Synthesis and antitumor evaluation of novel Benzo[d]pyrrolo[2,1-b]thiazole derivatives

Article abstract of DOI:10.1016/j.ejmech.2012.03.030

A series of novel 2,3-bis(hydroxymethyl)benzo[d]pyrrolo[2,1-b]thiazoles and their bis(alkylcarbamate) derivatives were synthesized starting from benzothiazole via reaction with dimethyl acetylenedicarboxylate (DMAD)/tetra-fluoro boric acid, catalytic hydr

Full text of DOI:10.1016/j.ejmech.2012.03.030

European Journal of Medicinal Chemistry 53 (2012) 28e40
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European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and antitumor evaluation of novel Benzo[d]pyrrolo[2,1-b]thiazole
derivatives
,
,
Ravi Chaniyara a b, Satishkumar Tala a, Chi-Wei Chen a c, Pei-Chih Lee a, Rajesh Kakadiya a, Huajin Dong d,
Bhavin Marvania a e, Ching-Huang Chen a, Ting-Chao Chou d, Te-Chang Lee a , Anamik Shah ,
,
,
b
*
,e,
**
Tsann-Long Su a
a Institute of Biomedical Sciences, Academia Sinica, Taipei 11529, Taiwan
b Department of Chemistry, Saurashtra University, Rajkot, Gujarat, India
c Institute of Biopharmaceutical Science, National Yang-Ming University, Taipei 11221, Taiwan
d Preclinical Pharmacology Core Laboratory, Molecular Pharmacology and Chemistry Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA
e Graduate Institute of Pharmaceutical Chemistry, China Medical University, Taichung, Taiwan
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of novel 2,3-bis(hydroxymethyl)benzo[d]pyrrolo[2,1-b]thiazoles and their bis(alkylcarbamate)
derivatives were synthesized starting from benzothiazole via reaction with dimethyl acetylenedi-
carboxylate (DMAD)/tetra-fluoro boric acid, catalytic hydrogenation, and alkylcarbamoylation. The anti-
proliferative activity of these agents against human leukemia and various solid tumor cell growth in vitro
was studied. The structureeactivity relationship studies revealed that the bis(alkylcarbamates) deriva-
tives are generally more cytotoxic than the corresponding bis(hydroxymethyl) congeners in inhibiting
human lymphoblastic leukemia CCRF-CEM and various human solid tumor cell growth in culture. These
agents have no cross-resistance to taxol or vinblastine. Studies on the therapeutic effect against human
breast carcinoma MX-1 xenograft showed that complete tumor remission (CR) were achieved by treating
with C1-40-F- or C1-40-Cl-Ph-bis(i-propylcarbamates) derivatives (19b and 19c, respectively) and more
than 99% tumor suppression by the corresponding bis(ethylcarbamates) 18b and 18c at the maximal
tolerated dose. Alkaline agarose gel shifting assay revealed that the newly synthesized compounds are
able to induce DNA interstrand cross-linking. The present studies generated a series of new potent DNA
interstrand cross-linking agents, which have potential for further antitumor drug development.
Ó 2012 Elsevier Masson SAS. All rights reserved.
Received 5 January 2012
Received in revised form
15 March 2012
Accepted 15 March 2012
Available online 29 March 2012
Keywords:
Antitumor agents
benzo[d]pyrrolo[2,1-b]thiazol
Cell cycle
Cytotoxicity
DNA cross-linking
1. Introduction
a bioreductive DNA bisalkylating agent [1e3]. This natural product
can induce DNA cross-linking via bioreactivation by reductase
The naturally occurring antibiotic antineoplastic Mitomycin C
(MMC 1, Fig. 1), isolated from Streptomyces caespitosis, is
under anaerobic conditions [4]. Thereby, this agent is effective for
treating hypoxia tumors. Another antitumor natural product, ret-
rorsine (2), which also contains pyrrolizidine pharmacophore, is
able to induce DNA cross-linking by a similar mechanism of action
as that of MMC [5]. However, the interaction of the pyrrolizidine
alkaloids with DNA double strands can proceed without the catalyst
of the reductase. Several synthetic compounds containing bis(hy-
droxymethyl)pyrrolidine or pyrrolizidine pharmacophore can
serve as DNA bifunctional alkylating agents. These agents are thi-
oimidazoles (i.e., carmethizole, 3) [6], 3,4-bis(methylcarbamate)
pyrroles (4) [7], 2,3-dihydroxy-6,7-bis(hydroxymethyl)pyrrolizines
(IPP, 5) [8], and their thio analogues 6,7-bis(hydroxymethyl)pyrrolo
[1,2-c]thiazole (6) [9], and dihydropyrrolo[2,1-b]thiazole (7) [10].
Among these derivatives, carmethizole hydrochloride (3) was
showed to have potent inhibitory activity against murine P388
leukemias. It is also reported that compound 5 has significant
Abbreviations: DMAD, dimethyl acetylenedicarboxylate; CR, complete tumor
remission; MMC, Mitomycin C; ATO, arsenic trioxide; CCRF-CEM, human lympho-
blastic leukemia; SAR, structureeactivity relationship; MX-1, human breast carci-
noma; SK-OV-3, ovarian adenocarcinoma; HCT-116, human colon carcinoma;
H1299, human lung cancer; PC-3, human prostate adenocarcinoma; OECM-1, oral
cancer; U87, glioma; H460, human large-cell lung carcinoma; (Q2D ꢀ 2), every
other days; two times, (Q2D ꢀ 3); every other days, three times; (Q2D ꢀ 4), every
other days; four times, (Q2D ꢀ 5); every other days, five times; DMF, N,N-dime-
thylformamide; THF, tetrahydrofuran; DMSO, dimethyl sulfoxide; PI, propidium
iodide; PBS, phosphate buffered saline.
* Corresponding author. Tel.: þ886 2 27899145; fax: þ886 2 27829142.
** Corresponding author. Institute of Biomedical Sciences, Academia Sinica, Taipei
11529, Taiwan. Tel.: þ886 2 27899045; fax: þ886 2 27829142.
E-mail addresses: bmtcl@ibms.sinica.edu.tw (T.-C. Lee), tlsu@ibms.sinica.edu.tw
(T.-L. Su).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.03.030

R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
29
Fig. 1. Chemical structure of some DNA bifunctional alkylating agents.
antitumor activity against a broad range of experimental marine
neoplasias and human tumor xenograft in nude athymic mice [8].
Lalezari et. al. reported that dihydropyrrolo[2,1-b]thiazole (7) was
as cytotoxic as IPP [10].
potent antitumor activity against human breast carcinoma MX-1 and
ovarian adenocarcinoma SK-OV-3 xenografts.
Study on the mechanism of action and the chemical properties of
thioimidazoles (e.g. 3, Fig. 1) or dihydropyrrolo[2,1-b]thiazole (7) sug-
gested that the sulfur atom participates in the expulsion of the
hydroxyl or carbamate moiety leading to the nucleophilic attack by
DNA [14]. To continue our research and development of new bifunc-
tional DNA-cross linking agents for antitumor application, we utilized
the known benzo[d]pyrrolo[2,1-b]thiazole diesters [15,16] to prepare
2,3-bis(hydroxymethyl)-4H-benzo[d]pyrrolo[2,1-b]thiazoles and their
bis(alkylcarbamate) derivatives (13, Fig.1) for antitumor studies. These
derivatives can be considered as the “benzologue” of dihydropyrrolo
[2,1-b]thiazole (7). One can expect that the sulfur atom of compound 13
may also involve in the DNA cross-linking as shown in thioimidazoles
(e.g. 3, Fig. 1) or dihydropyrrolo[2,1-b]thiazole (7). Herein, we report
the anti-proliferative activity against various human tumor cell growth
in vitro, therapeutic efficacy in xenograft model, as well as the mech-
anism of actions of the new benzo[d]pyrrolo[2,1-b]thiazole analogues.
Recently, we have synthesized a series of bis(hydroxymethyl)-8H-
3a-azacyclopenta[a]indene-1-yl and their bis(methylcarbamate)
derivatives, which can considered as a “benzologue” derivatives of
pyrrolizines (5) [11]. We have demonstrated that these agents
possess significant cytotoxicity in inhibiting human lymphoblastic
leukemia and a variety of human tumors in vitro and have potent
therapeutic efficacy in xenograft model. For example, the bis(hy-
droxymethyl) derivatives [BO-1090 (8) and BO-1099 (9)] and bis(-
methylcarbamates) derivatives, BO-1012 (10) and BO-1124 (11) are
able to induce complete tumor remission (CR) in nude mice bearing
human breast carcinoma MX-1 xenograft. Moreover, the bis(hy-
droxymethyl) derivatives, BO-1099 (9) significantly suppressed
(>95%) human prostate adenocarcinoma PC-3 xenograft. Remark-
ably, we found that the combination treatment of BO-1012 (10) with
arsenic trioxide (ATO, DNA repair inhibitor) achieved more than 82%
and 92% tumor suppression in nude mice bearing human large-cell
lung carcinoma H460 and cisplatin-resistant human bladder carci-
noma NTUB1/P xenografts, respectively [12]. More recently, we have
also synthesized a series of new bis(hydroxymethyl)pyrrolo[1,2-b]
isoquinolines and their bis(alkylcarbamate) derivatives for antitumor
evaluation [13]. Of these derivatives, BO-1107 (12) was shown to have
2. Results and discussion
2.1. Chemistry
The synthetic route for the preparation of (benzo[d]pyrrolo[2,1-b]
thiazole- 2,3-diyl)dimethanol (17aej) and their bis(alkylcarbamates)

30
R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
Scheme 1. Reaction Condition: (a) Acid Chloride/AlCl3, trimethylsilyl cynide/CH2Cl2, rt; (b) tetrafluoroboric acid in ether; (c) DMAD/DMF, ambient temperature; (d) LiAlH4, CH2Cl2/
ether 0 ꢁC; (e) R2NCO/TEA.
(18aej and 19aej) is shown in Scheme 1. Compound 15aej was
synthesized from the commercially available benzothiazole 14
according to the literature procedure [17]. By following the method
developed by Berrabah et al., [15], compounds 15aej were treated
with tetra-fluoro boric acid in ether, and directly further reacted with
dimethyl acetylenedicarboxylate (DMAD) toyieldthediester 16aejin
low yields. It should be noted that many attempts have been made to
synthesize compound 16 having a phenyl substituent at C1 (wherein
R1 ¼ Ph) under various conditions failed; a gummy polymer with
a complex mixture was obtained instead. The diester functions of
16aej were reduced to the corresponding bis-alcohol derivatives
17aej by reacting with LiAlH4 in a mixture of ether/CH2Cl2 in an ice
bath. Treatment of 17aej with ethyl or iso-propyl isocyanate in the
presenceoftriethylamineaffordedthedesiredbis(ethylcarbamate)or
Table 1
The cytotoxicity of newly synthesized 2,3-bis(hydroxymethyl)benzo[d]pyrrolo[2,1-b]thiazole (17aej) and their bis(alkylcarbamate) derivatives (18aej and 19aej) against
human lymphoblastic leukemia (CCRF-CEM), its drug-resistant sublines (CCRF-CEM/Taxol and CCRF-CEM/VBL).
Compd.
R1
R2
Cell growth inhibition (IC50
CCRF-CEM
m
M)a
CCRF-CEM/Taxolb
CCRF-CEM/VBLb
17a
17b
17c
17d
17e
17f
17g
17h
17i
Me
H
H
H
H
H
H
H
H
H
H
Et
Et
Et
Et
Et
Et
0.97 ꢂ 0.03
1.06 ꢂ 0.04
2.61 ꢂ 0.03
0.19 ꢂ 0.02
1.04 ꢂ 0.02
3.09 ꢂ 0.04
1.13 ꢂ 0.09
1.82 ꢂ 0.03
4.03 ꢂ 0.02
0.97 ꢂ 0.07
0.33 ꢂ 0.01
0.07 ꢂ 0.0002
0.21 ꢂ 0.01
0.21 ꢂ 0.01
0.38 ꢂ 0.001
0.38 ꢂ 0.01
0.19 ꢂ 0.002
0.49 ꢂ 0.003
0.32 ꢂ 0.001
0.17 ꢂ 0.03
0.13 ꢂ 0.01
0.05 ꢂ 0.001
0.08 ꢂ 0.0003
0.33 ꢂ 0.001
2.29 ꢂ 0.04
0.50 ꢂ 0.02
0.14 ꢂ 0.002
0.11 ꢂ 0.02
1.05 ꢂ 0.03
0.10 ꢂ 0.01
0.0007 ꢂ 0.0001
0.0012 ꢂ 0.0001
1.88 ꢂ 0.01 [1.93 ꢀ ]c
2.56 ꢂ 0.01 [2.41 ꢀ ]
5.22 ꢂ 0.03 [2.00 ꢀ ]
0.19 ꢂ 0.01 [1.00 ꢀ ]
NDd
1.86 ꢂ 0.01 [1.91 ꢀ ]
3.72 ꢂ 0.18 [3.50 ꢀ ]
4.09 ꢂ 0.03 [1.56 ꢀ ]
0.28 ꢂ 0.01 [1.47 ꢀ ]
ND
40-F-C6H4
40-Cl-C6H4
30,40-di-F-C6H3
30,40-di-Cl-C6H3
30-Clꢃ40ꢃF-C6H3
40-MeO-C6H4
30,40- di-MeO-C6H3
30,40,50-tri-MeO-C6H2
Cyclopropane
Me
ND
ND
2.17 ꢂ 0.02 [1.9 ꢀ ]
2.63 ꢂ 0.03 [2.30 ꢀ ]
ND
ND
ND
ND
17j
2.31 ꢂ 0.01 [2.37 ꢀ ]
0.45 ꢂ 0.01 [1.30 ꢀ ]
0.38 ꢂ 0.01 [5.42 ꢀ ]
0.32 ꢂ 0.01 [1.52 ꢀ ]
0.34 ꢂ 0.01 [1.61 ꢀ ]
ND
2.44 ꢂ 0.01 [2.51 ꢀ ]
0.43 ꢂ 0.01 [1.30 ꢀ ]
0.45 ꢂ 0.03 [6.42 ꢀ ]
0.39 ꢂ 0.01 [1.85 ꢀ ]
0.31 ꢂ 0.01 [1.47 ꢀ ]
ND
18a
18b
18c
18d
18e
18f
18g
18h
18i
40-F-C6H4
40-Cl-C6H4
30,40-di-F-C6H3
30,40-di-Cl-C6H3
30-Clꢃ40ꢃF-C6H3
40-MeO-C6H4
30,40- di-MeO-C6H3
30,40,50-tri-MeO-C6H2
Cyclopropane
Me
ND
ND
Et
Et
Et
Et
0.37 ꢂ 0.01 [1.90 ꢀ ]
0.42 ꢂ 0.01 [2.20 ꢀ ]
ND
ND
ND
ND
18j
0.68 ꢂ 0.01 [4.01 ꢀ ]
0.22 ꢂ 0.01 [1,69 ꢀ ]
0.16 ꢂ 0.01 [3.20 ꢀ ]
0.39 ꢂ 0.01 [4.87 ꢀ ]
0.19 ꢂ 0.01 [0.57 ꢀ ]
ND
0.54 ꢂ 0.01 [3.18 ꢀ ]
0.25 ꢂ 0.01 [1.92 ꢀ ]
0.19 ꢂ 0.01 [3,80 ꢀ ]
0.42 ꢂ 0.01 [5.25 ꢀ ]
0.30 ꢂ 0.01 [0.90 ꢀ ]
ND
19a
19b
19c
19d
19e
19f
19g
19h
19i
i-Pr
i-Pr
i-Pr
i-Pr
i-Pr
i-Pr
i-Pr
i-Pr
i-Pr
i-Pr
40-F-C6H4
40-Cl-C6H4
30,40-di-F-C6H3
30,40-di-Cl-C6H3
30-Clꢃ40ꢃF-C6H3
40-MeO-C6H4
30,40- di-MeO-C6H3
30,40,50-tri-MeO-C6H2
Cyclopropane
ND
ND
0.170.01 [1.17 ꢀ ]
0.25 ꢂ 0.01 [1.47 ꢀ ]
ND
ND
ND
ND
19j
Vinblastine
Taxol
0.73 ꢂ 0.01 [7.30 ꢀ ]
0.08 ꢂ 0.01 [106 ꢀ ]
0.43 ꢂ 0.04 [358 ꢀ ]
0.90 ꢂ 0.01 [9.00 ꢀ ]
0.50 ꢂ 0.12 [680 ꢀ ]
1.27 ꢂ 0.05 [980 ꢀ ]
a
Data represent the mean ꢂ SD of each compound from three to six independent experiments.
b
CCRF-CEM/VBL and CCRF-CEM/Taxol are sub-cell lines of CCRF-CEM cells with 680-fold resistance to vinblastine and 358-fold resistance to taxol, respectively, relative to
the IC50 value of the parent cell line.
c
Numbers in bracket are measures of cross-resistance as determined by comparison with the corresponding IC50 value of the parent cell line.
Not determined.
d

R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
31
Table 2
The cytotoxicity of benzo[d]pyrrolo[2,1-b]thiazole derivatives against human solid tumors (breast carcinoma MX-1, colon carcinoma HCT-116, lung carcinoma H1299, prostate
carcinoma PC-3, oral carcinoma OECM1 and glioma U87) cell growth in vitro.
Compd
IC50
(
m
M)a
MX-1
HCT-116
H1299
PC3
OECM-1
U87
17a
17b
17c
17d
17g
17j
18b
18c
18d
18g
18j
19a
19b
19c
19d
19g
19j
7.10 ꢂ 0.037
2.77 ꢂ 0.005
10.40 ꢂ 0.003
ND
ND
ND
0.48 ꢂ 0.005
0.91 ꢂ 0.01
1.49 ꢂ 0.04
ND
4.44 ꢂ 0.08
9.59 ꢂ 0.19
9.26 ꢂ 0.049
ND
ND
ND
0.52 ꢂ 0.025
0.70 ꢂ 0.008
1.10 ꢂ 0.015
ND
NDb
ND
ND
ND
9.27 ꢂ 1.69
12.43 ꢂ 0.44
21.35 ꢂ 3.74
9.94 ꢂ 1.87
13.32 ꢂ 2.23
13.55 ꢂ 2.03
4.33 ꢂ 0.45
10.33 ꢂ 1.11
8.85 ꢂ 1.53
20.64 ꢂ 1.89
9.70 ꢂ 1.10
33.03 ꢂ 3.20
8.75 ꢂ 0.88
30.75 ꢂ 6.92
8.19 ꢂ 1.74
20.44 ꢂ 1.65
16.53 ꢂ 0.90
13.78 ꢂ 2.15
20.41 ꢂ 3.43
15.22 ꢂ 1.86
8.81 ꢂ 1.20
10.74 ꢂ 2.29
23.70 ꢂ 3.05
13.85 ꢂ 3.78
19.27 ꢂ 3.44
17.90 ꢂ 3.59
24.19 ꢂ 2.96
ND
24.19 ꢂ 4.17
13.71 ꢂ 1.28
29.56 ꢂ 1.83
17.49 ꢂ 0.80
13.38 ꢂ 0.84
4.71 ꢂ 0.66
6.03 ꢂ 1.20
14.60 ꢂ 1.13
8.48 ꢂ 0.67
5.00 ꢂ 0.78
7.12 ꢂ 0.95
11.47 ꢂ 2.63
11.61 ꢂ 2.84
14.60 ꢂ 3.39
9.39 ꢂ 1.90
26.28 ꢂ 4.44
ND
11.89 ꢂ 2.90
7.54 ꢂ 0.95
15.10 ꢂ 1.34
7.23 ꢂ 1.57
9.67 ꢂ 1.06
2.44 ꢂ 0.53
15.46 ꢂ 1.31
20.66 ꢂ 2.26
17.36 ꢂ 1.86
18.10 ꢂ 3.71
28.70 ꢂ 4.32
29.24 ꢂ 2.63
26.01 ꢂ 2.27
38.48 ꢂ 7.21
29.47 ꢂ 4.86
74.50 ꢂ 8.70
ND
53.50 ꢂ 9.97
29.04 ꢂ 4.49
74.85 ꢂ 15.69
60.88 ꢂ 13.43
45.49 ꢂ 6.19
54.57 ꢂ 3.33
ND
ND
0.13 ꢂ 0.001
0.62 ꢂ 0.014
1.19 ꢂ 0.004
1.00 ꢂ 0.002
ND
0.97 ꢂ 0.018
0.35 ꢂ 0.010
1.70 ꢂ 0.006
0.41 ꢂ 0.005
ND
ND
ND
ND
ND
Cisplatin
a
Data represent the mean ꢂ SD of each compound from three to six independent experiments.
b
Not determined.
bis(iso-propylcarbamate) derivatives (18aej and 19aej, respectively)
compounds 18b, 19b, and 19c, were shown to have most significant
in good yield.
cytotoxicity with IC50 values of 0.07, 0.05 and 0.08
against CCRF-CEM cell growth in culture.
mM, respectively,
Our previous research on the SAR studies of 1,2-
2.2. Biological activity
bis(hydroxylmethyl)cyclopenta[a]indenes and their counterparts
2.2.1. In vitro cytotoxicity
In our anticancer agent screening program, we use human
lymphoblastic leukemia (CCRF-CEM) and its drug-resistant
sublines resistant to Taxol (CCRF-CEM/Taxol) and Vinblastine
(CCRF-CEM/VBL) cell lines for evaluating the primary anti-
proliferative activity and structureeactivity relationship (SAR)
studies of the tested compound. Those compounds warranted
in vitro cytotoxicity against these cancer cell lines are further
selected for evaluating their anti-proliferative activity against
other human solid tumor cell growth in culture. Table 1 shows
proliferative activities of the newly synthesized bis(hydrox-
ylmethyl)benzo[d]pyrrolo[2,1-b]thiazole
derivatives
(17aej),
bis(ethylcarbamates) (18aej), and bis(iso-propylcarbamates)
derivatives (19aej) against CCRF-CEM and CCRF-CEM/Taxol and
CCRF-CEM/VBL cell growth in vitro. Generally, the bis(alkylcarba-
mate) derivatives (18aej or 19aej) are more potent than the cor-
responding bis(hydroxymethyl) derivatives (17aej). In the series of
bis(hydroxymethyl) derivatives (17aej), compound having 30,40-di-
F-Ph (17d) at C1 is the most potent derivative with IC50 value
0.19 mM against CCRF/CEM cell growth in culture. While, C1-Me
(17a), 40-F-Ph (17b), 30,40-di-Cl-Ph (17e), 40-OMe-Ph (17g), cyclo-
propyl (17j) substituted derivatives are equally potent, but some-
what more cytotoxic than other phenyl substituted derivatives. In
the C1-halo substituted phenyl derivatives, 40-F-Ph (17b) is 2-fold
more potent than 40-Cl-Ph (17c), but the order of the potency for
di-halo substituted derivatives is 30,40-di-F-Ph (17d) > 30,40-di-Cl-
Ph (17e) > 30-Cl,40-F-Ph (17f). While, in the series of compounds
having an electron-donating MeO function on the C1-Ph deriva-
tives, the cytotoxicity decreases when the number of the MeO
function increases [e. g. 40-MeO-Ph (17g)
>
30,40-(MeO)2-Ph
(17h) > 30,40,50-(MeO)3-Ph (17i)].
In comparison with the cytotoxicity of bis(ethylcarbamates)
(18aej) and bis(iso-propylcarbamates) (19aej), one can see that the
latter congeners are more or equal cytotoxic than the former
derivatives. The SAR study of bis(alkylcarbamates) derivatives
reveals that the order of their potency is almost as same as that of
the bis(hydroxymethyl) derivatives. Of these derivatives,
Fig. 2. Therapeutic effects of 19b and 19c in nude mice bearing MX-1 human
mammary xenograft (i.v. inj., n ¼ 3, p < 0.01). A: average tumor size changes. B:
average body weight changes.

32
R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
properties of the substituent(s) on the phenyl ring have little
influence over their potency. The size of the substituent on C1
affects the cytotoxicity of the bis(hydroxymethyl) derivatives, but
this does not happen in the bis(alkylcarbamate) derivatives.
Our previous report demonstrated that cyclopenta[a]indenes
(e.g. 8e11) or bis(hydroxylmethyl)pyrrolo[1,2-b]isoquinolines (e.g.
12) did not have multi-drug resistance to antitumor agents such as
Taxol and Vinblastine [11,13]. To realize whether the newly
synthesized pyrrolo[1,2-b]isoquinoline derivatives are cross-
resistant to these two natural products, we evaluated their cyto-
toxicity against CCRF-CEM/Taxol and CCRF-CEM/VBL, which are
330-fold resistant to Taxol, and 680-fold resistant to Vinblastine,
respectively. As shown in Table 1, the newly synthesized benzo[d]-
pyrrolo[2,1-b]thiazole derivatives have no cross-resistance to either
Taxol or Vinblastine. This suggests that all derivatives are neither
a good substrate of membrane multi-drug resistance transporters
(i.e. p-glycoprotein) nor mutated tubulin.
On the basis of the in vitro anti-proliferative activity (IC50
values) against CCRF/CEM cell lines, the selected compounds were
further examined for their anti-proliferative activity against
a variety of human solid tumor cell growth in vitro as shown in
Table 2. The results showed that bis(alkylcarbamates) derivatives
are generally more active than bis(hydroxymethyl) derivatives
against human breast cancer MX-1 and colon cancer HCT-116. It
also revealed that the tested compounds are generally more cyto-
toxic against MX-1 and HCT-116 than other tested tumor cell lines.
Over all, we found that most of compounds are more potent than
the positive control cisplatin in inhibiting human lung cancer
H1299 and glioma U87 cell lines. It also showed that several
derivatives have significant anti-proliferative activity against
human lung H1299, prostate PC-3, oral cancer OECM-1, and glioma
U87 cell line growth in vitro. However, it is unable to conclude
a SAR among the tested compounds and tumor cell lines examined.
Fig. 3. Therapeutic effects of 18b and 18c in nude mice bearing MX-1 human
mammary xenograft (i.v. inj., n ¼ 3, p < 0.01). A: average tumor size changes. B:
average body weight changes.
2.2.2. In vivo antitumor activity
To further investigate the antitumor activity of benzo[d]pyrrolo
[2,1-b]thiazole derivatives, we selected compounds 18b, 18c, 19b
and 19c for evaluating their therapeutic efficacy in human tumor
xenograft models, since these agents exhibit a broad spectrum of
antitumor activity in vitro. The results showed that complete tumor
remission was observed in all three tested mice subcutaneously
implanted with MX-1 xenograft when treated with 19c at the
submaximal tolerated dose of 20 mg/kg, every other days, three
times (Q2D ꢀ 3) and then 25 mg/kg [Q2D ꢀ 5, intravenous injection
(iv. inj.)] on D24, D26, and D26, respectively, (Fig. 2A). When the
same tumor bearing mice were treated with 19b (20 mg/kg,
Q2D ꢀ 3 and then 25 mg/kg, Q2D ꢀ 4, iv. inj.), we found that
complete tumor remission was seen in two out of three mice on
1,2-bis(methylcarbamate) derivatives (8e11, Fig. 1) demonstrated
that the size and the electron property of the substituents at the C1
position affected the cytotoxicity of these agents [11]. More
recently, our studies on the SAR of pyrrolo[1,2-b]isoquinolines
reveal that their cytotoxicity is mainly affected by the size of the
substituents at the C1 position rather than the electron properties,
although the C1-methoxyphenyl derivatives are somewhat more
cytotoxic than the corresponding halophenyl compounds [13]. In
the present studies, however, we found that the halophenyl
derivatives are generally have better cytotoxicity than the MeO
substituted phenyl counterparts. It indicates that the electron
Fig. 4. Representative DNA cross-linking gel shift assay for bis(hydroxymethyl) derivatives (17b, 17c and 17g) and their corresponding bis(alkylcarbamate) derivatives (18b, 18c, 18g,
19b, 19c and 19g) at the concentrations of 1, 10, and 20 M. Melphalan (1 M) was used as a positive control. XL stands for cross-linking DNA, and SS refers to single strand DNA.
m
m

R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
33
D26 and 28 (Fig. 2A). In another experiment, it revealed that more
than 99% tumor suppression was achieved when the tumor bearing
mice were treated with 18b (10 mg/kg, Q2D ꢀ 4) or 18c (30 mg/kg,
Q2D ꢀ 4) via iv. inj. (Fig. 3). As shown in Fig. 2B and Fig. 3B, one can
see that the tested mice had about 20% average body weight loss
during the drug treatment, with an exception of 19c, which caused
about 10% body weight loss. However, the body weight recovered
after ceasing of treatment, indicating that the test compounds have
a little toxicity to the host.
100
80
60
40
20
2.2.3. DNA cross-linking study
Our previous reports demonstrated that bis(hydroxymethyl)-
8H-3a-azacyclopenta[a]indene-1-yl or bis(hydroxymethyl)pyrrolo
[1,2-b]isoquinolines and their bis(methylcarbamate) derivatives are
able to induce DNA interstrand cross-linking. To realize whether
the newly synthesized compounds are also capable of cross-linking
with DNA, linearized pBR322 DNA was treated with bis(hydrox-
ymethyl) derivatives (17b, 17c and 17g) and their corresponding
bis(alkylcarbamate) derivatives (18b, 18c, 18g, 19b, 19c and 19g) at
0
0
0
6.25
12.5
25
0
6.25
12.5
25
17c (µM), 24 h
17c (µM), 48 h
100
various concentrations (1, 10, and 20
mM) using alkaline agarose gel
shifting assay. Melphalan (1 M) was used as the positive control.
m
80
60
40
20
As revealed in Fig. 4, it revealed that bis(hydroxymethyl) deriva-
tives 17b, 17c and 17g are much less efficient DNA-cross-linking
agent than bis(alkylcarbamate) compounds 18b, 18c, 18g, 19b, 19c
and 19g, probably due to the latter derivatives exhibit better leaving
groups (e.g., alkylcarbamoyl function) than the former (e.g.,
hydroxymethyl group). It should be noted to show that 17b and 17c
are weak cross-linking agents, but they are as cytotoxic as 18g in
inhibiting various solid tumors in culture (see Table 2). These
observations suggests, in addition to DNA cross-linking, other
mechanism of action may be involved. Other possible mechanism
of action of bis(hydroxymethyl) derivatives (such as 17c) will be
elucidated latter.
2.25
4.5
9
0
2.25
4.5
9
18c (µM), 24 h
18c (µM), 48 h
100
2.2.4. Cell cycle inhibition
Previously, we demonstrated that 3a-aza-cyclopenta[a]indenes
are able to induce cell cycle perturbation in H1299 cells [11]. We
found that the S phase cell accumulation was observed when the
tested cells were treated with the bis(methylcarbamates) deriva-
tives at 24 h and followed by the increase of the G2/M phase at 48 h
and 72 h. Additionally, increased sub-G1 population were noticed
at 72 h. To investigate whether the newly synthesized benzo[d]
pyrrolo[2,1-b]thiazole derivatives are able to interfere with cell
cycle progression, H1299 cells were treated with compound 17c,
18c, and 19c at 0.5-, 1- and 2-fold of IC50 dose for 24 h and 48 h
respectively. Then the cell cycle distributions were analyzed by
flow cytometry. From the cell cycle data (Fig. 5), we observed that
80
60
40
20
4.5
9
18
0
4.5
9
18
17c at the highest dose used (25 mM) reduced the G1 population at
19c (µM), 24 h
19c (µM), 48 h
24 h, but resulted in increased accumulation of the G1 phase at
48 h. These results revealed that high dose of 17c slightly delay the
cell cycle progression at G2/M first and then resulted in G1 arrest.
While 18c treated cells notably accumulate in sub-G1 and G2/M
phase but reduced G1 population was observed after 24 h and 48 h
by a dose dependent manner. Similarly, it also revealed that, the cell
progression was arrested at G2/M phase after treating with the 19c
Fig. 5. Effect of compounds on cell cycle progression in human non-small cell lung
adenocarcinoma H1299 cells. The cells were cultured without compound (control),
with 17c (6.25
mM, 12.5 mM, 25 mM), 18c (2.25 mM, 4.5 mM, 9 mM), and 19c (4.5 mM,
9.0 M, 18 M) for 24 h and 48 h. Then cells were subjected for cell cycle analysis with
a flow cytometer.
m
m
compared to 18c and 19c suggests that 17c may have different
mode of action from compounds 18c and 19c.
after 24 h at doses of 4.5 and 9 mM. After 48 h, the G2/M population
was decreased whereas the sub-G1 population increased, indi-
cating that the progress of disturbed cell cycle resulted in cell death.
However, 19c at 18
m
M
apparently inhibited the cell cycle
3. Conclusions
progression after 24 h treatment, while the cell cycle progression
was resumed to result in increased G2/M population as well as sub-
G1 population. These results implicated that cell cycle disturbance
by 18c and 19c play crucial role in triggering cell death. Consistent
to our results shown in Fig. 4, the weak induction of DNA inter-
strand cross-linking as well as cell cycle progression of 17c as
In the present studies we have synthesized
substituted 2,3-bis(hydroxymethyl)-4H-benzo[d]pyrrolo[2,1-b]
thiazoles (17aej) and their bis(alkylcarbamate) derivatives (18aej
and 19aej) for antitumor studies. A variety of substituent(s) were
introduced to the C1 position of the pyrrole ring for studying their
a series of

34
R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
SAR. The preliminary antitumor studies revealed that these agents
exhibited significant antitumor activity in inhibiting various human
tumor cell growth in vitro with no or a little cross-resistance to
either Taxol or Vinblastine. Among the newly synthesized deriva-
tives, compounds 18b, 19b, and 19c were found to have potent
therapeutic efficacy against MX-1 in xenograft model, which was
used as a primary drug screening model for selecting lead
compounds for further evaluation of anticancer activity against
other solid tumor xenografts in our Drug Development Program.
Based on the results of preliminary therapeutic efficacy studies,
compound 19c will be selected as a lead compound for studying its
anticancer activity alone or in combination with DNA repair
inhibitors in inhibiting human solid tumors, which are resistant to
chemotherapeutic agents, in xenograft model and the results will
be reported in a due course.
DMSO):
d
¼ 54.5, 116.0, 116.2, 117.5, 118.7, 123.9, 125.9, 128.1, 129.7,
130.8,130.9,136.3,163.1,165.0,166.9 ppm;Anal. calcd for C15H9FN2OS:
C 63.37, H 3.19, N 9.85, S 11.28, found: C 63.02, H 3.27, N 9.81, S 11.31.
4.2.3. 3-(4-Chlorobenzoyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15c)
Compound 15c was prepared from 14 (5.4 g, 40.0 mmol), 4-
chlorobenzoyl chloride (7.2 g, 41.0 mmol), and trimethylsilylcya-
nide (4.1 g, 42.0 mmol); Yield: 10.1 g (84%); mp: 115e117 ꢁC (lit.:
115e118 ꢁC [18]); 1H NMR ([D6]DMSO):
d
¼ 6.75 (br s, 1H, C2-H),
6.85 (s, 1H, ArH), 7.05e7.09 (m, 1H, ArH), 7.13e7.17 (m, 1H, ArH),
7.52e7.54 (m, 1H, ArH), 7.58e7.61 ppm (m, 4H, 4 ꢀ ArH); 13C NMR
([D6]DMSO):
d
¼ 54.4, 117.5, 118.8, 124.0, 125.9, 126.0, 128.2, 129.1,
130.0, 132.0, 136.2, 136.8, 166.9 ppm.
4.2.4. 3-(3,4-Difluorobenzoyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15d)
4. Experimental protocols
Compound 15d was prepared from 14 (6.8 g, 50.0 mmol), 3,4-
difluorobenzoyl chloride (9.0 g, 51.0 mmol), and trimethylsilylcya-
nide (5.1 g, 52.0 mmol); Yield: 11.9 g (79%); mp: 149e150 ꢁC; 1H
4.1. Material and methods
All commercial chemicals and solvents were reagent grade and
were used without further purification unless otherwise specified.
Melting points were determined in open capillaries on a Fargo
melting point apparatus and are uncorrected. Thin-layer chroma-
tography was performed on silica gel G60 F254 (Merck) with short-
wavelength UV light for visualization. Elemental analysis was done
on a Heraeus CHNeO Rapid instrument. High-performance liquid
chromatography was performed on Elite instrument: column,
Mightysil RP-18 (250 ꢀ 4.6 mm). Compounds were detected by UV
at 260 nm. The mobile phase was MeCN/THF (50:50 v/v) with flow
rate of 1 mL/min. The purity of all tested compounds was ꢄ95%
based on analytical HPLC. 1H NMR spectra and 13C NMR spectra
were recorded on Bruker AVANCE 600 DRX and 400 MHz, Bruker
Top-Spin spectrometers in the solvents indicated. The Proton
NMR ([D6]DMSO):
d
¼ 6.77 (br s, 1H, C2eH), 6.84 (s, 1H, ArH),
7.06e7.10 (m, 1H, ArH), 7.13e7.17 (m, 1H, ArH), 7.45 (m, 1H, ArH),
7.52e7.54 (m, 1H, ArH), 7.58e7.65 (m, 1H, ArH), 7.73e7.76 ppm (m,
1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 54.3, 117.4, 117.9, 118.1, 118.3,
118.4, 118.8, 123.9, 125.6, 125.9, 126.0, 128.2, 130.7, 136.0, 148.3,
148.4, 150.3, 150.4, 150.5, 150.6, 152.5, 152.6, 165.7 ppm; Anal. calcd
for C15H8F2N2OS: C 59.60, H 2.67, N 9.27, S 10.61, found: C 59.24, H
2.90, N 9.17, S 10.53.
4.2.5. 3-(3,4-Dichlorobenzoyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15e)
Compound 15e was prepared from 14 (6.8 g, 50.0 mmol), 3,4-
dichlorobenzoyl chloride (10.4 g, 51.0 mmol), and trimethylsi-
lylcyanide (5.1 g, 52.0 mmol); Yield: 13.5 g (81%); mp: 166e167 ꢁC;
chemical shifts were reported in parts per million (
d
ppm) relative
1H NMR ([D6]DMSO):
d
¼ 6.84 (br s, 2H, C2eH and ArH), 7.08e7.11
to (CH3)4Si and coupling constants (J) in Hertz (Hz) and s, d, t, m, br
s, refer to singlet, doublet, triplet, multiplet, broad respectively.
(m, 1H, ArH), 7.14e7.18 (m, 1H, ArH), 7.52e7.54 (m, 2H, 2 ꢀ ArH),
7.80e7.82 (m, 1H, ArH), 7.88e7.90 ppm (m, 1H, ArH); 13C NMR ([D6]
DMSO):
d
¼ 54.3, 117.4., 118.9, 123.9, 125.9, 126.2, 128.1, 128.2, 130.2,
4.2. Chemistry: general methods
131.2, 131.8, 133.7, 134.7, 135.9, 165.5 ppm; Anal. calcd for
C15H8Cl2N2OS: C 53.75, H 2.41, N 8.36, S 9.57, found: C 53.55, H 2.50,
N 8.31, S 9.18.
4.2.1. 3-Acetyl-2,3-dihydrobenzo[d]thiazole-2-carbonitrile (15a)
Acetyl chloride (4.0 g, 51.0 mmol) was added drop wise to
a
stirring solution of benzothiazole (6.8 g, 50.0 mmol) in
4.2.6. 3-(3-Chloro-4-fluorobenzoyl)-2,3-dihydrobenzo[d]thiazole-
2-carbonitrile (15f)
Compound 15f was prepared from 14 (6.8 g, 50.0 mmol), 3-
chloro-4-fluorobenzoyl chloride (9.9 g, 51.0 mmol), and trime-
thylsilylcyanide (5.1 g, 52.0 mmol); Yield: 13.2 g (83%); mp:
dichloromethane (40 mL) under argon atmosphere. A catalytic
amount of AlCl3 and trimethylsilylcyanide (5.1 g, 52.0 mmol) were
then added into the reaction mixture. After being stirred for 17 h at
room temperature, the reaction mixture was evaporated to dryness
in vacuo and the residue was crystallized from ether to give 15a.
Yield: 8.2 g (80%); mp: 92e93 ꢁC; 1H NMR (400 MHz, [D6]DMSO):
187e188 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 6.81(br s, 1H, C2eH), 6.84 (s,
1H, ArH), 7.06e7.10 (m, 1H, ArH), 7.13e7.17 (m, 1H, ArH), 7.52e7.56
(m, 1H, ArH), 7.58e7.60 (m, 2H, 2 ꢀ ArH), 7.86e7.88 ppm (m, 1H,
d
¼ 2.39 (s, 3H, Me), 7.05 (s, 1H, ArH), 7.16e7.26 (m, 2H, 2 ꢀ ArH),
7.49e7.51 (m, 1H, ArH), 7.65 ppm (br s, 1H, C2eH); 13C NMR ([D6]
DMSO):
ArH); 13C NMR ([D6]DMSO):
120.9, 124.4, 126.4, 126.5, 128.7, 129.6, 129.7, 131.3, 131.5, 136.5,
158.6, 160.6, 166.1 ppm; Anal. calcd for C15H8ClFN2OS: C 56.52, H
2.53, N 8.79, S 10.06, found: C 56.41, H 2.92, N 8.78, S 9.69.
d
¼ 54.8, 117.8, 117.9, 118.1, 119.3, 120.8,
d
¼ 22.6, 52.6, 117.4, 118.4, 123.5, 125.8, 126.3, 131.2, 134.5,
168.8 ppm; Anal. calcd for C10H8N2OS: C 58.80, H 3.95, N 13.72, S
15.70, found: C 58.85, H 3.95, N 13.78, S 15.76.
By following the same synthetic procedure as that for 15a, the
following compounds were synthesized:
4.2.7. 3-(4-Methoxybenzoyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15g)
4.2.2. 3-(4-Fluorobenzoyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15b)
Compound 15b was prepared from 14 (5.4 g, 40.0 mmol), 4-
fluorobenzoyl chloride (6.5 g, 41.0 mmol), and trimethylsilylcyanide
(4.1 g, 42.0 mmol). Yield: 9.3 g (82%); mp: 150e151 ꢁC; 1H NMR ([D6]
Compound 15g was prepared from 14 (5.4 g, 40.0 mmol), 4-
methoxybenzoyl chloride (7.0 g, 41.0 mmol), and trimethylsi-
lylcyanide (4.1 g, 42.0 mmol); Yield: 8.2 g (70%); mp: 122e123 ꢁC
(lit.: 122e123.5 ꢁC [18]); 1H NMR ([D6]DMSO):
d
¼ 3.82 (s, 3H,
MeO), 6.72 (br s, 1H, C2eH), 6.84 (s, 1H, ArH), 7.01e7.14 (m, 4H,
DMSO):
d
¼ 6.71 (br s, 1H, C2eH), 6.85 (s, 1H, ArH), 7.04e7.07 (m, 1H,
4 ꢀ ArH), 7.52e7.55 ppm (m, 3H, 3 ꢀ ArH); 13C NMR ([D6]DMSO):
ArH), 7.12e7.16 (m, 1H, ArH), 7.38e7.40 (m, 2H, 2 ꢀ ArH), 7.52e7.54
d
¼ 54.8, 55.5, 114.2, 117.7, 118.6, 123.9, 124.9, 125.6, 125.8, 128.0,
(m, 1H, ArH), 7.64e7.67 ppm (m, 2H, 2 ꢀ ArH); 13C NMR ([D6]
130.3, 136.7, 162.3, 167.6 ppm .

R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
35
4.2.8. 3-(3,4-Dimethoxybenzoyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15h)
Compound 15h was prepared from 14 (6.8 g, 50.0 mmol), 3,4-
dimethoxybenzoyl chloride (10.3 g, 51.0 mmol), and trimethylsi-
lylcyanide (5.1 g, 52.0 mmol); Yield: 10.4 g (64%); mp: 219e220 ꢁC;
8.03e8.05 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 51.6, 51.9,
103.4, 113.8,115.7,115.9,120.1, 124.9,125.4, 126.3, 128.3, 131.4,133.0,
133.1, 133.7, 136.9, 161.9, 162.4, 163.8, 164.0 ppm; Anal. calcd for
C20H14FNO4S: C 62.65, H 3.68, N 3.65, S 8.36, found: C 62.68, H 3.46,
N 3.62, S 8.76.
1H NMR ([D6]DMSO):
d
¼ 3.76 (s, 3H, MeO), 3.84 (s, 3H, MeO), 6.82
(br s, 2H, C2eH and ArH), 7.07e7.11 (m, 2H, 2 ꢀ ArH), 7.12e7.17 (m,
4.2.13. Dimethyl 1-(4-chlorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-dicarboxylate (16c)
Compound 16c was prepared from 15c (9.1 g, 30.0 mmol), HBF4
(9 mL), and DMAD (12.7 g, 90.0 mmol); Yield: 2.5 g (21%); mp:
3H, 3 ꢀ ArH), 7.51e7.53 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 54.9, 55.5, 55.7,111.3,117.8,118.9,121.6,123.9,124.9,125.7,125.8,
128.0, 136.7, 148.6, 151.9, 167.7 ppm; Anal. calcd for C17H14N2O3S: C
62.56, H 4.32, N 8.58, S 9.82, found: C 62.31, H 4.25, N 8.39, S 9.68.
172e173 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 3.64 (s, 3H, COOMe), 3.83 (s,
3H, COOMe), 6.85e6.87 (m, 1H, ArH), 7.30e7.42 (m, 2H, ArH),
4.2.9. 3-(3,4,5-Trimethoxybenzoyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15i)
Compound 15i was prepared from 14 (6.8 g, 50.0 mmol), 3,4,5-
trimethoxybenzoyl chloride (11.5 g, 51.0 mmol), and trimethylsi-
lylcyanide (5.1 g, 52.0 mmol); Yield: 15.8 g (88%); mp: 189e190 ꢁC;
7.60e7.66 (m, 4H, 4 ꢀ ArH), 8.04e8.06 ppm (m, 1H, ArH); 13C NMR
([D6]DMSO):
d
¼ 51.7, 52.0, 103.5, 133.9, 120.3, 125.0, 125.4, 126.3,
127.9, 128.0, 128.9, 131.4, 132.4, 133.7, 134.6, 137.1, 162.4, 164.0 ppm;
Anal. calcd for C20H14ClNO4S$0.5H2O: C 58.75, H 3.70, N 3.43, S 7.84,
found: C 58.76, H 3.45, N 3.59, S 7.73.
1H NMR ([D6]DMSO):
d
¼
3.75 (s, 3H, MeO), 3.78 (s, 6H,
2 ꢀ MeO),6.81 (s, 1H, ArH), 6.91 (s, 2H, 2 ꢀ ArH), 7.02 (br s, 1H,
4.2.14. Dimethyl 1-(3,4-difluorophenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-dicarboxylate (16d)
C2eH), 7.13e7.20 (m, 2H, 2 ꢀ ArH), 7.51e7.54 ppm (m, 1H, ArH); 13C
NMR ([D6]DMSO):
d
¼ 54.8, 56.1, 60.2, 105.4, 117.9, 119.4, 123.7,
Compound 16d was prepared from 15d (10.2 g, 33.0 mmol),
HBF4 (10 mL), and DMAD (14.1 g, 99.0 mmol); Yield: 3.8 g (28%);
125.9, 126.1, 128.2, 128.5, 136.4, 140.3, 153.1, 167.7 ppm; Anal. calcd
for C18H16N2O4S: C 60.66, H 4.53, N 7.86, S 9.00, found: C 60.36, H
4.45, N 7.76, S 9.11.
mp: 183e184 ꢁC; 1H NMR ([D6]DMSO):
3.83 (s, 3H, COOMe), 6.86e6.88 (m, 1H, ArH), 7.31e7.35 (m, 1H,
ArH), 7.40e7.44 (m, 2H, 2 ꢀ ArH), 7.61e7.68 (m, 1H, ArH), 7.73e7.78
(m, 1H, ArH), 8.04e8.06 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 3.65 (s, 3H, COOMe),
4.2.10. 3-(Cyclopropanecarbonyl)-2,3-dihydrobenzo[d]thiazole-2-
carbonitrile (15j)
Compound 15j was prepared from 14 (6.8 g, 50.0 mmol),
cyclopropylcarbonyl chloride (5.2 g, 51.0 mmol), and trimethylsi-
lylcyanide (5.1 g, 52.0 mmol); Yield: 6.3 g (55%); mp: 113e114 ꢁC;
d
¼ 51.8, 52.4, 113.7, 118.6, 122.3, 122.5, 125.6, 125.9, 126.7, 127.0,
134.6, 135.6, 148.2, 149.6, 152.2, 156.4, 163.7, 165.0 ppm; Anal. calcd
for C20H13F2NO4S: C 59.85, H 3.26, N 3.49, S 7.99, found: C 59.56, H
3.53, N 3.32, S 8.12.
1H NMR ([D6]DMSO):
d
¼ 0.97 (m, 2H, CH2), 1.15 (m, 2H, CH2),
2.13 (m,1H, CH), 7.17e7.27 (m, 3H, 3 ꢀ ArH), 7.52e7.54 (m,1H, ArH),
7.74 ppm (br s, 1H, C2eH); 13C NMR ([D6]DMSO):
4.2.15. Dimethyl 1-(3,4-dichlorophenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-dicarboxylate (16e)
d
¼ 8.8, 9.8, 13.4,
52.8, 117.5, 118.4, 123.7, 125.7, 126.4, 128.2, 136.3, 171.4 ppm; Anal.
calcd for C12H10N2OS: C 62.59, H 4.38, N 12.16, S 13.92, found: C
62.56, H 4.29, N 12.26, S 13.93.
Compound 16e was prepared from 15e (12.1 g, 35.9 mmol), HBF4
(11 mL), and DMAD (15.2 g, 107.0 mmol); Yield: 5 g (32%); mp:
198e199 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 3.64 (s, 3H, COOMe), 3.81 (s,
3H, COOMe), 6.88e6.90 (m, 1H, ArH), 7.31e7.42 (m, 2H, 2 ꢀ ArH),
4.2.11. Dimethyl 1-methylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-
dicarboxylate (16a)
7.57e7.60 (m, 1H, ArH), 7.81e7.83 (m, 1H, ArH), 7.91 (s, 1H, ArH),
8.03e8.05 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 51.7, 52.0,
To a solution of 15a (5.0 g, 24.5 mmol) in dichloromethane
(50 mL) was added drop wise 7 mL of tetrafluoroboric acid (HBF4).
The solution was stirred for 3 h at room temperature. The brown
precipitates appeared were collected by filtration and the filter cake
was washed with ether to give hydrofluoroborate salt. The solid salt
was added to a solution of dimethyl acetylenedicarboxylate (10.4 g,
73.0 mmol) in DMF (30 mL) and then warmed at 35 ꢁC for 13 h. The
reaction mixture was concentrated in vacuo and the residue was
crystallized from methanol to give 16a. Yield: 1.7 g (23%); mp:
103.7, 114.0, 120.4, 124.8, 125.4, 126.4, 126.8, 129.6, 130.9, 131.4,
132.5, 132.7, 133.6, 137.5, 162.3, 163.8 ppm; Anal. calcd for
C20H13Cl2NO4S: C 55.31, H 3.02, N 3.23, S 7.38, found: C 55.36, H
3.02, N 3.13, S 7.35.
4.2.16. Dimethyl 1-(3-chloro-4-fluorophenyl)benzo[d]pyrrolo[2,1-
b]thiazole-2,3-dicarboxylate (16f)
Compound 16f was prepared from 15f (10.0 g, 31.4 mmol), HBF4
(9 mL), and DMAD (13.3 g, 94.0 mmol); Yield: 2.2 g (17%); mp:
143e144 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 2.85 (s, 3H, Me), 3.83 (s, 3H,
196e197 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 3.65 (s, 3H, COOMe), 3.83 (s,
COOMe), 3.85 (s, 3H, COOMe), 7.50 (t, J ¼ 7.6 Hz, 1H, ArH), 7.58 (t,
J ¼ 7.6 Hz, 1H, ArH), 8.07 (d, J ¼ 8.0 Hz, 1H, ArH), 8.12 ppm (d,
3H, COOMe), 6.83e6.85 (m, 1H, ArH), 7.32e7.43 (m, 2H, 2 ꢀ ArH),
7.62e7.64 (m, 2H, 2 ꢀ ArH), 7.89e7.90 (m, 1H, ArH), 8.05e8.07 ppm
J ¼ 8.0 Hz,1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 12.2, 51.4, 51.7, 102.9,
(m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 51.6, 51.9, 103.6, 114.0, 117.2,
114.7, 117.6, 124.5, 125.2, 126.4, 128.9, 131.2, 134.2, 136.1, 162.4,
164.6 ppm; Anal. calcd for C15H13NO4S: C 59.39, H 4.32, N 4.62, S
10.57, found: C 59.02, H 4.23, N 4.87, S 10.42.
117.4, 119.8, 119.9, 120.3, 124.9, 125.5, 126.4, 126.7, 128.8, 127.1, 131.3,
131.7, 131.8, 133.0, 133.7, 137.2, 157.2, 158.8, 162.4, 163.8 ppm; Anal.
calcd for C20H13ClFNO4S: C 57.49, H 3.14, N 3.35, S 7.67, found: C
57.79, H 3.16, N 3.55, S 7.41.
By following the same synthetic procedure as that for 16a, the
following compounds were synthesized:
4.2.17. Dimethyl 1-(4-methoxyphenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-dicarboxylate (16g)
4.2.12. Dimethyl 1-(4-fluorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-dicarboxylate (16b)
Compound 16g was prepared from 15g (5.3 g, 17.9 mmol), HBF4
(5 mL), and DMAD (7.6 g, 54.0 mmol); Yield: 1.7 g (24%); mp:
Compound 16b was prepared from 15b (8.1 g, 28.0 mmol), HBF4
(8 mL), and DMAD (12.2 g, 84.0 mmol); Yield: 2.5 g (23%); mp:
167e168 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 3.63 (s, 3H, COOMe), 3.82 (s,
186e187 ꢁC; 1H NMR ([D6]DMSO):
3H, COOMe), 6.78e6.80 (m, 1H, ArH), 7.28e7.30 (m, 1H, ArH),
7.30e7.44 (m, 3H, 3 ꢀ ArH), 7.62e7.66 (m, 2H, 2 ꢀ ArH),
d
¼ 3.63 (s, 3H, COOMe), 3.83 (s,
3H, COOMe), 3.86 (s, 3H, Me), 6.84e6.86 (m, 1H, ArH), 7.12 (d,
J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 7.28e7.32 (m, 1H, ArH), 7.37e7.40 (m, 1H,
ArH), 7.48 (d, J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 8.03e8.05 ppm (m, 1H, ArH);

36
R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
13C NMR ([D6]DMSO):
121.3, 125.3, 125.7, 126.6, 129.6, 131.8, 132.5, 134.3, 136.8, 160.6,
163.0, 164.7 ppm; Anal. calcd for C21H17NO5S: C 63.79, H 4.33, N
3.54, S 8.11, found: C 63.81, H 4.33, N 3.55, S 8.10.
d
¼ 52.0, 52.3, 55.7, 103.7, 114.3, 144.6, 120.4,
By following the same synthetic procedure as that for 17a, the
following compounds were synthesized:
4.2.22. [1-(4-Fluorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
dimethanol (17b)
4.2.18. Dimethyl 1-(3,4-dimethoxyphenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-dicarboxylate (16h)
Compound 17b was prepared from 16b (2.4 g, 6.6 mmol) and
LiAlH4 (0.6 g, 16.0 mmol); Yield: 1.5 g (76%); mp: 155e156 ꢁC; 1H
Compound 16h was prepared from 15h (5.8 g, 17.7 mmol), HBF4
(5 mL), and DMAD (7.5 g, 53.0 mmol); Yield: 2.3 g (30%); mp:
NMR ([D6]DMSO):
d
¼ 4.22 (d, J ¼ 5.1 Hz, 2H, CH2), 4.66 (d, J ¼ 5.1 Hz,
2H, CH2), 4.73 (t, J ¼ 5.1 Hz,1H, exchangeable, OH), 5.13 (t, J ¼ 5.1 Hz,
1H, exchangeable, OH), 6.83e6.85 (m, 1H, ArH), 7.15e7.19 (m, 2H,
2 ꢀ ArH), 7.37e7.41 (m, 2H, 2 ꢀ ArH), 7.53e7.57 (m, 2H, 2 ꢀ ArH),
226e227 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 3.65 (s, 3H, COOMe), 3.73 (s,
3H, Me), 3.82 (s, 3H, COOMe), 3.86 (s, 3H, Me), 6.93e6.95 (m, 1H,
ArH), 7.07e7.16 (m, 3H, 3 ꢀ ArH), 7.30e7.34 (m, 1H, ArH), 7.37e7.41
(m, 1H, ArH), 8.03e8.05 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
7.79e7.81 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 54.7, 56.1,
112.7, 114.7, 115.9, 123.9, 123.5, 124.4,124.7, 125.6, 126.8, 127.8, 131.8,
133.2, 133.3, 134.8 ppm; Anal. calcd for C18H14FNO2S: C 66.04, H
4.31, N 4.28, S 9.79, found: C 66.25, H 4.22, N 4.14, S 9.76.
d
¼ 51.6, 51.9, 55.5, 55.6, 103.2, 111.5, 113.9, 114.1, 119.9, 120.8, 123.2,
124.8, 125.3, 126.2, 129.2, 131.3, 133.9, 136.3, 148.4, 149.7, 162.5,
164.3 ppm; Anal. calcd for C22H19NO6S: C 62.11, H 4.50, N 3.29, S
7.54, found: C 62.12, H 4.44, N 3.38, S 7.48.
4.2.23. [1-(4-Chlorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]dimethanol (17c)
4.2.19. Dimethyl 1-(3,4,5-trimethoxyphenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-dicarboxylate (16i)
Compound 17c was prepared from 16c (2.1 g, 5.0 mmol) and
LiAlH4 (0.45 g, 12.5 mmol); Yield: 1.5 g (88%); mp: 151e152 ꢁC; 1H
Compound 16i was prepared from 15i (10.2 g, 28.0 mmol), HBF4
(8 mL), and DMAD (11.9 g, 84.0 mmol); Yield: 2.8 g (22%); mp:
NMR ([D6]DMSO):
d
¼ 4.22 (d, J ¼ 4.4 Hz, 2H, CH2), 4.66 (d, J ¼ 4.4 Hz,
2H, CH2), 4.76 (t, J ¼ 5.2 Hz,1H, exchangeable, OH), 5.14 (t, J ¼ 5.2 Hz,
1H, exchangeable, OH), 6.91e6.93 (m, 1H, ArH), 7.16e7.22 (m, 2H,
2 ꢀ ArH), 7.53 (d, J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 7.61 (d, J ¼ 8.4 Hz, 2H,
2 ꢀ ArH), 7.79e7.82 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
244e245 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 3.69 (s, 3H, COOMe), 3.76 (s,
6H, 2 ꢀ Me), 3.79 (s, 3H, Me), 3.83 (s, 3H, COOMe), 6.89 (s, 2H,
2 ꢀ ArH), 7.03e7.05 (m, 1H, ArH), 7.35e7.43 (m, 2H, 2 ꢀ ArH),
8.03e8.05 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 51.6, 51.9,
d
¼ 54.6, 56.1,112.9,114.9,124.0,124.2,124.8,125.5,125.7,127.0,129.1,
53.8, 56.1, 60.2, 103.3, 108.5, 114.3, 119.9, 123.9, 124.7, 125.3, 126.2,
129.0, 131.3, 133.8, 136.5, 138.4, 152.8, 162.4, 164.3 ppm; Anal. calcd
for C23H21NO7S: C 60.65, H 4.65, N 3.08, S 7.04; found: C 60.38, H
4.67, N 3.15, S 7.13.
130.3, 131.8, 132.7, 133.5, 134.8 ppm; Anal. calcd for C18H14ClNO2S: C
62.88, H 4.10, N 4.07, S 9.33, found: C 62.73, H 4.09, N 4.06, S 9.19.
4.2.24. [1-(3,4-Difluorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]dimethanol (17d)
4.2.20. Dimethyl 1-cyclopropylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-
dicarboxylate (16j)
Compound 17d was prepared from 16d (1.6 g, 4.0 mmol) and
LiAlH4 (0.37 g, 10.0 mmol); Yield: 1.1 g (83%); mp: 154e155 ꢁC; 1H
Compound 16j was prepared from 15j (5.6 g, 24.3 mmol), HBF4
(7 mL), and DMAD (10.3 g, 73.0 mmol); Yield: 1.8 g (22%); mp:
NMR ([D6]DMSO):
d
¼ 4.22 (d, 2H, J ¼ 4.6 Hz, CH2), 4.66 (d,
J ¼ 4.6 Hz, 2H, CH2), 4.76 (t, J ¼ 5.0 Hz, 1H, exchangeable, OH), 5.14
(t, J ¼ 5.0 Hz, 1H, exchangeable, OH), 6.91e6.93 (m, 1H, ArH),
7.18e7.21 (m, 2H, 2 ꢀ ArH), 7.37 (m, 1H, ArH), 7.58e7.65 (m, 2H,
2 ꢀ ArH), 7.81e7.83 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
157e158 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 0.71 (m, 2H, CH2), 1.13 (m,
2H, CH2), 2.26 (m, 1H, CH), 3.77 (s, 3H, COOMe), 3.82 (s, 3H,
COOMe), 7.45e7.49 (m, 1H, ArH), 7.55e7.59 (m, 1H, ArH), 8.03e8.05
(m, 1H, ArH), 8.36e8.38 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 54.6, 56.0, 112.9, 114.8, 118.1, 120.0, 123.3, 124.0, 124.7, 125.8,
d
¼ 7.1, 51.4, 52.1, 103.0, 115.2, 119.8, 124.4, 125.0, 126.4, 130.2, 131.2,
127.2, 128.1, 128.2, 128.8, 131.8, 134.7, 148.7, 150.7 ppm; Anal. calcd
for C18H13F2NO2S: C 62.60, H 3.79, N 4.06, S 9.28, found: C 62.46, H
3.84, N 3.90, S 9.16.
134.2, 135.2, 162.3, 165.2 ppm; Anal. calcd for C17H15NO4S: C 61.99,
H 4.59, N 4.25, S 9.74, found: C 61.78, H 4.48, N 4.29, S 9.66.
4.2.21. [1-Methylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
dimethanol (17a)
4.2.25. [1-(3,4-Dichlorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]dimethanol (17e)
A solution of 16a (2.0 g, 6.6 mmol) in anhydrous dichloro-
methane (30 mL) was added drop wise to a stirred mixture of
lithium aluminum hydride (0.6 g, 16.0 mmol) in anhydrous ether
(20 mL) at ꢃ5 ꢁC to 0 ꢁC. The reaction mixture was allowed to stir at
this temperature for 20 min. The excess hydride was decomposed
by adding water (1 mL) followed by NH4OH (1 mL) and water
(1 mL) at ꢃ5 ꢁC to 0 ꢁC. The mixture was filtered through a pad of
Celite, washed with several times with dichloromethane. The
combined filtrate and washings were washed successively with
brine solution and water. The organic layer was dried (Na2SO4) and
concentrated to dryness in vacuo. The residue was crystallizes from
ether to give 17a. Yield: 1.3 g (82%); mp: 138e139 ꢁC; 1H NMR ([D6]
Compound 17e was prepared from 16e (4.0 g, 9.2 mmol) and
LiAlH4 (0.85 g, 23.0 mmol); Yield: 2.8 g (84%); mp: 159e160 ꢁC; 1H
NMR ([D6]DMSO):
d
¼ 4.23 (s, 2H, CH2), 4.66 (s, 2H, CH2), 4.83 (br s,
1H, exchangeable, OH), 5.16 (br s, 1H, exchangeable, OH), 6.95e6.99
(m, 1H, ArH), 7.19e7.24 (m, 2H, 2 ꢀ ArH), 7.49e7.52 (m, 1H, ArH),
7.78e7.84 ppm (m, 3H, 3 ꢀ ArH); 13C NMR ([D6]DMSO):
d
¼ 54.5,
56.0, 113.0, 115.1, 122.9, 124.1, 124.8, 125.8, 126.2, 127.6, 131.0, 131.1,
131.3, 131.7, 131.8, 132.0, 132.3, 134.7 ppm; Anal. calcd for
C18H13Cl2NO2S: C 57.15, H 3.46, N 3.70, S 8.48, found: C 57.20, H
3.48, N 3.66, S 8.33.
4.2.26. [1-(3-chloro-4-fluorophenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-diyl]dimethanol (17f)
DMSO):
d
¼ 2.63 (s, 3H, Me), 4.39 (d, J ¼ 5.2 2H, Hz, CH2), 4.56 (d,
J ¼ 5.2 Hz, 2H, CH2), 4.60 (t, J ¼ 5.6 Hz, 1H, exchangeable, OH), 4.97
(t, J ¼ 5.6 Hz, 1H, exchangeable, OH), 7.21e7.25 (m, 1H, ArH),
7.34e7.38 (m, 1H, ArH), 7.76e7.78 (m, 1H, ArH), 7.83e7.85 ppm (m,
Compound 17f was prepared from 16f (2.0 g, 4.7 mmol) and
LiAlH4 (0.45 g, 12.0 mmol); Yield: 1.3 g (75%); mp: 178e179 ꢁC; 1H
NMR ([D6]DMSO):
d
¼ 4.22 (d, J ¼ 4.4 Hz, 2H, CH2), 4.65 (d,
1H, ArH); 13C NMR (125 MHz [D6]DMSO):
d
¼ 11.9, 54.4, 56.1, 113.0,
J ¼ 4.4 Hz, 2H, CH2), 4.78 (t, J ¼ 4.8 Hz, 1H, exchangeable, OH), 5.14
(t, J ¼ 4.8 Hz, 1H, exchangeable, OH), 6.89e6.91 (m, 1H, ArH),
7.17e7.23 (m, 2H, 2 ꢀ ArH), 7.51e7.53 (m, 1H, ArH), 7.57e7.62 (m,
1H, ArH), 7.75e7.77 (m, 1H, ArH), 7.80e7.82 ppm (m, 1H, ArH); 13C
113.6, 121.4, 122.3, 123.5, 124.4, 124.8, 125.9, 131.6, 135.6 ppm; Anal.
calcd for C13H13NO2S: C 63.13, H 5.30, N 5.66, S 12.97, found: C
62.94, H 5.24, N 5.83, S 12.90.

R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
37
NMR ([D6]DMSO):
d
¼ 54.6, 56.0, 112.8, 114.8, 117.6, 120.0, 123.1,
the reaction mixture was evaporated to dryness in vacuo. The
desired product was obtained by crystallization.
124.0, 124.8, 125.6, 125.8, 127.3, 129.2, 131.8, 132.8, 134.7 ppm; Anal.
calcd for C18H13ClFNO2S: C 59.75, H 3.62, N 3.87, S 8.86, found: C
59.72, H 3.61, N 3.86, S 8.81.
4.2.32. [1-Methylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
bis(methylene)bis(ethylcarbamate) (18a)
4.2.27. [1-(4-Methoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]dimethanol (17g)
Compound 18a was prepared from 17a (0.24 g, 1.0 mmol), Et3N
(0.5 mL), and ethylisocyanate (0.28 g, 4.0 mmol); Yield: 0.3 g (85%);
Compound 17g was prepared from 16g (1.7 g, 4.3 mmol) and
mp: 158e159 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 0.98 (t, J ¼ 7.0 Hz, 6H,
LiAlH4 (0.4 g, 10.0 mmol); Yield: 1.3 g (91%); mp: 156e157 ꢁC; 1H
2 ꢀ Me), 2.67 (s, 3H, Me), 2.98 (q, J ¼ 7.0 Hz, 4H, CH2), 5.01 (s, 2H,
CH2), 5.03 (s, 2H, CH2), 7.02e7.05 (br s, 2H, exchangeable, NH), 7.29
(t, J ¼ 8.0 Hz,1H, ArH), 7.41 (t, J ¼ 8.0 Hz,1H, ArH), 7.84 (d, J ¼ 8.0 Hz,
1H, ArH), 7.91 ppm (d, J ¼ 8.0 Hz, 1H, ArH); 13C NMR ([D6]DMSO):
NMR ([D6]DMSO):
d
¼ 3.85 (s, 3H, MeO), 4.22 (d, J ¼ 4.8 Hz, 2H,
CH2), 4.66 (m, 3H, CH2 and exchangeable OH), 5.09 (t, J ¼ 4.8 Hz,1H,
exchangeable, OH), 6.85e6.87 (m, 1H, ArH), 7.08e7.19 (m, 4H,
4 ꢀ ArH), 7.39e7.42 (m, 2H, 2 ꢀ ArH), 7.76e7.79 ppm (m, 1H, ArH);
d
¼ 11.4, 14.9, 34.9, 35.0, 56.2, 57.0, 108.3, 113.2, 120.4, 123.1, 123.7,
13C NMR ([D6]DMSO):
d
¼ 54.8, 55.7, 56.2, 112.8, 114.4, 114.5, 123.5,
124.0, 125.0, 125.7, 130.6, 134.8, 155.9, 156.1 ppm; Anal. calcd for
C19H23N3O4S: C 58.59, H 5.95, N 10.79, S 8.23, found: C 58.26, H
5.62, N 10.46, S 8.60.
123.8, 124.3, 124.6, 125.5, 125.6, 126.4, 131.8, 132.4, 135.0,
159.7 ppm; Anal. calcd for C19H17NO3S: C 67.24, H 5.05, N 4.13, S
9.45, found: C 67.54, H 5.00, N 4.00, S 9.30.
4.2.33. [1-(4-Fluorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
bis(methylene)bis(ethylcarbamate) (18b)
Compound 18b was prepared from 17b (0.33 g, 1.0 mmol), Et3N
(0.5 mL), and ethylisocyanate (0.28 g, 4.0 mmol); Yield: 0.36 g
4.2.28. [1-(3,4-Dimethoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-diyl]dimethanol (17h)
Compound 17h was prepared from 16h (2.2 g, 5.0 mmol) and
LiAlH4 (0.5 g, 13.0 mmol); Yield: 1.7 g (91%); mp: 215e216 ꢁC 1H
(79%); mp: 172e173 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 0.98 (t, J ¼ 7.0 Hz,
NMR ([D6]DMSO):
d
¼ 3.74 (s, 3H, OMe), 3.85 (s, 3H, MeO), 4.25
6H, 2 ꢀ Me), 2.98 (q, J ¼ .0 Hz, 4H, CH2), 4.81 (s, 2H, CH2), 5.12 (s, 2H,
CH2), 6.79e6.81(m, 1H, ArH), 7.05e7.11 (br s, 2H, exchangeable,
NH), 7.19e7.26 (m, 2H, 2 ꢀ ArH), 7.38e7.43 (m, 2H, 2 ꢀ ArH),
7.56e7.59 (m, 2H, 2 ꢀ ArH), 7.86e7.88 ppm (m, 1H, ArH); 13C NMR
(s, 2H, CH2), 4.66 (br s, 3H, CH2 and exchangeable OH), 5.09 (br s,
1H, exchangeable, OH), 6.93e6.95 (m, 1H, ArH), 7.01e7.06 (m, 2H,
2 ꢀ ArH), 7.10e7.18 (m, 3H, 3 ꢀ ArH), 7.76e7.78 ppm (m, 1H,
ArH); 13C NMR ([D6]DMSO):
d
¼ 54.9, 55.9, 56.0, 56.2, 112.0, 113.0,
([D6]DMSO):
d
¼ 14.9, 15.0, 34.9, 35.0, 56.7, 57.0, 109.2, 112.7, 115.6,
114.5, 114.7, 123.5, 123.6, 123.8, 124.3, 124.6, 125.5, 125.7, 126.4,
131.8, 135.0, 148.9, 149.4 ppm; Anal. calcd for C20H19NO4S$0.5H2O:
C 63.47, H 5.33, N 3.70, S 8.47, found: C 63.37, H 5.13, N 3.67, S
8.28.
115.8, 122.4, 124.0, 124.4, 125.6, 126.4, 127.3, 130.8, 132.8, 132.9,
134.1, 155.7, 156.1, 161.4, 163.3 ppm; Anal. calcd for C24H24FN3O4S: C
61.39, H 5.15, N 8.95, S 6.83, found: C 61.36, H 5.26, N 8.94,S 7.14.
4.2.34. [1-(4-Chlorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(ethylcarbamate) (18c)
Compound 18c was prepared from 17c (0.25 g, 0.7 mmol), Et3N
(0.4 mL), and ethylisocyanate (0.23 g, 3.0 mmol); Yield: 0.28 g
4.2.29. [1-(3,4,5-Trimethoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-diyl]dimethanol (17i)
Compound 17i was prepared from 16i (2.4 g, 5.2 mmol) and
LiAlH4 (0.5 g, 13.0 mmol); Yield: 1.5 g (74%); mp: 160e161 ꢁC; 1H
(78%); mp: 190e191 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.00 (t, J ¼ 7.2 Hz,
NMR ([D6]DMSO):
d
¼ 3.76 (s, 9H, 3 ꢀ MeO), 4.26 (d, J ¼ 4.4 Hz, 2H,
6H, 2 ꢀ Me), 2.99 (q, J ¼ 7.2 Hz, 4H, CH2), 4.83 (s, 2H, CH2), 5.13 (s,
2H, CH2), 6.87e6.89 (m, 1H, ArH), 7.07e7.14 (br s, 2H, exchangeable,
NH), 7.21e7.29 (m, 2H, 2 ꢀ ArH), 7.55 (d, J ¼ 8.4 Hz, 2H, 2 ꢀ ArH),
7.63 (d, J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 7.88e7.90 ppm (m, 1H, ArH); 13C
CH2), 4.66 (d, J ¼ 4.4 Hz, 2H, CH2), 4.71 (t, J ¼ 4.8 Hz, 1H,
exchangeable, OH), 5.09 (t, J ¼ 4.8 Hz, 1H, exchangeable, OH), 6.80
(s, 2H, 2 ꢀ ArH), 7.03e7.05 (m, 1H, ArH), 7.17e7.22 (m, 2H, 2 ꢀ ArH),
7.77e7.79 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 54.9, 56.1,
NMR ([D6]DMSO):
d
¼ 14.9, 15.0, 34.9, 35.0, 56.6, 57.0, 109.4, 112.8,
56.5, 60.7, 108.5, 113.2, 114.6, 123.9, 124.6, 124.7, 125.6, 125.8, 126.5,
126.6, 131.8, 134.9, 137.9, 153.2 ppm; Anal. calcd for C21H21NO5S: C
63.14, H 5.30, N 3.51, S 8.03, found: C 63.17, H 5.35, N 3.49, S 8.14.
122.6, 124.1, 124.4, 125.3, 125.5, 127.7, 128.7, 128.9, 130.7, 132.3,
133.6, 134.0, 155.7, 156.0 ppm; Anal. calcd for C24H24ClN3O4S: C
59.31, H 4.98, N 8.65, S 6.60, found: C 59.44, H 4.95, N 8.52, S 6.72.
4.2.30. [1-Cyclopropylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
dimethanol (17j)
4.2.35. [1-(3,4-Difluorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(ethylcarbamate) (18d)
Compound 17j was prepared from 16j (1.8 g, 5.4 mmol) and
Compound 18d was prepared from 17d (0.2 g, 0.6 mmol), Et3N
(0.3 mL), and ethylisocyanate (0.17 g, 2.4 mmol). Yield, 0.24 g (85%);
LiAlH4 (0.5 g, 13.0 mmol); Yield: 1.0 g (67%); mp: 153e154 ꢁC; 1H
NMR ([D6]DMSO):
d
¼ 0.71 (m, 2H, CH2), 1.12 (m, 2H, CH2), 2.04 (m,
mp: 167e168 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.00 (t, J ¼ 7.2 Hz, 6H,
1H, CH), 4.47 (d, J ¼ 4.8 Hz, 2H, CH2), 4.55 (m, 3H, CH2 and
exchangeable OH), 4.96 (t, J ¼ 4.8 Hz, 1H, exchangeable, OH), 7.23 (t,
J ¼ 7.6 Hz, 1H, ArH), 7.38 (t, J ¼ 7.6 Hz, 1H, ArH), 7.77 (d, J ¼ 8 Hz, 1H,
2 ꢀ Me), 2.98 (q, J ¼ 7.2 Hz, 4H, CH2), 4.84 (s, 2H, CH2), 5.12 (s, 2H,
CH2), 6.87e6.88 (m, 1H, ArH), 7.06e7.13 (br s, 2H, exchangeable,
NH), 7.20e7.28 (m, 2H, 2 ꢀ ArH), 7.37e7.38 (m, 1H, ArH), 7.59e7.68
(m, 2H, 2 ꢀ ArH), 7.87e7.89 ppm (m, 1H, ArH); 13C NMR ([D6]
ArH), 8.20 ppm (d, J ¼ 8 Hz, 1H, ArH); 13C NMR ([D6]DMSO):
¼ 6.8,
d
7.6, 54.7, 56.0, 113.6, 114.0, 122.7, 123.5, 124.2, 125.8, 126.1,
126.6131.6, 135.5 ppm; Anal. calcd for C15H15NO2S: C 65.91, H 5.53,
N 5.12, S 11.73, found: C 65.63, H 5.42, N 5.10, S 11.54.
DMSO):
d
¼ 14.9, 15.0, 34.9, 35.0, 56.5, 57.0, 109.3, 112.9, 117.8, 119.8,
122.8, 124.1, 124.4, 125.6, 127.5, 127.8, 130.7, 134.0, 148.3, 148.9,
150.2, 150.8, 155.7, 156.0 ppm; Anal. calcd for C24H23F2N3O4S: C
59.13, H 4.76, N 8.62, S 6.58, found: C 59.36, H 4.65, N 8.32, S 6.80.
4.2.31. General procedure for preparation of bis(alkylcarbamate)
derivatives (18 and 19)
4.2.36. [1-(3,4-Dichlorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(ethylcarbamate) (18e)
A solution of bis(hydroxymethyl) derivatives (17aej, 1.0 equiv-
alent) in anhydrous THF was treated with excess triethylamine (6.0
equivalent) followed by excess alkylisocyanate (4.0 equivalent). The
reaction mixture was stirred at ambient temperature under an
argon atmosphere. After completion of the reaction (for 4e20 h),
Compound 18e was prepared from 17e (0.3 g, 0.8 mmol), Et3N
(0.4 mL), and ethylisocyanate (0.23 g, 3.0 mmol); Yield: 0.37 g
(90%); mp: 178e179 ꢁC; 1H NMR ([D6]DMSO):
6H, 2 ꢀ Me), 2.99 (q, J ¼ 7.0 Hz, 4H, CH2), 4.85 (s, 2H, CH2), 5.13 (s,
d
¼ 1.00 (t, J ¼ 7.0 Hz,

38
R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
2H, CH2), 6.93e6.95 (m, 1H, ArH), 7.08e7.14 (br s, 2H, exchangeable,
NH), 7.23e7.30 (m, 2H, 2 ꢀ ArH), 7.52e7.54 (m, 1H, ArH), 7.81e7.83
(m, 2H, 2 ꢀ ArH), 7.88e7.90 ppm (m, 1H, ArH); 13C NMR ([D6]
4.2.41. [1-Cyclopropylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
bis(methylene)bis(ethylcarbamate) (18j)
Compound 18j was prepared from 17j (0.2 g, 0.66 mmol), Et3N
(0.5 mL), and ethylisocyanate (0.18 g, 2.6 mmol); Yield: 0.23 g
DMSO):
d
¼ 14.9,15.0, 35.0, 35.1, 56.5, 56.9,109.6,112.9,123.2,123.9,
124.2,124.5,125.6,128.4,130.7,130.8,131.4,131.6,132.1,134.0,155.7,
156.0 ppm; Anal. calcd for C24H23Cl2N3O4S: C 55.39, H 4.45, N 8.07,
S 6.16, found: C 55.27, H 4.45, N 8.09, S 6.00.
(75%); mp: 187e188 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 0.76 (m, 2H, CH2),
0.99 (t, J ¼ 7.2 Hz, 6H, 2 ꢀ Me), 1.16 (m, 2H, CH2), 2.10 (m, 1H, CH),
3.00 (q, J ¼ 7.2 Hz, 4H, CH2), 5.00 (s, 2H, CH2), 5.07 (s, 2H, CH2),
7.03e7.08 (br s, 2H, exchangeable, NH), 7.30 (t, J ¼ 8.4 Hz, 1H, ArH),
7.43 (t, J ¼ 8.4 Hz, 1H, ArH), 7.84 (d, J ¼ 8 Hz, 1H, ArH), 8.27 ppm (d,
4.2.37. [1-(3-chloro-4-fluorophenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-diyl]bis(methylene)bis(ethylcarbamate) (18f)
Compound 18f was prepared from 17f (0.36 g, 1.0 mmol), Et3N
(0.5 mL), and ethylisocyanate (0.28 g, 4.0 mmol); Yield: 0.41 g
J ¼ 8 Hz,1H, ArH); 13C NMR ([D6]DMSO):
¼ 6.2, 7.1, 15.0, 34.9, 35.0,
d
56.8, 57.0, 108.2, 114.1, 121.5, 123.7, 123.9, 125.5, 125.6, 127.9, 130.6,
134.8, 155.9, 156.1 ppm; Anal. calcd for C21H25N3O4S$0.5H2O: C
59.92, H 6.13, N 9.98, S 7.62, found: C 59.60, H 5.87, N 9.89, S 7.53.
(82%); mp: 158e159 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.00 (t, J ¼ 7.2 Hz,
6H, 2 ꢀ Me), 2.99 (q, J ¼ 7.2 Hz, 4H, CH2), 4.84 (s, 2H, CH2), 5.13 (s,
2H, CH2), 6.86e6.88 (m, 1H, ArH), 7.01e7.12 (br s, 2H, exchangeable,
NH), 7.21e7.29 (m, 2H, 2 ꢀ ArH), 7.54e7.56 (m, 1H, ArH), 7.59e7.63
(m, 1H, ArH), 7.77e7.79 (m, 1H, ArH), 7.88e7.90 ppm (m, 1H, ArH);
4.2.42. [1-Methylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
bis(methylene)bis(iso-propylcarbamate) (19a)
Compound 19a was prepared from 17a (0.24 g, 1.0 mmol), Et3N
(0.5 mL), and isopropylisocyanate (0.34 g, 4.0 mmol); Yield: 0.33 g
13C NMR: ([D6]DMSO):
d
¼ 14.9, 15.0, 35.0, 35.1, 56.5, 57.0, 109.4,
112.8, 117.1, 119.8, 122.9, 124.1, 124.4, 125.6, 127.8, 130.7, 131.5, 131.6,
132.6, 134.0, 155.7, 156.0, 156.5, 158.5 ppm; Anal. calcd for
C24H23ClFN3O4S: C 57.20, H 4.60, N 8.34, S 6.36, found: C 57.26, H
4.47, N 8.04, S 6.50.
(79%); mp: 172e173 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.03 (d, J ¼ 6.4 Hz,
12H, 4 ꢀ Me), 2.67 (s, 3H, Me), 3.59 (m, 2H, CH), 5.02 (s, 2H, CH2),
5.04 (s, 2H, CH2), 6.97e6.99 (br s, 2H, exchangeable, NH), 7.29 (t,
J ¼ 8.0 Hz, 1H, ArH), 7.41 (t, J ¼ 8.0 Hz, 1H, ArH), 7.84 (d, J ¼ 8.0 Hz,
1H, ArH), 7.91 ppm (d, J ¼ 8.0 Hz, 1H, ArH); 13C NMR ([D6]DMSO):
4.2.38. [1-(4-Methoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(ethylcarbamate) (18g)
Compound 18g was prepared from 17g (0.2 g, 0.6 mmol), Et3N
(0.3 mL), and ethylisocyanate (0.17 g, 2.4 mmol); Yield: 0.24 g
d
¼ 11.4, 22.5, 42.2, 42.3, 56.1, 56.9, 66.9, 108.3, 113.1, 120.5, 123.1,
123.7, 124.0, 125.1, 125.7, 130.6, 134.8, 155.3, 155.4 ppm; Anal. calcd
for C21H27N3O4S: C 60.41, H 6.52, N 10.06, S 7.68, found: C 60.08, H
6.24, N 9.76, S 7.97.
(85%); mp: 146e147 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.00 (t, J ¼ 7.2 Hz,
6H, 2 ꢀ Me), 2.99 (q, J ¼ 7.2 Hz, 4H, CH2), 3.87 (s, 3H, OMe), 4.80 (s,
2H, CH2), 5.11 (s, 2H, CH2), 6.83e6.85 (m, 1H, ArH), 7.04e7.10 (br s,
2H, exchangeable, NH), 7.11e7.13 (m, 2H, 2 ꢀ ArH), 7.18e7.26 (m,
2H, 2 ꢀ ArH), 7.41e7.43 (m, 2H, 2 ꢀ ArH), 7.84e7.86 ppm (m, 1H,
4.2.43. [1-(4-Fluorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
bis(methylene)bis(iso-propylcarbamate) (19b)
Compound 19b was prepared from 17b (0.33 g, 1.0 mmol), Et3N
(0.5 mL), and isopropylisocyanate (0.34 g, 4.0 mmol); Yield: 0.47 g
ArH); 13C NMR ([D6]DMSO):
109.0, 112.7,114.1, 121.9, 122.0,123.9, 124.3,125.4, 126.7, 130.7, 132.0,
134.2, 155.8, 156.1, 159.6 ppm; Anal. calcd for C25H27N3O5S: C 62.35,
H 5.65, N 8.73, S 6.66, found: C 62.19, H 5.55, N 8.75, S 7.03.
d
¼ 14.9,15.0, 34.9, 35.0, 56.1, 56.9, 57.1,
(95%); mp: 174e175 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.04 (d, J ¼ 6.6 Hz,
12H, 4 ꢀ Me), 3.59 (m, 2H, CH), 4.81 (s, 2H, CH2), 5.12 (s, 2H, CH2),
6.79e6.81 (m, 1H, ArH), 6.99e7.05 (br s, 2H, exchangeable, NH),
7.18e7.26 (m, 2H, 2 ꢀ ArH), 7.38e7.42 (m, 2H, 2 ꢀ ArH), 7.55e7.58
(m, 2H, 2 ꢀ ArH), 7.86e7.88 ppm (m, 1H, ArH); 13C NMR ([D6]
4.2.39. [1-(3,4-Dimethoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-diyl]bis(methylene)bis(ethylcarbamate) (18h)
Compound 18h was prepared from 17h (0.2 g, 0.5 mmol), Et3N
(0.3 mL), and ethylisocyanate (0.14 g, 2.0 mmol); Yield: 0.25 g
DMSO):
d
¼ 22.5, 42.2, 42.3, 56.5, 56.9, 109.2, 112.7, 115.6, 115.8,
122.4, 124.1, 124.4, 125.5, 125.6, 126.4, 127.4, 130.8, 132.9, 134.1,
155.1, 155.4, 161.4, 163.3 ppm; Anal. calcd for C26H28FN3O4S: C
62.76, H 5.67, N 8.44, S 6.44, found: C 62.46, H 5.61, N 8.30, S 6.48.
(90%); mp: 145e146 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.00 (t, J ¼ 7.0 Hz,
6H, 2 ꢀ Me), 2.99 (q, J ¼ 7.0 Hz, 4H, CH2), 3.74 (s, 3H, MeO), 3.87 (s,
3H, MeO), 4.85 (s, 2H, CH2), 5.12 (s, 2H, CH2), 6.93e6.95 (m, 1H,
ArH), 7.03e7.08 (br s, 2H, exchangeable, NH), 7.09e7.11 (m, 2H,
2 ꢀ ArH), 7.13e7.15 (m, 1H, ArH), 7.21e7.26 (m, 2H, 2 ꢀ ArH),
4.2.44. [1-(4-Chlorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(iso-propylcarbamate) (19c)
Compound 19c was prepared from 17c (0.17 g, 0.5 mmol), Et3N
(0.3 mL), and isopropylisocyanate (0.17 g, 2.0 mmol); Yield: 0.2 g
7.84e7.86 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 15.0, 34.9,
(78%); mp: 171e172 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.04 (d, J ¼ 6.4 Hz,
35.0, 55.4, 57.0, 57.1, 109.1, 111.6, 113.0, 114.0, 121.8, 122.0, 123.1,
124.0, 124.3, 125.4, 126.8, 127.0, 130.8, 134.3, 148.5, 149.2, 155.9,
156.1 ppm; Anal. calcd for C26H29N3O6S: C 61.04, H 5.71, N 8.21, S
6.27, found: C 60.81, H 5.71, N 8.11, S 6.19.
12H, 4 ꢀ Me), 3.59 (m, 2H, CH), 4.82 (s, 2H, CH2), 5.12 (s, 2H, CH2),
6.87e6.89 (m, 1H, ArH), 6.97e7.05 (br s, 2H, exchangeable, NH),
7.20e7.28 (m, 2H, 2 ꢀ ArH), 7.54 (d, J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 7.63 (d,
J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 7.87e7.89 ppm (m, 1H, ArH); 13C NMR ([D6]
DMSO):
d
¼ 22.5, 42.2, 42.3, 56.4, 56.8, 109.4, 112.8, 122.6, 124.1,
4.2.40. [1-(3,4,5-Trimethoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-diyl]bis(methylene)bis(ethylcarbamate) (18i)
Compound 18i was prepared from 17i (0.39 g, 1.0 mmol), Et3N
(0.5 mL), and ethylisocyanate (0.28 g, 4.0 mmol); Yield: 0.48 g
124.4, 125.3, 125.5, 127.8, 128.7, 128.9, 130.8, 132.3, 133.6, 134.0,
155.0, 155.4 ppm; Anal. calcd for C26H28ClN3O4S$0.5H2O: C 59.70, H
5.59, N 8.03, S 6.13, found: C 59.38, H 5.32, N 7.87, S 6.50.
(89%); mp: 196e197 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 0.99 (t, J ¼ 7.2 Hz,
4.2.45. [1-(3,4-Difluorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(iso-propylcarbamate) (19d)
Compound 19d was prepared from 17d (0.17 g, 0.5 mmol), Et3N
(0.3 mL), and isopropylisocyanate (0.17 g, 2.0 mmol); Yield: 0.2 g
6H, 2 ꢀ Me), 2.99 (q, J ¼ 7.2 Hz, 4H, CH2), 3.76 (s, 6H, 2 ꢀ OMe), 3.78
(s, 3H, MeO), 4.85 (s, 2H, CH2), 5.12 (s, 2H, CH2), 6.80 (s, 2H,
2 ꢀ ArH), 7.01e7.03 (m, 1H, ArH), 7.07e7.12 (br s, 2H, exchangeable,
NH), 7.24e7.27 (m, 2H, 2 ꢀ ArH), 7.85e7.87 ppm (m, 1H, ArH); 13C
(79%); mp: 173e174 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.03 (d, J ¼ 6.6 Hz,
NMR ([D6]DMSO):
d
¼ 14.9, 15.0, 34.9, 35.0, 55.9, 57.0, 60.1, 107.9,
12H, 4 ꢀ Me), 3.58 (m, 2H, CH), 4.83 (s, 2H, CH2), 5.12 (s, 2H, CH2),
6.87e6.89 (m, 1H, ArH), 6.98e7.04(br s, 2H, exchangeable, NH),
7.20e7.28 (m, 2H, 2 ꢀ ArH), 7.38 (m, 1H, ArH), 7.59e7.67 (m, 2H,
2 ꢀ ArH), 7.87e7.89 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
109.3, 113.2,121.9, 124.1, 124.3, 125.1, 125.4, 127.0, 127.2, 130.8, 134.2,
137.7, 152.8, 155.8, 156.1 ppm; Anal. calcd for C27H31N3O7S: C 59.87,
H 5.77, N 7.76, S 5.92, found: C 59.49, H 5.72, N 7.44, S 5.78.

R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
39
d
¼ 22.5, 42.2, 42.3, 56.3, 56.8, 109.4, 112.9, 117.8, 117.9, 119.8, 119.9,
(88%); mp: 195e196 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.04 (d, J ¼ 6.4 Hz,
122.9, 124.1, 124.4, 125.6, 127.5, 127.9, 130.8, 134.0, 148.3, 150.3,
155.0, 155.4 ppm; Anal. calcd for C26H27F2N3O4S: C 60.57, H 5.28, N
8.15, S 6.22, found: C 60.22, H 5.02, N 7.94, S 6.36.
12H, 4 ꢀ Me), 3.59 (m, 2H, CH), 3.77 (s, 6H, 2 ꢀ MeO), 3.79 (s, 3H,
MeO), 4.85 (s, 2H, CH2), 5.12 (s, 2H, CH2), 6.80 (s, 2H, 2 ꢀ ArH),
7.00e7.02 (m, 1H, ArH), 7.03e7.08 (br s, 2H, exchangeable, NH),
7.24e7.27 (m, 2H, 2 ꢀ ArH), 7.86e7.88 ppm (m, 1H, ArH); 13C NMR
4.2.46. [1-(3,4-Dichlorophenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(iso-propylcarbamate) (19e)
([D6]DMSO):
d
¼ 22.5, 23.2, 42.2, 42.3, 55.9, 56.8, 56.9, 60.1, 107.8,
107.9, 109.4, 113.2, 121.9, 124.1, 124.3, 125.1, 125.4, 127.0, 127.3, 130.8,
134.2, 137.7, 152.8, 155.2, 155.4 ppm; Anal. calcd for C29H35N3O7S: C
61.14, H 6.19, N 7.38, S 5.63, found: C 60.77, H 6.06, N 7.00, S 5.78.
Compound 19e was prepared from 17e (0.25 g, 0.66 mmol), Et3N
(0.4 mL), and isopropylisocyanate (0.22 g, 2.6 mmol); Yield: 0.32 g
(88%); mp: 181e182 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.04 (d, J ¼ 6.6 Hz,
12H, 4 ꢀ Me), 3.59 (m, 2H, CH), 4.85 (s, 2H, CH2), 5.13 (s, 2H, CH2),
6.94e6.96 (m, 1H, ArH), 7.00e7.06 (br s, 2H, exchangeable, NH),
7.23e7.30 (m, 2H, 2 ꢀ ArH), 7.52e7.54 (m, 1H, ArH), 7.81e7.83(m,
2H, 2 ꢀ ArH), 7.88e7.91 ppm (m, 1H, ArH); 13C NMR ([D6]DMSO):
4.2.51. [1-Cyclopropylbenzo[d]pyrrolo[2,1-b]thiazole-2,3-diyl]
bis(methylene)bis(iso-propylcarbamate) (19j)
Compound 19j was prepared from 17j (0.13 g, 0.5 mmol), Et3N
(0.4 mL), and isopropylisocyanate (0.17 g, 2.0 mmol); Yield: 0.17 g
d
¼ 22.5, 42.2, 42.3, 56.3, 56.7, 109.6, 112.9, 123.2, 123.9, 124.2, 124.4,
(77%); mp: 215e216 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 0.77 (m, 2H, CH2),
125.6, 128.4, 130.7, 130.8, 131.4, 131.6, 132.0, 134.0, 155.0, 155.4 ppm;
Anal. calcd for C26H27Cl2N3O4S: C 56.94, H 4.96, N 7.66, S 5.85,
found: C 56.66, H 4.93, N 7.54, S 5.68.
1.04 (d, J ¼ 6.4 Hz, 12H, 4 ꢀ Me), 1.16 (m, 2H, CH2), 2.10 (m, 1H, CH),
3.60 (m, 2H, CH), 5.01 (s, 2H, CH2), 5.07 (s, 2H, CH2), 6.95 (br s, 2H,
exchangeable, NH), 7.31 (t, J ¼ 7.6 Hz,1H, ArH), 7.44 (t, J ¼ 7.6 Hz,1H,
ArH), 7.85 (d, J ¼ 7.6 Hz, 1H, ArH), 8.27 ppm (d, J ¼ 7.6 Hz, 1H, ArH);
4.2.47. [1-(3-chloro-4-fluorophenyl)benzo[d]pyrrolo[2,1-b]
thiazole-2,3-diyl]bis(methylene)bis(iso-propylcarbamate) (19f)
Compound 19f was prepared from 17f (0.18 g, 0.5 mmol), Et3N
(0.4 mL), and isopropylisocyanate (0.17 g, 2.0 mmol); Yield: 0.2 g
13C NMR ([D6]DMSO):
121.6, 123.7, 123.9, 125.5, 125.6, 127.9,130.6,134.8,155.3,155.4 ppm;
Anal. calcd for C23H29N3O4S: C 62.28, H 6.59, N 9.47, S 7.23, found: C
62.02, H 6.68, N 9.22, S 7.40.
d
¼ 6.2, 7.1, 22.5, 42.2, 56.6, 56.8, 108.2, 114.1,
(75%); mp: 178e179 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.04 (d, J ¼ 6.6 Hz,
12H, 4 ꢀ Me), 3.60 (m, 2H, CH), 4.84 (s, 2H, CH2), 5.13 (s, 2H, CH2),
6.86e6.88 (m, 1H, ArH), 6.99e7.06 (br s, 2H, exchangeable, NH),
7.21e7.29 (m, 2H, 2 ꢀ ArH), 7.53e7.55 (m,1H, ArH), 7.62e7.64 (m,1H,
ArH), 7.77e7.79 (m, 1H, ArH), 7.88e7.90 ppm (m, 1H, ArH); 13C NMR
4.3. Biological assays
4.3.1. Cytotoxicity assays
([D6]DMSO):
d
¼ 22.5, 23.2, 42.2, 42.3, 56.3, 56.8, 109.4, 112.8, 117.3,
The cytotoxic effects of the newly synthesized compounds were
determined in T-cell acute lymphocytic leukemia (CCRF-CEM) and
their resistant sub-cell lines (CCRF-CEM/Taxol and CCRF-CEM/VBL)
by the XTT assay [19] and human solid tumor cells (i.e. breast
carcinoma MX-1 and colon carcinoma HCT-116) by the SRB assay
[20] in a 72 h incubation using a microplate spectrophotometer as
previously described [21]. After the addition of phenazine
methosulfate-XTT solution, incubated at 37 ꢁC for 6 h and absor-
bance at 450 and 630 nm was detected on a microplate reader (EL
340). The cytotoxicity of the newly synthesized compounds against
non-small cell lung carcinoma H1299, prostate cancer PC-3, oral
cancer OECM-1 and glioma U87 were determined by the Alamar
blue assay [22] in a 72 h incubation using a microplate spectro-
photometer as previously described. After the addition of Alamar
blue solution, it was incubated at 37 ꢁC for 6 h. Absorbance at 570
and 600 nm was detected on a microplate reader. IC50 values were
determined from dose-effect relationship at six or seven concen-
trations of each drug using the CompuSyn software by Chou and
Martin [23] based on the median-effect principle and plot [24,25].
Ranges given for the values of IC50 were mean ꢂ SE (n ¼ 4).
119.9, 123.0, 124.1, 124.4, 125.6, 127.9, 130.8, 131.4, 131.5132.6, 134.0,
155.0, 155.4, 156.5, 158.5 ppm; Anal. calcd for C26H27ClFN3O4S: C
58.70, H 5.12, N 7.90, S 6.03, found: C 58.80, H 5.20, N 7.88, S 5.73.
4.2.48. [1-(4-Methoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-2,3-
diyl]bis(methylene)bis(iso-propylcarbamate) (19g)
Compound 19g was prepared from 17g (0.25 g, 0.7 mmol), Et3N
(0.4 mL), and isopropylisocyanate (0.24 g, 2.8 mmol); Yield: 0.32 g
(86%); mp: 141e142 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.04 (d, J ¼ 6.6 Hz,
12H, 4 ꢀ Me), 3.59 (m, 2H, CH), 3.87 (s, 3H, MeO), 4.81 (s, 2H, CH2),
5.11 (s, 2H, CH2), 6.83e6.85 (m, 1H, ArH), 6.97e7.04 (br s, 2H,
exchangeable, NH), 7.12 (d, J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 7.17e7.25 (m,
2H, 2 ꢀ ArH), 7.42 (d, J ¼ 8.4 Hz, 2H, 2 ꢀ ArH), 7.84e7.86 ppm (m,
1H, ArH); 13C NMR ([D6]DMSO):
d
¼ 22.5, 42.2, 42.3, 55.2, 56.7, 57.0,
109.1,112.7,114.1,122.0, 123.9, 124.3,125.4,126.7,130.8,132.0,134.3,
155.1, 155.4, 159.6 ppm; Anal. calcd for C27H31N3O5S: C 63.63, H
6.13, N 8.25, S 6.29, found: C 63.38, H 6.06, N 8.05, S 6.61.
4.2.49. [1-(3,4-Dimethoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-diyl]bis(methylene)bis(iso-propylcarbamate) (19h)
Compound 19h was prepared from 17h (0.18 g, 0.5 mmol), Et3N
(0.4 mL), and isopropylisocyanate (0.17 g, 2.0 mmol); Yield: 0.23 g
4.3.2. In vivo studies
Athymic nude mice bearing the nu/nu gene were obtained from
NCI, Frederick, MD and used for all human tumor xenografts. Male
nude mice 6 weeks or older weighing 20e24 g or more were used.
Compounds were administered via the tail vein for i.v. injection as
described previously [21]. Tumor volume was assessed by
measuring length ꢀ width ꢀ height (or width) by using a caliper.
(89%); mp: 145e146 ꢁC; 1H NMR ([D6]DMSO):
d
¼ 1.04 (d, J ¼ .6 Hz,
12H, 4 ꢀ Me), 3.59 (m, 2H, CH), 3.74 (s, 3H, MeO), 3.86 (s, 3H, MeO),
4.85 (s, 2H, CH2), 5.12 (s, 2H, CH2), 6.94e6.96 (m, 1H, ArH), 6.96e7.01
(br s, 2H, exchangeable, NH), 7.02e7.06(m, 2H, 2 ꢀ ArH), 7.14e7.17 (m,
1H, ArH), 7.19e7.29 (m, 2H, 2 ꢀ ArH), 7.85e7.87 ppm (m,1H, ArH); 13C
NMR ([D6]DMSO):
d
¼ 22.5, 23.2, 42.2, 42.3, 55.4, 56.9, 109.1, 111.6,
Vehicle used was 50 mL DMSO and 40 mL Tween 80 in 160 mL saline.
113.0, 114.0, 121.8, 122.1, 123.1, 124.0, 124.3, 125.4, 126.8, 127.0, 130.8,
134.3, 148.5, 149.2, 155.2, 155.4 ppm; Anal. calcd for C28H33N3O6S: C
62.32, H 6.16, N 7.79, S 5.94, found: C 62.11, H 5.90, N 7.45, S 6.15.
The maximal tolerate dose of the tested compound was determined
and applied for the in vivo therapeutic efficacy assay. For tumor-
bearing nude mice during the course of the experiment, the body
weight refers to total weight minus the weight of the tumor. All
animal studies were conducted in accordance with the guidelines
for the National Institute of Health Guide for the Care and Use of
Animals and the protocol approved by the Institutional Animal Care
and Use Committee.
4.2.50. [1-(3,4,5-Trimethoxyphenyl)benzo[d]pyrrolo[2,1-b]thiazole-
2,3-diyl]bis(methylene)bis(iso-propylcarbamate) (19i)
Compound 19i was prepared from 17i (0.2 g, 0.5 mmol), Et3N
(0.4 mL), and isopropylisocyanate (0.17 g, 2.0 mmol); Yield: 0.25 g

40
R. Chaniyara et al. / European Journal of Medicinal Chemistry 53 (2012) 28e40
4.3.3. Alkaline agarose gel shift assay
obtained at the High-Field Biomolecular NMR Core Facility sup-
ported by the National Research Program for Genomic Medicine
(Taiwan). We thank Dr. Shu-Chuan Jao at the Institute of Biological
Chemistry (Academia Sinica) for providing NMR service and the
NationalCenter for High-Performance Computing for computer
time and facilities.
Formation of DNA cross-linking was analyzed by alkaline
agarose gel electrophoresis as previously described [13]. In brief,
purified pEGFP-N1 plasmid DNA (1500 ng) was mixed with various
concentrations (1e20
mM) of 17b, 18b, 19b, 17c, 18c, 19c, 17g, 18g,
and 19g in 40 L binding buffer (3 mM sodium chloride/1 mM
m
sodium phosphate, pH 7.4, and 1 mM EDTA). The reaction mixture
was incubated at 37 ꢁC for 2 h. At the end of reaction, the plasmid
DNA was linearized by digestion with BamHI and followed by
precipitation with ethanol. The DNA pellets were dissolved and
denatured in alkaline buffer (0.5 N NaOH-10 mM EDTA). An aliquot
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Acknowledgements
We are grateful for support from the National Science Council,
Taiwan, Republic of China (NSC 99-2320-B-001-014 and NSC 100-
2325-B-001-003) and Academia Sinica, Taipei, Taiwan (AS-100-TP-
B13). Dr. T.-C. Chou was supported by the Sloan-Kettering Institute
General Fund. NMR spectra of synthesized compounds were