Synthesis of novel isatin-thiazoline and isatin-benzimidazole conjugates as anti-breast cancer agents

Article abstract of DOI:10.1007/s12272-011-1005-3

A series of new isatin-thiazoline 3a-h and isatin-benzimidazole 4a-h derivatives were synthesized via condensation of isatin Mannich bases 2a-h with either 2-aminothiazoline or 2-aminobenzimidazole. The structures of the newly synthesized compounds were c

Full text of DOI:10.1007/s12272-011-1005-3

Arch Pharm Res Vol 34, No 10, 1615-1621, 2011
DOI 10.1007/s12272-011-1005-3
Synthesis of Novel Isatin-Thiazoline and Isatin-Benzimidazole
Conjugates as Anti-Breast Cancer Agents
Azza T. Taher, Nadia A. Khalil, and Eman M. Ahmed
Department of Organic Chemistry, Faculty of Pharmacy, Cairo University, Cairo, Egypt
(Received May 4, 2011/Revised June 15, 2011/Accepted June 21, 2011)
A series of new isatin-thiazoline 3a
-
h
and isatin-benzimidazole 4a
-h derivatives were synthe-
sized via condensation of isatin Mannich bases 2a
-h
with either 2-aminothiazoline or 2-ami-
nobenzimidazole. The structures of the newly synthesized compounds were characterized by
spectral data. The anti-breast cancer activity of some of the synthesized compounds was
assessed in the MCF-7 human breast cancer cell line. The results showed that compounds 4b
,
4d and 4g possess significant antiproliferative activity against MCF-7 cells.
Key words: Isatin, Benzimidazole, Thiazoline, Breast cancer, Anticancer activity
angiogenic properties of SU-5416 (Semaxanib) and
SU-11248 (Sunitinib) (Matesic et al., 2008) (Fig. 1).
INTRODUCTION
Cancer is a major health concern worldwide. The The latter compound has received FDA approval for
relative mortality rate caused by cancer is very high the treatment of gastrointestinal stromal tumors and
in developing countries. Furthermore, breast carcin- advanced renal cell carcinoma (Motzer et al., 2006).
oma is the most common cancer in women worldwide
Besides the exploitation of new targets, there is an-
and remains the most frequent cause of malignancy- other approach that combines two or more pharmaco-
associated deaths among women (Smith et al., 2009). phores with anticancer activity, each with a different
Although chemotherapy is the mainstay of cancer mode of action, into a single molecule (Matesic et al.,
therapy, the use of available chemotherapeutics is often 2008; Mayur et al., 2009). In our ongoing search for
limited due to undesirable side effects as well as the new pharmacophores as anti-breast cancer agents,
increasing incidence of drug resistance to cancer che- benzimidazole-based compounds (I, II, Fig. 2) demon-
motherapeutic agents, which is a serious medical pro- strated high cytotoxic activity against hormone-depend-
blem (Rojo et al., 2008). Therefore, there is an urgent ent breast cancer cells (MCF-7) (Ramla et al., 2006).
need to develop new classes of chemotherapeutic agents Benzimidazole derivatives selectively inhibit endothelial
to treat cancer. Isatin (1H-indole-2,3-dione) is one of cell growth and suppress angiogenesis in vitro and in
the most promising heterocyclic molecules that have vivo (Hori et al., 2002). Furthermore, other authors
interesting active profiles, and importantly, it is well have reported antiproliferative or anticancer potential
tolerated by humans (Pandeya et al., 2005; Vine et al., of certain thiazolinederivatives (Rostom, 2006; Johnson
2009). Numerous publications have reported that the
isatin scaffold shows anticancer activity against various
human tumor cell lines (Bramson et al., 2001; Solomon
et al., 2010). Significant interest in 2-oxoindole deriva-
tives as kinase inhibitors came after the disclosure of
the tyrosine kinase inhibitory properties and anti-
Correspondence to: Azza T. Taher, Department of Organic
Chemistry, Faculty of Pharmacy, Cairo University, Cairo, Egypt
Tel: 2-10-147-0792
E-mail: azzataher2005@yahoo.com
Fig. 1. Chemical structures of isatin analogues reported for
anticancer activity
1615

1616
A. T. Taher et al.
Fig. 2. Structures of previously synthesized benzimidazole analogues with anti-breast cancer activity
et al., 2007).
Due to the great potential of both moieties, and en- with either benzimidazole scaffold or thiazoline moiety
couraged by these previous reports, we postulated that by a Schiff base reaction 3a and 4a (Scheme 1),
Herein we report the synthesis of isatin conjugates
-h
-h
a compound containing both isatin and benzimidazole and the in vitro cytotoxic effects of some of the syn-
or thiazoline pharmacophores could have synergistic thesized compounds on a human breast carcinoma cell
effects as an anticancer agent.
line (MCF-7).
Scheme 1. Synthesis of isatin Mannich bases and Schiff base analogues. Reagents and conditions: (a) Mannich reaction
(HCHO and secondary amine, ethanol); (b) 2-Aminothiazoline, ethanol, AcOH, 13-15 h; (c) 2-Aminobenzimidazole, ethanol,
AcOH, 13-15 h

Novel Isatin-Thiazoline and Isatin-Benzimidazole Conjugates
1617
2CH2 piperazinyl), 3.05-3.11 (m, 4H, 2CH2 piperazinyl),
3.77 (s, 3H, OCH3), 4.55 (s, 2H, CH2), 6.82-6.94 (m, 4H,
Ar-H), 7.15 (d, 2H, Ar-H), 7.62 (d, 2H, Ar-H). Anal.
MATERIALS AND METHODS
Chemistry
All melting points were determined on a Stuart ap- calcd for C20H21N3O3 (351.40): C, 68.36; H, 6.02; N,
paratus and are uncorrected. The IR spectra were re- 11.96. Found: C, 68.55; H, 6.18; N, 11.70.
corded on a Perkin-Elmer spectrophotometer using
1
potassium bromide discs. H-NMR spectra were re- 5-Bromo-1-[4-(4-methoxyphenyl)piperazin-1-yl]
corded using a Varian Gemini 200 MHz Spectrophoto- methyl-1H-indole-2,3-dione (2f)
meter and Varian Mercury-300 (300 MHz) Spectropho- Yield 68%; m.p. 130-131oC; IR (KBr, cm−1): 3062, 3050
tometer using TMS as an internal standard. Chemical (C-H aromatic), 2947, 2831 (C-H aliphatic), 1750, 1732
1
shift values were recorded in ppm on
δ
scale, Micro- (C=O); H-NMR 300 MHz (CDCl3): 2.78-2.83 (m, 4H,
analytical Center, Cairo University, Egypt. Mass spec- 2CH2 piperazinyl), 3.05-3.12 (m, 4H, 2CH2 piperazinyl),
tra were recorded on a GCMP-QP1000EX Mass spec- 3.77 (s, 3H, OCH3), 4.53 (s, 2H, CH2), 6.83-7.06 (m,
trometer, Microanalytical Center, Cairo University, 5H, Ar-H), 7.71-7.74 (m, 2H, Ar-H). Anal. calcd for
Egypt. Elemental analyses were performed on a Perkin- C20H20BrN3O3 (430.30): C, 55.83; H, 4.68; N, 9.77. Found:
Elmer 2400 C, H, N analyzer at the Microanalytical C, 55.60; H, 4.55; N, 9.96.
Center, Cairo University, Egypt, and values were
within ± 0.4 of theoretical percentages. Progress of the
reactions was monitored by TLC. TLC sheets were
precoated with UV florescent silica gel Merck 60 F
254; CHCl3-CH3OH (9.5:0.5) was used as a solvent and
products were visualized using UV lamp.
General synthetic procedure for the synthesis
of 3-(4,5-dihydro[1,3]thiazol-2-ylimino)-1-(substi-
tuted-1-ylmethyl)indol-2(3H)-ones (3a-h) and 3-
(1H-benzimidazol-2-ylimino)-1-(substituted-1-yl-
methyl)indol-2(3H)-ones (4a-h)
Equimolar quantities of an appropriate Mannich
base 2a-h and 2-amino-4,5-dihydrothiazole or 2-amino-
1H-benzo[d]imidazole (3.65 mmol each) were dissolv-
General procedure for the preparation of 1-(sub-
stitutedmethyl)-1H-indole-2,3-diones (2a-h)
To a solution of isatin or 5-bromoisatin (4.08 mmol) ed in absolute ethanol (50 mL) containing glacial
in absolute ethanol (10 mL) was added a mixture of acetic acid (0.5 mL). The reaction mixture was heated
secondary amino compound (1.35 mmol) and forma- under reflux for 13-15 h, concentrated under reduced
line 37% (7 mmol) in absolute ethanol (15 mL). The pressure and left to cool. The precipitated solid was
reaction mixture was stirred for 6 h at 50oC and then filtered and crystallized from ethanol (70%) to give 3a
-
refrigerated for 48 h. The crystalline products were
separated by filtration, washed with cold methanol,
h or 4a-h.
dried and recrystallized from ethanol. Compounds 2a
, and were prepared as reported (Solomon et 1-ylmethyl) indol-2(3H)-one (3a)
al., 2009).
Yield 70%; m.p. 160-161oC; IR (KBr, cm−1): 3062, 3050
,
3-(4,5-Dihydro[1,3]thiazol-2-ylimino)-1-(piperidin-
b,
c,
g
h
(C-H aromatic), 2932, 2855 (C-H aliphatic), 1728 (C=O),
1
1-[4-(2-Methoxyphenyl)piperazin-1-yl]methyl-1H-
indole-2,3-dione (2d)
1616 (C=N); H-NMR 300 MHz (DMSO-d6): 1.05-1.46
(m, 6H, 3CH2 piperidinyl), 2.76-2.84 (m, 4H, 2CH2 pi-
Yield 76%; m.p. 144-145oC; IR (KBr, cm−1): 3062, 3050 peridinyl), 3.55 (t, 2H, CH2 thiazoline), 4.36 (t, 2H,
(C-H aromatic), 2943, 2831 (C-H aliphatic), 1750, 1735 CH2 thiazoline), 4.69 (s, 2H, CH2), 6.59-7.58 (m, 4H,
1
(C=O); H-NMR 300 MHz (DMSO-d6): 2.71-2.74 (m, Ar-H). Anal. calcd for C17H20N4OS (328.43): C, 62.17;
4H, 2CH2 piperazinyl), 2.92-2.94 (m, 4H, 2CH2 pipera- H, 6.14; N, 17.06. Found: C, 62.38; H, 6.18; N, 17.45.
zinyl), 3.77 (s, 3H, OCH3), 4.48 (s, 2H, CH2), 6.85-6.92
(m, 4H, Ar-H), 7.15-7.17 (m, 1H, Ar-H), 7.30-7.33 (m, 3-(4,5-Dihydro[1,3]thiazol-2-ylimino)-1-(morpholin-
1H, Ar-H), 7.55-7.58 (m, 1H, Ar-H), 7.67-7.68 (m, 1H, 1-ylmethyl)indol-2(3H)-one (3b)
Ar-H). Anal. calcd for C20H21N3O3 (351.40): C, 68.36; Yield 65%; m.p. 172-173oC; IR (KBr, cm−1): 3101, 3050
H, 6.02; N, 11.96. Found: C, 68.15; H, 5.92; N, 11.75.
(C-H aromatic), 2955, 2897 (C-H aliphatic), 1732 (C=O),
1612 (C=N); H-NMR 200 MHz (DMSO-d6): 2.41-2.49
1
1-[4-(4-Methoxyphenyl)piperazin-1-yl]methyl-1H- (m, 4H, 2CH2 morpholinyl), 2.56 (t, 2H, CH2 thiazoline),
indole-2,3-dione (2e)
3.35-3.45 (m, 4H, 2CH2 morpholinyl), 3.53 (t, 2H, CH2
Yield 82%; m.p. 170-171oC; IR (KBr, cm−1): 3086, 3035 thiazoline), 4.39 (s, 2H, CH2), 6.90-7.71 (m, 4H, Ar-H);
(C-H aromatic), 2947, 2823 (C-H aliphatic), 1745, 1728 MS (EI) m/z (% rel. Int.): 330 (45.97, M+ ), 331 (16.93,
.
1
(C=O); H-NMR 300 MHz (CDCl3): 2.80-2.85 (m, 4H, M+H). Anal. calcd for C16H18N4O2S (330.40): C, 58.16;

1618
A. T. Taher et al.
H, 5.49; N, 16.96. Found: C, 58.25; H, 5.32; N, 16.68.
3-(4,5-Dihydro[1,3]thiazol-2-ylimino)-1-(N,N-diphen-
ylamino-1-ylmethyl)indol-2(3H)-one (3g)
3-(4,5-Dihydro[1,3]thiazol-2-ylimino)-1-[(4-phenyl- Yield 73%; m.p. 152-153oC; IR (KBr, cm−1): 3100, 3062
piperazin-1-yl)methyl]indol-2(3H)-one (3c)
(C-H aromatic), 2927, 2858 (C-H aliphatic), 1732 (C=O),
1
Yield 60%; m.p. 142-143oC; IR (KBr, cm−1): 3086, 3059 1616 (C=N); H-NMR 300 MHz (DMSO-d6): 2.88 (t,
(C-H aromatic), 2924, 2855 (C-H aliphatic), 1728 (C=O), 2H, CH2 thiazoline), 3.47 (t, 2H, CH2 thiazoline), 4.66
1616 (C=N); 1H-NMR 300 MHz (DMSO-d6): 2.73 (t, 2H, (s, 2H, CH2), 6.89-7.61 (m, 14H, Ar-H). Anal. calcd for
CH2 thiazoline), 2.82-2.88 (m, 4H, 2CH2 piperazinyl), C24H20N4OS (412.51): C, 69.88; H, 4.89; N, 13.58. Found:
3.00-3.10 (m, 4H, 2CH2 piperazinyl), 3.55 (t, 2H, CH2 C, 69.68; H, 4.62; N, 13.72.
thiazoline), 4.75 (s, 2H, CH2), 6.73-7.71 (m, 9H aroma-
tic). Anal. calcd for C22H23N5OS (405.52): C, 65.16; H, 3-(4,5-Dihydro[1,3]thiazol-2-ylimino)-1-[(4-methyl-
5.72; N, 17.27. Found: C, 64.95; H, 5.55; N, 17.05.
piperazin-1-yl)methyl]indol-2(3H)-one (3h)
Yield 65%; m.p. 138-139oC; IR (KBr, cm−1): 3120, 3050
(C-H aromatic), 2947, 2877 (C-H aliphatic), 1728 (C=O),
1604 (C=N); 1H-NMR 300 MHz (CDCl3): 2.65-2.83 (m,
4H, 2CH2 piperazinyl), 2.90-3.15 (m, 4H, 2CH2 pipera-
3-(4,5-Dihydro[1,3]thiazol-2-ylimino)-1-{[2-(meth-
oxyphenyl)piperazin-1-yl]methyl}indol-2(3H)-one
(3d)
Yield 60%; m.p. 100-101oC; IR (KBr, cm−1): 3080, 3059 zinyl), 3.36 (t, 2H, CH2 thiazoline), 3.76 (s, 3H, CH3),
(C-H aromatic), 2935, 2831 (C-H aliphatic), 1728 (C=O), 3.84 (t, 2H, CH2 thiazoline), 4.54 (s, 2H, CH2), 6.84-
1616 (C=N); 1H-NMR 300 MHz (CDCl3): 2.77-2.81 (m, 7.61 (m, 4H, Ar-H). Anal. calcd for C17H21N5OS (343.45):
4H, 2CH2 piperazinyl), 2.89 (t, 2H, CH2 thiazoline), C, 59.45; H, 6.16; N, 20.39. Found: C, 59.35; H, 6.44;
2.95-3.04 (m, 4H, 2CH2 piperazinyl), 3.52 (t, 2H, CH2 N, 20.15.
thiazoline), 3.76 (s, 3H, OCH3), 4.53 (s, 2H, CH2), 6.84-
7.27 (m, 8H, Ar-H). Anal. calcd for C23H25N5O2S (435.54): 3-(1H-Benzimidazol-2-ylimino)-1-(piperidin-1-yl-
C, 63.43; H, 5.79; N, 16.08. Found: C, 63.15; H, 5.88; methyl)indol-2(3H)-one (4a)
N, 15.90.
Yield 74%; m.p. 220-221oC; IR (KBr, cm−1): 3174, 3059
(C-H aromatic), 2931, 2858 (C-H aliphatic), 1735 (C=O),
1616 (C=N); H-NMR 200 MHz (DMSO-d6): 1.24-1.65
(m, 6H, 3CH2 piperidinyl), 2.95-3.05 (m, 4H, 2CH2 pi-
peridinyl), 4.60 (s, 2H, CH2), 6.98-7.89 (m, 8H aroma-
1
3-(4,5-Dihydro[1,3]thiazol-2-ylimino)-1-{[4-(meth-
oxyphenyl)piperazin-1-yl]methyl}indol-2(3H)-one
(3e)
Yield 68%; m.p. 150-151oC; IR (KBr, cm−1): 3086, 3059 tic), 11.06 (br s, 1H, NH, D2O exchangeable); MS (EI)
(C-H aromatic), 2935, 2832 (C-H aliphatic), 1728 (C=O), m/z (% rel. Int.): 359 (33, M+.). Anal. calcd for C21H21N5O
1
1616 (C=N); H-NMR 300 MHz (DMSO-d6): 2.60-2.68 (359.42): C, 70.17; H, 5.89; N, 19.48. Found: C, 70.25;
(m, 4H, 2CH2 piperazinyl), 2.72 (t, 2H, CH2 thiazoline), H, 5.99; N, 19.30.
2.85-2.98 (m, 4H, 2CH2 piperazinyl), 3.43 (t, 2H, CH2
thiazoline), 3.66 (s, 3H, OCH3), 4.45 (s, 2H, CH2), 6.58- 3-(1H-Benzimidazol-2-ylimino)-1-(morpholin-1-yl-
7.45 (m, 8H, Ar-H). Anal. calcd for C23H25N5O2S (435.54): methyl)indol-2(3H)-one (4b)
C, 63.43; H, 5.79; N, 16.08. Found: C, 63.12; H, 5.88; Yield 67%; m.p. 168-169oC; IR (KBr, cm−1): 3109, 3050
N, 16.35.
(C-H aromatic), 2951, 2854 (C-H aliphatic), 1732 (C=O),
1612 (C=N); H-NMR 200 MHz (DMSO-d6): 2.40-2.56
1
5-Bromo-3-(4,5-dihydro[1,3]thiazol-2-ylimino)-1-
{[4-(4-methoxyphenyl)piperazin-1-yl]methyl}indol-
2(3H)-one (3f)
(m, 4H, 2CH2 morpholinyl), 3.50-3.65 (m, 4H, 2CH2
morpholinyl), 4.39 (s, 2H, CH2), 6.93-7.70 (m, 8H, Ar-
H), 11.02 (br s, 1H, NH, D2O exchangeable). Anal. calcd
Yield 73%; m.p. 120-121oC; IR (KBr, cm−1): 3097, 3066 for C20H19N5O2 (361.40): C, 66.47; H, 5.30; N, 19.38.
(C-H aromatic), 2931, 2831 (C-H aliphatic), 1724 (C=O), Found: C, 66.28; H, 5.45; N, 19.55.
1
1612 (C=N); H-NMR 300 MHz (DMSO-d6): 2.62 (t,
2H, CH2 thiazoline), 2.68-2.71 (m, 4H, 2CH2 piperazin- 3-(1H-Benzimidazol-2-ylimino)-1-[(4-phenylpipera-
yl), 2.90-2.95 (m, 4H, 2CH2 piperazinyl), 3.43 (t, 2H, zin-1-yl)methyl]indol-2(3H)-one (4c)
CH2 thiazoline), 3.67 (s, 3H, OCH3), 4.44 (s, 2H, CH2), Yield 65%; m.p. 244-245oC; IR (KBr, cm−1): 3059 (C-H
6.58-7.45 (m, 7H, Ar-H); MS (EI) m/z (% rel. Int.): 515 aromatic), 2927, 2881 (CH aliphatic), 1735 (C=O), 1620
(1.71, M+2). Anal. calcd for C23H24BrN5O2S (514.44): (C=N); 1H-NMR 200 MHz (DMSO-d6): 2.65-2.80 (m, 4H,
C, 53.70; H, 4.65; N, 13.61. Found: C, 53.55; H, 4.70; 2CH2 piperazinyl), 3.05-3.18 (m, 4H, 2CH2 piperazin-
N, 13.88.
yl), 4.47 (s, 2H, CH2), 6.60-7.60 (m, 13H, Ar-H), 11.05
(br s, 1H, NH, D2O exchangeable); MS (EI) m/z (%

Novel Isatin-Thiazoline and Isatin-Benzimidazole Conjugates
1619
1
rel. Int.): 434 (54.03, M
-2). Anal. calcd for C26H24N6O 1616 (C=N); H-NMR 300 MHz (DMSO-d6): 2.80-2.92
(436.51): C, 71.54; H, 5.54; N, 19.25. Found: C, 71.25; (m, 4H, 2CH2 piperazinyl), 3.15-3.22 (m, 4H, 2CH2 pi-
H, 5.36; N, 19.15.
perazinyl), 3.77 (s, 3H, CH3), 4.56 (s, 2H, CH2), 6.83-7.62
(m, 8H, Ar-H), 11.10 (br s, 1H, NH, D2O exchangeable).
Anal. calcd for C21H22N6O (374.44): C, 67.36; H, 5.92;
N, 22.44. Found: C, 67.15; H, 5.85; N, 22.65.
3-(1H-Benzimidazol-2-ylimino)-1-{[4-(2-methoxy-
phenyl)-piperazin-1-yl]methyl}indol-2(3H)-one
(4d)
Yield 58%; m.p. 160-161oC; IR (KBr, cm−1): 3066 (C-H
aromatic), 2935, 2827 (C-H aliphatic), 1743 (C=O), 1608
Cytotoxic activity studies
Anticancer activity studies were carried out at Cairo
(C=N); 1H-NMR 300 MHz (DMSO-d6): 2.64-2.73 (m, 4H, University, National Cancer Institute, Cancer Biology
2CH2 piperazinyl), 2.96-3.04 (m, 4H, 2CH2 piperazinyl), Department, Pharmacology Unit. All chemicals and
3.79 (s, 3H, OCH3), 3.84 (s, 2H, CH2), 6.85-7.51 (m, 12H, solvents were purchased from Sigma and Aldrich.
Ar-H), 11.01 (br s, 1H, NH, D2O exchangeable). Anal. Eleven of the synthesized compounds 3a
calcd for C27H26N6O2 (466.53): C, 69.51; H, 5.62; N, 3g 4a 4b 4c 4d 4e and 4g were tested at concen-
trations between 1 and 10 mg/mL using the Sulfo-
, 3b, 3c, 3d,
,
,
,
,
,
18.01. Found: C, 69.32; H, 5.44; N, 18.28.
Rhodamine-B (SRB) assay for cytotoxic activity against
a human breast adenocarcinoma cell line (MCF7).
3-(1H-Benzimidazol-2-ylimino)-1-{[4-(4-methoxy-
phenyl)-piperazin-1-yl]methyl}indol-2(3H)-one (4e)
Yield 72%; m.p. 185-186oC; IR (KBr, cm−1): 3059 (C-H
aromatic), 2931, 2850 (C-H aliphatic), 1732 (C=O), 1616
Measurement of cytotoxicity by SRB assay
The cytotoxic activity of the newly synthesized com-
(C=N); 1H-NMR 200 MHz (DMSO-d6): 2.68-2.81 (m, 4H, pounds was measured in vitro using the SRB assay
2CH2 piperazinyl), 2.90-3.15 (m, 4H, 2CH2 piperazinyl), according to the method of Skehan et al. (1990). Cells
3.67 (s, 3H, OCH3), 4.47 (s, 2H, CH2), 6.79-7.59 (m, 12H, were plated in 96-multiwell microtiter plates (104 cells
Ar-H), 11.05 (br s, 1H, NH, D2O exchangeable). Anal. /well) for 24 h before treatment with the compound(s)
calcd for C27H26N6O2 (466.53): C, 69.51; H, 5.62; N, to allow attachment of cells to the walls of the plates.
18.01. Found: C, 69.64; H, 5.35; N, 18.15.
Test compounds were dissolved in DMSO and diluted
with saline to the appropriate volume. Different con-
centrations of the test compounds (0, 1, 2.5, 5, and 10
3-(1H-Benzimidazol-2-ylimino)-5-bromo-1-{[4-(4-
methoxyphenyl)piperazin-1-yl]methyl}indol-2(3H)-
one (4f)
µg/mL) were added to the cell monolayer. Triplicate
wells were prepared for each individual dose. Mono-
Yield 72%; m.p. 134-135oC; IR (KBr, cm−1): 3066 (C-H layer cells were incubated with the compound(s) for 48
aromatic), 2935, 2831 (C-H aliphatic), 1743 (C=O), h at 37oC in an atmosphere of 5% CO2. After 48 h,
1
1612 (C=N); H-NMR 300 MHz (DMSO-d6): 2.67-2.71 cells were fixed, washed and stained for 30 min with
(m, 4H, 2CH2 piperazinyl), 2.95-3.17 (m, 4H, 2CH2 pi- 0.4% (wt/vol) with SRB dissolved in 1% acetic acid.
perazinyl), 3.69 (s, 3H, OCH3), 4.45 (s, 2H, CH2), 6.71- Unbound dye was removed by four washes with 1%
7.72 (m, 11H, Ar-H), 11.15 (br s, 1H, NH, D2O exchange- acetic acid, and attached stain was recovered with
able). Anal. calcd for C27H25BrN6O2 (545.43): C, 59.46; Tris EDTA buffer. Color intensity was measured in an
H, 4.62; N, 15.41. Found: C, 59.65; H, 4.45; N, 15.65.
ELISA reader. The relationship between surviving
fraction and drug concentration was plotted to get the
survival curve. The concentration required for 50%
inhibition of cell viability (IC50) was calculated. The
3-(1H-Benzimidazol-2-ylimino)-1-[N,N-diphenyl-
amino-1-yl]-methyl}indol-2(3H)-one (4g)
Yield 75%; m.p. 174-175oC; IR (KBr, cm−1): 3062 (C-H results are given in Table I and presented graphically
aromatic), 2943, 2827 (C-H aliphatic), 1743 (C=O), 1608 in Table I. The survival curves for the most potent
(C=N); 1H-NMR 300 MHz (DMSO-d6): 4.38 (s, 2H, CH2), cytotoxic compounds, 4b and 4g, against MCF-7 cells
6.77-8.10 (m, 18H, Ar-H), 11.18 (br s, 1H, NH, D2O ex- are presented in Fig. 3.
changeable). Anal. calcd for C28H21N5O (443.50): C,
75.83; H, 4.77; N, 15.79. Found: C, 75.68; H, 4.52; N,
16.05.
RESULTS AND DISCUSSION
Compounds 2a
outlined in Scheme 1. The isatin Mannich bases 2a
were prepared by condensing isatin with formaline
-h, 3a-h and 4a-h were prepared as
3-(1H-Benzimidazol-2-ylimino)-1-[4-(4-methylpi-
perazin)-1-ylmethyl]indol-2(3H)-one (4h)
-h
Yield 62%; m.p. 166-167oC; IR (KBr, cm−1): 3059 (C-H and secondary amine of the amino compound. The 1H-
aromatic), 2924, 2854 (C-H aliphatic), 1732 (C=O), NMR spectra of the new Mannich bases 2d, 2e, 2f

1620
A. T. Taher et al.
Table I. In vitro cytotoxic activity of some of the synthe-
sized compounds against the human breast cancer cell
line (MCF-7)
Compound No.
MCF7 (IC50)a,b nM
3a
3b
3c
3d
3g
4a
4b
4c
59.67
47.81
38.22
50.97
42.62
34.50
24.95
64.14
26.36
42.86
22.59
05.46
4d
4e
4g
Dox.
aIC50: dose of the compound that inhibits tumor cell proli-
feration by 50%; bvalues are means of three experiments.
(compounds
isatin conjugates with thiazoline pharmacophore (com-
pounds ). The best activity was obtained with com-
pounds 4b 4d and 4g. On the other hand, from the
isatin-linked thiazoline Schiff base series, compound
demonstrated the characteristic singlet signals of CH2 3c displayed the highest anti-breast cancer activity
in the region of 4.48-4.55 ppm. Compounds 3a and with IC50 = 38.22 nM.
4a were synthesized by condensation of the Mannich
In summary, the synthesis and characterization of
base analogues 2a with either 2-aminothiazoline or two series of isatin conjugates, isatin-linked thiazoline
2-aminobenzimidazole by refluxing in ethanol in the Schiff base analogues 3a and isatin-linked benzimi-
presence of glacial acetic acid. IR spectra in both dazole Schiff base analogues 4a , has been described.
derivatives 3a and 4a showed one strong band All the tested compounds displayed antitumor activity
corresponding to carbonyl in the range of 1724-1743 against MCF-7. Some of these derivatives exhibited
cm−1. 1H-NMR spectra of 3a
exhibited two triplets of promising anti-breast cancer activity; in particular,
two CH2 protons of thiazoline in the range of 2.56-3.55 the compounds 3-(1 -benzimidazol-2-ylimino)-1-(mor-
and 3.43-4.36 ppm. Compounds 4a showed an ex- pholin-1-ylmethyl)indol-2(3H)-one (4b), 3-(1 -benzi-
changeable signal of NH proton around 11 ppm. All midazol-2-ylimino)-1-{[4-(2-methoxyphenyl)-piperazin-
other aromatic and aliphatic protons were observed at 1-yl]methyl}indol-2(3H)-one (4d), and 3-(1 -benzimi-
dazol-2-ylimino)-1-[N,N-diphenylamino-1-yl]-methyl}
4) showed higher cytotoxic activity than
Fig. 3. Effect of different concentrations of compounds 4b
and 4g on the viability of the MCF-7 cell line
3
,
-h
-h
-h
-h
-h
-h
-h
-h
H
-h
H
H
the expected regions.
indol-2(3H)-one (4g) emerged as promising compounds
and could therefore serve as lead compounds for fur-
ther modification to render them clinically useful agents.
Cytotoxic activity
The cytotoxicity of compounds 3a
, 3b, 3c, 3d, 3g,
4a 4b 4c 4d, 4e, and 4g was evaluated against a To further improve the efficacy and specificity for
,
,
,
human breast adenocarcinoma cell line (MCF-7). For cancer cell killing, additional structural modifications
comparison purposes, the cytotoxicity of doxorubicin, a of isatin conjugates are in progress.
standard antitumor drug, was evaluated under the
same conditions. The IC50 (the concentration required
for 50% inhibition of cell viability) was calculated for
each compound and the results are given in Table I.
ACKNOWLEDGEMENTS
We are grateful to Prof. Dr. Samia Shouman, Pro-
Our results revealed that the title compounds showed fessor of Pharmacology, and all members of the de-
inhibition of proliferation of MCF-7 human breast partment of Cancer Biology, National Cancer Institute,
cancer cells with IC50 range from 22.59-64.14 nM (Table Cairo, Egypt, for carrying out the cytotoxicity testing.
I). Generally, the isatin conjugates with benzimidazole

Novel Isatin-Thiazoline and Isatin-Benzimidazole Conjugates
1621
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