Synthesis and antitumor activities of unsymmetrically disubstituted acylthioureas fused with hydrophenanthrene structure

Article abstract of DOI:10.1007/s00044-010-9303-8

A series of novel unsymmetrically disubstituted acylthioureas fused with hydrophenanthrene structure were synthesized from Δ⁸- dihydroabietic and dehydroabietic acid, respectively. Their structures were characterized by IR, ¹H-, and

Full text of DOI:10.1007/s00044-010-9303-8

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2011) 20:333–338
DOI 10.1007/s00044-010-9303-8
ORIGINAL RESEARCH
Synthesis and antitumor activities of unsymmetrically
disubstituted acylthioureas fused with hydrophenanthrene
structure
•
Xiao-Ping Rao Yong Wu Zhan-Qian Song
•
•
•
Shi-Bin Shang Zong-De Wang
Received: 9 June 2009 / Accepted: 13 January 2010 / Published online: 7 March 2010
Ó Springer Science+Business Media, LLC 2011
Abstract A series of novel unsymmetrically disubstituted
acylthioureas fused with hydrophenanthrene structure were
synthesized from D8-dihydroabietic and dehydroabietic
acid, respectively. Their structures were characterized by IR,
1H-, and 13C-NMR spectroscopy. The antitumor activities of
the title compounds against SMMC7721 and A549 tumor
cells were evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-
2,5-diphenyltetrazolium bromide) method. The results
showed that three compounds (4c1, 4d1, and 4d2) exhibited
highly effective activities against SMMC7721 and A549
cells, their IC50 values are between 1.87–12.67 lM for
SMMC7721 cells and 2.20–6.79 lM for A549 cells,
respectively. Structure–activity relationship indicating that
acylthioureas that furan group fused with D8-dihydroabietyl
group, trihydroxymethyl fused with D8-dihydroabietyl and
dehydroabietyl could generate enhance activities of this kind
of compounds against SMMC7721 and A549 cells.
Especially in the area of cancer diseases, there are up to
60% of these drugs were shown to be of natural origin
during past decades (Cragg et al., 1997). New compounds
having antitumor activity have still been attracted great
attention (Son et al., 2005). Rosin is an abundantly
available natural product, which is mainly composed of a
series of diterpenic monocarboxic acids (Wang et al.,
2008), in which dehydroabietic acid (DHA) and dihy-
droabietic acid are more stable to air oxidation than
abietic acid due to change of conjugate double bonds
(Chyu et al., 2005). The potential of abietane acids as
source for bioactive compounds was reviewed by San
Felician et al. (1993). It has been found that pine resin
acids and their derivatives have shown a broad spectrum
of biological activities such as antitumor (Rao et al.,
2008a, b), antimicrobial (Savluchinske et al., 1999),
antiinflammatory (Fernandez et al., 2001), and gastro-
protective (Sepulveda et al., 2005) effects. Rosin acids
were reported to have properties of enhancing the
inhibitory activity of an anticancer drug in cervical car-
cinoma cells, hepatocellular carcinoma cells, or breast
cancer cells (Lin and Chuang, 2003).
Keywords Unsymmetrically disubstituted acylthioureas ꢀ
Hydrophenanthrene structure ꢀ D8-Dihydroabietic acid ꢀ
Dehydroabietic acid ꢀ Antitumor activities
In our previous study, we had successfully obtained a
series of unsymmetrically N,N0-substituted ureas from
DHA, the cytotoxicity activity of these novel ureas
against SMMC7721 cells was also investigated (Rao
et al., 2008b). Although much attention has been paid to
the biological activities of nitrogen derivatives of pine
resin acids, the antitumor activities of unsymmetrically
disubstituted acylthioureas fused with hydrophenanthrene
structure have not yet been dealt with. In this article, we
report the synthesis and antitumor activities of novel
unsymmetrically acylthioureas fused with hydrophe-
nanthrene structure from D8-dihydroabietic and dehydro-
abietic acid.
Natural products are a rich source for drugs discovery
and development process (Newman et al., 2003).
X.-P. Rao (&) ꢀ Y. Wu ꢀ Z.-Q. Song ꢀ S.-B. Shang ꢀ
Z.-D. Wang
Institute of Chemical Industry of Forest Products,
Chinese Academy of Forestry, Nanjing 210042,
People’s Republic of China
e-mail: rxping2001@163.com
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Med Chem Res (2011) 20:333–338
Experimental
Materials and methods
Table 1 Structures of the title compounds
Compd
R1
R2
R3
4a1
4b1
4c1
A
A
A
NH
O
All solvents were of analytical grade and the reagents were
used as purchased. Infrared spectra was recorded as KBr
pellets on a Bio-Rad FTS-185 IR spectrophotometer.
Melting points were determined by XT5 digital melting-
O
1
point apparatus and were uncorrected. H- and 13C-NMR
H
spectra were recorded on a DPX-300 Bruker AVANCE
300 spectrometer (CDCl3 as solvent).
-C(CH2OH)3
-CH2CH2OH
4d1
4e1
4f1
A
A
A
H
H
H
-CH2CHOHCH3
General procedure for the synthesis of unsymmetrically
disubstituted acylthioureas
4g1
A
H
D8-Dihydroabietic (A) or dehydroabietic (B) acid (100 g)
dissolved in 150 ml CHCl3 was gradually added to a
solution of 36.5 ml SOCl2, the reaction was performed for
5 h at 40–45°C. Then cooled to room temperature and the
solvent was distilled off under vacuum. The synthesized
chloride was slowly added to the 38.5 g KSCN in 80 ml
CH3CN solution, the mixture was quickly stirred for 1.5 h
under reflux, and then the precipitates were filtered.
50 mmol acylthioisocyanate and 50 mmol amine were
added to 30 ml CH2Cl2 solution, the mixture was stirred at
40°C for 6 h, the solvents was distilled off under vacuum
and crystals were crystallized from ethanol to give acy-
lthioureas. The reaction process are shown in Scheme 1,
the structures of the title compounds are shown in Table 1.
4a1: C25H41N3OS, m.p. 136.9–137.8, yield: 59.9%; IR
(cm-1): 3419 (N–H), 2929 2870 (–CH3,–CH2), 1701
([C=O), 1523 (O=C–N), 1234 ([C=S), 1191 (S=C–N),
4a2
4b2
4c2
B
B
B
NH
O
O
H
-C(CH2OH)3
-CH2CH2OH
4d2
4e2
4f2
B
B
B
H
H
H
-CH2
CHOHCH
3
4g2
B
H
A=
B=
1
1017 (C–N); H-NMR: (CDCl3, d/ppm, 300 MHz), 8.06
(1H O=C–NH), 4.23–3.69 (4H S=C–N(CH2)2), 2.01 (1H
CH2–NH), 2.04–1.86 (4H N–CH2), 1.44 (1H CH(CH3)),
1.63 (2H[CH–), 1.78–1.34 (16H –CH2–), 1.26–0.77 (12H
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 180.3 (1C,
C=S), 175.4 (1C, C=O), 137.3, 126.2 (2C, C=C); 56.1 (2C,
–N–CH2), 50.6, 50.3 (2C, –NH–CH2); 47.3, 35.0 (2C, C–
C); 48.5, 36.9, 32.5 (3C, CH–C); 40.4, 37.2, 34.3, 31.7,
27.0, 24.6, 18.0, 16.0 (8C, CH2–C); 21.5, 19.8, 19.5, 19.3
(4C, CH3).
(16H –CH2–), 1.30–0.76 (12H –CH3). 13C-NMR: (CDCl3,
d/ppm, 300 MHz), 180.2 (1C, C=S), 175.1 (1C, C=O),
137.0, 125.0 (2C, C=C); 66.1 (2C, –O–CH2), 52.4, 51.6
(2C, –N–CH2); 47.2, 35.2 (2C, C–C); 48.4, 37.1, 32.7 (3C,
CH–C); 40.3, 37.8, 34.2, 31.6, 27.0, 24.3, 18.1, 16.4 (8C,
CH2–C); 21.5, 20.6, 19.7, 19.3 (4C, CH3).
4c1: C26H38N2O2S, m.p. 93.3–94.8, yield: 30.2%; IR
(cm-1): 3434 3234 (N–H), 2952 2930 2871 (–CH3, –CH2),
1672 ([C=O), 1543 1512 (O=C–N), 1315 ([C=S), 1149
(S=C–N), 1013 (C–O). 1H-NMR: (CDCl3, d/ppm,
300 MHz), 10.94 (1H O=C–NH), 8.60 (1H S=C–NH),
7.39–6.33 (3H=CH–), 4.22 (2H NH–CH2–), 1.43 (1H
CH(CH3)), 1.62 (2H [CH–), 2.04–1.24 (16H –CH2–),
4b1: C25H40N2O2S, m.p. 128.8–131.4, yield: 63.2%; IR
(cm-1): 3417 3364 (N–H), 2933 2870 (–CH3, –CH2), 1691
([C=O), 1524 (O=C–N), 1244 ([C=S), 1144 (S=C–N),
1
1045 (C–O). H-NMR: (CDCl3, d/ppm, 300 MHz), 8.00
(1H O=C–NH), 4.16–3.50 (4H S=C–N(CH2)2), 3.79 (4H
O–CH2–), 1.43 (1H CH(CH3)), 1.62 (2H[CH–), 2.04–1.34
Scheme 1 Synthetic scheme of
SOCl2
O
S
O
R3R2NH
KSCN
unsymmetrically disubstituted
R1COOH
R1COCl
2
R1CNCS
3
R1CNHCNR2R3
4
acylthioureas
1
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Med Chem Res (2011) 20:333–338
335
1.20–0.76 (12H –CH3). 13C-NMR: (CDCl3, d/ppm,
300 MHz), 179.9 (1C, C=S), 179.7 (1C, C=O), 149.2,
142.6, 137.2, 126.1, 110.4, 108.5 (6C, C=C); 46.8, 35.4
(2C, C–C); 48.5, 36.9, 32.5 (3C, CH–C); 42.6, 40.3, 37.0,
34.2, 31.6, 27.0, 24.0, 18.0, 16.0 (9C, CH2–C); 21.2, 20.6,
19.8,19.3 (4C, CH3).
179.0 (1C, C=O), 137.3, 136.2, 132.6, 130.4, 127.1, 126.1,
125.6, 124.9 (8C, C=C); 46.8, 34.9 (2C, C–C); 36.9, 32.5
(3C, CH–C); 40.3, 37.0, 48.5, 34.1, 31.5, 26.9, 24.4, 17.7,
16.0 (8C, CH2–C); 21.2, 19.8, 19.3, 19.2, 17.9 (5C, CH3).
4a2: C25H37N3OS, m.p. 143.2–143.6, 52.3%; IR (cm-1):
3417 (N–H), 2955 2930 2868 (–CH3, –CH2), 1701
([C=O), 1523 (O=C–N), 1285 ([C=S), 1189 (S=C–N),
1017 (C–N), 826 (Ar). 1H-NMR: (CDCl3, d/ppm,
300 MHz), 8.04 (1H O=C–NH), 7.25–6.88 (3H C=CH–),
4.22–3.69 (4H S=C–N(CH2)2), 2.96–2.79 (4H N–CH2),
2.35–2.33 (1H CH(CH3)), 2.08 (1H CH2–NH), 1.77 (1H
[CH–), 2.35–1.48 (10H –CH2–), 1.34–1.20 (12H –CH3).
13C-NMR: (CDCl3, d/ppm, 300 MHz), 180.3 (1C, C=S),
174.9 (1C, C=O), 146.3, 146.0, 134.3, 128.9, 127.0, 124.0
(6C, C=C); 58.3 (2C, –N–CH2), 50.3, 49.9 (2C, –NH–
CH2); 45.7, 36.9 (2C, C–C); 48.5, 33.4 (2C, CH–C); 37.8,
37.1, 29.8, 18.6, 16.2 (5C, CH2–C); 25.1, 24.0, 21.7 (4C,
CH3).
4d1: C25H42N2O4S, m.p. 138.6–140.4, yield: 28.5%; IR
(cm-1): 3427 (N–H), 3384 3021 (O–H), 2932 2872 (–CH3,
–CH2), 1676 ([C=O), 1549 1520 (O=C–N), 1237 ([C=S),
1
1167 (S=C–N), 1101 1055 (C–O). H-NMR: (CDCl3, d/
ppm, 300 MHz), 11.49 (1H O=C–NH), 8.49 (1H S=C–
NH), 3.91–3.89 (6H –CH2–OH), 3.27–3.25 (3H –OH), 1.45
(1H CH(CH3)), 1.61 (2H[CH–), 2.00–1.25 (16H –CH2–),
1.20–0.85 (12H –CH3). 13C-NMR: (CDCl3, d/ppm,
300 MHz), 179.8 (1C, C=S), 178.9 (1C, C=O), 137.2,
126.2 (2C, C=C); 64.1 (1C, NHC), 64.1 (3C, CH2OH),
46.8, 35.1 (2C, C–C); 48.8, 37.0, 32.5 (3C, CH–C); 40.3,
37.1, 34.2, 31.6, 27.0, 24.6, 18.0, 16.0 (8C, CH2–C); 21.3,
19.8, 19.5,19.3 (4C, CH3).
4b2: C25H36N2O2S, m.p. 118.2–119.3, 90.1%; IR
(cm-1): 3417 (N–H), 2957 2930 2866 (–CH3, –CH2), 1698
([C=O), 1529 (O=C–N), 1275 ([C=S), 1141 (S=C–N),
4e1: C23H38N2O2S, m.p. 128.8–130.0, 48.3%; IR
(cm-1): 3433 (N–H), 3330 3247 (O–H), 2931 2871 (–CH3,
–CH2), 1679 ([C=O), 1557 1513 (O=C–N), 1214 ([C=S),
1167 (S=C–N), 1060 (C–O). 1H-NMR: (CDCl3, d/ppm,
300 MHz), 11.00 (1H O=C–NH), 8.62 (1H S=C–NH),
3.91–3.85 (4H –CH2–CH2–OH), 2.93 (1H –OH), 1.41 (1H
CH(CH3)), 1.62 (2H [CH–), 2.17–1.25 (16H –CH2–),
1.20–0.98 (12H –CH3). 13C-NMR: (CDCl3, d/ppm,
300 MHz), 180.7 (1C, C=S), 179.8 (1C, C=O), 137.2,
126.2 (2C, C=C); 47.7 (1C, NHC), 60.8 (1C, CH2OH),
46.8, 35.0 (2C, C–C); 48.6, 36.9, 32.5 (3C, CH–C); 40.3,
37.0, 34.2, 31.6, 27.0, 24.5, 18.0, 16.0 (8C, CH2–C); 21.3,
19.8, 19.5, 19.3 (4C, CH3).
1
1034 (C–O). H-NMR: (CDCl3, d/ppm, 300 MHz), 7.97
(1H O=C–NH), 7.16–6.88 (3H C=CH–), 4.16–3.51 (4H
S=C–N(CH2)2), 3.79 (4H O–CH2–), 2.35–2.32 (1H
CH(CH3)), 1.77 (1H [CH–), 2.16–1.49 (10H –CH2–),
1.34–1.21 (12H –CH3). 13C-NMR: (CDCl3, d/ppm,
300 MHz): 179.7 (1C, C=S), 174.7 (1C, C=O), 146.4,
145.9, 134.3, 126.9, 124.0 (6C, C=C); 66.2 (2C, –O–CH2),
52.4, 51.6 (2C, –N–CH2); 45.8, 36.9 (2C, C–C); 48.4, 33.4
(2C, CH–C); 37.8, 37.1, 29.8, 18.6, 16.5 (5C, CH2–C);
25.1, 24.0, 23.9, 21.6 (4C, CH3).
4c2: C26H34N2O2S, m.p. 109.8–110.0, 50.2%; IR
(cm-1): 3435 3239 (N–H), 2956 2927 2871 (–CH3, –CH2),
1675 ([C=O), 1546 1515 (O=C–N), 1312 ([C=S), 1158
(S=C–N), 1016 (C–O), 826 (Ar). 1H-NMR: (CDCl3, d/
ppm, 300 MHz), 10.90 (1H O=C–NH), 8.59 (1H S=C–
NH), 7.38–6.33 (6H C=CH–), 4.86–4.79 (2H NH–CH2–),
2.79–2.84 (1H CH(CH3)), 1.77 (1H [CH–), 2.34–1.47
(10H –CH2–), 1.33–1.20 (12H –CH3). 13C-NMR: (CDCl3,
d/ppm, 300 MHz), 180.0 (1C, C=S), 179.4 (1C, C=O),
149.2, 146.3, 146.0, 142.7, 134.2, 126.9, 124.1, 124.0,
110.5, 108.6 (10C, C=C); 42.6 (1C NH–CH2–), 45.3, 37.0
(2C, C–C); 48.5, 33.5 (2C, CH–C); 37.7, 37.1, 29.7, 18.5,
16.2 (5C, CH2–C); 25.1, 24.0, 24.0, 21.4 (4C, CH3).
4d2: C25H38N2O4S, m.p. 163.6–164.6, 67.0%; IR
(cm-1): 3427 (N–H), 3384 3045 (O–H), 2955 2934 2871
(–CH3, –CH2), 1677 ([C=O), 1551 1520 (O=C–N), 1233
([C=S), 1169 (S=C–N), 1101 1052 (C–O). 1H-NMR:
(CDCl3, d/ppm, 300 MHz), 11.44 (1H O=C–NH), 8.49 (1H
S=C–NH), 7.16–6.87 (3H C=CH–), 3.88–3.87 (6H –CH2–
OH), 3.72–3.33 (3H –OH), 2.92 (1H CH(CH3)), 1.77 (1H
[CH–), 2.17–1.68 (10H –CH2–), 1.34–1.19 (12H –CH3).
13C-NMR: (CDCl3, d/ppm, 300 MHz), 179.4 (1C, C=S),
4f1: C24H40N2O2S, m.p. 132.8–134.9, 34.8%; IR
(cm-1): 3434 (N–H), 3321 3255 (O–H), 2929 2870 (–CH3,
–CH2), 1680 ([C=O), 1557 1510 (O=C–N), 1215 ([C=S),
1171 (S=C–N), 1057 (C–O). 1H-NMR: (CDCl3, d/ppm,
300 MHz), 11.00 (1H O=C–NH), 8.60 (1H S=C–NH),
4.15–3.49 (3H –CH2–CH–OH), 2.90 (1H –OH), 1.41 (1H
CH(CH3)), 1.62 (2H [CH–), 2.05–1.34 (16H –CH2–),
1.28–0.83 (15H –CH3). 13C-NMR: (CDCl3, d/ppm,
300 MHz): 180.6 (1C, C=S), 179.8 (1C, C=O), 137.2,
126.2 (2C, C=C), 66.3 (1C, NHC), 52.6 (1C, CH2OH),
46.8, 35.1 (2C, C–C); 48.6, 36.9, 32.5 (3C, CH–C); 40.3,
37.0, 34.2, 31.6, 27.0, 24.5, 18.0, 16.0 (8C, CH2–C); 21.2,
19.8, 19.7, 19.4, 19.3 (5C, CH3).
4g1: C28H40N2OS, m.p. 155.5–156.0, 72.6%; IR (cm-1):
3400 (N–H), 2946 2867 (–CH3, –CH2), 1670 ([C=O),
1552 1519 (O=C–N), 1252 ([C=S), 1144 (S=C–N), 1107
1
(Ar–N), 830 (Ar). H-NMR: (CDCl3, d/ppm, 300 MHz),
12.27 (1H O=C–NH), 8.72 (1H S=C–NH), 7.80–7.17 (4H
C=CH–), 2.30 (3H Ar–CH3), 1.42 (1H CH(CH3)), 1.64 (2H
[CH–), 2.01–1.28 (16H –CH2–), 1.22–0.82 (12H –CH3).
13C-NMR: (CDCl3, d/ppm, 300 MHz): 180.0 (1C, C=S),
123

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Med Chem Res (2011) 20:333–338
178.9 (1C, C=O), 146.3, 146.0, 134.1, 126.9, 124.0, 123.9
(6C, C=C); 64.0 (1C, NHC), 62.2 (3C, CH2OH), 45.2, 37.0
(2C, C–C); 48.7, 33.4 (2C, CH–C); 37.6, 29.6, 18.4, 16.0
(5C, CH2–C); 25.0, 23.9, 21.4 (4C, CH3).
maintained at 37°C in a humidified 5% CO2, and incubated
for 48 h, then 20 ll MTT solution was added and the plate
was incubated for additional 4 h. Supernatant from each
well was drawn out carefully and 150 ml DMSO was
added to each well and subjected to vibration on ELISA
spectrophotometer at 570 nm. IC50 and 95% confidence
intervals values were calculated by GWBASIC software
(Lokhande et al., 2006).
4e2: C23H34N2O2S, m.p. 181.7–182.2, 76.4%; IR
(cm-1): 3433 (N–H), 3322 3256 (O–H), 2956 2932 2871
(–CH3, –CH2), 1683 ([C=O), 1558 1510 (O=C–N), 1226
([C=S), 1171 (S=C–N), 1060 (C–O). 1H-NMR: (CDCl3, d/
ppm, 300 MHz), 10.96 (1H O=C–NH), 8.60 (1H S=C–
NH), 7.24–6.87 (3H C=CH–), 3.91–3.85 (4H –CH2–OH),
2.91 (1H –OH), 2.89 (1H CH(CH3)), 1.77 (1H [CH–),
2.31–1.67 (10H –CH2–), 1.49–1.46 (12H –CH3). 13C-
NMR: (CDCl3, d/ppm, 300 MHz), 180.7 (1C, C=S), 179.4
(1C, C=O), 146.2, 146.0, 134.2, 126.9, 124.1, 123.9 (6C,
C=C); 60.8 (1C, NHC), 47.7 (1C, CH2OH), 45.2, 37.0 (2C,
C–C); 48.5, 33.4 (2C, CH–C); 37.6, 29.7, 18.5, 16.1 (5C,
CH2–C); 25.1, 23.9, 21.4 (4C, CH3).
Results and discussion
Synthesis and spectra
As shown in Scheme 1, a series of unsymmetrically
disubstituted acylthioureas fused with hydrophenthrane
structure were synthesized through three steps of reactions.
Chemical modification of acid group of rosin acids often
need high temperature and pressure (Kushnir et al., 2003),
modify the acid group to active group can avoid these
shortcomings, acyl chloride, anhydride, and isocyanate of
rosin acids were proved to be very active intermediates
(Zhang et al., 2005). Although high steric hinderance of
tricyclo structure to the carbonyl group, acyl chloride can
be easily obtained by the reaction of rosin acids with
SOCl2, and acyl chloride can be converted to isothiocya-
nate with KSCN, both intermediates exhibited strong
activities and afford good yields of final products at mild
reaction conditions.
4f2: C24H36N2O2S, m.p. 155.6–165.5, 58.8%; IR
(cm-1): 3319 (N–H), 3199 (O–H), 2960 2928 2851 (–CH3,
–CH2), 1686 ([C=O), 1562 1539 (O=C–N), 1226 ([C=S),
1177 (S=C–N), 1054 (C–O). 1H-NMR: (CDCl3, d/ppm,
300 MHz), 10.96 (1H O=C–NH), 8.58 (1H S=C–NH),
7.17–6.87 (3H C=CH–), 3.54–3.52 (3H –CH2– CH–OH),
2.90 (1H –OH), 2.84 (1H CH(CH3)), 1.75 (1H [CH–),
2.31–1.45 (10H –CH2–), 1.34–1.19 (15H –CH3). 13C-
NMR: (CDCl3, d/ppm, 300 MHz): 180.6 (1C, C=S), 179.4
(1C, C=O), 146.2, 146.0, 134.2, 126.9, 124.0, 123.9 (6C,
C=C); 64.4 (1C, NHC), 52.6 (1C, CH2OH), 45.2, 37.0 (2C,
C–C); 48.5, 33.4 (2C, CH–C); 37.6, 29.7, 18.5, 16.1 (5C,
CH2–C); 25.0, 23.9, 21.4, 21.2 (5C, CH3).
1
The acylthioureas were characterized by IR, H-NMR,
and 13C-NMR spectra. In the IR spectra, characteristic
4g2: C28H36N2OS, m.p. 144.4–144.6, 77.4%; IR (cm-1):
3399 (N–H), 2950 2867 (–CH3, –CH2), 1669 ([C=O),
1552 1518 (O=C–N), 1253 ([C=S), 1139 (S=C–N), 1107
banks at 1,669–1,701 cm-1 for [C=O group and
1
1,205–1,315 cm-1 for [C=S group appeared. In the H-
1
(Ar–N), 824 (Ar). H-NMR: (CDCl3, d/ppm, 300 MHz),
NMR spectra, chemical shift of hydrogen of NH group
appeared near 8.59–10.90 ppm. In the 13C-NMR spectra,
chemical shift of C=O and C=S appeared at near 179.0 and
180.0 ppm, respectively. The vibration of all other func-
tional group are present, which indicating the formation of
acylthioureas.
12.25 (1H O=C–NH), 8.72 (1H S=C–NH), 7.79–6.88 (7H
C=CH–), 2.83 (1H CH(CH3)), 2.30 (3H Ar–CH3), 1.73 (1H
[CH–), 2.36–1.52 (10H –CH2–), 1.39–1.21 (12H –CH3).
13C-NMR: (CDCl3, d/ppm, 300 MHz): 179.6 (1C, C=S),
179.1 (1C, C=O), 146.9, 134.1, 130.6, 127.3, 126.8, 126.2,
125.7, 124.0, 123.1 (12C, C=C); 45.2, 36.3 (2C, C–C);
48.5, 33.3 (2C, CH–C); 37.6, 36.9, 29.6, 18.4, 16.1 (5C,
CH2–C); 24.9, 23.8, 21.4, 17.8 (5C, CH3).
Antitumor activities
Pharmacology
In order to investigate the antitumor activities of synthe-
sized unsymmetrically disubstituted acylthioureas,
SMMC7721, and A549 cells were selected as tumor cell
lines. All compounds were subjected to a preliminary
screening for antitumor activities at a fixed sample con-
centration of 10 lM, using the MTT (3-(4,5-dimethylthi-
azol-2-yl)-2,5-diphenyltetrazolium bromide) assay, respec-
tively. The inhibition ratios of the title compounds against
SMMC7721 and A549 cells for incubation of 48 h are
listed in Table 2. As can be concluded from the results,
SMMC7721 or A549 cells were propagated continuously
in culture and grown in RPMI 1640 medium with 10%
inactivated fetal calf serum and antibiotics. The cells har-
vested from exponential phase were seeded equivalently
into 96-well plates and incubated for 8 h, then the selected
compounds were added in concentration gradient. The final
concentrations were maintained at 10 lM. Control was
added the same amount of solvent. The plates were
123

Med Chem Res (2011) 20:333–338
337
Table 2 Inhibition ratios of compounds against SMMC7721 and A549 cells at 10 lM for 48 h
Cells
Inhibition ratios
4a1
4b1
4c1
4d1
4e1
4f1
4g1
4a2
4b2
4c2
4d2
4e2
4f2
4g2
SMMC7721
A549
19.73
19.52
33.70
26.20
93.47
92.64
34.14
80.86
13.46
6.14
34.40
12.69
18.56
13.09
23.21
28.71
42.09
19.00
14.21
0.00
61.72
82.89
48.68
35.44
40.29
10.20
24.27
8.91
change the substituted groups on N atoms leads to increase
in antitumor activities. Compounds that furan group fused
with D8-dihydroabietyl group (compound 4c1) exhibited
highest activities both against SMMC7721 and A549 cells,
the inhibition ratios reached 93.47 and 92.64%, respec-
tively. However, compounds that furan group fused with
dehydroabietyl group (compound 4c2) exhibited weak
activities against SMMC7721 or A549 cells. Compounds
that trihydroxymethyl fused with D8-dihydroabietyl (com-
pound 4d1) and dehydroabietyl (compound 4d2) exhibited
higher activities against A549 than SMMC7721, the inhi-
bition ratios were 80.86, 82.89, 34.14, and 61.72%,
respectively.
enhancement to antitumor activity except of compounds
4c1 and 4c2. Compound that furan group fused with D8-
dihydroabietyl group exhibited high activity, however,
compound that the furan group fused with dehydroabietyl
group exhibited low activity. The antitumor activities of
acylthioureas against SMMC7721 cells were compared
with ureas, which were reported previously (Rao et al.,
2008b), it has been found that they exhibited the same level
of antitumor activities against SMMC7721 cells. The
analysis of structure–activity relationships suggested that
compounds that furan group fused with D8-dihydroabietyl
group, trihydroxymethyl fused with D8-dihydroabietyl, and
dehydroabietyl group could generate enhance activities of
this kind of compounds against SMMC7721 and A549
cells.
The inhibition ratios at different concentrations were
evaluated for three most active compounds (compounds
4c1, 4d1, and 4d2). The IC50 and 95% confidence interval
values were calculated by GWBASIC software. The IC50
and 95% confidence interval data were listed in Table 3.
The IC50 values of three most active compounds are
between 1.87–12.67 lM against SMMC7721 tumor cells
and 2.20–6.79 lM against A549 tumor cells, respectively.
The antitumor activity against SMMC7721 of this work
were compared with that of N,N0-disubstituted ureas of our
previous work with IC50 values between 8.8 and 14.2 lM
(Rao et al., 2008b). The antitumor activity of acylthioureas
and ureas are in the same order of magnitude with 10-6 M,
change substituted groups can slightly improve the antitu-
mor activity.
Possible mechanism
We think the unsymmetrically acylthioureas fused with
hydrophenanthrene possess the same mechanism as imines
that previously reported (Li et al., 2008). Its mechanism
may have been related to the decrease in the expression of
anti-apoptotic protein, Bcl-2, accompanied by a drop in the
mitochondrial membrane potential and the activation of
caspase-3, that led to apoptotic body formation and finally
apoptosis.
Conclusions
Structure–activity relationship
In summary, a series of unsymmetrically disubstituted a-
cylthioureas were synthesized from D8-dihydroabietic and
dehydroabietic acid. Their structures were characterized by
From the results of antitumor activities of the title com-
pounds, the introduced substituted groups have important
effects to the antitumor activities. The change of the tri-
cyclic hydrophenanthrene structure has no obvious
1
IR, H-, and 13C-NMR spectroscopy. Some of these com-
pounds showed efficacious antitumor activities against
Table 3 IC50 and 95% confidence interval data of three compounds against SMMC-7721 and A-549 cells
Compound
SMMC7721
A549
IC50 (lM)
95% confidence interval
IC50 (lM)
95% confidence interval
4c1
4d1
4d2
1.87
12.67
6.84
1.30–2.70
10.68–15.04
6.08–7.69
2.20
6.79
6.44
1.51–3.21
5.15–8.94
5.68–7.29
123

338
Med Chem Res (2011) 20:333–338
dehydroabietylamine derivative, on human hepatocellular carci-
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SMMC7721 and A549 cells. Three compounds (4c1, 4d1,
and 4d2) exhibited highly effective activities against
SMMC7721 and A549 cells, their IC50 values are between
1.87–12.67 lM for SMMC7721 cells and 2.20–6.79 lM
for A549 cells, respectively. The analysis of structure–
activity relationships suggested that acylthioureas that
furan group fused with D8-dihydroabietyl group, trihydr-
oxymethyl fused with D8-dihydroabietyl, and dehydroa-
bietyl could generate enhance activities of this kind of
compounds against SMMC7721 and A549 cells.
Acknowledgments This research was financially supported by
grants from National Natural Science Foundation of China (No.
30771690) and Forestry Commonwealth Industry Special Foundation
of China (No. 200704008).
Rao XP, Song ZQ, He L, Jia WH (2008b) Synthesis, structure
analysis and cytotoxicity studies of novel unsymmetrically N,
N’-substituted ureas from dehydroabietic acid. Chem Pharm Bull
56(11):1575–1578
San Felician A, Gordaliza M, Salinero MA, Miguel del Corral JM
(1993) Abietane acids: sources, biological activities, and ther-
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