Design, synthesis, anti-schistosomal activity and molecular docking of novel 8-hydroxyquinoline-5-sufonyl 1,4-diazepine derivatives

Article abstract of DOI:10.1016/j.bioorg.2012.10.003

Schistosomiasis remains one of the most prevalent parasitic infections and has significant public health consequences. Praziquantel (PZQ) is the only drug currently administrated to treat this disease. However, praziquantel-resistant parasites have been identified in endemic areas and can be generated in the laboratory. Therefore, it is essential to find new therapeutics. Herein we report a series of novel 8-hydroxyquinoline-5-sufonyl 1,4-diazepine derivatives, which were synthesized, characterized and tested as anti-schistosomal agents in vitro. Among all tested compounds, compounds 4a, 5b, and 7b at different tested concentrations (50, 100, and 200 μg/mL) showed the highest schistosomicidal activity. Among those 3 compounds, compound 7b was the most potent anti-schistosomal one. Moreover, all tested compound, at 50 μg/mL concentration, significantly reduced oviposition of adult worms in vitro. Furthermore, both compound 4a and 7b, as well as compound 6a, completely diminished egg deposition. To clarify the possible mechanism by which novel 8-hydroxyquinoline-5-sufonyl 1,4-diazepine derivatives act as anti-schistosomal agents, molecular docking of all new compounds was carried out using Molsoft ICM pro 3.5-0a to investigate the binding affinity and binding mode to thioredoxin glutathione reductase enzyme (TGR), a potential drug target for anti-schistosomal agents. The docking results revealed moderate to high affinity of the new compounds towards TGR. Compound 7b scored the highest binding energy (-101.13 kcal/mol) against TGR crystal structure forming eight hydrogen bonds with the amino acid residues at the binding site of the receptor. This result indicates that compound 7b could exert its effect through inhibition of TGR, which is a vital enzyme for schistosome survival.

Full text of DOI:10.1016/j.bioorg.2012.10.003

Bioorganic Chemistry 46 (2013) 17–25
Contents lists available at SciVerse ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Design, synthesis, anti-schistosomal activity and molecular docking of novel
8-hydroxyquinoline-5-sufonyl 1,4-diazepine derivatives
Ahmad F. Eweas a,b, , Gamal Allam c,d, Abdelaziz S.A. Abuelsaad c,d, Abdul Hamid ALGhamdi c,
⇑
Ibrahim A. Maghrabi b
a Department of Medicinal Chemistry, National Research Center, Dokki, Cairo, Egypt
b College of Pharmacy, Taif University, Taif, Saudi Arabia
c College of Medicine, Taif University, Taif, Saudi Arabia
d Department of Zoology, Faculty of Science, Beni-Suef University, Beni-Suef, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 31 July 2012
Available online 8 November 2012
Schistosomiasis remains one of the most prevalent parasitic infections and has significant public health
consequences. Praziquantel (PZQ) is the only drug currently administrated to treat this disease. However,
praziquantel-resistant parasites have been identified in endemic areas and can be generated in the
laboratory. Therefore, it is essential to find new therapeutics. Herein we report a series of novel
8-hydroxyquinoline-5-sufonyl 1,4-diazepine derivatives, which were synthesized, characterized and
tested as anti-schistosomal agents in vitro. Among all tested compounds, compounds 4a, 5b, and 7b at
Keywords:
8-Hydroxyquinoline
1,4-Diazepines
Anti-schistosoma
Molecular docking
Thioredoxin-glutatione reductase (TGR)
different tested concentrations (50, 100, and 200
Among those 3 compounds, compound 7b was the most potent anti-schistosomal one. Moreover, all
tested compound, at 50 g/mL concentration, significantly reduced oviposition of adult worms in vitro.
lg/mL) showed the highest schistosomicidal activity.
l
Furthermore, both compound 4a and 7b, as well as compound 6a, completely diminished egg deposition.
To clarify the possible mechanism by which novel 8-hydroxyquinoline-5-sufonyl 1,4-diazepine deriva-
tives act as anti-schistosomal agents, molecular docking of all new compounds was carried out using
Molsoft ICM pro 3.5-0a to investigate the binding affinity and binding mode to thioredoxin glutathione
reductase enzyme (TGR), a potential drug target for anti-schistosomal agents. The docking results
revealed moderate to high affinity of the new compounds towards TGR. Compound 7b scored the highest
binding energy (ꢀ101.13 kcal/mol) against TGR crystal structure forming eight hydrogen bonds with the
amino acid residues at the binding site of the receptor. This result indicates that compound 7b could exert
its effect through inhibition of TGR, which is a vital enzyme for schistosome survival.
Ó 2012 Elsevier Inc. All rights reserved.
1. Introduction
populations [3]. Praziquantel (PZQ, Fig. 1) is the only drug of choice
for treatment of schistosomiasis. PZQ is administered to millions of
Human schistosomiasis, or bilharzia, is considered one of the
most significant and neglected tropical diseases in the world. This
parasitic disease ranks second after malaria in terms of its public
health importance [1]. Different approaches have been used to
control schistosomiasis transmission including educational pro-
grams, mass chemotherapy, biological and chemical control for
the snail intermediate host. However, despite these control pro-
grams, the disease still affects more than 200 million people world-
wide and approximately 800 million people remain under
infection risk [2]. Moreover, schistosomiasis continues to spread
to new geographic areas due to environmental changes that result
from the development of water resources, growth and migration of
people yearly [4,5]. They are rapidly re-infected and must be re-
treated on an annual or semiannual basis. If praziquantel-resistant
parasites develop, treatment for schistosomiasis will be in a crisis
state [6,7]. Furthermore, the use of such drug for treatment is com-
plicated due to the difficulties and expenses involved in a long-
lasting maintenance of these programs [8]. These limitations, in
combination with a considerable concern about the development
of PZQ resistance, have motivated the scientific community to call
for research and development of novel and inexpensive drugs
against schistosomiasis [9–11]. A reliable alternative to PZQ does
not exist at the moment. Oxamniquine (Fig. 1), the only other drug
commercially available, is expensive and is active against only one
of the main three schistosome species capable of infecting humans,
Schistosoma (S.) mansoni. Artemisinins, although safe, are active
only against the immature stages of the parasites. A treatment
strategy that relies on just a single drug is at risk given the high
⇑
Corresponding author at: Department of Medicinal Chemistry, National
Research Center, Dokki, Cairo, Egypt.
E-mail address: eweas1@gmail.com (A.F. Eweas).
0045-2068/$ - see front matter Ó 2012 Elsevier Inc. All rights reserved.
http://dx.doi.org/10.1016/j.bioorg.2012.10.003

18
A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
Fig. 1. Anti-schistosomal drugs.
probability that drug-resistant parasites will emerge [12,13]. Based
on these findings, the identification of new and effective schistos-
omicidal compounds is in great demand.
protection from oxidative stress, makes it a promising drug target
[25]. Therefore, TGR is an essential protein for the survival of S.
mansoni and that it meets all the major criteria of an important
target for anti-schistosomal chemotherapy development. There-
fore the current study was conducted to synthesis a more potent
anti-schistosomal 8-hydroxyquinoline-5-sufonyl 1,4-diazepine
derivatives which may have a high binding energy with TGR.
8-Hydroxyquinoline derivatives are well known as powerful
iron chelatores with antioxidant property. 8-Hydroxyquinoline
derivatives are important constituents in a variety of pharmaceuti-
cally important compound classes. They have become of interest as
a new class of potent HIV-1 integrase inhibitors [14] in modeling of
the inhibition of retroviral integrases [15], protein tyrosine kinase
inhibitors, anti-protozoal and retroviral co-infections [16], anti-
HIV-1 agents [17], anti-leishmanial [18] and anti-filarial agents
[19].
2. Results and discussion
2.1. Chemistry
It has been reported that 8-hydroxyquinoline derivatives have
anti-schistosomal activity [20,21]. In our earlier study [22], two
novel 8-hydroxyqinoline-5-sulfonamido derivatives (3-amino-1-
((8-hydroxyquinolin-5-yl)sulfonyl)-1H-pyrazol-5(4H)-one and1-
((8-hydroxyquinolin-5-yl)sulfonyl)-3-methyl-1H-pyrazol-5(4H)-one)
The study started with protection of 8-hydroxyquinoline-5-sul-
fonic acid with tert-butyl dimethyl silyl chloride (TBDMS) in pres-
ence of imidazole to afford the protected compound 1 in high yield.
Compound 1 was then reacted with thionyl chloride in presence of
anhydrous DMF to afford the corresponding sulfonyl chloride 2.
Compound 2 was then reacted with b-diketones namely acetyl ace-
tone and 1,3-diphenylpropane-1,3-dione in presence of sodium
methoxide which yielded corresponding protected substituted
b-diketones 3a and b (scheme 1).
Compounds 3a and b was then deprotected using 6N HCl to
afford the free 8-hydroxyquinoline 5-sulfonyl-b-diketones 4a and
b in high yield. The structure of compounds 4a and b was con-
firmed via 1H NMR showing signals at d 2.13 (s, 6H) dimethyl
groups and d 5.62 (s, 1H) SO2–CH for 4a and d 6.43 (s, 1H)
SO2–CH for 4b. Compounds 4a and b were then condensed with
diamines namely o-phenylendiamine, ethylenediamine and
3,4-diaminopyridine in ethyl acetate in presence of silica sulfuric
acid (SSA) [26] (Scheme 2) to afford the corresponding 8-hydroxy-
quinoline-5-sufonyl 1,4-diazepine derivatives 5a and b, 6a and b
and 7a and b respectively.
at 200
activity against S. mansoni adult worms in vitro. However, at lower
concentrations (50 and 100 g/mL) both compounds showed a
lg/mL concentration showed powerful anti-schistosomal
l
weak schistosomicidal activity in comparison with PZQ. The anti-
schistosomal activity of such compounds was attributed to inhibi-
tion of thioredoxin glutathione reductase (TGR) enzyme. In Schisto-
soma, thiol redox homeostasis is completely dependent on the TGR
enzyme, which directs the NADPH reducing equivalents to both
GSH and thioredoxin [23]. This reliance on a single enzyme for
both glutathione disulfide and thioredoxin reduction suggests that
the parasite’s redox systems are subject to a bottleneck depen-
dence on TGR. Given the importance of cellular redox systems
and the biochemical differences between the redox metabolism
of S. mansoni and its human host, it was hypothesized that TGR
could be an essential parasite protein and a potentially important
drug target. The significance of S. mansoni TGR as a putative drug
target was first demonstrated using an RNA interference approach,
which killed 90% of treated parasites in vitro [24]. Moreover, treat-
ment of S. mansoni with a TGR inhibitor, auranofin, resulted in
100% mortality [13]. It was demonstrated that TGR activity is
inhibited by two schistosomicidal drugs used in the past to fight
the infection, antimonyl potassium tartrate and oltipraz, suggest-
ing that the enzyme is the main target of these compounds [24].
The dependence of S. mansoni on a single protein, TGR, for its
2.2. Molecular docking study
In S. mansoni, thiol redox homeostasis is completely dependent
on the enzyme thioredoxin-glutathione reductase (TGR), which di-
rects the NADPH reducing equivalents to both GSH and thioredoxin
[13,24,25]. Characterization of the parasite TGR revealed enzy-
matic properties that differed from those of mammalian TGR,

A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
19
Scheme 1. Reagents and conditions: (a) TBDMS, imidazole, room temp., (b) thionyl chloride, DMF, reflux; (c) NaOMe, b-diketones, stirring, 50 °C; and (d) 6N HCl, reflux.
Scheme 2.
thioredoxin (TrxR) and glutathione reductase (GR) enzymes. TGR
was also identified as a multifunctional oxidoreductase with a
remarkably wide substrate specificity, capable of directly reducing
peroxides, selenium-containing compounds, and several important
low-molecular-weight antioxidants, as well as Trx, 5,50-dithiobis 2-
nitrobenzoic acid (DTNB), reduces glutathione disulfide (GSSG),

20
A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
Table 1
The docking energy scores of compounds 4a–7b with the amino acid residues forming hydrogen bonds in comparison with reference ligand praziquantel.
Cpd. no.
Docking score (kcal/mol)
No. of hydrogen bonds
3
Amino acid residues forming hydrogen bonds in A0
Ligand (praziquantel)
ꢀ84.13
S117 hg – m M o2: 2.54 A
R393 he – m M o1: 2.02 A
R393 hh22 – m M o1: 1.83 A
4a
ꢀ93.36
4
T153 hg1 – m M o3: 2.74 A
K227 hz1 – m M o1: 2.58 A
K227 hz2 – m M o1: 2.14 A
R260 hh11 – m M o2: 2.33 A
5a
5b
ꢀ91.74
ꢀ88.85
2
5
T153 hg1 – m M n2: 1.97 A
R260 hh11 – m M o2: 2.49 A
G116 hn – m M o1: 2.42 A
S117 hn – m M o1: 2.42 A
Y138 hn – m M n2: 2.17 A
T153 hn – m M n1: 2.38 A
G115 o – m M h6: 2.66 A
6a
6b
ꢀ86.71
ꢀ99.01
3
5
T153 hn – m M n3: 2.78 A
T153 hg1 – m M n3: 2.41 A
E140 oe1 – m M h17: 2.33 A
E140 hn – m M n2: 2.75 A
T153 hn – m M o1: 1.64 A
T153 hg1 – m M o1: 1.15 A
R260 hh11 – m M n2: 2.07 A
T153 og1 – m M h6: 1.31 A
7a
7b
ꢀ100.21
ꢀ101.13
5
8
Y138 hn – m M o2: 2.18 A
E140 hn – m M o3: 1.57 A
G228 hn – m M n1: 2.04 A
R260 hh11 – m M n3: 2.26 A
G228 o – m M h6: 2.02 A
Y138 hn – m M o2: 2.45 A
Y138 hn – m M n4: 2.15 A
E140 hn – m M o3: 1.43 A
K227 hz1 – m M n1: 2.38 A
K227 hz1 – m M o1: 2.10 A
K227 hz2 – m M n1: 2.46 A
K227 hz2 – m M o1: 2.54 A
G228 o – m M h6: 2.37 A
and glutathione–b-hydroxyethyl disulfide (GSH–HED) [24]. This
dependence of S. mansoni on TGR for its protection from oxidative
stress, makes it a promising drug target [25]. Based on these find-
ing we decided to dock our new 8-hydroxyquinoline derivatives
against TGR and compare it with the reference drug praziquantel.
Molecular docking studies were carried out using Mol soft ICM
3.5-0a. The aim of the flexible docking calculations is the predic-
tion of correct binding geometry for each binder. The scoring func-
tions and hydrogen bonds formed with the surrounding amino
acids of the receptor TGR are used to predict tested compounds
binding modes. We evaluated the new compounds 4a, 5b, and 7b
(Table 1) through molecular modeling and docking techniques
against TGR crystal structure which was downloaded from PDB
website PDB id (2V6O).
score of ꢀ101.13 kcal/mol forming eight hydrogen bonds with TGR
binding site at Tyr-138, Glu-140, Lys-227 and Gly-228.
2.3. Anti-schistosomal activity
2.3.1. Effect of 8-hydroxyquinoline derivatives on adult schistosomes
survival
The survival of 56-day-old adult worms of S. mansoni was as-
sessed in vitro by incubation with different concentrations of
new 8-hydroxyquinoline derivatives. The effect of 8-hydroxyquin-
oline derivatives on the mortality rate of both male and female
adult worms was analyzed with respect to the concentration and
incubation time. Compounds 4b, 5a, 6a, 6b, and 7a at 50, 100,
and 200
al effect until 96 h post-incubation. However, only compounds 4b,
5a, and 6a at 200 g/mL concentration caused 50% death of both
lg/mL concentrations did not showed any schistosomicid-
Fig. 2 shows binding mode of the reference drug (Praziquantel),
4a, 5b, and 7b into its binding site of TGR. Praziquantel ‘‘the refer-
ence drug’’ docking results to TGR reveals docking score of
l
male and female worms after 120 h post-incubation. On the other
hand, as depicted in Table 2, 4a caused 100% death of all worms at
D
G = ꢀ84.13 kcal/mol and three hydrogen bonds to the active site
with Ser-117 and Arg-393. on the other hand compound 4a dock-
ing result against TGR shows docking score of ꢀ93.36 kcal/mol
forming four hydrogen bonds with TGR binding site at Thr-153,
Lys-227 and Arg-260. On the other hand compound 5b docking re-
sults revealed a binding energy of ꢀ88.85 kcal/mol, forming five
hydrogen bonds with binding site of TGR at Gly-115, Gly-116,
Ser-117, Tyr-138 and Thr-153.
a concentration of 100 and 200
lg/mL after 96 and 72 h, respec-
tively. However, the 50 g/mL concentration of 4a caused only
l
50% and 40% death of male and female worms, respectively, after
120 h of incubation. Interestingly, both 5b and 7b at 50 and
100
lg/mL concentrations caused 100% death of all worms after
72 h and 48 h, respectively. Moreover, the 200
lg/mL concentra-
tion of 7b resulted in 75% and 100% death of male and female
While compound 6a docking result against TGR shows docking
score of ꢀ86.71 kcal/mol forming three hydrogen bonds with TGR
binding site at Glu-140 and Thr-153. Compound 7b showed the
highest binding energy among all tested compounds with docking
worms, respectively, after 24 h of incubation.
Subsequently, the effect of new 8-hydroxyquinoline derivatives
on the motor activity of the worms was examined, and a dose-
dependent reduction of activity was observed in compounds 4a,

A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
21
Fig. 2. Molecular docking and binding mode of tested compounds PZQ (reference drug), 4a, 5b, and 7b to the active binding site of TGR receptor.
5b, and 7b (Table 2). However, 4b, 5a, 6a, 6b, and 7a did not
showed any significant decrease in motor activity of adult parasites
until 72 h post-incubation.
In addition to the mortality rate and changes in the motor
capacity of S. mansoni adults, the results highlighted the effect of
8-hydroxyquinoline derivatives on the parasite’s tegument. As
shown in Table 2, only partial morphological alterations of the teg-
ument occurred in a dose-dependent manner after incubation with
compounds 4b and 7b. No extensive tegumental alterations were
observed in all worms incubated with any compound. In contrast,
which has the highest binding energy with TGR, is more effective
in killing of S. mansoni adult worms than other compounds. Sec-
ond: it is well documented that 8-hydroxyquinoline is a potent
chelator to various metals, especially iron [27,28]. Iron (Fe) is an
important trace element found in nearly all organisms, and is used
as a cofactor in many biological reactions. Schistosomes require Fe
for development and maintain their life [29]. Lipid solubility of the
neutral iron-8-hydroxyquinoline complex ensures penetration of
epithelial cells and distribution within the cells [30]. It was shown
that iron bound to 8-hydroxyquinoline causes oxidative damages
(via free radical lipid peroxidation) in living cells [30,31] as well
as DNA strand breakage in cultured cells [31]. The Fungicidal and
bactericidal action of 8-hydroxyquinoline derivatives has been
attributed in part to its ability to chelate essential trace metals
[32]. Taken together, our new 8-hydroxyquinoline derivatives
could exert its schistosomicidal effect through inhibition of TGR
activity and Fe chelation.
10 lg/mL PZQ group had tegumental alteration in all the worms
and severe tegumental alteration were more pronounced in male
than in female adults. Meanwhile, no tegumental changes in adult
worms were observed in the negative control group.
From previously mentioned data, compounds 4a, 5b, and 7b
have a potent schistosomicidal activity, and compound 7b is the
most efficient one. The exact mechanism by which such com-
pounds exert their in vitro schistosomicidal effect is not clear.
However, there are two possible mechanisms; first: compounds
7b has a high binding energy (
D
G = ꢀ101.33 kcal/mol), and eight
2.3.2. Effect of 8-hydroxyquinoline derivatives on the reproductive
fitness of S. mansoni
In order to evaluate the egg production by adult worms of S.
mansoni, 8-hydroxyquinoline derivatives were tested at 50 lg/mL
concentration. As shown in Fig. 3, worm pairs incubated with 5b,
6a, and 7b did not produce any eggs until 72 h post-incubation.
However, worm pairs incubated with 4a, 4b, 5a, and 7a showed
about 50% significant reduction in oviposition after 24 h, 48, and
72 h of incubation in comparison with the negative control group.
Unexpected, worm pairs incubated with 6b showed significant
hydrogen bonds at the active binding site of TGR, as mentioned
previously. Such binding could inhibit the activity of TGR, schisto-
some antioxidant enzyme, which in turn lead to inhibition of the
antioxidant response of the parasite that can lead to parasite death.
This assumption supported by the previous studies that showed
silencing of TGR expression lead to death of S. mansoni in vitro
within 4 days [24]. Moreover, treatment of S. mansoni with a TGR
inhibitor resulted in 100% mortality of the parasite after 9 h of
exposure in vitro [13]. This finding explains why compound 7b,

22
A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
Table 2
In vitro effects of 8-hydroxyquinoline derivatives against 56-day-old adult S. mansoni.
Groups
Drug concentration
Incubation period (h)
Dead worms (%)
Motor activity reduction (%)
Worms with tegumental alteration (%)
Slight
M
Significant
Partial
M
Extensive
M F
M
F
F
M
F
F
Control
RPMI 1640 + 1% DMSO
24
48
72
96
120
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
PZQ
10
50
l
g/mL
g/mL
24
100
100
0
0
0
10
30
30
0
100
100
20
35
80
65
4a
l
24
48
72
96
120
0
0
0
25
50
0
0
0
0
40
10
20
20
25
10
0
0
30
50
100
0
0
10
40
90
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
100
200
l
g/mL
g/mL
24
48
72
96
0
25
50
0
0
40
20
50
0
10
20
20
0
30
50
50
20
40
80
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
100
100
0
100
100
l
24
48
72
0
60
100
0
80
100
50
0
0
0
100
100
50
100
100
100
0
0
0
0
0
0
0
0
0
0
0
0
0
5b
50
lg/mL
24
48
72
0
40
100
0
45
100
55
0
0
45
0
0
20
100
100
30
100
100
0
0
0
0
0
0
0
0
0
0
0
0
100
200
l
g/mL
g/mL
24
48
0
100
0
100
30
0
10
0
70
100
90
100
10
15
10
25
0
0
0
0
l
24
48
0
100
0
100
0
0
0
0
100
100
100
100
10
20
15
20
0
0
0
0
7b
50
lg/mL
24
48
72
10
35
100
10
70
100
40
20
0
55
0
0
20
80
100
45
100
100
0
10
20
0
15
20
0
0
0
0
0
0
100
200
l
g/mL
g/mL
24
48
20
100
50
100
30
0
20
0
70
100
80
100
10
20
10
20
0
0
0
0
l
24
48
75
100
100
100
10
0
0
0
90
100
100
100
10
20
20
20
0
0
0
0
Worm motor activity and survival of the parasites was monitored under a stereomicroscope. Tegumental alterations were monitored using an inverted microscope. The
effects of the compounds on motor activity and tegumental alterations of 20 adult S. mansoni worms were assessed qualitatively from five separate experiments.
increase in oviposition after 48 h and 72 h of incubation in compar-
ison with the negative control group.
Considering the strong lethal effect of 8-hydroxyquinoline
derivatives on adult schistosomes, the in vitro oviposition was con-
tinually monitored to assess the sexual fitness of treated worms.
study is required to evaluate schistosomicidal effect of compound
4a, 5b, and 7b on S. mansoni adult worms and their induced
pathology.
From the aforementioned data, 50 lg/mL concentration of all com-
3. Experimental
pounds, except compound 6b, showed a potent reduction in ova
deposition. This effect could be attributed to chelation of Fe by 8-
hydroxyquinoline derivatives. Schistosomes store abundant Fe in
vitelline (eggshell-forming) cells of the female system. Schisto-
somes synthesize eggshells from among a range of precursor’s pro-
teins belonging to tyrosine-rich proteins, the P14, P19, and P48
proteins [33]. The eggshell is formed by enzymatic oxidation of
the tyrosine residues to dihydroxyphenylalanine-quinones, which
then bind lysine residues of adjacent proteins [34,35] to form a
rigid structure. Fe plays an important role in cross-linking and
stabilization of dihydroxyphenylalanine-rich proteins [29,36,37].
8-hydroxyquinoline and its derivatives have been reported as
potent Fe chelator [38]. Consequently, depravation of Fe leads to
disruption of egg formation. However, mechanism by which com-
pound 6b increases oviposition is unclear and needs further study.
In conclusion, our results indicate that 8-hydroxyquinoline
derivatives possess in vitro schistosomicidal activity against
S. mansoni adult worms and may be novel broad-spectrum
anti-schistosomal drug candidates. For future work, an in vivo
3.1. Chemistry
All starting materials and reagents were purchased from Sig-
ma–Aldrich, BDH and Fluka and used without further purification.
All solvents were either of analytical grades or dried and distilled
immediately prior to use. All of the reactions were performed using
oven-dried glassware. Melting points were measured on a Stuart-
SMP10 melting point apparatus and are uncorrected. TLC was per-
formed using Merck precoated Silica gel 60 F254 aluminum sheets
(20 ꢁ 20 cm, layer thickness 0.2 mm) and Merck precoated Silica
gel aluminum sheets (20 ꢁ 20 cm, layer thickness 0.2 mm) and
spots were visualized by UV (254 nm), KMNO4 solution and/or
charring with H2SO4–EtOH (5% v/v). All reaction products were
stored refrigerated under 4 °C. The 1H NMR spectra were recorded
(DMSO-d6) with Varian 400-MR system, Chemical shifts are
expressed in parts per million (ppm) and reported either relative
to an internal tetramethyl-silane standard (TMS d = 0.0) or relative
to solvent peaks. High resolution mass spectra (HRMS) were

A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
23
Fig. 3. In vitro effect of 8-hydroxyquinoline derivatives on Schistosoma mansoni oviposition. Adult worm couples were incubated with 50
l
g/mL of either compound 4a, 4b,
5a, 5b, 6a, 6b, 7a, or 7b and at the indicated time periods, the cumulative number of eggs per worm couple was assessed and scored using an inverted microscope. Values are
means SD (bars) of 10 worm couples. ꢂP < 0.05 and ꢂꢂP < 0.001 compared with control group (RPMI 1640 + 1% DMSO).
recorded on Bruker Maxis spectrometer. Elemental analyses were
recorded with EuroVector EA3000 Elemental Analyzer.
reaction, the mixture was cooled and toluene was removed under
reduced pressure. The reaction mixture was extracted using chlo-
roform and washed with water. The chloroform layer was dried
using anhydrous sodium sulfate and distilled to yield the solid
compound. The product was purified by column chromatography
over silica gel using pet ether:ethyl acetate (1:2) as an eluent.
3.1.1. 8-((tert-Butyldimethylsilyl)oxy)quinoline-5-sulfonic acid 1
A 100-mL flask was successively loaded with dry CH2Cl2
(50 mL), 8-hydroxy-5-sulfonic acid (7.2 g, 0.032 mol), imidazole
(2.29 g, 0.034 mol, 1.05 equiv), and tert-butyldimethylsilyl chloride
(5.31 g, 0.035 mol, 1.1 equiv). The solution was stirred at room
temperature for 10 h, diluted with Et2O (500 mL), washed with
aqueous HCl (0.1 M, 50 mL), brine (100 mL), water (100 mL), dried
over K2CO3 and evaporated under reduced pressure to afford
10.32 g (95%) of protected product as a colorless oil. 1H NMR
(400 MHz, DMSO-d6) d 0.29 (s, 6H); d 1.09 (s, 9H); 7.23 (d, 1H,
J = 8.3); 7.66–7.73 (m, 2H); 7.94 (dd, 1H, J = 1.3, 5.3); 8.71 (dd,
1H, J = 1.3, 8.4); HRMS (M)+ calcd for C15H21NO4SSi 339.0960.
Found: 339.1034. Elemental analysis, calcd: C, 53.07; H, 6.24; N,
4.13; O, 18.85; S, 9.45. Found: C, 53.09; H, 6.25; N, 4.15; O,
18.87; S, 9.43.
3.2.1. 2-((8-((tert-Butyldimethylsilyl)oxy)quinolin-5-yl)sulfonyl)-1,3-
dimethylpropane-1,3-dione 3a
Colorless oil, yield 72%; 1H NMR (400 MHz, DMSO-d6) d 0.29 (s,
6H); d 1.09 (s, 9H); d 2.13 (s, 6H); 5.62 (s, 1H); 7.31–8.89 (m, 5H
aromatic protons); HRMS (M)+ calcd for C20H27NO5SSi 421.1379.
Found: 421.1425. Elemental analysis, calcd: C, 56.98; H, 6.46; N,
3.32; O, 18.98; S, 7.61. Found: C, 56.95; H, 6.48; N, 3.31; O,
18.95; S, 7.65.
3.2.2. 2-((8-((tert-Butyldimethylsilyl)oxy)quinolin-5-yl)sulfonyl)-1,3-
diphenylpropane-1,3-dione 3b
3.1.2. 8-((tert-Butyldimethylsilyl)oxy)quinoline-5-sulfonyl chloride 2
In a 100 mL round-bottom flask fitted with magnetic stirrer,
condenser and N2 inlet were placed 8-((tert-butyldimethylsi-
lyl)oxy)quinoline-5-sulfonic acid 1 (10.32 g, 0.03 mol), 60 mL of
SOCl2, and 1 mL of anhydrous DMF. The suspension was heated
to reflux for 3 h, and complete dissolution was achieved after
30 min. After 1 h, product began to precipitate from the reaction
mixture. After 3 h, the reaction mixture was cooled and transferred
to a 250 mL flask before stripping off the SOCl2. The remaining
SOCl2 was removed by successive azeotropic distillations with CH2-
Cl2 and toluene on a rotary evaporator. The crude sulfonyl chloride
was used for next reaction without further purification.
Colorless oil, yield 62%;1H NMR (400 MHz, DMSO-d6) d 0.29 (s,
6H); d 1.09 (s, 9H); d 6.43 (s, 1H); d 7.31–8.89 (m, 15H, aromatic
protons); HRMS (M)+ calcd for C30H31NO5SSi 545.1692. Found:
545.1632. Elemental analysis, calcd: C, 66.03; H, 5.73; N, 2.57; O,
14.66; S, 5.88. Found: C, 65.98; H, 5.76; N, 2.58; O, 14.62; S, 5.89.
3.3. General procedure for the preparation of 2-((8-hydroxyquinolin-
5-yl)sulfonyl)-1,3-dimethyl/1,3-diphenylpropane-1,3-dione 4a,b
The tert-butyl ester 3a and b (0.01 mol) was refluxed in 6N HCl
(20 mL) under nitrogen until TLC analysis of the solution showed
complete deprotection (ca. 5 h). The hydrolyzed mixture was
cooled, diluted with 1 M HCl (25 mL), extracted with EtOAc
(2 ꢁ 50 mL), and concentrated to dryness under reduced pressure
to afford the deprotected products 4a and b.
3.2. General procedure for the preparation of 2-((8-((tert-
butyldiphenylsilyl)oxy)quinolin-5-yl)sulfonyl)-1,3-dimehyl/1,3-
diphenyl propane-1,3-dione 3a,b
Sodium methoxide (0.81 g, 0.015 mol) and b-diketones
(0.015 mol) were placed in a dried round bottom flask and stirred
for 1 h on a magnetic stirrer at 50 °C, after which a creamy mass
was obtained. The 8-((tert-butyldimethylsilyl)oxy)quinoline-5-sul-
fonyl chloride 2 (5.37 g, 0.015 mmol) was taken in dry toluene
(15 mL) and added drop by drop in above reaction mass. The reac-
tion mixture was refluxed for 7 h at 100 °C with stirring. The pro-
gress of the reaction was monitored by TLC. After completion of the
3.3.1. 3-((8-Hydroxyquinolin-5-yl)sulfonyl)pentane-2,4-dione 4a
M.p. 198 °C, yield 59%; 1H NMR (400 MHz, DMSO-d6) d 2.12 (s,
6H); d 5.62 (s, 1H); d 7.31–8.89 (m, 5H aromatic); d 8.34 (s, 1H,
broad, OH); HRMS (M)+ calcd for C14H13NO5S 307.0514. Found:
307.0872. Elemental analysis, calcd: C, 54.71; H, 4.26; N, 4.56; O,
26.03; S, 10.43. Found: C, 54.74; H, 4.23; N, 4.57; O, 26.01; S, 10.45.

24
A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
3.3.2. 2-((8-Hydroxyquinolin-5-yl)sulfonyl)-1,3-diphenylpropane-1,3-
dione 4b
3.4.5. 5-((2,4-Dimethyl-3H-pyrido[3,4-b][1,4]diazepin-3-
yl)sulfonyl)quinolin-8-ol 7a
M.p. 203 °C, yield 65%; 1H NMR (400 MHz, DMSO-d6) d 6.43 (s,
1H); d 7.31–8.89 (m, 15H, aromatic protons); d 8.34(s, 1H, broad,
OH); HRMS (M)+ calcd for C24H17NO5S 431.0827. Found:
431.1037. Elemental analysis, calcd: C, 66.81; H, 3.97; N, 3.25; O,
18.54; S, 7.43. Found: C, 66.83; H, 3.95; N, 3.28; O, 18.51; S, 7.43.
M.p. 153 °C, yield 61%; 1H NMR (400 MHz, DMSO-d6) d 2.22 (s,
3H); d 2.25 (s, 3H); d 4.98 (s, 1H); d 7.31–8.89 (m, 8H, aromatic pro-
tons); 13C NMR (DMSO) d 24.8, 61.8, 110.8, 117.0, 118.6, 126.4,
127.1, 130.0, 135.9, 138.3, 142.2, 143.3, 143.8, 144.9, 155.0, 161.1,
161.4, 162.1; HRMS (M)+ calcd for C19H16N4O3S 380.0943. Found:
380.1034; Elemental analysis, calcd: C, 59.99; H, 4.24; N, 14.73;
O, 12.62; S, 8.43. Found: C, 60.01; H, 4.19; N, 14.74; O, 12.65; S, 8.41.
3.4. General procedure for the preparation of 1H-1,4-diazepine 5a-b,
3H-1,5-benzodiazepine 6a-b and 3H-pyrido[3,4-b][1,4]diazepine
derivatives
3.4.6. 5-((2,4-Diphenyl-3H-pyrido[3,4-b][1,4]diazepin-3-
yl)sulfonyl)quinolin-8-ol 7b
M.p. 217 °C, yield 58%; 1H NMR (400 MHz, DMSO-d6) d 6.14 (s,
1H); d 7.31–8.89 (m, 18H, aromatic protons); 13C NMR (DMSO) d
68.1, 110.0, 119.3, 120.1, 126.4, 127.8, 129.1, 129.5, 130.2, 133.9,
133.6, 134.2, 135.3, 136.2, 142.7, 143.3, 143.9, 145.6, 155.3,
161.1, 162.7; HRMS (M)+ calcd for C29H20N4O3S 504.1256. Found:
504.1202. Elemental analysis, calcd: C, 69.03; H, 4.00; N, 11.10;
O, 9.51; S, 6.36. Found: C, 69.01; H, 4.01; N, 11.11; O, 9.53; S, 6.34.
An equimolar ratio of b-diketones (10 mmol) 3a–d, and EDA/o-
PDA/3,4-DAP (10.0 mmol) in ethyl acetate (50 mL) in the presence
of silica sulfuric acid (10.0 mmol) was stirred 50 °C for 2 h. The
progress of reaction was monitored by TLC using 7:2:1
(benzene:ethanol:ammonia) upper layer as mobile phase. Upon
completion of reaction, the mixture was extracted with ethyl ace-
tate (2 ꢁ 25 mL) and the solvent was removed. The crude product
was washed with dry ether and recrystallized from petroleum
ether:ethyl acetate (1:1). The product was purified by column
chromatography over silica gel using pet ether:ethyl acetate
(40:60) as an eluent.
4. Molecular docking studies
All docking studies were performed using ‘Internal Coordinate
Mechanics (Molsoft ICM 3.5-0a).
3.4.1. 5-((2,4-Dimethyl-3H-benzo[b][1,4]diazepin-3-
yl)sulfonyl)quinolin-8-ol 5a
4.1. Preparation of small molecule
M.p. 143 °C, yield 78%; 1H NMR (400 MHz, DMSO-d6) d 2.13 (s,
6H); d 5.62 (s, 1H); d 7.31–8.89 (m, 9H, aromatic protons); d 8.34 (s,
1H, broad, OH); 13C NMR (DMSO) d 24.8, 61.8, 110.8, 117, 124.6,
126.4, 127.1, 130.3, 133.2, 138.3, 142.3, 143.3, 161.11, 162.1;
HRMS (M)+ calcd For C20H17N3O3S 379.0991. Found: 379.1045.
Elemental analysis, calcd: C, 63.31; H, 4.52; N, 11.07; O, 12.65; S,
8.45. Found: C, 63.28; H, 4.54; N, 11.08; O, 12.63; S, 8.47.
Novel 8-hydroxyquinoline-5-(1,4-diazepin-6-yl)sulfonyl deriv-
atives were synthesized and tested as anti-schistosomal agents
and compiled using ChemDraw. 3D structures were constructed
using Chem 3D ultra 12.0 software [Molecular Modeling and
Analysis; Cambridge Soft Corporation, USA (2010)], and then they
were energetically minimized by using MOPAC (semi-empirical
quantum mechanics), Jop Type with 100 iterations and minimum
RMS gradient of 0.01, and saved as MDL MolFile (ꢂ.mol).
3.4.2. 5-((2,4-Diphenyl-3H-benzo[b][1,4]diazepin-3-
yl)sulfonyl)quinolin-8-ol 5b
M.p. 187 °C, yield 64%; 1H NMR (400 MHz, DMSO-d6) d 6.43 (s,
1H); d 7.31–8.89 (m, 19H, aromatic protons); d 8.34 (s, 1H, broad,
OH); 13C NMR (DMSO) d 68.2, 110.0, 120.1, 124.6, 126.1, 127.8,
129.1, 129.5, 130.2, 130.9, 132.9, 133.2, 133.9, 135.5, 142.7,
143.3, 161.1, 162.6; HRMS (M)+ calcd for C30H21N3O3S 503.1304.
Found: 503.1296. Elemental analysis, calcd: C, 71.55; H, 4.20; N,
8.34; O, 9.53; S, 6.37. Found: C, 71.58; H, 4.18; N, 8.35; O, 9.50;
S, 6.39.
4.2. Generation of ligand and enzyme structures
The crystal structure of target enzyme thioredoxin-glutathione
reductase TGR (2V6O) is was retrieved from the Protein Data Bank
(http://www.rcsb.org/pdb/welcome.do). All bound waters ligands
and cofactors were removed from the enzyme.
4.3. Docking using Molsoft ICM 3.4-8C program
3.4.3. 5-((5,7-Dimethyl-3,6-dihydro-2H-1,4-diazepin-6-
yl)sulfonyl)quinolin-8-ol 6a
1. Convert our PDB file into an ICM object: This conversion
involves addition of hydrogen bonds, assignment of atoms
types, and charges from the residue templates.
2. To perform ICM small molecule docking:
M.p. 132 °C, yield 81%; 1H NMR (400 MHz, DMSO-d6) d 2.13 (s,
6H); 3.61 (m, 4H, N–CH2–CH2–N); d 5.23 (s, 1H); d 7.31–8.89 (m,
5H, quinolone aromatic Protons); 13C NMR (DMSO) d 23.5, 47.7,
61.12, 110.8, 120.6, 126.5, 127.8, 130.3, 138.3, 141.6, 143.3,
160.5, 161.1; HRMS (M)+ calcd for C16H17N3O3S 331.0991. Found:
331.1094. Elemental analysis, calcd: C, 57.99; H, 5.17; N, 12.68;
O, 14.48; S, 9.68. Found: C, 58.02; H, 5.15; N, 12.69; O, 14.43; S,
9.71.
(a) Setup docking project:
(1)
(2)
(3)
(4)
Set project name
Setup the receptor
Review and adjust binding site
Make receptor maps
3.4.4. 5-((5,7-Diphenyl-3,6-dihydro-2H-1,4-diazepin-6-
yl)sulfonyl)quinolin-8-ol 6b
(b) Start docking simulation:
3. Display the result: ICM stochastic global optimization algorithm
attempts to find the global minimum of the energy function
that include five grid potentials describing interaction of the
flexible ligand with the receptor and internal conformational
energy of the ligand, during this process a stack of alternative
low energy conformations is saved (Table 1).
M.p. 171 °C, yield 74%; 1H NMR (400 MHz, DMSO-d6) d 3.68 (m,
4H, N–CH2–CH2–N); d 6.25 (s, 1H); d 7.31–8.89 (m, 15H, aromatic
protons); 13C NMR (DMSO) d 50.7, 67.4, 110.0, 121.9, 126.4, 128.5,
128.7, 129.1, 130.1, 131.3, 133.3, 135.4, 142.0, 143.3, 156.9, 161.1;
HRMS (M)+ calcd for C26H21N3O3S 455.1304. Found: 455.1267. Ele-
mental analysis, calcd: elemental analysis, calcd: C, 68.55; H, 4.65;
N, 9.22; O, 10.54; S, 7.04. Found: C, 68.52; H, 4.68; N, 9.19; O,
10.57; S, 7.04.
All inhibitors were compared according to the best binding free
energy (minimum) obtained among all the run.

A.F. Eweas et al. / Bioorganic Chemistry 46 (2013) 17–25
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The statistical tests were performed with the SPSS (version 16)
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Acknowledgment
This work was financially supported from Taif University, Taif,
Saudi Arabia by a Grant No. 2-433-1222.
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