Synthesis and antitumor activity of 5′-deoxy-4′-thio-l- nucleosides

Article abstract of DOI:10.1111/j.1747-0285.2010.00967.x

A series of novel 5′-deoxy-4′-thio-l-nucleosides was designed and synthesized. The absolute configuration of the target compound 23α was confirmed by X-ray crystallography. The antitumor activities of the target compounds were tested against the growth of human carcinoma of colon (LOVO), human leukemia cell line (CEM) and human breast cancer cell line (MDA-MB-435) cells in vitro. 6-cyclopentylamino and 6-cyclohexylamino purine compounds 26 and 27, both in α-configuration and in β-form, exhibited strong inhibition to CEM.

Full text of DOI:10.1111/j.1747-0285.2010.00967.x

ª 2010 John Wiley & Sons A/S
doi: 10.1111/j.1747-0285.2010.00967.x
Chem Biol Drug Des 2010; 75: 619–627
Research Article
Synthesis and Antitumor Activity of 5¢-deoxy-4¢-
thio-L-nucleosides
Materials and Methods
Liqing Cong*, Weicheng Zhou, Dongzhe
Jin, Juan Wang and Xiuhua Chen
Reagents and analysis
Melting points were determined on an Electrothermal Melting Point
Apparatus and were uncorrected. 1H NMR spectra were recorded
on 400 MHz instruments in CDCl3 or DMSO-d6 with tetramethylsi-
lane as an internal standard. Chemical shifts are reported in parts
per million (d), and signals are expressed as s (singlet), d (doublet),
t (triplet), m (multiplet), b (broad).
State Key Laboratory of New Drug & Pharmaceutical Process,
Shanghai Institute of Pharmaceutical Industry, 1111 Zhongshanbeiyi
Road, Shanghai 200437, China
*Corresponding author: Liqing Cong, cong75@139.com
A series of novel 5¢-deoxy-4¢-thio-L-nucleosides was
designed and synthesized. The absolute configura-
tion of the target compound 23a was confirmed by
X-ray crystallography. The antitumor activities of
the target compounds were tested against the
growth of human carcinoma of colon (LOVO),
human leukemia cell line (CEM) and human breast
cancer cell line (MDA-MB-435) cells in vitro. 6-cy-
clopentylamino and 6-cyclohexylamino purine
compounds 26 and 27, both in a-configuration and
in b-form, exhibited strong inhibition to CEM.
Synthesis
(2S,3S,4R)-2-tert-butyldimethylsilyloxymethyl-3,4-di-
hydroxy-3,4-O-isopropylidene-tetrahydrothiophene (3)
A solution of dimesylate 2 (32.1 g, 70 mmol) and Na2SÆ9H2O (20 g,
80 mmol) in DMF (700 mL) was stirred at 100 ꢀC for 5 h. After
being cooled to room temperature, H2O was added. The mixture
was extracted with Et2O (5 · 200 mL). The organic layer was dried
with MgSO4 and concentrated in vacuo. The residue was purified
by silica gel column chromatography to afford 3 as the syrup
(13.8 g, yield = 65%).
Key words: antitumor activity, L-thionucleosides, X-ray crystallo-
graphy
Received 25 October 2009, revised 9 February 2010 and accepted for
publication 14 February 2010
[a]D = +194 (C = 1.0, MeOH), ESI-MS m ⁄ z 305 (M++1), 1H NMR
25
(CDCl3, 400 MHz, dppm); 0.89 (s, 9H), 0.08 (s, 6H), 1.29 (s, 3H), 1.41
(s, 3H), 2.86 (t, J = 2.0, J = 1.6, 2H), 3.32–3.37 (m, 1H), 3.78 (dd,
J = 7.2, J = 10.0, 1H), 4.03 (dd, J = 7.6, J = 10.4, 1H), 4.69–4.72
(m, 1H), 4.84–4.87 (m, 1H).
4¢-Thionucleosides may be considered as the analogues by the
replacement of the lactol oxygen of nucleosides with a sulfur atom
and exhibit many biological activities, for example antiviral (1) and
antitumor (2). Moreover, they have some inherent advantages over
4¢-oxonucleosides that show a more stable glycosyl bond and
increased metabolic stability against various viral and cellular
enzymes (3).
(2S,3S,4R)-2-hydroxymethyl-3,4-dihydroxy-3,4-O-iso-
propylidene-tetrahydrothiophene (3¢)
Compound 3 (1 g, 3.3 mmol) was dissolved in tetrahydrofuran (THF)
(10 mL), and then TBAFÆ3H2O (1.3 g, 4.1 mmol) was added. The mix-
ture was stirred at room temperature for 3 h. After the solvent was
removed under reduced pressure, the residue was partitioned
between ethyl acetate and H2O. The organic layer was washed with
Some 5¢-Deoxy nucleosides (e.g., doxifluridine and capecitabine)
posses superior antitumor activity against various experimental
tumors in animals and cause considerably less host toxicity com-
pared to the well-known clinically used 5-FU, 5-fluorodeoxyuridine
(FdUrs), and Ftorafur (Ft) (4,5).
B
S
S
So far, no reports on 5¢-deoxy-4¢-thionucleosides have been found
in literatures, but it is logical to assume that such nucleosides
of both purines and pyrimidines would have stable glycosyl bond
and superior antitumor activity. Here, we report the synthesis
and the antitumor activities in cell culture of a series of
5¢-deoxy-4¢-thio-L-a-nucleosides and 5¢-deoxy-4¢-thio-L-b-nucleosides
(Figure 1).
B
H3C
H3C
OH OH
OH OH
α
5′-deoxy-4′-thio-L- -nucleosides
5′-deoxy-4′-thio-L- -nucleosides
β
B = pyrimidines or N6-substituented-2-chloro-purines
Figure 1: Designed compounds of 5¢-deoxy-4¢-thio-L-nucleosides.
619

Cong et al.
[a]D = )179.2 (C = 1.0, MeOH), ESI-MS m ⁄ z 381 (M++23), 1H
25
H2O followed by brine. The organic layer was dried with MgSO4 and
concentrated in vacuo. The residue was crystallized from acetonitrile
to obtain crystal 3¢.
NMR (CDCl3, 400 MHz, dppm); 1.33 (s, 3H), 1.50 (s, 3H), 2.05 (s,
3H), 3.29–3.33 (m, 1H), 3.46 (t, J = 10.0, J = 9.6, 1H), 3.95–3.97 (m,
1H), 4.84–4.89 (m, 2H), 6.00 (s, 1H).
25
[a]D = +62.2 (C = 1.0, MeOH), mp 70–72 ꢀC, ESI-MS m ⁄ z 191
1
(M++1), H NMR (CDCl3, 400 MHz, dppm); 1.28 (s, 3H), 1.47 (s, 3H),
(3R,4S,5S)-2-acetyloxy-3,4-dihydroxy-3,4-O-isopropy-
lidene-5-methyl-tetrahydrothiophene (8)
2.62–2.89 (m, 2H), 3.30–3.34 (m, 1H), 3.82–3.86 (m, 1H), 3.89–3.92
(m, 1H), 4.76–4.79 (m, 1H), 4.83–4.87 (m, 1H).
Compound 7 (6.2 g, 17.3 mmol) and AIBN (1.42 g, 8.65 mmol) were
dissolved in toluene (100 mL) in a three-necked flask. N2 was bub-
bled through the reaction mixture for 15 min, and tributhyltrin
hydride (0.72 mL, 26.0 mmol) was added. The reaction mixture was
heated in an oil bath at 80 ꢀC for 30 min. The reaction was cooled
to room temperature, washed with water, dried by MgSO4 and fil-
tered. The filtrate was evaporated to dryness, and the residue was
purified by silica gel column chromatography to afford 8 as white
solid (2.67 g, yield = 67%).
Crystal data: C8H14O3S, monoclinic, space group P21, unit cell
dimensions a = 12.6283(5), b = 5.7460(2), c = 13.2081(5) ꢀ, a = c =
90.00ꢀ, b = 94.112(2)ꢀ, V = 955.94(6) ꢀ3, Z = 2, 1790 reflections
were measured, and 1782 independent reflections were used for
the structure solution. Final R values are as follows: wR2 = 0.1035,
R1 = 0.0389, S = 1.004.
(2S,3S,4R)-2-tert-butyldimethylsilyloxymethyl-3,4-dih-
ydroxy-3,4-O-isopropylidene-tetrahydrothiophene-1-
oxide (4)
25
[a]D = )212.6 (C = 1.0, MeOH), mp 44–46 ꢀC, ESI-MS m ⁄ z 255
(M++23), 1H NMR (CDCl3, 400 MHz, dppm); 1.32 (s, 3H), 1.50 (s,
3H), 1.38 (d, J = 6.8, 3H), 2.05 (s, 3H), 3.69–3.72 (m, 1H), 4.70–4.73
(m, 1H), 4.77–4.78 (m, 1H), 5.94 (s, 1H).
Ozone gas was bubbled through a clear solution of 3 (7.4 g,
24.3 mmol) in CH2Cl2 (100 mL) at )78 ꢀC. The reaction was
completed in 30 min, as indicated by persistence of a blue color.
Nitrogen gas was bubbled through the solution to remove excess
ozone until the blue color vanished. The reaction mixture was
allowed to warm to room temperature and concentrated under
reduced pressure. The residue was purified by column chromatogra-
phy to give sulfoxide 4 as a white solid (3.97 g, 51%).
2,6-dichloro-9-((3R,4S,5S)-3,4-dihydroxy-3,4-O-isopro-
pylidene-5-methyl-tetrahydrothiophene-2-yl)purine (9)
To a stirred solution of compound 8 (3 g, 12.9 mmol) and 2,6-
dichloro-purine (6.11 g, 32.3 mmol) in dried acetonitrile (30 mL)
was dropped EtAlCl2 (17.7 mL, 15.9 mmol) at 0 ꢀC. The reaction
mixture was heated at 60 ꢀC for 3 h, cooled to 0 ꢀC and then
saturated aqueous solution of NaHCO3 was added. The mixture
was extracted with methylene dichloride three times. The com-
bined organic layer was washed with H2O and brine, dried with
MgSO4 and filtered. The filtrate was concentrated under reduced
pressure to give 9 (4.5 g). It was used in the next substitution
without purification.
25
1
[a]D = +153 (C = 1.0, MeOH), ESI-MS m ⁄ z 343 (M++23), H NMR
(CDCl3, 400 MHz, dppm); 0.93 (s, 9H), 0.12 (s, 6H), 1.30 (s, 3H), 1.49
(s, 3H), 2.85 (dd, J = 6.0, J = 13.6, 1H), 3.12–3.17 (m, 1H), 3.64 (dd,
J = 1.6, J = 13.6, 1H).
(3R,4S,5R)-2-acetyloxy-3,4-dihydroxy-3,4-O-isopropy-
lidene-5-iodiomethyl-tetrahydrothiophene (7)
The stirred mixture of sulfoxide 4 (28.9 g, 90 mmol) and Ac2O
(50 mL) was heated at 100 ꢀC for 3 h. The reaction mixture was
concentrated in vacuo to get 5 (22 g, 60 mmol). The crude 5 was
dissolved in THF (500 mL), and then TBAFÆ3H2O (23 g, 72 mmol)
was added. The mixture was stirred at room temperature for 3 h.
After the solvent was removed under reduced pressure, the resi-
due was partitioned between ethyl acetate and H2O. The organic
layer was washed with H2O followed by brine. Then, the organic
layer was dried with MgSO4, filtered and concentrated to afford
2-Chloro-6-amino-9-((3R,4S,5S)-3,4-dihydroxy-3,4-O-
isopropylidene-5-methyl-tetrahydrothiophene-2-yl)pur-
ine (10)
Ammonia was bubbled through a solution of 9 (2.0 g, 5.6 mmol) in
ethanol (20 mL) at 60 ꢀC until TLC showed 9 disappeared. The
reaction mixture was concentrated under reduced pressure to give
crude product 10 (2.2 g). It was used in the next deprotection with-
out purification.
6. The crude
6 (12 g, 50 mmol), triphenylphosphine (32 g,
125 mmol), iodine (25 g, 100 mmol) and imidazole (10.2 g,
150 mmol) were stirred in toluene (500 mL) under reflux for
20 min. When the reaction mixture was cooled, saturated aqueous
solution of NaHCO3 was added, and the mixture was stirred for
5 min. Excess of iodine was removed by the addition of Na2S2O3.
The organic layer was washed with water, dried with MgSO4, fil-
tered and concentrated. Triphenylphosphine oxide was precipitated
in diethyl ether, and the filtrate was concentrated and then sub-
jected to column chromatography to give 7 as yellow oil (13 g,
yield 42% for three steps).
2-Chloro-6-methylamino-9-((3R,4S,5S)-3,4-dihydroxy-3,
4-O-isopropylidene-5-methyl-tetrahydrothiophene-2-yl)
purine (11)
9 (2.0 g, 5.6 mmol) and 30% aqueous methylamine (1 mL) was stir-
red in THF at room temperature until TLC showed compound 9 dis-
appeared. The reaction mixture was concentrated under reduced
pressure to give crude product 11 (2.5 g).
Compounds 12–20 were obtained by the same procedure.
Chem Biol Drug Des 2010; 75: 619–627
620

Antitumor Activity of Nucleosides
1-[(3R,4S,5S)-3,4-dihydroxy-3,4-O-isopropylidene-5-
methyl-tetrahydrothiophene-2-yl]-5-fluorouracil (32)
(DMSO-d6, 400 MHz, dppm); 1.48 (d, J = 7.2, 3H), 2.94 (s, 3H),
3.29–3.38 (m, 1H), 3.97 (d, J = 3.2, 1H), 4.62 (d, J = 3.6, 1H), 5.30
(d, J = 4.4, 1H), 5.56 (d, J = 5.6, 1H), 5.78 (d, J = 5.6, 1H), 8.20 (b,
1H), 8.40 (s, 1H).
A mixture of 5-fluorouracil (500 mg, 3.8 mmol) and Hexamethyl disi-
lazane (3 mL) was stirred at 120 ꢀC for 5 h. The mixture was con-
centrated under reduced pressure to afford protected 5-fluorouracil
as white solid. Dried acetonitrile (5 mL), compound 8 (357 mg,
1.5 mmol) in dried acetonitrile (5 mL) were added at room tempera-
ture. Then the TMSOTf (0.70 mL. 3.8 mmol) was dropped at 0 ꢀC.
The reaction mixture was stirred at room temperature for 30 min,
and then saturated aqueous solution of NaHCO3 was added. The
mixture was extracted with methylene dichloride three times. The
combined organic layer was washed with H2O and brine, dried with
MgSO4 and filtered. The filtrate was concentrated under reduced
pressure to give crude product 32. It was used in the next depro-
tection without further purification.
2-Chloro-6-ethylamino-9-((2R,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (23a)
Yield = 56% (based on compound 8), white solid, mp 180 ꢀC (dec.),
[a]D = )108 (C = 1.0, MeOH), ESI-MS m ⁄ z 330 (M++1), 1H NMR
25
(DMSO-d6, 400 MHz, dppm); 1.19 (d, J = 7.2, 3H), 1.12–1.16
(m, 3H), 3.38–3.44 (m, 2H), 3.90–3.95 (m, 1H), 3.99 (s, 1H), 4.76
(d, J = 7.6, 1H), 5.39 (b, 1H), 5.54 (b, 1H), 5.84 (d, J = 8.8, 1H), 8.31
(b, 1H), 8.45 (s, 1H).
Crystal data: C12H16O2N5Cl1S1, triclinic system, space group P1, unit
cell dimensions a = 8.7829(4), b = 9.2313(4), c = 9.8800(5) ꢀ,
a = 94.110(3), b = 99.528(3), c = 109.544(2)ꢀ, V = 737.49(6) ꢀ3, Z =
1, 1955 reflections were measured, and 1942 independent reflec-
tions were used for the structure solution. Final R values are as
follows: wR2 = 0.0903, R1 = 0.0341, S = 1.056.
Compounds 33 and 34 were obtained by the same procedure.
2-Chloro-6-amino-9-((2R,3R,4S,5S)-3,4-dihydroxy-
5-methyl-tetrahydrothiophene-2-yl)purine (21a) and
2-Chloro-6-amino-9-((2S,3R,4S,5S)-3,4-dihydroxy-
5-methyl-tetrahydrothiophene-2-yl)purine (21b)
2-Chloro-6-ethylamino-9-((2S,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (23b)
A solution of 10 (2.5 g, 7.3 mmol) in 85% aqueous formic acid
(3 mL) was stirred at room temperature for 12 h. The mixture was
concentrated under reduced pressure, and the residue was purified
by silica gel chromatography to give 21a and 21b.
Yield = 18% (based on compound 8), white solid, mp 197 ꢀC (dec.),
[a]D = )15 (C = 1.0, MeOH), ESI-MS m ⁄ z 330 (M++1), 1H NMR
25
(DMSO-d6, 400 MHz, dppm); 1.46 (d, J = 7.2, 3H), 1.13 (t, J = 7.2,
J = 7.6, 3H), 3.41 (t, J = 7.2, J = 6.4, 2H), 1.55–1.61 (m, 2H), 3.30–3.33
(m, 1H), 3.93 (d, J = 3.6, 1H), 4.55 (d, J = 3.2, 1H), 5.34 (d, J = 4.8, 1H),
5.59 (d, J = 6.0, 1H), 5.75 (d, J = 6.0, 1H), 8.34 (b, 1H), 8.42 (s, 1H).
21a: Yield = 47% (based on compound 8), white solid, mp 167 ꢀC
(dec.), [a]D = )113 (C = 1.0, DMSO), ESI-MS m ⁄ z 302 (M++1), H
NMR (CD3OD, 400 MHz, dppm); 1.35 (d, J = 7.2, 3H), 4.09–4.11 (m,
1H), 4.17–4.19 (m, 1H), 4.83–4.86 (m, 1H), 6.06 (d, J = 8.0, 1H),
8.46 (s, 1H).
25
1
2-Chloro-6-propylamino-9-((2R,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (24a)
21b: Yield = 15% (based on compound 8), white solid, mp 87 ꢀC
Yield = 59% (based on compound 8), white solid, mp 173 ꢀC (dec.),
25
25
(dec.), [a]D = )41 (C = 1.0, DMSO), ESI-MS m ⁄ z 302 (M++1), 1H
[a]D = )93 (C = 1.0, DMSO), ESI-MS m ⁄ z 342 (M+-1), 1H NMR
NMR (CD3OD, 400 MHz, dppm); 1.55 (d, J = 7.2, 3H), 3.48–3.51 (m,
1H), 3.98–4.00 (m, 1H), 4.56–4.58 (m, 1H), 5.85 (d, J = 4.0, 1H),
8.45 (s, 1H).
(DMSO-d6, 400 MHz, dppm); 1.22 (d, J = 6.8, 3H), 0.87–0.92
(m, 2H), 1.57–1.64 (m, 2H), 3.37 (b, 2H), 3.94–3.97 (m, 1H), 4.03
(d, J = 2.8, 1H), 4.78 (b, 1H), 5.30 (b, 1H), 5.46 (d, J = 6.0, 1H), 5.88
(d, J = 8.4, 1H), 8.22 (b, 1H), 8.44 (s, 1H).
The other target compounds 22–31 (a and b) and 35–37 (a and
b) were obtained by the same procedure.
2-Chloro-6-propylamino-9-((2S,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (24b)
2-Chloro-6-methylamino-9-((2R,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (22a)
Yield = 20% (based on compound 8), white solid, mp 166 ꢀC (dec.),
25
[a]D = )31 (C = 1.0, DMSO), ESI-MS m ⁄ z 342 (M+-1), 1H NMR
Yield = 60% (based on compound 8), white solid, mp 140 ꢀC (dec.),
(DMSO-d6, 400 MHz, dppm); 1.46 (d, J = 7.2, 3H), 0.87 (t, J = 7.2,
J = 7.6, 3H), 1.55–1.61 (m, 2H), 3.33–3.36 (m, 2H), 3.29–3.33 (m,
1H), 3.39–3.96 (m, 1H), 4.59 (d, J = 3.2, 1H), 5.23 (d, J = 4.8, 1H),
5.55 (d, J = 5.6, 1H), 5.75 (d, J = 5.6, 1H), 8.23 (b, 1H), 8.38 (s, 1H).
25
1
[a]D = )85 (C = 1.0, DMSO), ESI-MS m ⁄ z 338 (M++23), H NMR
(DMSO-d6, 400 MHz, dppm); 1.22 (d, J = 6.8, 3H), 2.93 (s, 3H), 3.95–
3.97 (m, 1H), 4.04 (d, J = 3.6, 1H), 4.79 (d, J = 6.0, 1H), 5.37 (d, J = 4.8,
1H), 5.51 (d, J = 6.8, 1H), 5.88 (d, J = 8.4, 1H), 8.20 (b, 1H), 8.43 (s, 1H).
2-Chloro-6-cyclopropylamino-9-((2R,3R,4S,5S)-3,4-
dihydroxy-5-methyl-tetrahydrothiophene-2-yl)purine
(25a)
2-Chloro-6-methylamino-9-((2S,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (22b)
Yield = 21% (based on compound 8), white solid, mp 200 ꢀC (dec.),
Yield = 60% (based on compound 8), white solid, mp 168 ꢀC (dec.),
25
1
25
1
[a]D = )23 (C = 1.0, DMSO), ESI-MS m ⁄ z 338 (M++23), H NMR
[a]D = )89.5 (C = 1.0, MeOH), ESI-MS m ⁄ z 342 (M++1), H NMR
Chem Biol Drug Des 2010; 75: 619–627
621

Cong et al.
(CD3OD, 400 MHz, dppm); 1.32 (d, J = 6.8, 3H), 0.61–0.66 (m, 2H),
0.84–0.90 (m, 2H), 3.04 (b, 1H), 4.03–4.09 (m, 1H), 4.14–4.16 (m,
1H), 4.79–4.82 (m, 1H), 6.02 (d, J = 8.0, 1H), 8.27 (s, 1H).
2-Chloro-6-benzylamino-9-((2R,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (28a)
Yield = 50% (based on compound 8), white solid, mp 163 ꢀC (dec.),
25
1
2-Chloro-6-cyclopropylamino-9-((2S,3R,4S,5S)-3,4-di-
hydroxy-5-methyl-tetrahydrothiophene-2-yl)purine
(25b)
[a]D = )103 (C = 1.0, DMSO), ESI-MS m ⁄ z 414 (M++23), H NMR
(DMSO-d6, 400 MHz, dppm); 1.22 (d, J = 5.6, 3H), 7.21–7.35 (m,
5H), 4.65 (d, J = 1.6, 2H), 3.95–3.97 (m, 1H), 4.02–4.04 (m, 1H),
4.78 (b, 1H), 5.29 (d, J = 4.4, 1H), 5.76 (d, J = 6.8, 1H), 5.89 (d,
J = 8.4, 1H), 8.76 (b, 1H), 8.47 (s, 1H).
Yield = 18% (based on compound 8), white solid, mp 168 ꢀC
(dec.), [a]D = )23 (C = 1.0, MeOH), ESI-MS m ⁄ z 342 (M++1), 1H
25
NMR (CD3OD, 400 MHz, dppm); 1.57 (d, J = 6.8, 3H), 0.64–0.68
(m, 2H), 0.87–0.92 (m, 2H), 3.04 (b, 1H), 3.49–3.55 (m, 1H),
4.00–4.02 (m, 1H), 4.57–4.58 (m, 1H), 5.86 (d, J = 4.0, 1H), 8.33
(s, 1H).
2-Chloro-6-benzylamino-9-((2S,3R,4S,5S)-3,4-dihydr-
oxy-5-methyl-tetrahydrothiophene-2-yl)purine (28b)
Yield = 17% (based on compound 8), white solid, mp 166 ꢀC (dec.),
[a]D = )56 (C = 1.0, DMSO), ESI-MS m ⁄ z 414 (M++23), 1H NMR
25
2-Chloro-6-cyclopentylamino-9-((2R,3R,4S,5S)-3,4-di-
hydroxy-5-methyl-tetrahydrothiophene-2-yl)purine
(26a)
(DMSO-d6, 400 MHz, dppm); 1.48 (d, J = 7.2, 3H), 7.21–7.35 (m,
5H), 4.65 (b, 2H), 3.95–3.99 (m, 1H), 5.26 (d, J = 4.8, 1H), 5.53 (d,
J = 6.0, 1H), 5.79 (d, J = 6.0, 1H), 8.80 (b, 1H), 8.43 (s, 1H).
Yield = 42% (based on compound 8), white solid, mp 143 ꢀC (dec.),
2-Chloro-6-(a-methylbenzylamino-yl)-9-((2R,3R,4S,5S)
-3,4-dihydroxy-5-methyl-tetrahydrothiophene-2-yl)
purine (29a)
25
[a]D = )89 (C = 1.0, MeOH), ESI-MS m ⁄ z 368 (M+-1), 1H NMR
(DMSO-d6, 400 MHz, dppm); 1.21 (d, J = 6.8, 3H), 1.55–1.94 (m,
8H), 4.42 (b, 1H), 3.94–3.98 (m, 1H), 4.03 (d, J = 3.2, 1H), 4.76 (b,
1H), 5.29 (d, J = 4.4, 1H), 5.45 (d, J = 6.8, 1H), 5.88 (d, J = 8.0,
1H), 8.13 (d, J = 8.0, 1H), 8.43 (s, 1H).
Yield = 45% (based on compound 8), white solid, mp 162 ꢀC (dec.),
25
[a]D = )53 (C = 1.0, DMSO), ESI-MS m ⁄ z 404 (M++1), 1H NMR
(DMSO-d6, 400 MHz, dppm); 1.22 (d, J = 6.8, 3H), 7.18–7.45 (m,
5H), 5.29 (b, 1H), 1.54 (d, J = 6.8, 3H), 3.93–3.96 (m, 1H), 4.03 (b,
1H), 4.77 (b, 1H), 5.29 (d, J = 3.2, 1H), 5.45 (d, J = 6.4, 1H), 5.88 (d,
J = 8.8, 1H), 8.68 (d, J = 8.0 1H), 8.47 (s, 1H).
2-Chloro-6-cyclopentylamino-9-((2S,3R,4S,5S)-3,4-di-
hydroxy-5-methyl-tetrahydrothiophene-2-yl)purine
(26b)
Yield = 13% (based on compound 8), white solid, mp 158 ꢀC (dec.),
2-Chloro-6-(a-methylbenzylamino-yl)-9-((2S,3R,4S,5S)
-3,4-dihydroxy-5-methyl-tetrahydrothiophene-2-yl)
purine (29b)
25
1
[a]D = )26.5 (C = 1.0, MeOH), ESI-MS m ⁄ z 368 (M+-1), H NMR
(DMSO-d6, 400 MHz, dppm); 1.48 (d, J = 6.8, 3H), 1.54–1.93 (m,
8H), 4.42 (b, 1H), 3.30–3.36 (m, 1H), 3.96 (d, J = 3.2, 1H), 4.61 (d,
J = 3.6, 1H), 5.27 (d, J = 5.2, 1H), 5.53 (d, J = 5.2, 1H), 5.77 (d,
J = 5.6, 1H), 8.18 (d, J = 7.6, 1H), 8.40 (s, 1H).
Yield = 14% (based on compound 8), white solid, mp 150 ꢀC (dec.),
[a]D = )20 (C = 1.0, DMSO), ESI-MS m ⁄ z 404 (M++1), 1H NMR
25
(DMSO-d6, 400 MHz, dppm); 1.48 (d, J = 6.4, 3H), 7.20–7.45 (m,
5H), 5.42 (b, 1H), 1.54 (d, J = 5.6, 3H), 3.96 (b, 1H), 4.62 (b, 1H),
5.24 (b, 1H), 5.51 (d, J = 4.8, 1H), 5.77 (d, J = 4.8, 1H), 8.71 (d,
J = 7.2 1H), 8.43 (s, 1H).
2-Chloro-6-cyclohexylamino-9-((2R,3R,4S,5S)-3,4-di-
hydroxy-5-methyl-tetrahydrothiophene-2-yl)purine
(27a)
Yield = 57% (based on compound 8), white solid, mp 127 ꢀC (dec.),
2-Chloro-6-(2-fluorobenzylamino-yl)-9-((2R,3R,4S,5S)-
3,4-dihydroxy-5-methyl-tetrahydrothiophene-2-yl)pur-
ine (30a)
[a]D = )85.6 (C = 1.0, MeOH), ESI-MS m ⁄ z 384 (M++1), H NMR
(DMSO-d6, 400 MHz, dppm); 1.22 (d, J = 6.8, 3H), 1.22–1.87 (m,
10H), 3.95–3.99 (m, 1H), 4.03 (b, 1H), 4.78 (b, 1H), 5.28 (d, J = 2.8,
1H), 5.45 (b, 1H), 5.88 (d, J = 8.8, 1H), 8.02 (d, J = 7.6, 1H), 8.44 (s,
1H).
25
1
Yield = 51% (based on compound 8), white solid, mp 164 ꢀC (dec.),
25
[a]D = )91 (C = 1.0, DMSO), ESI-MS m ⁄ z 408 (M++1), 1H NMR
(DMSO-d6, 400 MHz, dppm); 1.22 (d, J = 6.8, 3H), 7.11–7.37 (m,
4H), 4.71 (b, 2H), 3.94–3.97 (m, 1H), 4.02–4.04 (m, 1H), 4.77 (b, 1H),
5.28 (d, J = 4.4, 1H), 5.44 (d, J = 6.8, 1H), 5.89 (d, J = 8.8, 1H),
5.89 (d, J = 8.8, 1H), 8.78 (b, 1H), 8.46 (b, 1H)., 8.48 (s, 1H).
2-Chloro-6-cyclohexylamino-9-((2S,3R,4S,5S)-3,4-di-
hydroxy-5-methyl-tetrahydrothiophene-2-yl)purine
(27b)
Yield = 22% (based on compound 8), white solid, mp 125 ꢀC (dec.),
2-Chloro-6-(2-fluorobenzylamino-yl)-9-((2S,3R,4S,5S)-
3,4-dihydroxy-5-methyl-tetrahydrothiophene-2-yl)pur-
ine (30b)
25
[a]D = )24 (C = 1.0, MeOH), ESI-MS m ⁄ z 406 (M++23), 1H NMR
(DMSO-d6, 400 MHz, dppm); 1.48 (d, J = 6.8, 3H), 1.23–1.68 (m,
10H), 3.14–3.20 (m, 1H), 4.03 (d, J = 3.6, 1H), 4.60 (b, 1H), 5.26 (d,
J = 5.2, 1H), 5.52 (d, J = 5.2, 1H), 5.77 (d, J = 5.6, 1H), 8.04 (d,
J = 8.4, 1H), 8.39 (s, 1H).
Yield = 16% (based on compound 8), white solid, mp 162 ꢀC (dec.),
25
[a]D = )55 (C = 1.0, DMSO), ESI-MS m ⁄ z 408 (M++1), 1H NMR
622
Chem Biol Drug Des 2010; 75: 619–627

Antitumor Activity of Nucleosides
(DMSO-d6, 400 MHz, dppm); 1.49 (d, J = 7.2, 3H), 7.13–7.39
(m, 4H), 4.71 (b, 2H), 3.33–3.39 (m, 1H), 3.98 (d, J = 3.6, 1H), 4.64
(d, J = 3.2, 1H), 5.27 (d, J = 5.2, 1H), 5.54 (d, J = 6.0, 1H), 5.80
(d, J = 5.2 1H), 8.79 (b, 1H), 8.46 (s, 1H).
(CD3OD, 400 MHz, dppm); 1.50 (d, J = 7.2, 3H), 3.37–3.42 (m, 1H),
3.85–3.87 (m, 1H), 4.23–4.25 (m, 1H), 5.80 (d, J = 8.4, 1H), 6.22 (d,
J = 5.6, 1H), 8.02 (d, J = 8.4, 1H).
1-[(2R,3R,4S,5S)-3,4-dihydroxy-5-methyl-tetrahydro-
thiophene-2-yl]-thymine (37a)
2-Chloro-6-(4-fluorobenzylamino-yl)-9-((2R,3R,4S,5S)-
3,4-dihydroxy-5-methyl-tetrahydrothiophene-2-yl)pur-
ine (31a)
Yield = 55% (based on compound 8), white solid, mp 158 ꢀC (dec.),
[a]D = )150 (C = 1.0, MeOH), ESI-MS m ⁄ z 259 (M++1), 1H NMR
25
Yield = 51% (based on compound 8), white solid, mp 173 ꢀC (dec.),
(CD3OD, 400 MHz, dppm); 1.30 (d, J = 6.4, 3H), 1.95 (s, 3H), 3.93–
3.96 (m, 1H), 4.07–4.08 (m, 1H), 4.32–4.35 (m, 1H), 6.25 (d, J = 9.2,
1H), 7.83 (s, 1H).
25
[a]D = )89 (C = 1.0, DMSO), ESI-MS m ⁄ z 408 (M++1), 1H NMR
(DMSO-d6, 400 MHz, dppm); 1.22 (d, J = 6.4, 3H), 7.10–7.14 (m,
2H), 7.36–7.39 (m, 2H), 4.62 (b, 2H), 3.94–3.97 (m, 1H), 4.03 (b, 1H),
4.77 (b, 1H), 5.30 (d, J = 3.2, 1H), 5.46 (b, 1H), 5.88 (d, J = 8.8, 1H),
8.77 (b, 1H), 8.47 (s, 1H).
1-[(2S,3R,4S,5S)-3,4-dihydroxy-5-methyl-tetrahydro-
thiophene-2-yl]-thymine (37b)
2-Chloro-6-(4-fluorobenzylamino-yl)-9-((2S,3R,4S,5S)-
3,4-dihydroxy-5-methyl-tetrahydrothiophene-2-yl)pur-
ine (31b)
Yield = 19% (based on compound 8), white solid, mp 167 ꢀC (dec.),
25
[a]D = )55 (C = 1.0, MeOH), ESI-MS m ⁄ z 259 (M++1), 1H NMR
(CD3OD, 400 MHz, dppm); 1.44 (d, J = 7.2, 3H), 1.87 (s, 3H), 3.30–
3.34 (m, 1H), 3.82–3.84 (m, 1H), 4.20–4.23 (m, 1H), 6.00 (d, J = 6.0,
1H), 7.69 (s, 1H).
Yield = 18% (based on compound 8), white solid, mp 154 ꢀC (dec.),
25
[a]D = )50 (C = 1.0, DMSO), ESI-MS m ⁄ z 408 (M++1), 1H NMR
(DMSO-d6, 400 MHz, dppm); 1.48 (d, J = 7.6, 3H), 7.10–7.17 (m,
2H), 7.36–7.40 (m, 2H), 4.60–4.64 (m, 1H), 5.25 (d, J = 4.8, 1H),
5.51 (d, J = 5.6, 1H), 5.78 (d, J = 5.2, 1H), 8.80 (b, 1H), 8.43 (s, 1H).
Biological assay
Cell viability was measured by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenylte-
trazolium bromide (MTT) assay to assess the chemosensitivity of tumor
cells. The growth inhibitory activity of target compounds was deter-
mined on cell lines LOVO (human carcinoma of colon), MDA-MB-435
(human breast cancer cell line) and CEM (human leukemia cell line)
using a modified version of the microculture tetrazolium assay. All the
cell lines tested were cultured in our pharmacology department.
1-[(2R,3R,4S,5S)-3,4-dihydroxy-5-methyl-tetrahydro-
thiophene-2-yl]-5-fluorouracil (35a)
Yield = 56% (based on compound 8), white solid, mp 196 ꢀC (dec.),
[a]D = )112 (C = 1.0, MeOH), ESI-MS m ⁄ z 285 (M++23), H NMR
(CD3OD, 400 MHz, dppm); 1.30 (d, J = 6.4, 3H), 3.93–3.95 (m, 1H),
4.06–4.08 (m, 1H), 4.27–4.30 (m, 1H), 6.24 (d, J = 8.4, 1H), 8.23 (d,
J = 6.8, 1H).
25
1
Once the cells reached 90% confluency, a cell suspension was
prepared by trypsinization of monolayer cultures. Cell counts were
performed, and the suspensions were diluted accordingly to give
4–5 · 104 cells ⁄ mL with the appropriate medium. Aliquots (10 lL)
of the cell suspension were added to each well in a 96-well microti-
tre plate. The cells were incubated for 24 h (37 ꢀC, 5% CO2). Stock
solutions of the test compound were prepared in dimethyl sulfoxide
(DMSO), and serial dilutions were made with medium to over a 100-
fold concentration range. Not more than 1% DMSO (final concentra-
tion) was present in each well. The test sample was incubated with
the cells for 72 h. Wells without cells and those with cells in culture
medium ⁄ DMSO were examined in parallel. At the end of the incuba-
tion period, the medium was decanted and replaced with 20 lL
MTT solution (5 mg ⁄ mL). The cells were incubated for another 4 h,
after which the medium was removed from each well by pipetting.
DMSO (100 lL) was added to each well to lyse the cells. The OD
values were measured within 30 min at 570 nm on an elisa reader
(WellscanMK-2). IC50 values were determined from logarithmic plots
of the % absorbance versus concentration generated.
1-[(2S,3R,4S,5S)-3,4-dihydroxy-5-methyl-tetrahydro-
thiophene-2-yl]-5-fluorouracil (35b)
Yield = 15% (based on compound 8), white solid, mp 180 ꢀC (dec.),
25
[a]D = )28 (C = 1.0, MeOH), ESI-MS m ⁄ z 285 (M++23), 1H NMR
(CD3OD, 400 MHz, dppm); 1.51 (d, J = 7.2, 3H), 3.38–3.41 (m, 1H),
3.85–3.87 (m, 1H), 4.24–4.26 (m, 1H), 6.03 (d, J = 5.2, 1H), 8.13 (d,
J = 7.2, 1H).
1-[(2R,3R,4S,5S)-3,4-dihydroxy-5-methyl-tetrahydro-
thiophene-2-yl]-uracil (36a)
Yield = 47% (based on compound 8), white solid, mp 167 ꢀC (dec.),
25
[a]D = )152 (C = 1.0, MeOH), ESI-MS m ⁄ z 245 (M++1), 1H NMR
(CD3OD, 400 MHz, dppm); 1.30 (d, J = 6.4, 3H), 3.92–3.94 (m, 1H),
4.07–4.09 (m, 1H), 4.30–4.33 (m, 1H), 5.81 (d, J = 8.0, 1H), 6.24 (d,
J = 8.4, 1H), 8.02 (d, J = 8.0, 1H).
1-[(2S,3R,4S,5S)-3,4-dihydroxy-5-methyl-tetrahydro-
thiophene-2-yl]-uracil (36b)
Results and Discussion
Synthesis
Yield = 16% (based on compound 8), white solid, mp 154 ꢀC (dec.),
Our strategy to the desired 5¢-deoxy-4¢-thio-L-nucleosides is to syn-
thesize the glycosyl donor and then to condense the donor with the
25
[a]D = )47 (C = 1.0, MeOH), ESI-MS m ⁄ z 245 (M++1), 1H NMR
Chem Biol Drug Des 2010; 75: 619–627
623

Cong et al.
HO
S
OMs
TBDMSO
S
O
OH
a
OMs
d
TBDMSO
HO
O
O
O
O
OH OH
D-ribose
1
O
O
3
3′
2
b
O
S
S
S
OAc
OAc
d
c
HO
TBDMSO
O
O
TBDMSO
O
O
O
O
O
Scheme 1: Reagents
and
conditions: (a) Na2S, DMF, heat,
65%. (b) m-CPBA, CH2CL2, )78,
51%. (c) Ac2O, heat. (d) TBAF,
THF, r.t. (e) Ph3P, imidazole, I2,
toluene, heat, 42% (three stepts)
(f) Bu3SnH, azobisisobutyronitrile,
toluene. heat, 67%.
6
4
5
e
S
S
OAc
OAc
f
H3C
I
O
O
O
8
7
nucleosidic bases. The synthesis of the key L-glyxosyl donor 8
started from D-ribose, as shown in Scheme 1.
major problem was shown by the over-oxidation to give sulfone.
Even when the quantity of m-CPBA was decreased to 0.9 eq., the
reaction was not improved. We therefore decided to replace
m-CPBA with ozone (10,11). Ozonization of the thioether 3 at
)78 ꢀC gave the desired (2S,3S,4R)-2-tert-butyldimethylsilyloxy-
methyl-3,4-dihydroxy-3,4-O-isopropylidene-tetrahydrothiophene-1-oxide
(sulfoxide 4) in 78% yield, followed by heating the sulfoxide 4
with acetic anhydride afforded (3R,4S,5S)-2-acetyloxy-5-tert-butyl-
dimethylsilyloxymethyl-3,4-dihydroxy-3,4-O-isopropylidene-tetrahydro-
thiophene (5). Treatment of 5 with tetrabutylammonium fluoride
trihydrate gave (3R,4S,5S)-2-acetyloxy -3,4-dihydroxy-3,4-O-isopro-
pylidene-5-hydroxymethyl-tetrahydrothiophene (6). Iodination was
performed on the 5-position by heating 6 with imidazole, triphenyl-
phosphine and iodine in toluene produced (3R,4S,5R)-2-acetyloxy-
3,4-dihydroxy-3,4-O-isopropylidene-5-iodiomethyl-tetrahydrothiophene
(7), which was converted to the key intermediate, the glycosyl
donor, (3R,4S,5S)-2-acetyloxy-3,4-dihydroxy-3,4-O-isopropylidene-5-
methyl-tetrahydrothiophene (8) by treating with tributyltin hydride
and azobisisobutyronitrile (AIBN) in toluene at 100 ꢀC in 67% yield.
D-ribose 1 was converted to 5-O-tert-butyldimethylsilyl-1,4-O-dime-
syl-2,3-O-isopropylidene-D-ribitol (2) in four steps according to the
published procedures (6–8). The cyclization of 2 with sodium sulfide
smoothly proceeded upon heating in N, N-dimethylformamide (DMF)
to yield the (2S,3S,4R)-2-tert-butyldimethylsilyloxymethyl-3,4-dihydr-
oxy-3,4-O-isopropylidene-tetrahydrothiophene (thioether 3) in
a
yield of 65%. To verify the absolute configuration of the thioether
3, it was deprotected with tetrabutylammonium fluoride trihydrate
(TBAFÆ3H2O) to obtain (2S,3S,4R)-2-hydroxymethyl-3,4-dihydroxy-
3,4-O-isopropylidene-tetrahydrothiophene (3¢). Compound 3¢ was
crystallized from acetonitrile, and its X-ray crystallographic analysis
showed that the configuration is 2S, 3S, 4R. The three-dimensional
structure of 3¢ was illustrated in Figure 2.
To achieve the synthesis of the nucleosidic base, we chose to
synthesize an acetate derivative through the Pummerer reaction.
Initially,thioether 3 was treated with 1.0 eq. m-chloroperoxybenzoic
acid (m-CPBA) at )78 ꢀC as reported by Jeong et al. (9), but the
sulfoxide (4) was obtained only in a low yield (15%), while the
The gylcosyl donor 8 was condensed with 2,6-dichloropurine in the
presence of EtAlCl2 in acetonitrile at room temperature to give the
nucleoside derivative 9. 1H NMR of 9 showed two sets of peak.
For example, there were two singlets, respectively, at 8.20 ppm and
8.38 ppm corresponding to H(8) in purine. The ratio of the two sing-
lets was approximately 3:1. So, the nucleoside derivative 9 might
be the mixture of a-isomer and b-isomer with the approximate ratio
of 3:1. They were used in next substitution without separation. Sub-
stitution of N6-position of 9 with various amines gave the isomeric
N6-substituted purine nucleoside derivatives 10–20 (Scheme 2).
The treatment of 10–20 with 85% aqueous formic acid at room
temperature offered the deprotection products 21–31 (a and b),
and the separation of the isomers was achieved by column chroma-
tography on silica gel. The ratio of the isomers obtained was approx-
imately 3:1 as confirmed the by 1HMR analysis of compound 9.
Condensation of the acetate 8 with persilylated pyrimidine deriva-
tives under Vorbrꢁggen conditions with trimethylsilyl trifluorome-
thanesulfonate (TMSOTf) as a catalyst was depicted in Scheme 3.
Figure 2: X-ray crystal structure of 3¢.
624
Chem Biol Drug Des 2010; 75: 619–627

Antitumor Activity of Nucleosides
N
N
Cl
S
8
S
OAc
N
a
b
H3C
N
H3C
O
S
O
O
O
Cl
9
NHR
S
N
N
N
N
NHR
H3C
HO
N
N
N
c
N
N
Cl
OH
N
H3C
S
+
N
NHR
O
O
H3C
Cl
N
HO
OH
Cl
Scheme 2: (a)
2,6-dichloro-
purine, EtAlCl2, CH3CN, r.t. (b)
Appropriate amine, r.t. THF. (c)
85% aqueous HCOOH, r.t.
OTMS
R
N
H
N
O
TMSO
a
O
N
S
S
OAc
N
b
H3C
H3C
R
O
O
O
O
8
32 R = F
33 R = H
34 R = CH3
O
N
R
NH
S
O
S
H3C
N
R
+
OH OH
O
H3C
N
H
OH OH
O
35 R = F
36 R = H
37 R = CH3
35 R = F
36 R = H
37 R = CH3
Figure 3: X-ray crystal structure of 23a.
Scheme 3: (a) trimethylsilyl trifluoromethanesulfonate, CH3CN,
r.t. (b) 85% aqueous HCOOH, r.t.
The absolute configuration of 23a was determined on the basis of
X-ray crystallography (in Figure 3). It verified that the structure of
23a was 2-chloro-6-ethylamino-9-((2R,3R,4S,5S)-3,4-dihydroxy-5-
methyl-tetrahydrothiophene-2-yl)purine. The result of X-ray crystal-
The isomeric mixtures of 32–34 were formed in moderate yield.
After deprotection with 85% aqueous formic acid at room tempera-
ture, the isomers (35a–37a and 35b–37b) were separated by
chromatography on silica gel.
1
lography confirmed the postulate from H NMR.
Biological activity
Structure analysis
All the target compounds were tested for the inhibition of the
growth of LOVO human carcinoma of colon, CEM human leukemia
and MDA-MB-435 human breast adenocarcinoma cells in culture
with 5-FU as the reference (Table 1).
The structural assignment of the target compounds were made on
1
the basis of H NMR studies. The magnitude of the coupling con-
stant (J-value) provided the important structural information. In the
target compounds of our report, the J2¢3¢ = 8–9 Hz for H(2¢)
(d = 5.80–6.30 ppm, d) on the main products was larger than the
J2¢3¢ = 4–6 Hz of minor products (d = 5.70–6.00 ppm, d). Based on
the fact that trans-configuration gave larger vicinal coupling con-
stant than the cis-configuration, we postulated the main products
were the a-isomers (2¢R), and the minor products were the b-iso-
mers (2¢S).
From the data in Table 1, it was seen that purine nucleosides gave
moderate or strong inhibition on the growth of CEM cells. For exam-
ple, compounds 22 (a, IC50 = 43.01; b, IC50 = 40.32), 23 (a,
IC50 = 37.20; b, IC50 = 43.62), 24 (a, IC50 = 40.52; b, IC50 = 36.24)
and 25 (a, IC50 = 35.95) shown comparable activity to 5-FU
(IC50 = 103.4), 26–31 (a and b) shown greater inhibition than 5-FU.
Chem Biol Drug Des 2010; 75: 619–627
625

Cong et al.
Table 1: Inhibition against human tumor cells in vitro
H
O
N
N
N
NHR
O
S
S
N
N
N
H3C
HO
H3C
R
OH
OH OH
Cl
21–31
35–37
CEM
LOVO
MDA-MB-435
IC% at
IC% at
IC% at
Compound No.
R
IC50 (lM)
100 lg ⁄ mL
IC50 (lM)
100 lg ⁄ mL
IC50 (lM)
100 lg ⁄ mL
21b
21a
22b
22a
23b
23a
24b
24a
25b
25a
26b
26a
27b
27a
28b
28a
29b
29a
30b
30a
31b
31a
35b
35a
36b
36a
37b
37a
5-FU
H
H
14.8
153.8
40.32
43.01
43.62
37.20
36.24
40.52
10.94
35.95
0.4932
0.2336
0.1757
0.1875
2.711
2.685
10.72
12.10
7.506
7.384
7.873
8.85
98.22
69.81
88.10
93.99
91.95
96.94
96.60
90.70
99.21
86.41
93.86
99.76
98.32
99.64
98.92
99.88
89.54
89.98
96.44
98.66
98.89
98.66
14.57
20.16
23.33
32.99
37.36
31.33
60.68
183.3
>240
>240
>240
>240
ꢀ240
280.3
>240
71.70
>240
43.01
52.47
ꢀ240
190.47
110.7
43.84
>240
>240
>240
>240
>240
>240
>240
>240
>240
>240
>240
>240
>240
64.96
4.17
44.83
19.81
30.44
49.63
51.68
47.23
72.93
44.93
81.30
90.48
49.01
52.41
63.97
100
7.45
46.78
31.91
42.49
22.00
23.65
0
67.50
316.8
237.9
219.8
>240
70.17
50.58
53.53
58.21
48.92
52.26
52.19
51.89
78.73
53.41
72.86
71.16
88.17
87.43
79.89
84.92
92.17
38.69
77.53
87.10
82.59
89.64
18.58
16.97
14.34
28.61
26.49
19.67
77.17
Methyl
Methyl
Ethyl
Ethyl
Propyl
Propyl
Cyclopropyl
Cyclopropyl
Cyclopentyl
Cyclopentyl
Cyclohexyl
Cyclohexyl
Benzyl
254.4
247.9
257.4
51.58
231.4
76.69
89.19
41.30
47.94
52.30
44.07
35.95
>240
52.40
47.65
63.79
48.63
>240
>240
>240
>240
>240
Benzyl
a-Methylbenzyl
a-Methylbenzyl
2-Fluorobenzyl
2-Fluorobenzyl
4-Fluorobenzyl
4-Fluorobenzyl
F
F
H
H
>240
>240
>240
>240
>240
>240
103.4
0
0
0
0
Methyl
Methyl
–
0
>240
31.54
42.28
6-cyclopentylamino and 6-cyclohexylamino derivatives 26 (a,
IC50 = 0.2336; b, IC50 = 0.4932) and 27 (a, IC50 = 0.1875; b,
IC50 = 0.1757) shown the best inhibition to CEM human leukemia
cells in vitro. But they gave weaker inhibition to LOVO and MDA-MB-
435 than 5-FU. Pyrimidine nucleosides 35–37 (a and b) show no sig-
nificant effect on the growth of LOVO, CEM or MDA-MB-435 cells.
benzylamino. The alkylamino groups only gave moderate activity.
Generally, the epimers at position 2¢ (a or b form) had no signifi-
cant difference on the antitumor activity.
Conclusion
In summary, we have synthesized a series of novel 5¢-deoxy-4¢-thio-
L-nucleosides and verified the configuration of the target com-
pounds by MS, 1H-NMR and X-ray crystallography. The antitumor
activities of the target compounds were tested against the growth
of LOVO, CEM and MDA-MB-435 cells in vitro. It showed that
6-cyclopentylamino and 6-cyclohexylamino purine compounds 26
and 27, both in a-configuration and in b-form, shown strong inhibi-
tion to CEM.
Structure–activity relationship
The target compounds showed the better inhibition against CEM
cell than against LOVO or MDA-MB-435 in vitro. The inhibitory
activity of 5¢-deoxy-4¢-thio-L-purine nucleosides was better than that
of 5¢-deoxy-4¢-thio-L-pyrimidine nucleosides. Among the 5¢-deoxy-4¢-
thio-L-purine nucleosides, the best substituted group in position 6 in
purine was cycloalkylamino (e.g. compounds 26 or 27), followed by
626
Chem Biol Drug Des 2010; 75: 619–627

Antitumor Activity of Nucleosides
6. Kim D.K., Kim G., Kim Y.W. (1996) Preparation of optically
active 3-substituted piperidines via ring expansion: synthesis of
4-amino- and 4-fluoro-1,4,5-trideoxy-1,5-imino-D-ribitol and
1,5-dideoxy-l,5-imino-D-ribitol. J Chem Soc Perkin 1;803–808.
7. Schmidt R.R., Karg J., Guilliard W. (1977) Glycosylhydrazine, 2.
Aufbauende Synthese von Pyrazolnucleosiden ꢁber Ribosylhydr-
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Acknowledgments
The work was supported by ``New Drug Innovation 2009X09301-
007'' by Ministry of Science and Technology of China.
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