Synthesis and biological evaluation of new pyrrolopyrazinone compounds as potential antitumor agents

Article abstract of DOI:10.1016/j.cclet.2013.04.020

A series of pyrrolo[1,2-a]pyrazinone compounds (5a-9f) were synthesized, and their cytotoxic activity against SKOV-3, A549, HeLa cells in vitro were evaluated by the MTT method. Some of the compounds showed potential antitumor activity against three tumor cell lines. Among them, compounds 9c and 9d showed the most potent cytotoxic activity. The preliminary mechanism of action was discussed.

Full text of DOI:10.1016/j.cclet.2013.04.020

Chinese Chemical Letters 24 (2013) 619–621
Contents lists available at SciVerse ScienceDirect
Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
Original article
Synthesis and biological evaluation of new pyrrolopyrazinone compounds as
potential antitumor agents
b,
Ying Meng a, Guan Wang b, Yue Li a, Kuan Hou a, Yue Yuan a, Li-Juan Zhang a, Hong-Rui Song a, , Wei Shi
*
*
a Key Laboratory of Structure-Based Drug Design and Discovery, Ministry of Education, Shenyang Pharmaceutical University, Shenyang 110016, China
b Key Laboratory for Molecular Enzymology and Engineering, Ministry of Education, Jilin University, Changchun 130012, China
A R T I C L E I N F O
A B S T R A C T
Article history:
A series of pyrrolo[1,2-a]pyrazinone compounds (5a–9f) were synthesized, and their cytotoxic activity
against SKOV-3, A549, HeLa cells in vitro were evaluated by the MTT method. Some of the compounds
showed potential antitumor activity against three tumor cell lines. Among them, compounds 9c and 9d
showed the most potent cytotoxic activity. The preliminary mechanism of action was discussed.
ß 2013 Hong-Rui Song and Wei Shi. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All
rights reserved.
Received 6 March 2013
Received in revised form 24 March 2013
Accepted 28 March 2013
Available online 20 May 2013
Keywords:
Pyrrolopyrazinone
Antitumor
Synthesis
1. Introduction
by reacting with chloroacetone. The overall yield of these two steps
from 2 to 4 was 56%. Then compound 4 was converted to 5a–5b by
Modification of natural product leads is an important approach
to identify promising anticancer agents [1]. Pyrrolo[1,2-a]pyrazi-
none core is exclusively found in the natural products from marine
sponges, mainly from the families Agelasidae, Axinellidae [2].
Longamide B and Palau’amine are two examples (Fig. 1). Long-
amide B methyl ester [3], which was isolated initially in racemic
form from the sponge mentioned above in Japan, exhibits cytotoxic
activity against the same leukemia cell line in vitro [4].
Palau’amine, isolated from the sponge Stylotella aurantium by
Scheuer and co-workers [5], also exhibits cytotoxic properties [6].
Encouraged by the above results, recently we simplified the
structure of these natural product, carried out the synthesis of a
series of novel pyrrolo[1,2-a]pyrazinone compounds. We found
several compounds with potent cytotoxicity, which was worthy of
further investigation.
treatment with the corresponding amine, and 5a–5b were
converted to 6a–6d by treatment with the corresponding acyl
chloride in DCM.
Methyl 2-pyrrolecarboxylate 3 was converted to methyl 1-(2-
oxo-2-p-tolylethyl)-1H-pyrrole-2-carboxylate 7 by treatment with
a-bromo-4-methoxyacetophenone in 76% yield, then compound 7
was converted to 8a–8b by treatment with the corresponding
amine, and 8a–8b were converted to 9a–9d by treatment with the
corresponding acyl chloride in DCM. The structures of compounds
5a–9f were characterized [7].
3. Results and discussion
All target compounds were evaluated for cytotoxicity against
three cells in vitro by the MTT method, using 5-Fu as a control.
Analysis of cytotoxicity presented in Table 1 for these compounds
[F(ig._1)TD$FIG]
2. Experimental
O
H N
Br
2
H
As depicted in Scheme 1, the starting material pyrrole 1 was
firstly converted to 2-(trichloroacetyl)pyrrole 2 by treatment with
trichloroacetyl chloride in 62% yield. Then esterification of 2 using
CH3ONa in CH3OH afforded methyl 2-pyrrolecarboxylate 3, which
was converted to 3-methyl-lH-pyrrolo[2,1-c][1,4]oxazin-l-one 4
N
H
O
Br
N
N
Cl
NH
NH
NH
HO
NH
N
2
N
H
COOCH
3
2
Palau'amine
Longamide B methyl ester
* Corresponding authors.
Fig. 1. The structure of Longamide B methyl ester and Palau’amine.
E-mail addresses: hongruisong@163.com (H.-R. Song), Shiw@jlu.edu.cn (W. Shi).
1001-8417/$ – see front matter ß 2013 Hong-Rui Song and Wei Shi. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
http://dx.doi.org/10.1016/j.cclet.2013.04.020

[
Y. Meng et al. / Chinese Chemical Letters 24 (2013) 619–621
O
O
n
O
O
n
R
1
a
e
N
N
b
d
c
N
NH
O
2
H
COCCl
COOCH
3
3
N
N
H
N
H
N
H
N
N
CH
3
CH
CH
3
3
2
1
3
4
6a-6d
5a-5b
O
n
n
O
O
R
COOCH
O
1
3
N
N
NH
2
N
N
g
H
f
h
COOCH
3
N
N
N
H
3
7
CH
8a-8b
CH
9a-9f
3
3
CH
3
5a 5b
6a
6b
2
H
6c
0
Cl
6d
8a
0
-
8b
2
-
9a
0
H
9b
2
H
9c
9d
9e
9f
n
R1
0
-
2
-
0
2
0
2
0
2
H
Cl
Cl
Cl
CH2Cl
CH2Cl
Scheme 1. Reagents and conditions: (a) trichloroacetyl chloride, ether, r.t., (b) CH3ONa, CH3OH, r.t., (c) chloroacetone, acetone, r.t., (d) corresponding amine, CH3CH2OH, r.f.,
(e) corresponding acyl chloride, DCM, 0 8C, (f)
chloride, DCM, 0 8C.
a-bromo-4-methoxyacetophenone, K2CO3, DMF, r.t., (g) corresponding amine, CH3CH2OH, r.f., and (h) corresponding acyl
[(Fig._2)TD$FIG]
Fig. 2. Effects of compound 9c on the induction of apoptosis in A549 cells. (A) Karyomorphism altered after treated with compound 9c as revealed by the DNA-binding AT-
specific fluorochrome 40-6-diamidino-2-phenylindole (DAPI) fluorescence staining. (B) Annexin V-FITC was proceeded and the lower right quadrant represents early
apoptosis.
and their structural features had revealed several patterns of
structure-activity relationship. (1) Compounds 5a, 5b and 6a–6d
exhibited weak or no cytotoxicity compared to 8a, 8b and 9a–9f,
suggesting that the presence of large or hydrophobic groups at C3
enhanced their antitumor activity significantly in all three cancer
cell lines. (2) Compounds with R1 = Cl (6c, 6d, 9c, and 9d) showed
highest cytotoxicity, The chlorine atom was three atoms away
from the nucleus in 6c and 9c and five atoms away from the
nucleus in 6d and 9d, while the chlorine atom in Palau’amine was 4
atoms away from the nucleus, Comparing to the structure of
Palau’amine a conclusion could be drawn that a certain length of
chain with a chlorine atom might be essential for the antitumor
activity. Moreover, it could be seen from Fig. 2 that compound 9c
inhibits cell proliferation by inducing apoptosis in A549 cells.
4. Conclusion
In summary, a series of pyrrolo[1,2-a]pyrazinone derivatives
were synthesized and evaluated for their in vitro cytotoxicity
against SKOV-3, A549 and HeLa cells. DAPI staining and Annexin V/
PI experiments showed that compound 9c inhibits cell prolifera-
tion by inducing apoptosis in A549 cells. The study will provide
valuable information for further research on the pyrazinone
antitumor analogs.
Table 1
IC50s (mmol/L) of compounds against three tumor cell lines. The IC50 values of
compounds against SKOV-3, A549, HeLa cells were evaluated by methylthiazolte-
trazolium (MTT) assay. ND: not determined.
Compounds
SKOV-3
A549
HeLa
5a
5b
6a
6b
6c
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
43.803
39.118
22.107
27.314
>50
49.239
44.117
References
6d
8a
8b
9a
9b
9c
41.109
26.551
23.477
40.554
29.332
30.736
[1] F. Deng, J.J. Lu, H.Y. Liu, Synthesis and antitumor activity of novel salvicine
analogues, Chin. Chem. Lett. 22 (2011) 25–28.
>50
>50
8.967
>50
>50
5.551
[2] E.N. Delphine, M. Peter, L. Thomas, Chiroptical analysis of marine sponge alkaloids
sharing the pyrrolopyrazinone core, Chem. Eur. J. 10 (2004) 1141–1148.
[3] A. Umeyama, S. Ito, E. Yuasa, et al., A new bromopyrrole alkaloid and the optical
resolution of the racemate from the marine sponge Homaxinella sp., J. Nat. Prod. 61
(1998) 1433–1434.
[4] G.B. Martin, M.B. Andrew, C.W. Anthony, First syntheses of the pyrroloketopiper-
azine marine natural products (ꢀ)-longamide, (ꢀ)-longamide B, (ꢀ)-longamide B
methyl ester and (ꢀ)-hanishin, New J. Chem. 23 (1999) 687–690.
>50
6.908
5.438
34.157
40.126
ND
9d
9e
9f
12.184
29.639
38.965
86.934
10.532
35.682
43.683
61.438
5-Fu

Y. Meng et al. / Chinese Chemical Letters 24 (2013) 619–621
621
[5] R.B. Kinnel, H.P. Gehrken, R. Swali, Palau’amine and its congeners: a family of
bioactive bisguanidines from the marine sponge Stylotella aurantium 1, J. Org.
Chem. 63 (1998) 3281–3286.
(300 MHz, DMSO-d6): d 7.11 (d, 2H, J = 8.2 Hz), 7.03 (d, 2H, J = 8.2 Hz), 6.72–
6.67 (m, 1H), 6.56 (dd, 1H), 5.97 (dd, 1H), 4.55 (d, 1H), 4.30 (t, 1H), 4.23 (d,
1H), 3.54 (dt, 1H), 3.47–3.35 (m, 1H), 2.18 (s, 3H), 1.02 (t, 2H). 9a: Yield 29%. Mp:
111–112 8C; ESI-MS: m/z 282 [M+H]+; 1H NMR (300 MHz, DMSO-d6): d 10.52 (s,
1H), 7.45 (dd, 1H), 7.37 (s, 1H), 7.27 (d, 2H, J = 8.1 Hz), 7.19 (d, 2H, J = 8.1 Hz), 6.96
(d, 1H, J = 3.9 Hz), 6.57 (dd, 1H, J = 3.9, 2.6 Hz), 2.31 (s, 3H), 1.71 (s, 3H). 9b: Yield
31%. Mp: 194–197 8C; ESI-MS: m/z 310 [M+H]+; 1H NMR (300 MHz, DMSO-d6): d
7.18 (d, 2H, J = 8.3 Hz), 7.13–7.09 (m, 1H), 7.06 (d, 2H, J = 8.3 Hz), 6.60 (dd, 1H), 6.09
(dd, 1H), 5.62 (d, 1H, J = 13.0 Hz), 4.34 (d, 1H, J = 13.0 Hz), 4.22–4.10 (m, 1H), 3.94
(dd, 1H), 3.88–3.77 (m, 1H), 3.62 (dt, 1H), 2.21 (s, 3H), 2.04 (s, 3H). 9c: Yield 23%.
Mp: 110–113 8C; ESI-MS: m/z 316 [M+H]+; 1H NMR (300 MHz, DMSO-d6): d 7.90 (d,
2H, J = 8.2 Hz), 7.35 (d, 2H, J = 8.2 Hz), 7.13–7.08 (m, 1H), 6.87 (dd, 1H), 6.14 (dd,
1H), 5.81 (s, 2H), 3.57 (s, 2H), 2.37 (s, 3H). 9d: Yield 23%. Mp: 191–193 8C; ESI-MS:
m/z 343.9 [M+H]+; 1H NMR (300 MHz, DMSO-d6): d 7.15 (d, 2H, J = 8.3 Hz), 7.12–
7.09 (m, 1H), 7.05 (d, 2H, J = 8.3 Hz), 6.59 (dd, 1H), 6.07 (dd, 1H), 5.60 (d, 1H), 4.40
(dt, 3H), 4.16 (dt, 1H), 4.03–3.80 (m, 2H), 3.62 (dt, 1H), 2.18 (s, 3H). 9e: Yield 32%.
Mp: 123–125 8C; ESI-MS: m/z 330 [M+H]+; 1H NMR (300 MHz, DMSO-d6): d 10.71
(s, 1H), 7.49–7.33 (m, 2H), 7.25 (d, 2H, J = 7.9 Hz), 7.17 (d, 2H, J = 7.9 Hz), 6.97 (s,
1H), 6.57 (s, 1H), 3.57 (t, 2H), 2.70–2.52 (m, 2H), 2.28 (s, 3H). 9f: Yield 27%. Mp:
192–194 8C; ESI-MS: m/z 357.5 [M+H]+, 380 [M+Na]+; 1H NMR (300 MHz, DMSO-
d6): d 7.18 (d, 2H, J = 8.3 Hz), 7.12 (d, 1H), 7.04 (d, 2H, J = 8.3 Hz), 6.71–6.51 (m, 2H),
6.26–6.00 (m, 2H), 5.82–5.52 (m, 2H), 4.33 (m, 1H), 4.22–4.09 (m, 1H), 4.04 (m, 1H),
3.99–3.89 (m, 1H), 3.73 (dt, 1H), 3.68–3.54 (m, 1H), 2.18 (s, 3H).
[6] L. Thomas, J.E.N. Delphine, Z. Michael, Study on the absolute configuration of (ꢁ)-
palau’amine, Tetrahedron Lett. 51 (2010) 6353–6355.
[7] Data for new compounds. 5a: Yield 62%. Mp: 147–150 8C; ESI-MS: m/z 163.9
[M+H]+; 1H NMR (300 MHz, DMSO-d6): d 6.95 (s, 1H), 6.63 (d, 1H), 6.19–6.12 (m,
1H), 4.83 (s, 1H), 4.67 (s, 2H), 1.13 (s, 3H). 5b: Yield 67%. Mp: 153–155 8C; ESI-MS:
m/z 192.2 [M+H]+; 1H NMR (300 MHz, DMSO-d6): d 6.99–6.93 (m, 1H), 6.60 (dd,
1H), 6.13 (dd, 1H), 4.24 (d, 1H), 3.89 (d, 1H), 3.51 (dt, 1H), 3.36–3.24 (m, 2H), 3.13
(m, 2H), 1.17 (s, 3H). 6a: Yield 31%. Mp: 144–146 8C; ESI-MS: m/z 205.9 [M+H]+; 1
H
NMR (300 MHz, DMSO-d6): d 7.32 (dd, 1H), 7.24 (s, 1H), 6.83–6.80 (m, 1H), 6.50
(dd, 1H), 5.37 (s, 1H), 2.20 (s, 3H), 1.13 (s, 3H). 6b: Yield 27%. Mp: 182–185 8C; ESI-
MS: m/z 234.2 [M+H]+; 1H NMR (300 MHz, DMSO-d6): d 7.33 (m, 1H), 7.25 (dd, 1H),
6.84 (dd, 1H), 6.50 (s, 1H), 4.06 (m, 2H), 3.50 (m, 2H), 2.24 (s, 3H), 1.23 (s, 3H). 6c:
Yield 36%. Mp: 176–178 8C; ESI-MS: m/z 261.9 [M+Na]+; 1H NMR (300 MHz,
DMSO-d6): d 7.33 (dd, 1H), 7.24 (s, 1H), 6.84–6.79 (m, 1H), 6.50 (dd, 1H), 5.39
(s, 2H), 2.21 (s, 3H). 6d: Yield 34%. Mp: 200–202 8C; ESI-MS: m/z 267.9 [M+H]+; 1
H
NMR (300 MHz, DMSO-d6): d 7.11–7.05 (m, 1H), 6.66 (dd, 1H), 6.14 (dd, 1H), 4.94
(d, 1H), 4.94 (d, 1H), 4.51–4.29 (m, 1H), 4.07 (dt, 1H), 3.99 (d, 1H), 3.95–3.78 (m,
1H), 3.73 (dd, 1H), 3.45 (dt, 1H), 1.40 (s, 3H). 8a: Yield 73%. Mp: 139–141 8C; ESI-
MS: m/z 240 [M]+; 1H NMR (300 MHz, DMSO-d6): d 7.44 (d, J = 8.2 Hz, 1H), 7.09 (s,
1H), 7.03 (d, J = 8.2 Hz, 2H), 6.66 (d, 1H), 6.55 (s, 1H), 5.93–5.88 (m, 1H), 5.50 (s, 2H),
2.20 (s, 3H). 8b: Yield 78%. Mp: 191–193 8C; ESI-MS: m/z 268 [M]+, 1H NMR