K. N. Kim et al. / Bioorg. Med. Chem. Lett. 15 (2005) 77–79
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Barry, D. W.; Broder, S. Proc. Natl. Acad. Sci. U.S.A.
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Jones, A. S.; Walker, R. T. Proc. Natl. Acad. Sci. U.S.A.
1
1
20
00
1
8
9
1
8
6
4
2
0
0
0
0
0
1
979, 76, 2947.
10. DeClercq, E.; DeGreef, H.; Wildiers, J.; De Jonge, G.;
Drochmans, A.; Descamps, J.; De somer, P. Br. Med. J.
1980, 281, 1178.
1. Kauhman, H. E.; Heidelberger, C. Science 1964, 145,
585.
1
0
5
10
15
20
Concentration (µM)
12. Heidelberger, C.; Griesbach, L.; Cruz, O.; Schnitzer, R. J.;
Grunberg, E. Proc. Soc. Exp. Biol. Med. 1958, 97, 470.
13. Mukherjee, K. L.; Boohar, J.; Wentand, D.; Ans-fied, F.
J.; Heidelberger, C. Cancer Res. 1963, 23, 49.
14. Anderson, A. S.; Hwang, J. T.; Greenberg, M. M. J. Org.
Chem. 2000, 65, 4648.
Figure 2. Cell viability assay. 4a (5–20lM) was applied to the cell and
incubated for up to 72h. The cells were then stained with trypan blue
and counted by hemocytometer.
1
5. Synthetic procedure for 4a; (compound Ac-4a); Com-
pound 3 (370mg, 0.959mmol) was dissolved in DMF
of HeLa cell with IC50 value of 15lM, suggesting that
the increased hydrophobic property at 5-methyl group
of pyrimidine may contribute to enhance biological
activity of the compound. From this result, we speculate
that the position of bromide in phenoxy ring and hydro-
phobic property of the substitution can affect the biolog-
ical activity of analogues. In addition, it would be
interesting to isolate the specific binding protein for
(
5mL, dry) containing K CO (132.5mg, 0.959mmol), 3-
2 3
bromo-phenol (112.8lL, 0.913mmol) at 0ꢁC. The reac-
tion mixture was preceded at room temperature for 5h.
The reaction mixture was diluted with ethyl acetate
(20mL), washed with brine, dried (MgSO ), and concen-
4
trated in vacuo. The crude product was purified by flash
column chromatography (1.2:1 = n-hexane–EtOAc) to
give Ac-4a as a solid in 21% yield. (Compound 4a);
Compound Ac-4a was dissolved in CHCl2 (3mL) and
MeOH (2mL) containing NaOMe (12mg, 0.153mmol) at
4
the effect on the viability of the tumor cells using the try-
a. The selected candidate, 4a, was further studied for
1
9,20
pan blue exclusion method.
0
ꢁC. After 24h at room temperature, the reaction mixture
was neutralized at 0ꢁC using 10% HOAc in MeOH and
concentrated in vacuo. The crude product was directly
At the end of the assay the relative cell viability was cal-
culated by comparison with total cells counted. Com-
pound 4a showed no cytotoxicity against HeLa cells
even at the concentration ranges in which the prolifera-
tion of the cells was inhibited by the compound (Fig. 2).
purified
(20:1 = CH Cl –MeOH) to give 4a as a solid in 97% yield.
by
flash
column
chromatography
2
2
1
16. Analytical data for Ac-4a; H NMR (200MHz, CDCl ) d
3
7.69 (s, 1H), 7.11 (m, 3H), 6.80 (m, 1H), 6.24 (dd,
J = 5.8Hz), 4.79 (q, 2H, J = 1.0Hz), 4.34–4.26 (m, 4H),
2
HRMS (FAB, M+Na) Calcd C H O N Br: 519.0379,
.61–1.99 (m, 2H), 1.22 (dd, 6H, J = 3.6Hz, J = 14Hz).
In conclusion, we synthesized a new thymidine analogue
with increased hydrophobic property at 5-methyl group
of pyrimidine and this compound will be a promising
candidate for development of thymidine based anti-
tumor agent. Target identification using biotinylated
2
0
21
8
1
2
found = 519.0381: 4a; mp 74–76ꢁC. H NMR (200
MHz, DMSO-d ) d 8.54 (s, 1H), 7.64–7.60 (m, 2H), 7.52
d, 1H, J = 6.0Hz), 7.38 (dd, 1H, J = 1.0Hz, J = 4.0Hz),
6
(
4
.17 (q, 1H, J = 2.0Hz), 3.95 (s, 1H), 2.9–2.7 (m, 2H).
4a will be our next challenge for deciphering its mode
of action in anti-proliferative activity of tumor cells.
HRMS (FAB, M+Na) Calcd C16
found = 435.167.
17 6 2
H O N Br: 435.017,
1
1
7. Shim, J. S.; Kim, D. H.; Jung, H. J.; Kim, J. H.; Ahn, J.
W.; Yoo, J. S.; Rho, J. R.; Shin, J. H.; Kwon, H. J. Bioorg.
Med. Chem. 2002, 10, 2987.
Acknowledgements
8. HeLa (cervical carcinoma) cells were maintained in
DMEM and were grown at 37ꢁC in a humidified
This study was supported by the National Research Lab-
oratory Grant from the Ministry of Science and Techno-
logy, Republic of Korea and the Brain Korea 21 Project.
2
atmosphere of 5% CO . Cell proliferation was measured
by using MTT assay. Briefly, HeLa cells were inoculated
3
at a density of 4 · 10 cells/well in 96-well plates. Various
doses of thymidine analogues were added to each well and
inoculated for 72h. MTT (50lM, 2mg/mL) was added
and the plate was incubated for 4h. The absorbance of
MTT-formazan was measured using a 540nm filter-
equipped mircroplate reader (Bio-Tek instruments, Inc.,
Winooski, VT).
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at a density of 5 · 10 cells/well in a 24-well plate. The cells
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trypan blue and counted by hemocytometer.
1
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