Synthesis and anti-tumor activities of novel [1,2,4]triazolo[1,5-a] pyrimidines

Article abstract of DOI:10.3390/12051136

A series of novel [1,2,4]triazolo[1,5-a]pyrimidine derivatives has been designed and synthesized in order to find novel anti-tumor compounds. The structures of all the compounds were confirmed by IR, ¹H-NMR, MS and elemental analysis. Their ant

Full text of DOI:10.3390/12051136

Molecules 2007, 12, 1136-1146
molecules
ISSN 1420-3049
© 2007 by MDPI
www.mdpi.org/molecules
Full Research Paper
Synthesis and Anti-tumor Activities of Novel
[1,2,4]triazolo[1,5-a]pyrimidines
Xiang-Lin Zhao †, Yan-Fang Zhao ‡, Shu-Chun Guo, Hai-Sheng Song, Ding Wang and Ping
Gong*
School of Pharmaceutical Engineering, Shenyang Pharmaceutical University, 103 Wenhua Road,
Shenhe District, 110016 Shenyang, Liaoning, P. R. China; Fax: (+86) 24-23986426/23882925,
Phone: (+86) 24-23986426; E-mail: † smoothdagger@126.com; ‡ yanfangzhao@126.com
* Author to whom correspondence should be addressed; E-mail: gongpinggp@126.com
Received: 8 May 2007 / Accepted: 24 May 2007 / Published: 25 May 2007
Abstract: A series of novel [1,2,4]triazolo[1,5-a]pyrimidine derivatives has been designed
and synthesized in order to find novel anti-tumor compounds. The structures of all the
compounds were confirmed by IR, 1H-NMR, MS and elemental analysis. Their anti-tumor
activities against cancer cell lines (HT-1080 and Bel-7402) were tested by the MTT
method in vitro. Among them, compound 19 displayed the best anti-tumor activity with
IC50 values of 12.3 µM and 6.1 µM against Bel-7402 and HT-1080 cell lines respectively.
Keywords: [1,2,4]triazolo[1,5-a]pyrimidines, Synthesis, antitumor activities
Introduction
Purine analogs are widely used against various diseases, particularly cancer. The clinical
application of 6-mercaptopurine [1] and thioguanine [2] in cancer treatment and the development of
potent purine based CDK inhibitors, such as Purvalanols [3, 4], Olomoucine [3] and Roscovitine [5],
together with other findings based on the purine scaffold, have largely inspired and directed parallel
developments in the chemistry and anti-tumor research of related heterocyclic analogs, including
pyrrolo-pyrimidines [6], pyrazolo-pyrimidines [7, 8], imidazo-pyridines [9], triazolo-pyrimidines [10],
pyrazolo-pyridazines [11] and imidazo-pyrazines [12].
[1,2,4]Triazolo[1,5-a]pyrimidines, a subtype of purine bioisosteric analogs, were also reported to
possess potential anti-tumor activities, especially those bearing functional groups at C-5, C-6 or C-7
positions [13-15]. However, substitutions at the C-2 position seemed to be less attractive. Recently,

Molecules 2007, 12
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some [1,2,4]triazolo[1,5-a]pyrimidines bearing functional group at C-2 and C-7 positions, were
disclosed for their good antiproliferative ability [16, 17], which greatly encouraged us to explore more
potent analogs of such structures. Therefore, we carried out a series of modifications upon the
[1,2,4]triazolo[1,5-a]pyrimidine scaffold by introducing functional groups into C-2 and C-7 positions.
Herein, we report the synthesis and in vitro anti-tumor evaluation of a new series of 7-anilino-5-
methyl-2-(3-((5-(substituted-aminomethyl)furan-2-yl)methylthio)propyl)[1,2,4]triazol[1,5-
a]pyrimidines (6-20).
Results and Discussion
Chemistry
The synthetic route for the target compounds is shown in Scheme 1.
Scheme 1. Synthesis of target compounds.
OH
OH
O
N
b
N
N
N
N
N
a
c
N
N
OH
OH
OAc
O
+
N
H
H2N
N
N
2'
2
1
R1
N
R1
Cl
N
HN
N
HN
N
OH
N
N
N
N
N
N
N
d
N
N
f
N
e
Cl
S
Cl
OH
O
N
N
R1NH2
3
2
5a: R1=phenyl
4a: R1=phenyl
5b: R1=3-fluorophenyl
4b: R1=3-fluorophenyl
4c: R1=4-fluoro-3-(trifluoro
methyl)phenyl
5c: R1=4-fluoro-3-(trifluoro
methyl)phenyl
6: R1= phenyl, R2R3N=morpholino
7: R1= phenyl, R2R3N=piperidin-1-yl
8: R1= phenyl, R2R3N=pyrrolidin-1-yl
9: R1= phenyl, R2R3N=4-methylpiperazin-1-yl
10: R1= 3-fluorophenyl, R2R3N=morpholino
R1
HN
N
11: R1= 3-fluorophenyl, R2R3N=piperidin-1-yl
R2
R3
g
12: R1= 3-fluorophenyl, R2R3N=pyrrolidin-1-yl
N
N
N
N
S
13: R1= 3-fluorophenyl, R2R3N=4-methylpiperazin-1-yl
14: R1= 3-fluorophenyl, R2R3N=dimethylamino
R2R3NH
O
15 R1= 4-fluoro-3-(trifluoromethyl)phenyl, R2R3N=morpholino
16: R1= 4-fluoro-3-(trifluoromethyl)phenyl, R2R3N=piperidin-1-yl
17: R1= 4-fluoro-3-(trifluoromethyl)phenyl, R2R3N=pyrrolidin-1-yl
18: R1= 4-fluoro-3-(trifluoromethyl)phenyl, R2R3N=4-methylpiperazin-1-yl
19: R1= 4-fluoro-3-(trifluoromethyl)phenyl, R2R3N=diethylamino
20: R1= 4-fluoro-3-(trifluoromethyl)phenyl, R2R3N=dimethylamino
Reagents and conditions: a) aminoguanidine, γ-butyrolactone, Py, reflux, 10 h; b) ethyl acetoacetate, acetic acid, reflux, 30
i
h, c) methanolic ammonia, room temperature, 24 h; d) POCl3, reflux, 3 h; e) R1NH2, PrOH, 50°C, 3 h; f) furan-2-
ylmethanethiol, NaH, DMF, 50°C, 0.5 h; g) HCHO, R2R3NH, AcOH, 50°C, 4 h.
Commercially available γ-butyrolactone was first transformed into the substituted [1,2,4]triazole 1
with aminoguanidine carbonate in the presence of pyridine in 40% yield. Following the procedure
reported by Okabe [18], condensation of 1 with ethyl acetoacetate was carried out in acetic acid at
reflux to give a mixture of 2 and 2′(the acetylated product of 2), which further treated with methanolic

Molecules 2007, 12
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ammonia to give 2 in 88% yield. Subsequent treatment of 2 with phosphorus oxychloride afforded 3 in
94% yield. Displacement of the chloro group of 3 with various anilines provided the intermediate 4a-
4c in 87-93% yields.
Etherification of 4a-4c with (furan-2-yl)methanethiol in the presence of sodium hydride provided
intermediate 5a-5c. Subsequent aminomethylation of 5a-5c with various secondary amines and
formaldehyde, followed by chromatographic purification gave the final products (6-20) in 45-80%
yields.
Anticancer activities
All target compounds (6-20) were evaluated for their anti-tumor activity in vitro by the MTT
method. The IC50 values against Bel-7402 (Human Liver Cancer Cell Lines) and HT-1080 (Human
Fibro Sarcoma Cell Lines) cell lines are summarized in Table 1.
Table 1. The anticancer activities of compounds 6-20.
IC50 (µM)
Bel-7402
IC50 (µM)
Bel-7402
Compd.
Compd.
HT-1080
>300
HT-1080
30.8
>300
72.5
67.3
>300
>300
45.8
77.6
>300
54.1
44.6
42.1
13.9
22.1
12.3
22.2
35.5
6
7
14
15
52.1
57.4
237
36.3
32.2
28.4
29.3
6.1
8
16
9
17
280
10
11
12
13
18
35.1
41.4
261.9
19
14.8
22.7
20
cisplatin
Table 1 shows that both substituents on the aniline ring and variations on 5"-furylmethyl of the
side-chain have substantial influence on the anti-tumor activity. 3-Fluoro-phenylamino substituted
series showed comparable or better anti-tumor activities than phenylamino substituted series
(comparison of 7 and 11, 8 and 12). Morever, introduction of 4-fluoro-3-(trifluoromethyl)phenylamino
group into the C-7 position was found to be quite favorable for increasing anti-tumor activity
(comparison of 8 and 17, 14 and 20). Among this series, compound 19 showed the best anti-tumor
activity with IC50 values of 12.3 µM and 6.1 µM against Bel-7402 and HT-1080 cell lines respectively,
which was 4 times more potent than cisplatin. On the other hand, among the hydrophilic groups
introduced into the 5"-furylmethyl of the side-chain at C-2 position of the scaffold, piperidinyl,
pyrrolidinyl, diethylamino and dimethylamino groups rendered better anti-tumor activity, while
morpholino, 4-methylpiperizinyl resulted in drastic decrease in anti-tumor potency (comparison of 6, 9
and 7, 8). However, despite the negative effects of morpholino and 4-methylpiperizinyl substitutions,
anti-tumor potency could be retained in the 4-fluoro-3-(trifluoromethyl)phenyl amino series as
illustrated by the comparison of 6 and 15, 13 and 18, suggesting that the contribution of 4-fluoro-3-

Molecules 2007, 12
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(trifluoromethyl)phenyl amino groups to the potency might partly compensate the negative effect of
morpholino and 4-methylpiperizinyl substitution.
Conclusions
In conclusion, a novel series of [1,2,4]triazolo[1,5-a]pyrimidines were synthesized and found to be
active against tumor cell lines. Compounds with disubstituents on the aniline ring and piperidinyl,
pyrrolidinyl, diethylamino or dimethylamino on the 5"-furylmethyl of the side-chain showed better
anti-tumor activities than other series. Among this series, compound 19 showed the best anti-tumor
activity and will be considered in a further study.
Experimental Section
General
Melting points were determined by the capillary tube method, and the thermometer was
1
uncorrected. Mass spectra were obtained on an Agilent 1100 HPLC-MS instrument. H-NMR spectra
were run in DMSO-d6, CDCl3, with TMS at the internal standard, on a Bruker ARX-300 instrument
operating at 300 MHz. IR spectra (KBr disks) were recorded on a Bruker IFS 55 instrument. Elemental
analysis was performed with a Carlo-Erba 1106 Elemental analysis instrument.
5-amino-3-(3-hydroxypropyl)-4H-[1,2,4]triazole (1)
A mixture of aminoguanidine (65 g, 0.55 mol), γ-butyrolactone (43 g, 0.50 mol) and pyridine (900
mL) was heated to reflux for 10 h. The resultant mixture was concentrated and filtered to give a pale
solid. The mass product was recrystallized from absolute ethanol (150 mL) to obtain 1 (28 g, 40%) as a
1
white solid, m. p.146-147 °C, (lit. 149-150 °C [19]). MS [MH+] (m/z): 142.1; H-NMR(DMSO-d6)δ:
1.71 (pentad, 2H, CH2), 2.44 (t, 2H, J = 6.0 Hz, CH2), 3.41 (t, 2H, J = 6.2 Hz, CH2), 4.45 (s, 1H, OH),
5.55 (br s, 2H, NH2), 11.51 (br s, 1H, NH); Anal. calcd. for C5H10N4O: C 42.24, H 7.09, N 39.41;
Found: C 42.01, H 7.01, N 39.23.
7-hydroxyl-2-(3-hydroxypropyl)-5-methyl-[1,2,4]triazolo[1,5-a]pyrimidine (2)
A mixture of 1 (28 g, 0.20 mol), ethyl acetoacetate (30.8 g, 0.24 mol) and acetic acid (300 mL) was
heated to reflux for 30 h and then concentrated, filtered to give a mixture of 2 and 2′ as a pale yellow
solid. Then the solid was added into the saturated ammonia methanol (300 mL) and stirred at room
temperature for 24 h. The mixture was evaporated and water was added, which was then acidified with
dilute hydrochloride solution to pH 5-6, filtered and recrystallized from ethanol to give the product 2
1
(37 g, 88%), m. p. 204-206 °C. MS [MH+] (m/z): 208.1; H-NMR(DMSO-d6) δ: 1.85 (pentad, 2H,
CH2), 2.30 (s, 1H, CH3), 2.72 (t, 2H, J = 6.0 Hz, CH2), 3.45 (t, 2H, J = 6.1 Hz, CH2), 4.49 (br s, 1H,
OH), 5.76 (s, 1H, C6-H), 12.98 (br s, 1H, C7-OH); Anal. calcd. for C9H12N4O2: C 51.92, H 5.81, N
26.91; Found: C 51.86, H 5.68, N 26.79.

Molecules 2007, 12
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7-chloro-2-(3-chloropropyl)-5-methyl-[1, 2, 4]triazolo[1,5-a]pyrimidine (3)
A mixture of 2 (37 g, 0.18 mol) and phosphorus oxychloride (265 g, 1.70 mol) was heated to reflux
for 3 h, and then was concentrated in vacuo to result a red oil, which was poured into water, and
extracted with chloroform (250 mL×3). The organic layer was then washed with water three times,
dried with anhydrous magnesium sulfate and concentrated to give 3 (41 g, 94%) as a yellow solid (LC
purity: 96%, MS [MH+] (m/z): 245.2), which was directly used in next step without purification.
General procedure for the synthesis of 7-anilino-5methyl-2-(3-chloroproyl)-[1,2,4]triazolo[1,5-
a]pyrimidine (4a-4c)
A mixture of R1NH2 (44 mmol), 3 (10 g, 40 mmol) and isopropanol (100 mL) was heated at 50°C
for 3 h. After cooling to room temperature and the following filtration, a yellow solid was obtained.
Recrystallization of the solid from methanol gave 4a-4c as slight yellow powder (87-93%).
General procedure for the synthesis of 7-anilino-5-methyl-2-(3-(furan-2-ylmethylthio)propyl)-
[1,2,4]triazolo [1,5-a]pyrimidine (5a-5c)
Furan-2-ylmethanethiol (8.6 g, 75 mmol) was added dropwise into a suspension of NaH (1.8 g 75
mmol) and dry DMF (80 mL) at room temperature. After the addition, the mixture was stirred for 5
min, and then 4a-4c (25 mmol) was added. The mixture was heated at 50°C for 30 min and was then
poured into water. The brown oil which separated from water was triturated with diethyl ether, then
filtered and recrystallized from ethyl acetate/cyclohexane to afford 5a-5c as gray solid (80%-85%).
General procedure for the synthesis of (6-20)
Formaldehyde (0.6 g, 8 mmol) was added into the solution of R2R3NH (13 mmol) in acetic acid (20
mL). The mixture was stirred at 30°C for 10 min, and then 5a-5c (5 mmol) was added. After stirring at
50°C for 4 h, the mixture was concentrated in vacuo. The residue was taken up in water (50 mL),
basified with concentrated aqueous sodium hydroxide to pH 9-10, extracted with methylene dichloride,
and then dried over magnesium sulfate. Evaporation of the solvent provided an oil residue, which was
purified by chromatography to give the final products 6-20.
7-phenylamino-5-methyl-2-(3-((5-(morpholinomethyl)furan-2-yl)methylthio)propyl)-[1,2,4]triazolo
[1,5-a]pyrimidine (6)
Yield: 60%; m.p. 167-168 °C; MS [MH+] (m/z): 479.2; IR (KBr) cm-1: 3445.5 (υNH), 2963.4 (υCH3),
1
1609.3, 1576.3 (υC=C, υC=N), 1479.8 (δCH2), 1328.5 (υC-N). H-NMR(DMSO-d6)δ: 2.02 (pentad, 2H,
CH2), 2.51 (s, 3H, CH3), 2.59 (m, 6H, 3×CH2), 2.88 (t, 2H, J = 7.2 Hz, CH2), 3.61 (br s, 4H, 2×CH2),
3.73 (s, 2H, CH2), 3.77 (s, 2H, CH2), 6.23 (d, 1H, J =2.6 Hz , furyl-H), 6.27 (d, 1H, J =2.7 Hz , furyl-
H), 6.34 (s, 1H, C6-H), 7.29~7.48 (m, 5H, Ph-5×H); Anal. calcad. for C25H30N6O2S : C 62.74, H 6.32,
N 17.56 Found: C 62.59 H 6.25, N 17.45.

Molecules 2007, 12
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7-phenylamino-5-methyl-2-(3-((5-(piperidin-1-ylmethyl)furan-2-yl)methylthio)propyl)-[1,2,4]triazolo
[1,5-a]pyrimidine (7)
Yield: 64%; m.p. 169-170°C; MS [MH+] (m/z): 477.1; IR (KBr) cm-1: 3447.2 (υNH), 2966.5 (υCH3),
1609.1, 1575.9 (υC=C, υC=N), 1478.0 (δCH2), 1329.6 (υC-N). 1H-NMR(CDCl3)δ: 1.42 (m, 2H, CH2), 1.64
(m, 4H, 2×CH2), 2.16 (pentad, 2H, CH2), 2.49 (br s, 4H, 2×CH2), 2.52 (s, 3H, CH3), 2.64 (t, 2H, J =
7.1 Hz, CH2), 2.99 (t, 2H, J = 7.3 Hz, CH2), 3.57 (s, 2H, CH2), 3.72 (s, 2H, CH2), 6.11 (d, 1H, J =3.0
Hz , furyl-H), 6.14 (d, 1H, J =3.0 Hz , furyl-H), 6.32 (s, 1H, C6-H), 7.35~7.50 (m, 5H, Ph-5×H); Anal.
calcad. for C26H32N6OS : C 65.52, H 6.77, N 17.63 Found: C 65.44, H 6.63, N 17.60.
7-phenylamino-5-methyl-2-(3-((5-(pyrrolidin-1-ylmethyl)furan-2-yl)methylthio)propyl)-[1,2,4]triazolo
[1,5-a]pyrimidine (8)
Yield: 64%; m.p. 162-164°C; MS [MH+] (m/z): 463.2; IR (KBr) cm-1: 3440.5 (υNH), 2965.2 (υCH3),
1
1611.5, 1576.1 (υC=C, υC=N), 1478.3 (δCH2), 1329.8 (υC-N). H-NMR(CDCl3)δ: 1.82 (m, 4H, 2×CH2),
2.15 (pentad, 2H, CH2), 2.52 (s, 3H, CH3), 2.64 (t, 2H, J = 7.2 Hz, CH2), 2.69 (br s, 4H, 2×CH2), 2.99
(t, 2H, J = 7.3 Hz, CH2), 3.71 (s, 2H, CH2), 3.72 (s, 2H, CH2), 6.11 (d, 1H, J =3.0 Hz , furyl-H), 6.17
(d, 1H, J =2.9 Hz, furyl-H), 6.32 (s, 1H, C6-H), 7.35~7.50 (m, 5H, Ph-5×H); Anal. calcad. for
C25H30N6OS : C 64.91, H 6.54, N 18.17 Found: C 64.79, H 6.43, N 18.11.
7-phenylamino-5-methyl-2-(3-((5-((4-methylpiperazin-1-yl)methyl)furan-2-yl)methylthio)propyl)-
[1,2,4] triazolo[1,5-a]pyrimidine (9)
Yield: 73%; m.p. 176-179°C; MS [MH+] (m/z): 492.2; IR (KBr) cm-1: 3442.6 (υNH), 2966.8 (υCH3),
1
1609.3, 1576.8 (υC=C, υC=N), 1479.2 (δCH2), 1329.0 (υC-N). H-NMR(DMSO-d6)δ: 2.01 (pentad, 2H,
CH2), 2.41 (s, 3H, CH3), 2.61 (t, 2H, J = 6.9 Hz, CH2), 2.74 (br s, 7H, CH3, 2×CH2), 3.17 (br s, 4H,
2×CH2), 3.72 (s, 2H, CH2), 3.77 (s, 2H, CH2), 6.22 (m, 2H, furyl-2×H), 6.36 (s, 1H, C6-H), 7.29~7.48
(m, 5H, Ph-5×H); Anal. calcad. for C26H33N7OS : C 63.52, H 6.77, N 19.94 Found: C 63.47, H 6.85, N
19.82.
7-(3-fluorophenylamino)-5-methyl-2-(3-((5-(morpholinomethyl)furan-2-yl)methylthio)propyl)-[1,2,4]
triazolo[1,5-a]pyrimidine (10)
Yield: 80%; m.p. 173-175°C; MS [MH+] (m/z): 497.1; IR (KBr) cm-1: 3442.8 (υNH), 2970.5 (υCH3),
1
1608.0, 1575.2 (υC=C, υC=N), 1478.5 (δCH2), 1329.4 (υC-N). H-NMR(DMSO-d6)δ: 2.03 (pentad, 2H,
CH2), 2.44 (s, 3H, CH3), 2.61 (t, 2H, J = 7.1 Hz, CH2), 2.70 (br s, 4H, 2×CH2), 2.90 (t, 2H, J = 7.3 Hz,
CH2), 3.66 (br s, 4H, 2×CH2), 3.79 (s, 2H, CH2), 3.83 (s, 2H, CH2), 6.26 (d, 1H, J =2.9 Hz , furyl-H),
6.33 (d, 1H, J =2.8 Hz , furyl-H), 6.52 (s, 1H, C6-H), 7.13 (dd, 1H, JFH=8.4 Hz, JHH=7.7 Hz, Ph-4H),
7.34 (br d, 1H, JFH=7.7 Hz, Ph-2H), 7.34 (br d, 1H, JHH=7.7 Hz, Ph-6H), 7.50 (dd, 1H, J1=7.8 Hz,
J2=7.7 Hz, Ph-5H); Anal. calcad. for C25H29FN6O2S : C 55.28, H 5.89, N 16.92 Found: C 55.21, H
5.78, N 16.79.

Molecules 2007, 12
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7-(3-fluorophenylamino)-5-methyl-2-(3-((5-(piperidin-1-ylmethyl)furan-2-yl)methylthio)propyl)-
[1,2,4] triazolo[1,5-a]pyrimidine (11)
Yield: 68%; m.p. 170-172 °C; MS [MH+] (m/z): 495.1; IR (KBr) cm-1: 3444.5 (υNH), 2963.5 (υCH3),
1609.5, 1576.5 (υC=C, υC=N), 1479.8 (δCH2), 1329.1 (υC-N). 1H-NMR(DMSO-d6)δ: 1.48 (br s, 2H, CH2),
1.71 (br s, 2H, 2×CH2), 2.02 (pentad, 2H, CH2), 2.45 (s, 3H, CH3), 2.63 (t, 2H, J = 7 Hz, CH2), 2.91 (t,
2H, J = 7.1 Hz, CH2), 3.03 (br s, 4H, 2×CH2), 3.82 (s, 2H, CH2), 4.26 (s, 2H, CH2), 6.35 (br s, 1H,
furyl-H), 6.53 (br s, 2H, C6-H, furyl-H), 7.14 (dd, 1H, JFH=8.5 Hz, JHH=7.7 Hz, Ph-4H), 7.34 (br d, 1H,
JFH=7.9 Hz, Ph-2H), 7.34 (br d, 1H, JHH=7.9 Hz, Ph-6H), 7.52 (dd, 1H, J1=7.7 Hz, J2=7.7 Hz, Ph-5H);
Anal. calcad. for C26H31FN6OS : C 63.13, H 6.32, N 16.99 Found: C 63.05, H 6.21, N 16.88.
7-(3-fluorophenylamino)-5-methyl-2-(3-((5-(pyrrolidin-1-ylmethyl)furan-2-yl)methylthio)propyl)-
[1,2,4] triazolo[1,5-a]pyrimidine (12)
Yield: 65%; m.p. 166-168 °C; MS [MH+] (m/z): 481.1; IR (KBr) cm-1: 3442.8 (υNH), 2968.5 (υCH3),
1
1610.2, 1574.2 (υC=C, υC=N), 1478.2 (δCH2), 1328.9 (υC-N). H-NMR(CDCl3)δ: 1.82 (m, 4H, 2×CH2),
2.14 (pentad, 2H, CH2), 2.55 (s, 3H, CH3), 2.64 (t, 2H, J = 7.2 Hz, CH2), 2.69 (br s, 4H, 2×CH2), 2.99
(t, 2H, J = 7.2 Hz, CH2), 3.71 (s, 4H, 2×CH2), 6.11 (br s, 1H, furyl-H), 6.16 (br s, 1H , furyl-H), 6.38
(s, 1H, C6-H), 7.03 (dd, 1H, JFH=8.1 Hz, JHH=7.0 Hz, Ph-4H), 7.14 (br d, 1H, JFH=7.1 Hz, Ph-2H), 7.14
(br d, 1H, JHH=7.1 Hz, Ph-6H), 7.45 (dd, 1H, J1=7.2 Hz, J2=7.0 Hz, Ph-5H); Anal. calcad. for
C25H29FN6OS : C 62.48, H 6.08, N 17.49 Found: C 62.38, H 5.99, N 17.41.
7-(3-fluorophenylamino)-5-methyl-2-(3-((5-((4-methylpiperazin-1-yl)methyl)furan-2-yl)methylthio)pro
pyl)-[1,2,4]triazolo[1,5-a]pyrimidine (13)
Yield: 77%; m.p. 179-180 °C; MS [MH+] (m/z): 510.2; IR (KBr) cm-1: 3445.1 (υNH), 2967.8 (υCH3),
1
1608.5, 1575.9 (υC=C, υC=N), 1479.5 (δCH2), 1329.1 (υC-N). H-NMR(DMSO-d6)δ: 2.02 (pentad, 2H,
CH2), 2.43 (s, 3H, CH3), 2.57~2.62 (m, 6H, 3×CH2), 2.73 (s, 3H, CH3), 2.89 (t, 2H, J = 6.9 Hz, CH2),
3.16 (br s, 4H, 2×CH2), 3.56 (s, 2H, CH2), 3.76 (s, 2H, CH2), 6.22 (m, 2H, furyl-2×H), 6.51 (s, 1H, C6-
H), 7.12 (dd, 1H, JFH=8.6 Hz, JHH=7.5 Hz, Ph-4H), 7.33 (br d, 1H, JFH=7.8 Hz, Ph-2H), 7.33 (br d, 1H,
JHH=7.8 Hz, Ph-6H), 7.51 (dd, 1H, J1=7.5 Hz, J2=7.5 Hz, Ph-5H); Anal. calcad. for C26H32FN7OS : C
61.27, H 6.33, N 19.24 Found: C 61.15, H 6.25, N 19.12.
7-(3-fluorophenylamino)-5-methyl-2-(3-((5-((dimethylamino)methyl)furan-2-yl)methylthio)propyl)-
[1,2,4]triazolo[1,5-a]pyrimidine (14)
Yield: 52%; m.p. 156-157 °C; MS [MH+] (m/z): 455.2; IR (KBr) cm-1: 3443.5 (υNH), 2964.5 (υCH3),
1
1608.2, 1575.4 (υC=C, υC=N), 1479.8 (δCH2), 1329.5 (υC-N). H-NMR(CDCl3)δ: 2.15 (pentad, 2H, CH2),
2.30 (s, 6H, 2×CH3), 2.55 (s, 3H, CH3), 2.64 (t, 2H, J = 7.1 Hz, CH2), 2.99 (t, 2H, J = 7.2 Hz, CH2),
3.51 (s, 2H, CH2), 3.72 (s, 2H, CH2), 6.13 (br s, 2H, furyl-2×H), 6.40 (s, 1H, C6-H), (dd, 1H, JFH=8.3
Hz, JHH=7.1 Hz, Ph-4H), 7.15 (br d, 1H, JFH=7.3 Hz, Ph-2H), 7.15 (br d, 1H, JHH=7.3 Hz, Ph-6H), 7.45
(dd, 1H, J1=7.2 Hz, J2=7.1 Hz, Ph-5H); Anal. calcad. for C23H27FN6OS : C 60.77, H 5.99, N 18.49
Found: C 60.69, H 5.78, N 18.38.

Molecules 2007, 12
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7-(4-fluoro-3-(trifluoromethyl)phenylamino)-5-methyl-2-(3-((5-(morpholinomethyl)furan-2-yl)methyl
lthio)propyl)-[1,2,4]triazolo[1,5-a]pyrimidine (15)
Yield: 75%; m.p. 190-192 °C; MS [MH+] (m/z): 565.1; IR (KBr) cm-1: 3446.5 (υNH), 2870.3 (υCH3),
1622.3, 1581.2, 1505.3 (υC=C, υC=N), 1478.1 (δCH2), 1323.5 (υC-N). 1H-NMR(DMSO-d6)δ: 2.02 (pentad,
2H, CH2), 2.42 (s, 3H, CH3), 2.61 (t, 2H, J = 7.2 Hz, CH2), 2.75 (br s, 4H, 2×CH2), 2.90 (t, 2H, J = 7.1
Hz, CH2), 3.11 (br s, 4H, 2×CH2), 3.65 (br s, 2H, CH2), 3.79(br s, 2H, CH2), 6.26 (d, 1H, J =3.0 Hz,
furyl-H), 6.33 (d, 1H, J =2.9 Hz, furyl-H), 6.41 (s, 1H, C6-H), 7.64 (dd, 1H, JFH=10.0 Hz, JHH=9.5 Hz,
Ph-5H), 7.83 (m, 2H, Ph-2, 6-2×H); Anal. calcad. for C26H28F4N6O2S : C 55.31, H 5.00, N 14.88
Found: C 55.19, H 4.79, N 14.72.
7-(4-fluoro-3-(trifluoromethyl)phenylamino)-5-methyl-2-(3-((5-(piperidin-1-ylmethyl)furan-2-yl)meth
ylthio)propyl)-[1,2,4]triazolo[1,5-a]pyrimidine (16)
Yield: 54%; m.p. 182-183 °C; MS [MH+] (m/z): 563.1; IR (KBr) cm-1: 3443.6 (υNH), 2953.7 (υCH3),
1620.4, 1575.8, 1505.3 (υC=C, υC=N), 1476.7 (δCH2), 1323.0 (υC-N). 1H-NMR(DMSO-d6)δ: 1.44 (m, 2H,
CH2), 1.66 (m, 4H, 2×CH2), 2.02 (pentad, 2H, CH2), 2.41 (s, 3H, CH3), 2.61 (t, 2H, J = 6.9 Hz, CH2),
2.89 (t, 2H, J = 7.1 Hz, CH2), 3.00 (br s, 4H, 2×CH2), 3.79 (s, 2H, CH2), 4.09 (s, 2H, CH2), 6.30 (br s,
1H, furyl-H), 6.40 (s, 1H, C6-H), 6.47 (br s, 1H, furyl-H), 7.63 (dd, 1H, JFH=9.9 Hz, JHH=9.0 Hz, Ph-
5H), 7.79~7.83 (m, 2H, Ph-2, 6-2×H); Anal. calcad. for C27H30F4N6OS : C 57.64, H 5.37, N 14.94
Found: C 57.49, H 5.15, N 14.87.
7-(4-fluoro-3-(trifluoromethyl)phenylamino)-5-methyl-2-(3-((5-(pyrrolidin-1-ylmethyl)furan-2-yl)
methylthio)propyl)-[1,2,4]triazolo[1,5-a]pyrimidine (17)
Yield: 54%; m.p. 179-180 °C; MS [MH+] (m/z): 549.1; IR (KBr) cm-1: 3445.1 (υNH), 2953.7 (υCH3),
1
1620.4, 1505.3 (υC=C, υC=N), 1479.5 (δCH2), 1323.0 (υC-N). H-NMR(CDCl3)δ: 1.81 (m, 4H, 2×CH2),
2.13 (pentad, 2H, CH2), 2.53 (s, 3H, CH3), 2.62 (t, 2H, J = 7.2 Hz, CH2), 2.67 (br s, 4H, 2×CH2), 2.99
(t, 2H, J = 7.3 Hz, CH2), 3.71 (s, 2H, CH2), 3.72 (s, 2H, CH2), 6.10 (d, 1H, J =3.1 Hz , furyl-H), 6.14
(d, 1H, J =3.0 Hz , furyl-H), 6.16 (s, 1H, C6-H), 7.36 (dd, 1H, JFH=9.2 Hz, JHH=9.0 Hz, Ph-5H), 7.61
(m, 2H, Ph-2, 6-2×H); Anal. calcad. for C26H28F4N6OS : C 56.92, H 5.14, N 15.32 Found: C 56.85, H
5.21, N 15.18.
7-(4-fluoro-3-(trifluoromethyl)phenylamino)-5-methyl-2-(3-((5-((4-methylpiperazin-1-yl)methyl)furan-
2-yl)methylthio)propyl)-[1,2,4]triazolo[1,5-a]pyrimidine (18)
Yield: 79%; m.p. 179-180 °C; MS [MH+] (m/z): 578.0; IR (KBr) cm-1: 3444.7 (υNH), 2965.7 (υCH3),
1
1610.2, 1575.8 (υC=C, υC=N), 1478.7 (δCH2), 1329.5 (υC-N). H-NMR(DMSO-d6)δ: 2.02 (pentad, 2H,
CH2), 2.41 (s, 3H, CH3), 2.59 (t, 2H, J = 7.1 Hz, CH2), 2.72 (br s, 7H, CH3, 2×CH2), 2.89 (t, 2H, J =
7.1 Hz, CH2), 3.15 (br s, 4H, 2×CH2), 3.55 (s, 2H, CH2), 3.76 (s, 2H, CH2), 6.21 (m, 2H, furyl-2×H),
6.40 (s, 1H, C6-H), 7.63 (dd, 1H, JFH=9.9 Hz, JHH=9.2 Hz, Ph-5H), 7.83 (m, 2H, Ph-2, 6-2×H); Anal.
calcad. for C27H31F4N7OS : C 56.14, H 5.41, N 16.97 Found: C 56.21, H 5.29, N 16.86.

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7-(4-fluoro-3-(trifluoromethyl)phenylamino)-5-methyl-2-(3-((5-((diethylamino)methyl)furan-2-yl)meth
yllthio)propyl)-[1,2,4]triazolo[1,5-a]pyrimidine (19)
Yield: 45%; m.p. 172-174 °C; MS [MH+] (m/z): 551.2; IR (KBr) cm-1: 3445.2 (υNH), 2970.1 (υCH3),
1575.6, (υC=C, υC=N), 1479.5 (δCH2), 1329.5 (υC-N). 1H-NMR(CDCl3)δ: 1.11 (t, 6H, J = 6.9 Hz,, 2×CH3),
2.15 (pentad, 2H, CH2), 2.54 (s, 3H, CH3), 2.61 (m, 6H, 3×CH2), 2.99 (t, 2H, J = 7.4 Hz, CH2), 3.72 (s,
4H, 2×CH2), 6.12 (m, 2H, furyl-2H), 6.18 (s, 1H, C6-H), 7.36 (dd, 1H, JFH=9.0 Hz, JHH=9.0 Hz, Ph-
5H), 7.64 (m, 2H, Ph-2, 6-2×H); Anal. calcad. for C26H30F4N6OS : C 56.71, H 5.49, N 15.26 Found: C
56.57, H 5.39, N 15.12.
7-(4-fluoro-3-(trifluoromethyl)phenylamino)-5-methyl-2-(3-((5-((dimethylamino)methyl)furan-2-
yl)methylthio)propyl)-[1,2,4]triazolo[1,5-a]pyrimidine (20)
Yield: 47%; m.p. 166-168 °C; MS [MH+] (m/z): 523.1; IR (KBr) cm-1: 3443.9 (υNH), 2966.3 (υCH3),
1
1609.5, 1575.3 (υC=C, υC=N), 1479.8 (δCH2), 1328.2 (υC-N). H-NMR(CDCl3)δ: 2.14 (pentad, 2H, CH2),
2.33 (s, 6H, 2×CH3), 2.54 (s, 3H, CH3), 2.63 (t, 2H, J = 7.1 Hz, CH2), 2.99 (t, 2H, J = 7.2 Hz, CH2),
3.56 (s, 2H, CH2), 3.72 (s, 2H, CH2), 6.13 (br s, 1H, furyl-H), 6.17 (br s, 2H, furyl-H , C6-H), 7.36 (dd,
1H, JFH=9.0 Hz, JHH=8.7 Hz, Ph-5H), 7.64 (m, 2H, Ph-2, 6-2×H); Anal. calcad. for C24H26F4N6OS : C
55.16, H 5.01, N 16.08 Found: C 55.03, H 4.92, N 16.15.
Pharmacology
The anticancer activities of compounds 6-20 were evaluated in vitro on Bel-7402 (Human Liver
Cancer cell lines) and HT-1080 (Human Fibro Sarcoma cell lines) by measuring cell viability by the
MTT method, with cisplatin as the positive control. The cells were seeded in RPM I 1640 medium
(100 µL) in a 96-well plate at a concentration of 4000 cells per well. After culturing for 12 h at 37 °C
and 5% CO2, cells were incubated with various concentrations of the samples for 24 h. MTT was
added at a terminal concentration of 5µg/mL and incubated with the cells for 4 h. The formazan
crystals were dissolved in DMSO (100 µL) in each well and the optical density was measured at 492
nm (for the absorbance of MTT formazan) and 630 nm (for the reference wavelength). The IC50 was
calculated using the Bacus Laboratories Incorporated Slide Scanner (Bliss) software. Each compound
was tested in triplicate at every concentration.
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Sample Availability: Samples of the compounds mentioned above are available from authors
© 2007 by MDPI (http://www.mdpi.org). Reproduction is permitted for noncommercial purposes.