Synthesis and antitumor screening of new 1,7-diphenyl-3-(1,3-disubstituted- 1H-pyrazole-4-carbonyl)-[1,2,4]triazolo[4,3-a]pyrimidin-5(1H)-ones

Article abstract of DOI:10.1007/s12272-010-2224-8

A new series of 3-(1,3-disubstituted-1H-pyrazole-4-carbonyl)-1,7-diphenyl- [1,2,4]triazolo[4,3-a]pyrimidin-5(1H)-ones 4 was prepared by reaction of the enaminone 2 with hydrazonoyl halides 3. The preliminary screening for antitumor activity of the synthes

Full text of DOI:10.1007/s12272-010-2224-8

Arch Pharm Res Vol 33, No 1, 55-60, 2010
DOI 10.1007/s12272-010-2224-8
Synthesis and Antitumor Screening of New 1,7-Diphenyl-3-(1,3-di-
substituted-1H-pyrazole-4-carbonyl)-[1,2,4]triazolo[4,3-a]pyrimidin-
5(1H)-ones
Ahmad S. Shawali1, Sherif M. Sherif1, Manal A. A. Darwish1, and Mahmoud M. El-merzabani2
2
1Department of Chemistry, Faculty of Science, University of Cairo, Giza, Egypt and Department of Experimental Oncol-
ogy, National Cancer Institute, University of Cairo, Giza, Egypt
(Received July 28, 2009/Revised October 28, 2009/Accepted November 1, 2009)
A new series of 3-(1,3-disubstituted-1H-pyrazole-4-carbonyl)-1,7-diphenyl-[1,2,4]triazolo[4,3-
a
]pyrimidin-5(1H)-ones
halides
carried out against Ehrlich Ascites Carcinoma tumor cells. The results revealed that the stud-
ied compounds have low or no antitumor activity towards EAC tumor cells
4 was prepared by reaction of the enaminone 2 with hydrazonoyl
3
. The preliminary screening for antitumor activity of the synthesized compounds was
4
.
Key words : Enaminone, Hydrazonoyl halides, Pyrazole, [1,2,4]Triazolo[4,3-a]pyrimidine
compounds which incorporate both the pyrazole and
1,2,4-triazolo[4,3-
]pyrimidine rings and screen their
a]pyrimidine- antitumor activity against Ehrlich Ascites Carcinoma
INTRODUCTION
a
Various derivatives of 1,2,4-triazolo[4,3-
5(1H)-one were reported to be useful as calcium-channel- tumor cells. The latter type of cells were used in this
blocking vasodilators, some other have antihyperten- study because they are the only tumor cells that grow
sive, cardiovascular and anxiolytic activities and others in mice available in Egypt. The target compounds have
are used as components in photographic materials not been reported hitherto. In continuation of our pro-
(Shawali et al., 2007; Barthelmy et al., 1985; Bru- gram aimed at exploring the synthetic potentialities of
Magniez et al., 1995; Albright et al., 1985; Nakamura hydrazonoyl halides
et al., 1991). Also, many -arylpyrazole derivatives were et al., 1980, 1995, 1980, 2005, 2006, 2007), we in-
reported to have remarkable pharmacological activities vestigated reactions of the latter with the enaminone
as antimicrobial, hypoglycemic, tumor necrosis inhi- in an attempt to synthesize the target compounds.
3 (Shawali, 1983, 1993; Shawali
N
2
bitors, antithromboembolic disorder, antiangiogeric Although there are many reports dealing with the
agents, A3 Adenosine receptor antagonist, neuropep- reactions of hydrazonoyl halides with various ena-
tide Y Y5 receptor antagonists, kinase inhibitor for minones (Shawali et al., 2006), the reactions described
treatment of type 2 diabetes, hyperlipidemia and herein have not been reported hitherto.
obesity, insecticides, thrombopiotinmimetics and anti-
inflammatory (Abdel-Gawad et al., 2003; Maekawa et
al., 2004; Mercep et al., 2004; Qiao et al., 2004; Baraldi
et al., 2003; Stamford et al., 2004; Brown et al., 2004;
MATERIALS AND METHODS
Melting points were determined on an electrother-
Stevensons et al., 2004; Heerding, 2004; Cardia et al., mal Gallenkamp apparatus. The IR spectra were
1998; Mullican et al., 1993). These findings attracted recorded in KBr discs using Pye Unicam SP-1000
our attention towards the synthesis of the title Spectrophotometer. 1H NMR spectra were recorded in
CDCl3 and DMSO-d6 using a Varian Em-200 MH
z
Correspondence to: Ahmad S. Shawali, Department of Chemis-
try, Faculty of Science, University of Cairo, Giza, Egypt
Tel: 25084164
Spectrometer, and TMS as internal reference. Mass
spectra were recorded on AEI MS 30 mass spectro-
meter operating at 70 eV. Elemental analyses were
carried out by the Microanalytical Center of Cairo
E-mail: as_shawali@mail.com
55

56
A. S. Shawali et al.
University, Giza, Egypt. 3-Acetyl-1,7-diphenyl-[1,2,4] 3-[3-Acetyl-1-(4-methylphenyl)pyrazole-4-carbonyl]-
triazolo[4,3- ]pyrimidine-5(1H)-one (Shawali et al., 1,7diphenyl-[1,2,4]triazolo[4,3- ]pyrimidin-5(1H)-
2002) and hydrazonoyl halides were prepared follow- one (4Ab)
ing literature procedures (Shawali et al., 2002; Shawali Pale yellow crystals (1.95 g, 76%), m. p 200oC. IR (KBr):
a
1
a
3
et al., 1971; Shawali et al., 1973; Wolkoff, 1975; Eweiss
et al., 1980).
ν
max/cm-1 1692, 1675, 1660. 1H NMR (CDCl3)
3H, CH3), 2.56 (s, 3H, CH3), 6.56 (s, 1H, pyrimidine H),
7.27-7.60 (m, 10H, ArH), 7.68 (d, = 8 Hz, 2H, ArH),
δ 2.46 (s,
J
3-[3-(Dimethylamino)acryloyl]-1,7-diphenyl[1,2,4]
8.31 (d, J = 8Hz, 2H, ArH), 8.75 (s, 1H, pyrazole-H).
triazolo-[4,3-a]pyrimidin-5(1H)-one (2)
MS m/z (%) 515 (M++ 1, 11) 514 (M+, 29), 367 (13), 227
A mixture of compound
1
(6.6 g, 0.02 mole) and DMF- (51), 129 (28), 102 (25), 98 (71), 91 (38), 77 (100). Anal.
DMA (2.68 g, 0.023 mole) in dry toluene (20 mL) was Calcd. for C30H22N6O3 (514.55) C, 70.03; H, 4.31; N,
refluxed for 6 h then left to cool to room temperature. 16.33. Found C, 70.21; H, 4.03; N, 16.02%.
The precipitate was filtered off, washed with ether,
dried and crystallized from ethanol to give the 3-[3-Acetyl-1-(4-chlorophenyl)pyrazole-4-carbony]-
enaminone
mp. 212oC. IR (KBr):
(CDCl3) 2.91 (s, 3H, N-CH3), 3.06 (s, 3H, N-CH3), 5.36 Pale yellow crystals (2.27g, 85%) m.p 250oC. IR (KBr):
max/cm-1, 1691, 1685, 1671. 1H NMR(CDCl3)
(d, = 13 H , 1H, =CH), 6.63 (s, 1H, pyrimidine H), 2.50 (s,
7.42-8.17 (m, 10H, ArH), 8.18 (d, = 13 Hz, 1H, =CH). 3H, CH3), 6.72 (s, 1H, pyrimidine H), 7.55-8.25(m, 14H,
2 as orange crystals (6.16 g, 80% yield), 1,7-diphenyl-[1,2,4]triazolo[4,3-a]pyrimidin-5(1H)-
1
ν
max/cm-11685, 1650; H NMR one (4Ac)
J
z
ν
δ
J
MS: m/z (%) 385 (M+, 10), 367 (22), 366 (12), 105 (3), ArH), 9.54 (s, 1H, pyrazole H). MS m/z (%) 536 (M++ 2,
98 (100), 91 (3), 89 (7), 77 (26), 70 (16). Anal. Calcd for 0.2) 535 (M++1, 0.7) 534 (M+, 0.7), 249 (15), 246 (66),
C22H19N5O2 (385.43) C, 68.56; H, 4.97; N, 18.17. 204 (16), 128 (22), 111 (18), 103 (19), 77 (96), 76 (100).
Found: C, 68.38; H, 4.69; N, 18.00%.
Anal. Calcd. for C29H19ClN6O3 (534.97) C, 65.11, H,
3.58, N, 15.71. Found C, 65.45, H, 3.80, N, 15.59%.
1,7-Diphenyl-3-(1,3-disubstituted-1H-pyrazole-
1,7-Diphenyl-3-[3-ethoxycarbonyl-1-phenylpyra-
4-carbonyl)-[1,2,4]triazolo[4,3-a]pyrimidin-5(1H)-
zole-4-carbonyl]-[1,2,4]traizolo[4,3-a]pyrimidin-5
ones (4A-G)
General Procedure
appropriate hydrazonoyl halide
enaminone
- To a stirred solution of the (1H)-one (4Ba)
3
(0.005 mol) and the Yellow crystals (2.1g, 79%), m.p 178-180oC. IR (KBr):
2
(1.93 g, 0.005 mole) in dioxane (20 mL) 1.16 (t,
max/cm-1, 1695, 1671, 1652. 1H NMR (CDCl3)
was added triethylamine (0.7 mL) dropwise with = 7 Hz, 3H, CH3), 4.15 (q, = 7 Hz, 2H, CH2), 6.63
ν
J
δ
J
stirring at room temperature for 6 h. The precipitated (s, 1H, pyrimdine H), 7.27-8.34 (m, 15H, ArH), 8.75
triethylamine hydrochloride was filtered off and the (s,1H, pyrazole H). MS m/z (%): 531 (M++ 1, 9) 530
filtrate was evaporated under reduced pressure. The (M+, 26), 367 (27), 215 (29), 178 (100), 129 (19), 111
residue was triturated with ethanol where the product (11), 104 (28), 95 (15), 83 (22), 77 (33). Anal. Calcd. for
precipitated. The latter was filtrered, and crystallized C30H22N6O4 (530.55) C, 67.92, H, 4.18, N, 15.84.
form EtOH/dioxane to give the respective product
4.
Found C, 67.70, H, 3.98, N, 16.10%.
The compounds 4A-G prepared are listed below
together with their physical constants.
1,7-Diphenyl-3-[3-ethoxycarbonyl-1-(4-methyl-
phenyl) pyrazole-4-carbonyl]-[1,2,4]triazolo[4,3-
3-[3-Acetyl-1-phenylpyrazole-4-carbonyl]-1,7-di-
a
]pyrimidin-5(1H)-one (4Bb)
Pale yellow crystals (2.23 g, 82%) m.p 206-208oC. IR
(KBr):
max/cm-1, 1705, 1698, 1669; 1H NMR (CDCl3)
= 7 Hz, 3H, CH3), 2.44 (s, 3H, CH3), 4.16 (q,
= 7 Hz, 2H, CH2) 6.63 (s, 1H, pyrimidine H), 7.27-
3H, CH3), 6.58 (s, 1H, pyrimidine H) 7.27–8.34 (m, 7.58 and 8.09-8.12 (m, 10H, ArH), 7.66 (d, = 9 Hz,
, 2H, ArH), 8.70 (s, 1H,
phenyl[1,2,4]-triazolo[4,3-a]pyrimidin-5(1H)-one
(4Aa)
ν
J
δ
Yellow crystals (1.98g, 79%), m.p 249-250oC. IR (KBr): 1.16 (t,
ν
max/cm-1, 1705, 1689, 1665. 1H NMR (CDCl3)
δ
2.57 (s,
J
J
15H, ArH), 8.79 (s, 1H, pyrazole H). MS m/z (%) 502 2H, ArH), 8.29 (d,
J = 9 Hz
(M++ 2, 6) 501 (M++1, 29), 500 (M+, 81), 457 (27), 367 pyrazole H). MS m/z (%): 546 (M++2, 8) 545 (M++1,
(16), 213 (100), 178 (24), 129 (22), 116 (12), 104 (25), 22), 544 (M+, 22), 228 (44), 128 (24), 118 (88), 103 (23),
98 (34), 77 (80). Anal. Calcd. for C29H20N6O3 (500.52) 91 (89), 77 (100). Anal. Calcd. for C31H24N6O4 (544.57)
C, 69.59; H, 4.03; N, 16.79. Found C, 69.81; H, 3.90; N, C, 68.37; H, 4.44; N, 15.43. Found C, 68.13; H, 4.70; N,
16.77%.
15.71%.

Synthesis and Antitumor Screening of 1,2,4-Triazolo[4,3-a]pyrimidinones
57
1,7-Diphenyl-3-[3-ethoxycarbonyl-1-(4-chloro-
1,7-Diphenyl-3-[3-(N-phenylaminocarbonyl)-1-(4-
phenyl)pyrazole-4-carbonyl]-[1,2,4]triazolo[4,3-
a]
methylphenyl-)pyrazole-4-carbonyl][1,2,4]triazolo
pyrimidin-5(1H)-one (4Bc)
[4,3-a]pyrimidin-5(1H)-one (4Db)
Pale yellow crystals (2.23g, 79%) m.p 185oC IR (KBr): Dark yellow crystals (2.22g, 75%), m.p 296-298oC. IR
1
ν
max/cm-1, 1710, 1692, 1640; H NMR (DMSO-d6)
δ
(KBr):
ν
max/cm-1 3105, 1698, 1682, 1651; 1H NMR (DMSO-
1.18 (t,
CH2), 6.63 (s, 1H, pyrimidine H), 7.49-7.71 (m, 10H, (d,
ArH), 7.94 (d, = 9 Hz, 2H, ArH), 8.19 (d,
J
= 7 Hz, 3H, CH3), 4.24 (q,
J
= 7 Hz, 2H, d6)
δ
2.38 (s, 3H, CH3), 6.69 (s, 1H, pyrimidine H), 7.39
= 9 Hz, 2H, ArH), 7.53-7.62 and 8.13-8.16 (m,
J
J
J
= 9 Hz, 15H, ArH), 7.89 (d,
J
= 9 Hz, 2H, ArH), 9.47 (s, 1H,
2H, ArH), 9.49 (s, 1H, pyrazole H). MS m/z (%) 565 pyrazole H), 10.72 (s, 1H, NH). MS m/z (%) 592 (M++1,
(M++1, 0.42), 564 (M+, 0.35) 451 (100), 450 (66), 422 16), 591 (M+, 38), 397 (33), 303 (88), 288 (44), 287 (59),
(26), 233 (11), 163 (7), 103 (28), 91 (50), 77 (24). Anal. 259 (29), 171 (19), 145 (22), 129 (38) 118 (66), 105 (26),
Calcd. for C30H21ClN6O4 (564.99 ) C, 63.78; H, 3.75; N, 103 (29), 91 (100), 77 (64). Anal. Calcd. For C35H25N7O3
14.87. Found C, 63.56; H, 3.63; N, 15.08%.
(591.63) C, 71.06; H, 4.26; N, 16.57. Found C, 71.17 H,
3.89, N, 16.81%.
1,7-Diphenyl-3-[3-ethoxycarbonyl-1-(4-nitrophenyl)
pyrazole-4-carbonyl][1,2,4]triazolo[4,3-
a
]pyrimidin- 1,7-Diphenyl-3-[3-(
chlorophenyl)-pyrazole-4-carbonyl][1,2,4]triazolo
]pyrimidine-5(1H)-one (4Dc)
1.18 (t, Yellow crystals (2.51g, 82%), m.p 340oC. IR (KBr):
= 7 Hz, 2H, CH2), 6.62 6.69
max/cm-1, 3105, 1684, 1653; 1H NMR (DMSO-d6)
(s, 1H, pyrimidine H), 7.27-8.11 (m, 10H, ArH), 8.28 (s, 1H, pyrimidine H), 7.21 – 7.64 (m, 15H, ArH), 7.67
N-phenylaminocarbonyl)-1-(4-
5(1H)-one (4Bd)
Yellow crystals (2.36 g, 82%), m.p 325oC. IR (KBr): [4,3-
a
ν
J
max/cm-1 ,1719, 1681, 1666; 1H NMR (CDCl3)
= 7 Hz, 3H, CH3), 4.16 (q,
δ
J
ν
δ
(d,
J = 7 Hz, 2H, ArH), 8.41 (d, J = 7 Hz, 2H, ArH), (d, J = 9 Hz, 2H, ArH), 8.06 (d, J = 9 Hz, 2H, ArH), 9.54
8.90 (s, 1H, pyrazole H). MS m/z (%) 576 (M++1, 11), (s, 1H, pyrazole H), 10.74 (s, 1H, NH). MS m/z (%) 613
575 (M+, 21) 475 (12), 261 (11), 259 (39), 213 (28), 148 (M++2, 3), (612 (M++1, 3), 611 (M+, 6), 367 (16), 325
(20), 128 (27), 117 (11), 103 (52), 91 (20), 77 (83), 76 (11), 324 (11), 323 (22), 262 (12), 204 (11), 138 (31),
(100). Anal. Clacd. for C30H21N7O6 (575.55) C, 62.61, 129 (38), 116 (15), 105 (35), 98 (85), 91 (35), 77 (100).
H, 3.68, N, 17.04. Found C, 62.45, H, 3.89, N, 17.06%.
Anal. Calcd. For C34H22ClN7O3 (612.05) C, 66.72, H,
3.62, N, 16.02. Found C, 66.61, H, 3.99, N, 15.96%.
3-[3-Benzoyl-1-phenylpyrazole-4-carbonyl]-1,7-
diphenyl[1,2,4]-triazolo[4,3-a]pyrimidin-5(1H)-
1,7-Diphenyl-3-[3-(2-thiophenyl)-1-(4-nitrophenyl)
one (4Ca)
pyrazole-4-carbonyl][1,2,4]triazolo[4,3-a]pyrimi-
Orange crystals (2.28 g, 81%), m.p 190oC. IR (KBr) ): din-5(1H)-one (4Ed)
ν
max/cm-1, 1705, 1690, 1670; 1H NMR (CDCl3) 6.48 (s, Yellow crystals (2.49g, 85%), m.p > 340oC. IR (KBr):
δ
1H, pyrimidine H), 6.61-8.13 (m, 20H, ArH), 8.85 (s, 6.72
max/cm-1, 1697, 1687, 1667; 1H NMR (DMSO-d6)
1H, pyrazole H). MS m/z (%) 563 (M++1, 2), 562 (M+, (s, 1H, pyrimidine H), 7.26-780 and 8.20-8.32 (m, 13H,
3) 534 (40), 491 (13), 274 (23), 104 (43), 77 (87), 76 ArH), 8.10 (d, = 9 Hz, 2H, ArH), 8.40 (d, = 9 Hz,
ν
δ
J
J
(100). Anal. Calcd. for C34H22N6O3 (562.59) C, 72.59; 2H, ArH), 9.54 (s, 1H, pyrazole-H). MS m/z (%) 587
H, 3.94; N, 14.94. Found C, 72.44; H, 3.76; N, 15.18%. (M++2, 21), 586 (M++1, 44), 585 (M+, 84), 556 (33), 480
(50), 323 (84), 298 (100), 253 (18), 252 (92), 150 (14),
3-[3-Benzoyl-1-(4-chlorophenyl)pyrazole-4-car-
129 (26), 122 (22), 103 (48), 91 (20), 77 (62). Anal. Clacd.
bonyl]-1,7-diphenyl-[1,2,4]triazolo[4,3-
5(1H)-one (4Cc)
a]pyrimidin- for C31H19N7O4S (585.61) C, 63.58; H, 3.27; N, 16.74.
Found C, 63.40; H, 3.35; N, 16.59%.
Yellow crystals (2.48 g, 83%), m. p 220oC. IR (KBr):
1
ν
max/cm-1, 1716, 1676, 1660; H NMR (DMSO-d6)
δ
1,7-Diphenyl-3-[3-(2-naphthoyl)-1-phenylpyrazole-
6.69 (s, 1H, pyrimidine H), 7.44-7.70 (m, 15H, ArH), 4-carbonyl]-[1,2,4]triazolo[4,3-
7.93 (d, = 8 Hz, 2H, ArH), 8.13 (d, = 8 Hz, 2H, one (4Fa)
ArH), 9.60 (s, 1H, pyrazole H). MS m/z (%) 598 (M++2, Orange crystals (2.42 g, 79%), m.p 230oC IR (KBr):
21), 597 (M++1, 18) 596 (M+, 67), 491 (34), 309 (72), max/cm-1, 1697, 1668, 1622; 1H NMR (DMSO-d6)
6.62
a]pyrimidin-5(1H)-
J
J
ν
δ
287 (25), 143 (18), 138 (25), 128 (15), 118 (15), 111 (s, 1H, pyrimidine H), 7.43-8.14 (m, 22H, ArH), 8.63
(22), 103 (24), 77 (100). Anal. Calcd. for C34H21ClN6O3 (s, 1H, pyrazole H). MS m/z (%) 613 (M++1, 21), 612
(597.04) C, 68.40; H, 3.55; N, 14.08. Found C, 68.32 H, (M+, 45), 457 (35), 325 (68), 288 (12), 155 (46), 129
3.37, N, 13.89%.
(18), 127 (100), 103 (14), 89 (11), 77 (74). Anal. Calcd.
for C38H24N6O3 (612.65), C, 74.50; H, 3.95; N, 13.72.

58
A. S. Shawali et al.
Table I. In vitro cytotoxic activity of newly synthesized
compounds and doxorubicin against (EAC)
Found C, 74.35; H, 3.89; N, 13.98%.
1,7-Diphenyl-3-[3-(2-naphthoyl)-1-(4-methylphenyl)
Non-viable cells (%)
Compound
pyrazole-4-carbonyl]-[1,2,4]triazolo[4,3-
a]pyrimi-
Concentration (µg/mL)
No.
din-5(1H)-one (4Fb)
100
50
25
Red crystals (2.50 g, 80%), m.p 240-242oC IR (KBr):
4Aa
4Ab
10
8
6
2
2
1
1
1
1
0
1
1
3
1
ν
max/cm-1 1697, 1680; H NMR (DMSO-d6)
δ
2.35 (s,
J = 9 Hz,
3H, CH3), 6.66 (s, 1H, pyrimidine H), 7.35 (d,
4Ac
4Ba
10
8
2
3
2H, ArH), 7.55-7.65 and 7.98-8.16 (m, 17H, ArH), 7.77
(d,
J = 9 Hz, 2H, ArH), 9.52 (s, 1H, pyrazole H). MS m/
4Bc
6
2
z (%) 627 (M++1, 47), 626 (M+, 100) 471 (74), 339 (56),
311 (53), 145 (15), 129 (25), 127 (69), 106 (20), 104 (24),
91 (81), 77 (12). Anal. Clacd. for C39H26N6O3 (626.68)
C, 74.75; H, 4.18; N, 13.41. Found C, 74.69; H, 4.11; N,
13.11%.
4Ca
6
3
4Cc
8
5
4Db
7
4
4Dc
6
5
4Ed
4Fa
4Ga
10
10
8
5
3
3
2
1
2
1,7-Diphenyl-3-[1,3-diphenylpyrazole-4-carbonyl]-
[1,2,4]triazolo [4,3-
a
]-pyrimidin-5(1H)-one (4Ga)
Cell control
Doxorubicin
0
100
0
55
0
20
Dark white crystals (2.1 g, 78%), m. p 200oC. IR (KBr):
1
ν
max/cm-1, 1685, 1670; H NMR (DMSO-d6)
δ
6.63 (s,
1H, pyrimidine H), 7.42-8.27 (m, 20H, ArH), 9.37 (s, where N is the number of non-viable cells counted, Nt
1H, pyrazole H). MS m/z (%) 535 (M++1, 5), 534 (M+, is the total number of cells. The test was repeated
22), 533 (17), 506 (10), 505 (20), 429 (22), 267 (19), 247 four times for each compound. The results are
(100), 129 (18), 104 (20), 89 (16), 77 (68). Anal. Clacd. summarized in Table I.
for C33H22N6O2 (534.58) C, 74.15; H, 4.15; N, 15.72.
Found C, 74.37; H, 4.35; N, 15.49%.
RESULTS AND DISCUSSION
The required new enaminone
with DMF-DMA in toluene (Scheme
1). Its structure was confirmed by its spectroscopic
2 was prepared in this
Pharmacology
Cytotoxic activity against Ehrlich Ascites Ca- study by refluxing
cinoma cell in vitro
1
1
Ehrlich Ascites Cacinoma cell (EAC), used in this data (MS, H NMR and IR) and elemental analysis
study, was kindly supplied by National Institute of (see Experimental section). Its 1H NMR spectrum
Cancer in Netherland since 1952 and mentained in showed two doublet signals at
Swiss albino mice by serial IP transplantation, RPMI = 13 Hz assignable to the two olefinic protons. This
1640 medium (Sigma), Ehrlich Ascites Cacinoma cell finding indicates that this enaminone exists in the
(EAC) suspension (2.5
105/mL), and Trypan blue dye. depicted trans configuration (Scheme 1).
A stock solution was prepared by dissolving one gram Reaction of with each of the hydrazonoyl halides
δ 5.36 and 8.18 with J
2
×
2
of the dye in distilled water (100 mL). The working 3A-G in dioxane in the presence of triethylamine
solution was then prepared by diluting (1 mL) of the gave, in each case, one isolable product as evidenced
stock solution with 9 mL of distilled water. The stain by tlc analysis. Both mass spectra and elemental
was then used for staining the dead EAC cells (El- analysis data of the isolated products were consistent
Merzabani et al., 1979; El-Merzabani et al., 1979).
A set of sterile test tubes was used, where 2.5
tumor cells/mL were suspended in phosphate - buffer- however, the isolated products were assigned struc-
ed saline. Then 25, 50, 100 g/mL from tested com- ture
. The 1H NMR spectra of the products revealed,
pound were added to the suspension, kept at 37oC for in each case, a singlet signal for the pyrazole ring pro-
with either one of the two isomeric structures
4
or
5
1
×
105 (Scheme 1). On the basis of the H NMR spectra,
4
µ
2 h. Trypan blue dye exclusion test was then carried ton in the region
δ 8.63-9.60 assignable to pyrazole-
out to calculate the percentage of nonviable cells. 5H. Literature reports indicate that the 1H NMR
Doxorubicin (Adriablastina) is taken as a positive spectra of 5- and 4-unsubstituted pyrazoles exhibit the
control. The percentage of the non-viable cells were characteristic singlet signals of 5-CH and 4-CH pro-
calculated by the following equation :
tons at
spectral simulation for 5- and 4-unsubstituted pyrazoles
showed the H-5 and 4-H signals at 8.60 and 6.50,
δ 8.35 and 7.30, respectively. Furthermore,
% of non-viable cells = 100 (N/Nt)
δ

Synthesis and Antitumor Screening of 1,2,4-Triazolo[4,3-a]pyrimidinones
59
Scheme 1.
respectively (Shawali et al., 1988). In addition, litera- toward EAC tumor cells as compared with the
ture reports (Shawali et al., 2006) indicate that reac- reference drug Doxorubicin
.
tion of hydrazonoyl halides with various enaminones
are regioselective and lead to the formation of 5-
unsubstituted pyrazole derivatives.
CONCLUSION
To account for the formation of the products
suggested that the studied reactions of with
ceed via initial dehydrohalogenation of the hydrazonoyl enaminone
halides to give the respective nitrilimines (Shawali strated. The structures of the newly synthesized
et al., 2006), which in turn cycloadd regioselectively to compounds and were confirmed by spectral data
the enaminone . The resulting cycloadducts II undergo and elemental analyses. The results of antitumor
in situ elimination of dimethylamine under the em- activity screening of compounds in series revealed
as end that all such compounds exhibited little or no activity
against Ehrlich Ascites Carcinoma cells in relation to
the reference drug Doxorubicin
4, it is
In conclusion, a simple method for the synthesis of
2
3, pro- the title heterocyclic derivatives starting from the new
2
and hydrazonoyl halides is demon-
3
3
I
2
4
2
4
ployed reaction conditions to give compounds
products (Scheme 1).
4
.
Antitumor screening tests
The in vitro cytotoxic effects of the newly synthesiz-
ed compounds and positive control drug doxorubicin
Adriablastina) were evaluated at three different con-
centrations of each against Ehrlich Ascites Carcinoma
tumor cells (EAC) in the National Institute of Cancer.
The results are summarized in Table I. As shown, all
compounds have low or no in vitro antitumor activity
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