Synthesis of 1,6,7,8-tetrahydro-naphtho[2,3-d]-azepino[4,5-b]indole-9,14-diones and their inhibitory effects on pro-inflammatory cytokines

Article abstract of DOI:10.1016/j.bmcl.2009.07.154

A rapid route to a series of naphthoquinone-fused indole derivatives via irradiation in a modified commercial domestic microwave is reported. The desired products were produced in high yields and short reaction times. The naphthoquinone-fused indole deriv

Full text of DOI:10.1016/j.bmcl.2009.07.154

Bioorganic & Medicinal Chemistry Letters 19 (2009) 5753–5756
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis of 1,6,7,8-tetrahydro-naphtho[2,3-d]-azepino[4,5-b]indole-9,14-
diones and their inhibitory effects on pro-inflammatory cytokines
a
Waya S. Phutdhawong a, , Wanwikar Ruensamran , Weerachai Phutdhawong b,c, Thongchai Taechowisan d
*
a Department of Chemistry, Faculty of Science, Silpakorn University, Nakorn Pathom 73000, Thailand
b Department of Chemistry, Faculty of Liberal Arts and Science, Kasetsart University, Kampang Saen Campus, Nakhon Pathom 73140, Thailand
c Research Promotion and Technology Transfer Center, Faculty of Liberal Arts and Science, Kasetsart University, Kampang Saen Campus, Nakhon Pathom 73140, Thailand
d Department of Microbiology, Faculty of Science, Silpakorn University, Nakhon Pathom 73000, Thailand
a r t i c l e i n f o
a b s t r a c t
Article history:
A rapid route to a series of naphthoquinone-fused indole derivatives via irradiation in a modified com-
mercial domestic microwave is reported. The desired products were produced in high yields and short
reaction times. The naphthoquinone-fused indole derivatives were evaluated for their pro-inflammatory
cytokines responses using lipopolysaccharide (LPS)-stimulated RAW264.7 murine macrophages. The
results showed that most of the tested compounds inhibit the production of nitric oxide (NO), prostaglan-
Received 25 June 2009
Revised 20 July 2009
Accepted 31 July 2009
Available online 4 August 2009
din (PG)E2, tumour necrosis factor (TNF)-
LPS.
a
, interleukin (IL)-6 and IL-1b in RAW264.7 cells treated with
Keywords:
Aminonapthoquinones
Naphthoquinone derivatives
Indole derivatives
Azepine
Ó 2009 Elsevier Ltd. All rights reserved.
Pro-inflammatory cytokines
Quinones are common in a variety of natural products and have
been reported to possess a broad range of biological activities
including anticancer,1–5 antibacterial,1,6–9 antimalarial,10–12 anti-
fungal9,13 and anti-inflammatory14,15 activities. Also aminonaph-
thoquinones were claimed to show promising antimalarial,
require forcing conditions or prolonged reaction times, a wide vari-
ety of organic reactions can be conducted very rapidly and in high
yields. On the basis of the above consideration, we now wish to re-
port the synthesis of naphtho[2,3-d]-azepino[4,5-b]indole-9,14-
dione derivatives (Fig. 1) via microwave assisted intramolecular
cyclisation and the evaluation of pro-inflammatory activities on
lipopolysaccharide (LPS)-induced RAW264.7 cells.
The substituted aminonaphthoquinones (3) required for cycli-
sation were prepared under irradiation using a modified commer-
cial domestic microwave oven.20 Nucleophilic substitution of
naphthoquinones (1a–c) with tryptamine derivatives (2a–b) affor-
ded the corresponding substituted aminonaphthoquinones (3a–e)
in moderate to high yields21 by heating in a microwave reactor
at 850 W for 4 min.20 The palladium catalysed intermolecular reac-
tion of 3a and 3b under microwave irradiation at 850 W for 6 min
failed to yield the cyclised products. However, under the same
antiplatelet, anti-inflammatory and antiallergenic activities.16,17
A
number of naphthoquinones fused with heterocyclic rings have
been synthesised and it was found that the presence of a heterocy-
cle fused to the quinone moiety also played a crucial role in the
biological activity of the compounds.18,19 As part of our research
program to synthesise novel bioactive compounds, we became
interested in naphtho[2,3-d]-azepino[4,5-b]indole-9,14-dione, in
which seven-membered ring heterocyclic naphthoquinones are
fused to the indole nucleus, since indoles also possess a wide vari-
ety of biological properties. We anticipated that the combined
chemical motif of a naphthoquinone, an indole group and an aze-
pine moiety in the same molecule would markedly enhance the
anti-inflammatory activity as each compound is predominantly
seen in a wide range of anti-inflammatory agents. To the authors’
best knowledge, there is no report concerning the synthesis and
biological activities of 1,4-naphthoquinones fused through the qui-
none nucleus to the azepino-indole moiety. With the increased role
of microwave chemistry, especially when conventional methods
R3
O
N
R1
O
HN
R2
R1, R2= H, OMe
R3= H, CH3, C3H7, Allyl, Bn
* Corresponding author. Tel.: +66 34255797.
E-mail address: sengwaya@su.ac.th (W.S. Phutdhawong).
Figure 1. General structure of naphtho[2,3-d]-azepino[4,5-b]indole-9,14-dione.
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.07.154

5754
W. S. Phutdhawong et al. / Bioorg. Med. Chem. Lett. 19 (2009) 5753–5756
R3
NH2
N
H
O
O
O
O
2a; R3 = H
O
O
H
N
H
N
R2
R2
2b; R3= OMe
R3
b
R2
a
R1
R1
N
H
R1
HN
1a; R1 = R2 = H
R3
3a; R1 = R2 = R3 = H (40% yield)
1b; R1 = H, R2= Br
1c; R1 = OMe, R2 = Br
3b; R1 = R2 = H, R3 = OMe (38% yield)
3c; R1 = H, R2 = Br, R3 = H (89% yield)
3d; R1 = OMe, R2 = Br, R3 = H (53% yield)
3e; R1 = H, R2 = Br, R3 = OMe (75% yield)
4a; R1 = R3 = H (94% yield)
4b; R1 = OMe, R3 = H (72% yield)
4c; R1 = H, R3 = OMe (49% yield)
Scheme 1. Reagents and conditions: (a) C2H5OH, microwave, 850 W, 4 min (b) 0.1 equiv Pd(OAc)2, 0.2 equiv Ph3P, 3 equiv K2CO3, microwave, 850 W, DMF, 6 min.
R4
N
bility was observed. In the measurement of NO production
stimulated by LPS (1 g/mL), treatment of the tested compounds
(1–5 g/mL) inhibits the production of NO in a dose-dependent
O
O
O
O
H
N
l
a
l
manner (Fig. 2).
R1
R1
HN
HN
The effect of the tested compounds on LPS (1 lg/mL) induced
production of PEG2 in RAW 264.7 was shown in Figure 3. The
tested compounds could dramatically inhibit the production of
R3
R3
4a; R1 = R3 = H
5a; R1 = R3 = H, R4 = Me (96% yield)
5b; R1 = R3 = H, R4 = C3H7 (36% yield)
5c; R1= R3 = H, R4 = Allyl (50% yield)
5d; R1= R3 = H, R4 = Benzyl (43% yield)
PGE2 at a concentration of 5
c and 5d slightly inhibited the production of PGE2 at a concentra-
tion of 1 g/mL.
Further studies on the pro-inflammatory cytokines such as
TNF-
, IL-1b and IL-624–26 that are thought to be interlinked in a
lg/mL, however only compound 4b–
l
Scheme 2. Reagents and conditions: (a) R4X, K2CO3, microwave, 850 W, DMF,
8 min.
a
cascade, being produced serially by macrophages during inflam-
matory response demonstrated that the tested compounds have
inhibitory effects on the production of TNF-
LPS-stimulated RAW 264.7 macrophages. As shown in Figures 4–
6, LPS-induced production of TNF- was significantly inhibited
by the tested compounds in a concentration-dependent manner.
However, all of the tested compounds mildly affected the produc-
tion of IL-1b and IL-6.
In summary, syntheses of the indole-fused azepinonaphthoqui-
nones (4a–c) were achieved with an intramolecular palladium-cat-
alysed cyclisation of aminonaphthoquinones bearing an indole
nucleus as the key step. In addition, N-alkylation of the NH group
on the severn-membered rings afforded the corresponding deriva-
tives 5a–d. The indole-fused azepinonaphthoquinones (4a–c and
5a–d) were tested for their pro-imflammatory activities. Most of
the tested compounds inhibited the production of NO, PGE2,
a, IL-1b and IL-6 in
reaction conditions, brominated aminonaphthoquinones (3c–e)
were cyclised to afford the indole fused seven-membered ring het-
erocyclic naphthoquinones (4a–c) (Scheme 1) in high yields unac-
companied by any reduced products.22
The effect on activity of eliminating the NH group of the aze-
pines was evaluated by selective N-alkylation of azepine moiety
4a using various alkyl halides and potassium carbonate under
microwave irradiation at 850 W for 8 min (Scheme 2). Compounds
5a–d were obtained in a short reaction time and in moderate to
high yields.23
a
Compounds 4a–c and 5a–d were tested in LPS (1 lg/mL)-stim-
ulated RAW264.7 macrophages. The cytotoxic effects of these com-
pounds were evaluated in the presence or absence of LPS using the
MTT assay, and cell viabilities were observed to be 58–87% for all
tested compounds with the exception of 5c for which 24% cell via-
TNF-a
,
IL-1b and IL-6 in macrophages stimulated with LPS
80
70
60
1 ug/mL
*
2.5 ug/mL
5 ug/mL
*
*
50
**
***
40
***
***
30
***
***
5c
***
***
***
***
20
***
10
*** ***
4c
***
***
***
0
blank
ctrl
4a
4b
5a
5b
5d
LPS (1μg/mL)
Figure 2. Evaluation of nitric oxide production by RAW 264.7 cells stimulated for 24 h with LPS alone or in combination with increasing concentrations (1.0–5.0
lg/ml) of
*
**
***
heterocyclic naphthoquinones derivatives. The values are the means of at least three determinations SD. Probability level (student’s t-test): p <0.05, p <0.01, p <0.001
versus LPS-treated group.

W. S. Phutdhawong et al. / Bioorg. Med. Chem. Lett. 19 (2009) 5753–5756
5755
1 ug/mL
40
30
20
10
0
2.5 ug/mL
*
5 ug/mL
**
*
***
***
***
***
***
***
***
***
***
***
***
***
***
***
***
blank
ctrl
4a
4b
4c
5a
5b
5c
5d
LPS (1μg/mL)
Figure 3. Effect of heterocyclic naphthoquinones derivatives on PGE2 production in LPS-induced RAW 264.7 macrophage for 24 h. The values are the means of at least three
*
**
***
determinations SD. Probability level (student’s t-test): p <0.05, p <0.01, p <0.001 versus LPS-treated group.
1 ug/mL
2.5 ug/mL
40
5 ug/mL
*
*
*
**
*
30
20
10
0
***
***
***
***
***
***
***
***
***
***
***
***
***
***
blank
ctrl
4a
4b
4c
5a
5b
5c
5d
LPS (1μg/mL)
a production by RAW 264.7 cells. The values are the means of at least three
Figure 4. Effect of heterocyclic naphthoquinones derivatives on LPS-induced TNF-
*
**
***
determinations SD. Probability level (student’s t-test): p <0.05, p <0.01, p <0.001 versus LPS-treated group.
1 ug/mL
40
2.5 ug/mL
5 ug/mL
*
30
20
10
0
**
**
***
***
***
***
***
***
***
***
***
***
***
***
***
***
***
blank
ctrl
4a
4b
4c
5a
5b
5c
5d
LPS (1μg/mL)
Figure 5. Effect of heterocyclic naphthoquinones derivatives on LPS-induced IL-1b production by RAW 264.7 cells. The values are the means of at least three
determinations SD. Probability level (student’s t-test): *p <0.05, **p <0.01,***p <0.001 versus LPS-treated group.
in vitro. We conclude from our investigation that the series
of 1,6,7,8-tetra hexahydro-naphtho[2,3-d]-azepino[4,5-b]indole-
9,14-diones could be potential useful for the development of
anti-inflammatory agents.

5756
W. S. Phutdhawong et al. / Bioorg. Med. Chem. Lett. 19 (2009) 5753–5756
1 ug/mL
2.5 ug/mL
5 ug/mL
30
20
10
0
*
***
**
***
***
***
***
***
***
***
***
***
***
***
***
***
blank
ctrl
4a
4b
4c
5a
LPS (1μg/mL)
5b
5c
5d
Figure 6. Effect of heterocyclic naphthoquinones derivatives on LPS-induced IL-6 production by RAW 264.7 cells. .The values are the means of at least three
** ***
*
determinations SD. Probability level (student’s t-test): p <0.05, p <0.01, p <0.001 versus LPS-treated group.
20. Phutdhawong, W.; Buddhasukh, D. Molecules 2005, 10, 1409–1412.
21. Typical procedure: the preparation of 2-(1H-indol-3-yl)ethylamino-1,4-
naphthoquinone (3a). To solution of 1,4 naphthoquinones (1a) (50 mg,
Acknowledgements
a
We gratefully acknowledge support from The Thailand Re-
search Fund (Research grant No. MRG5080169) and the Depart-
ment of Chemistry, Silpakorn University for a scholarship (to W.R.).
0.32 mmol) in DCM (0.5 mL) was added a solution of tryptamine (2a) (61 mg,
0.38 mmol) in EtOH (3 mL) at room temperature under argon. The mixture was
irradiated in a microwave reactor at 850 W for 4 min and then cooled. The
mixture was concentrated in vacuo, and the residue was purified by column
chromatography (silica gel, hexane/EtOAc (1:1) to give 3a as a red solid with an
Rf value of 0.57 (40 mg, 40%), mp 184–185 °C; IR (CH2Cl2)
mmax: 1509, 1575,
Supplementary data
1609, 1678, 3382, 3469 cmÀ1 1H-NMR (CDCl3) d 3.16 (t, J = 6.8 Hz, 2H), 3.52 (q,
;
J = 6.4 Hz, 2H), 5.80 (s, 1H), 6.03 (br s, 1H), 7.09 (s, 1H), 7.15 (td, J = 0.8, 7.3 Hz,
1H), 7.23 (td, J = 1.1, 7.7 Hz, 1H), 7.39 (d, J = 8.0 Hz, 1H), 7.60 (td, J = 1.0, 7.6 Hz,
1H), 7.61 (d, J = 8.0 Hz, 1H), 7.72 (td, J = 1.2, 7.5 Hz, 1H), 8.01 (dd, J = 1.0, 7.6 Hz,
1H), 8.10 (dd, J = 0.8, 7.6 Hz, 1H), 8.15 (br s, 1H); 13C-NMR (CDCl3) d 24.2, 42.5,
100.9, 111.5, 112.7, 118.5, 119.7, 122.2, 122.5, 126.2, 126.3, 127.0, 130.5, 132.0,
133.7, 134.8, 136.5, 147.9, 181.0, 183.0; HRES-MS m/z calcd for [M+H]+
C20H17N2O2: 317.1285, found: 317.1251.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2009.07.154.
References and notes
1. Tandon, V. K.; Singh, R. V.; Yadav, D. B. Bioorg. Med. Chem. Lett. 2004, 14, 2901–
2904.
2. Yamashita, M.; Kaneko, M.; Iida, A.; Tokuda, H.; Nishimura, K. Bioorg. Med.
Chem. Lett. 2007, 17, 6417–6420.
3. Miguel del Corral, J. M.; Castro, M. A.; Oliveira, A. B.; Gualberto, S. A.; Cuevas, C.;
Feliciano, A. S. Bioorg. Med. Chem. 2006, 14, 7231–7240.
4. Miguel del Corral, J. M.; Castro, M. A.; Gordaliza, M.; Martin, M. L.; Gamito, A.
M.; Cuevas, C.; Feliciano, A. S. Bioorg. Med. Chem. 2006, 14, 2816–2827.
5. Miguel del Corral, J. M.; Castro, M. A.; Gordaliza, M.; Martn, M. L.; Gualberto, S. A.;
Gamito, A. M.; Cuevas, C.; Feliciano, A. S. Bioorg. Med. Chem. 2005, 13, 631–644.
6. Mahapatra, A.; Mativandlela, S. P. N.; Binneman, B.; Fourie, P. B.; Hamilton, C. J.;
Meyer, J. J. M.; van der Kooy, F.; Houghton, P.; Lall, N. Bioorg. Med. Chem. 2007,
15, 7638–7646.
7. Tandon, V. K.; Yadav, D. B.; Singh, R. V.; Chaturvedi, A. K.; Shukla, P. K. Bioorg.
Med. Chem. Lett. 2005, 15, 5324–5328.
8. Machado, T. B.; Pinto, A. V.; Pinto, M. C. F. R.; Leal, I. C. R.; Silva, M. G.; Amaral, A.
C. F.; Kuster, R. M.; Netto-dosSantos, K. R. Int. J. Antimicrob. Agents 2003, 21,
279–284.
9. Gafner, S.; Wolfender, J.-L.; Nianga, M.; Stoeckli-Evans, H.; Hostettmann, K.
Phytochemistry 1996, 42, 1315–1320.
10. Likhitwitayawuid, K.; Kaewamatawong, R.; Ruangrungsi, N.; Krungkrai, J.
Planta Med. 1998, 64, 237–241.
22. Typical procedure: the preparation of 1,5,6,7,8,14b-hexahydro-naphtho[2,3-d]-
azepino[4,5-b]indole-9,14-dione (4a). A mixture of aminonaphthoquinone (3a)
(46 mg, 0.12 mmol), Pd(OAc)2 (3 mg, 0.012 mmol), P(Ph)3 (7 mg, 0.024 mmol)
and K2CO3 (48 mg, 0.35 mmol) in DMF (4.0 ml) was irradiate in the modified
domestic microwave reactor at 850 W for 6 minutes under an Ar atmosphere.
After cooling, the DMF was evaporated, and then EtOAc (15 mL) was added and
washed several times with water. The EtOAc layer was washed with brine,
dried, evaporated and subjected to flash column chromatography (silica gel,
2:1 hexane/EtOAc) to give 4a with an Rf value of 0.73 (35 mg, 94%) as a blue
solid, mp 218–219 °C; IR (CH2Cl2) mmax
: 1519, 1566, 1599, 1630, 1668,
3347 cmÀ1 1H NMR (CDCl3) d 3.16 (t, J = 4.5 Hz, 2H), 3.65 (q, J = 4.5 Hz, 2H),
;
7.00 (td, J = 1.0, 7.5 Hz, 1H), 7.10 (td, J = 1.0, 7.5 Hz, 1H), 7.31 (d, J = 8.0 Hz, 1H),
7.40 (d, J = 8.0 Hz, 1H), 7.50 (td, J = 1.2, 7.5 Hz, 1H), 7.62 (td, J = 1.2, 7.5 Hz, 1H),
7.91 (dd, J = 1.0, 7.5 Hz, 1H), 8.05 (dd, J = 1.0, 7.5 Hz, 1H), 11.46 (br s, 1H); 13C
NMR (CDCl3) d 26.4, 44.1, 105.0, 110.1, 114.4, 116.6, 118.2, 121.5, 125.0, 125.7,
126.2, 128.7, 128.8, 131.4, 133.1, 133.9, 134.5, 143.6, 180.1, 183.6; HRES-MS m/
z cacld for [M+H]+ C20H15N2O3: 315.1128, found: 315.1032.
23. Typical procedure: the preparation of 1,5,6,7,8,14b-hexahydro-naphtho[2,3-d]-
N-methyl-azepino[4,5-b]indole-9,14-dione (5a). To a solution of heterocyclic
naphthoquinones 4a (45 mg, 0.14 mmol) and K2CO3 (48 mg, 0.35 mmol) in dry
DMF (4 mL) was stirred at 0–5 °C for 30 minutes under Argon atmosphere was
added methyl iodide (0.03 mL, 0.43 mmol) and the mixture irradiated in the
microwave reactor at 850 W for 8 minutes. Water (15 ml) was then added to
the reaction mixture and the mixture was extracted with EtOAc (2 Â 10 ml).
The EtOAc layer was dried, evaporated and subjected to flash column
chromatography (silica gel, 4:1 hexane/EtOAc) to give 5a with an Rf value of
11. dos Santos, E. V. M.; Carneiro, J. W. d. M.; Ferreira, V. F. Bioorg. Med. Chem. 2004,
12, 87–93.
12. Kittakoop, P.; Punya, J.; Kongsaeree, P.; Lertwerawat, Y.; Jintasirikul, A.;
Tanticharoen, M.; Thebtaranonth, Y. Phytochemistry 1999, 52, 453–457.
13. Tandon, V. K.; Maurya, H. K.; Tripathi, A.; ShivaKeshava, G. B.; Shukla, P. K.;
Srivastava, P.; Panda, D. Eur. J. Med. Chem. 2009, 44, 1086–1092.
14. Checker, R.; Sharma, D.; Sandur, S. K.; Khanam, S.; Poduval, T. B. Int.
Immunopharmacol. 2009, 9, 949–958.
15. Ferreira, M. A. D.; Nunes, O. D. R. H.; Fontenele, J. B.; Pessoa, O. D. L.; Lemos, T. L.
G.; Viana, G. S. B. Phytomedicine 2004, 11, 315–322.
16. Kapadia, G.; Azuine, M. A.; Balasuramanian, V.; Sridhar, R. Pharmaco. Res. 2001,
43, 363–367.
0.60 (45 mg, 96%) as a blue solid, mp 218–219 °C; IR (CH2Cl2)
mmax: 1542, 1595,
1634, 1664, 3379 cmÀ1 1H NMR (CDCl3) d 3.16 (t, J = 4.9 Hz, 2H), 3.27 (t,
;
J = 4.9 Hz, 2H), 3.29 (s, 3H), 7.03 (td, J = 1.0, 7.4 Hz, 1H), 7.15 (td, J = 1.0, 7.4 Hz,
1H), 7.33 (d, J = 8.0 Hz, 1H), 7.49 (d, J = 8.0 Hz, 1H), 7.56 (td, J = 1.5, 7.5 Hz, 1H),
7.62 (td, J = 1.5, 7.5 Hz, 1H), 7.92 (dd, J = 1.3, 7.5 Hz, 1H), 8.07 (dd, J = 1.3,
7.5 Hz, 1H), 11.32 (br s, 1H).; 13C NMR (CDCl3) d 24.3, 42.3, 53.2, 110.0, 112.7,
116.2, 117.6, 118.3, 122.2, 124.9, 125.4, 126.6, 128.9, 130.5, 131.8, 132.3, 132.8,
134.6, 149.7, 181.8, 185.5; HRES-MS m/z calcd for [M+H]+ C21H16N2O2:
329.1290, found: 329.1247.
17. Lien, J.-C.; Huang, L.-J.; Wang, J.-P.; Teng, C.-M.; Lee, K.-H.; Kuo, S.-C. Bioorg.
Med. Chem. 1997, 5, 2111–2120.
18. Wang, X. L.; Zheng, X. F.; Liu, R. H.; Reiner, J.; Chang, J. B. Tetrahedron 2007, 63,
3389–3394.
24. Chang, H. P.; Huang, S. Y. J. Agric. Food Chem. 1994, 53, 2530–2534.
25. Feldmann, M.; Brennan, F. M.; Maina, R. N. Annu. Rev. Immunol. 1996, 14, 397–
440.
19. SungKim, J.; Lee, H.-J.; Suh, M.-E.; Choo, H.-Y. P.; Lee, S. K.; Park, H. J.; Kim, C.;
Park, S. W.; Lee, C.-O. Bioorg. Med. Chem. 2004, 12, 3683–3686.
26. Hasko, G.; Nemeth, Z. H.; Szabo, C. Brain Res. Bull. 1998, 45, 183–187.