Quinazoline-tyrphostin as a new class of antitumor agents, molecular properties prediction, synthesis and biological testing

Article abstract of DOI:10.1016/j.ejmech.2012.03.044

A new series of substituted quinazolin-4-(3H)-one-tyrphostin derivatives was prepared and screened for their cytotoxic activity against three tumor cell lines, namely human breast cancer cell line (MCF-7), human cervical cancer cell line (HeLa) and human

Full text of DOI:10.1016/j.ejmech.2012.03.044

European Journal of Medicinal Chemistry 53 (2012) 133e140
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Quinazolineetyrphostin as a new class of antitumor agents, molecular properties
prediction, synthesis and biological testing
,
b
c
d
e
Ahmed M. Alafeefy a , Saleh I. Alqasoumi , Abdelkader E. Ashour , Vijay Masand , Nabila A. Al-Jaber ,
*
Taibi Ben Hadda f, Menshawy A. Mohamed a
,
g
a Department of Pharmaceutical Chemistry, College of Pharmacy, Salman Bin Abdulaziz University, P.O. Box 173, Alkharj 11942, Saudi Arabia
b Department of Pharmacognosy, College of Pharmacy, King Saud University, P.O. Box 2457, Riyadh 11451, Saudi Arabia
c Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, P.O. Box 2457, Riyadh 11451, Saudi Arabia
d Department of Chemistry, Vidya Bharati College, Amravati, Maharashtra 444602, India
e Women Students-Medical Studies and Sciences Sections, Chemistry Department, College of Science, King Saud University, P.O. Box 22452, Riyadh 11495, Saudi Arabia
f Laboratoire Chimie des Matériaux, Faculté des Sciences, Université Mohammed Premier, 60000 Oujda, Maroc
g Department of Organic Chemistry, Faculty of Pharmacy, Al-Azhar University, Cairo 11884, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
A new series of substituted quinazolin-4-(3H)-one-tyrphostin derivatives was prepared and screened for
their cytotoxic activity against three tumor cell lines, namely human breast cancer cell line (MCF-7),
human cervical cancer cell line (HeLa) and human hepatocellular liver carcinoma cell line (HepG2) using
the colorimetric MTT assay. Among the current series, 10 compounds exhibited remarkable in vitro
antiproliferative activity against the three tested cell lines with the IC50 values ranging from 0.009 to
0.015 mM. All the compounds showed suitable drug like characteristics according to Lipinski’s rule.
Ó 2012 Elsevier Masson SAS. All rights reserved.
Received 9 January 2012
Received in revised form
23 March 2012
Accepted 26 March 2012
Available online 5 April 2012
Keywords:
Quinazolin-4-(3H)-one
Tyrphostin
MTT assay
Cytotoxicity
POM analysis
1. Introduction
inhibitors were generated and the term tyrphostins was coined
for such compounds by Levitki and Mishani [6]. In spite of the
Cancer is one of the most challenging disorders in the world
and chemotherapy regimens led to unsatisfactory results. Future
improvements are likely to come from novel agents targeting
molecular pathways that promote tumor cell growth and
survival [1]. Protein tyrosine kinases (PTK) play important roles
in regulating most cellular functions including cell cycle, prolif-
eration, metabolism, survival, apoptosis, DNA repair and
response to the microenvironment [2,3]. The development of
PTK inhibitors has renovated the tactic to cancer therapy and is
likely to affect other fields of medicine. The first step in the
development of PTK inhibitors was from the family of benzene
malononitrile (Fig. 1) [4,5]. Since then, many hundreds of such
weaknesses among PTKs, one can develop small molecules that
antagonize the activity of certain PTKs and that display much
less toxicity than the currently available chemotherapeutic
agents [6].
On the other hand, quinazoline containing compounds form an
important class among heterocyclic pharmaceuticals and represent
an attractive scaffold for designing anticancer agents. Research on
quinazoline has led to the development and marketing of a new
series of antitumor agents [7].
We thought that incorporating the tyrphostin structural feature
on a quinazoline backbone might lead to interesting antitumor
agents. Thus, in the present work certain new quinazoline bearing
2-cyano-3-substituted acrylamide derivatives, as tyrphostin
analogs and/or their isosteres, were prepared and screened for their
antitumor activity against three tumor cell lines. We anticipated
that, these compounds would to inhibit tumor growth in a similar
manner to, or even better than other tyrosine kinase inhibitors.
* Corresponding author. Tel.: þ966 507069896; fax: þ966 15454573.
E-mail
addresses:
ahmed.alafeefy@yahoo.com,
aalafeefy@yahoo.com
(A.M. Alafeefy).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.03.044

134
A.M. Alafeefy et al. / European Journal of Medicinal Chemistry 53 (2012) 133e140
CN
HO
HO
CONH2
CN
O
C6H5
CN
N
Q
Tyrphostin AG 99
NH2
15
CN
Fig. 1.
NH2
CN
O
Q
C6H5
i
5
S
ii
H
N
iii
N
2. Results and discussion
CO2Et
4
iv
Q
NH2
16
NH2
O
2.1. Chemistry
17
NH2
CO2Et
S
H
The synthetic strategies adopted for the synthesis of the inter-
mediates and target compounds are depicted in Schemes 1 and 2. In
Scheme 1, the 2,6-dimethyl-3-(2-cyanoacetylamino)-quinazoline-
4(3H)-one 4 was prepared from the 3-aminoquinazoline derivative
N
Q
NH2
vi
O
v
18
4
S
S
S
O
NH
O
3
according to a reported procedure [8,9]. IR spectrum of
N
R
NH
Q
Q
compound 4 showed the presence of two carbonyl groups at 1689
and 1678 cmꢀ1, in addition to another band at 2225 cmꢀ1 due to the
cyano group. 1H NMR spectrum of compound 4 showed the pres-
ence of two singlets corresponding to 2CH3 groups, a singlet at
3.31 ppm for CH2 group, in addition to the aromatic protons and
a singlet at
8.10 ppm for an NH group. Moreover, the 13C NMR data
showed the presence of the three aliphatic, seven aromatic and two
carbonyl carbon atoms at the appropriate values. Further eluci-
H2N
vii
H2N
19
20: R: C6H5.
21:R=CH2C6H5.
CN
O
H
N
Q=
Q
N
d
N
O
HN
d
N
22: R=H
23: R=SO2NH2.
Reagents and conditions:
i: malononitrile, ethanol, triethylamine/heat.
ii: ethyl cyanoacetate, ethanol, triethylamine/heat.
R
d
dation for the structure of 4 was obtained through studying its
chemical reactivity with some chemical reagents. Thus, the reaction
of 4 with certain aromatic aldehydes gave the corresponding
iii: malononitrile, dioxane, triethylamine/elemental sulfur/heat.
iv: ethyl cyanoacetate, dioxane, triethylamine/elemental sulfur/heat.
v: ethanol, cyclohexanone, triethylamine/elemental sulfur/heat.
vi: ethanol, RNCS, triethylamine/elemental sulfur/heat.
vii: aryldiazonium chloride/ethanol/0-5 °C/stirring/1 hr.
O
O
O
NH2
Scheme 2.
OH
O
N
i
ii
NH2
N
N
3
benzylidene derivatives, tyrphostin 5e11. Analytical and spectral
data for the later compounds were in agreement with the proposed
structures. Generally, the IR spectra for these derivatives showed
the presence of the NH group at around 3280 cmꢀ1, cyano group at
around 2224 cmꢀ1 and two carbonyl groups at around 1680 and
1670 cmꢀ1. Whereas the NMR spectra showed the absence of the
active methylene protons in the 1H spectrum and the appearance of
2
1
iii
CN
CN
H
H
N
N
iv
Q
Q
R
O
O
4
5-11
the produced arylidene carbon at 107.0 d
value in the 13C spectrum.
5: R= H. 6: R= 4-CH3. 7: R= 4-Cl. 8: R= 4-NO2.
9: R= 3-OH. 10: R= 4-OH. 11: R= 3,4(OH)2.
v
On the other hand, the iminochromene derivative 12 was obtained
through reaction of 4 with salicylaldehyde. The reaction goes in
analogy with the reported literature [10]. The IR and 13C NMR
spectra of compound 12 were devoid of the cyano group and were
consistent with the proposed structure. The reaction of 4 or 5 with
either malononitrile or ethyl cyanoacetate gave the 2-pyridinone
derivatives 13e16. Analytical and spectral data are consistent
with the proposed structures where the amino, cyano and carbonyl
groups were common features of compounds 13e16 in the IR and
NMR spectra.
The substituted thiophene derivatives 17 and 18 were obtained
through reaction of compound 4 with either malononitrile or ethyl
cyanoacetate and elemental sulfur in the presence of triethylamine
as presented in Scheme 2. The reaction goes in parallel to the
reported Gewald’s thiophene synthesis [11]. Similarly, the reaction
of 4 with cyclohexanone and elemental sulfur gave the 4,5,6,7-
tetrahydrobenzothiophene derivative 19. The later reaction took
place in a similar way to the reported reactions of cyclohexanone
with methylene reagents and elemental sulfur [12].
HN
O
O
Q=
H
N
N
Q
12
O
N
CN
CN
O
Q
NH2
O
Q
NH2
vi
vii
4
N
N
OH
14
NH2
13
Reagents and conditions:
i: Ac2O/reflux.
ii: N2H4.H2O/reflux.
iii: ethyl cyanoacetate/reflux.
iv: aldehyde, dioxane, piperidine/heat.
v: salicylaldehyde, dioxane, piperidine/heat.
vi: malononitrile, dioxane, triethylamine/heat.
vii: ethyl cyanoacetate, dioxane, triethylamine/heat.
On the other hand the reaction of 4 with elemental sulfur and
phenyl or benzylisothiocyanate, gave the thiazole derivative 20, 21,
Scheme 2. Formation of the later products took place in parallel to
Scheme 1.

A.M. Alafeefy et al. / European Journal of Medicinal Chemistry 53 (2012) 133e140
135
a reported reaction [13]. The reaction of 4 with either benzene
diazonium chloride, 4-aminosulfonyl benzene diazonium chloride
gave the aryl hydrazone derivatives 22 and 23. Structures of all the
new compounds were based on analytical and spectral data and
were in accordance with the proposed structures.
affect the activity except for compound 15 which exhibited activity
better than that of the standard drug against MCF-7 and HepG2,
IC50 ¼ 0.008 “and” 0.005 mM, respectively, whereas compounds
12e14 and 16,17 showed considerable activity against the three cell
lines at somewhat higher IC50 values ranging from 0.010 to
0.026 mM. Conversely, cyclization with modification of the cyano
group led to less active and more selective compounds. Compounds
19e21 were active only against human breast cancer cell line MCF-
7 with IC50 0.010e0.018 mM compared to DOX IC50 (0.009 mM).
Compound 22 was active against MCF-7 and HepG2 while its
sulfonamide congener 23 was active against the three tested cell
lines mostly due to the impact of the well-known antitumor
sulfonamide. This moiety was shown to selectively concentrate in
tumor tissues in addition to their unique role in carbonic anhydrase
inhibition [14,15].
2.2. In vitro anticancer screening
The newly synthesized compounds 4e23 were initially screened
at the single concentration of 0.06 mM using the colorimetric MTT
[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] to
test their in vitro cytotoxicity against human breast cancer cell line
(MCF-7), cervix cancer (HeLa) and hepatocellular liver carcinoma
(HepG2). Doxorubicin was used as the reference drug in this study.
The cytotoxicity of the tested compounds was estimated in terms of
percent growth inhibition compared to untreated control cells. All
the compounds effected ꢁ70% inhibition and were retested by
serial dilution from 0.06 to 0.002 mM. The results were expressed
as IC50 (inhibitory concentration 50%), the concentration of the
compound which inhibits the tumor cell growth by 50% and the
data are presented in Table 1.
Except for compounds 5e11 almost all the active compounds
were devoid of the catechol moiety but had a considerable number
of hydrogen bond donor and acceptor groups comparable to the
model tyrphostin AG99. Such observation renders this family of
quinazoline containing tyrphostin analogs worthy for further
research and development.
Close inspection of cytotoxic data revealed that most of the
tested compounds showed remarkable antiproliferative activity
against the three tested cell lines. This observation could be
attributed to the synergistic effect that may result from combining
the quinazoline core with the typical tyrphostin structural feature,
2-cyano-3-substituted acrylamide (Fig. 1).
3. POM virtual screenings and molecular properties
calculations [16e23]
3.1. Molinspiration calculations
MiLogP (octanol/water partition coefficient) is calculated by the
methodology developed by Molinspiration as a sum of fragment-
based contributions and correction factors (Tables 2and 3). The
method is very robust and is able to process practically all organic
and most organometallic molecules. Total Polar Surface Area (TPSA)
is calculated as a sum of fragment contributions [16]. O- and N-
centered polar fragments are considered. PSA has been shown to be
a very good descriptor characterizing drug absorption, including
intestinal absorption, bioavailability, Caco-2 permeability and
bloodebrain barrier penetration. Prediction results of compounds
4e23 molecular properties (TPSA, GPCR ligand and ICM) are valued
(Tables 2and 3). Lipophilicity (logP) and polar surface area (PSA)
values are two important properties for the prediction of oral
bioavailability of drug molecules [16e23]. Therefore logP and PSA
values for compounds 4e23 we calculated using molinspiration
software programs and compared with the values obtained for
standard drug doxorubicin (DOX). For all the compounds, without
exception, the calculated clogP values were around ꢀ1.5e2.3 (<5),
which is the upper limit for the drugs to be able to penetrate
through bio-membranes according to Lipinski’s rule. Thus, these
compounds, 4e23, are expected to exhibit good bioavailability.
The lowest degree of lipophilicity among all the compounds was
exhibited by compounds 4e23, which is an indication for good
water solubility. The (PSA) is calculated from the surface areas that
are occupied by oxygen and nitrogen atoms and by hydrogen atoms
attached to them. Thus, the PSA is closely related to the hydrogen
bonding potential of a compound [16e23]. Molecules with PSA
values around of 160 Å2 or more are expected to exhibit poor
intestinal absorption. Table 3 shows that all the compounds are
within this limit. It has to be kept in mind that logP and PSA values
are the most important two features, although not sufficient
criteria for predicting oral absorption of a drug. It is worth to
mention that all the compounds have zero violation of the rule of 5.
Two or more violations suggest the probability of problems in
bioavailability. Drug likeness of compounds 4e23 is tabulated in
Table 3. Drug likeness may be defined as a complex balance of
various molecular properties and structure features which deter-
mine whether particular molecule is similar to the known drugs.
Most of the compounds showed efficacy against human breast
cancer cell line MCF-7 with IC50 values range of 0.06e0.002 mM
compared to DOX IC50 (0.009 mM) Table 1. Human hepatocellular
liver carcinoma cell line, HepG2 and human cervical cancer cells,
HeLa proved to be sensitive toward compounds 5e11, 13, 15, 17 and
23 with IC50 concentration range of 0.009e0.017 mM compared to
DOX (IC50 ¼ 0.008, 0.009 mM, respectively).
Evidently, compounds comprising the frank tyrphostin feature
(5e11), open chain, retained the highest activity against the three
cell lines with IC50 ranging from 0.009 to 0.019 mM. However,
rigidifying the structure and keeping the cyano group do not much
Table 1
In vitro antitumor activity of the designed quinazoline derivatives 4e23.
Compd. No.
IC50(mM)
MCF-7a
HeLab
HepG2c
4
5
6
7
8
9
0.040
0.009
0.010
0.012
0.012
0.015
0.014
0.008
0.017
0.010
0.026
0.008
0.017
0.010
0.016
0.018
0.013
0.010
0.019
0.009
0.009
0.030
0.013
0.009
0.012
0.015
0.014
0.013
0.008
0.038
0.015
0.045
0.005
0.021
0.015
0.023
0.031
0.035
0.026
0.079
0.012
0.008
0.055
0.014
0.012
0.009
0.010
0.014
0.013
0.008
0.023
0.017
0.041
0.010
0.052
0.015
0.022
0.022
0.027
0.032
0.012
0.009
0.009
10
11
12
13
14
15
16
17
18
19
20
21
22
23
DOX
a
Human breast cancer cell line (MCF-7).
Human cervical cancer cell line (HeLa).
Human hepatocellular liver carcinoma cell line (HepG2).
b
c

136
A.M. Alafeefy et al. / European Journal of Medicinal Chemistry 53 (2012) 133e140
Table 2
These properties, mainly hydrophobicity, electronic distribution,
Osiris calculations of compounds 4e23.
hydrogen bonding characteristics, molecule size and flexibility and
presence of various pharmacophoric features influence the
behavior of molecule in a living organism, including bioavailability,
transport properties, affinity to proteins, reactivity, toxicity, meta-
bolic stability and many others. Activity of all the 20 compounds
and standard drug were rigorously analyzed under four criteria of
known successful drug activity in the areas of GPCR ligand activity,
ion channel modulation, kinase inhibition activity, and nuclear
receptor ligand activity. Results are shown for all compounds in
Table 3 by means of numerical assignment. Likewise all the
compounds have consistent negative values in all categories and
numerical values comparable to that of standard drug used for
comparison. Therefore it is readily seen that all the compounds are
expected to have close similar activity to standard drug used based
upon these four rigorous criteria (GPCR ligand, ion channel
modulator, kinase inhibitor and nuclear receptor ligand).
Compd. Toxicity Risksa
Bioavailability and Drug Scoreb
MUT TUM IRRIT RE MW CLP
S
DL
D-S
4
256
344
358
377
389
360
360
376
360
322
323
408
455
354
401
368
423
437
360
439
0.37 ꢀ2.81
1.99 ꢀ4.2
2.30 ꢀ4.55
2.60 ꢀ4.94
ꢀ4.62 0.22
ꢀ1.65 0.45
ꢀ1.98 0.32
0.13 0.40
5
6
7
8
1.86 ꢀ4.66 ꢀ10.80 0.21
9
2.30 ꢀ4.55
1.69 ꢀ3.91
1.39 ꢀ3.61
0.72 ꢀ3.08
ꢀ1.51 ꢀ3.89
ꢀ0.98 ꢀ3.81
0.66 ꢀ5.11
0.64 ꢀ4.86
0.27 ꢀ4.65
0.87 ꢀ4.32
2.22 ꢀ4.85
1.78 ꢀ6.43
1.90 ꢀ5.91
2.10 ꢀ3.67
0.71 ꢀ3.57
ꢀ0.79 0.47
ꢀ0.41 0.27
0.27 0.63
10
11
12
13
14
15
16
17
18
19
20
21
22
23
2.97 0.84
ꢀ2.47 0.45
ꢀ2.35 0.45
ꢀ0.19 0.37
ꢀ3.72 0.25
ꢀ1.72 0.42
ꢀ1.30 0-44
ꢀ2.78 0.36
4.64 0.50
3.2. Osiris calculations
Structure based design is now fairly routine but many potential
drugs fail to reach the clinic because of ADME-Tox liabilities. One
very important class of enzymes, responsible for many ADMET
problems, is the cytochromes P450. Inhibition of these or produc-
tion of unwanted metabolites can result in many adverse drug
reactions. With our recent work on the drug design by combination
of various pharmacophoric sites using heterocyclic structure, it is
now possible to predict activity and/or inhibition with increasing
success in two targets (bacteria and HIV). This was done using
5.36 0.53
a
combined electronic/structure docking procedure and an
ꢀ1.07 0.24
1.44 0.33
example will be given here. The remarkably well behaved muta-
genicity of diverse synthetic molecules classified in data base of
Celeron Company of Swiss can be used to quantify the role played
by various organic groups in promoting or interfering with the way
a drug can associate with DNA. The Osiris calculations are tabulated
in Table 2. Toxicity risks (mutagenicity, tumorogenicity, irritation,
reproduction) and physicochemical properties (miLogP, solubility,
drug likeness and drug score) of compounds 4e23 were calculated
DOX
545
ꢀ0.95 ꢀ3.72
7.38 0.61
a
MUT: mutagenic; TUM: tumorigenic; IRRIT: irritant; RE: reproductive effective.
CLP: cLogP, S: Solubility, DL: Drug likeness, DS: Drug Score.
b
Table 3
Drug likeness calculation of compounds 4e23 by using molinspiration.
Compd.
Molinspiration calculationsa
Calculation of Bioactivity Scoresb
cLogP
0.103
TPSA
OHeNH Interract
Volume
GPCRL
ICM
KI
NRL
PI
EI
4
5
6
7
8
9
88
88
88
88
134
88
108
128
101
133
127
131
133
140
142
90
1
1
1
1
1
1
2
3
2
4
3
2
2
5
5
3
3
3
2
4
7
226
308
325
322
332
325
316
324
312
277
274
354
398
294
339
323
351
368
317
359
465
ꢀ0.49
ꢀ0.46
ꢀ0.45
ꢀ0.45
ꢀ0.55
ꢀ0.44
ꢀ0.41
ꢀ0.41
ꢀ0.66
ꢀ0.04
ꢀ0.06
ꢀ0.12
ꢀ0.19
ꢀ0.37
ꢀ0.44
ꢀ0.25
ꢀ0.55
ꢀ0.46
ꢀ0.45
ꢀ0.47
0.02
ꢀ1.08
ꢀ0.71
ꢀ0.68
ꢀ0.69
ꢀ0.68
ꢀ0.68
ꢀ0.64
ꢀ0.64
ꢀ1.10
ꢀ0.18
ꢀ0.16
ꢀ0.29
ꢀ0.25
ꢀ1.11
ꢀ1.03
ꢀ0.83
ꢀ1.21
ꢀ1.17
ꢀ0.89
ꢀ0.86
ꢀ0.16
ꢀ0.57
ꢀ0.29
ꢀ0.28
ꢀ0.30
ꢀ0.39
ꢀ0.28
ꢀ0.24
ꢀ0.25
ꢀ0.71
0.14
ꢀ1.36
ꢀ0.71
ꢀ0.68
ꢀ0.70
ꢀ0.73
ꢀ0.67
ꢀ0.55
ꢀ0.56
ꢀ1.19
ꢀ0.61
ꢀ0.35
0.45
ꢀ0.37
ꢀ1.28
ꢀ1.21
ꢀ1.05
ꢀ1.36
ꢀ1.18
ꢀ1.26
ꢀ1.29
0.14
ꢀ0.99
ꢀ0.65
ꢀ0.63
ꢀ0.67
ꢀ0.72
ꢀ0.62
ꢀ0.62
ꢀ0.62
ꢀ1.07
ꢀ0.34
ꢀ0.32
ꢀ0.32
ꢀ0.32
ꢀ0.89
ꢀ0.90
ꢀ0.68
ꢀ0.90
ꢀ0.74
ꢀ0.78
ꢀ0.55
0.34
ꢀ0.66
ꢀ0.45
ꢀ0.44
ꢀ0.47
ꢀ0.51
ꢀ0.44
ꢀ0.39
ꢀ0.38
ꢀ0.44
ꢀ0.07
ꢀ0.06
ꢀ0.12
ꢀ0.18
ꢀ0.66
ꢀ0.69
ꢀ0.42
ꢀ0.62
ꢀ0.48
ꢀ0.54
ꢀ0.37
0.51
2.719
3.17
3.40
2.68
3.14
2.24
1.75
1.26
0.60
0.90
2.59
3.05
0.957
1.41
2.84
2.27
10
11
12
13
14
15
16
17
18
19
20
21
22
23
DOXc
0.11
0.15
ꢀ0.00
ꢀ0.36
ꢀ0.52
ꢀ0.28
ꢀ0.55
ꢀ0.45
ꢀ0.58
ꢀ0.53
ꢀ0.22
95
95
112
172
206
2059
3.36
2.05
ꢀ0.657
a
TPSA: Total polar surface area.
b
c
GPCRL: GPCR ligand; ICM: Ion channel modulator; KI: Kinase inhibitor; NRL: Nuclear receptor ligand; PI: Protease inhibitor; EI: Enzyme inhibitor.
DOX: Doxorubicin.

A.M. Alafeefy et al. / European Journal of Medicinal Chemistry 53 (2012) 133e140
137
by the methodology developed by Osiris. The toxicity risk predictor
locates fragments within a molecule, which indicate a potential
toxicity risk. Toxicity risk alerts are an indication that the drawn
structure may be harmful concerning the risk category specified.
From the data evaluated in Table 2 it is obvious that, 15 of 20
structures are supposed to be non-mutagenic, non-irritating with
no reproductive effects when run through the mutagenicity
assessment system in comparison with the standard drug. The logP
value of a compound, which is the logarithm of its partition coef-
ficient between n-octanol and water, is a well-established measure
of the compound’s hydrophilicity. Low hydrophilicities and there-
fore high logP values may cause poor absorption or permeation. It
has been shown that for compounds to have a reasonable proba-
bility of good absorption, their logP value must not be greater than
5.0. On this basis, all the compounds 4e23 possessed logP values in
the acceptable range. Along with this, compound 11, which showed
good antitumor screening results (IC50 ¼ 0.008 mM against the
three tested cell lines), is having the same logP value as compared
to other compounds in the series.
spectrometer (Shimadzu, Tokyo, Japan). 1H NMR spectra (DMSO-d6)
were recorded on Bruker AC-300 Ultra Shield NMR spectrometer
(Bruker, Flawil, Switzerland, d
ppm) at 300 MHz for 1H and 75 MHz
for 13C, using TMS as internal standard and peak multiplicities are
designated as follows: s, singlet; d, doublet; t, triplet; m, multiplet.
Electron Impact Mass Spectra were recorded on a Shimadzu GC-
MS-QP 5000 instrument (Shimadzu, Tokyo, Japan). Elemental
analyses were performed, on Carlo Erba 1108 Elemental Analyzer
(Heraeus, Hanau, Germany), at the Microanalytical Unit, Faculty of
Science, Cairo University, Cairo, Egypt, and the found results were
within ꢃ0.4% of the theoretical values.
4.1.1. 2-Cyano-N-(2,6-dimethyl-4-oxoquinazolin-3(4H)-yl)
acetamide 4
This compound was prepared according to a reported procedure
[8,9].
4: Yield: 88%, mp: 185e187 ꢂC (toluene). IR (KBr, cmꢀ1): 3281
(NH), 3089 (CH arom.), 2989, 2886 (CH aliph.), 2225 (C^N), 1689,
1678 (2C]O), 1624 (C]N). 1H NMR:
d 1.30 (s, 3H, CH3), 2.36 (s, 3H,
CH3), 3.31 (s, 2H, CH2CN), 7.44 (d, J ¼ 7.0 Hz, 1H, AreH), 7.64 (d,
J ¼ 7.0 Hz, 1H, AreH), 7.82 (s, 1H, AreH), 8.10 (bs, 1H, NH, D2O
3.3. The aqueous solubility
exchangeable). 13C NMR:
d 19.0 (CH3), 24.2 (CH3), 28.0 (CH2), 115.1
(C^N), 124.1, 126.4, 130.4, 134.2, 135.2, 144.8, 158.8 (AreC), 164.6,
168.2 (2C]O). MS m/z (Rel. Int.): 257 (Mþ þ 1, 56), 105 (100).
Analysis for C13H12N4O2.
The aqueous solubility of a compound significantly affects its
absorption and distribution characteristics. Typically, a low solu-
bility goes along with a bad absorption and therefore the general
aim is to avoid poorly soluble compounds. Our estimated logS value
is a unit stripped logarithm (base 10) of a compound’s solubility
measured in mol/liter. There are more than 80% of the drugs on the
market have a (estimated) logS value greater than ꢀ4. In case of
compounds 4e23, values of logS are around ꢀ4. Further, Table 3
shows drug likeness of compounds 4e23 which is in the compa-
rable zone with that of standard drug used for comparison. We have
calculated overall drug score (DS) for the compounds 4e23 and
compared with that of standard drug doxorubicin, Table 2. The drug
score combines drug likeness, miLogP, logS, molecular weight and
toxicity risks in one handy value than may be used to judge the
compound’s overall potential to qualify for a drug. This value is
calculated by multiplying contributions of the individual properties
with the equation (1):
4.1.2. 2-Cyano-N-(2,6-dimethyl-4-oxoquinazolin-3(4H)-yl)-3-(4-
substituted phenyl) acrylamide 5e11and N-(2,6-dimethyl-4-
oxoquinazolin-3(4H)-yl)-2-imino-2H-chromene-3-carboxamide 12
General procedure: equimolecular mixture of
4 (2.56 g,
0.01 mol) and the selected aldehyde (0.01 mol), in 1,4-dioxane
(20 mL) containing piperidine (0.5 mL) was heated under reflux
for 3 h. The reaction mixture was left to cool then poured onto ice/
water containing few drops of hydrochloric acid and the formed
solid product was collected by filtration.
5: Yield: 82%, mp: 198e200 ꢂC (dioxane). IR (KBr, cmꢀ1): 3283
(NH), 3092 (CH arom.), 2985, 2880 (CH aliph.), 2223 (C^N), 1688,
1678 (2C]O), 1629 (C]N). 1H NMR:
d 1.60 (s, 3H, CH3), 2.35 (s, 3H,
CH3), 7.34 (bs, 1H, NH, D2O exchangeable), 7.28 (d, J ¼ 6.5 Hz, 2H,
AreH), 7.32 (t, J ¼ 6.0 Hz, 1H, AreH), 7.39 (d, J ¼ 6.2 Hz, 2H, AreH),
7.44 (d, J ¼ 6.0 Hz, 1H, AreH), 7.68 (d, J ¼ 6.1 Hz, 1H, AreH), 7.78 (s,
Q
Q
DS ¼ ð1=2 þ 1=2SiÞ ti
(1)
1H, AreH), 8.1 (s, 1H, vinyliceH). 13C NMR:
d 17.1 (CH3), 22.2 (CH3),
where; S ¼ 1/1 þ eapþb
)
107.1 (arylidene carbon), 116.0 (C^N), 123.8, 126.4, 126.9, 127.5,
128.0, 130.3, 134.2, 135.0, 135.3, 135.7, 144.8, 153.6, 158.8 (AreC),
164.5, 168.2 (2C]O). MS m/z (Rel. Int.): 345 (Mþ þ 1, 34), 105
(100). Analysis for C20H16N4O2.
DS is the drug score. Si is the contributions calculated directly
from miLogP; logS, molecular weight and drug likeness (pi) via the
second equation, which describes a spline curve. Parameters a and
b are (1, ꢀ5), (1, 5), (0.012, ꢀ6) and (1, 0) for miLogP, logS, molecular
weight and drug likeness, respectively. The ti is the contributions
taken from the four toxicity risk types and the values are 1.0, 0.8
and 0.6 for no risk, medium risk and high risk, respectively. The
reported compounds 4e23 showed moderate to good drug score as
compared with standard drug used.
6: Yield: 88%, mp: 192e194 ꢂC (acetic acid). IR (KBr, cmꢀ1): 3274
(NH), 3091 (CH arom.), 2984, 2880 (CH aliph.), 2222 (C^N), 1685,
1677 (2C]O), 1622 (C]N). 1H NMR:
d 1.58 (s, 3H, CH3), 2.35 (s, 3H,
CH3), 2.41 (s, 3H, CH3), 7.33 (bs, 1H, NH, D2O exchangeable), 7.40 (d,
J ¼ 7.1 Hz, 2H, AreH), 7.43 (d, J ¼ 7.0 Hz, 2H, AreH), 7.47 (d,
J ¼ 7.3 Hz, 1H, AreH), 7.65 (d, J ¼ 7.5 Hz, 1H, AreH), 7.82 (s, 1H,
AreH), 8.0 (s, 1H, vinyliceH). 13C NMR:
d 17.2 (CH3), 22.2 (2CH3),
4. Experimental protocols
107.2 (arylidene carbon), 116.2 (C^N), 123.6, 126.3, 126.9, 127.5,
128.0, 130.3, 134.2, 135.0, 135.3, 135.7, 144.8, 153.6, 158.8 (AreC),
164.5, 168.2 (2C]O). MS m/z (Rel. Int.): 359 (Mþ þ 1, 52). Anal-
ysis for C21H18N4O2.
4.1. Chemistry
Melting points (ꢂC, uncorrected) were determined in open
capillaries on a Gallenkamp melting point apparatus (Sanyo Gal-
lenkamp, Southborough, UK) and were uncorrected. Precoated
silica gel plates (silica gel 0.25 mm, 60G F254; Merck, Germany)
were used for thin layer chromatography, dichloromethane/meth-
anol (9.5:0.5) mixture was used as a developing solvent system and
the spots were visualized by ultraviolet light and/or iodine. Infra
red spectra were recorded in KBr discs using IR-470 Shimadzu
7: Yield: 83%, mp: 189e191 ꢂC (acetic acid). IR (KBr, cmꢀ1): 3277
(NH), 3092 (CH arom.), 2986, 2880 (CH aliph.), 2221 (C^N), 1689,
1671 (2C]O), 1620 (C]N). 1H NMR:
d 1.60 (s, 3H, CH3), 2.35 (s, 3H,
CH3), 7.39 (d, J ¼ 7.1 Hz, 2H, AreH), 7.41 (d, J ¼ 7.0 Hz, 2H, AreH),
7.45 (d, J ¼ 7.3 Hz, 1H, AreH), 7.66 (d, J ¼ 7.5 Hz, 1H, AreH), 7.82
(s, 1H, AreH), 8.01 (bs, 1H, NH, D2O exchangeable), 8.2 (s, 1H,
vinyliceH). 13C NMR:
d
18.3 (CH3), 23.6 (CH3), 106.7 (arylidene
carbon), 115.0 (C^N), 123.4, 126.0, 126.5, 127.8, 128.6, 130.2, 133.4,

138
A.M. Alafeefy et al. / European Journal of Medicinal Chemistry 53 (2012) 133e140
135.3, 135.6, 147.8, 153.6, 158.8 (AreC), 164.3, 168.1 (2C]O). MS m/z
was heated under reflux for 5 h. The reaction mixture was left to
cool and evaporated under vacuum. The remaining product was
triturated with ethanol and the formed solid product was collected
by filtration.
(Rel. Int.): 380 (Mþ
C20H15ClN4O2.
þ
2, 24), 378 (Mþ, 75). Analysis for
8: Yield: 75%, mp: 194e196 ꢂC (acetic acid). IR (KBr, cmꢀ1): 3275
(NH), 3091 (CH arom.), 2988, 2881 (CH aliph.), 2220 (C^N), 1687,
13: Yield: 67%, mp: 203e205 ꢂC (ethanol). IR (KBr, cmꢀ1): 3381,
3355, 3326 (2NH2), 3065 (CH arom.), 2884 (CH aliph.), 2225 (C^N),
1670 (2C]O), 1623 (C]N). 1H NMR:
d 1.58 (s, 3H, CH3), 2.36 (s, 3H,
CH3), 7.33 (bs, 1H, NH, D2O exchangeable), 7.44 (d, J ¼ 7.3 Hz, 1H,
AreH), 7.57 (d, J ¼ 7.4 Hz, 2H, AreH), 7.62 (d, J ¼ 7.0 Hz, 1H, AreH),
7.83 (s, 1H, AreH), 8.02 (d, J ¼ 6.0 Hz, 2H, AreH), 8.13 (s, 1H,
1677,1661 (2C]O),1620 (C]N). 1H NMR:
d 1.46 (s, 3H, CH3), 2.35 (s,
3H, CH3), 4.37 (s,1H, pyridineeH), 7.27 (d, J ¼ 6.5 Hz,1H, AreH), 7.30
(d, J ¼ 6.5 Hz, 1H, AreH), 7.81 (s, 1H, AreH), 8.45 (bs, 4H, 2NH2, D2O
vinyliceH). 13C NMR:
d
17.8 (CH3), 24.0 (CH3), 106.6 (arylidene
exchangeable). 13C NMR:
d 19.5 (CH3), 23.7 (CH3), 70.0 (CeCN), 79.3
carbon), 114.8 (C^N), 123.4, 126.0, 126.5, 127.9, 128.2, 130.2, 134.0,
135.1, 135.3, 135.6, 147.8, 153.6, 158.8 (AreC), 164.3, 168.1 (2C]O).
MS m/z (Rel. Int.): 390 (Mþ þ 1, 19). Analysis for C20H15N5O4.
9: Yield: 75%, mp: 208e210 ꢂC (acetic acid). IR (KBr, cmꢀ1): 3376
(OH), 3282 (NH), 3084 (CH arom.), 2980, 2884 (CH aliph.), 2225
(pyridineeCH), 115.7 (C^N), 121.0, 123.5, 128.4, 133.9, 136.4, 145.3
(AreC), 162.7, 166.5 (2C]O), 168.0, 175.7 (2CeNH2). MS m/z (Rel.
Int.): 323 (Mþ þ 1, 13), 322 (Mþ, 25). Analysis for C16H14N6O2.
14: Yield: 65%, mp: 179e181 ꢂC (ethanol). IR (KBr, cmꢀ1): 3416
(OH), 3307, 3304 (NH2), 3089 (CH arom.), 2884, 2867 (CH aliph.),
(C^N), 1681, 1672 (2C]O), 1627 (C]N). 1H NMR:
d
1.58 (s, 3H,
1677, 1661 (2C]O), 2220 (C^N), 1620 (C]N). 1H NMR:
d 1.50 (s,
CH3), 2.34 (s, 3H, CH3), 6.86 (s, 1H, AreH), 6.90 (d, J ¼ 6.4 Hz, 1H,
AreH), 6.99 (d, J ¼ 6.5 Hz, 1H, AreH), 7.03 (t, J ¼ 6.6 Hz, 1H, AreH),
7.41 (d, J ¼ 7.0 Hz, 2H, AreH), 7.90 (s, 1H, AreH), 8.03 (s, 1H,
vinyliceH), 8.11 (bs, 1H, NH, D2O exchangeable), 10.50 (bs, 1H, OH,
3H, CH3), 2.34 (s, 3H, CH3), 5.02 (bs, 2H, NH2, D2O exchangeable),
6.37 (s, 1H, pyridineeH), 7.25 (d, J ¼ 6.5 Hz, 1H, AreH), 7.31 (d,
J ¼ 6.5 Hz, 1H, AreH), 7.80 (s, 1H, AreH), 12.80 (bs, 1H, OH, D2O
exchangeable). 13C NMR:
d 19.8 (CH3), 23.6 (CH3), 71.8 (CeCN), 74.5
D2O exchangeable). 13C NMR:
d
18.9 (CH3), 24.0 (CH3), 107.7 (ary-
(pyrimidineeCH), 115.8 (C^N), 121.2, 123.7, 128.5, 133.4, 136.9,
145.0 (AreC), 162.5, 165.8 (2C]O), 171.0 (CeOH), 175.7 (CeNH2).
MS m/z (Rel. Int.): 324 (Mþ þ 1, 12), 323 (Mþ, 29). Analysis for
C16H13N5O3.
lidene carbon), 116.2 (C^N), 123.8, 126.4, 126.8, 127.2, 128.2, 131.6,
134.7, 135.3, 136.8, 145.8, 153.7, 158.5 (AreC), 164.3, 168.0 (2C]O).
MS m/z (Rel. Int.): 360 (Mþ, 64). Analysis for C20H16N4O3.
10: Yield: 75%, mp: 214e216 ꢂC (ethanol). IR (KBr, cmꢀ1): 3370
(OH), 3278 (NH), 3095 (CH arom.), 2981, 2881 (CH aliph.), 2220
4.1.4. 6-Amino-1-(2,6-dimethyl-4-oxoquinazolin-3(4H)-yl)-2-oxo-
4-phenyl-1,2-dihydro pyridine-3,5-dicarbonitrile 15, ethyl 2-amino-
5-cyano-1-(2,6-dimethyl-4-oxoquinazolin-3(4H)-yl)-6-oxo-4-
phenyl-1,6-dihydropyridine-3-carboxylate 16
(C^N), 1687, 1672 (2C]O), 1621 (C]N). 1H NMR:
d 1.59 (s, 3H,
CH3), 2.35 (s, 3H, CH3)), 6.70 (d, J ¼ 6.5 Hz, 2H, AreH), 7.0 (d,
J ¼ 6.5 Hz, 2H, AreH), 7.34 (d, J ¼ 7.0 Hz, 2H, AreH), 7.95 (s, 1H,
AreH), 8.12 (s, 1H, vinyliceH), 8.40 (bs, 1H, NH, D2O exchangeable),
To a solution of compound 5 (3.44 g, 0.01 mol) in 1,4-dioxane
(40 mL) containing triethylamine (1 mL) the appropriate active
methylene derivative (0.01 mol) was added. The reaction mixture
was heated under reflux for 6 h then poured onto ice/water and the
separated solid was filtered, washed with water, dried and
crystallized.
10.26 (bs, 1H, OH, D2O exchangeable). 13C NMR:
d 19.1 (CH3), 24.5
(CH3), 108.3 (arylidene carbon), 116.5 (C^N), 123.2, 126.4, 126.1,
127.4, 128.2, 130.5, 134.0, 135.2, 135.3, 136.1, 144.3, 153.7, 159.0
(AreC), 164.0, 166.2 (2C]O). MS m/z (Rel. Int.): 361 (Mþ þ 1, 16).
Analysis for C20H16N4O3.
11: Yield: 89%, mp: 251e253 ꢂC (ethanol). IR (KBr, cmꢀ1): 3382,
3376 (2OH), 3276 (NH), 3095 (CH arom.), 2980, 2886 (CH aliph.), 2228
15: Yield: 67%, mp: 196e198 ꢂC (ethanol). IR (KBr, cmꢀ1): 3352,
3329 (NH2), 3091 (CH arom.), 2888, 2865 (CH aliph.), 2228, 2225
(C^N), 1683, 1669 (2C]O), 1625 (C]N). 1H NMR:
d
1.54 (s, 3H, CH3),
(2C^N), 1676, 1665 (2C]O), 1621 (C]N). 1H NMR:
d 1.50 (s, 3H,
2.35 (s, 3H, CH3)), 6.79 (d, J ¼ 7.40 Hz, 1H, AreH), 7.24 (s, 1H, AreH),
7.35 (d, J ¼ 7.0 Hz, 1H, AreH), 7.45 (d, J ¼ 7.0 Hz, 1H, AreH), 7.66 (d,
J ¼ 7.0 Hz, 1H, AreH), 7.94 (s, 1H, AreH), 8.15 (s, 1H, vinyliceH), 8.47
(bs, 1H, NH, D2O exchangeable), 10.27 (bs, 2H, 2OH, D2O exchange-
CH3), 2.33 (s, 3H, CH3), 7.0 (d, J ¼ 7.0 Hz, 2H, AreH), 7.14 (t, J ¼ 7.0 Hz,
1H, AreH), 7.27 (d, J ¼ 6.5 Hz, 1H, AreH), 7.40 (t, J ¼ 7.0 Hz, 2H,
AreH), 7.52 (d, J ¼ 6.5 Hz,1H, AreH), 7.81 (s,1H, AreH), 8.61 (bs, 2H,
NH2, D2O exchangeable). 13C NMR:
d 20.0 (CH3), 23.9 (CH3), 72.0
able). 13C NMR:
d
19.6 (CH3), 22.9 (CH3), 107.8 (arylidene carbon),
(CeChN), 115.0 (CeChN), 115.3, 115.9 (2C^N), 120.0, 121.3, 126.4,
128.3, 128.4, 129.3, 132.9, 133.6, 137.2, 144.5, 157.0, 158.7, 159.1
(AreC), 162.5, 165.8 (2C]O). MS m/z (Rel. Int.): 409 (Mþ þ 1, 18),
408 (Mþ, 65). Analysis for C23H16N6O2.
115.0, 116.2 (C^N), 117.5, 121.0, 123.5, 125.5, 127.1, 129.6, 134.3, 136.9,
144.5, 145.4, 146.9, 151.4, 156.3 (AreC), 161.0, 164.7 (2C]O). MS m/z
(Rel. Int.): 377 (Mþ þ 1, 18). Analysis for C20H16N4O4.
12: Yield: 72%, mp: 229e231 ꢂC (ethanol). IR (KBr, cmꢀ1): 3272
(NH), 3255 (NH), 3095 (CH arom.), 2886, 2871 (CH aliph.), 1671,
16: Yield: 70%, mp: 170e172 ꢂC (ethanol). IR (KBr, cmꢀ1): 3341
(NH2), 3090 (CH arom.), 2889, 2863 (CH aliph.), 2226 (C^N), 1729,
1668 (2C]O), 1622 (C]N). 1H NMR:
d
1.60 (s, 3H, CH3), 2.34 (s, 3H,
1676, 1665 (3C]O), 1621 (C]N). 1H NMR:
d 1.0 (s, 3H, CH3), 2.13 (t,
CH3), 5.70 (bs, 1H, ¼NH, D2O exchangeable), 6.89 (t, J ¼ 6.2 Hz, 1H,
AreH), 7.04 (t, J ¼ 6.3 Hz, 1H, AreH), 7.18 (d, J ¼ 6.5 Hz, 1H, AreH),
7.22 (d, J ¼ 6.4 Hz, 1H, AreH), 7.35 (d, J ¼ 6.5 Hz, 1H, AreH), 7.59 (d,
J ¼ 6.5 Hz, 1H, AreH), 7.70 (s, 1H, AreH), 8.32 (s, 1H, AreH), 8.90 (bs,
3H, CH3), 2.36 (s, 3H, CH3), 4.63 (q, 2H, CH2), 6.94 (d, J¼6.6 Hz, 2H,
AreH), 7.12 (t, 1H, J ¼ 7.1 Hz, AreH), 7.28 (d, J ¼ 6.5 Hz, 1H, AreH),
7.41 (t, J¼6.5 Hz, 2H, AreH), 7.49 (d, J ¼ 6.5 Hz, 1H, AreH), 7.81 (s,
1H, AreH), 8.45 (bs, 2H, NH2, D2O exchangeable). 13C NMR:
d 15.2
1H, NH, D2O exchangeable). 13C NMR:
d
19.9 (CH3), 23.2 (CH3),115.9,
(CH3), 19.8 (CH3), 24.0 (CH3), 87.0 (CeCO2Et), 110.8 (CeChN), 116.5
(C^N), 120.3, 122.1, 126.7, 128.4, 128.9, 129.5, 132.2, 133.5, 138.0,
144.7, 156.8, 158.6, 159.4 (AreC), 161.7, 163.5, 168.0 (3C]O). MS m/z
(Rel. Int.): 456 (Mþ þ 1, 18), 455 (Mþ, 65). Analysis for C25H21N5O4.
117.0, 122.0, 123.4, 126.0, 126.5, 127.8, 128.6, 130.2, 131.8, 133.4,
135.3, 135.6, 147.8, 153.6 (AreC), 158.1, 162.5, 165.8 (2C]O and C]
N). MS m/z (Rel. Int.): 361 (Mþ þ 1, 14). Analysis for C20H16N4O3.
4.1.3. 4,6-Diamino-1,2-dihydro-1-(2,6-dimethyl-4-oxoquinazolin-
3(4H)-yl)-3-cyano-2-oxopyridine 13 and 4-amino-1,2-dihydro-6-
hydroxy-1-(2,6-dimethyl-4-oxoquinazolin-3(4H)-yl)-3-cyano-2-
oxopyridine 14
4.1.5. 3,5-Diamino-4-cyano-N-(2,6-dimethyl-4-oxoquinazolin-
3(4H)-yl) thiophene-2-carboxamide 17 and ethyl 5-(2,6-dimethyl-
4-oxoquinazolin-3(4H)-ylcarbamoyl)2,4diaminothiophene-3-
carboxylate 18
Equimolecular amounts of compound 4 (2.56 g, 0.01 mol) and
either malononitrile (0.66 g, 0.01 mol) or ethyl cyanoacetate (1.13 g,
0.01 mol) in 1,4-dioxane (30 mL) containing triethylamine (1.5 mL)
4.1.5.1. General procedure. To a solution of compound 4 (2.56 g,
0.01 mol) in absolute ethanol (50 mL) containing triethylamine
(1.0 mL) either malononitrile (0.66 g, 0.01 mol), ethyl cyanoacetate

A.M. Alafeefy et al. / European Journal of Medicinal Chemistry 53 (2012) 133e140
139
(1.13 g, 0.01 mol) or cyclohexanone (0.98 g, 0.01 mol) together with
elemental sulfur (0.32 g, 0.01 mol) were added. The whole reaction
mixture was heated under reflux for 1 h then poured onto ice/water
mixture and the formed solid product, in each case, was collected
by filtration.
CH3), 2.33 (s, 3H, CH3), 4.59 (s, 2H, CH2), 6.80 (bs, 2H, NH2, D2O
exchangeable), 7.20 (d, J ¼ 7.0 Hz, 2H, AreH), 7.24 (t, J ¼ 7.0 Hz, 1H,
AreH), 7.27 (d, J ¼ 6.5 Hz, 1H, AreH), 7.41 (t, J¼7.0 Hz, 2H, AreH),
7.55 (d, J ¼ 6.5 Hz, 1H, AreH), 7.81 (s, 1H, AreH), 9.15 (bs, 1H, NH,
D2O exchangeable). 13C NMR:
d 20.1 (CH3), 24.5 (CH3), 50.5 (CH2),
17: Yield: 76%, mp: 221e223 ꢂC (ethanol). IR (KBr, cmꢀ1): 3374,
3298, 3275 (2NH2), 3091 (CH arom.), 2888, 2865 (CH aliph.), 2226
77.5, 120.7, 127.5, 129.2, 132.7, 133.5, 135.0, 138.1, 143.6, 154.0, 156.9
(AreC), 160.4, 162.9 (2C]O), 175.0 (C]S). MS m/z (Rel. Int.): 438
(Mþ þ 1, 3). Analysis for C21H19N5O2S2.
(C^N), 1676, 1665 (2C]O), 1621 (C]N). 1H NMR:
d 2.16 (s, 3H,
CH3), 2.37 (s, 3H, CH3), 6.48, 6.69 (2bs, 4H, 2NH2, D2O exchange-
able), 7.24 (d, J ¼ 7.0 Hz, 1H, AreH), 7.27 (d, J ¼ 7.0 Hz, 1H, AreH),
7.80 (s, 1H, AreH), 8.10 (bs, 1H, NH, D2O exchangeable). 13C NMR:
4.1.7. 2-(2-Phenylhydrazono)-2-cyano-N-(2,6-dimethyl-4-
oxoquinazolin-3(4H)-yl)acetamide 22 and 2-(2-(4-
d
20.0 (CH3), 23.8 (CH3), 82.5 (CeChN), 116.3 (C^N), 121.0, 122.8,
aminosulphonylphenyl)hydrazono)-2-cyano-N-(2,6-dimethyl-4-
oxoquinazolin-3(4H)-yl)acetamide 23
127.9, 133.7, 136.5, 137.3, 144.9 (AreC), 155.0, 158.6 (2CeNH2), 161.7,
162.5 (2C]O). MS m/z (Rel. Int.): 355 (Mþ þ 1, 20), 354 (Mþ, 47).
Analysis for C16H14N6O2S.
4.1.7.1. General procedure. To a solution of compound 4 (2.56 g,
0.01 mol) in ethanol (40 mL) containing sodium acetate (10.0 g),
benzene diazonium chloride (1.40 g, 0.01 mol) and/or 4-
sulphonylaminoebenzeneediazoniume chloride (2.19 g, 0.01 mol)
with continuous stirring at 0e5 ꢂC for 1 h. The solid product, formed
in each case, was collected by filtration and crystallized from
ethanol.
18: Yield: 76%, mp: 200e2002 ꢂC (ethanol). IR (KBr, cmꢀ1): 3365,
3307, 3285 (2NH2, NH), 3089 (CH arom.), 2984, 2867 (CH aliph.),
1721,1672,1666 (3C]O),1620 (C]N). 1H NMR:
d
1.03 (t, J ¼ 7.36 Hz,
3H, CH3), 2.25 (s, 3H, CH3), 2.33 (s, 3H, CH3), 4.25 (q, J ¼ 7.60 Hz, 2H,
CH2), 5.74, 6.80 (2bs, 4H, 2NH2, D2O exchangeable), 7.25 (d,
J ¼ 7.0 Hz, 1H, AreH), 7.32 (d, J ¼ 7.0 Hz, 1H, AreH), 7.82 (s, 1H,
22: Yield: 73%, mp: 207e209 ꢂC (ethanol). IR (KBr, cmꢀ1): 3317,
3283 (2NH), 3087 (CH arom.), 2878, 2869 (CH aliph.), 2226 (C^N)
AreH), 11.13 (s, 1H, NH, D2O exchangeable). 13C NMR:
d 15.20 (CH3),
19.9 (CH3), 24.2 (CH3), 60.7 (CH2), 121.2, 122.7, 127.9, 130.6, 131.5,
135.0, 137.3, 138.0, 144.5, 158.6 (AreC), 161.2, 163.0, 167.20 (3C]O).
MS m/z (Rel. Int.): 402 (Mþ þ 1, 12), 401 (Mþ, 24). Analysis for
C18H19N5O4S.
1677, 1661 (2C]O), 1622 (C]N),1375, 1190 (SO2). 1H NMR:
d 2.17 (s,
3H, CH3), 2.33 (s, 3H, CH3), 6.25 (s, 1H, NH, D2O exchangeable), 6.88
(t, J ¼ 6.5 Hz, 1H, AreH), 7.10 (d, J ¼ 7.0 Hz, 2H, AreH), 7.24 (d,
J ¼ 7.0 Hz, 2H, AreH), 7.27 (d, J ¼ 7.0 Hz, 1H, AreH), 7.32 (d,
J ¼ 7.0 Hz, 1H, AreH), 7.82 (s, 1H, AreH), 11.20 (s, 1H, NH, D2O
4.1.6. 3-Amino-4,5,6,7-tetrahydro-N-(2,6-dimethyl-4-oxoquinazolin-
3(4H)-yl)benzo[b]thiophene-2-carboxamide 19, 4-amino-2,3-
dihydro-N-(2,6-dimethyl-4-oxoquinazolin-3(4H)-yl)-3-phenyl-2-
thioxothiazole-5-carboxamide 20 and 4-amino-N-(2,6-dimethyl-4-
oxoquinazolin-3(4H)-yl)-3-benzyl-2-thioxo-2,3-dihydrothiazole-5-
carboxamide 21
exchangeable). 13C NMR:
d 19.2 (CH3), 24.5 (CH3), 107.0 (CeCN),
115.6 (C^N), 117.0, 119.5, 121.0, 122.4, 128.9, 129.6, 133.0, 138.0,
144.9, 148.3, 154.0 (AreC), 161.2, 163.0 (2C]O). MS m/z (Rel. Int.):
361 (Mþ þ 1, 3), 360 (Mþ, 7). Analysis for C19H16N6O2.
23: Yield: 58%, mp: 256e258 ꢂC (ethanol). IR (KBr, cmꢀ1): 3332,
3275, 3251 (NH2, 2NH), 3087 (CH arom.), 2880, 2864 (CH aliph.),
2227 (C^N) 1674, 1667 (2C]O), 1630 (C]N), 1371, 1192 (SO2). 1H
4.1.6.1. General procedure. To a solution of compound 4 (2.56 g,
0.01 mol) in absolute ethanol (50 mL) containing triethylamine
(1.0 mL) cyclohexanone (0.98 g, 0.01 mol) and/or the isothiocyanate
derivative together with elemental sulfur (0.32 g, 0.01 mol) were
added. The whole reaction mixture was heated under reflux for 3 h
then poured onto ice/water mixture and the formed solid product,
in each case, was collected by filtration and crystallized from
ethanol.
NMR:
d
2.13 (s, 3H, CH3), 2.40 (s, 3H, CH3), 6.97 (d, J ¼ 7.0 Hz, 2H,
AreH), 7.37 (d, J ¼ 7.0 Hz, 1H, AreH), 7.42 (d, J ¼ 7.0 Hz, 1H, AreH),
7.64 (d, J ¼ 7.0 Hz, 2H, AreH), 7.76 (s, 1H, AreH), 7.85 (s, 2H, NH2,
D2O exchangeable), 8.20 (s, 1H, NH, D2O exchangeable), 11.70 (s, 1H,
NH, D2O exchangeable). 13C NMR:
d 20.0 (CH3), 24.7 (CH3), 107.1
(CeChN), 115.0 (C^N), 117.0, 121.0, 122.4, 128.9, 129.5, 130.0, 133.4,
137.5, 144.6, 148.5, 154.2 (AreC), 161.5, 163.0 (2C]O). MS m/z (Rel.
Int.): 440 (Mþ þ 1, 5), 439 (Mþ, 9). Analysis for C19H17N7O4S.
19: Yield: 66%, mp: 192e194 ꢂC (ethanol). IR (KBr, cmꢀ1): 3377,
3292, 3245 (NH2, NH), 3095 (CH arom.), 2962, 2880 (CH aliph.),
1686, 1675 (2C]O), 1638 (C]N). 1H NMR:
d 1.45 (s, 3H, CH3),
1.70e1.80 (t, J ¼ 7.40, 4H, 2CH2), 2.24e2.25 (m, 4H, 2CH2), 2.30 (s,
3H, CH3), 6.49 (bs, 2H, NH2, D2O exchangeable), 7.23 (d, J ¼ 7.0 Hz,
1H, AreH), 7.27 (d, J ¼ 7.0 Hz, 1H, AreH), 7.80 (s, 1H, AreH), 11.80
4.2. In vitro anticancer screening
The stock solutions of the tested compounds were prepared in
DMSO and were used for serial dilutions in culture medium.
Human breast cancer (MCF-7), cervical cancer (HeLa) and hepato-
cellular liver carcinoma (HepG2) cell lines were grown in RPMI-
1640 medium supplemented with 10% calf serum. For growth
assays, exponentially growing cells were suspended in the above-
mentioned medium at a density of 4 ꢄ 104 cells per mL, seeded
(bs, 1H, NH, D2O exchangeable). 13C NMR:
d 19.6 (CH3), 23.2 (CH3),
17.3, 21.0, 23.5, 25.2 (4CH2), 120.7, 122.9, 126.8, 127.9, 133.5, 136.6,
137.4, 139.0, 144.1, 147.3, 159.0 (AreC), 161.7, 162.5 (2C]O). MS m/z
(Rel. Int.): 369 (Mþ þ 1, 14), 368 (Mþ, 33). Analysis for C19H20N4O2S.
20: Yield: 73%, mp: 211e213 ꢂC (ethanol). IR (KBr, cmꢀ1): 3325,
3289, 3252 (NH2, NH), 3089 (CH arom.), 2888, 2865 (CH aliph.),
1676, 1661 (2C]O), 1640 (C]N), 1269 (C]S). 1H NMR:
d
2.15 (s, 3H,
onto 96-well plates well (200 m
L/well), and incubated at 37 ꢂC in
CH3), 2.34 (s, 3H, CH3), 6.72 (bs, 2H, NH2, D2O exchangeable), 7.08
(d, J ¼ 7.0 Hz, 2H, AreH), 7.22 (t, J ¼ 7.0 Hz, 1H, AreH), 7.28 (d,
J ¼ 6.5 Hz, 1H, AreH), 7.35 (t, J ¼ 7.0 Hz, 2H, AreH), 7.56 (d,
J ¼ 6.5 Hz, 1H, AreH), 7.80 (s, 1H, AreH), 9.55 (bs, 1H, NH, D2O
a humidified 5% CO2 atmosphere for 24 h. After that, the cell
medium in test wells was changed to new culture medium con-
taining different concentrations of the tested compounds, while the
cell medium in control wells was changed to new culture medium
containing an equivalent volume of solvent. After incubation at
exchangeable). 13C NMR:
d 20.0 (CH3), 24.6 (CH3), 78.0, 120.3, 127.7,
129.3, 132.1, 133.9, 136.7, 140.1, 143.5, 154.8, 158.1 (AreC), 161.7,
166.8 (2C]O), 177.2 (C]S). MS m/z (Rel. Int.): 424 (Mþ þ 1, 11), 423
(Mþ, 42). Analysis for C20H17N5O2S2.
37 ꢂC in a humidified 5% CO2 atmosphere for 3 d, 100
mL of MTT (3-
(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide,
0.5 mg/mL) in serum-free medium was added to each well and
21: Yield: 81%, mp: 211e213 ꢂC (ethanol). IR (KBr, cmꢀ1): 3330,
3282, 3257 (NH2, NH), 3089 (CH arom.), 2890, 2868 (CH aliph.),
incubated at 37 ꢂC for an additional 4 h. Then, 100
mL of DMSO was
added to each well and mixed thoroughly to dissolve the resulting
formazan product. The cell viability was evaluated by measurement
1669, 1663 (2C]O), 1640 (C]N), 1265 (C]S). 1H NMR:
d 2.15 (s, 3H,

140
A.M. Alafeefy et al. / European Journal of Medicinal Chemistry 53 (2012) 133e140
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percentage of cell growth inhibition was calculated as follows:
%Inhibition ¼ ðMeanOD control ꢀ MeanOD testÞ=MeanOD control
ꢄ 100%
The dose response curves of the compounds effecting ꢁ70%
inhibition in one-dose prescreening for each cell line were
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5. Conclusion
The present work, through simple synthetic approaches, led to
the development of novel hybrids of quinazoline containing tyr-
phostin pharmacophore that exhibited remarkable anti-
proliferative activities against three tumor cell lines. The
compounds showed suitable drug like properties and are expected
to present good bioavailability profile. As new class of tyrphostin
analogs, 10 of the obtained new compounds showed remarkable
antiproliferative activity against three tumor cell lines. Most of the
active compounds were devoid of the typical catechol feature of
tyrphostin and so the activity could be attributed to some sort of
synergism between quinazoline and tyrphostin coupled molecule.
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Acknowledgments
[17] A. Rauf, F. Ahmed, A.M. Qureshi, Aziz-ur-Rehman, A. Khan, M.I. Qadir,
M.I. Choudhary, Z.H. Chohan, M.H. Youssoufi, T. Ben Hadda, Synthesis and
urease inhibition studies of barbituric and thiobarbituric acid derived sul-
phonamides, J. Chin. Chem. Soc. 58 (4) (2011) 1e10.
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T. Ben Hadda, Synthesis, biopharmaceutical characterization, antimicrobial
This work was supported by a grant from the National Plan of
Science, Technology and Innovation (Grant No. 10-MED1188-02)
King Saud University, Riyadh, Saudi Arabia.
and antioxidant activities of 1-(4’-O-b-D-glucopyranosyloxy-2’-hydrox-
yphenyl)-3-aryl-propane-1,3-diones, Eur. J. Med. Chem. 46 (2011)
1390e1399.
Appendix A. Supplementary data
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and experimental verification of the antibacterial potential of some mono-
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physicochemical properties and verification by experimental correlations of
novel coumarin derivatives produced via Betti’s protocol, Eur. J. Med. Chem.
45 (9) (2010) 4370e4378.
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.ejmech.2012.03.044. These data
include MOL files and InChIKeys of the most important compounds
described in this article.
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