Synthesis, molecular docking and preliminary in-Vitro cytotoxic evaluation of some substituted tetrahydronaphthalene (2',3',4',6'-tetra-O-acetyl-β-D- gluco-/galactopyranosyl) derivatives

Article abstract of DOI:10.3390/molecules17044717

A facile, convenient and high yielding synthesis of novel S-glycosides and N-glycosides incorporating 1,2,3,4-tetrahydronaphthalene and or 1,2-dihydropyridines moieties has been described. The aglycons 2, 4, and 7 were coupled with different activated halosugars in the presence of basic and acidic medium. The preliminary in-vitro cytotoxic evaluation revealed that compounds 3c, 3f, 5c and 7b show promising activity. A molecular docking study was performed against tyrosine kinase (TK) (PDB code: 1t46) by Autodock Vina. The docking output was analyzed and some compounds have shown hydrogen bond (H-B) formation with reasonable distances ranged from 2.06 A to 3.06 A with Thr 670 and Cys 673 residues found in the specified pocket. No hydrogen bond was observed with either Glu 640 nor Asp 810 residues, as was expected from pdbsum.

Full text of DOI:10.3390/molecules17044717

Molecules 2012, 17, 4717-4732; doi:10.3390/molecules17044717
OPEN ACCESS
molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Article
Synthesis, Molecular Docking and Preliminary in-Vitro
Cytotoxic Evaluation of Some Substituted Tetrahydro-
naphthalene (2',3',4',6'-Tetra-O-Acetyl-β-D-Gluco-/
Galactopyranosyl) Derivatives
Maha S. Al-Mutairi 1, Ebtehal S. Al-Abdullah 1, Mogedda E. Haiba 1,2, Mohammed A. Khedr 3
and Wafaa A. Zaghary 1,3,
*
1
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, King Saud University,
Riyadh 11451, Saudi Arabia
2
3
Department of Medicinal Chemistry, National Research Center, Dokki, Cairo 12622, Egypt
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Helwan University,
Ain Helwan, Cairo 11795, Egypt
* Author to whom correspondence should be addressed; E-Mail: wzaghary@ksu.edu.sa;
Tel.: +966-1-291-4843; Fax: +966-1-206-4501.
Received: 14 February 2012; in revised form: 9 April 2012 / Accepted: 13 April 2012 /
Published: 23 April 2012
Abstract: A facile, convenient and high yielding synthesis of novel S-glycosides and
N-glycosides incorporating 1,2,3,4-tetrahydronaphthalene and or 1,2-dihydropyridines
moieties has been described. The aglycons 2, 4, and 7 were coupled with different
activated halosugars in the presence of basic and acidic medium. The preliminary in-vitro
cytotoxic evaluation revealed that compounds 3c, 3f, 5c and 7b show promising activity. A
molecular docking study was performed against tyrosine kinase (TK) (PDB code: 1t46) by
Autodock Vina. The docking output was analyzed and some compounds have shown
hydrogen bond (H-B) formation with reasonable distances ranged from 2.06 A° to 3.06 A°
with Thr 670 and Cys 673 residues found in the specified pocket. No hydrogen bond was
observed with either Glu 640 nor Asp 810 residues, as was expected from pdbsum.
Keywords: tetrahydronaphthalene; pyridine; glycoside; cytotoxic; molecular docking

Molecules 2012, 17
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1. Introduction
Tetrahydronaphthalene derivatives have attracted significant attention in the field of drug discovery
because of their wide array of pharmacological activities, including action as non-steroidal glucocorticoid
receptor modulators, anticancer, analgesic, anti-inflammatory, antimicrobial, antiplatelet aggregation,
hypotensive, antiarrhythmic and anti-HIV effects [1–11]. Additionally, pyridine analogs are very
important class of heterocyclic compounds that have remarkable pharmacological activities as PDE4
inhibitors, analgesic, antifungal, antimalarial, anti-inflammatory, antibacterial, anti-HIV, antitumor and
antiviral properties [12–17].
On the other hand recent advances have implicated the role of tyrosine kinases (TK) in the
physiology of cancer. Though their activity is tightly regulated in normal cells, they may acquire
transforming functions due to mutation(s), over expression leading to malignancy [18,19]. The
activation of cancer cells can be blocked by selective tyrosine kinase inhibitors and thus considered to
be a promising target for anticancer drug discovery. Tyrosine kinases are important mediators of the
signal transduction process, leading to cell proliferation, differentiation, migration, metabolism and
apoptosis. Tyrosine kinases are a family of enzymes, which catalyzes phosphorylation of select tyrosine
residues in target proteins, using ATP. They are implicated in several steps of neoplastic development
and progression. Tyrosine kinase signaling pathways normally prevent deregulated proliferation or
contribute to sensitivity towards apoptotic stimuli. These signaling pathways are often genetically or
epigenetically altered in cancer cells to impart a selection advantage to the cancer cells [20–22].
Regarding the above mentioned findings and our previous reports [23] and [24], the main goal of
the present work was to design, synthesize and investigate the anticancer activity of some novel
pyridine-tetrahydronaphthalene derivatives carrying carbohydrate residues attached through S-glycosidic
or N-glycosidic bond formation. A molecular docking study was performed in order to interpret the
activity expressed by compounds 3c, 3f, 5b, 5c, 7a and 7b.
2. Results and Discussion
2.1. Chemistry
A series of S-glycosides and N-glycosides was designed in such a way that they incorporate a
tetrahydronaphthalene derivative attached to a glycosyl moiety through a S-linkage or N-linkage.
Synthesis of our desired compounds was achieved by allowing the acetyl derivative 1-(1,2,3,4-
tetrahydronaphthaline-6-yl)ethanone (1) to react directly with the appropriate aromatic aldehyde
and cyanothioacetamide in the presence of ammonium acetate, to afford the corresponding
2-thioxo(1H)pyridines 2a–c, respectively (Scheme 1, Table 1). The coupling between the aglycon and
the activated cyclic sugars (2',3',4',6'-O-acetyl--D-gluco- (or galato-) pyranosyl bromide was achieved
in a basic medium to give the corresponding thioglycoside derivatives 3a–f in a good yields,
respectively (Scheme 1, Table 1). The structures of thioglycosides 3a–f were established and
confirmed on the basis of their elemental analysis and spectral data (IR, 1H-NMR, 13C-NMR and MS,
cf. Experimental).

Molecules 2012, 17
Scheme 1. Synthetic pathways to compounds 2a–c, 3a–f, 4a,b and 5a–d.
4719
Ar
Ar
CN
NH
S
O
CN
ArCHO/
NH2
CN
/ CH COONH4/EtOH
3
CH3
N
H
CN
ArCHO/
CN /CH3COONH4/EtOH
N
H
S
1 : 1 : 8
1 : 1 : 8
1
2a-c
4a,b
F
F
Cl
Cl
b) Ar =
NO 2
OH
a) Ar =
b) Ar =
a) Ar =
O
c) Ar =
OC2H5
AcO
R1
AcO
R1
O
O
Br
Br
OAc
OAc
R2
R2
OAc
OAc
Ar
N
CN
S
Ar
N
CN
NH
AcO
R1
O
AcO
OAc
O
R1
R2
OAc
OAc
R2
3a-f
OAc
R1
R1
R2
R2
H
5a-d
OAc
OAc
d) Ar =
e) Ar =
f) Ar =
H
H
a) Ar =
b) Ar =
OAc
OAc
R1
R2
F
R1
R2
O
O
Cl
OAc
H
OAc
a) Ar =
b) Ar =
H
NO
c) Ar =
d) Ar =
H
H
NO2
2
F
F
Cl
Cl
OAc
OH
c) Ar =
OH
OAc
H
OAc
OAc
H
H
OC2H5
OC2H5
Cl
F
Table 1. Physicochemical properties of the newly synthesized compounds 2a, 3a–f, 5a–d,
7a, 7b, 8a and 8b.
Comp
No.
2a
Yield
%
Molecular Formula
Ar
R1
R2
MP (°C)
Cryst.Solv.
(Mol. Wt.)
-C4H3O
-C4H3O
---
H
---
OAc
OAc
OAc
H
245–247
194–196
172–174
118–120
215–217
140–142
175–177
192–194
205–207
107–109
132–134
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
80
76
75
68
75
76
79
65
70
66
72
C20H16N2OS (332)
C34H34N2O10S (662)
C36H35N3O11S (717)
C38H40N2O11S (732)
C34H34N2O10S (662)
C36H35N3O11S (717)
C38H40N2O11S (732)
C36H35Cl2N3O9 (724)
C36H35Cl2N3O9 (724)
C36H35F2N3O9 (691)
C36H35F2N3O9 (691)
3a
3b
3c
4-NO2-C6H5
3-EtO,4-OH-C6H4
-C4H3O
H
H
3d
3e
OAc
OAc
4-NO2-C6H5
H
3f
3-EtO-4-OH-C6H4 OAc
H
5a
2,6-Cl2-C6H4
2,6-Cl2-C6H4
2,6-F2-C6H4
2,6-F2-C6H4
H
OAc
H
5b
5c
OAc
H
OAc
H
5d
OAc

Molecules 2012, 17
4720
Table 1. Cont.
Comp
Ar
Yield
%
Molecular Formula
R1
R2
MP (°C)
Cryst.Solv.
No.
(Mol. Wt.)
7a
7b
8a
8b
2,6-Cl2-C6H4
---
---
---
---
H
180–182
170–172
230–232
155–157
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
Ethanol/H2O
95
C22H18Cl2N2O2 (413)
C22H18F2N2O2 (380)
C36H36Cl2N2O11 (743)
C36H36F2N2O11 (710)
2,6-F2-C6H4
2,6-Cl2-C6H4
2,6-F2-C6H4
92
OAc
OAc
86
H
85
Also, one-step synthesis of 2-imino (1H)pyridines 4a,b was accomplished by heating the acetyl
derivative 1 with malononitrile and the appropriate aldehyde in the presence of ammonium acetate
(Scheme 1, Table 1). The authors used these intermediate aglycones for the preparation of the new
glycosides 5a–d (Scheme 1, Table 1). The formation of N-glycosides 5a–d was proven using spectral
data (IR, 1H-NMR, 13C-NMR and MS).
On the other hand, the one pot reaction of compound 1 with the appropriate aromatic aldehyde and
ethyl cyanoacetate in the presence of ammonium acetate, afforded the corresponding pyridones 6a,b,
respectively (Scheme 2, Table 1). Hydrolysis of 6a,b in the presence of conc. sulphuric acid afforded
the amide derivatives 7a,b in good yield. Finally the coupling between the aglycons 7a,b and the
activated cyclic sugar 2',3',4',6'-tetra-O--D-galactopyranosyl bromide gave the corresponding
N-glycosides 8a,b in a good yield respectively. The structures of N-glycosides 8a,b were established
and confirmed for the reaction products on the basis of their spectral data.
Scheme 2. Synthetic pathways to compounds 6a,b, 7a,b and 8a,b.
Ar
CN
O
CH3
ArCHO/ CNCH2COOC2H5 / CH3COONH4/EtOH
1 : 1 : 8
N
H
O
6a,b
1
Cl
Cl
F
a) Ar =
b) Ar =
F
H2SO4
Ar
O
NH
AcO
R1
N
H
O
O
O
Ar
O
O
AcO
R1
Br
OAc
R2
NH2
OAc
OAc
N
H
R2
OAc
8a,b
7a,b
R1
R2
H
Cl
Cl
F
F
Cl
a) Ar=
b) Ar=
a) Ar =
OAC
OAC
Cl
F
b) Ar =
H
F

Molecules 2012, 17
4721
2.2. Molecular Modeling Study
Docking Using Autodock Vina
The tyrosine kinase (TK) crystal structure (PBD code = 1t46) was downloaded (Figure 1). In order
to find the important residues that form interactions with that inhibitor, the pdbsum server was used. It
was found that Cys 673, Thr 670, Glu 640 and Asp 810 formed some interactions with different
moieties of the inhibitor (Figure 2).
Figure 1. Showing the active site at which the inhibitor complexed.
Figure 2. A representation image simulating the interactions that was found in pdbsum,
showing the most visible interactions that were formed with the complexed ligand.
Cys 673
O
N
N
Thr 670
N
O
O
O
N
N
O
N
Asp 810
N
O
O
N
N
O
O
O
Glu 640
N
N
The affinity of any small molecule can be considered to be a unique tool in the field of drug design,
as there is a relationship between the affinity of organic molecules and the free binding energy and this

Molecules 2012, 17
4722
can contribute in prediction and interpretation of the activity of organic compounds toward a specific
target protein. Here we used the Autodock Vina [25,26] program for docking and to obtain the
affinities and RMSD (root mean square deviation) of all compounds.
The most visible feature that was observed in the docking output that all compounds fitted well
inside the same docking site and hence, the next step was to compare their affinities and try to rank
them as observed in the previous table and then to measure the ligands’ interaction to find out which
ones interact well and correlate that to the activity.
The six compounds with the highest calculated affinities were found to be compounds 7b, 3f, 5c,
5b, 3c and 7a, respectively (−11.6, −11.4, −11.2, −11.0, −10.7 and −10.3 Kcal/mol). Compounds 3c
and 7b were found to have low RMSD values that reflect their better fit as they were expected to have
a conformation with a low deviation from the expected best binding mode. No clear correlation
between the biological activity and the calculated affinity was observed. The only observation that was
found that the active compounds have the best affinities regardless of their ranking in activity.
Regarding the best mode of interaction for each compound of these six compounds, the docking
output was visualized to determine these interactions. It was important to concentrate on the previously
mentioned residues (Cys 673, Thr 670, Glu 640 and Asp 810) whether they were involved in these
interactions or not. For compound 3c THR 670 (Figure 3) was the major residue for making hydrogen
bond with one of the acetyl groups found in the side chains of this compound.
Figure 3. Compound 3c showing hydrogen bond with Thr 670 with the -CH3CO of the compound.
Compound 3f shows two hydrogen bonds with Thr 670 and Cys 673 (Figure 4). Compounds 5c, 5b,
and 7b have shown the same hydrogen bond with the previously mentioned residues as well
(Thr 670, and Cys 673), as shown in Figures 5–7.
Figure 4. Compound 3f hydrogen bond formation with Thr 670 and Cys 673.

Molecules 2012, 17
Figure 5. Compound 5c hydrogen bond formation with Thr 670 and Cys 673.
4723
Figure 6. Compound 5b hydrogen bond formation with Thr 670 and Cys 673.
Figure 7. Compound 7b hydrogen bond formation with Cys 673.
2.3. In Vitro Cytotoxic Screening
Some of the newly synthesized compounds were tested at the Department of Tumor Pathology,
National Cancer Institute, Cairo, Egypt. Ehrlich Ascites Carcinoma (EAC) cells were used for
cytotoxic activity screening. The selected synthesized analogues 2b, 3a–f, 5b, 5c, 7a and 7b were
tested for in vitro cytotoxic activity at two different concentrations. The results (Table 2) are expressed
in the form of the percentage of non-viable cells [27]. The in vitro evaluation revealed that 3c, 3f,
5c and 7b show promising activity. This provides a new class of antitumor agents for further
biological evaluation.

Molecules 2012, 17
Table 2. The effect of the tested compounds on the viability of tumor cell in vitro. The
4724
calculated affinities and RMSD for some of the newly synthesized compounds.
Anticancer effect %
Affinity
RMSD Deviation
inhibition of cell viability
Compound
Kcal/mol
from best mode
100 µg/mL
50 µg/mL
2b
3a
3b
3c
3d
3e
3f
0
0
0
0
−9.5
−8.3
7.07
9.5
0
0
−7.7
11.6
4.02
10.3
9.2
70%
0
50%
0
−10.7
−6.6
0
0
−7.4
10%
0
0
−11.4
−11.0
−11.2
−10.3
−11.6
9.0
5b
5c
7a
7b
0
9.3
40%
0
7%
0
9.5
7.2
20%
15%
4.6
3. Experimental
3.1. Chemistry
3.1.1. General
Melting points (°C, uncorrected) were determined in open glass capillaries using a Barnstead 9001
Electrothermal melting point apparatus. Infrared (IR) spectra were recorded on a Perkin Elmer FT-IR
Spectrum BX Spectrometer at cm−1 scale using KBr discs. 1H-NMR and 13C-NMR were recorded on a
JEOL 300 MHz Spectrometer; chemical shifts are expressed in δ (ppm) with reference to TMS. The
mass spectra were run at 70 ev with Packard 5890 Thewlett GC/MS Spectrometer, using the Electron
Ionization (EI) technique. Elemental analyses (C, H, N) were carried out at the Microanalytical Data
Center, Faculty of Science, Cairo, Egypt, and were in full agreement with the proposed structures
within ±0.4% of the theoretical values. Thin layer chromatography was performed on precoated
(0.75 mm) silica gel GF254 plates (E. Merck, Germany). Visualization was performed by illumination
with UV light source (254 nm). Compounds 2b, 2c, 4a, 4b, 6a and 6b were prepared using the
previously published procedures [10] and [11] as described below (Schemes 1 and 2).
3.1.2. Synthesis of 4-Aryl-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-thioxo-1,2-dihydropyridine-3-
carbonitriles 2a–c
A mixture of 6-acetyltetraline 1 (0.3 g, 0.0017 mol), 2-cyanothioacetamide (0.17 g, 0.0017 mol),
the an appropriate aromatic aldehyde, namely 2-furfural, p-nitrobenzaldehyde, and/or 3-ethoxy-4-
hydroxybenzaldehyde (0.00172 mol) and excess ammonium acetate (1.05 g, 0.0137 mol) was gently
refluxed in absolute ethanol (15 mL) for 3–8 h. On cooling, the separated solid was filtered off,
washed successively with water, and then with ether and recrystallized with aqueous ethanol to give
2a–c respectively (Table 1).

Molecules 2012, 17
4725
4-(Furan-2-yl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-thioxo)-1,2-dihydropyridine-3-carbonitrile (2a).
IR: (υ, cm−1) 3,300 (NH), 2,200 (CN) and 1,206 (C-O). 1H-NMR (DMSO-d6): δ 1.76 (m, 4H, -CH2-CH2-),
2.78 [m, 4H, 2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 6.89–8.61 (m, 7H,
aromatic protons & pyridine-H) and 13.9 (NH). 13C-NMR (DMSO-d6): δ 22.9 (-CH2-CH2-), 29.2
[2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 114.2 (CN), 106.2, 107.6, 117.7,
117.9, 125.66, 128.7, 129.3, 129.8, 137.8, 141.5, 142.1, 147.5, 148.3, 152.6, 180.4 (Ar-C), MS, m/z
(%): (M+, 332, 0.05%).
3.1.3. Synthesis of 4-Aryl-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-gluco
and galactopyranosyl thio)pyridine-3-carbonitriles 3a–f
A mixture of 2a–c (0.01 mol) in ethanol (10 mL) and potassium hydroxide (0.56 g, 0.01 mol) in
distilled water (6 mL) was added to a solution of 2,3,4,6-tetra-O-acetyl--D-gluco- or galactopyranosyl
bromide (4.10 g, 0.01 mol) in acetone (30 mL). The reaction mixture was stirred at room temperature
until completion (TLC) 20–24 h. and then the solution was evaporated and poured onto ice/cold water.
The formed solid product was collected by filtration. The residue was washed with distilled water to
remove the formed potassium bromide. The resulting solid product was dried and crystallized from
dilute ethanol (Table 1).
4-(Furan-2-yl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-glucopyranosylthio)
pyridine-3-carbonitrile (3a). IR: (υ, cm−1) 2937 (CH, alkane), 2,216 (CN), 1,748 (C=O) and 1,230 (C-O).
1H-NMR (DMSO-d6): δ 1.78 (m, 4H, -CH2-CH2-), 2.02–2.03 (4s, 12H, 4 × CH3CO), 2.88 [m, 4H,
2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 4.02–4.06 (m, 2H, 6'-H2) 4.31 (m, 1H,
5'-H), 5.03 (t, 1H, 4'-H) 5.26 (t, 1H, 3'-H), 5.71 (t, 1H, 2'-H), 6.24 (d, 1H, 1'-H) and 6.84–8.13 (m, 7H,
aromatic protons and pyridine-H). 13C-NMR (DMSO-d6): δ 22.9 (-CH2-CH2-), 29.2 [2(CH2)] attached
to the aromatic ring of tetrahydronaphthalene), 115.7 (CN), 62.29 (CH2, C-6'), 68.20 (C-4'), 69.76 (C-2'),
73.1 (C-3'), 75.17 (C-5'), 83.90 (C-1'), 106–180 (Ar-C) and 169.5–170.3 (4 × CO). MS, m/z (%):
(M+, 662, 0.03%).
4-(4-Nitrophenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-glucopyranosylthio)
pyridine-3-carbonitrile (3b). IR: (υ, cm−1) 2,930 (CH, alkane), 2,217 (CN), 1,752 (C=O) and 1,348 &
1
1,528 (NO2). H-NMR (DMSO-d6): δ 1.75 (m, 4H, -CH2-CH2-), 2.04 (4s, 12H, 4 × CH3CO), 2.76
[m, 4H, 2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 4.04–4.06 (m, 2H, 6'-H2) 4.35
(m, 1H, 5'-H), 5.05 (t, 1H, 4'-H) 5.24 (t, 1H, 3'-H), 5.74 (t, 1H, 2'-H), 6.31 (d, 1H, 1'-H) and 7.1–8.4
13
(m, 8H, aromatic protons and pyridine-H). C-NMR (DMSO-d6): δ 22.18 (-CH2-CH2-), 19.62–19.90
(4 × -COCH3), 28.65 [2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 61.5 (CH2, C-6'),
67.2 (C-4'), 69.5 (C-2'), 72.9 (C-3'), 74.6 (C-5'), 79.6 (C-1'), 115.46 (CN), 102–147 (Ar-C), 169.5 (4 × CO).
MS, m/z (%): (M+, 717, 1.34%).
4-(3-Ethoxy-4-hydroxyphenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-
glucopyranosyl thio)pyridine-3-carbonitrile (3c). IR: (υ, cm−1) 3,458 (OH), 2,937 (CH, alkane), 2,216
1
(CN), 1,754 (C=O) and 1,125 (C-O). H-NMR (DMSO-d6): δ 1.36 (t, 3H, OCH2CH3), 1.78 (m, 4H,
-CH2-CH2-), 1.9–2.03 (4s, 12H, 4 × CH3CO), 2.87 [m, 4H, 2(CH2)] attached to the aromatic ring of

Molecules 2012, 17
4726
tetrahydronaphthalene), 4.01–4.35 (m, 5H, 6'-H2, 5'-H & OCH2CH3), 4.9 (t, 1H, 4'-H) 5.2 (t, 1H, 3'-H),
5.7 (t, 1H, 2'-H), 6.29 (d, 1H, 1'-H) 6.94–8.1 (m, 7H, aromatic protons & pyridine-H) and 9.6 (OH).
13C-NMR (DMSO-d6): δ 14.49 (-OCH2CH3), 20.28 (4 × -COCH3), 22.49 (-CH2-CH2-), 28.63 [2(CH2)]
attached to the aromatic ring of tetrahydronaphthalene), 61.54 (CH2, C-6'), 63.89 (-O-CH2-CH3), 67.52
(C-4'), 69.50 (C-2'), 72.89 (C-3'), 74.63 (C-5'), 79.60 (C-1'), 115.75 (CN), 113–158 (Ar-C) and 169.45
(4 × CO). MS, m/z (%): (M+, 732, 12.15%).
4-(Furan-2-yl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-galactopyranosyl
thio)pyridine-3-carbonitrile (3d). IR: (υ, cm−1) 2,936 (CH, alkane), 2,216 (CN), 1,754 (C=O) and
1,239 (C-O). 1H-NMR (DMSO-d6) δ 1.78 (m, 4H, -CH2-CH2-), 1.96–2.14 (4s, 12H, 4 × CH3CO), 2.88
[m, 4H, 2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 3.95 (m, 2H, 6'-H2), 4.10 (m,
1H, 5'-H), 4.53 (t, 1H, 4'-H) 5.25 (t, 1H, 3'-H), 5.41 (t, 1H, 2'-H), 6.22 (d, 1H, 1'-H), 6.8–8.1 (m, 7H,
aromatic protons and pyridine-H). 13C-NMR (DMSO-d6): δ 22.9 (-CH2-CH2-), 29.2 (2(CH2) attached
to the aromatic ring of tetrahydronaphthalene), 115.7 (CN), 61.70 (CH2, C-6'), 67.74 (C-4'), 69.45 (C-2'),
72.64 (C-3'), 74.97 (C-5'), 84.74 (C-1'), 106–180 (Ar-C) and 169.03–169.87 (4 × CO). MS, m/z (%):
(M+, 662, 0.71%).
4-(4-Nitrophenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-galactopyranosyl
thio)pyridine-3-carbonitrile (3e). IR: (υ, cm−1) 2,932 (CH, alkane), 2,217 (CN), 1751 (C=O) and 1,348
1
& 1,528 (NO2). H-NMR (DMSO-d6): δ 1.76 (m, 4H, -CH2-CH2-), 1.97–2.14 (4s, 12H, 4 × CH3CO),
2.77 [m, 4H, 2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 3.94 (m, 2H, 6'-H2), 4.08
(m, 1H, 5'-H), 4.55 (t, 1H, 4'-H) 5.30 (t, 1H, 3'-H), 5.40 (t, 1H, 2'-H), 6.25 (d, 1H, 1'-H) and 7.21–8.43
(m, 8H, aromatic protons and pyridine-H). 13C-NMR (DMSO-d6): δ 22.2 (-CH2-CH2-), 19.6–19.9
(4 × -COCH3), 28.82 (2(CH2) attached to the aromatic ring of tetrahydronaphthalene), 61.28 (CH2, C-6'),
65.23 (C-4'), 66.67 (C-2'), 71.34 (C-3'), 74.46 (C-5'), 80.75 (C-1'), 115.46 (CN), 102–147 (Ar-C),
169.56–169.70 (4 × -CO of COCH3 groups). MS, m/z (%): (M+ −1,716, 8.9%).
4-(3-Ethoxy-4-hydroxyphenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-
galactopyranosyl thio)pyridine-3-carbonitrile (3f). IR: (υ, cm−1) 3,458 (OH), 2,217 (CN), 1,752 (C=O)
and 1,056 (C-O). 1H-NMR (DMSO-d6): δ 1.39 (t, 3H, OCH2CH3), 1.80 (m, 4H, -CH2-CH2-), 1.96–2.06
(4s, 12H, 4 × CH3CO), 2.79 [m, 4H, 2(CH2)] attached to the aromatic ring of tetrahydronaphthalene),
4.08 (m, 2H, 6'-H2) 4.11 (m, 1H, 5'-H), 4.13 (t, 2H, -OCH2CH3) 4.5 (t, 1H, 4'-H) 5.26–5.65 (t, 2H, 3'-H
& 2'-H), 6.25 (d, 1H, 1'-H) and 6.98–8.08 (m, 7H, aromatic protons and pyridine-H). 13C-NMR
(DMSO-d6): δ 13.96 (-OCH2CH3) 19.62–19.85 (4 × -COCH3), 22.17 (-CH2-CH2-), 28.81 [2(CH2)]
attached to the aromatic ring of tetrahydronaphthalene), 61.25 (CH2, C-6'), 64.06 (-O-CH2-CH3),
65.27 (C-4'), 66.73 (C-2'), 71.40(C-3'), 74.34 (C-5'), 80.69 (C-1'), 115.6 (CN), 110–147 (Ar-C) and
168.56–169.52 (4 × CO). MS, m/z (%): (M+, 732, 6.9%).
3.1.4. Synthesis of 4-Aryl-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-gluco
and galactopyranosyl imino)pyridine-3-carbonitriles 5a–d
To a solution of 4a and/or 4b (0.001 mol) in acetone (10 mL), 2,3,4,6-tetra-O-acetyl--D-gluco- or
galactopyranosyl bromide (4.10 g, 0.01 mol) was added. A few drops of acetic acid were added and the

Molecules 2012, 17
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reaction mixture was heated with stirring on a water bath at 40–50 °C for 25 h. The reaction mixture
was cooled and then poured onto ice water. The formed precipitate was washed for several times with
water and recrystallized from aqueous ethanol (Table 1).
4-(2,6-Dichlorophenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-gluco-
pyranosyl imino)pyridine-3-carbonitrile (5a). IR: (υ, cm−1) 3,450 (NH), 2,990 (CH2), 2,216 (CN),
1
1,750 (C=O). H-NMR (DMSO-d6): δ 1.79 (m, 4H, tetrahydronaphthalene CH2), 1.98–2.29 (4s, 12H,
4 × CH3CO) 2.81 (m, 4H, tetrahydronaphthalene CH2), 4.09 (s, 2H, 6'-H), 4.54 (m, 1H, 5'-H), 4.64 (m,
2H, 4'-H, 3'-H), 5.46 (t, 1H, 2'-H), 5.95 (d, 1H, 1'H), 7.14–7.84 (m, 7H, Ar-H and pyridine-H).
13C-NMR (DMSO-d6): 20.65–20.87 (4 × -COCH3), δ 22.2 (-CH2-CH2-), 28.7 (2(CH2) attached to the
aromatic ring of tetrahydronaphthalene), 61.9 (CH2, C-6'), 68.4 (C-4'), 68.8 (C-2'), 73.9 (C-3'), 75.25 (C-5'),
81.9 (C-1'), 115 (CN), 106–176 (Ar-C & pyridine-C), 169.5–170.3 (4 × CO). MS, m/z (%): (M+ +1,
725, 4.47%).
4-(2,6-Dichlorophenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-galactopy
ranosyl imino)pyridine-3-carbonitrile (5b). IR: (υ, cm−1) 3,360 (NH), 2,929 (CH2), 2,212 (CN), 1,737
(C=O). 1H-NMR (DMSO-d6): δ 1.74 (m, 4H, tetrahydronaphthalene CH2), 2–2.01 (4s, 12H, 4 × CH3CO)
2.76 (m, 4H, tetrahydronaphthalene CH2), 4.01–4.06 (s, 2H, 6'-H), 4.51 (m, 1H, 5'-H), 4.71 (t, 1H, 4'),
5.47 (t, 1H, 3'-H), 5.62 (t, 1H, 2'-H), 6.53 (d, 1H, 1'H), 7.14–7.84 (m, 7H, Ar-H and pyridine-H).
13C-NMR (DMSO-d6): 20.69–19.89 (4 × -COCH3), 22.17 (-CH2-CH2-), 28.81 (2(CH2) attached to the
aromatic ring of tetrahydronaphthalene), 61.94 (CH2, C-6'), 66.22 (C-4'), 68.01 (C-2'), 71.73(C-3'),
74.33 (C-5'), 82.28 (C-1'), 115.6 (CN), 111–180 (Ar-C & pyridine C) and 169.56–170.33(4 × CO).
MS, m/z (%): (M+ +1, 725, 3%).
4-(2,6-Difluorophenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-glucopy
ranosyl imino)pyridine-3-carbonitrile (5c). IR: (υ, cm−1) 3,359 (NH), 2,932 (CH2), 2,214 (CN),
1
1,750 (C=O). H-NMR (DMSO-d6): δ 1.75 (m, 4H, tetrahydronaphthalene CH2), 2–2.02 (4s, 12H,
4 × CH3CO) 2.77 (m, 4H, tetrahydronaphthalene CH2), 4.02 (s, 2H, 6'-H), 4.18 (m, 1H, 5'-H), 4.70 (t,
1H, 4'), 4.92 (t, 1H, 3'-H), 5.25 (t, 1H, 2'-H), 6.62 (d, 1H, 1'H), 7.15–8.55 (m, 7H, Ar-H and pyridine-H).
MS, m/z (%):13C-NMR (DMSO-d6): 20.65–20.87 (4 × -COCH3), δ 22.4 (-CH2-CH2-), 28.91 (2(CH2)
attached to the aromatic ring of tetrahydronaphthalene), 61.9 (CH2, C-6'), 68.4 (C-4'), 68.8 (C-2'), 73.9
(C-3'), 75.25 (C-5'), 81.9 (C-1'), 115.8 (CN), 111.52–174.1 (Ar-C and pyridine-C), 169.5–170.3 (4 × CO).
MS, m/z (%): (M+, 691, 16%).
4-(2,6-Difluorophenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-(2',3',4',6'-tetra-O-acetyl-β-D-galactopyranosyl
imino)pyridine-3-carbonitrile (5d). IR: (υ, cm−1) 3,430 (NH), 2,990 (CH2), 2,217 (CN), 1,750 (C=O).
1H-NMR (DMSO-d6): δ 1.75 (m, 4H, tetrahydronaphthalene CH2), 1.99–2.01 (4s, 12H, 4 × CH3CO)
2.76 (m, 4H, tetrahydronaphthalene CH2), 4.05 (s, 2H, 6'-H), 4.2 (m, 1H, 5'-H), 4.68 (t, 1H, 4'), 4.95
(t, 1H, 3'-H), 5.37 (t, 1H, 2'-H), 6.65 (d, 1H, 1'H), 7.15–9.54 (m, 7H, Ar-H and pyridine-H). 13C-NMR
(DMSO-d6): 20.69–19.89 (4 × -COCH3), 22.17 (-CH2-CH2-), 28.81 (2(CH2) attached to the aromatic
ring of tetrahydronaphthalene), 61.94 (CH2, C-6'), 66.22 (C-4'), 68.01 (C-2'), 71.73(C-3'), 74.33 (C-5'),
82.28 (C-1'), 115 (CN), 111–176 (Ar-C and pyridine C) and 169.56–170.33 (4 × CO). MS, m/z (%):
(M+, 691, 2%).

Molecules 2012, 17
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3.1.5. Synthesis of 4-Aryl-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-oxo-1,2-dihydropyridine-3-
carboxamides 7a,b
A mixture of 6a,b (0.01 mol) and conc. sulphuric acid (15 mL) was stirred at room temperature 6 h,
and then poured on cold water. The formed solid product was collected by filtration and recrystallized
from ethanol to give 7a,b (Table 1).
4-(2,6-Dichlorophenyl-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-oxo)-1,2-dihydropyridine-3-carboxamide
(7a). IR: (υ, cm−1) br. 3,382 (NH & NH2), 2,931 (CH2) and 1,668 (C=O). 1H-NMR (DMSO-d6): δ 1.69
(m, 4H, -CH2-CH2-), 2.70 [m, 4H, 2(CH2)] attached to the aromatic ring of tetrahydronaphthalene),
4.03 (br. s, 2H, NH2) and 6.40–8.89 (m, 7H, aromatic protons and pyridine-H). 13C-NMR (DMSO-d6):
δ 22.97 (-CH2-CH2-), 29.23 [2(CH2)] attached to the aromatic ring of tetrahydronaphthalene), 107.17,
118.39, 124.78, 128.05, 128.42, 129.20, 129.56, 131.71, 137.98, 140.12, 140.68, 149.39, 153.11 (Ar-C),
163.88, 164.97 (2C=O). MS, m/z (%): (M+, 413, 14%).
4-(2,6-Difluorophenyl)-6-(1,2,3,4-tetrahydronaphthalen-6-yl)-2-oxo-1,2-dihydropyridine-3-carboxamide
(7b). IR: (υ, cm−1) br. 3,380 (NH & NH2), 2,932 (CH2) and 1,666 (C=O). 1H-NMR (DMSO-d6): δ 1.69
(m, 4H, -CH2-CH2-), 2.71 (m, 4H, 2(CH2) attached to the aromatic ring of tetrahydronaphthalene),
13
3.34 (br. s, 2H, NH2), 6.55–8.72 (m, 7H, aromatic protons & pyridine-H) and 12.5 (NH). C-NMR
(DMSO-d6): δ 22.98 (-CH2-CH2-), 29.18 (2(CH2) attached to the aromatic ring of tetrahydronaphthalene,
108.16, 111.59, 112.45, 118.37, 120.53, 123.07, 124.80, 128.41, 129.45, 130.15, 137.91, 140.51, 145.09,
148.75, 157.87, 159.81 (Ar-C), 163.20, 165.54 (2C=O). MS, m/z (%): (M+, 380, 23%).
3.1.6. Synthesis of 4-Aryl-N-(2',3',4',6'-tetra-O-acetyl-β-D-galactopyranosyl)-6-(1,2,3,4-tetrahydro-
naphthalen-6-yl)-2-oxo-1,2-dihydro pyridine-3-carboxamides 8a,b
To a solution of 7a and/or 7b (0.001 mol) in acetone (10 mL), 2,3,4,6-tetra-O-acetyl--D-gluco- or
galactopyranosyl bromide (4.10 g, 0.01 mol) was added. Few drops of acetic acid were added and the
reaction mixture was heated with stirring on a water bath at 40–50 °C for 25 h. The reaction mixture
was cooled and then poured onto ice water. The formed precipitate was washed for several times with
water and recrystallized from ethanol (Table 1).
4-(2,6-Dichlorophenyl-N-(2',3',4',6'-tetra-O-acetyl-β-D-galactopyranosyl)-6-(1,2,3,4-tetrahydronaphthalen-
1
6-yl)-2-oxo)-1,2-dihydropyridine-3-carboxamide (8a). IR: (υ, cm−1) H-NMR (DMSO-d6): δ 1.73 (m,
4H, -CH2-CH2-), 1.90–2.08 (4s, 12H, 4 × CH3CO), 2.75 [m, 4H, 2(CH2)] attached to the aromatic ring
of tetrahydronaphthalene), 3.93 (m, 2H, 6'-H2) 4.08 (m, 1H, 5'-H), 4.20 (t, 1H, 4'-H) 5.15–5.17 (t, 2H,
3'-H & 2'-H), 5.35 (d, 1H, 1'-H), 6.42–8.99 (m, 7H, aromatic protons and pyridine-H) and 12.67 (NH).
13C-NMR (DMSO-d6): δ 19.85–20.04 (4 × -COCH3), 22.98 (-CH2-CH2-), 29.14 [2(CH2)] attached to
the aromatic ring of tetrahydronaphthalene), 61.25 (CH2, C-6'), 65.88 (C-4'), 67.34 (C-2'), 72.1 (C-3'),
74.34 (C-5'), 80.23 (C-1'), 107.17–153.09 (Ar-C) and 163.87–190.99 (6 × CO). MS, m/z (%): (M+ +1,
744, 10.96).
4-(2,6-Difluorophenyl-N-(2',3',4',6'-tetra-O-acetyl-β-D-galactopyranosyl)-6-(1,2,3,4-tetrahydronaphthalen-
1
6-yl)-2-oxo)-1,2-dihydropyridine-3-carboxamide (8b). IR: (υ, cm−1). H-NMR (DMSO-d6): δ 1.73 (m,

Molecules 2012, 17
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4H, -CH2-CH2-), 1.99–2.08 (4s, 12H, 4 × CH3CO), 2.74 (m, 4H, 2(CH2) attached to the aromatic ring
of tetrahydronaphthalene), 3.92 (m, 2H, 6'-H2) 4.08 (m, 1H, 5'-H), 4.56 (t, 1H, 4'-H) 5.15–5.17 (t, 2H,
3'-H & 2'-H), 5.40 (d, 1H, 1'-H), 6.59–9.48 (m, 7H, aromatic protons and pyridine-H) and 12.66 (NH).
13C-NMR (DMSO-d6): δ 19.85–20.04 (4 × -COCH3), 22.98 (-CH2-CH2-), 29.14 (2(CH2) attached to
the aromatic ring of tetrahydronaphthalene), 61.25 (CH2, C-6'), 65.88 (C-4'), 67.34 (C-2'), 72.1(C-3'),
74.34 (C-5'), 80.23 (C-1'), 108.17–159.79 (Ar-C) and 163.22–170.33 (6 × CO). MS, m/z (%): (M+,
710, 8.66).
3.2. Molecular Modelling
Autodock vina allows the flexible docking of ligands into their site of action. It has the ability to use
all the rotable bonds of the ligands to give a number of conformations from which the best mode could
be deduced. The crystal structure of tyrosine kinase was downloaded from protein data bank [28]. Pdb
code = 1t46 was written in search area of the pdbsum by clicking on find tap the complexed inhibitor
will be viewed showing all possible interactions with the binding site [29]. All compounds were built
and saved as pdbqt format and docking with Autodock Vina was performed according to the specified
condition in which the grid box was adjusted to have center x = 24.15, center_y = 22.8, and center_z = 12.9
by these centres the pocket with the main residues were involved inside the box.
3.3. In Vitro Cytotoxic Screening
3.3.1. Tumor
Ehrlich Ascites Carcinoma (EAC) cells was kindly provided by the National Cancer Institute (Cairo
University, Cairo, Egypt). The tumor line was maintained in female Swiss albino mice by weekly
intra-peritoneal injection of 2.5 × 105 cells/mouse according to the method recommended by the
Egyptian National Cancer Institute, Cairo University. Such developed tumor is characterized by its
moderate rapid growth which could not kill the animal due to the accumulation of ascites before about
14 days after transplantation. Cells were counted before injection using the bright line haemocytometer
and dilutions made by physiological saline. The desired number of cells was injected in a volume of
0.5 mL/mouse. Solid Ehrlich carcinoma was induced by inoculation of 2.5 × 105 cells in the back
between the thighs of each animal [30].
3.3.2. Animals
Adult female Swiss albino mice, obtained from the animal house of Cairo Cancer Institute (Cairo
University, Egypt) weighing 18–22 g were used. The local committee approved the design of the
experiments, and the protocol conforms to the guidelines of the National Institutes of Health (NIH).
The animals were maintained on a standard pellet diet and tap water ad libitum.
3.3.3. In Vitro Test for Cytotoxic Effect
A set of sterile test tubes were used, where 2.5 × 105 tumor cells per ml were suspended in
phosphate buffer saline, then 25, 50, 100 µg/mL of the test compound were added to the suspension,

Molecules 2012, 17
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kept at 37 °C for 2 h. Trypan blue dye exclusion test was then carried out to calculate the percentage of
non-viable cells [27].
4. Conclusions
Some substituted tetrahydronaphthalene (2,3,4,6-tetra-O-acetyl-β-D-gluco/-galactopyranosyl)
derivatives were synthesized and tested for their anticancer activity. The preliminary in vitro
cytotoxicity evaluation revealed that five compounds show promised activity. This encouraged us to
do a molecular docking study to find out a suitable binding mode for these compounds. The docking
was done against TK and the docking output was analyzed. The compounds have shown hydrogen
bond formation with a reasonable distance ranging from 2.06 A° to 3.06 A° with Thr 670 and Cys 673
residues. No hydrogen bond was observed with neither Glu 640 nor Asp 810 residues as was expected
from pdbsum. From that we can conclude that substituted tetrahydronaphthalene (2,3,4,6-tetra-O-
acetyl-β-D-gluco-/galactopyranosyl) compounds may provide more activity by further study trying to
modify their substitutions to give new derivatives with expected improved anticancer activity and that
may be the subject of our future work.
Acknowledgments
This research project was supported by a grant from the “Research Center of the Center for Female
Scientific and Medical Colleges”, Deanship of Scientific Research, King Saud University.
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Sample Availability: Samples of the compounds 1, 2 and 4 are available from the authors.
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