A Pd-based regioselective strategy to indole-1,2-fused 8- and 9-membered rings: Their evaluation as potential scaffolds for apoptosis in zebrafish

Article abstract of DOI:10.1039/c4ob00140k

A strategy based on Pd-mediated ring closure of 1,2-disubstituted indoles containing an unactivated olefin leading to indole-1,2-fused 8- and 9-membered rings has been developed for the identification of new and potential scaffolds for apoptosis. A large number of fused indole derivatives containing an endocyclic double bond were synthesized using this robust methodology. A representative compound showed promising apoptotic properties in zebrafish embryos. This journal is the Partner Organisations 2014.

Full text of DOI:10.1039/c4ob00140k

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A Pd-based regioselective strategy to indole-1,2-
fused 8- and 9-membered rings: their evaluation
as potential scaffolds for apoptosis in zebrafish†
Cite this: DOI: 10.1039/c4ob00140k
Received 19th January 2014,
Accepted 20th February 2014
Bagineni Prasad,a B. Yogi Sreenivas,a Araka Sushma,a Swapna Yellanki,a,b
DOI: 10.1039/c4ob00140k
Raghavender Medisetti,a,b Pushkar Kulkarnia,b and Manojit Pal*a
www.rsc.org/obc
A strategy based on Pd-mediated ring closure of 1,2-disubstituted hydro-1H-[1,4]diazepino[1,7-a]indole (C, Fig. 1) possessing
indoles containing an unactivated olefin leading to indole-1,2- antipressant2 and CNS related activities,3 indolo-1,5-
fused 8- and 9-membered rings has been developed for the benzodiazocine (D, Fig. 1) having CNS related activities,4
identification of new and potential scaffolds for apoptosis. A large 2,3,4,5-tetrahydro-1H-[1,4]diazepino[1,2-a]indole (E, Fig. 1)
number of fused indole derivatives containing an endocyclic possessing 5-HT antagonistic properties5 are mention worthy.
double bond were synthesized using this robust methodology. A
Surprisingly, while compounds containing an indole-2,3-
representative compound showed promising apoptotic properties fused 7- or 8-membered ring system have been explored for
in zebrafish embryos.
potential anticancer/antitumor activities6–9 their indole-1,2-
fused analogues remain less or unexplored for this purpose.
More importantly, reports describing the pharmacological pro-
perties of compounds containing indole-2,3-fused 9 membered
ring system are not common in the literature perhaps due to
the cumbersome or non accessibility of this class of indoles. It
was therefore necessary to access and evaluate compounds
containing indole-1,2-fused 8- or 9-membered ring for their
potential anticancer especially apoptotic properties. Notably,
cancer is associated with a lower degree of apoptosis and most
of the cytotoxic anticancer agents are known to induce apopto-
sis. Apoptosis10 that occurs in physiological and pathological
conditions is an ordered and orchestrated cellular process.
The complex mechanism of apoptosis involves many pathways.
Defects in apoptotic pathways are thought to contribute to a
number of human diseases, ranging from neurodegenerative
disorders to malignancy.10 The design of our target molecules
I was prompted by known cytotoxic agents F containing an
indole-2,3-fused 7-membered ring system6 (Fig. 2). Thus the
generic structure I was drawn from F via some necessary struc-
tural manipulations through G and then H as shown in Fig. 2.
A number of methods have been reported for the synthesis
of indole-fused 7- and 8-membered rings at the 1,2 position
that can be classified according to the substrates used and the
bond formed in the ring-closing step.11 These includes either
indole ring formation on a preformed 7- or 8-membered ring
or ring closure of (i) N-substituted indoles via bond formation
to C-2 position, (ii) 2-substituted indoles via bond formation
to N-1 position and (iii) 1,2-disubstituted indoles via bond for-
mation between the substituents. While many of these
methods are very effective and elegant to access a particular
class of fused indole derivatives some of them suffer from
Compounds containing 6-, 7- or 8-membered ring fused
with an indole framework at the 1,2-position (A, Fig. 1) are not
only of immense importance in medicinal chemistry and
pharmacology but also common targets in synthetic organic
chemistry. For example, synthesis of indolo-diazepines
(B, Fig. 1) possessing antiserotonin activities,1 2,3,4,5-tetra-
Fig. 1 Compounds containing 6- or 7- or 8-membered ring fused with
indole at the 1,2-position.
aDr Reddy’s Institute of Life Sciences, University of Hyderabad Campus, Gachibowli,
Hyderabad 500 046, India. E-mail: manojitpal@rediffmail.com;
Tel: +91 40 6657 1500
bZephase Therapeutics (an incubated company at the Dr Reddy’s Institute of Life
Sciences), University of Hyderabad Campus, Gachibowli, Hyderabad 500046, India
†Electronic supplementary information (ESI) available: Experimental pro-
cedures, spectral data for all new compounds, copies of spectra. See DOI:
10.1039/c4ob00140k
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Table 1 Optimization of reaction conditionsa
Yieldb (%)
Entry
Catalyst/additive
Time (h)
2a
2aa
Fig. 2 Design of novel and potential apoptotic agents I from F.
1
2
Pd(PPh3)2Cl2
Pd(PPh3)4
Pd/C/PPh3
Cu(OAc)2
4
4
12
12
63
54
51
0
30
32
30
0
3
being not general and versatile in nature. Additionally, con-
struction of an indole-fused 9-membered ring at the 1,2 posi-
tion appeared as difficult or not feasible by using some of
these methodologies. We therefore required a general and
robust method to prepare our target compound I. Herein, we
report an effective method for the direct access to I (or 2) via
Pd-mediated intramolecular ring closure of 1,2-disubstituted
indoles 1 (Scheme 1).
4c
a Reactions were carried out using 1a (0.2 mmol), catalyst (5 mmol%)
and Et3N (0.4 mmol) in DMF (2 mL) at 110 °C. b Isolated yield.
c 1 mmol of catalyst used.
treated with Pd(PPh3)2Cl2 and Et3N in DMF at 110 °C for 4 h,
when the desired product 2a containing an endocyclic double
bond was isolated in 63% yield (entry 1, Table 1). While the
cyclopenta[b]indole15 2aa was isolated as a side product in this
case the formation of no isomeric product containing exocyclic
double bond was detected. The use of other catalysts e.g.
Pd(PPh3)4 (entry 2, Table 1), Pd/C/PPh3 (entry 3, Table 1) and
Cu(OAc)2 (entry 4, Table 1) was examined when yield of 2a was
not improved in first two cases and the reaction did not
proceed in the last case.
Since its discovery in 1977,12 the intramolecular Heck reac-
tion has become a powerful tool for the quick construction of
carbocyclic/heterocyclic rings.13 Intramolecular12a Heck reac-
tions are generally more efficient and regioselective than their
intermolecular12b version. While ring closure was found to be
highly exo selective for 5-, 6- and 7-membered rings and endo
selective for 13-membered and larger rings a mixture of exo
and endo products were formed in the closure of medium
sized, i.e. 8–12 membered, rings. Nevertheless, we adopted a
similar Pd-mediated strategy to prepare our target compound
2. The key starting material 1 required was prepared via C-2
A variety of compounds containing indole-1,2-fused 8-mem-
bered rings were prepared by using the optimized reaction
conditions in acceptable yields (Table 2). A number of ana-
logues containing indole-1,2-fused 9-membered rings were
also prepared using this methodology (Table 3). All the syn-
thesized compounds were well characterized by spectral data
(NMR, IR & MS). The coupling constant (J = 11.2–11.5 Hz) of
the Hb proton appeared in the region 6.8–6.9 ppm in the
1HNMR spectra of compounds 2a–r and this indicated that a
cis geometry of the double bond was present in these com-
pounds. This was further supported by the interaction of Ha
proton (6.00 ppm) with Hb (6.8 ppm) (being at the same side
of the double bond) upon irradiation in a 1D NOE experiment
performed using the compound 2b (Fig. 3). However, the coup-
ling constants of the olefinic protons could not be measured
in the case of compounds 2s–x due to the complex nature of
amination of indoles
3 using N-sulfonyl arylamines 4
(Scheme 2).14 We then performed the Pd-mediated intramole-
cular reaction of 1a under a variety of conditions (Table 1) to
establish the optimized reaction conditions. Initially, 1a was
Scheme 1 Synthesis of compound 2 via Pd-mediated intramolecular
ring closure of 1,2-disubstituted indoles 1.
1
these signals in their H NMR spectra (see ESI†). The 1D NOE
experiment performed in the case of compound 2s was not
conclusive (see ESI†). Notably, the DEPT experiment
(13C NMR) of 2s confirmed the presence of an endocyclic (not
exocyclic) double bond in the 9-membered ring. It is worthy to
mention that the endo mode of the intramolecular Heck cycli-
zation has been reported to be favored for substrates that
contain a Michael-type olefinic fragment (termed as electronic
Scheme 2 Iodine-mediated synthesis of sulfonamide 1.
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Table 2 Pd-mediated synthesis of indole-1,2-fused 8-membered rings
(2a–r)a,b
Table 3 Pd-mediated synthesis of indole-1,2-fused 9-membered rings
(2s–x)a,b
a All the reactions were performed using 1s–x (0.2 mmol), PdCl2(PPh3)2
(5 mol%) and Et3N (0.4 mmol) in DMF (2 mL) at 110 °C for 2–6 h
under N2. b Figures indicate % yield and reaction time.
Based on our experimental observations a proposed reac-
tion mechanism is presented in Scheme 3. While the reaction
seems to follow a classical Heck coupling pathway12b i.e. via
the generation of intermediate X (but not Y) the regioselectivity
of the double bond formation is the key feature of the present
process. The higher stability of the resultant conjugated
double bond (i.e. the styrene moiety) over the isolated one
perhaps aided the regioselective elimination of H-Pd species
from X leading to the product 2. The flexible geometry due to
a All the reactions were performed using 1a–r (0.2 mmol), PdCl2(PPh3)2
(5 mol%) and Et3N (0.4 mmol) in DMF (2 mL) at 110 °C for 2–6 h
under N2. b Figures indicate % yield and reaction time.
reasons)13a leading to 7-, 8- or 9-membered rings.16a This is in
sharp contrast to our observations where such electronic
reasons did not aid the endocyclization process. Additionally,
we were able to achieve this mode of cyclization using unacti-
vated olefin as one of the reactant moieties.
Fig. 3 1D NOE experiment of compound 2b.
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Fig. 4 The percentage induction of apoptosis caused by compound 2f
at different concentrations along with methotrexate. All the statistical
analyses were performed using GraphPad Prism® software.
Scheme 3 The proposed reaction mechanism leading to 2.
the large ring size of X could be the other reason for affording
the cis olefin rather than the trans product usually observed in
the case of intermolecular Heck reaction.16b
All the synthesized compounds were tested for their apopto-
tic activities initially at 30 µM using Zebrafish embryos. Zebra-
fish (or Danio rerio), a small pet-shop fish, is being explored as
a tool for enhancing interdisciplinary studies in biology and
chemistry as well as in drug discovery.17 Indeed, Zebrafish pro-
vides an inexpensive, reliable and efficient first-level screening
model for testing toxicity, efficacy, and tissue-targeting for a
large number of new chemical entities (NCEs). Our continued
interest18–24 in Zebrafish as a screening model for NCEs
prompted us to assess the potential pharmacological effects of
the present class of molecules in Zebrafish and/or their
embryos. The most active compound e.g. 2f was tested at 1, 3,
10, and 30 µM along with a standard drug methotrexate. The
percentage induction of apoptosis caused by compound 2f at
different concentrations along with methotrexate is shown in
Fig. 4 and the representative images of embryos are shown in
Fig. 5. Compound 2f showed dose-dependent increase in apop-
totic activity along with promising activities both at 10 and
30 µM. The EC50 and the percentage induction of apoptosis of
the compounds were calculated. The EC50 of compound 2f was
found to be 8.88. Pro-apoptotic chemotherapeutic drugs
provide an approach to overcoming the clinical problem of
Fig. 5 Representative images of the embryos treated with methotrexate
and compound 2f assayed for apoptosis.
drug resistance25 and hence the present class of molecules cyclic double bond, the geometry of which was assigned as cis
may have potential medicinal value. in the case of compounds containing indole-1,2-fused 8-mem-
In conclusion, we have described a strategy based on Pd- bered rings. A large number of fused indole derivatives were
mediated intramolecular endo-trig ring closure of 1,2-disubsti- synthesized using this robust methodology and a representa-
tuted indoles leading to indole-1,2-fused 8- and 9-membered tive compound showed promising apoptotic properties when
rings for the identification of new and potential scaffolds for tested in zebrafish embryos. As most of the cytotoxic anti-
apoptosis. The methodology involved the use of unactivated cancer agents are known to induce apoptosis the present class
olefin and proceeded with regioselective formation of an endo- of indoles seemed to possess potential medicinal value. The
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strategy presented here could therefore be useful for the
design and discovery of potential new drugs.
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