Design and synthesis of thiazol derivatives with biological evaluations as antitumor agents

Abstract

The synthesis of 2-cyanomethyl-thiazole-4-one (1) was obtained via the reaction of malononitrile with thioglycolic acid, then the formed product was directed toward the reaction with either acetoacetanlide or 4- chloro acetoacetanlide to produce compounds 2, 3. The latter compounds were reacted with either aromatic aldehyde or diazonium chloride derivatives to produce compounds 10, 11, 16-21 and 28-33. Finally compound 14 and 15 were obtained through the reaction of compound 10 with either malononitrile or ethylcyanoacetate. The newly synthesized compounds were evaluated for antitumor activity.

Full text

Research J. Pharm. and Tech. 8(5): May 2015
520
ISSN 0974-3618 (Print) www.rjptonline.org
0974-360X (Online)
RESEARCH ARTICLE
Design and synthesis of thiazol derivatives with biological evaluations as
antitumor agents
Karam A. El-Sharkawy1,2*, Mohammed M. A. El-Brrati,1Ibrahim A. Ghardaly,3and
Maksood Ali4
1Pharmaceutical Chemistry Department, Pharmacy Collage, Jazan University.
Jazan City, Kingdom of Saudi Arabia.
2Chemistry Department, Faculty of Biotechnology, October University for Modern Sciences and Arts(MSA),
El-Wahat Road, 6 October City, Egypt.
3Poison Control and Medical Forensic Chemistry Center, Jazan Health, Jazan City, Kingdom of Saudi Arabia.
4Pharmacognosy Department, Pharmacy Collage, Jazan University, Jazan City, Kingdom of Saudi Arabia.
*Corresponding Author E-mail: karamsyn@yahoo.com
ABSTRACT:
The synthesis of 2-cyanomethyl-thiazole-4-one (1) was obtained via the reaction of malononitrile with
thioglycolic acid, then the formed product was directed toward the reaction with either acetoacetanlide or 4-
chloro acetoacetanlide to produce compounds 2, 3. The latter compounds were reacted with either aromatic
aldehyde or diazonium chloride derivatives to produce compounds 10, 11, 16-21 and 28-33. Finally compound
14 and 15 were obtained through the reaction of compound 10 with either malononitrile or ethylcyanoacetate.
The newly synthesized compounds were evaluated for antitumor activity.
KEYWORDS: Thiazol, thiophene, pyridazine, antitumor activity.
1. INTRODUCTION:
Thiazoles and thiazolidones[1] derivatives exhibit an
interesting numbers of biological properties such as
antimicrobial[2-4], antiprotozoal[5,6], anti-
inflammatory, antiallergic[7,8], anticonvulsant[9],
cardiotonic[10], analgesic and antithermic[11] and
anticancer[12], among others. In this article we have
synthesized new thiazol-4-one derivatives purposely to
try to improve the antitumor activity against three
different cell lines. The structures of the newly
synthesized compounds were established using IR, NMR
and Mass spectrometry techniques.
Received on 29.01.2015 Modified on 21.02.2015
Accepted on 25.02.2015 © RJPT All right reserved
Research J. Pharm. and Tech. 8(5): May, 2015; Page 520-528
DOI: 10.5958/0974-360X.2015.00087.6
2. MATERIAL AND METHODS:
2.1. Experimental
All melting points were determined in open capillaries
and are uncorrected. Elemental analyses were performed
on a Yanaco CHNS Corder elemental analyzer (Japan).
IR spectra were measured using KBr discs on a Pye
Unicam SP-1000 spectrophotometer. 1H NMR spectra
were measured on a Varian EM 390-200 MHz
instrument in CD3SOCD3as solvent using TMS as
internal standard and chemical shifts were expressed as δ
ppm. Mass spectra were recorded on Kratos (75 eV) MS
equipment (Germany).
2.1.1. General procedures for the synthesis of 2-
cyanomethylenothiazol-4-one (1):
The titled compound 1obtained via the reaction of
malononitrile with thioglycolic acid in the presence of
glacial acetic acid according to procedures described
before[13].

Research J. Pharm. and Tech. 8(5): May 2015
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2.1.2. General procedures for the synthesis of:-
3-Methyl-4-phenylformamido-α-yl-2-(thiazolo-4'-one-
2'-yl) crotononitrile (2):
Equimolecular amounts of compound 1(1.4 g, 0.01 mol)
and acetoacetanilide (1.7 g, 0.01 mol), in presence of
catalytic amount of ammonium acetate were heated
under reflux at 140˚C for 1h. The reaction mixture was
then evaporated in vacuum. The remaining product was
triturated with ethanol and the solid product formed was
collected by filtration.
Compound 2: Pale yellow crystals from 1,4-dioxane,
yield: 83.4 % (2.5 g); mp: 60-62ºC. IR (KBr): υ/cm-1 =
3460-3380 (NH), 3057 (CH-aromatic), 2988 (CH3),
2875 (CH2), 2222 (CN), 1715, 1680 (2CO), 1658 (C=N).
1H NMR (DMSO-d6): δ = 2.94 (s, 3H, CH3), 5.34 (s, 2H,
CH2), 6.82 (s, 2H, thiazol CH2), 7.32-7.37 (m, 5H,
C6H5), 8.94 (s, 1H, NH, D2O-exchangeable). MS
(relative intensity) m/z: 299 (M+, 34%). Analysis for
C15H13N3O2S Calcd: C, 60.14; H, 4.73; N, 14.08, S,10.7
Found: C, 60.3; H, 4.57; N, 14.03, S, 10.88 %.
3-Methyl-4-(4'-Cl-phenyl) formamido-α-yl-2-(thiazole
- 4'-one-2'-yl) crotononitrile (3):
Equimolar amounts of compound 1(1.4 g, 0.01 mol) and
4-chloroacetoacetanilide (2.12 g, 0.01 mol), in the
presence of catalytic amount of ammonium acetate were
heated under reflux at 140˚C for 1.5 hr. The solid
product formed upon neutralization with ice/water
mixture containing few drops of hydrochloric acid was
collected by filtration.
Compound 3: Brown crystals from 1,4-dioxane , yield:
83.3 % (2.78 g); mp: 160-162 ºC. IR (KBr): υ/cm-1 =
3456-3374 (NH), 3060 (CH-aromatic), 2980 (CH3),
2865 (CH2), 2220 (CN), 1710, 1684 (2CO), 1640 (C=C).
1H NMR (DMSO-d6): δ = 2.93 (s, 3H, CH3), 5.3 (s, 2H,
CH2), 6.8 (s, 2H, thiazol CH2), 7.33-7.36 (m, 4H, C6H4),
9.24 (s, 1H, NH, D2O-exchangeable). MS (relative
intensity) m/z: 334 (M+, 21%). Analysis for
C15H12N3O2SCl Calcd: C, 53.93; H, 3.62; N, 12.63, S,
9.59 Found: C, 53.7; H, 3.8; N, 12.81, S, 9.72 %.
2.1.3. General procedure for the synthesis of
compounds:- 3-Methyl-4-phenylformamido-α-yl-2-
(thiazole-4'-one-2'-yl-)-5-phenyl-2,4-dienovaleronitrile
(10) and 3-Methyl-4-phenyl- formamido-α-yl-2-
(thiazole-4'-one-2'-yl)-5-(2'-hydroxy phenyl )-2,4-
dienovaleronitrile (11):
A mixture of compounds 2(2.9 g, 0.01 mol) and either
benzaldehyde (1.08 g, 0.01 mol) or salicylaldehyde (1.2
g, 0.01 mol), in dimethylformamide (40 ml) containing a
catalytic amount of piperidine (0.5 ml) was heated under
reflux for 2hrs. The reaction mixture was cooled at room
temperature and poured onto ice/water mixture
containing few drops of hydrochloric acid the solid
products, so formed, were filtered off and dried.
Compound 10: Pale brown crystals from 1,4-dioxane ,
yield: 77.3 % (2.99 g); mp: 98-100 ºC. IR (KBr): υ/cm-1
= 3455-3375 (NH), 3055 (CH-aromatic), 2975(CH3),
2863 (CH2), 2220 (CN), 1693, 1683 (2CO), 1658 (C=N),
1636 (C=C). 1H NMR (DMSO-d6): δ = 2.91 (s, 3H,
CH3), 6.83 (s, 2H, thiazol CH2), 7.01 (s,1H, CH=C),
7.32-7.39 (m, 10H, 2C6H5), 8.91 (s, 1H, NH, D2O-
exchangeable). MS (relative intensity) m/z: 387 (M+,
27%). Analysis for C22H17N3O2S Calcd: C, 68.16; H,
4.42; N, 10.88, S, 8.27 Found: C, 68.2; H, 4.6; N, 10.9,
S, 8.41 %.
Compound 11: Brown crystals from 1,4-dioxane , yield:
74.8 % (3.02 g); mp: 70-73 ºC. IR (KBr): υ/cm-1 = 3520-
3360 (OH, NH), 3050 (CH-aromatic), 2966 (CH3), 2861
(CH2), 2222 (CN), 1700, 1680 (2CO), 1660 (C=N), 1634
(C=C). 1H NMR (DMSO-d6): δ = 2.95 (s, 3H, CH3), 6.68
(s, 2H, thiazol CH2), 6.98 (s, 1H, CH=C), 7.36-7.42 (m,
9H, C6H5, C6H4), 8.89 (s, 1H, NH, D2O-exchangeable),
10.36 (s, 1H, OH). MS (relative intensity) m/z: 403 (M+,
20%). Analysis for C22H17N3O3S Calcd: C, 65.46; H,
4.24; N, 10.45, S, 7.94 Found: C, 65.6; H, 4.42; N,
10.59, S, 7.99 %.
2.1.4. General procedure for the synthesis of:- 1-
Amino-6-cyano-3-methyl-5-phenyl-4-
phenylformamido-α-yl-2-(thiazole-4'-one-2'-yl-)
benzene (14) and Ethyl-1-amino-6-cyano -3-methyl-5-
phenyl-4-phenylformamido-α-yl-2-(thiazole-4'-one-2'-
yl) benzoate (15):
To a solution of compound 10 (3.8 g, 0.01 mol) in
dimethylformamide (50 ml) containing a catalytic
amount of triethylamine (0.5 ml), either malononitrile
(0.66 g, 0.01 mol) or ethyl cyanoacetate (1.13 g, 0.01
mol) was added. The reaction mixture was heated under
reflux for 2hrs. Then it was poured onto ice/water
mixture containing few drops of hydrochloric acid. The
formed solid product was collected by filtration.
Compound 14: Yellow crystals from 1,4-dioxane , yield:
71.6 % (3.055 g); mp: 210-212 ºC. IR (KBr): υ/cm-1 =
3460-3345 (NH2, NH), 3058 (CH-aromatic), 2983
(CH3), 2869 (CH2), 2225 (CN), 1664 (C=N), 1638
(C=C). 1H NMR (DMSO-d6): δ = 3.11 (s, 3H, CH3), 5.48
(s, 2H, NH2, D2O-exchangeable), 6.83 (s, 2H, thiazol
CH2), 7.32-7.37 (m, 10H, 2C6H5), 8.81 (s, 1H, NH, D2O-
exchangeable). MS (relative intensity) m/z: 426 (M+,
16%). Analysis for C24H18N4O2S Calcd: C, 67.55; H,
4.25; N, 13.18, S, 7.51 Found: C, 67.76; H, 4.41; N,
13.29, S, 7.38 %.
Compound 15: Yellowish brown crystals from 1,4-
dioxane , yield: 67.5 % (3.197 g); mp: 140-142 ºC. IR
(KBr): υ/cm-1 = 3470-3325 (NH2, NH), 3060 (CH-
aromatic), 2979 (CH3), 2883 (CH2), 1705, 1690-1683
(3C=O), 1655 (C=N), 1637 (C=C). 1H NMR (DMSO-
d6): δ = 1.51 (t, 3H, CH3), 3.19 (s, 3H, CH3), 4.32 (q, 2H,

Research J. Pharm. and Tech. 8(5): May 2015
522
CH2), 5.36 (s, 2H, NH2, D2O-exchangeable), 6.81 (s, 2H,
thiazol CH2), 7.31-7.44 (m, 10H, 2C6H5), 8.89 (s, 1H,
NH, D2O-exchangeable). MS (relative intensity) m/z:
473 (M+, 23%). Analysis for C26H23N3O4S Calcd: C,
65.92; H, 4.89; N, 8.90, S, 6.76 Found: C, 65.81; H,
4.66; N, 8.72, S, 6.58 %.
2.1.5. General procedure for the synthesis of
compounds:- 3-Methyl-4-phenylformamido-α-yl-4-
phenylhydrazono-2-(thiazole-4'-one-2'-yl) crotono-
nitrile (16), 3-Methyl-4-(hydrazo-3-cyano-4',5',6',7'-
tetrahydrobenzo[b] thiophenophenylformamido-α-yl-
)-2-(thiozole-4'-one-2'-yl) crotononitrile (17) and 4-
[Ethyl 2–hydrazo (4',5',6',7'-tetrahydrobenzo[b]
thiopheno-3-carboxylate)phenylforma-mido-α-yl] - 3-
methyl-2-(thiozole-4'-one-2'-yl) crotononitrile (18):
To a cold solution of compound 2(2.9 g, 0.01 mol), in
ethanol (50 ml) containing sodium acetate (0.82 g, 0.01
mol), either benzene diazonium chroride (0.9 g, 0.01
mol), or 1-cyano-2-diazo-4,5,6,7- tetrahydrobenzo
[b]thiophene (1.7 g, 0.01 mol), or ethyl 2-diozo- 4,5,6,7-
tetrahydrobenzo [b]thiophencarboxylate (2.25 g, 0.01
mol) was added respectively with continuous stirring.
The reaction mixture was stirred at room temperature for
2hrs and the formed solid product was collected by
filtration.
Compound 16: Brown crystals from ethanol, yield: 81.7
% (3.299 g); mp: 175-177 ºC. IR (KBr): υ/cm-1 = 3480-
3410 (2NH), 3060 (CH-aromatic), 2977 (CH3), 2880
(CH2), 2227(CN), 1690, 1684 (2C=O), 1658 (C=N),
1634 (C=C). 1H NMR (DMSO-d6): δ = 2.94 (s, 3H,
CH3), 6.84 (s, 2H, thiazol CH2), 7.29-7.36 (m, 10H,
2C6H5), 8.44, 8.78 (2s, 2H, 2NH, D2O-exchangeable).
MS (relative intensity) m/z: 403 (M+, 24%). Analysis for
C21H17N5O2S Calcd: C, 62.47; H, 4.24; N, 17.41, S, 7.94
Found: C, 62.71; H, 4.16, N, 17.67, S, 7.68 %.
Compound 17: Dark brown crystals from 1,4-dioxane ,
yield: 71.5 % (3.496 g); mp: 233-235 ºC. IR (KBr):
υ/cm-1 = 3474-3370 (2NH), 3051 (CH-aromatic), 2984
(CH3), 2890 (CH2), 2226, 2223 (2CN), 1694, 1685
(2C=O), 1665 (C=N), 1643 (C=C). 1H NMR (DMSO-
d6): δ = 2.25-2.28 (m, 4H, 2CH2), 2.31-2.35 (m, 4H,
2CH2), 2.94 (s, 3H, CH3), 6.78 (s, 2H, thiazol CH2),
7.28-7.35 (m, 5H, C6H5), 8.41, 8.79 (2s, 2H, 2NH, D2O-
exchangeable). MS (relative intensity) m/z: 489 (M+,
31%). Analysis for C24H20N6O2S2Calcd: C, 58.95; H,
4.12; N, 17.26; S, 13.11 Found: C, 58.82; H, 4.06; N,
17.44; S, 13.34 %.
Compound 18: Brown crystals from 1,4-dioxane , yield:
59.3 % (3.176 g); mp: 170-172 ºC. IR (KBr): υ/cm-1 =
3478-3324 (2NH), 3048 (CH-aromatic), 2987 (CH3),
2890 (CH2), 2221 (CN), 1693, 1688, 1680 (3C=O), 1670
(C=N), 1642 (C=C). 1H NMR (DMSO-d6): δ = 1.14 (t,
3H, CH3), 2,24-2.27 (m, 4H, 2CH2), 2.34-2.37 (m, 4H,
2CH2), 2.90 (s, 3H, CH3), 4.25 (q, 2H, CH2), 6.45 (s, 2H,
thiazol CH2), 7.32-7.39 (m, 5H, C6H5), 8.51, 8.76 (2s,
2H, 2NH, D2O-exchangeable). MS (relative intensity)
m/z: 535 (M+, 21%). Analysis for C26H25N5O4S2Calcd:
C, 58.30; H, 4.66; N, 13.06; S, 11.94 Found: C, 58.03;
H, 4.70; N, 13.24; S, 11.74 %.
2.1.6. General procedure for the synthesis of
compounds:- 3-Methyl-4-(hydrazophenyl–4'–
chlorophenyl- formamido-α-yl)-2-(thiazole-4'-one-2'-
yl)- crotononitrile (19), 3-Methyl–4–[hydrazo–3'–
cyano–4-(4',5',6',7'-tetrahydro- benzo[b] thiophene–4-
chloro phenyl formamido–α-yl]–2-(thiazole–4'-one–
2'–yl) crotononitrile (20) and 4-[Ethyl -2–
hydrazo(4',5',6',7'-tetrahydrobenzo [b]- thiopheno-3-
carboxylate) 4-chlorophenyl- formamido-α-yl]-3-
methyl-2-(thiozole-4'-one-2'-yl) crotononitrile (21):
To a cold solution of compound 3(3.3 g, 0.01 mol) in
ethanol (50 ml) containing sodium acetate (0.82 g, 0.01
mol), either benzene diazonium chloride (0.9 g, 0.01
mol), or 1-cyano-2-diazo–4,5,6,7–
tetrahydrobenzo[b]thiophene (1.7 g, 0.01 mol), or ethyl
2-diazo–4,5,6,7–terahydrobenzo[b]thiophene
carboxylate (2.25 g, 0.01 mol) was added respectively
with continuous stirring. The reaction mixture was
stirred at room temperature for 3hrs the formed solid
product in each case was collected by filtration.
Compound 19: Deep brown crystals from 1,4-dioxane ,
yield: 66.1 % (2.897 g); mp: 222-225 ºC. IR (KBr):
υ/cm-1 = 3465-3340 (2NH), 3060 (CH-aromatic), 2980
(CH3), 2877 (CH2), 2224 (CN), 1691, 1683 (2C=O),
1660 (C=N), 1644 (C=C). 1H NMR (DMSO-d6): δ =
2.89 (s, 3H, CH3), 6.83 (s, 2H, thiazol CH2), 7.27-7.38
(m, 9H, C6H5, C6H4), 8.48, 8.82 (2s, 2H, 2NH, D2O-
exchangeable). MS (relative intensity) m/z: 438 (M+,
17%). Analysis for C21H16N5O2SCl Calcd: C, 57.55; H,
3.68; N, 16.04; S, 7.31 Found: C, 57.76; H, 3.97; N,
16.23; S, 7.12 %.
Compound 20: Brown crystals from acetic acid, yield:
63 % (3.298 g); mp: 158-159 ºC. IR (KBr): υ/cm-1 =
3458-3347 (2NH), 3050 (CH-aromatic), 2993 (CH3),
2866 (CH2), 2225, 2222 (2CN), 1692, 1677 (2C=O),
1660 (C=N), 1645 (C=C). 1H NMR (DMSO-d6): δ =
2.12-2.19 (m, 4H, 2CH2), 2.25-2.38 (m, 4H, 2CH2), 2.88
(s, 3H, CH3), 6.81 (s, 2H, thiazol CH2), 7.34-7.45 (d.d,
4H, C6H4), 8.39, 8.68 (2s, 2H, 2NH, D2O-
exchangeable). MS (relative intensity) m/z: 523 (M+,
27%). Analysis for C24H19N6O2SCl Calcd: C, 55.07; H,
3.65; N, 16.12; S, 12.25 Found: C, 55.28; H, 3.91; N,
16.34; S, 12.09 %.
Compound 21: Brown crystals from acetic acid, yield:
64.8 % (3.696 g); mp: 140-142 ºC. IR (KBr): υ/cm-1 =
3460-3330 (2NH), 3061 (CH-aromatic), 2984 (CH3),

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2870 (CH2), 2220 (CN), 1705, 1691, 1684, 1680
(3C=O), 1656 (C=N), 1641 (C=C). 1H NMR (DMSO-
d6): δ = 1.16 (t, 3H, CH3), 1.89-1.95 (m, 4H, 2CH2),
2.11-2.22 (m, 4H, 2CH2), 2.95 (s, 3H, CH3), 4.21 (q, 2H,
CH2), 6.73 (s, 2H, thiazol CH2), 7.37-7.54 (m, 5H,
C6H4), 8.33, 8.79 (2s, 2H, 2NH, D2O-exchangeable).
MS (relative intensity) m/z: 570 (M+, 29%). Analysis for
C26H24N5O4S2Cl Calcd: C, 54.74; H, 4.24; N, 12.33; S,
11.24 Found: C, 54.91; H, 4.37; N, 12.12; S, 11.03 %.
2.1.7. General procedure for the synthesis of
compounds:- 3-Phenylformamido-α-yl-6-oxo-4-
methyl-5-(thiazole-4'-one-2'-yl)-1-phenylpyridazine
(28), 3-Phenylforma-mido-α-yl-6-oxo-4-methyl-5-
(thiazole-4'-one-2'-yl)-1-(3'-cyano-4'-[4',5',6',7'-tetra-
hydrobenzo[b]- thiopheno)]pyridazine (29), 1-(Ethyl
4'-(4, 5,6,7–tetrahydrobenzo [b] thiopheno) –3'–
carboxylate–3–phenylformamido–α-yl–6–oxo–4-
methyl–5-(thiazole–4'-one-2'-yl -pyridazine (30).
A suspension of each compound of 16–18 (0.01 mol) in
sodium ethoxide solution (40 ml), was heated in a
boiling water bath for 2hrs, and then left to cool. The
solid product formed upon pouring onto ice/water
mixture containing few drops of hydrochloric acid was
collected by filtration.
Compound 28: Yellowish brown crystals from
dimethylformamide, yield: 69.1 % (2.796 g); mp: 216-
218 ºC. IR (KBr): υ/cm-1 = 3360-3315 (NH), 3054 (CH-
aromatic), 2978 (CH3), 2885 (CH2), 1705, 1695, 1682,
(3C=O), 1656 (C=N), 1638 (C=C). 1H NMR (DMSO-
d6): δ = 2.99 (s, 3H, CH3), 6.82 (s, 2H, thiazol CH2),
7.21-7.45 (m, 10H, 2C6H5), 8.49 (s, 1H, NH, D2O-
exchangeable). MS (relative intensity) m/z: 404 (M+,
29%). Analysis for C21H17N5O2S Calcd: C, 62.32; H,
3.98; N, 13.90, S, 7.92 Found: C, 62.61; H, 4.16, N,
13.73, S, 7.65 %.
Compound 29: Brown crystals from acetic acid, yield:
71.4 % (3.498 g); mp: 180-182 ºC. IR (KBr): υ/cm-1 =
3415-3384 (NH), 3053 (CH-aromatic), 2983 (CH3),
2897 (CH2), 2220 (CN), 1700, 1684 (2C=O), 1660
(C=N), 1648 (C=C). 1H NMR (DMSO-d6): δ = 2.05-2.14
(m, 4H, 2CH2), 2.26-2.37 (m, 4H, 2CH2), 2.88 (s, 3H,
CH3), 6.82 (s, 2H, thiazol CH2), 7.37-7.48 (m, 5H,
C6H5), 8.73 (s, 1H, NH, D2O-exchangeable). MS
(relative intensity) m/z: 489 (M+, 22%). Analysis for
C24H19N5O3S2Calcd: C, 58.84; H, 3.90; N, 14.35; S,
13.09 Found: C, 58.65; H, 4.08; N, 14.14; S, 13.32 %.
Compound 30: Brown crystals from acetic acid, yield:
78.2 % (4.198 g); mp: 165-167 ºC. IR (KBr): υ/cm-1 =
3438-3369 (NH), 3056 (CH-aromatic), 2974 (CH3),
2886 (CH2), 1712, 1693, 1681, 1674 (4C=O), 1655
(C=N), 1646 (C=C). 1H NMR (DMSO-d6): δ = 1.21 (t,
3H, CH3), 2,23-2.29 (m, 4H, 2CH2), 2.32-2.38 (m, 4H,
2CH2), 3.08 (s, 3H, CH3), 4.22 (q, 2H, CH2), 6.74 (s, 2H,
thiazol CH2), 7.36-7.44 (m, 5H, C6H5), 8.42 (s, 1H, NH,
D2O-exchangeable). MS (relative intensity) m/z: 536
(M+, 17%). Analysis for C26H24N4O5S2Calcd: C, 58.16;
H, 4.50; N, 10.48; S, 11.94 Found: C, 58.42; H, 4.71; N,
10.21; S, 11.76 %.
2.1.8. General procedure for the synthesis of
compounds:-3-(4'-Chlorophenyl- formamido -α-yl)-6-
oxo-4-methyl -5-(thiazole-4'-one-2'-yl)pyridazine (31),
1-[3'-Cyano-4'(4,5,6,7-tetra-hydrobenzo[b]thiopheno)]
-3-(4'-chloro phenylformamido-α-yl)-4-methyl-6-oxo-
5-(thiazole-4'-one-2'-yl)-pyridazine (32) and 1-[Ethyl
4-(4',5',6',7'- tetrahydro-benzo[b] thiopheno)-3-
carboxylate]-3(4'-chlorophenyl-formamido-α-yl)-4-
methyl-6-oxo-5-(thiazol -4'-one-2'–yl)pyridazine (33).
A suspension of each compound of 19–21 (0.01 mol) in
sodium ethoxide solution (40 ml), was heated in a
boiling water bath for 3hrs, and then left to cool. The
solid product formed upon pouring onto ice/water
mixture containing few drops of hydrochloric acid was
collected by filtration.
Compound 31: Brown crystals from dimethylformamide,
yield: 75.1 % (3.298 g); mp: 266-268 ºC. IR (KBr):
υ/cm-1 = 3463-3398 (NH), 3064 (CH-aromatic), 2975
(CH3), 2863 (CH2), 1703, 1688, 1677 (3C=O), 1655
(C=N), 1641 (C=C). 1H NMR (DMSO-d6): δ = 2.98 (s,
3H, CH3), 6.78 (s, 2H, thiazol CH2), 7.35-7.42 (m, 9H,
C6H5, C6H4), 8.81 (s, 1H, NH, D2O-exchangeable). MS
(relative intensity) m/z: 439 (M+, 14%). Analysis for
C21H15N4O3SCl Calcd: C, 57.43; H, 3.44; N, 12.81; S,
7.30 Found: C, 57.64; H, 3.22; N, 12.62; S, 7.13 %.
Compound 32: Brown crystals from acetic acid, yield:
66.3 % (3.477 g); mp: 221-223 ºC. IR (KBr): υ/cm-1 =
3455-3390 (NH), 3056 (CH-aromatic), 2990 (CH3),
2863 (CH2), 2225 (CN), 1698, 1681, 1673 (3C=O), 1654
(C=N), 1648 (C=C). 1H NMR (DMSO-d6): δ = 1.86-1.97
(m, 4H, 2CH2), 2.08-2.15 (m, 4H, 2CH2), 2.77 (s, 3H,
CH3), 6.89 (s, 2H, thiazol CH2), 7.29-7.41 (d.d, 4H,
C6H4), 8.91, (s, 1H, NH, D2O-exchangeable). MS
(relative intensity) m/z: 524 (M+, 33%). Analysis for
C24H18N5O3S2Cl Calcd: C, 54.97; H, 3.46; N, 13.41; S,
12.22 Found: C, 55.08; H, 3.64; N, 13.34; S, 12.42 %.
Compound 33: Brown crystals from dimethyformamide,
yield: 64.8 % (3.702 g); mp: 232-235 ºC. IR (KBr):
υ/cm-1 = 3451-3386 (NH), 3046 (CH-aromatic), 2994
(CH3), 2883 (CH2), 1710, 1693, 1685, 1678 (4C=O),
1660 (C=N), 1643 (C=C). 1H NMR (DMSO-d6): δ =
1.19 (t, 3H, CH3), 1.84-1.93 (m, 4H, 2CH2), 2.11-2.19
(m, 4H, 2CH2), 2.83 (s, 3H, CH3), 4.31 (q, 2H, CH2),
6.89 (s, 2H, thiazol CH2), 7.31-7.52 (d.d, 4H, C6H4),
8.95 (s, 1H, NH, D2O-exchangeable). MS (relative
intensity) m/z: 571 (M+, 16%). Analysis for
C26H23N4O5S2Cl Calcd: C, 54.65; H, 4.05; N, 9.84; S,
11.22 Found: C, 54.84; H, 4.31; N, 9.76; S, 11.01 %.

Research J. Pharm. and Tech. 8(5): May 2015
524
3. RESULTS AND DISCUSSION:
2-Cyanomethylenothiazol-4-one (1) obtained via the
reaction of malononitrile with thioglycolic acid in
presence of glacial acetic acid[13] compound 1
containing two methylene groups capable for many
chemical transformations. It has been reported that when
it coupled with either acetoacetanilide and p-
chloroacetoacetanilide in the presence of ammonium
acetate at 140 oC gave a single product with molecular
formula C15H13N3SO2.
Four different pairs isomeric structures were considered
2–9, structures 4,5,6,7and 8,9were ruled out on the
basis of IR spectrum of the reaction product which
showed the presence of one CN group stretching at -
2222 cm-1 and absence of any NH2group stretching
which might be expected to appear if structure 6,7is
considered and one OH group stretching if structure 8,9
is considered. Further the confirmations for structures 2,
3are obtained through studying their reactivity towards
chemical reagents. Thus, compound 2(as an example)
reacts with aryl aldehydes namely benzaldehyde or
salicyladehyde to give the arylidene derivatives 10 and
11.(Scheme.1)
The reaction of 10 with either malononitrile or ethyl
cyanoacetate gave the polyfunctionally substituted
benzene derivatives 14 and 15 respectively through the
intermediate formation of 12,13 followed by cyclization
and HCN elimination.
The reactivity of either 2or 3towards aryl and
heterocyclic diazonium salts was studied to form
hydrazones capable for cyclization. Thus, the reaction of
either compound 2or 3with benzenediazonium chloride
or 2-diazo-4, 5, 6, 7-tetrahydrobenzo[b]thiophene
derivatives gave the corresponding hydrazone
derivatives 16-21. The structures of the latter products
were based on analytical and spectral data. (Scheme.2)
Compounds 16-21 underwent ready cyclization when
heated in sodium ethoxide solution to give the
corresponding 6-oxo-pyridazine derivatives 28-33.
Formation of the latter products is assumed to take place
via the intermediate formation of the 6-iminopyridazine
derivatives 22-27 followed by hydrolysis of the imino
group into the oxo group through ammonia
liberation[14]. The IR and 1H NMR spectra of the
products are in-consistent with the assigned structures.
(Scheme.3)
H2CCN
CN
+CH2COOH
SH
AcOH N
S
O
NC
1
1+H3C NHAr
O O
Ar=Ph, p-Cholrophenyl
N
S
O
NC
ArHN CH3
O
2, Ar=Ph, 3, Ar= p-Cholrophenyl
N
S
O
N
S
O
N
S
O
NN
NH
O
NH
O
Ar
Ar
CH3CH3CH3
NH2
Ar
HO
4, Ar=Ph, 5, Ar= p-Cholrophenyl 6, Ar=Ph, 7, Ar= p-Cholrophenyl 8, Ar=Ph, 9, Ar= p-Cholrophenyl
2+ ArCHO Piperidine
N
S
O
NC
PhHN CH3
O
CHAr
Ar=Ph, 2-OH-C6H4
10, Ar=Ph, 11, Ar= 2-OH-C6H4
Scheme 1. Synthesis of compounds 1--3, 10, 11

Research J. Pharm. and Tech. 8(5): May 2015
525
10 +H2CCN
X
X=CN, COOEt
N
S
O
NC
PhHN CH3
O
PhHC CH CN
12, X=CN, 13, X=COOEt
X
2,3+ R-N=NCl
_
+_EtOH, 0-5C 0
NaOH
N
S
O
NC
PhHN CH3
O
N-NHR
-HCN
N
S
O
NH2
CH3
Ph
X
NHPhO 14, X=CN, 15, X=COOEt
Comp.No Ar R
16
17
18
19
20
21
Ph
Ph
Ph
4-Cl-C6H4
4-Cl-C6H4
4-Cl-C6H4
Ph
Ph
S
CN
S
CN
S
COOEt
S
COOEt
Scheme 2. Synthesis of compounds 14-21.

Research J. Pharm. and Tech. 8(5): May 2015
526
N
S
O
N
N
NH
CH3
R
NHArO
Comp.No Ar R
28
29
30
31
32
33
Ph
Ph
Ph
4-Cl-C6H4
4-Cl-C6H4
4-Cl-C6H4
Ph
Ph
S
CN
S
CN
S
COOEt
S
COOEt
16-21 NaOEt
Heat
N
S
O
N
N
NH
CH3
R
NHArO
-H2O
-NH3
22-27 28-33
Scheme 3. Synthesis of compounds 28-33.
3.1 Antitumor activity tests:
Reagents: Fetal bovine serum (FBS) and L-glutamine
from Gibco Invitrogen Co. (Scotland, UK). RPMI-1640
medium from Cambrex (New Jersey, USA). Dimethyl
sulfoxide (DMSO), doxorubicin, penicillin, streptomycin
and sulforhodamine B (SRB) were from Sigma
Chemical Co. (Saint Louis, USA).
Cell cultures: Three human tumor cell lines, MCF-7
(breast adenocarcinoma), NCI-H460 (non-small cell lung
cancer), and SF-268 (CNS cancer) were used. MCF-7
was obtained from the European Collection of Cell
Cultures (ECACC, Salisbury, UK) and NCI-H460, SF-
268 and normal fibroblast cells (WI 38) were kindly
provided by the National Cancer Institute (NCI, Cairo,
Egypt). They grow as monolayer and routinely
maintained in RPMI-1640 medium supplemented with
5% heat inactivated FBS, 2 mM glutamine and
antibiotics (penicillin 100 U/mL, streptomycin 100
µg/mL), at 37oC in a humidified atmosphere containing
5% CO2. Exponentially growing cells were obtained by
plating 1.5 X 105cells/mL for MCF-7 and SF-268 and
0.75 X 104cells/mL for NCI-H460, followed by 24 h of
incubation. The effect of the vehicle solvent (DMSO) on
the growth of these cell lines was evaluated in all the
experiments by exposing untreated control cells to the
maximum concentration (0.5%) of DMSO used in each
assay.
Tumor cell growth assay: The effects of 2–33 on the in
vitro growth of human tumor cell lines were evaluated
according to the procedure adopted by the National
Cancer Institute (NCI, USA) in the ‘In vitro Anticancer
Drug Discovery Screen’ that uses the protein-binding
dye sulforhodamine B to assess cell growth. Briefly,
exponentially, cells growing in 96-wellplates were then
exposed for 48 h to five serial concentrations of each
compound[15], starting from a maximum concentration
of 150 µM. Following this exposure period adherent
cells were fixed, washed, and stained. The bound stain
was solubilized and the absorbance was measured at 492
nm in a plate reader (Bio-Tek Instruments Inc., Power
wave XS, Wincoski, USA). For each test compound and
cell line, a dose–response curve was obtained and the
growth inhibition of 50% (GI50), corresponding to the
concentration of the compounds that inhibited 50% of
the net cell growth, was calculated as described
elsewhere[16]. Doxorubicin was used as a positive
control and it was tested in the same manner.

Research J. Pharm. and Tech. 8(5): May 2015
527
Table 1. Effect of compounds 2-33 on the growth of three human tumor cell lines
__________________________________________________________
Compound GI50 (mol L-1)
_____________________________
MCF-7 NCI-H460 SF-268 WI38
221.7 ± 6.9 19.9 ± 4.7 22.5 ± 5.3 na
321.2 ± 5.6 18.0 ± 3.9 17.7 ± 4.7 na
10 20.6 ± 3.8 17.1 ± 2.9 21.3 ±2.5 75.2±13.5
11 41.7 ± 7.9 32.2 ± 6.8 24 ± 7.8 na
14 39.2 ± 6.8 37.3 ± 6.4 35.9 ± 6.9 >100
15 46.2± 7.8 33.1 ± 8.4 28.4 ± 5.8 >100
16 38.0 ± 7.5 33.0 ± 8.9 32.5 ± 7.3 >100
17 11.8 ± 1.6 10.5 ± 2.2 11.4 ± 2.1 58.4 ± 9.6
18 22.0 ± 7.7 20.6 ± 6.3 22.4 ± 8.2 885.1 ± 12.3
19 44.4 ± 6.4 42.1 ± 8.7 38.3 ± 6.3 na
20 2.6 ± 0.07 2.4± 0.06 2.1 ± 0.08 53.7± 9.3
21 12.3 ±1.2 11.6 ± 0.9 9.7 ± 0.7 69.2 ± 11.5
28 21.3 ± 2.4 14.9 ± 2.8 12.6 ± 3.6 78.9 ± 12.9
29 13.1± 1.7 14.5 ±1.9 12.3 ± 2.2 80.2 ± 14.6
30 13.0 ± 0.9 12.0 ± 1.4 12.5 ± 1.8 77.6 ± 13.5
31 31.5 ± 7.5 29.2 ± 8.2 27.0 ± 6.1 83.8 ± 14.8
32 1.4 ± 0.03 1.6 ± 0.04 0.09 ± 0.02 42.7± 8.8
33 24.0 ± 1.8 24.0 ± 0.8 10.5 ± 1.1 49.7 ± 10.1
Doxorubicin 0.04 ±0.008 0.09±0.008 0.09±0.007 >100
Results are given in concentrations that were able to cause 50 % of cell growth inhibition (GI50) after a continuous exposure of 48 h and show
means ± SEM of three-independent experiments performed in duplicate.
The inhibitory effect of compounds 2-33 was evaluated
on the in vitro growth of three human tumor cell lines
representing different tumor types, namely, breast
adenocarcinoma (MCF-7), non-small cell lung cancer
(NCI-H460) and CNS cancer (SF-268) after a
continuous exposure for 48h. All of the tested
compounds were able to inhibit the growth of the tested
human tumor cell lines in a dose-dependent manner (data
not shown). The results indicated through Table 1
represented that hydrazothieno-thiazol derivatives 20
and thiazolopyridazine derivatives 32 showed the highest
inhibitory effect against all the three tumor cell lines
corresponding to reference standard material
(Doxorubicin), also compounds 17, 21, 29 and 30
showed the highest inhibitory effect against all the three
tumor cell lines corresponding to the remaining
synthesized compounds. On the other hand for
compounds 2, 3, 10, 18, 28 and 33 showed moderate
inhibitory effects against the three cancer cell lines. The
rest of the compounds 11, 14, 15, 16, 19 and 31 showed
a low growth inhibitory effect.
Comparing phenylformamido thiazole derivatives
derivatives 2and 3it was found that the two compounds
may be nearly the same effect regarding the presence of
p-chlorophenyl group in compound 3instead of phenyl
group in compound 2, on the other hand the dieno-
thiazole derivatives 10 and 11 it was found that
compound 10 with phenyl group it has higher effect than
that of compound 11 which it is containing p-
chlorophenyl group, comparing anilinothiazole
derivatives derivatives 14 and 15 it was clear that the
two compounds may be nearly the same effect although
they are containing two different groups, compound 14
containing CN group and compound 15 containing
COOEt.
Also comparing the hydrazothieno-thiazol derivatives
16-21 it was found that compound 20 with the Ar = 4-
Cl-C6H4, R= 3-Cyanotetrahydrobenzo[b]thieno group
showed the highest inhibitory effect among the six
compounds and then compound 17 which it has Ar = Ph
instead of 4-Cl-C6H4as in compound 20 has highest
inhibitory effect against all the three tumor cell lines
corresponding to the remaining synthesized group.
Finally comparing compounds 28-33 it is obvious that
compound 32 with the Ar = 4-Cl-C6H4, R= 3-
Cyanotetrahydrobenzo[b]thieno group showed the
highest inhibitory effect among the six compounds and
then compound 30 has highest inhibitory effect against
all the three tumor cell lines corresponding to the
remaining synthesized group which it has Ar = Ph
instead of 4-Cl-C6H4and R= 3- Ethylcaboxylate-
tetrahydrobenzo[b]thieno group instead of 3-
Cyanotetrahydrobenzo[b]thieno group as in compound
32.
4. CONCLUSIONS:
In this article the newly synthesized compounds thiazol-
4-one derivatives were investigated to detect their
antitumor activity against three different cell lines
corresponding to reference standard “doxorubicin”.
Among the newly synthesized products hydrazothieno-
thiazol derivatives 20 and thiazolopyridazine derivatives
32 showed the highest inhibitory effect against all the
three tumor cell lines, also compounds 17, 21, 29 and 30
showed the highest inhibitory effect against all the three
tumor cell lines corresponding to the remaining
synthesized compounds.

Research J. Pharm. and Tech. 8(5): May 2015
528
5. ACKNOWLEDGMENT:
The authors would like to thank the research group
working at the Medicinal Department at the National
Research Center, Dokki, Egypt, for recording the
pharmacological data of the synthesized products. More
over the Poison Control and Medical Forensic Chemistry
Center team. Jazan HealthJazan City, Kingdom of Saudi
Arabia, for recording the analytical and spectral data of
the newly synthesized compounds.
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