Synthesis of miltirone analogues as inhibitors of Cdc25 phosphatases

Article abstract of DOI:10.1016/j.bmcl.2005.12.080

Miltirone analogues were synthesized and evaluated for inhibitory activity against Cdc25 and PTP1B. Most of the compounds demonstrated potent Cdc25 inhibitory activity, and several exhibited higher selectivity for Cdc25 than for PTP1B. In a cytotoxic assay, most of the compounds displayed cytotoxicity against the tumor cell lines A549 and HCT-116, producing IC₅₀ values in the micromolar range.

Full text of DOI:10.1016/j.bmcl.2005.12.080

Bioorganic & Medicinal Chemistry Letters 16 (2006) 1905–1908
Synthesis of miltirone analogues as inhibitors
of Cdc25 phosphatases
Weigang Huang,b Jingya Li,b Wei Zhang,b Yueyang Zhou,b Chuanming Xie,b
Yu Luo,b Yunfei Li,b Jingli Wang,a Jia Lib,* and Wei Lua,*
aDepartment of Chemistry, East China Normal University, Shanghai 200062, China
bThe National Center for Drugs Screening, Shanghai Institute of Materia Medica, SIBS,
Graduate School of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai 201203, China
Received 31 October 2005; revised 3 December 2005; accepted 24 December 2005
Available online 24 January 2006
Abstract—Miltirone analogues were synthesized and evaluated for inhibitory activity against Cdc25 and PTP1B. Most of the
compounds demonstrated potent Cdc25 inhibitory activity, and several exhibited higher selectivity for Cdc25 than for PTP1B. In
a cytotoxic assay, most of the compounds displayed cytotoxicity against the tumor cell lines A549 and HCT-116, producing IC50
values in the micromolar range.
Ó 2006 Elsevier Ltd. All rights reserved.
The loss of cell cycle control leading to deregulated cell
proliferation is one of the hallmarks of cancer. The
Cdc25 phosphatases function as key regulators of the cell
cycle during normal eukaryotic cell division and as medi-
ators of the checkpoint response in cells with DNA dam-
age.1 Three Cdc25 homologues exist in humans: Cdc25A,
Cdc25B, and Cdc25C.1–4 Overexpression of Cdc25A and
B occurs in various forms of cancer and is strongly asso-
ciated with tumor aggressiveness and poor prognosis,5–8
making Cdc25 an attractive drug target for cancer thera-
py. Over the last few years, some small molecule Cdc25
inhibitors have been described.9–34
corresponding aniline to the solution of miltirone in
methanol. Treatment of miltirone with the correspond-
ing thiophenol in CHCl3/MeOH produced sulfides
2a–5a in 50–80% yield. Compound 10a was obtained
by the reaction of miltirone with benzene in acetic acid,
with Pd(OAc)2 as the catalyst under reflux conditions.
In our previous study, some tanshinones without the A
ring displayed more potent antitumor activities.35 Here,
we prepared a series of analogues without the A ring.
We started the synthesis of the important intermediate
16, also a miltirone analogue, from compound 11
(Scheme 2), which we prepared as reported previously.38
Reduction of 11 with NaBH4 produced the alcohol 12,
which was directly treated with 20% H2SO4 to give 13,
a colorless oil, in 77% yield (from 11). Dehydrogenation
of 13 with DDQ in toluene afforded 2-isopropyl-3-meth-
oxy naphthalene 14 in 100% yield. Treatment of 14 with
boron tribromide and oxidation with Dess–Martin peri-
odinane gave 16, a red solid. Compounds 2b–10b were
prepared according to the procedure described above
for compounds 2a–10a.
In our program of extensive screening for inhibitors of
Cdc25, we have found that some tanshinones demon-
strate inhibitory activity for Cdc2535 and cytotoxicity
against a number of cultured human tumor cell lines.36
We chose miltirone, one of these tanshinones,37 as the
pharmacophore from which we synthesized 19 ana-
logues and evaluated their antitumor activity as Cdc25
phosphatase inhibitors.
As shown in Scheme 1, the N-substituted series ana-
logues, 6a–9a were prepared by addition of the
Miltirone and its analogues were evaluated for their
antitumor activities39,40 and the results are summarized
in Table 1. Miltirone demonstrated no activity for
Cdc25A and Cdc25B. All of its analogues, except 7a,
were potent inhibitors of Cdc25 phosphatases in the
micromolar range. We speculated that substitution of
the 1-position of miltirone or the lack of the A ring
Keywords: Miltirone; Cdc25 phosphatases; Cancer.
*
Corresponding authors. Tel./fax: +86 21 62602475 (W.L.); fax: +86
21 50801552 (J.L.); e-mail addresses: jli@mail.shcnc.ac.cn; wlu@
chem.ecnu.edu.cn
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.12.080

1906
W. Huang et al. / Bioorg. Med. Chem. Lett. 16 (2006) 1905–1908
O
O
S
a
O
O
Miltrone 1
c
b
R1
2a, 3a, 4a, 5a
O
O
O
O
R1= H,
2a
Br,
F,
3a
4a
NH
NHAc, 5a
R2= H,
6a
7a
8a
9a
R2
6a, 7a, 8a, 9a
10a
NO2,
CH3,
OMe,
Scheme 1. Reagents: (a) different thiophenol, MeOH/CHCl3, 50–80%; (b) different aniline, MeOH, 50–60%; (c) benzene, Pd(OAc)2, HOAc, 35%.
O
OH
b
OMe
a
c
OMe
OMe
11
12
13
O
OMe
OH
d
e
O
16
14
15
f
g
h
O
S
O
O
O
R1= H,
2b
O
O
Br,
F,
3b
4b
NHAc, 5b
NH
R2= H,
6b
7b
8b
9b
R1
NO2,
CH3,
OMe,
R2
2b, 3b, 4b, 5b
10b
6b, 7b, 8b, 9b
Scheme 2. Reagents: (a) NaBH4, MeOH; (b) aq H2SO4, reflux, 76.77% from 11; (c) DDQ, toluene; (d) BBr3, CH2Cl2; (e) Dess–Martin periodinane,
CH2Cl2, 42% two steps from 14; (f) different thiophenol, MeOH/CHCl3, 50–80%; (g) different aniline, MeOH, 50–60%; (h) benzene, Pd(OAc)2,
HOAc, 75%.
increases the inhibitory activity for Cdc25. However,
contrary to our expectation, compounds 2b–10b, which
lack the A ring, did not display more potent Cdc25
inhibitory activity than 2a–10a, suggesting that the A
ring is not necessary for the Cdc25 inhibitory activity.
To investigate the relationship between the electronic ef-
fect on the aromatic ring and antitumor activities, we
introduced different substituents. However, we observed
no obvious relationship. To study the specificity for
Cdc25, we also tested the inhibition of PTP1B, another
protein-tyrosine phosphatase. Compounds 16, 6a, 6b,
7b, 8a, 8b, 9a, 9b, and 10b, which inhibit Cdc25 phos-
phatases in the micromolar range, demonstrated no
activity for PTP1B, indicating the high selectivity of
these compounds. We note that the rest of these com-
pounds inhibited PTP1B, producing IC50 values of
4.11–13.66 lM.
We evaluated the cytotoxicity of all compounds using
the A549 and HCT-116 cell lines. Miltirone was cytotox-
ic against the A549 and HCT-116 cell lines, producing
IC50 values of 24.59 0.42 and 12.39 0.75 lM, respec-
tively, which are consistent with previous results.36
Compounds 4b and 10b displayed similar or inferior
cytotoxicity to that produced by miltirone. The rest of
the compounds displayed more potent cytotoxicity.
Our findings suggest that the A ring is also unnecessary
for their cytotoxicity.

W. Huang et al. / Bioorg. Med. Chem. Lett. 16 (2006) 1905–1908
1907
Table 1. Inhibitory activity for Cdc25A, Cdc25B, and PTP1B phosphotases and cytotoxicity against A549 and HCT-116 cell lines
Compounds
Cdc25A
IC50 SDa (lM)
Cdc25B
IC50 SD (lM)
PTP1B
IC50 SD (lM)
A549
IC50 SD (lM)
HCT-116
IC50 SD (lM)
Miltirone 5
NAb
NA
NA
NA
24.59 0.42
4.14 0.18
26.64 5.34
7.59 0.11
5.07 0.04
5.93 0.18
1.25 0.54
20.42 0.97
15.43 0.87
7.36 0.86
19.06 0.51
25.01 0.56
5.34 0.05
6.59 0.28
4.25 2.45
8.20 3.19
9.19 4.53
87.01 20.80
6.24 0.17
24.71 0.46
12.39 0.75
4.00 0.03
5.75 0.03
8.03 0.07
4.94 0.07
5.85 0.15
5.71 0.13
20.78 0.55
5.18 0.04
6.32 0.08
6.25 0.01
8.22 0.15
7.10 1.24
2.39 0.03
24.94 1.10
6.16 2.03
5.51 0.09
4.04 1.23
6.28 0.12
29.60 4.36
16
2a
2b
3a
3b
4a
4b
5a
5b
6a
6b
7a
7b
8a
8b
9a
9b
10a
10b
11.18 1.38
3.97 0.63
5.86 1.79
20.31 6.41
5.43 1.11
22.06 9.03
12.75 1.32
5.96 0.73
21.87 0.28
8.81 0.38
10.88 0.96
NA
24.10 1.37
5.07 0.84
10.86 1.95
3.78 1.38
3.63 0.70
2.58 0.66
7.55 2.14
4.85 1.88
13.82 1.27
3.99 0.08
8.71 0.79
NA
10.40 2.086
13.66 0.50
7.19 0.75
7.57 1.35
4.55 0.14
7.22 1.13
9.05 1.01
7.63 0.54
27.16 4.67
NA
NA
19.08 5.33
NA
6.87 1.11
27.58 6.12
7.06 0.29
11.82 1.69
4.90 0.23
3.96 0.63
7.83 0.59
5.33 0.33
7.54 2.43
4.53 0.24
4.09 0.59
4.63 0.13
3.16 0.22
3.23 0.31
19.59 3.45
NA
NA
4.11 0.40
NA
a The IC50 values are means of three distinct experiments.
b NA: not active, IC50 > 20 lg/mL.
8. Hernandez, S.; Bessa, X.; Bea, S.; Hernmandez, L.; Nadal,
A.; Mallofre, C.; Muntane, J.; Castells, A.; Fernandez, P.
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In conclusion, we synthesized analogues of miltirone
and evaluated their antitumor activities as Cdc25 inhib-
itors. As expected for Cdc25 inhibitors, most of these
compounds were potent Cdc25 inhibitors and exhibited
cytotoxic activity against A549 and HCT-116 cell lines
in the micromolar range. Some compounds exhibited
higher selectivity for Cdc25 than for PTP1B. We con-
clude that these miltirone analogues might be attractive
lead molecules for developing drugs against the Cdc25
phosphatase.
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Acknowledgment
Financial supports from the National Natural Science
Foundation of China (30572232) and Shanghai Science
and Technology Mission (054319902) are gratefully
acknowledged.
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carried out in
.
The typical inhibition assay for cdc25s was
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a
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200–300 nM GST–cdc25A or 40–80 nM GST–cdc25B.
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MOPs, pH 6.5, 2 mM DTT, 1 mM EDTA, 2 mM
pNPP, 2% DMSO, and 30–40 nM GST–PTP1B. (a)
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concentration compounds for subsequent 72 h incuba-
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