In vitro anticancer screening and radiosensitizing evaluation of some new quinolines and pyrimido[4,5-b]quinolines bearing a sulfonamide moiety

Article abstract of DOI:10.1016/j.ejmech.2010.05.014

Sulfonamide bearing compounds posses many types of biological activities and have recently been reported to show substantial antitumor activity in vitro and/or in vivo. There are a variety of mechanisms for the anticancer activity, and the most prominent

Full text of DOI:10.1016/j.ejmech.2010.05.014

European Journal of Medicinal Chemistry 45 (2010) 3677e3684
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
In vitro anticancer screening and radiosensitizing evaluation of some new
quinolines and pyrimido[4,5-b]quinolines bearing a sulfonamide moiety
,
b
c
b
c
Mostafa M. Ghorab a , Fatma A. Ragab , Helmy I. Heiba , Reem K. Arafa , Ebaa M. El-Hossary
*
a Medicinal, Aromatic and Poisonous Plants Research Center, College of Pharmacy, King Saud University, P.O.Box 2457, Riyadh 11451, Saudi Arabia
b Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Cairo University, Cairo, Egypt
c Department of Drug Radiation Research, National Centre for Radiation Research and Technology, Atomic Energy Authority, Nasr City, Cairo, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
Sulfonamide bearing compounds posses many types of biological activities and have recently been
reported to show substantial antitumor activity in vitro and/or in vivo. There are a variety of mechanisms
for the anticancer activity, and the most prominent mechanism is the inhibition of carbonic anhydrase
(CA) isozymes. The present work reports the synthesis of twenty novel quinoline and pyrimido[4,5-b]
quinoline derivatives bearing a sulfonamide moiety. The new synthesized compounds were designed in
compliance with the general pharmacophoric requirements for CA inhibiting anticancer drugs, as this
may play a role in their anticancer activity. All the newly synthesized compounds were evaluated for
their in vitro anticancer activity against human breast cancer cell line (MCF7). Compounds 6, 9 and 18
Received 26 October 2009
Received in revised form
6 May 2010
Accepted 7 May 2010
Available online 12 May 2010
Keywords:
Quinoline
Pyrimido[4,5-b]quinoline
Sulfonamide
Carbonic anhydrase II
Anticancer
showed IC50 values (72.9
m
M, 72.1
M). On the other hand, compound 8 exhibited better activity than doxorubicin
M. Additionally, the most potent compounds 8 and 18 were evaluated for
-radiation.
Ó 2010 Elsevier Masson SAS. All rights reserved.
mM and 71.9 mM, respectively) comparable to that of the reference drug
doxorubicin (IC50 ¼ 71.8
m
with an IC50 value of 64.5
m
their ability to enhance the cell killing effect of
g
g-Radiation
1. Introduction
been discovered in higher vertebrates [12e15]. CAs are involved in
pH regulation, secretion of electrolytes, respiration [17,18],
biosynthetic reactions which require CO2/bicarbonate as substrate
such as gluconeogenesis, lipogenesis, ureagenesis, and pyrimidines
synthesis among others [19]. Other roles for these enzymes were
highlighted, such as calcification and bone resorption [19].
On the other hand, quinoline derivatives are important biolog-
ically active compounds showing anticancer activity [20e22]. In
the light of these facts, and as a continuation of our previous
reported work [23,24], we planned to synthesize a novel series of
quinoline and pyrimido[4,5-b]quinoline derivatives bearing
a sulfonamide moiety, in order to study their structure activity
relationship and hoping that the new compounds might show
significant anticancer activity. Moreover, we also aimed to evaluate
these new compounds for their in vitro anticancer activity in
Sulfonamides posses many types of biological activities and
many of them are widely used in therapy as antibacterial [1],
hypoglycemic [2], diuretic [3,4], anti-carbonic anhydrase [3,5] and
antithyroid agents [6]. Recently, a host of structurally novel
sulfonamide derivatives have been reported to show substantial
antitumor activity in vitro and/or in vivo [7e11].
It has been known that aryl/heteroaryl sulfonamides may act as
antitumor agents through a variety of mechanisms such as cell
cycle perturbation in the G1 phase, disruption of microtubule
assembly, angiogenesis inhibition, and functional suppression of
the transcriptional activator NF-Y. Moreover, following an exten-
sive evaluation, numerous sulfonamides were found to act as
carbonic anhydrase (CA) inhibitors [12e15]. The most prominent
mechanism was the inhibition of carbonic anhydrase isozymes
(CA) [16].
combination with
g-radiation, to evaluate their ability to enhance
the cytotoxic activity of
g-radiation.
In brief, the
a-CAs are a family of metalloenzymes involved in
the catalysis of an important physiological reaction, which is the
2. Results and discussion
hydration of carbon dioxide to bicarbonate and a proton (CO2
þ
H2O @ HCOꢀ3 þ Hþ). At least 13 enzymatically active isoforms have
A general pharmacophore (Fig. 1) for the compounds acting as
carbonic anhydrase inhibitors has been reported by Thiry et al. [25],
from the analysis of the CA active site and from the structure of
inhibitors described in the literature [15].
* Corresponding author. Tel.: þ966 534292860; fax: þ966 (01) 4670560.
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.2010.05.014

3678
M.M. Ghorab et al. / European Journal of Medicinal Chemistry 45 (2010) 3677e3684
Pro
Scaffold
Val
Leu
Phe
O
O
S
Hydrophobic
moiety
Thr
N
H2N
N
Cl
Hydrophobic
moiety
Glu
O
O
S
H2N
CN
Sulfonamide
Scaffold
H
Hydrophilic
link
O
Hydrophilic link
O
Glu
Zn
His
His
Scaffold
His
Gln
Asn
Fig. 1. Structural elements of CA inhibitors in the CA enzymatic active site.
O
S
O
Hydrophobic
moiety
Cl
N
N
H2N
This pharmacophore includes the structural elements that are
required to be present in the compounds in order to act as CA
inhibitors. This includes the presence of a sulfonamide moiety
which coordinates with the zinc ion of the active site of the CA and
the sulfonamide is attached to a scaffold, which is usually a benzene
ring. The side chain might posses a hydrophilic link able to interact
with the hydrophilic part of the active site and a hydrophobic
moiety which can interact with the hydrophobic part of the CA
active site.
O
NH
Sulfonamide
Hydrophilic link
Fig. 2. Representative examples of the synthesized compounds showing the compli-
ance with the general pharmacophore of sulfonamide compounds acting as CA
inhibitors.
Fig. 2 illustrates two representative examples of the newly
synthesized compounds showing compliance with the above-
mentioned pharmacophore model.
19 was obtained by the treatment of compound 6 with ethylene
diamine in the presence of carbon disulfide. The reaction proceeded
via intramolecular cyclization through the elimination of 1 mole of
ammonia. Also, the quinoline derivative 20 was obtained by the
reaction of compound 6 with ethyl acetoacetate (Scheme 4).
Treatment of compound 7 with phosphorus pentasulfide in
pyridine afforded the 4-thioxo-pyrimido[4,5-b]quinoline derivative
21. While, the 4-chloro-pyrimido[4,5-b]quinoline derivative 22 was
obtained by refluxing compound 7 in thionyl chloride for 2 h.
Compound 23 was obtained by the treatment of compound 7 with
ethyl bromoacetate in dry acetone in the presence of anhydrous
2.1. Chemistry
Enaminone 3 was obtained from condensation of 5,5-dimethyl-
1,3-cyclohexandione 1 with sulfanilamide 2 [23]. Treatment of
enaminone
3 with 2-(4-chlorobenzylidene)malononitrile 4 in
ethanol containing a catalytic amount of triethylamine, as a base
catalyst, yielded the corresponding hexahydroquinoline derivative
6, via the formation of the intermediate Michael type product 5,
followed by intramolecular cyclization (Scheme 1).
The pyrimido[4,5-b]quinoline derivative 7 was obtained by
refluxing compound 6 in formic acid. When compound 6 was
refluxed in acetic anhydride for 5 hours and 10 hours, the mono-
acetyl derivative 8 and the diacetyl derivative 9 were obtained,
respectively. While refluxing compound 6 in acetic anhydride for 15
hours yielded the fused pyrimido[4,5-b]quinoline system 10. Also,
the pyrimido[4,5-b]quinoline derivative 11 was obtained by the
reaction of compound 6 with formamide (Scheme 2).
O
O
NH2
SO2NH2
Ethanol
O
N
H
SO2NH2
(3)
(2)
(1)
Treatment of compound 6 with triethylorthoformate in the
presence of acetic anhydride yielded the quinoline derivative 12.
Reaction of compound 6 with chloroacetyl chloride at room
temperature afforded the quinoline derivative 13. While refluxing
compound6withchloroacetylchlorideindimethylformamidefor16
hours, yielded the 2-chloromethyl-pyrimido[4,5-b]quinoline deriv-
ative 14. Fusion of compound 6 with ethyl cyanoacetate yielded the
corresponding 2-(cyanomethyl)-pyrimido[4,5-b]quinoline deriva-
tive 15. The formation of compound 15 was assumed to proceed via
elimination of 1 mol of ethanol followed by intramolecular cycliza-
tion. The thioureido derivative 16 was obtained by the reaction of
compound 6 with ethyl isothiocyanate in dimethyl formamide con-
taining a catalytic amount of triethylamine (Scheme 3).
NC
CN
EtOH / TEA
Ar
(4)
O
Ar
O
Ar
CN
N
CN
NH2
C
NH
N
SO2NH2
SO2NH2
Fusion of compound 6 with succinic anhydride yielded the 2,5-
dioxopyrrolidinyl derivative 17. Stirring of compound 6 in conc.
H2SO4 caused partial hydrolysis of the cyano group yielding the
carboxamide derivative 18. Additionally, the imidazolyl derivative
(6)
(5)
Ar= 4-Chlorophenyl
Scheme 1.

M.M. Ghorab et al. / European Journal of Medicinal Chemistry 45 (2010) 3677e3684
3679
O
Ar
N
CN
O
Ar
N
O
Ar
N
O
N
CN
N(COCH3)2
NH
O
N
H
SO2NH2
(9)
(10)
(8)
SO2NH2
SO2NH2
5 h
10 h
15 h
O
Ar
N
O
O
Ar
NH2
N
NH
Ac2O
N
N
N
HCONH2
HCOOH
(6)
SO2NH2
SO2NH2
Ar= 4-Chlorophenyl
(7)
(11)
Scheme 2.
potassium carbonate. The 4-hydrazinyl derivative 24 and the
4-isothiocyanato derivative 25 were obtained by the reaction of the
4-chloro-pyrimido[4,5-b]quinoline derivative 22 with hydrazine
hydrate and ammonium thiocyanate, respectively (Scheme 5).
synthesized compounds where some compounds exhibited
significant activity compared to the reference drug.
From the results in Table 1, it was found that the quinoline
derivative 8 (IC50 ¼ 64.5
screening, and exhibited a higher cytotoxic activity when compared
M). Compounds
mM) was the most potent compound in this
with the reference drug doxorubicin (IC50 ¼ 71.8
m
6, 9 and 18 are nearly as active as doxorubicin, while compounds 7,
10e13, 16, 17, 22, 24 and 25 showed lower IC50 values than that of
2.2. In vitro anticancer screening
the reference drug, ranging from 76e99.6 mM.
The newly synthesized compounds were evaluated for their in
vitro cytotoxic activity against human breast cancer cell line, MCF7.
Doxorubicin which is one of the most effective anticancer agents
was used as the 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
2.3. Radiosensitizing activity
The rationale for combining chemotherapy and radiotherapy is
based mainly on two ideas, one being spatial cooperation, which is
effective if chemotherapy is sufficiently active to eradicate
subclinical metastases and if the primary local tumor is effectively
treated by radiotherapy. In this regard, no interaction between
O
Ar
N
O
NH
O
Ar
N
O
N
O
Ar
N
Cl
N
CN
NH
O
CN
Cl
N
H
SO2NH2
(14)
O
SO2NH2
SO2NH2
NC
Stirring
at R.T.
6 h
O
(13)
reflux
16 h
(15)
Fusion
O
O
Ar
O
Ar
N
DMF
Cl
CN
S
CN
N
Cl
N
N
H
N
H
O
HC(OCH2CH3)3
N
C S
(6)
DMF/TEA
Ar= 4-Chlorophenyl
SO2NH2
SO2NH2
(16)
(12)
Scheme 3.

3680
M.M. Ghorab et al. / European Journal of Medicinal Chemistry 45 (2010) 3677e3684
O
Ar
N
O
Ar
N
N
CONH2
NH2
N
H
NH2
SO2NH2
SO2NH2
(19)
(18)
Stirring at
R. T. for 6 h
NH2
CS2
Conc. H2SO4
O
Ar
N
H2N
O
Ar
N
CN
O
O
CN
N
O
O
O
O
O
N
H
O
O
(6)
O
Fusion
Fusion
SO2NH2
Ar= 4-Chlorophenyl
SO2NH2
(17)
(20)
Scheme 4.
radiotherapy and chemotherapy is required. The other idea is the
enhancement of radiation effects.
Similarly, compound 18 showed IC50 value of 71.9
alone, as shown in Table 1. The IC50 value was decreased to 44.4
when the cells were treated with compound 18 in combination
with -radiation (Fig. 4).
From these results, we can conclude that using the combination
of compound or 18 and ionizing radiation synergistically
m
M when used
mM,
Cytotoxic agents can enhance radiation effects by direct
enhancement of the initial radiation damage by incorporating
drugs into DNA, inhibiting cellular repair, accumulating cells in
a radiosensitive phase or eliminating radioresistant phase cells,
eliminating hypoxic cells or inhibiting the accelerated repopulation
of tumor cells. Virtually, all chemotherapeutic agents have the
ability to sensitize cancer cells to the lethal effects of ionizing
radiation [26].
g
8
enhanced growth inhibition on breast cancer cells, compared with
each agent alone.
3. Conclusion
Consequently, the ability of the most two active compounds,
compounds 8 and 18, to enhance the cell killing effect of
ation was studied. From the results obtained in Table 1, compound 8
showed an in vitro cytotoxic activity with IC50 value of 64.5 M,
when the cells were subjected to different concentrations of the
compound alone. While when the cells were subjected to the same
concentrations of compound 8, and irradiated with a single dose of
g-irradi-
The objective of the present study was to synthesize and
investigate the anticancer activity of some new quinoline and
pyrimidoquinoline derivatives bearing a free sulfonamide moiety.
Some of these new compounds exhibited significant anticancer
activity, when compared to doxorubicin as a reference drug. Since it
was reported that compounds bearing a free sulfonamide group
may exhibit potent carbonic anhydrase inhibition activity, which is
considered to be an interesting target for the design of anticancer
m
g-radiation at a dose level of 8 Gy, as shown in Table 2, the IC50
value was synergistically decreased to 33.1
mM (Fig. 3).
O
Ar
N
Cl
N
O
Ar
N
NCS
O
Ar
N
NHNH2
N
N
N
N
N
NH2NH2.H2O
NH4SCN
SO2NH2
SO2NH2
SO2NH2
(25)
(22)
(24)
O
Ar
O
N
O
Ar
N
S
SOCl2
O
N
NH
O
O
N
N
Br
P2S5
O
(7)
Ar= 4-Chlorophenyl
SO2NH2
SO2NH2
(21)
(23)
Scheme 5.

M.M. Ghorab et al. / European Journal of Medicinal Chemistry 45 (2010) 3677e3684
3681
dioxane to give 6: Yield, 85%; m.p. 280e282 ꢂC; IR, cmꢀ1: 3467, 3346,
3199 (2NH2), 3065 (CH arom.), 2969, 2879 (CH aliph.), 2174 (ChN),
d: 0.7, 0.9 [2s, 6H,
2CH3], 1.6e2.3 [m, 4H, 2CH2], 4.5 [s, 1H, CH], 5.5 [s, 2H, NH2,
Table 1
In vitro anticancer screening of the synthesized compounds against human breast
cell line (MCF7).
1645 (C¼O), 1374, 1162 (SO2). 1H NMR (DMSO-d6)
Compound Compound concentration (
10 25
m
M)
50
IC50
M)
(
m
exchangeable with D2O], 7.3e8.0 [m, 10H, Ar-H
þ
SO2NH2,
100
exchangeable with D2O]. 13C NMR (CDCl3)
d: 27.3, 33.4, 38.5, 43.3,
Surviving fraction (Means ꢁ SE)#
51.0, 57.9, 112.0, 117.1, 128.5, 130.4, 142.2, 143.9, 153.0, 167.5, 194.0.
MS, m/z (%): 482 [Mþ] (27.47), 371 (100). Anal. Calcd. For
C24H23ClN4O3S: C, 59.68; H, 4.80; N, 11.60. Found: C, 59.92; H, 5.00;
N, 11.33.
Doxorubicin 0.721 ꢁ 0.02 0.546 ꢁ 0.02 0.461 ꢁ 0.01 0.494 ꢁ 0.03
71.8
6
7
8
9
0.915 ꢁ 0.04 0.612 ꢁ 0.01 0.429 ꢁ 0.04 0.484 ꢁ 0.01
0.891 ꢁ 0.03 0.676 ꢁ 0.01 0.499 ꢁ 0.01 0.468 ꢁ 0.01
0.948 ꢁ 0.01 0.779 ꢁ 0.03 0.446 ꢁ 0.01 0.324 ꢁ 0.01
0.895 ꢁ 0.04 0.773 ꢁ 0.02 0.504 ꢁ 0.01 0.414 ꢁ 0.03
0.976 ꢁ 0.02 0.721 ꢁ 0.04 0.450 ꢁ 0.05 0.557 ꢁ 0.04
0.886 ꢁ 0.02 0.612 ꢁ 0.05 0.495 ꢁ 0.03 0.602 ꢁ 0.03
0.803 ꢁ 0.04 0.551 ꢁ 0.05 0.498 ꢁ 0.04 0.607 ꢁ 0.01
0.986 ꢁ 0.01 0.736 ꢁ 0.01 0.523 ꢁ 0.04 0.502 ꢁ 0.06
72.9
76.8
64.5
72.1
85.9
96.6
97.2
84.3
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
4.1.3. 4-(5-(4-Chlorophenyl)-8,8-dimethyl-4,6-dioxo-3,4,6,7,8,9-
hexahydropyrimido[4,5-b]quinolin-10(5H)-yl)benzenesulfonamide (7)
A solution of compound 6 (0.482 g, 1 mmol) in formic acid (30
mL) was refluxed for 5 h, the reaction mixture was cooled and then
poured onto cold water, the obtained solid was recrystallized from
0.914 ꢁ 0.03 0.775 ꢁ 0.05 0.731 ꢁ 0.05 0.625 ꢁ 0.04 >100
0.971 ꢁ 0.01 0.901 ꢁ 0.01 0.774 ꢁ 0.04 0.525 ꢁ 0.01 >100
0.916 ꢁ 0.01 0.837 ꢁ 0.02 0.594 ꢁ 0.06 0.561 ꢁ 0.04
0.930 ꢁ 0.02 0.813 ꢁ 0.02 0.584 ꢁ 0.02 0.549 ꢁ 0.02
0.922 ꢁ 0.02 0.662 ꢁ 0.01 0.445 ꢁ 0.01 0.452 ꢁ 0.01
99.6
95.2
71.9
dioxane to give compound 7: Yield, 78%; m.p. 162e164 ꢂC; IR, cmꢀ1
:
3390, 3359, 3263 (NH, NH2), 3072 (CH arom.), 2960, 2885 (CH
aliph.), 1708, 1644 (2 C¼O), 1627 (C¼N), 1372, 1165 (SO2). 1H NMR
0.939 ꢁ 0.01 0.763 ꢁ 0.01 0.568 ꢁ 0.03 0.621 ꢁ 0.03 >100
0.897 ꢁ 0.01 0.820 ꢁ 0.03 0.648 ꢁ 0.05 0.549 ꢁ 0.06 >100
0.927 ꢁ 0.03 0.910 ꢁ 0.02 0.884 ꢁ 0.04 0.814 ꢁ 0.01 >100
(DMSO-d6) d: 0.9, 1.0 [2s, 6H, 2CH3], 1.9e2.3 [m, 4H, 2CH2], 4.6 [s,
1H, CH], 7.3e7.6 [m, 10H, Ar-H þ SO2NH2], 7.9 [s, 1H, NH], 8.0 [s, 1H,
CH¼N]. MS, m/z (%): 495 [M-CH3] (2.25), 483 (100). Anal. Calcd. For
C25H23ClN4O4S: C, 58.76; H, 4.54; N, 10.96. Found: C, 58.94; H, 4.38;
N, 11.18.
0.843 ꢁ 0.05 0.696 ꢁ 0.03 0.476 ꢁ 0.03 0.546 ꢁ 0.05
86.5
0.954 ꢁ 0.01 0.880 ꢁ 0.01 0.653 ꢁ 0.03 0.563 ꢁ 0.03 >100
0.940 ꢁ 0.02 0.788 ꢁ 0.01 0.672 ꢁ 0.03 0.358 ꢁ 0.03
0.972 ꢁ 0.01 0.721 ꢁ 0.01 0.419 ꢁ 0.01 0.515 ꢁ 0.04
76
79.6
#
Each value is the mean of three values ꢁ Standard Error.
4.1.4. N-(4-(4-chlorophenyl)-3-cyano-7,7-dimethyl-5-oxo-1-
(4-sulfamoylphenyl)-1,4,5,6,7,8-hexahydroquinolin-2-yl)acetamide (8)
A solution of compound 6 (0.482 g, 1 mmol) in acetic anhydride
(20 mL) was refluxed for 5 h, the reaction mixture was then
concentrated, the solid separated was recrystallized from ethanol
to give compound 8: Yield, 82%; m.p. 149e151 ꢂC; IR, cmꢀ1: 3453,
3231, 3208 (NH, NH2), 3050 (CH arom.), 2955, 2875 (CH aliph.),
2213 (ChN), 1720, 1649 (2 C¼O), 1371, 1163 (SO2). 1H NMR (DMSO-
agents, the results obtained from the anticancer screening may give
a suggestion that the synthesized compounds may act as carbonic
anhydrase inhibitors and this may contribute in part to their anti-
cancer activity. Moreover, the most two active compounds showed
the ability to sensitize cancer cells to the lethal effects of ionizing
radiation.
d6) d: 0.8, 0.9 [2s, 6H, 2CH3], 1.5 [s, 3H, COCH3], 1.9e2.2 [m, 4H,
4. Experimental
2CH2], 4.8 [s, 1H, CH], 7.3e8.0 [m, 8H, Ar-H þ SO2NH2], 9.9 [s, 1H,
NH]. Anal. Calcd. For C26H25ClN4O4S: C, 59.48; H, 4.80; N, 10.67.
Found: C, 59.31; H, 5.05; N, 10.49.
4.1. Chemistry
Melting points are uncorrected and were determined on a Stuart
melting point apparatus (Stuart Scientific, Redhill, UK). Elemental
analysis (C, H, N) were performed on Perkin-Elmer 2400 analyser
(Perkin-Elmer, Norwalk, CT, USA) at the microanalytical laborato-
ries of the Faculty of Science, Cairo University. All compounds were
within ꢁ0.4% of the theoretical values. The IR spectra (KBr) were
measured on Shimadzu IR 110 spectrophotometer (Shimadzu,
Koyoto, Japan), 1H NMR spectra were obtained on a Bruker proton
NMR-Avance 300 (300 MHz) (Bruker, Munich, Germany), in DMSO-
d6 as a solvent, using tetramethylsilane (TMS) as internal standard.
Mass spectra were run on HP Model MS-5988 (Hewlett Packard,
Palo, Alto, California, USA). All reactions were monitored by thin
layer chromatograph (TLC) using precoated Aluminium sheets
Silica gel Merck 60 F254 and were visualized by UV lamp (Merck,
Darmstadt, Germany).
4.1.5. N-acetyl-N-(4-(4-chlorophenyl)-3-cyano-7,7-dimethyl-5-
oxo-1-(4-sulfamoylphenyl)-1,4,5,6,7,8-hexahydroquinolin-2-yl)
acetamide (9)
A solution of compound 6 (0.482 g, 1 mmol) in acetic anhy-
dride (20 mL) was refluxed for 10 h, the reaction mixture was
then concentrated, the solid separated was recrystallized from
ethanol to give compound 9: Yield, 91%; m.p. 182e184 ꢂC; IR,
cmꢀ1: 3447, 3260, 3212 (NH, NH2), 3098 (CH arom.), 2959, 2873
(CH aliph.), 2213 (ChN), 1728, 1655 (3C¼O), 1370, 1164 (SO2). 1H
NMR (DMSO-d6) d: 0.7, 0.9 [2s, 6H, 2CH3], 1.8e2.3 [m, 4H, 2CH2],
2.4 [s, 6H, 2COCH3], 4.8 [s, 1H, CH], 7.2e8.0 [m, 10H, Ar-H þ
SO2NH2]. MS, m/z (%): 566 [M-1] (0.27), 90 (100). Anal. Calcd. For
C28H27ClN4O5S: C, 59.31; H, 4.80; N, 9.88. Found: C, 59.52; H,
4.58; N, 10.07.
4.1.6. 4-(5-(4-Chlorophenyl)-2,8,8-trimethyl-4,6-dioxo-3,4,6,7,8,9-
hexahydropyrimido[4,5-b]quinolin-10(5H)-yl)benzenesulfonamide (10)
A solution of compound 6 (0.482 g, 1 mmol) in acetic anhydride
(20 mL) was refluxed for 15 h, the reaction mixture was then
concentrated, the solid separated was recrystallized from ethanol
to give compound 10: Yield, 65%; m.p. 212e214 ꢂC; IR, cmꢀ1: 3466,
3290, 3200 (NH, NH2), 3062 (CH arom.), 2959, 2874 (CH aliph.),
1724, 1656 (2 C¼O), 1595 (C¼N), 1370e1163 (SO2). 1H NMR (DMSO-
4.1.1. 4-(5,5-Dimethyl-3-oxocyclohex-1-enylamino)
benzenesulfonamide (3)
Prepared according to the previously reported procedure [23].
4.1.2. 4-(2-Amino-4-(4-chlorophenyl)-3-cyano-7,7-dimethyl-5-
oxo-5,6,7,8-tetrahydroquinolin-1(4H)-yl)benzenesulfonamide (6)
A mixture of 4-(5,5-dimethyl-3-oxocyclohex-1-enylamino)ben-
zenesulfonamide 3 (2.94 g, 10 mmol) and 2-(4-chlorobenzylidene)
malononitrile 4 (1.88 g,10 mmol) in EtOH (20 mL) containing 3 drops
of triethylamine was refluxed for 6 h. The reaction mixture was
filtered while hot and the solid obtained was recrystallized from
d6) d: 0.7, 0.9 [2s, 6H, 2CH3], 1.9e2.2 [m, 4H, 2CH2], 2.4 [s, 3H, CH3],
4.9 [s, 1H, CH], 7.3e8.0 [m, 11H, Ar-H þ NH þ SO2NH2]. Anal. Calcd.
For C26H25ClN4O4S: C, 59.48; H, 4.80; N, 10.67. Found: C, 59.68; H,
4.66; N, 10.91.

3682
M.M. Ghorab et al. / European Journal of Medicinal Chemistry 45 (2010) 3677e3684
Table 2
In vitro anticancer screening of compounds 8 and 18 against human breast cell line (MCF7) in combination with g-radiation.
Compd. No.
Control
Irradiated (8 Gy)
Compound concentration (
m
M) þ Irradiation (8 Gy)
IC50 (mM)
10
25
50
100
Surviving fraction (Means ꢁ SE)#
8
18
1.000
1.000
0.927 ꢁ 0.02*
0.427 ꢁ 0.02*
0.653 ꢁ 0.02*
0.503 ꢁ 0.02*
0.517 ꢁ 0.02*
0.310 ꢁ 0.01*
0.281 ꢁ 0.01*
0.137 ꢁ 0.01*
0.282 ꢁ 0.01*
33.1
44.4
0.927 ꢁ 0.02*
# Each value is the mean of three values ꢁ Standard Error.
*
Significant difference from control group at p < 0.001.
4.1.7. 4-(4-Amino-5-(4-chlorophenyl)-8,8-dimethyl-6-oxo-6,7,8,9-
tetrahydropyrimido[4,5-b]quinolin-10(5H)-yl)benzenesulfonamide
(11)
from dioxane to give compound 13: Yield, 91%; m.p.164e166 ꢂC; IR,
cmꢀ1: 3371, 3300, 3251 (NH, NH2), 3096 (CH aliph.), 2959, 2882 (CH
aliph.), 2213 (ChN), 1705, 1644 (2 C¼O), 1371, 1162 (SO2). 1H NMR
A solution of compound 6 (0.482 g, 1 mmol) in formamide (30
mL) was refluxed for 5 h, the reaction mixture was cooled and then
poured onto cold water, the obtained solid was recrystallized from
dioxane to give compound 11. Yield, 49%; m.p. 149e151 ꢂC; IR,
cmꢀ1: 3421, 3338, 3263 (NH2), 3052 (CH arom.), 2952, 2862 (CH
aliph.), 1645 (C¼O), 1598 (C¼N), 1376, 1155 (SO2). MS, m/z (%): 510
[Mþ] (0.46), 87 (100). Anal. Calcd. For C25H24ClN5O3S: C, 58.88; H,
4.74; N, 13.73. Found: C, 59.11; H, 4.59; N, 13.92.
(DMSO-d6) d: 0.7, 0.9 [2s, 6H, 2CH3], 1.8e2.2 [m, 4H, 2CH2], 4.2 [s,
1H, CH], 4.7 [s, 2H, CH2Cl], 7.4e7.9 [m, 10H, Ar-H þ SO2NH2], 10.2 [s,
1H, NH]. MS, m/z (%): 559 [Mþ] (1.43), 63 (100). Anal. Calcd. For
C26H24Cl2N4O4S: C, 55.82; H, 4.32; N,10.01. Found: C, 55.92; H, 4.60;
N, 9.80.
4.1.10. 4-(2-(Chloromethyl)-5-(4-chlorophenyl)-8,8-dimethyl-4,6-
dioxo-3,4,6,7,8,9-hexahydropyrimido[4,5-b]quinolin-10(5H)-yl)
benzenesulfonamide (14)
4.1.8. Ethyl N-4-(4-chlorophenyl)-3-cyano-7,7-dimethyl-5-oxo-1-
(4-sulfamoylphenyl)-1,4,5,6,7,8-hexahydroquinolin-2-
ylformimidate (12)
A solution of compound 6 (0.482 g, 1 mmol) in triethylortho-
formate (30 mL) containing 3 drops of acetic anhydride was refluxed
for 8 h, the reaction mixture was cooled and then poured onto cold
water, the obtained solid was recrystallized from methanol to give
compound 12. Yield, 98%; m.p. 154e156 ꢂC; IR, cmꢀ1: 3354, 3251
(NH2), 3071 (CH arom.), 2966, 2886 (CH aliph.), 2194 (ChN), 1642
A mixture of compound 6 (0.482 g, 1 mmol) and chloroacetyl
chloride (0.112 g, 1 mmol) in Dimethyl formamide (20 mL) was
refluxed for 16 h. The reaction mixture was cooled and then poured
onto cold water and the solid obtained was recrystallized from
dioxane to give compound 14: Yield, 64%; m.p. >300 ꢂC; IR, IR,
cmꢀ1: 3460, 3359, 3249 (NH, NH2), 3063 (CH arom.), 2939, 2879
(CH aliph.), 1671, 1648 (2 C¼O), 1572 (C¼N), 1395, 1156 (SO2). 1H
NMR (DMSO-d6) d: 0.8, 0.9 [2s, 6H, 2CH3], 1.9e2.2 [m, 4H, 2CH2], 4.6
[s, 1H, CH], 5.1 [s, 2H, CH2Cl], 7.3e7.9 [m, 10H, Ar-H þ SO2NH2], 8.2
[s, 1H, NH]. Anal. Calcd. For C26H24Cl2N4O4S: C, 55.82; H, 4.32; N,
10.01. Found: C, 56.01; H, 4.19; N, 9.78.
(C¼O), 1590 (C¼N), 1378, 1175 (SO2). 1H NMR (DMSO-d6)
d: 0.8, 0.9
[2s, 6H, 2CH3],1.2 [t, 3H, CH3 ethyl], 1.9e2.2 [m, 4H, 2CH2], 4.3 [q, 2H,
CH2 ethyl], 4.6 [s, 1H, CH], 7.4e8.0 [m, 10H, Ar-H þ SO2NH2], 8.7 [s,
1H, N¼CH]. 13C NMR (CDCl3)
d: 15.2, 27.4, 33.2, 39.9, 44.0, 51.9, 63.5,
4.1.11. 4-(5-(4-Chlorophenyl)-2-(cyanomethyl)-8,8-dimethyl-4,6-
dioxo-3,4,6,7,8,9-hexahydropyrimido[4,5-b]quinolin-10(5H)-yl)
benzenesulfonamide (15)
76.6, 112.1, 117.2, 128.9, 131.1, 142.9, 153.5, 155.7, 161.6, 194.2. MS, m/z
(%): 539 [Mþ] (0.52), 90 (100). Anal. Calcd. For C27H27ClN4O4S: C,
60.16; H, 5.05; N, 10.39. Found: C, 59.95; H, 5.12; N, 10.59.
A mixture of compound 6 (0.482 g, 1 mmol) and ethyl cyanoa-
cetate (10 mL) was refluxed together for 5 h. The formed solid mass
was collected and recrystallized from dioxane to give compound
15: Yield, 82%; m.p. >300 ꢂC; IR, cmꢀ1: 3370, 3255, 3208 (NH, NH2),
3038 (CH arom.), 2962, 2890 (CH aliph.), 2214 (ChN), 1686, 1645 (2
4.1.9. 2-Chloro-N-(4-(4-chlorophenyl)-3-cyano-7,7-dimethyl-5-
oxo-1-(4-sulfamoylphenyl)-1,4,5,6,7,8-hexahydroquinolin-2-yl)
acetamide (13)
A mixture of compound 6 (0.482 g, 1 mmol) and chloroacetyl
chloride (0.112 g, 1 mmol) in Dimethyl formamide (20 mL) was
stirred at room temperature for 6 h. The reaction mixture was
poured onto cold water and the solid obtained was recrystallized
C¼O), 1621 (C¼N), 1365, 1159 (SO2). 1H NMR (DMSO-d6)
d: 0.8, 0.9
[2s, 6H, 2CH3], 1.9, 2.2 [2s, 4H, 2CH2], 4.1 [s, 1H, CH], 4.7 [s, 2H,
CH2CN], 7.3-8.0 [m, 9H, Ar-H þ NH], 12.5 [s, 2H, SO2NH2]. Anal.
1
1
0.922
0.653
Compound 18
1
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0
1
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0
0.948
Compound 8
Compound 18 + 8 Gy
Compound 8 + 8 Gy
0.779
0.503
0.662
0.517
0.452
0.282
0.445
0.281
0.446
0.427
20
0.324
0.137
0.31
0
40
60
80
100
0
20
40
Concentration (µM)
60
80
100
Concentration (µM)
Fig. 3. Survival curve for MCF7 cell line for compound 8 alone or in combination with
-irradiation (8 Gy).
Fig. 4. Survival curve for MCF7 cell line for compound 18 alone or in combination with
g-irradiation (8 Gy).
g

M.M. Ghorab et al. / European Journal of Medicinal Chemistry 45 (2010) 3677e3684
3683
Calcd. For C27H24ClN5O4S: C, 58.96; H, 4.40; N, 12.73. Found: C,
59.13; H, 4.19; N, 13.02.
was collected and recrystallized from dioxane to give compound
20: Yield, 90%; m.p. 138e140 ꢂC; IR, cmꢀ1: 3366, 3307, 3253 (NH,
NH2), 3095 (CH arom.), 2961, 2896 (CH aliph.), 2183 (ChN), 1727,
4.1.12. 4-(4-(4-Chlorophenyl)-3-cyano-2-(3-ethylthioureido)-7,7-
dimethyl-5-oxo-5,6,7,8-tetrahydroquinolin-1(4H)-yl)
benzenesulfonamide (16)
1702, 1644 (C¼O), 1338, 1166 (SO2). 1H NMR (DMSO-d6)
d: 0.7, 0.9
[2s, 6H, 2CH3], 1.9e2.1 [m, 4H, 2CH2], 2.2 [s, 3H, COCH3], 4.5 [s, 1H,
CH], 4.8 [s,1H, COCH2], 7.2e8.0 [m,10H, Ar-H þ SO2NH2],10.1 [s,1H,
NH]. Anal. Calcd. For C28H27ClN4O5S: C, 59.31; H, 4.80; N, 9.88.
Found: C, 59.52; H, 4.66; N, 9.58.
A mixture of compound 6 (0.482 g, 1 mmol) and ethyl iso-
thiocyanate (0.086 g, 1 mmol) in dimethyl formamide (20 mL) con-
taining 3 drops of triethylamine was refluxed for 10 h. The reaction
mixture was cooled and then poured onto cold water, the solid
obtained was recrystallized from ethanol to give compound 16: Yield,
94%;m.p.182e184 ꢂC;IR, cmꢀ1: 3412, 3347, 3281(NH, NH2), 3097 (CH
arom.), 2963, 2878 (CH aliph.), 2181 (ChN), 1645 (C¼O), 1256 (C¼S),
4.1.17. 4-(5-(4-Chlorophenyl)-8,8-dimethyl-6-oxo-4-thioxo-
3,4,6,7,8,9-hexahydropyrimido[4,5-b]quinolin-10(5H)-yl)
benzenesulfonamide (21)
A mixture of compound 7 (0.51 g, 1 mmol) and phosphorus
pentasulfide (0.22 g, 1 mmol) in pyridine (20 mL) was refluxed for 8
h, the reaction mixture was cooled and then poured onto cold
water, then acidified with dil HCl. The solid obtained was recrys-
tallized from ethanol to give compound 21: Yield, 81%; m.p.
182e184 ꢂC; IR, cmꢀ1: 3384, 3317, 3255 (NH, NH2), 3074 (CH arom.),
2959, 2875 (CH aliph.), 1660 (C¼O), 1252 (C¼S), 1374, 1165 (SO2).
MS, m/z (%): 527 [Mþ] (0.21), 458 (100). Anal. Calcd. For
C25H23ClN4O3S2: C, 56.97; H, 4.40; N, 10.63. Found: C, 56.80; H,
4.58; N, 10.89.
1369, 1136 (SO2). 1H NMR (DMSO-d6)
d: 0.7, 0.9 [2s, 6H, 2CH3], 1.2 [t,
3H, CH3 ethyl],1.8e2.2 [m, 4H, 2CH2], 3.2 [q, 2H, CH2 ethyl], 4.4 [s,1H,
CH], 7.2e7.8 [m, 10H, Ar-H þ SO2NH2], 8.2, 9.0 [2s, 2H, 2NH]. MS, m/z
(%): 570 [Mþ] (0.7), 78 (100). Anal. Calcd. For C27H28ClN5O3S2: C,
56.88; H, 4.95; N, 12.28. Found: C, 56.68; H, 4.75; N, 12.51.
4.1.13. 4-(4-(4-Chlorophenyl)-3-cyano-2-(2,5-dioxopyrrolidin-1-
yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydroquinolin-1(4H)-yl)
benzenesulfonamide (17)
A mixture of compound 6 (0.482 g, 1 mmol) and succinic anhy-
dride (0.15 g,1.5 mmol) was fused together in an oil bath at 250 ꢂC for
15 min, the fused mass was dissolved in dimethyl formamide and
poured onto cold water, the solid obtained was recrystallized from
4.1.18. 4-(4-Chloro-5-(4-chlorophenyl)-8,8-dimethyl-6-oxo-6,7,8,9-
tetrahydropyrimido[4,5-b]quinolin-10(5H)-yl)benzenesulfonamide (22)
A solution of compound 7 (0.51 g, 1 mmol) in thionyl chloride
(10 mL) was refluxed for 2 h, the thionyl chloride was then
removed by distillation, the obtained solid was washed twice
with benzene and recrystallized from dioxane to give compound
22: Yield, 98%; m.p. 228e230 ꢂC; IR, cmꢀ1: 3345, 3260 (NH2),
3090 (CH arom.), 2979, 2883 (CH aliph.), 1673 (C¼O), 1339, 1162
(SO2). MS, m/z (%): 531 [Mþ2] (0.8), 87 (100). Anal. Calcd. For
C25H22Cl2N4O3S: C, 56.71; H, 4.19; N, 10.58. Found: C, 56.92; H,
4.38; N, 10.79.
ethanol to give compound 17: Yield, 55%; m.p.194e196 ꢂC; IR, cmꢀ1
:
3360, 3249 (NH2), 3070 (CH arom.), 2959, 2876 (CH aliph.), 2213
(ChN), 1734, 1655 (3C¼O), 1369, 1165 (SO2). 1H NMR (DMSO-d6)
d:
0.7, 0.8 [2s, 6H, 2CH3],1.9e2.2 [m, 4H, 2CH2], 4.8 [s,1H, CH], 5.1 [t, 4H,
2CH2 pyrrolidine], 7.0e8.0 [m, 10H, Ar-H þ SO2NH2]. MS, m/z (%):
564 [M-1](0.6), 78 (100). Anal. Calcd. ForC28H25ClN4O5S:C, 59.52;H,
4.46; N, 9.92. Found: C, 59.35; H, 4.71; N, 9.62.
4.1.14. 2-Amino-4-(4-chlorophenyl)-7,7-dimethyl-5-oxo-1-(4-
sulfamoylphenyl)-1,4,5,6,7,8-hexahydroquinoline-3-carboxamide (18)
A solution of compound 6 (0.482 g, 1 mmol) in conc. H2SO4 (10
mL) was stirred for 6 h at room temperature, then the reaction
mixture was poured onto cold water. The obtained solid was
recrystallized from ethanol to give 18: Yield, 72%; m.p. 256e258 ꢂC;
IR, cmꢀ1: 3444, 3339, 3240 (NH2), 3069 (CH arom.), 2961, 2884 (CH
aliph.), 1706, 1677 (2 C¼O), 1376, 1165 (SO2). MS, m/z (%): 501 [Mþ]
(1.41), 458 (100). Anal. Calcd. For C24H25ClN4O4S: C, 57.54; H, 5.03;
N, 11.18. Found: C, 57.70; H, 4.81; N, 11.44.
4.1.19. Ethyl 2-(5-(4-chlorophenyl)-8,8-dimethyl-4,6-dioxo-10-(4-
sulfamoylphenyl)-6,7,8,9-tetrahydropyrimido[4,5-b]quinolin-3
(4H,5H,10H)-yl)acetate (23)
A mixture of compound 7 (0.51 g, 1 mmol), ethyl bromoacetate
(0.167 g, 1 mmol) and anhydrous potassium carbonate (1.5 g) was
refluxed in dry acetone for 24 h. The reaction mixture was filtered
while hot, and the filtrate was cooled then poured onto cold water.
The solid obtained was recrystallized from ethanol to give
compound 23: Yield, 85%; m.p. 88e90 ꢂC; IR, cmꢀ1: 3438, 3360
(NH2), 3092 (CH arom.), 2963, 2890 (CH aliph.), 1738, 1642 (C¼O),
4.1.15. 4-(2-Amino-4-(4-chlorophenyl)-3-(4,5-dihydro-1H-
imidazol-2-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydroquinolin-1
(4H)-yl)benzenesulfonamide (19)
1602 (C¼N), 1364, 1168 (SO2). 1H NMR (DMSO-d6)
d: 0.9, 1.0 [2s, 6H,
2CH3], 1.3 [t, 3H, CH3 ethyl], 1.9e2.2 [m, 4H, 2CH2], 4.1 [q, 2H, CH2
ethyl], 4.2 [s, 2H, CH2CO], 4.7 [s, 1H, CH], 7.1e7.9 [m, 10H, Ar-H þ
SO2NH2], 8.0 [s, 1H, N¼CH]. Anal. Calcd. For C29H29ClN4O6S: C,
58.34; H, 4.90; N, 9.38. Found: C, 58.60; H, 5.06; N, 9.09.
A mixture of compound 6 (0.482 g, 1 mmol) and ethylene
diamine (7 mL) was refluxed in carbon disulfide (7 mL) for 6 h. The
reaction mixture was cooled and then poured onto cold water. The
solid obtained was recrystallized from dioxane to give compound
19: Yield, 44%; m.p. 184e186 ꢂC; IR, cmꢀ1: 3400, 3319, 3200 (NH,
NH2), 3048 (CH arom.), 2930, 2867 (CH aliph.), 1643 (C¼O), 1591
4.1.20. 4-(5-(4-Chlorophenyl)-4-hydrazinyl-8,8-dimethyl-6-oxo-
6,7,8,9-tetrahydropyrimido[4,5-b]quinolin-10(5H)-yl)
benzenesulfonamide (24)
(C¼N), 1346, 1166 (SO2). 1H NMR (DMSO-d6)
d
: 0.9, 1.0 [2s, 6H,
A mixture of compound 22 (0.529 g, 1 mmol) and hydrazine
hydrate (0.1 mol) was refluxed in ethanol for 5 h. The reaction
mixture was cooled and poured onto cold water. The solid obtained
was recrystallized from dioxane to give compound 24: Yield, 69%;
m.p. 170e172 ꢂC; IR, cmꢀ1: 3359, 3322, 3253 (NH, NH2), 3092 (CH
arom.), 2968, 2890 (CH aliph.), 1655 (C¼O), 1590 (C¼N), 1334, 1160
2CH3], 1.9e2.2 [m, 4H, 2CH2], 4.5 [s, 1H, CH], 5.6 [s, 2H, NH2],
6.4e6.6 [m, 4H, 2CH2 imidazole], 7.1e8.0 [m, 10H, Ar-H þ SO2NH2],
8.3 [t, 1H, NH]. Anal. Calcd. For C26H28ClN5O3S: C, 59.36; H, 5.36; N,
13.31. Found: C, 59.18; H, 5.55; N, 13.50.
4.1.16. N-(4-(4-chlorophenyl)-3-cyano-7,7-dimethyl-5-oxo-1-(4-
sulfamoylphenyl)-1,4,5,6,7,8-hexahydroquinolin-2-yl)-3-
oxobutanamide (20)
A mixture of compound 6 (0.482 g, 1 mmol) and ethyl acetoa-
cetate (10 mL) was refluxed together for 5 h. The formed solid mass
(SO2). 1H NMR (DMSO-d6)
d: 0.9, 1.0 [2s, 6H, 2CH3], 1.9e2.2 [m, 4H,
2CH2], 4.5 [s, 1H, CH], 6.4 [s, 2H, NH2], 7.1e7.9 [m, 10H, Ar-H þ
SO2NH2], 8.0 [s, 1H, NH], 8.3 [s, 1H, N¼CH]. Anal. Calcd. For
C25H25ClN6O3S: C, 57.19; H, 4.80; N, 16.01. Found: C, 57.47; H, 4.51;
N, 16.23.

3684
M.M. Ghorab et al. / European Journal of Medicinal Chemistry 45 (2010) 3677e3684
4.1.21. 4-(5-(4-Chlorophenyl)-4-isothiocyanato-8,8-dimethyl-6-
oxo-6,7,8,9-tetrahydropyrimido[4,5-b]quinolin-10(5H)-yl)
benzenesulfonamide (25)
evaluate the ability of these compounds to enhance the cell killing
effect of -radiation.
Cells were subjected to a single dose of
g
g
-radiation at a dose
A mixture of compound 22 (0.529 g, 1 mmol) and Ammonium
thiocyanate (0.076 g, 1 mmol) was refluxed in dry acetone for 1 h.
The reaction mixture was cooled and poured onto cold water. The
solid obtained was recrystallized from ethanol to give compound
25: Yield, 52%; m.p. 200e202 ꢂC; IR, cmꢀ1: 3345, 3200 (NH2), 3094
(CH arom.), 2976, 2883 (CH aliph.), 2023 (N¼C¼S), 1666 (C¼O),
1610 (C¼N), 1341, 1163 (SO2). MS, m/z (%): 553 [Mþ1] (0.85), 90
(100). Anal. Calcd. For C26H22ClN5O3S2: C, 56.57; H, 4.02; N, 12.69.
Found: C, 56.71; H, 4.30; N, 12.48.
level of 8 Gy with a dose rate of 2 Gy/min. Irradiation was per-
formed in the National Cancer Institute, Cairo University, using
Gamma cell-40 (60Co) source.
The surviving fractions were expressed as means ꢁ standard
error. The results were analyzed using 1-way ANOVA test and given
in Table 2.
References
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4.2. In vitro anticancer screening
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Human tumor breast cell line (MCF7) was used in this study. The
cytotoxic activity was measured in vitro for the newly synthesized
compounds using the Sulfo-Rhodamine-B stain (SRB) assay using
the method of Skehan et al. [27]. The in vitro anticancer screening
was done by the pharmacology unit at the National Cancer Insti-
tute, 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. Test compounds were dissolved in dimethyl
sulfoxide. Different concentrations of the compound under test (10,
25, 50, and 100 mM) were added to the cell monolayer. Triplicate
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Targets 2 (2002) 55e75.
wells were prepared for each individual concentration. Monolayer
cells were incubated with the compound(s) for 48 h at 37 ꢂC and in
atmosphere of 5% CO2. After 48 h, cells were fixed, washed and
stained for 30 min with 0.4% (wt/vol) SRB dissolved in 1% acetic
acid. Excess unbound dye was removed by four washes with 1%
acetic acid and attached stain was recovered with TriseEDTA buffer.
Color intensity was measured in an ELISA reader. The relation
between surviving fraction and drug concentration is plotted to get
the survival curve for breast tumor cell line after the specified time.
The molar concentration required for 50% inhibition of cell viability
(IC50) was calculated and compared to the reference drug doxoru-
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The most potent compounds resulted from the in vitro anti-
cancer screening; the quinoline derivatives 8 and 18, were selected
to be evaluated again for their in vitro anticancer activity alone and
in combination with
g-radiation. This study was conducted to