Anti-breast cancer activity of some novel 1,2-dihydropyridine, thiophene and thiazole derivatives

Article abstract of DOI:10.1016/j.ejmech.2010.10.024

A variety of novel 1,2-dihydropyridines 10-17, thiophenes 18-21 and thiazole 22 having a biologically active sulfone moiety were obtained via the reaction of 2-cyano-N′-[1-(4-(piperidin-1-ylsulfonyl) phenyl) ethylidene] acetohydrazide 3 with a variety of

Full text of DOI:10.1016/j.ejmech.2010.10.024

European Journal of Medicinal Chemistry 46 (2011) 137e141
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Anti-breast cancer activity of some novel 1,2-dihydropyridine, thiophene and
thiazole derivatives
,
Mansour S. Al-Said a, Mahmoud S. Bashandy b, Saleh I. Al-qasoumi c, Mostafa M. Ghorab a
*
a Medicinal, Aromatic and Poisonous Plants Research Center (MAPPRC), College of Pharmacy, King Saud University. P.O. Box 2457, Riyadh 11451. Saudi Arabia
b Department of Chemistry, Faculty of Science, Al-Azhar University, Nasr City, Cairo, Egypt
c Department of Pharmacognosy, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia
a r t i c l e i n f o
a b s t r a c t
Article history:
A variety of novel 1,2-dihydropyridines 10e17, thiophenes 18e21 and thiazole 22 having a biologically
active sulfone moiety were obtained via the reaction of 2-cyano-N0-[1-(4-(piperidin-1-ylsulfonyl)
phenyl) ethylidene] acetohydrazide 3 with a variety of reagents. The structures of the newly synthesized
compounds were confirmed by elemental analysis, IR, 1H NMR, 13C NMR and mass spectral data. All the
newly synthesized compounds were evaluated for their in-vitro anticancer activity against human breast
Received 30 August 2010
Received in revised form
21 October 2010
Accepted 22 October 2010
Available online 29 October 2010
cancer cell line (MCF7). Compounds 15 and 11 with IC50 values (20.6, 25.5
m
M) exhibited better activity
than Doxorubicin as a reference drug with IC50 value (32.02
as Doxorubicin as positive control.
mM), while compound 14 is nearly as active
Keywords:
Sulfones
1,2-Dihydropyridines
Thiophenes
Ó 2010 Elsevier Masson SAS. All rights reserved.
Thiazole
Anticancer activity
1. Introduction
2. Results and discussion
Pyridine derivatives are currently an important group of organic
compounds that are used as bactericides [1], fungicides [2], anti-
cancer agents [3e6]. In addition thiophene and thiazole derivatives
are known to possess interesting biological properties that show
anticancer [7e9], and antimicrobial activities [10e12]., in previous
work [13e22], we studied the anticancer activity of sulfonamides
containing compounds, where the nitrogen of eSO2NHegroup is
either free or substituted and these compounds proved a significant
activity, in this study, we employed the sulfonamide group but in
this case the nitrogen is emerged in piperidine ring to study the
effect of this cyclized form on activity, furthermore, different
substituents were introduced on the benzene ring such as cya-
noacetohydrazide and hydrazide bearing either pyridine, thiazole,
thiophene or benzothiophene moieties due to the well-docu-
mented anticancer activity of these moieties to study their SAR and
their anti-breast cancer activity.
2.1. Chemistry
The present work was to design and synthesis of some new 1,2-
dihydropyridine, thiophene and thiazole derivatives carrying bio-
logically active sulfone moiety expected to have anticancer activity.
2-Cyanoacetohydrazide 2 reacts with 1-[4-(piperidin-1-ylsulfonyl)
phenyl] ethanone 1 [23], to give hydrazide hydrazone derivative 3.
The structure of compound 3 was proved on the basis of analytical
and spectral data. Thus, IR spectrum showed bands at 3190 cmꢀ1
(NH), 2264 cmꢀ1 (C^N), 1683 cmꢀ1 (C]O) and 1384,1141 (SO2). 1H
NMR spectrum revealed the presence of a singlet at
corresponding to CH3 group, a singlet at 4.24 ppm for CH2 group,
two duplets at 7.71,8.02 ppm for the aromatic protons and
a singlet at 11.22 ppm for an (NH) group. Moreover, the 13C NMR
data exhibited the presence of 13.79 (CH3), 24.88 (CH2), 166.02
d 1.22 ppm
d
d
d
(C]O). Further evidence for the structure of 3 was obtained
through studying its chemical reactivity via some chemical
reagents. Thus, the reaction of compound 3 with cinnamonitrile
derivatives was studied. Thus, the reaction of 3 with ethyl a-cya-
nocinnamate furnished the pyridine derivatives 10e15, respec-
tively. The reaction took place through the intermediate formation
of 4e9. Compounds 10e15 were confirmed from their
* Corresponding author. Tel.: þ20 966 534292860; fax: þ20 966 014670560.
E-mail address: mmsghorab@yahoo.com (M.M. Ghorab).
0223-5234/$ e see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.10.024

138
M.S. Al-Said et al. / European Journal of Medicinal Chemistry 46 (2011) 137e141
microanalytical and spectral data. IR spectra showed bands at 3392,
3115 cmꢀ1 (NH2), 2218-2208 cmꢀ1 (C^N), and 1690-1670 cmꢀ1
(C]O) of ester group. 1H NMR spectrum of 10, revealed a triplet at
d
0.71 ppm for CH3 of ethyl group, a quartet at
d
4.34 ppm due to
CH2 of ethyl group and a singlet at
(Scheme 1).
d 7.11 ppm for NH2 group,
The reaction of 3 with either malononitrile or ethyl cyanoacetate
furnished the pyridine-2-one derivatives 16 and 17, respectively.
Analytical and spectral data are consistent with the proposed
structures. The reaction of 3 with either malononitrile or ethyl
cyanoacetate and elemental sulfur in the presence of triethylamine
gave the corresponding thiophene derivatives 18 and 19, respec-
tively. The reaction goes in parallel to the reported Gewald’s thio-
phene synthesis [24], (Scheme 2).
Similarly, the reaction of 3 with either cyclopentanone or
cyclohexanone and elemental sulfur yielded the corresponding
thiophene derivatives 20 and 21, respectively. Formation of 20 and
21 took place according the similar reported reactions of cyclo-
hexanone with methylene reagents and elemental sulfur [25]. On
the other hand, the reaction of 3 with elemental sulfur and phenyl
isothiocyanate afforded the thiazole derivative 22. The formation of
the latter product took place in parallel to the reported Hanzesch
reported reaction [26], (Scheme 3). Structures of compound 20, 21
and 22 were based on analytical and spectral data.
Scheme 2. Reagent and conditions: i, malononitrile or ethyl cyanoacetate/1,4-dioxane/
TEA; ii, malononitrile or ethyl cyanoacetate and sulfur/ethanol/TEA.
equipotent than the reference drug (IC50 ¼ 32.02
mM), considering
the SAR for the tested compounds, it was found that introduction of
the 1,2-dihydropyridine moiety resulted in an increase in the
activity when compared to the starting material 3 (IC50 ¼ 44.2
m
m
M)
M)
especially for the 4-hydroxyphenyl derivative 11 (IC50 ¼ 25.5
and the 4-chlorophenyl derivative 15 (IC50 ¼ 20.6
mM) which
were the most potent compounds in this screening and they
showed higher activity than doxorubicin, then compounds 10
2.2. In-vitro anticancer screening
(IC50 ¼ 37.9
m
m
M), 13 (IC50 ¼ 38.7
m
M), 14 (IC50 ¼ 34.4
mM) and 16
(IC50 ¼ 36.4
M) showed nearly similar activity than doxorubicin.
The newly synthesized compounds were evaluated for their in-
vitro cytotoxic activity against human breast cancer cell line
(MCF7). Doxorubicin HCl is one of the most effective anticancer
agents was used as a reference drug in this study. The relationship
between surviving fraction and drug concentration was plotted to
obtain the survival curve of breast cancer cell line (MCF7). The
response parameter calculated was the IC50 value, which corre-
sponds to the concentration required for 50% inhibition of cell
viability. (Table 1), shows the in-vitro cytotoxic activity of the newly
synthesized compounds where some compounds revealed signifi-
cant activity compared to Doxorubicin as a reference drug. It was
observed from the results obtained from Table 1 that most of the
tested compounds showed significant anti-breast cancer activity,
some of them were more potent and the others were found to
Introduction of thiophene, benzothiophene or thiazole moieties in
compounds 18-22 showed a decrease in the activity with IC50
values ranging from 49.3 to 87.8 mM.
These preliminary results of biologically screening of the tested
compounds give an idea about the possible importance of the 1,2-
dihydropyridine moiety in the compounds acting as anti-breast
cancer and give an encouraging framework in this field that may
lead to the discovery of potent anticancer agent.
3. Conclusion
The synthesized compounds were evaluated for their in-vitro
cytotoxicity against human breast cancer cell line (MCF7) and some
Scheme 1. Reagent and conditions: i, ethyl
a-cyanocinnamate derivatives/1,4-dioxane/TEA.

M.S. Al-Said et al. / European Journal of Medicinal Chemistry 46 (2011) 137e141
139
Switzerland,
d ppm) at 300 MHz, using TMS as internal standard.
Electron impact Mass Spectra were recorded on a Shimadzu Gc-Ms-
Qp 5000 instrument (Shimadzu, Tokyo, Japan). Elemental analysis
was performed on Carlo Erba 1108 Elemental Analyzer (Heraeus,
Hanau, Germany). All compounds were within ꢂ0.4% of the theo-
retical values.
4.1.1. 2-Cyano-N’-(1-(4-(piperidin-1-ylsulfonyl) phenyl) ethylidene)
acetohydr-azide (3)
To a solution of 2-cyanoacetohydrazide (1.0 g, 0.01 mol) in 1,4-
dioxane (20 mL), 1-[4-(piperidine-1-ylsulfonyl) phenyl] ethanone 1
(2.67 g, 0.01 mol) was added. The reaction mixture was heated
under reflux for 2 h. then left to cool. The solid product formed
upon pouring onto ice/water was collected by filtration and
recrystallized from ethanol to give 3. Yield, 94%; m.p. 215e217 ꢁC IR
(KBr, cmꢀ1): 3190 (NH), 3095 (CH arom.), 2983,2943,2852 (CH
aliph.), 2264 (C^N), 1683 (C]O), 1616 (C]N), 1384,1141 (SO2). 1H
NMR (DMSO-d6): 1.22 [s, 3H, CH3], 1.31e1.62 [m, 6H, 3CH2 cyclo],
2.94 [s, 4H, CH2eNeCH2 cyclo], 4.24 [s, 2H, CH2CN], 7.71, 8.02 [2d,
4H, Ar-H, AB system, J ¼ 7.6 Hz], 11.22 [s, 1H, NH]. 13C NMR (DMSO-
d6): 13.79, 22.80, 24.62, 24.88, 46.51 (2C), 116.04, 126.92, 127.12
(2C), 127.40 (2C), 141.81, 147.38, 159.61, 166.02 MS, m/z (%): 348
[Mþ] (8.4), 83 (100). Anal. Calcd. For C16H20N4O3S: C, 55.16; H, 5.79;
N, 16.08. Found: C, 55.50; H, 5.40; N, 16.30.
Scheme 3. Reagent and conditions: i, cyclopentanone and sulfur/ethanol/TEA; ii, cyclo-
hexanone and sulfur/ethanol/TEA; iii, phenyl isothiocyanate and sulfur/ethanol/TEA.
of the tested compounds were equipotent, while the others were
more potent compared with Doxorubicin. Compounds 15 and 11
showed a significant cytotoxic activity which was even higher than
that of reference drug Doxorubicin, while compound 14 is nearly as
active as Doxorubicin. Compound 19 showed no activity.
4.1.2. 6-Amino-2-oxo-1-(1-(4-(piperidin-1-ylsulfonly) phenyl)
ethylidene-amino)-4-substituted-1,2-dihydropyridine (10e15)
General procedure: To a solution of compound 3 (3.4 g, 0.01 mol)
in 1,4-dioxane (30 mL) containing triethylamine (1 mL), ethyl
benzalacetate derivatives (0.01 mol) was added. The reaction
mixture was heated under reflux for 4 h. then poured onto ice/
water and the solid obtained was recrystallized from ethanol to
give 10e15, respectively.
4. Experimental
4.1. Chemistry
Melting points (ꢁC, uncorrected) were determined in open
capillaries on a GallenKemp melting point apparatus (Sanyo Gal-
lenKemp, Southborough, UK). Precoated silica gel plates (silica gel
0.25 mm, 60 G F 254; Merck, Germany) were used for thin layer
chromatography, dichloromethane/methanol (9.5:0.5 mL) mixture
was used as a developing solvent system and the spots were
recorded in KBr discs using IR-470 Shimadzu spectrometer (Shi-
madzu, Tokyo, Japan). 1H NMR spectra (in DMSO-d6) were recorded
on Bruker Ac-300 ultra shield NMR spectrometer (Bruker, Flawil,
4.1.3. 2-Amino-5-cyano-6-[oxo-1-{1-[4-(piperidine-1-sulfonyl)
phenyl]ethylidene-amino}-4-p-tolyl-1,6-dihydropyridine-3-
carboxylic acid ethyl ester (10)
Yield, 87%; m.p. 115e117 ꢁC; IR (KBr, cmꢀ1): 3414, 3329 (NH2),
2980, 2939, 2852 (CH aliph.), 2214 (C^N), 1710, 1662 (2C]O), 1598
(C]N),1340,1166 (SO2). 1H NMR (DMSO-d6): 0.71 [t, 3H, CH3 ethyl],
1.22 [s, 3H, CH3],1.33e1.52 [m, 6H, 3CH2 cyclo], 2.30 [s, 3H, CH3 tolyl],
2.93 [s, 4H, CH2eNeCH2 cyclo], 4.34 [q, 2H, CH2 ethyl], 7.11 [s, 2H,
NH2 exchangeable with D2O], 7.21e8.10 [m, 8H, Ar-H]. 13C NMR
Table 1
In-vitro anticancer screening of the synthesized compounds against human breast cell line (MCF7).
Cpd. No.
Compound concentration (
10 ( M)
mM)
IC50 (mM)
m
25 (
m
M)
50 (
m
M)
100 (mM)
Surviving fraction (mean ꢂ SE)a
DOX
3
0.721 ꢂ 0.020
0.944 ꢂ 0.002
0.911 ꢂ 0.058
0.925 ꢂ 0.032
0.972 ꢂ 0.085
0.921 ꢂ 0.008
0.861 ꢂ 0.017
0.818 ꢂ 0.044
0.371 ꢂ 0.074
0.748 ꢂ 0.025
0.588 ꢂ 0.021
1.925 ꢂ 0.032
0.942 ꢂ 0.062
0.971 ꢂ 0.008
0.861 ꢂ 0.017
0.546 ꢂ 0.020
0.666 ꢂ 0.009
0.677 ꢂ 0.026
0.647 ꢂ 0.024
0.657 ꢂ 0.016
0.694 ꢂ 0.033
0.745 ꢂ 0.015
0.723 ꢂ 0.012
0.298 ꢂ 0.033
0.540 ꢂ 0.003
0.507 ꢂ 0.058
0.947 ꢂ 0.024
0.857 ꢂ 0.016
0.994 ꢂ 0.033
0.745 ꢂ 0.015
0.461 ꢂ 0.010
0.440 ꢂ 0.028
0.652 ꢂ 0.030
0.681 ꢂ 0.023
0.452 ꢂ 0.022
0.621 ꢂ 0.032
0.557 ꢂ 0.003
0.622 ꢂ 0.014
0.416 ꢂ 0.004
0.433 ꢂ 0.076
0.451 ꢂ 0.004
0.981 ꢂ 0.023
0.552 ꢂ 0.022
0.621 ꢂ 0.032
0.557 ꢂ 0.003
0.494 ꢂ 0.030
0.511 ꢂ 0.047
0.581 ꢂ 0.022
0.641 ꢂ 0.015
0.375 ꢂ 0.043
0.478 ꢂ 0.081
0.407 ꢂ 0.027
0.553 ꢂ 0.065
0.394 ꢂ 0.021
0.437 ꢂ 0.003
0.406 ꢂ 0.016
0.841 ꢂ 0.015
0.375 ꢂ 0.043
0.558 ꢂ 0.081
0.407 ꢂ 0.027
32.02
44.2
37.9
25.5
51.2
38.7
34.4
20.6
36.4
74.6
49.3
NA
10
11
12
13
14
15
16
17
18
19
20
21
22
75.3
87.8
60.1
NA: Compound having IC50 value > 100
m
M.
a
Each value is the mean of three experiments ꢂ standard error.

140
M.S. Al-Said et al. / European Journal of Medicinal Chemistry 46 (2011) 137e141
(DMSO-d6): 12.72, 13.88, 22.80, 24.64 (3C), 25.71, 46.51 (2C), 62.22,
80.24, 115.66, 117.02, 126.89, 127.61 (2C), 129.21 (2C), 130.92 (2C),
137.70 (2C), 139.74, 144.31, 154.82, 156.01, 161.94, 163.82, 165.81. MS
m/z (%): 561 [Mþ] (0.7), 172 (100). Anal. Calcd. for C29H31N5O5S: C,
62.02; H, 5.56; N, 12.47. Found: C, 62.40; H, 5.20; N, 12.10.
4.1.8. 2-Amino-4-(4-chlorophenyl)-5-cyano-6-oxo-1-{1-[4-
(piperidine-1-sulf-onyl)phenyl]ethylideneamino}-1,6-
dihydropyridine-3-carboxylic acid ethyl ester (15)
Yield, 84%; m.p. 133e135 ꢁC; IR [(KBr, cmꢀ1): 3431, 3327 (NH2),
2980, 2939, 2852 (CH aliph.), 2214 (C^N), 1700, 1676 (2C]O), 1595
(C]N), 1396, 1166 (SO2), 715 (C-Cl). 1H NMR (DMSO-d6): 1.10 [t, 3H,
CH3 ethyl], 1.21 [s, 3H, CH3], 1.31e1.62 [m, 6H, 3CH2 cyclo], 2.94 [s, 4H,
CH2eNeCH2 cyclo], 4.21 [q, 2H, CH2 ethyl], 7.22e8.01 [m, 10H, Ar-
HþNH2]. MS m/z(%): 581[Mþ](0.6),83(100). Anal.Calcd. forC28H28Cl
N5O5S: C, 57.78; H, 4.85; N, 12.03. Found: C, 57.40; H, 4.50; N, 12.40.
4.1.4. 2-Amino-5-cyano-4-(4-hydroxyphenyl)-6-oxo-1-{1-[4-
(piperidine-1-sulf-onyl)phenyl]ethylideneamino}-1,6-
dihydropyridine-3-carboxylic acid ethyl ester (11)
Yield: 73%; m.p. 85e87 ꢁC; IR (KBr, cmꢀ1): 3460 (OH), 3288,
3255 (NH2), 3072 (CH arom.), 2983, 2939, 2854 (CH aliph.) 2218
(C^N), 1732, 1714 (2C]O), 1610 (C]N), 1396, 1168 (SO2). 1H NMR
(DMSO-d6): 1.11 [t, 3H, CH3 ethyl], 1.21 [s, 3H, CH3], 1.51e1.60 [m,
6H, 3CH2 cyclo], 2.84 [s, 4H, CH2eNeCH2 cyclo], 4.31 [q, 2H, CH2
ethyl], 6.74 [s, 2H, NH2 exchangeable with D2O], 6.91e8.04 [m, 8H,
Ar-H], 9.77 [s, 1H, OH]. 13C NMR (DMSO-d6): 13.71, 17.54, 24.02 (3C),
25.42, 46.51 (2C), 61.83, 96.90, 115.21, 115.71 (2C), 125.92, 127.62
(2C), 129.43 (2C), 130.01 (2C), 139.71, 141.74, 156.52, 157.92, 162.58,
163.00, 165.91, 166.32. MS m/z (%): 563 [Mþ], (1.4), 42 (100). Anal.
Calcd. For C28H29N5O6S: C, 59.67; H, 5.19; N, 12.43. Found: C, 59.30;
H, 5.50; N, 12.10.
4.1.9. 4,6-Diamino-2-oxo-1-[1-(1-(4-(piperidin-1-ylsulfonyl)
phenyl)ethylidene-amino]-1,2-dihydropyridine-3-carbonitrile (16),
and 4-amino-6-hydroxy-2-oxo-1-[1-(4-(piperidin-1-ylsulfonyl)
phenyl)ethylideneamino)-1,2-dihydropyridine-3-carbonitrile (17)
Equimolecular amounts of compound 3 (3.4 g, 0.01 mol) and
either malononitrile (0.66 g, 0.01 mol) or ethyl cyanoacetate (1.13 g;
0.01 mol) in dioxane (20 mL) containing triethylamine (1 mL) was
heated under reflux for 6 h. The reaction mixture was left to cool
and evaporated under vacuum. The remaining product was tritu-
rated with ethanol and the obtained solid was recrystallized from
dioxane to give 16 and 17, respectively.
4.1.5. 2-Amino-5-cyano-4-(4-methoxyphenyl)-6-oxo-1-{1-[4-
(piperidine-1-sulf-onyl)phenyl]ethylideneamino}-1,6-
dihydropyridine-3-carboxylic acid ethyl ester (12)
16: Yield, 81%; m.p. 295e297 ꢁC; IR (KBr, cmꢀ1): 3412, 3334,
3248, 3178 (2NH2), 2918, 2860 (CH aliph.), 2214 (C^N), 1653 (C]
O), 1579 (C]N), 1380, 1170 (SO2). 1H NMR (DMSO-d6): 1.21 [s, 3H,
CH3], 1.31e1.52 [m, 6H, 3CH2 cyclo], 2.91 [s, 4H, CH2eNeCH2 cyclo],
4.10 [s, 3H, CH þ NH2], 7.01e8.02 [m, 6H, Ar-H þ NH2]. 13C NMR
(DMSO-d6): 14.22, 24.64, 25.82 (2C), 46.51 (2C), 70.90, 81.31, 116.30,
127.01 (2C), 127.42 (2C), 137.81, 141.22, 143.11, 161.50, 186.81, 197.33.
MS m/z (%): 414 [Mþ] (5.5), 47(100). Anal. Calcd. for C19H22N6O3S: C,
55.06; H, 5.35; N, 20.28. Found: C, 55.40; H, 5.00; N, 20.50.
17: Yield, 73%; m.p. 220e222 ꢁC; IR (KBr, cmꢀ1): 3412 (OH),
3367, 3190 (NH2), 3074 (CH arom.), 2943, 2843 (CH aliph.), 2264
(C^N), 1685 (C]O), 1360, 1168 (SO2). 1H NMR (DMSO-d6): 1.21 [s,
3H, CH3], 1.30e1.54 [m, 6H, 3CH2 cyclo], 2.80 [s, 4H, CH2eNeCH2
cyclo], 4.21 [s, 2H, NH2 exchangeable with D2O], 4.40 [s, 1H, CH],
7.71,8.02 [2d, 4H, Ar-H, AB system, J ¼ 7.1 Hz], 11.21 [s, 1H, OH]. 13C
NMR (DMSO-d6): 14.20, 24.62, 24.91 (2C), 46.51 (2C), 66.33, 84.64,
116.01, 127.12 (2C), 127.42 (2C), 141.86, 147.42, 151.31, 159.62, 166.01,
174.62. MS m/z (%): 415 [Mþ] (2.1), 78 (100). Anal. Calcd. for
C19H21N5O4S: C, 54.93; H, 5.09; N,16.86. Found: C, 54.60; H, 4.70; N,
16.50.
Yield, 89%; m.p. 153e155 ꢁC; IR (KBr, cmꢀ1): 3414, 3387 (NH2),
2939, 2852 (CH aliph.), 2216 (C^N), 1707, 1662 (2C]O), 1608 (C]
N),1340,1166 (SO2). 1H NMR (DMSO-d6): 1.21 [t, 3H, CH3 ethyl],1.30
[s, 3H, CH3], 1.51e1.72 [m, 6H, 3CH2 cyclo], 2.94 [s, 4H, CH2eNeCH2
cyclo], 3.83 [s, 3H, OCH3], 4.21 [q, 2H, CH2 ethyl], 6.60 [s, 2H, NH2
exchangeable with D2O], 7.02e8.01 [m, 8H, Ar-H]. MS m/z (%): 577
[Mþ] (1.2), 84 (100). Anal. Calcd. for C29H31N5O6S: C, 60.30; H, 5.41;
N, 12.12. Found: C, 60.00; H, 5.10; N, 12.50.
4.1.6. 2-Amino-5-cyano-4-(4-hydroxy-3-methoxyphenyl)-6-oxo-1-
{1-[4-(piperidine-1-sulfonyl)phenyl]ethylideneamino}-1,6-
dihydropyridine-3-carboxylic acid ethyl ester (13)
Yield, 67% m.p. 132e134 ꢁC; IR (KBr, cmꢀ1): 3275 (OH), 3199,
3113 (NH2), 2980, 2923, 2843 (CH aliph.), 2227 (C^N), 1718, 1689
(2C]O), 1577 (C]N), 1398, 1192 (SO2). 1H NMR (DMSO-d6): 1.31 [t,
3H, CH3 ethyl], 1.40 [s, 3H, CH3], 1.51e1.72 [m, 6H, 3CH2 cyclo], 2.84
[s, 4H, CH2eNeCH2 cyclo], 3.82 [s, 3H, OCH3], 4.21 [q, 2H, CH2
ethyl], 6.82 [s, 2H, NH2 exchangeable with D2O], 7.51e8.10 [m, 7H,
Ar-H], 11.22 [s, 1H, OH]. 13C NMR (DMSO-d6): 13.81, 14.18, 24.40,
24.92 (2C), 46.53 (2C), 55.51, 61.82, 96.61, 113.94, 115.03, 115.81,
116.61, 122.02; 126.87, 127.21 (2C), 128.90 (2C), 135.73, 141.71,
147.81, 153.02, 154.81, 159.62, 162.61, 164.22, 166.02. MS m/z (%):
593 [Mþ] (3.2), 84 (100). Anal. Calcd. for C29H31N5O7S: C, 58.67; H,
5.26; N, 11.80. Found: C, 58.30; H, 5.50; N, 12.10.
4.1.10. 3,5-Diamino-4-cyano-N`-(1-(4-(piperidin-1-ylsulfonyl)
phenyl)ethylidene] thiophene-2-carbohydrazide (18) and ethyl-2,4-
diamino-5-[2-(1-(4-piperidin-1-ylsulfonyl)phenyl)ethylidene)
hydrazinecarbonyl]thiophene-3-carboxylate (19)
To a solution of compound 3 (3.4 g, 0.01 mol) in absolute ethanol
(30 mL) containing triethylamine (1 mL) either malononitrile
(0.66 g, 0.01 mol), or ethyl cyanoacetate (1.13 g, 0.01 mol) together
with elemental sulfur (0.32 g, 0.01 mol) were added. The reaction
mixture was heated under reflux for 1h. then poured onto ice/water
and the solid obtained was recrystallized from dioxane to give 18
and 19, respectively.
4.1.7. 2-Amino-5-cyano-4-(4-dimethylaminophenyl)-6-oxo-1-{1-
[4-(piperidine-1-sulfonyl)phenyl]ethylideneamino}-1,6-
dihydropyridine-3-carboxylic acid ethyl ester (14)
Yield, 76%; m.p. 205e207 ꢁC; IR (KBr, cmꢀ1): 3197, 3111 (NH2),
2922, 2841 (CH aliph.), 2209 (C^N), 1707, 1689 (2C]O), 1610 (C]
N),1350,1166 (SO2). 1H NMR (DMSO-d6): 1.32 [t, 3H, CH3 ethyl], 1.41
[s, 3H, CH3], 1.51e1.62 [m, 6H, 3CH2 cyclo], 2.82 [s, 4H, CH2eNeCH2
cyclo], 3.31 [s, 6H, N(CH3)2], 4.20 [q, 2H, CH2 ethyl], 6.82 [s, 2H, NH2
exchangeable with D2O], 7.21e8.20 [m, 8H, Ar-H]. 13C NMR (DMSO-
d6): 13.81, 14.22, 24.21, 24.80 (2C), 39.88 (2C), 46.51 (2C), 61.41,
92.02, 111.61 (2C), 115.90, 117.41, 118.22, 126.90 (2C), 127.11 (2C),
127.32 (2C), 135.94, 141.71, 147.41, 151.22, 154.04, 159.61, 163.42,
166.01. MS m/z (%): 590 [Mþ] (0.6), 83(100). Anal. Calcd. for
C30H34N6O5S: C, 61.00; H, 5.80; N, 14.23. Found: C, 61.30; H, 5.75; N,
14.13.
18: Yield, 88%; m.p. >300 ꢁC; IR (KBr, cmꢀ1): 3446, 3385, 3346
(NH, NH2), 2939, 2852 (CH aliph.), 2204 (C^N), 1680 (C]O), 1610
(C]N), 1370, 1165 (SO2). 1H NMR (DMSO-d6): 1.10 [s, 3H, CH3],
1.31e1.52 [m, 6H, 3CH2 cyclo], 3.02 [s, 4H, CH2eNeCH2 cyclo], 6.41
[s, 2H, NH2 exchangeable with D2O], 7.01e8.02 [m, 6H, Ar-H þ NH2],
8.30 [s, 1H, NH]. MS m/z (%): 446 [Mþ] (1.9), 84 (100). Anal. Calcd.
for C19H22N6O3S2: C, 51.10; H, 4.97; N, 18.82. Found: C, 51.40; H,
4.60; N, 18.50.
19: Yield, 69% m.p. 230e232 ꢁC; IR (KBr, cmꢀ1): 3412, 3348, 3192
(NH, NH2), 3095 (CH arom.), 2943, 2852 (CH aliph), 1700, 1647
(2C]O), 1616 (C]N), 1382, 1166 (SO2). 1H NMR (DMSO-d6): 1.03 [t,

M.S. Al-Said et al. / European Journal of Medicinal Chemistry 46 (2011) 137e141
141
3H, CH3 ethyl], 1.22 [s, 3H, CH3], 1.31e1.50 [m, 6H, 3CH2 cyclo], 2.84
[s, 4H, CH2eNeCH2 cyclo], 4.02 [q, 2H, CH2 ethyl], 4.41 [s, 2H, NH2
exchangeable with D2O], 7.42 [s, 2H, NH2 exchangeable with D2O],
7.71e8.02 [2d, 4H, Ar-H, AB system J ¼ 7.8 Hz], 11.21 [s, 1H, NH]. 13C
NMR (DMSO-d6): 13.83, 14.21, 24.64, 24.91 (2C), 46.52 (2C), 56.02,
126.86, 127.10 (2C), 127.41 (2C), 135.92, 141.71, 141.90, 147.44, 151.21,
159.62, 166.02, 173.41. MS m/z (%): 493 [Mþ] (1.7), 83 (100). Anal.
Calcd. for C21H27N5O5S2: C, 51.10; H, 5.51; N, 14.19. Found: C, 51.40;
H, 5.20; N, 14.50.
screening was done by the pharmacology unit at the National
Cancer Institute, Cairo University.
Cells were plated in 96-multiwell plate (104 cells/well) for 24 h.
before treatment with the compound(s) to allow attachment of cell
to the wall of the plate. Tested compounds were dissolved in
dimethyl sulfoxide. Different concentration of the compound under
test (10, 25, 50 and 100 mM) were added to the cell monolayer.
Triplicate wells were prepared for each individual concentration.
Monolayer cells were incubated with the compound(s) for 48 h. at
37 ꢁC and in atmosphere of 5% CO2. After 48 h. cells were fixed,
washed and stained for 30 min with 0.4% (W/V) SRB dissolved in 1%
acetic acid. Excess unbound dye was removed by four washes with
1% acetic acid and attached stain was recovered with Tris-EDTA
buffer. Color intensity was measured in an ELISA reader. The rela-
tion between surviving fraction and drug concentration is plotted
to get the survival curve for breast tumor cell line after the specified
time [27]. The molar concentration required for 50% inhibition of
cell viability (IC50) was calculated and compared to the reference
drug Doxorubicin (CAS, 25316-40-9). The surviving fractions were
expressed as means ꢂ standard error and the results are given in
(Table 1).
4.1.11. 3-Amino-N0-[1-(4-(piperidin-1-ylsulfonyl)phenyl)
ethylidene]-5,6-dihy-dro-4H-cyclopenta[b]thiophene-2-
carbohydrazide (20), and 3-amino-N0-[1-(4-(piperidin-1-ylsulfonyl)
phenyl)ethylidene-4,5,6,7-tetrahydrobenzo[b]-thiophene-2-
carbohydrazide (21)
To a solution of compound 3 (3.4 g, 0.01 mol) in absolute ethanol
(30 mL) containing triethylamine (1 mL) either cyclopentanone or
cyclohexanone (0.01 mol) together with elemental sulfur (0.32 g,
0.01 mol) were added. The reaction mixture was refluxed for 2 h.
then poured onto ice/water and the solid obtained was recrystal-
lized from dioxane to give 20 and 21, respectively.
20: Yield, 66%; m.p. 222e224 ꢁC; IR (KBr, cmꢀ1): 3421, 3387, 3327
(NH, NH2), 2937, 2850 (CH aliph.), 1676 (C]O), 1325, 1165 (SO2). 1H
NMR (DMSO-d6): 1.11 [s, 3H, CH3], 1.30e1.52 [m, 6H, 3CH2 cyclo],
1.80e2.51 [m, 6H, 3CH2 cyclopenta], 2.94 [s, 4H, CH2eNeCH2 cyclo],
7.21e8.02 [m, 6H, Ar-H þ NH2], 9.41 [s,1H, NH].13C NMR (DMSO-d6):
17.91, 22.82, 24.62, 27.04 (2C), 28.31, 37.51, 46.52 (2C),126.74,126.82
(2C),127.41 (2C),135.22,138.91,142.03,142.31,147.81,166.92,193.90.
MS m/z (%): 446 [Mþ] (6.2), 83 (100). Anal. Calcd. for C21H26N4O3S2:
C, 56.48; H, 5.87; N, 12.55. Found: 56.80; H, 5.50; N, 12.20.
21: Yield, 87%; m.p. 232e234 ꢁC; IR (KBr, cmꢀ1): 3446, 3390,
3265 (NH, NH2), 2927, 2854 (CH aliph.), 1662 (C]O), 1608 (C]N)
1350, 1175 (SO2). 1H NMR (DMSO-d6): 0.81 [s, 3H, CH3], 1.51e2.02
[m, 6H, 3CH2 cyclo], 2.11e2.53 [m, 8H, 4CH2 benzothiophene], 2.90
[s, 4H, CH2eNeCH2 cyclo], 6.61 [br, 2H, NH2], 7.02e7.91 [m, 4H, Ar-
H], 10.91 [s, 1H, NH]. MS m/z (%): 460 [Mþ] (1.6), 84 (100). Anal.
Calcd. for C22H28N4O3S2: C, 57.37; H, 6.13; N, 12.16. Found: C, 57.60;
H, 5.80; N, 12.40.
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4.1.12. 4-Amino-3-phenyl-N0-(1-(4-(piperidin-1-ylsulfonyl)phenyl)
ethylidene)-2-thioxo-2,3-dihydrothiazole-5-carbohydrazide (22)
A mixture of compound 3 (3.4 g, 0.01 mol) and elemental sulfur
(0.32 g, 0.01 mol), phenyl isothiocyanate (1.3 g, 0.01 mol) in ethanol
(50 mL) containing triethylamine (1 mL) was refluxed for 3h. then left
to cool. The solid product formed upon pouring onto ice/water was
recrystallized from ethanol to give 22. Yield, 91%; m.p. 280e282 ꢁC; IR
(KBr, cmꢀ1): 3421, 3311, 3163 (NH, NH2), 3100 (CH arom.), 2940, 2895
2837 (CH aliph),1635 (C]O),1585 (C]N),1388,1165 (SO2),1280 (C]
S). 1H NMR (DMSO-d6): 1.20 [s, 3H, CH3], 1.31e1.62 [m, 6H, 3CH2
cyclo], 2.90 [s, 4H, CH2eNeCH2 cyclo], 7.01e8.03 [m, 11H, Ar-
H þ NH2], 10.82 [s, 1H, NH]. 13C NMR (DMSO-d6): 14.41, 22.82, 24.62
(2C), 46.53 (2C), 79.54, 125.01 (2C), 127.20, 127.52 (2C), 128.91 (2C),
130.11 (2C), 134.82, 135.74, 142.02, 154.41, 163.34, 164.92, 189.75. MS
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53.57; H, 4.89; N, 13.58. Found: C, 53.20; H, 4.50; N, 13.90.
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study. The cytotoxic activity was measured in-vitro for the newly
synthesized compounds using the Sulforhodamine B stain (SRB)
assay using the method of Skehan et al. [27]. The in-vitro anticancer