of the nitrogen (instead of the oxygen) atom may
result in a distinctly different biological “chemistry” and hence
activity, for instance regarding electrostatic interactions and
metal binding.
(2) Initial screening for possible biological activity
In the next step, we have therefore studied some of the most
obvious potential activities of 1, such as (a) antioxidant activity
(which is oen associated with avones), (b) aromatase inhi-
bition (as found for coumestrol, see ref. 6) and cytotoxicity
against (c) nematodes and (d) against a specic leukemia cell
Results and discussion
(1) Isolation of a natural chromeno[4,3-b]indole derivative
from the roots of Robinia pseudoacacia
As part of our ongoing research into natural products associated
with a distinct biological activity, we have isolated a rather
unusual compound 1 (Scheme 1) of the tetrahydro-chromeno-
[4,3-b]indole type, a nitrogen-analogue of medicarpin (3), from
aInstitute for Organic Chemistry, Saarland University, D-66123 Saarbruecken,
Germany. E-mail: anspeich@mx.uni-saarland.de; Fax: +49 681 302 2029; Tel: +49
681 302 2749
bDivision of Bioorganic Chemistry, School of Pharmacy, Saarland University, D-66123
Saarbruecken, Germany
† Electronic supplementary information (ESI) available: Isolation of compound 1
from a natural source, details of chemical syntheses, NMR spectra and details of in
vitro activity assays. See DOI: 10.1039/c3md00213f
Scheme 1 Chemical structures of the novel tetrahydro chromeno-[4,3-b]indole 1,
its chemical dihydro precursor 2 and its known oxygen analogue medicarpin (3).
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line (human promyelocytic leukemia cells, HL-60). The results
obtained for compound 1 in these assays have been mostly
disappointing, with low or no activities observed in the DPPH
antioxidant assay (IC50 > 300 mM), the aromatase inhibition
assay (IC50 > 300 mM) and the Steinernema feltiae nematode
assay (no signicant toxicity observed at concentrations up to
200 mM and 48 h of incubation). At closer consideration, these
results may not be too surprising, however, as compound 1 does
not possess the typical kind of hydroquinone ring responsible
for the antioxidant activity of many avonoids and also lacks the
two free hydroxyl-groups of coumestrol required for interac-
tions within the estrogen pathway.
Compound 1, however, did exhibit an interesting cytotoxicity
against HL-60 cells, with cell survival of 72% at 40 mM and just
27% at 200 mM of compound 1. Whilst this activity is rather
modest when compared to classical cytotoxic molecules, it is
still rather impressive for a natural product which has not yet
been optimized for a specic activity.
(3) Strategy to synthesize dihydrochromeno[4,3-b]indole
derivatives (compare 2) as precursors for 1
Scheme 3 Retrosynthesis for compound 2 and previous attempts for synthesis
(PG: protective group). Schemes 3/4 (old) combined, the text was adapted.
Due to the fact that the heterocyclic backbone of 1 is rather
unknown and that only small amounts of the natural
compound were available, a total synthesis of 1 seemed to be
worthwhile. Ultimately, this should allow us to better under-
stand the cytotoxic effects associated with compound 1, to
access the various derivatives of this novel class of natural
chromeno[4,3-b]indoles and, possibly, also derive at more active
compounds which ultimately may be useful for pharmacolog-
ical application. Hence a strategy has been developed to
synthesize these compounds 1/2 and to extend the initial
pharmacological studies of 1 and of similar compounds.
(to 6) resulted in low yields and efficiency (route 1). Attempts to
cyclize haloenamins 9 (available through enamine formation by
Buchwald–Hartwig coupling11 as an intermediate step) failed
(route 2).
Hence, an alternative strategy had to be employed. Several
methods used to prepare condensed indole compounds are
based on C–C-cross-coupling followed by cyclization. For
instance, Ullmann type coupling between an a-halo enone and
an a-halo nitroarene followed by hydrogenation to the corre-
sponding anilines and (spontaneous) ring closure leads to
indoles (similar to the “Reissert synthesis”).12 Based on this
strategy (route 3), the indole structure of 2 would be accessible
by such a sequence starting with a halochromenone 11 and an
a-halo nitroarene 12 and formation of 10 in the rst step.
For reasons of accessibility and stability we synthesized the
3-iodochromenone 16 from a mono-protected dihydroxy aceto-
phenone 14 by aldol condensation with dimethylformamide
dimethylacetal and subsequent iodo-induced cyclization13 in
37% overall yield. Ullmann type cross coupling with the bromo
nitroanisole 16 yielded the 3-(o-nitroaryl)chromenone 18
(Scheme 4).
The synthesis of the non-substituted dihydrochromeno-
8
¨
[4,3-b]indole skeleton 2a was described by Buu-Hoı using the
classical Fischer indole approach (Scheme 2). Besides the low
yield (15%), this route is impracticable for the synthesis of
substituted derivatives such as 2 because of the need for specic
chromanones (e.g. 4) as starting material and because of a
notable lack of regioselectivity in the indole ring formation with
substituted phenylhydrazines like 5.
Numerous additional methods have been described for the
synthesis of indole derivatives.9 For the highly condensed
heterocyclic system 2 we rst explored two routes following the
retrosynthesis depicted in Scheme 3. The Suzuki coupling10
between carbamoyl aryl halides 7 and an indole boronic ester 8
Several reaction conditions for a hydrogenation cascade
from 18 to the dihydrochromenoindol 2 were set up also
considering the solubility of 18 and some intermediates
(Scheme 5). With H2–Pd–C in MeOH or EtOAc12 mainly a
mixture of 20 and 21 was obtained whereas H2–Pt–C in THF14
gave 21 in high yield. But 21 could be debenzylated to 2 only
under harsh reaction conditions via catalytic transfer hydroge-
nation using 1,4-cyclohexadiene as a hydrogen donor.15 Due to
this additional cleavage step, we tried to proceed with the
synthesis without the phenol protecting group, but this
approach led to difficult work-up conditions in the rst step
already.
Scheme 2 Fischer approach to the dihydrochromeno[4,3-b]-indole skeleton 4.
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tetrahydrochromeno-indole 1 are cis annulated for reasons of
strong thermodynamical preference.20 The natural compound
was isolated as a racemate.
(4) Biological activity associated with compounds 1 and 2
Based on the literature describing various biological activities
associated with medicarpin and coumestrol, and taking into
account the notable activities initially seen for the isolated
compound, it was decided to take a closer look at possible – and
possibly useful – biological activities associated with synthetic,
chemically pure compound 1 and its unsaturated analogue 2.
Here, it was expected that the pure compound 1 – as well as its
“medicarpin-like” analogue 2 – may be more active compared to
the isolated material. Both compounds were considered
together, as they clearly differ in their three-dimensional shape
and also chemical reactivity (see Fig. 1). They were tested
against relevant targets, such as the human colon cancer cell
line HCT-116 (this cell line is more representative of a typical
cancer cell than HL-60).
Scheme 4 Synthesis of the indole precursor 18. Reaction conditions: (i) K2CO3,
BnCl, MeCN, reflux; (ii) 90 ꢀC; (iii) I2, CHCl3, r.t.; (iv) Cu, Pd2(dba)3, DMSO, 70 ꢀC.
Fig. 2 shows the biological activity of 1 and 2 in the HCT-116
cell culture assay. Whilst the natural substance 1 shows no
signicant toxicity up to a concentration of 100 mM (higher
concentrations are not considered as pharmaceutically relevant),
the unsaturated analogue 2 is fairly cytotoxic against these cancer
cells, with an IC50 of 66.1 mM. These results are rather intriguing.
Whilst they conrm that compound 1 is not particularly toxic
against human cancer cells (as already expected when using the
extracted compound), they also highlight the quite signicant
change of activity when moving from the 2-pyrroline to the
pyrrole ring which in 2 forms part of an indole moiety.
While these initial results highlight a notable activity asso-
ciated with the indole-containing compound 2, they cannot yet
explain why this compound is active whilst compound 1 is not.
One may speculate that 2, as a planar, tetracyclic molecule may
interact with DNA (e.g. via intercalation) or enzymes involved in
the synthesis and/or replication of DNA. Here, the indole ring
present in 2 may enhance binding to DNA, possibly by coordi-
nating to a shared Mg2+ cation. Indeed, indoles, such as the
amino acid tryptophan, are known to interact quite strongly
with metal ions, such as Mg2+, and, besides the tetracyclic
Scheme 5 Hydrogenation steps for the synthesis of 1 from 18. Reaction
conditions: (i) 10% Pt/C, THF, 15 bar H2; (ii) cyclohexadiene, EtOAc, Pd/C, 70 ꢀC;
(iii) 10% Pt/C, Pd(OH)2/C, THF, 5 bar H2; (iv) NaBH3CN, HOAc, r.t. Stereo formula
for 1 was added.
Finally, by adding Pearlman's catalyst [Pd(OH)2/C]16 to the plat-
inum catalyst, the cyclization as well as the deprotection occurred
in one step to give the dihydrochromenoindole 2 in high yield.
Since the reaction conditions for the catalytic hydrogenation
of indoles are drastic,14 we decided to reduce the double bond
using NaBH3CN in HOAc17 and obtained the desired indoline 1,
now synthesized for the rst time in a six-step synthesis and
16% overall yield. The spectroscopic data of the synthetic
sample are identical to those of the isolated compound (see
ESI†). As expected18 and clearly veried for the medicarpin
analogue 3,19 the chroman and indoline subunits of this Fig. 1 3D shapes of 1 and 2 calculated by AM1.21
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various analogues are now easily accessible via the synthetic route
described as part of this study, more detailed investigations are
now feasible and bode well for future drug development.
Notes and references
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in the structure of compound 2 is the apparent cation-p metal
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notably absent in medicarpin 3 and is also clearly different in
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¨ ¨
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In summary, the biological activities associated with compound 2
are rather interesting and may form the basis for more extensive 16 J. A. Smith, D. J. Maloney, S. M. Hecht and D. A. Lannigan,
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as far as metal binding is concerned. At the same time, the furan 18 T. G. van Aardt, H. van Rensburg and D. Ferreira,
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presence of an aromatic ring in this oxygen-analogue may result 19 A. Goel, A. Kumar, Y. Hemberger, A. Raghuvanshi, R. Jeet,
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activity (against different cell types), selectivity (e.g. for specic
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in more detail, as it is still unclear why compound 2 is rather 21 HyperChemꢀ Professional 7.52, Hypercube, Inc., 1115 NW
cytotoxic whilst compound 1 is not. As compounds 1, 2 and
4th Street, Gainesville, Florida 32601, USA.
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