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12. The physicochemical data of selected compounds: 1c, mp:
Figure 7. Binding model of 1h with the a,b-tubulin polypeptide whose
backbones are rendered as brown (b chain) and gray (a chain) ribbons.
Residue side chains at the colchicine site (Thr a179, Val a181, Cys
b241, Lys b254 Leu b255, Asn b258, Thr b314, Ala b316, Val b318,
and Ala b354), 1h is shown in stick with the carbon atoms of tubulin
colored green and the carbon atoms of the 1h cyan, nitrogen atoms are
colored blue, oxygen atoms red, and sulfur atoms yellow. White
dashed lines indicate potential intermolecular hydrogen bonds.
1
152–154 °C. H NMR (DMSO-d6, 300 MHz): d ppm 8.30
(d, 1H), 8.27 (d, J = 9.0, 1H), 7.79 (d, J = 9.0, 1H), 7.47
(m, 5H), 7.19 (s, 1H), 7.09 (d, J = 8.3, 1H), 6.90 (d, J = 8.3,
1H), 5.16 (s, 2H). MS (EI) m/z 422 (M+); Anal. Calcd for
(C21H15ClN4O4) C, H, N: 59.65, 3.58, 13.25. Found:
59.83, 3.42, 13.47. 1h, mp: 125–127 °C. 1HNMR (DMSO-
d6, 300 MHz): d ppm 8.22 (s, 1H), 8.14 (d, J = 8.3, 1h),
7.03 (d, J = 8.3, 1H), 6.28 (s, 2H), 3.65 (s, 6H), 3.58 (s,
3H). MS (EI) m/z 372 (M+); Anal. Calcd for
(C17H16N4O6). C, H, N: 54.84, 4.33, 15.05. Found:
54.76, 4.42, 15.14.
new class of anti-microtubule agents. The effect of the
30,40,50-substitution on the 5-anilino ring was investigat-
ed. The large alkoxyl substitution on the 40-position of
5-anilino portion is beneficial for the inhibitory activity
against the growth and mitosis of the tumor cells. How-
ever, the alkyl or amide substituent at this position is
disfavored. With respect to the antiproliferative effect
and anti-mitosis activity, this series is more potent
against HGC-27 tumor cells than A549 tumor cells. Sig-
nificantly, the 5-(30,40,50-trimethoxyl)anilino-8-nitroqui-
nazoline (1h) displays an overwhelming potency in
arresting the tumor cells at the G2/M phase of the cell
cycle, providing new templates for further development
of potent mitosis inhibitors with therapeutic potential in
treating cancer.
13. Muathen, H. A. Molecules 2003, 8, 593.
14. Cell Growth Inhibition Assay. Two human carcinoma cell
lines, A549 (lung adenocarcinoma) and HGC-27 (gastro-
intestinal carcinoma), were used for the cell proliferation
assay. A549 cells were cultured in RPMI-1640 medium
supplemented with 10% calf serum; HGC-27 cells were
cultured in DMEM supplemented with 10% calf serum.
The growth inhibition was examined by MTT assay.
15. Mossman, T. J. Immunol. Methods 1983, 65, 55.
16. Cell mitosis assay by flow cytometry analysis. Tumor
cells were incubated with indicated concentration of
synthetic inhibitor for 24 h. The harvested cells were
washed with PBS, resuspended in 1 mL of iced 75%
ethanol at ꢀ20 °C. After being left to stand overnight,
cell pellets were collected by centrifugation, resuspended
in 500 lL of hypotonic buffer (0.5% Triton X-100 in
PBS and 0.5 lg/mL RNase), and incubated at 37 °C for
30 min. Then 25 lL of propidium iodide solution (50 lg
/mL) was added, and the mixture was allowed to stand
on ice for 1 h. Fluorescence emitted from the propidi-
um iodide–DNA complex was quantitated after excita-
tion of the fluorescent dye by FAC-Scan cytometry.
The histogram of DNA distribution was modeled as a
Acknowledgment
Financial support from the Ministry of Science and
Technology of China (No. 2004CB518903) is greatly
appreciated.
sum of G1, G2-M,
S phase, and an aneuploid
References and notes
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