Novel nucleoside analogues: First synthesis of pyridine-4-thioglycosides and their cytotoxic evaluation

Article abstract of DOI:10.1080/15257770.2015.1071843

The reaction of sodium 2,2-dicyanoethene-1,1-bis(thiolate) with 2-cyano-N-arylacetamides afforded sodium pyridine-4-thiolates, coupling of the latters with 2,3,4,6-tetra-O-acetyl-D-gluco- and D-galactopyranosyl bromides, respectively, afforded new pyridine-4-thioglycosides. Ammonolysis of the latter compounds afforded the free thioglycosides. The antitumor activities of the synthesized compounds were tested against human tumor cell lines; lung (A549), colon (HCT116), liver (HEPG2), and prostate (PC3).

Full text of DOI:10.1080/15257770.2015.1071843

Nucleosides, Nucleotides and Nucleic Acids
ISSN: 1525-7770 (Print) 1532-2335 (Online) Journal homepage: http://www.tandfonline.com/loi/lncn20
Novel Nucleoside Analogues: First Synthesis of
Pyridine-4-Thioglycosides and Their Cytotoxic
Evaluation
Galal Elgemeie, Mamdouh Abou-Zeid, Shahinaz Alsaid, Ali Hebishy & Hanaa
Essa
To cite this article: Galal Elgemeie, Mamdouh Abou-Zeid, Shahinaz Alsaid, Ali Hebishy & Hanaa
Essa (2015) Novel Nucleoside Analogues: First Synthesis of Pyridine-4-Thioglycosides and
Their Cytotoxic Evaluation, Nucleosides, Nucleotides and Nucleic Acids, 34:10, 659-673, DOI:
10.1080/15257770.2015.1071843
To link to this article: http://dx.doi.org/10.1080/15257770.2015.1071843
Published online: 13 Oct 2015.
Submit your article to this journal
Article views: 6
View related articles
View Crossmark data
Full Terms & Conditions of access and use can be found at
http://www.tandfonline.com/action/journalInformation?journalCode=lncn20
Download by: [Simon Fraser University]
Date: 28 October 2015, At: 07:37

Nucleosides, Nucleotides and Nucleic Acids, 34:659–673, 2015
Copyright ꢀC Taylor and Francis Group, LLC
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770.2015.1071843
NOVEL NUCLEOSIDE ANALOGUES: FIRST SYNTHESIS OF
PYRIDINE-4-THIOGLYCOSIDES AND THEIR CYTOTOXIC EVALUATION
Galal Elgemeie,1 Mamdouh Abou-Zeid,2 Shahinaz Alsaid,1 Ali Hebishy,1
and Hanaa Essa1
1Chemistry Department, Faculty of Science, Helwan University, Helwan, Cairo, Egypt
2Green Chemistry Department, National Research Center, Dokki, Cairo, Egypt
The reaction of sodium 2,2-dicyanoethene-1,1-bis(thiolate) with 2-cyano-N-arylacetamides af-
2
forded sodium pyridine-4-thiolates, coupling of the latters with 2,3,4,6-tetra-O-acetyl-D-gluco- and
D-galactopyranosyl bromides, respectively, afforded new pyridine-4-thioglycosides. Ammonolysis of
the latter compounds afforded the free thioglycosides. The antitumor activities of the synthesized com-
pounds were tested against human tumor cell lines; lung (A549), colon (HCT116), liver (HEPG2),
and prostate (PC3).
Keywords Ketene dithioacetals; sodium pyridine-4-thiolates; pyridine-4-thioglycosides;
activated nitriles
1. INTRODUCTION
Thiosugars are well known for their diverse array of bioactivities.[1]
Thus, the new developments in the synthetic and medicinal chemistry of
thiosugars are important for carbohydrate drug design.[2] Till date various
methods have been developed for the synthesis of these very interesting
molecules.[3]
Recently, deazanucleoside analogues have been known to exhibit an-
titumor activity.[4] During our studies of nucleoside analogues with novel
H-bonding patterns, a route for the synthesis of S-glycosides bearing a sub-
stituted pyridine ring as the heterocyclic aglycone was desired.[5–8] Such a
route could provide access to a variety of analogues of pyrimidine nucle-
osides with novel H-bonding patterns.[9–11] Such molecules might display
pharmaceutically useful antimetabolite activity.[12–14] In recent reports from
Received 1 August 2014; accepted 27 June 2015.
Address correspondence to Galal Elgemeie, Chemistry Department, Faculty of Science, Helwan
University, Helwan, Cairo, Egypt. E-mail: elgemeie@hotmail.com
Color versions of one or more of the figures in the article can be found online at
www.tandfonline.com/lncn.
659

660
G. Elgemeie et al.
our laboratory, we described the preparation of different novel functional-
ized pyridine thioglycosides, which revealed antagonistic activity.[15] In an
earlier communication, we had already reported the use of dihydropyridine
thioglycosides as substrates or inhibitors of protein glycosylation.[16] These
common features encouraged us to develop a new, straightforward route
for the synthesis of pyridine thioglycosides. Continuing our efforts for the
development of simple, eco-friendly, and cost-effective methodologies, we
report here a novel synthesis of pyridine-4-thioglycosides. This new protocol
has been applied to a variety of substituted pyridine-4-thioglycosides with
excellent yield. As far as we know, this is the first report of a pyridine-4-
thioglycoside.
2. RESULTS AND DISCUSSION
2.1. Chemistry
It has been found that reaction of malononitrile with carbon disulfide
in the presence of sodium ethoxide gives the sodium 2,2-dicyanoethene-
1,1-bis(thiolate) 2. Compounds 2 are readily reacted with one equiva-
lent of 2-cyano-N -arylacetamides 1a–e in the presence of sodium ethox-
ide to give the corresponding sodium 6-amino-3,5-dicyano-2-oxo-1-aryl-1,2-
dihydropyridine-4-thiolate derivatives 3a–e in good yields. Compounds 3 re-
acted with 2,3,4,6-tetra-O-acetyl-α–D-gluco- and galacto-pyranosyl bromides
4a,b in dimethylformamide at room temperature to give in high yield the
corresponding S-glycosides 5a–j (Scheme 1). It is suggested that the cis-
(α)sugars donors react with 3 by a simple SN2 reaction to give the β-glycoside
products.[17] The structures of 5 were established on the basis of its elemental
analysis and spectral data (IR, 1H NMR, and 13C NMR). For example, the an-
alytical data for 5a revealed a molecular formula C27H26N4O10S, its 1H NMR
spectrum showing the anomeric proton as a doublet at δ 5.21–5.23 ppm with
a spin–spin coupling constant of 10.50 Hz indicating the β-configuration.
The other six glucose protons resonated at δ 3.78–5.01 ppm. When glyco-
sides 5 were treated with methanolic ammonia at room temperature for 10
minutes, the free glycoside derivatives 6a,b were obtained in almost quan-
titative yields (Scheme 2), the structures of which were established on the
basis of elemental analysis and spectral data. Thus, the analytical data for
1
6a reveal the molecular formula C19H18N4O6S. The H NMR spectrum
shows the anomeric proton as a doublet at δ 5.79–5.81 (J 1’-2’ 10.0 Hz),
indicating the presence of only the β-D-configuration. Encouraged by these
results, we decided to synthesize the pyridine-4-thioglycosides 5 using the
reaction of 1a–e with [bis(mercapto)methylene]malononitrile 7 and com-
paring the resulting products for stereochemical considerations. Thus, in
a simple experimental procedure, treatment of compounds 1a–e with 7 in

First Synthesis of Pyridine-4-Thioglycosides and Their Cytotoxic Evaluation
661
sodium ethoxide, followed by acidification afforded the corresponding 4-
mercaptopyridines 9. The latter compounds were treated with peracetylated
sugars 4 in sodium ethoxide-dimethylformamide (DMF) at room temper-
ature to afford the S-glycosyl compounds 5. The latter were shown to be
the same as those obtained from the reaction of 3 with 4 by comparison
of their melting points and spectral data. Upon methylation of compounds
9 with methyl iodide in sodium ethoxide gave the 4-methylsulfanylpyridine
products 10a–e. Compounds 10 can also be prepared by reaction of 1a-e
with [bis(methylthio)methylene]malononitrile 8 in sodium ethoxide. When
compounds 5 and 10 were subjected to the reaction with hydrazine, the
pyrazolo[4,3-c]pyridines 11 were obtained[18] (Scheme 2).
SCHEME 1 Synthetic pathway for pyridine-4-thioglycosides 5a-j, 6a,b.

662
G. Elgemeie et al.
SCHEME 2 Synthesized derivatives 9a-e, 10a-e, 11a-e.
2.2. Antitumor Activity
The 10 of newly synthesized pyridine and pyridine-4-glucopyranosylthio
5a–d, 5h, 6b, 9a,c,d, 10a were selected and evaluated for in vitro anticancer
activity against human cancer cells according to the standard procedures
described in experimental part. The tumor cell lines panel consisted of
four human tumor cell lines, namely, hepatocellular carcinoma HEPG2,
colon carcinoma HCT116, prostate carcinoma PC3, and lung carcinoma
A549, which were incubated with 10 concentrations (0.19–100 μM) for each
compound. Cell viability was assessed by the mitochondrial-dependent re-
duction of yellow MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium
bromide) to purple formazan. A probit analysis was carried for LC50 deter-
mination using SPSS 11 program. The antitumor drug doxorubicin was used

First Synthesis of Pyridine-4-Thioglycosides and Their Cytotoxic Evaluation
663
TABLE 1 Cytotoxicity of the synthesized candidates on A549, HCT116, HEPG2, and PC3 cancer cell
lines
LC50a (μM)
Compound no.
(A549)
(HCT116)
(HEPG2)
(PC3)
5h
6b
9c
—
—
—
—
—
—
73.3
45.6
82.5
37.8
83.3
67.6
—
Doxorubicin
48.8
65.1
41.1
LC50a is lethal concentration of the sample, which causes the death of 50% of cells in 48 hours. Cancer
cell lines A549 (lung cell line), HCT116 (colon cell line), HEPG2 (liver cell line), and PC3 (prostate cell
line).
as positive control. The results are presented in Table 1. LC50 (cytostatic ac-
tivity) values were based on dose–response curves (LC50 value, defined as the
concentration corresponding to 50% growth inhibition). The tested com-
pounds 5a–d, 9a,d, and 10a exhibited weak antitumor activity toward both
the four cell lines (at 100 μM the values of mortality of cancer cell lines range
between 15% and 58%). While compounds 5h, 6b, 9c proved to be selective
toward hepatocellular carcinoma HEPG2 cancer cell line with LC50 values
of 73.3, 45.6, 82.5, respectively (Figure 1). On the other hand, compounds
5h and 6b displayed a considerable antitumor activity toward prostate car-
cinoma PC3 with LC50 values of 83.3 and 67.6, respectively (Figure 2). This
means that these compounds displayed selectivity on particular cell lines
FIGURE 1 Representative graph showing observed responses of HEPG2 cell in the presence of increasing
concentrations of compounds 5h, 6b, and 9c.

664
G. Elgemeie et al.
FIGURE 2 Representative graph showing observed responses of PC3 cell in the presence of increasing
concentrations of compound 5h and 6b.
(hepatocellular carcinoma HEPG2 and prostate carcinoma PC3). Pyridine
carrying two substituents (5h and 6b) especially when contain thioglycosidic
linkage at C4 and Para methoxy phenyl at N showed more activity toward
HEPG2 and prostate carcinoma cell lines PC3. Substitution of electron re-
leasing groups such as methoxy on the benzene ring present at 4-position
increased the anticancer activity and has shown good activity, whereas the
introduction of electron attracting chloro, group on benzene ring decreased
the activity and has shown weak activity. Compound 6b has more cytotoxic
activity. The results suggested that, pyridine-N -Para methoxyphenyl with de-
protected thiogalactosidic linkage at C4 played a vital role in the modulation
of cytotoxic activity.
3. CONCLUSION
In conclusion, we have developed a new and simple method for the
synthesis of the first pyridine-4-thioglycosides. The mild reaction conditions,
clean reaction profiles, zero side product, and cost efficiency render this
approach as useful and innovative to the existing methods for glycoside
formation. Further studies on the application of this method for the synthesis
of other highly functionalized biologically active glycosides are underway.
The antitumor activities of the synthesized compounds were tested against
human tumor cell lines; lung (A549), colon (HCT116), liver (HEPG2), and
prostate (PC3).

First Synthesis of Pyridine-4-Thioglycosides and Their Cytotoxic Evaluation
665
4. EXPERIMENTAL PROTOCOLS
4.1. Chemistry
All melting points were measured on a Gallenkamp melting point appa-
ratus. The 1H NMR spectra were measured on a Jeol-500 MHz spectrometer
for solutions DMSO-d6 and CDCl3 using Si(CH3)4 as an internal standard at
National Research Center, Cairo, Egypt. Progress of the reactions was mon-
itored by thin-layer chromatography (TLC) using aluminum sheets coated
with silica gel F254 (Merck). Viewing under a short-wavelength UV lamp
(Spectronics Corporation, USA) effected detection.
4.1.1. General Procedure for the Synthesis of 3a–e
A mixture of 2-cyano-N -arylacetamide 1a–e (10 mmol) and sodium
2,2-dicyanoethene-1,1-bis(thiolate) 2 (10 mmol) in EtOH (30 ml), sodium
ethoxide (10 mmol) was refluxed for 4 hours. The reaction mixtures were
cooled to room temperature and triturated with MeOH to afford compounds
3a–e, which were collected by filtration and recrystallized from the appro-
priate solvent. Yield (90%).
4.1.1.1. Sodium 6-amino-3,5-dicyano-2-oxo-1-phenyl-1,2-dihydropyridine-4-thi-
olate (3a). Brown solid; yield (90%), mp > 300◦C; IR (KBr, cm−1) υ 3434,
3396 (NH2), 3089 (CH), 2207 (CN), 1692 (CO). C13H7N4NaOS.
4.1.1.2. Sodium 6-amino-3,5-dicyano-2-oxo-1-p-tolyl-1,2-dihydropyridine-4-thi-
olate (3b). Brown solid; yield (85%), mp > 300◦C; IR (KBr, cm−1) υ 3427,
3368 (NH2), 3008 (CH), 2213 (CN), 1689 (CO). C14H9N4NaOS.
4.1.1.3. Sodium 6-amino-3,5-dicyano-1-(4-methoxyphenyl)-2-oxo-1,2-dihydrop-
yridine-4-thiolate (3c). Brown solid; yield (85%), mp > 300◦C; IR (KBr,
cm−1) υ 3436, 3373 (NH2), 1387 (CH3), 2215 (CN), 1690 (CO).
C14H9N4NaO2S.
4.1.1.4. Sodium 6-amino-1-(4-chlorophenyl)-3,5-dicyano-2-oxo-1,2-dihydropyr-
idine-4-thiolate (3d). Brown solid; yield (85%), mp > 300◦C. C13H6ClN4
NaOS.
4.1.1.5. Sodium 6-amino-3,5-dicyano-1-(4-(naphthalen-2-yl)phenyl)-2-oxo-1,2
dihydropyridine-4-thiolate (3e). Brown solid; yield (83%), mp > 300◦C.
C17H9
N4NaOS.
4.1.2. General Procedures for the Synthesis of 5a–j
A solution of compound 3a–e (10 mmol) in dry DMF (20 ml) was stirred
at room temperature, then a solution of 2,3,4,6-tetra-O-acetyl-α-D-gluco- or
galactopyranosyl bromide 4 in DMF (10 ml) was dropped within 30 minutes
and the reaction mixture was stirred at room temperature until compilation
(TLC, 6–8 hours). After completion, the reaction mixture was poured onto

666
G. Elgemeie et al.
water. The resulting solid was filtered off, purified by column chromatog-
raphy (petroleum ether 40–60/ethyl acetate 3:1) and recrystallized from
ethanol to give the compounds (5a–j).
4.1.2.1. 6-Amino-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-glucopyranosylthio)-2-oxo-1-
phenyl-1,2-dihydropyridine-3,5-dicarbonitrile (5a). Yellow solid; (EtOH),
yield (86%), mp 210◦C, 1H NMR (500 MHz, DMSO): δ 1.95–2.46 (4s, 12H,
4xOAc), 3.87 (m, 2H, 2H-6’), 4.20 (m, 1H, H-5’), 4.53 (t, 1H, J = 9.5 Hz,
H-4’), 4.78 (t, 1H, J = 9.3 Hz, H-3’), 5.01(t, 1H, J = 9.5 Hz H-2’), 5.21–5.23
(d, 1H, J 1’-2’ = 10.50 Hz, H-1’), 6.32 (s, 2H, NH2), 7.47–7.70 (m, 5H, C6H5).
13C NMR: δ 21.25 (4CH3CO), 62.12 (C-6’), 68.64 (C-4’), 70.22 (C-2’), 73.22
(C-3’), 79.13 (C-5’), 81.23 (C-1’), 113.26 (CN), 116.11 (CN), 126.25 (Ar-C),
128.43 (2C, Ar-C), 129.75 (2C, Ar-C), 132.51 (Ar-C), 150.25 (C-5), 158.10
(C-3), 160.34 (pyridine C-4), 168.34 (CO), 171.52 (C-6), 176.52 (4CO).
UV (EtOH): 265, 311, 390 nm. Anal. Calcd. for C27H26N4O10S (598.58): C,
54.18; H, 4.38; N, 9.36; S, 5.36%. Found: C, 54.2; H, 4.2; N, 9.2; S, 5.4%.
4.1.2.2. 6-Amino-1-(4-methylphenyl)-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-glucopyra-
nosylthio)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile (5b). Yellow
solid;
(EtOH), yield (82%), mp 198◦C, 1H NMR (500 MHz, DMSO-d6): δ
1.93–1.95 (4s, 12H, 4xOAc), 2.60 (s, 3H, CH3), 3.89–3.92 (m, 2H, H-6’
H-6”), 4.11 (m, H, H-5’), 4.90 (t, 1H, J = 9.5 Hz, H-4’), 5.14 (t, 1H, J =
9.3 Hz, H-3’), 5.31 (t, 1H, J = 9.5 Hz, H-2’), 5.58–5.60 (d, 1H, J 1’-2’ = 6.90 Hz,
H-1’), 6.88 (s, 2H, NH2) 7.36–7.51 (m, 4H, C6H4). 13C NMR: δ 17.55 (CH3),
20.00 (4CH3CO), 60.10 (C-6’), 66.00 (C-4’), 69.15 (C-2’), 71.00 (C-3’), 76.10
(C-5’), 80.00 (C-1’), 110.20 (CN), 113.00 (CN), 122.20 (Ar-C), 125.21 (2C,
Ar-C), 126.23 (2C, Ar-C), 130.99 (Ar-C), 149.20 (C-5), 156.00 (C-3), 162.00
(pyridine C-4), 166.99 (CO), 170.77 (C-6), 178.92 (4CO). UV (EtOH): 260,
300, 399 nm. Anal. Calcd. for C28H28N4O10S (612.61): C, 54.90; H, 4.61; N,
9.15; S, 5.23%. Found: C, 54.9; H, 4.5; N, 9.2; S, 5.3%.
4.1.2.3. 6-Amino-1-(4-methoxyphenyl)-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-gluco-
pyranosylthio)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile (5c). Yellow solid;
1
(EtOH), yield (82%), mp 182◦C, H NMR (500 MHz, CDCl3): δ 2.14 (4s,
12H, 4xOAc), 2.99 (s, 3H, OCH3), 3.94 (m, 2H, 2H-6’), 4.14 (m, 1H, H-5’),
4.37 (t, 1H, J = 9.2 Hz, H-4’), 4.65 (t, 1H, J = 9.1 Hz, H-3’), 4.79 (t, 1H, J
= 9.3 Hz, H-2’), 5.09–5.10 (d, 1H, J 1’-2’ = 5.70 Hz, H-1’), 6.42–6.51 (s, 2H,
NH2), 7.25–7.33 (m, 4H, C6H4). Anal. Calcd. for C28H28N4O11S (628.61):
C, 53.50; H, 4.49; N, 8.91; S, 5.10%. Found: C, 53.4; H, 4.4; N, 8.8; S, 5.1%.
4.1.2.4. 6-Amino-1-(4-chlorophenyl)-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-glucopyra-
nosylthio)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile (5d). Yellow
solid;
(EtOH), yield (81%), mp 175◦C, 1H NMR (500 MHz, DMSO-d6): δ
1.84–1.96 (4s, 12H, 4xOAc), 3.91 (m, 2H, 2H-6’), 4.21 (m, 1H, H-5’),
5.22–5.30 (m, 2H, H-4’ and H-3’), 5.43 (t, 1H, J = 9.5 Hz, H-2’), 6.18–6.19
(d, 1H, J 1’-2’ = 5.00 Hz, H-1’), 6.66 (s, 2H, NH2), 7.50–7.81 (m, 4H, C6H4). 13C
NMR: δ 19.99 (4CH3CO), 61.77 (C-6’), 67.68 (C-4’), 69.99 (C-2’), 72.56
(C-3’), 78.89 (C-5’), 80.94 (C-1’), 112.73 (CN), 115.87 (CN), 124.89 (Ar-C),

First Synthesis of Pyridine-4-Thioglycosides and Their Cytotoxic Evaluation
667
127.93 (2C, Ar-C), 129.00 (2C, Ar-C), 131.93 (Ar-C), 151.84 (C-5), 156.83
(C-3), 161.74 (pyridine C-4), 169.93 (CO), 171.52 (C-6), 176.52 (4CO). UV
(EtOH): 270, 320, 390 nm. Anal. Calcd. for C27H25ClN4O10S (633.03): C,
51.23; H, 3.98; Cl, 5.60; N, 8.85; S, 5.07%. Found: C, 51.3; H, 3.9; N, 8.7; S,
5.1%.
4.1.2.5. 6-Amino-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-glucopyranosylthio)-1-(napht-
halen-2-yl)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile (5e). Yellow
solid;
(EtOH), yield (80%), mp 206◦C 1H NMR (500 MHz, DMSO-d6): δ 2.03–2.19
(4s, 12H, 4xOAc), 3.92 (m, 1H, H-5’), 4.25–4.26 (t, 1H, H-4’), 4.46–4.52
(m, 2H, 2H-6’), 4.72 (t, 1H, J = 9.3 Hz, H-3’), 5.02 (t, 1H, J = 9.5 Hz,
H-2’), 5.37–5.38 (d, 1H, J 1’-2’ = 7.00 Hz, H-1’), 6.24 (s, 2H, NH2), 7.37–7.69
(m, 7H, C10H7). UV (EtOH): 270, 319, 385 nm. UV (EtOH): 266, 317,
392 nm. Anal. Calcd. for C31H28N4O10S (648.64): C, 57.40; H, 4.35; N, 8.64;
S, 4.94%. Found: C, 57.5; H, 4.3; N, 8.5; S, 4.8%.
4.1.2.6. 6-Amino-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-galactopyranosylthio)-2-oxo-
1-phenyl-1,2-dihydropyridine-3,5-dicarbonitrile (5f). Yellow solid; (EtOH),
yield (83%), mp 212◦C, IR (KBr, cm−1) υ 3065, 3033 (NH2), 2956 (CH),
2225 (CN), 1601 (CO). 1H NMR (500 MHz, DMSO-d6): δ 1.96–2.12 (4s, 12H,
4xOAc), 3.73–3.81 (m, 1H, H-5’), 4.04 (t, 1H, J = 9.2 Hz, H-4’), 4.25 (t, 1H,
J = 9.5 Hz, H-3’), 4.46–4.50 (t, 1H, J = 9.3 Hz, H-2’), 4.82 (m, 2H, 2H-6’),
5.29–5.32 (d, 1H, J 1’-2’ = 11.25 Hz, H-1’), 6.26–6.31 (s, 2H, NH2) 7.13–7.66
(m, 5H, C6H5). 13C NMR: δ 19.00 (4CH3CO), 61.10 (C-6’), 67.00 (C-4’),
71.56 (C-2’), 73.00 (C-3’), 78.95 (C-5’), 80.83 (C-1’), 114.00 (CN), 117.34
(CN), 123.99 (Ar-C), 126.00 (2C, Ar-C), 128.76 (2C, Ar-C), 131.67 (Ar-C),
151.00 (C-5), 156.43 (C-3), 161.78 (pyridine C-4), 165.98 (CO), 170.26 (C-6),
175.00 (4CO). Anal. Calcd. for C27H26N4O10S (598.58): C, 54.18; H, 4.38;
N, 9.36; S, 5.36%. Found: C, 54.2; H, 4.3; N, 9.3; S, 5.2%.
4.1.2.7. 6-Amino-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-galactopyranosylthio)-2-oxo-
1-p-tolyl-1,2-dihydropyridine-3,5-dicarbonitrile (5g). Yellow solid; (EtOH),
yield (85%), mp 201◦C, 1H NMR (500 MHz, CDCl3): δ 2.21–2.34 (4s, 12H,
4xOAc), 2.41 (s, 3H, CH3), 3.90 (m, 1H, H-5’), 4.23 (m, 2H, 2H-6’), 4.72 (t,
1H, J = 9.6 Hz, H-4’), 4.92 (t, 1H, J = 9.4 Hz, H-3’), 5.12 (t, 1H, J = 9.5 Hz,
H-2’), 5.33–5.35 (d, 1H, J 1’-2’ = 10.50 Hz, H-1’), 6.52 (s, 2H, NH2) 7.51–7.61
(m, 4H, C6H4). Anal. Calcd. for C28H28N4O10S (612.61): C, 54.90; H, 4.61;
N, 9.15; S, 5.23%. Found: C, 54.9; H, 4.6; N, 9.2; S, 5.1%.
4.1.2.8. 6-Amino-1-(4-methoxyphenyl)-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-galact-
opyranosylthio)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile (5h). Yellow solid;
(EtOH), yield (84%), mp 195◦C, 1H NMR (500 MHz, DMSO-d6): δ 1.91–2.01
(4s, 12H, 4xOAc), 3.45 (s, 3H, OCH3), 3.96 (m, 1H, H-5’), 4.13–4.14 (m,
2H, 2H-6’), 4.60 (t, 1H, J = 9.5 Hz, H-4’), 4.89 (t, 1H, J = 9.4 Hz, H-3’),
5.12 (t, 1H, J = 9.5 Hz, H-2’), 5.41–5.43 (d, 1H, J 1’-2’ = 10.00 Hz, H-1’),
6.34 (s, 2H, NH2), 7.37–7.64 (m, 4H, C6H4). Anal. Calcd. for C28H28N4O11S
(628.61): C, 53.50; H, 4.49; N, 8.91; S, 5.10%. Found: C, 53.4; H, 4.3; N, 8.7;
S, 5.1%.

668
G. Elgemeie et al.
4.1.2.9. 6-Amino-1-(4-chlorophenyl)-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-galactopy-
ranosylthio)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile (5i). Yellow solid;
(EtOH), yield (86%), mp 188◦C purified by using column chromatography
(petroleum ether/ethyl acetate 3:1); 1H NMR (500 MHz, DMSO-d6): δ
1.97–2.13 (4s, 12H, 4xOAc), 3.93–4.08 (m, H, H-5’), 4.12–4.19 (m, 2H,
2H-6’), 4.72–4.82 (t, 1H, J = 9.5 Hz, H-4’), 4.85–5.16 (t, 1H, J = 9.4 Hz,
H-3’), 5.22 (t, 1H, J = 9.5 Hz, H-2’), 5.40–5.42 (d, 1H, J 1’-2’ = 9.70 Hz,
H-1’), 6.27 (s, 2H, NH2), 7.25–7.50 (m, 4H, C6H4). Anal. Calcd. for
C27H25ClN4O10S (633.03): C, 51.23; H, 3.98; Cl, 5.60; N, 8.85; S, 5.07%.
Found: C, 51.1; H, 3.7; Cl, 5.6; N, 8.7; S, 5.2%.
4.1.2.10.
6-Amino-4-(2’,3’,4’,6’-tetra-O-acetyl-β-D-galactopyranosylthio)-1-
(5j). Yellow
(nap-hthalen-2-yl)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile
solid; (EtOH), yield (85%), mp 205◦C, H NMR (500 MHz, DMSO-d6): δ
1.93–2.05 (4s, 12H, 4xOAc), 3.80–4.25 (m, 3H, 2H-6’ and H-5’), 4.79–5.28
(m, 2H, H-4’ and H-3’), 5.35 (t, 1H, J = 9.2 Hz, H-2’), 5.51–5.53 (d, 1H, J 1’-2’
= 10.00 Hz, H-1’), 6.18–6.20 (s, 2H, NH2), 7.16–8.11 (m, 7H, C10H7). Anal.
Calcd. for C31H28N4O10S (648.64): C, 57.40; H, 4.35; N, 8.64; S, 4.94%.
Found: C, 57.5; H, 4.2; N, 8.5; S, 4.8%.
1
4.1.3. Ammonolysis of 5a and 5h
To a solution of an individual nucleoside 5a or 5h (1 mmol) in anhydrous
MeOH (10 ml), the dry gaseous ammonia was passed at room temperature
for 10 minutes. The reaction mixture was stirred until the reaction was judged
complete by TLC using (petroleum ether/ethyl acetate) (Rf, 0.72–0.74). The
resulting mixture was then concentrated under reduced pressure to afford
a solid residue that was crystallized from appropriate solvent.
4.1.3.1. 6-Amino-4-(2’,3’,4’,6’-tetrahydroxy-β-D-glucopyranosylthio)-2-oxo-1-
phenyl-1,2-dihydropyridine-3,5-dicarbonitrile (6a). Yellow solid; (MeOH),
yield (86%), mp 195◦C; IR (KBr, cm−1) υ 3502 (OH), 3302–3219 (NH2),
2221 (CN), 1657 (CO). 1H NMR (500 MHz, DMSO-d6): δ 3.69–3.71 (m, 3H,
2H-6’ and H-5’), 3.90 (t, 1H, H-4’), 4.19 (s, 1H, OH-2’), 4.25–4.35 (m, 2H,
H-3’ and H-2’), 4.45 (s, 1H, OH-3’), 5.11–5.34 (m, 2H, OH-4’ and OH-6’),
5.79–5.81 (d, 1H, J 1’-2’ = 9.90 Hz, H-1’), 6.76 (s, 2H, NH2), 7.12–7.56 (m,
5H, C6H5). 13C NMR: δ 61.10 (C-6’), 67.73 (C-4’), 71.00 (C-2’), 74.09 (C-3’),
77.67 (C-5’), 80.63 (C-1’), 114.78 (CN), 116.78 (CN), 125.78 (Ar-C), 127.73
(2C, Ar-C), 130.00 (2C,Ar-C), 133.91 (Ar-C), 151.75 (C-5), 156.19 (C-3),
161.73 (pyridine C-4), 167.34 (CO), 172.00 (C-6). UV (EtOH): 255, 320,
380 nm. Anal. Calcd. for C19H18N4O6S (430.43): C, 53.02; H, 4.22; N, 13.02;
S, 7.45%. Found: C, 53.1; H, 4.1; N, 13.1; S, 7.3%.
4.1.3.2. 6-Amino-1-(4-methoxyphenyl)-4-(2’,3’,4’,6’-hydroxy-β-D-glucopyranos-
ylthio)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitrile
(6b). Yellow
solid;
1
(MeOH), yield (84%), mp 185◦C; H NMR (500 MHz, DMSO-d6): δ
3.18 (s, 3H, OCH3), 4.02–4.13 (m, 2H, 2H-6’), 4.34 (m, 2H, H-5’ and H-4’),
4.60 (s, 1H, OH-2’), 4.80 (s, 1H, OH-3’), 4.91–4.95 (m, 2H, H-3’ and H-2’),

First Synthesis of Pyridine-4-Thioglycosides and Their Cytotoxic Evaluation
669
5.15–5.38 (m, 2H, OH-4’ and OH-6’) 5.76–5.78 (d, 1H, J 1’-2’ = 10.30 Hz,
H-1’), 6.58–6.60 (s, 2H, NH2), 7.40–7.64 (m, 5H, C6H5). Anal. Calcd. for
C20H20N4O7S (460.46): C, 52.17; H, 4.38; N, 12.17; S, 6.96%. Found: C,
52.1; H, 4.2; N, 12.1; S, 6.8%.
4.1.4. General Procedure for the Synthesis of 9a–e
A mixture of each of 1a–e (10 mmol), 2-(dimercaptomethylene)
malononitrile) 7 (10 mmol) and sodium ethoxide (10 mmol) was dissolved in
EtOH (30 ml). The reaction mixture was refluxed for 3 hours. The progress
of the reaction was monitored by TLC until the reactants disappeared, then
the solution was concentrated, acidified with dilute HCl, and the remain-
ing residue was triturated with MeOH to afford 9a–e, which was purified
by column chromatography (petroleum ether 40–60/ethyl acetate 3:1) and
recrystallized from the appropriate solvent.
4.1.4.1. 6-Amino-4-mercapto-2-oxo-1-phenyl-1,2-dihydropyridine-3,5-dicarboni-
trile (9a). Brown solid; (MeOH), yield (89%), mp 185–186◦C, IR (KBr,
cm−1) υ 3437,3378 (NH2), 3049 (CH), 2217 (CN), 1640 (CO). 1H NMR
(500 MHz, DMSO-d6): δ 6.67 (s, 2H, NH2), 7.24–7.49 (m, 5H, C6H5), 11.67
(s, 1H, SH). 13C NMR: δ 61.00 (C-5), 113.09 (C-3), 115.87 (2C, Ar-c), 117.00
(2CN), 123.99 (Ar-C), 127.54 (2C, Ar-C), 158.00 (C-6), 160.00 (C-4), 162.33
(Ar-C), 166.00 (CO). Anal. Calcd. for C13H8N4OS (268.29): C, 58.20; H,
3.01; N, 20.88; S, 11.95%. Found: C, 58.4; H, 3.2; N, 20.8; S, 11.8%.
4.1.4.2. 6-Amino-4-mercapto-1-(4-methylphenyl)-2-oxo-1,2-dihydropyridine-3,5-
dicarbonitrile (9b). Brown solid; (MeOH), yield (83%), mp 198◦C, 1H NMR
(500 MHz, DMSO-d6) δ 2.36 (s, 3H, CH3), 6.32 (s, 2H, NH2), 7.20–7.41 (m,
4H, C6H4), 11.04 (s, 1H, SH); Anal. Calcd. for C14H10N4OS (282.32): C,
59.56; H, 3.57; N, 19.85; S, 11.36%. Found: C, 59.5; H, 3.6; N, 19.7; S, 11.2%.
4.1.4.3. 6-Amino-4-mercapto-1-(4-methoxyphenyl)-2-oxo-1,2-dihydropyridine-
1
3,5-dicarbonitrile (9c). Brown solid; (MeOH), yield (82%), mp 182◦C, H
NMR (500 MHz, DMSO-d6): δ 3.81 (s, 3H, OCH3), 6.60 (s, 2H, NH2),
7.33–7.90 (m, 4H, C6H4), 11.61 (s, 1H, SH). 13C NMR: δ 57.31 (CH3), 62.24
(C-5), 110.12 (C-3), 113.25 (2C, Ar-c), 115.22 (2CN), 122.43 (Ar-C), 129.14
(2C, Ar-C), 159.12 (C-6), 159.85 (C-4), 160.10 (Ar-C), 165.21 (CO). Anal.
Calcd. for C14H10N4O2S (298.32): C, 56.37; H, 3.38; N, 18.78; S, 10.75%.
Found: C, 56.4; H, 3.3; N, 18.6; S, 10.6%.
4.1.4.4. 6-Amino-1-(4-chlorophenyl)-4-mercapto-2-oxo-1,2-dihydropyridine-3,5
dicarbonitrile (9d). Brown solid; (MeOH), yield (80%), mp 179◦C, 1H NMR
(500 MHz, DMSO-d6): δ 6.81(s, 2H, NH2), 7.15–7.30 (m, 4H, C6H4), 13.15
(s, 1H, SH). Anal. Calcd. for C13H7ClN4OS (302.74): C, 51.58; H, 2.33;
Cl, 11.71; N, 18.51; S, 10.59%. Found: C, 56.3; H, 2.2; Cl, 11.7; N, 18.6; S,
10.5%.
4.1.4.5. 6-Amino-4-mercapto-1-(naphthalen-2-yl)-2-oxo-1,2-dihydropyridine-3,
5 dicarbonitrile (9e). Brown solid; (MeOH), yield (79%), mp 200◦C, IR
1
(KBr, cm−1) υ 3446, 3386 (NH2), 2963 (CH), 2189 (CN), 1626 (CO); H

670
G. Elgemeie et al.
NMR (500 MHz, DMSO-d6): δ 6.25 (s, 2H, NH2), 7.01–7.47 (m, 7H, C10H7),
11.82 (s, 1H, SH). Anal. Calcd. for C17H10N4OS (318.35): C, 64.14; H, 3.17;
N, 17.60; S, 10.07%. Found: C, 64.2; H, 3.2; N, 17.5; S, 10.1%.
4.1.5. General Procedure for the Synthesis of (10a–e)
4.1.5.1. Method A. A solution of compounds 1a–e (10 mmol), 2-
(bis(methylthio)methylene)malononitrile 8 (10 mmol), and sodium ethox-
ide (10 mmol) was dissolved in EtOH (20 ml). The reaction was refluxed
for 3 hours, monitored by TLC (petroleum ether 40–60–ethyl acetate 3:1)
until the reactants disappeared. The solvent was then evaporated and the
remaining residue was triturated with MeOH to afford compounds 10a–e,
which was separated by column chromatography and recrystallized from the
appropriate solvent.
4.1.5.2. Method B. A solution of compounds 9a–e (10 mmol) in sodium
ethoxide (10 mmol) was stirred at 50◦C for 30 minutes and allowed to cool
to room temperature and then methyl iodide (10 mmol) was slowly added to
the mixture. Then the reaction mixture was refluxed with stirring for 3 hours.
The formed solid product was collected by filtration and recrystallized from
appropriate solvent.
4.1.5.3. 6-Amino-4-(methylthio)-2-oxo-1-phenyl-1,2-dihydropyridine-3,5-dicarb-
onitrile (10a). Brown solid; (MeOH), yield (80%), mp 290–291◦C, IR (KBr,
cm−1) υ 3405, 3322 (NH2), 3044 (CH), 2947 (CH), 2207 (CN), 1650
(CO). 1H NMR (500 MHz, DMSO-d6): δ 2.69–2.71 (S, 3H, SCH3), 6.52 (s,
2H, NH2), 7.29–7.66 (m, 5H, C6H5). Anal. Calcd. for C14H10N4OS (282.32):
C, 59.56; H, 3.57; N, 19.85; S, 11.36%. Found: C, 59.5; H, 3.4; N, 19.7; S,
11.4%.
4.1.5.4. 6-Amino-4-(methylthio)-2-oxo-1-p-tolyl-1,2-dihydropyridine-3,5-dicarb-
onitrile (10b). Brown solid; (MeOH), yield (80%), mp 260◦C, IR (KBr,
cm−1) υ 3401, 3312 (NH2), 3034 (CH), 2947 (CH), 2207 (CN), 1645 (CO).
1H NMR (500 MHz, DMSO-d6): δ 2.38 (s, 3H, CH3), 2.78 (s, 3H, SCH3),
6.45 (s, 2H, NH2), 7.16–7.44 (m, 5H, C6H5). Anal. Calcd. for C15H12N4OS
(296.35): C, 60.79; H, 4.08; N, 18.91; S, 10.82%. Found: C, 60.8; H, 4.2; N,
18.8; S, 10.7%.
4.1.5.5. 6-Amino-1-(4-methoxyphenyl)-4-(methylthio)-2-oxo-1,2-dihydropyridi-
ne-3,5-dicarbonitrile (10c). Brown solid; (MeOH), yield (78%), mp 257◦C,
1H NMR (500 MHz, DMSO-d6): δ 2.35–2.43 (s, 3H, SCH3), 3.84 (s, 3H,
OCH3), 6.43 (s, 2H, NH2), 7.14–7.53 (m, 5H, C6H5). Anal. Calcd. for
C15H12N4O2S (312.35): C, 57.68; H, 3.87; N, 17.94; S, 10.27%. Found: C,
57.8; H, 3.7; N, 17.8; S, 10.1%.
4.1.5.6. 6-Amino-1-(4-chlorophenyl)-4-(methylthio)-2-oxo-1,2-dihydropyridine-
3,5-dicarbonitrile (10d). Brown solid; (MeOH), yield (79%), mp 282◦C
(MeOH); IR (KBr, cm−1) υ 3421, 3332 (NH2), 3044 (CH), 2949 (CH),
2219 (CN), 1696 (CO). Anal. Calcd. for C14H9ClN4OS (316.77): C, 53.08;

First Synthesis of Pyridine-4-Thioglycosides and Their Cytotoxic Evaluation
671
H, 2.86; Cl, 11.19; N, 17.69; S, 10.12%. Found: C, 53.1; H, 2.7; Cl, 11.2; N,
17.7; S, 10.1%.
4.1.5.7. 6-Amino-4-(methylthio)-1-(naphthalen-2-yl)-2-oxo-1,2-dihydropyridine-
3,5-dicarbonitrile (10e). Brown solid; (MeOH), yield (80%), mp 255◦C IR
(KBr, cm−1) υ 3407, 3314 (NH2), 3042 (CH), 2942 (CH), 2221 (CN), 1697
(CO). Anal. Calcd. for C18H12N4OS (332.38): C, 65.04; H, 3.64; N, 16.86; S,
9.65%. Found: C, 65.1; H, 3.5; N, 16.9; S, 9.6%.
4.1.6. General Procedure for the Synthesis of Pyrazolo[3,4-c]pyridines 11a–e
A mixture of 5a–j or 10a–e (10 mmol) and hydrazine hydrate (10 mmol)
was dissolved in ethanol (20 ml), and then a few drops of triethylamine were
added. The mixture was refluxed for 3 hours. The resulting precipitated
solid was filtered off and recrystallized from ethanol.
4.1.6.1. 3,5-Diamino-7-oxo-6-phenyl-6,7-dihydro-1H-pyrazolo[3,4-c]pyridine-4-
carbonitrile (11a). Yellow solid, mp > 300◦C; IR (KBr, cm−1) υ 3214 (NH,
NH2), 2198 (CN), and 1660 (CO). Anal. Calcd. for C13H10N6O (266.26): C,
58.7; H, 3.8; N, 31.6%. Found: C, 58.9; H, 4.0; N, 31.2%. M+, 266.
4.1.6.2. 3,5-Diamino-6-(4-methylphenyl)-7-oxo-6,7-dihydro-1H-pyrazolo[3,4-c]
pyridine-4-carbonitrile (11b). Yellow solid, mp 253◦C; IR (KBr, cm−1) υ
3500–3211 (NH, NH2), 2205 (CN), and 1690–1660 (CO). Anal. Calcd. for
C14H12N6O (280.28): C, 60.0; H, 4.3; N, 30.0%. Found: C, 59.6; H, 4.5; N,
30.4%. M+, 280.
4.1.6.3. 3,5-Diamino-6-(4-methoxyphenyl)-7-oxo-6,7-dihydro-1H-pyrazolo[3,4-
c]pyridine-4-carbonitrile (11c). Yellow solid, mp > 300◦C; IR (KBr, cm−1)
υ 3411, 3228 (NH, NH2), 2197 (CN), and 1680 (CN). Anal. Calcd. for
C14H12N6O2 (296.28): C, 56.8; H, 4.1; N, 28.4%. Found: C, 56.5; H, 4.3; N,
28.1%. M+, 296.
4.1.6.4. 3,5-Diamino-6-(4-chlorophenyl)-7-oxo-6,7-dihydro-1H-pyrazolo[3,4-c]
pyridine-4-carbonitrile (11d). Yellow solid, mp > 300◦C; IR (KBr, cm−1) υ
3401, 3328, 3230 (NH, NH2), 2202 (CN), and 1644 (CO). Anal. Calcd. for
C13H9ClN6O (300.70): C, 51.92; H, 3.02; N, 27.95%. Found: C, 51.8; H, 3.1;
N, 27.8%; M+, 300.
4.1.6.5. 3,4-Diamino-5-(naphthalen-2-yl)-6-oxo-5,6-dihydro-1H-pyrazolo[4,3-c]
pyridine-7-carbonitrile (11e). Yellow solid, mp > 300◦C; IR (KBr, cm−1) υ
3418, 3336, 3241 (NH, NH2), 2217 (CN), 1639 (CO). Anal. Calcd for
C17H12N6O (300.70): C, 64.55; H, 3.82; N, 26.57%. Found: C, 64.4; H, 3.7;
N, 26.5%.
4.2. Antitumor Screening
Cell viability was assessed by the mitochondrial-dependent reduction of
yellow MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide)
to purple formazan.[19]

672
G. Elgemeie et al.
Procedure: All the following procedures were done in a sterile area
using a Laminar flow cabinet biosafety class II level (Baker, SG403INT,
and Sanford, ME, USA). Cells were suspended in RPMI 1640 medium for
HEPG2-PC3 and HCT116–DMEM for A549. The media are supplemented
with 1% antibiotic–antimycotic mixture (10,000 U/ml Potassium Penicillin,
10,000 μg/ml Streptomycin Sulfate, and 25 μg/ml Amphotericin B), 1% L-
glutamine and 10% fetal bovine serum and kept at 37◦C under 5% CO2.
Cells were batch cultured for 10 days, then seeded at concentration of
10 × 103 cells/well in fresh complete growth medium in 96-well microtiter
plastic plates at 37◦C for 24 hours under 5% CO2 using a water jacketed
carbon dioxide incubator (Sheldon, TC2323, Cornelius, OR, USA). Media
was aspirated, fresh medium (without serum) was added, and cells were in-
cubated either alone (negative control) or with different concentrations of
sample to give a final concentration of (100-50-25-12.5-6.25-3.125-0.78 and
1.56 ug/ml). After 48 hours of incubation, medium was aspirated, 40 ul
MTT salt (2.5 μg/ml) were added to each well and incubated for further
4 hours at 37◦C under 5% CO2. To stop the reaction and dissolving the
formed crystals, 200 μl of 10% sodium dodecyl sulphate (SDS) in deionized
water was added to each well and incubated overnight at 37◦C. A positive
control composed of 100 μg/ml was used as a known cytotoxic natural agent
who gives 100% lethality under the same conditions. The absorbance was
then measured using a microplate multi-well reader (Bio-Rad Laboratories
Inc., model 3350, Hercules, California, USA) at 595 nm and a reference
wavelength of 620 nm. A statistical significance was tested between samples
and negative control (cells with vehicle) using independent t-test by SPSS
11 program. DMSO is the vehicle used for dissolution of plant extracts and
its final concentration on the cells was less than 0.2%. The percentage of
change in viability was calculated according to the formula: ((Reading of ex-
tract/Reading of negative control) − 1) × 100. A probit analysis was carried
for IC50 and IC90 determination using SPSS 11 program.
REFERENCES
1. Bozo, E.; Demeter, A.; Rilla, A.; Kuszmann, J. The behavior of two thiosugar thioglycosides towards
oxidation. Tetrahedron: Asymmetry 2001, 12(24), 3423–3433.
2. Despras, G.; Robert, R.; Sendid, B.; Machez, E.; Poulain, D.; Mallet, J. Biotin sulfone tagged oligo-
mannosides as immunogens for eliciting antibodies against specific mannan epitopes. Bioorg. Med.
Chem. 2012, 20, 1817–1831.
3. Suzuki, K.; Ito, Y.; Kanie, O. An improved method for the synthesis of protected glycosyl fluorides
from thioglycosides using N,N diethylaminosulfur trifluoride (DAST). Carbohydr. Res. 2012, 359,
81–91.
4. Komor, R.; Kasprzycka, A.; Gawołek, G.; Szeja, W. Simple synthesis of glycosylthiols and thioglycosides
by rearrangement of O-glycosyl thionocarbamates. Carbohydr. Res. 2014, 396, 37–42.
5. Elgemeie, G.H.; Elsayed, S.H.; Hassan, A.S. Direct route to a new class of acrylamide thioglycosides
and their conversions to pyrazole derivatives. Synth. Commun. 2008, 38, 2700–2706.
6. Elgemeie, G.H.; Elsayed, S.H.; Hassan, A.S. Design and synthesis of the first thiophene thioglycosides.
Synth. Commun. 2009, 39, 1781–1792.

First Synthesis of Pyridine-4-Thioglycosides and Their Cytotoxic Evaluation
673
7. Elgemeie, G.H.; Zaghary, W.A.; Amin, K.M.; Nasr, T.M. First synthesis of thiophene thioglycosides.
J. Carbohydr. Chem. 2009, 28, 161–172.
8. Elgemeie, G.H.; Eltamny, E.H.; Elgawad, I.I.; Mahmoud, N.M. Direct route to novel 2-(β-D-xylo- and
arabinopyranosylthio)dihydropyridine glycosides and their corresponding dehydrogenated forms.
Synth. Commun. 2009, 39, 443–458.
9. Elgemeie, G.H.; Hussein, M.M.; Jones, P.G. 2-(2’,3’,4’,6’-Tetra-O-acetyl-β-D-glucopyranosylthio)-
4-pyridin-4-yl 6,7,8,9-tetrahydro-5H-cyclohepta[b]pyridin-3-carbonitrile. Acta Cryst. 2002, E58,
1244–1245.
10. Elgemeie, G.H.; Attia, A.M.; Hussain, B.A. A synthetic strategy to a new class of cycloalkane ring-fused
pyridine nucleosides as potential anti-HIV agents. Nucleos. Nucleot. 1998, 17, 855–868.
11. Elgemeie, G.H.; Mansour, O.A.; Metwally, N.H. Synthesis and anti-HIV activity of different novel
nonclassical nucleosides. Nucleos. Nucleot. 1999, 18, 113–123.
12. Elgemeie, G.H.; Zaghary, W.A.; Amin, K.M.; Nasr, T.M. First synthesis of thienopyrazole thioglyco-
sides. J. Carbohydr. Chem. 2008, 27, 345–355.
13. Jandu, K.S.; Selwood, D.L. 4’-Thio-nucleosides via in situ pyranose-furanose rearrangements: a short
synthesis of the antiherpes agent 2’-deoxy-5-ethyl-4’-thiouridine via direct coupling of a silylated
pyrimidine base with a 4-thiopyranose sugar. J. Org. Chem. 1995, 60, 5170–5173.
14. Elgemeie, G.H.; Hussein, M.M.; Al-Khursani, S.A. A total synthesis of a new class of biazine thiogly-
cosides. J. Carbohydr. Chem. 2004, 23, 465–475.
15. Elgemeie, G.H.; Mahdy, E.M.; Elgawish, M.A.; Ahmed, M.M.; Shousha, W.G.; Eldin, M.E. A new class
of antimetabolites: pyridine thioglycosides as potential anticancer agents. Z. Naturforsch. 2010, 65c,
577–587.
16. Scala, S.; Akhmed, N.; Rao, U.S.; Paull, K.; Lan, L.; Dickstein, B.; Lee, J.; Elgemeie, G.H.; Stein, W.D.;
Bates, S.E. P-Glycoprotein substrates and antagonists cluster into two distinct groups. Mol. Pharmacol.
1977, 51, 1024–1033.
17. Elgemeie, G.H.; Attia, A. A new class of dihydropyridine thioglycosides via piperdinium salts. Synth.
Commun. 2003, 33, 2243–2255.
18. Elgemeie, G.H.; Ali, H.A.; Elghandour, A.H.; Abd-Elaziz, G.W. Synthesis of novel derivatives of 4-
methylthio-N-aryl-2-pyridone and deazapurine analogues: The reaction of ketene dithioacetals with
substituted acetanilides. Phosphorus, Sulfur Silicon 2000, 164, 189–197.
19. Mosmann, T. Rapid colorimetric assay for cellular growth and survival: Application to proliferation
and cytotoxicity assays. J. Immunol. Methods 1983, 65, 55–63.