Synthesis of the naphthalene-derived inhibitors against Cdc25A dual-specificity protein phosphatase and their biological activity

Article abstract of DOI:10.1016/j.bmcl.2004.10.034

The synthesis naphthoquinones exhibited stronger activity than the pyranonaphthoquinone-type compound having moderate inhibitory activity against Cdc25A protein phosphatase. The novel naphthalene-type analogues 14 and 18 and the naphthoquinone-type analog

Full text of DOI:10.1016/j.bmcl.2004.10.034

Bioorganic & Medicinal Chemistry Letters 15 (2005) 61–65
Synthesis of the naphthalene-derived inhibitors against
Cdc25A dual-specificity protein phosphatase and
their biological activity
Akiko Shimbashi,a Ayako Tsuchiya,b Masaya Imotob and Shigeru Nishiyamaa,*
aDepartment of Chemistry, Faculty of Science and Technology, Keio University, Hiyoshi 3-14-1,
Kohoku-ku, Yokohama 223-8522, Japan
bDepartment of Biosciences and Informatics, Faculty of Science and Technology, Keio University, Hiyoshi 3-14-1,
Kohoku-ku, Yokohama 223-8522, Japan
Received 18 August 2004; revised 11 September 2004; accepted 12 October 2004
Available online 28 October 2004
Abstract—The novel naphthalene-type analogues 14 and 18 and the naphthoquinone-type analogues, 8, 9, 15, 16, 19, 21, 22, and 23–
28 have been synthesized, and their in vitro Cdc25A phosphatase-inhibitory activity was examined. In assessment of the inhibitory
activity, it was revealed that the naphthoquinone core is contributed to the activity, rather than the alkyl side chain.
Ó 2004 Elsevier Ltd. All rights reserved.
1. Introduction
ity, their detailed structure–activity relationships has
been remained unclear, and no active substance carrying
both arrest of cells in G1/S phase and selectivity for
Cdc25A, has been reported. Against such backgrounds,
we envisaged a new approach to synthetic lead mole-
cules of Cdc25A inhibitors for anticancer chemother-
apy, and initiated synthetic studies on the novel
pyranonaphthoquinone derivatives 1–4,11 isolated from
Streptomyces sp. Although they exhibited no remark-
able inhibitory activity in comparison with other Cdc25A
inhibitors, the family shares the structurally unique
characteristics that the benzoquinone core attached with
the methylamino group is located at the side of the tri-
cyclic molecule (Fig. 1).
Cell cycle progression is controlled by the cyclin-
dependent kinases (CDKs), which are positively regu-
lated by association with cyclins and negatively
regulated by binding to inhibitory subunits.1 CDKs
are also inactivated by phosphorylation with the protein
kinases wee12 and activated by dephosphorylation with
the Cdc25 phosphatases. Mammalian cells contain three
Cdc25 genes, named Cdc25A, Cdc25B, and Cdc25C.3
Cdc25A, a dual-specificity protein phosphatase, acti-
vates cyclin-dependent kinase 2 (CDK2), promoting en-
try into the S phase of the cell cycle.4 Overexpression of
Cdc25A has been shown previously to have oncogenic
potential,5 making Cdc25A inhibitors candidates of
new lead molecule for anticancer chemotherapies.6
O
OH
R
2
R
1
O
There are several inhibitors of Cdc25A protein phospha-
tase in such natural products, as dysidiolide,7 glucolipsin
A,8 indolyldihydroxyquinone,9 and menadione.10
Among them, dysidiolide is the most famous substance
exhibiting the Cdc25A inhibitory activity.7 Although
there are a number of inhibitors exhibiting potent activ-
CO H
2
MeHN
O
1: R = H, R = n-butyl
1
2
2
2: R = H, R = i-pentyl
1
3, 4 : isomers at the C-26 position
CO HO
2
S
R
R
=
1
2
N
H
Me
26
Keywords: Cell cycle regulation; Dual-specificity phosphatase; Cdc25A
phosphatase inhibitor; Naphthoquinone; Biological activity.
= i-pentyl
*
Figure 1. Structure of Cdc25A phosphatase inhibitor possessing the
pyranonaphthoquinone core.
Corresponding author. Tel./fax: +81 45 566 1717; e-mail: nisiyama@
chem.keio.ac.jp
0960-894X/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2004.10.034

62
A. Shimbashi et al. / Bioorg. Med. Chem. Lett. 15 (2005) 61–65
In a previous investigation,12a,b we succeeded total
synthesis of ( )-1, carrying the fundamental framework
of the family. It was observed that the naphthoquinone
derivatives, produced as synthetic intermediates of 1,
exhibits more effective inhibitory activity against the
Cdc25A phosphatase than those of the natural pyrano-
naphthoquinones.12b In this context, our attention was
mainly focused on biological activity of other naphtho-
quinone derivatives. We describe herein synthesis of new
naphthoquinone analogues and their structure–activity
relationship, as part of our extensive investigation of
inhibitors against Cdc25A phosphatase.
CHO
OBn OH
ref.12
a, b
MeO
Br
MeO
CO Et
2
OMe
OMe
5
6
OBn OMe
O
4
OMe
5
c
2
OMe
OMe
MeO
MeO
OMe
O
7
8
2. Synthesis of the benzyl methyl ether analogues 8 and 9
O
O
OMe
As can be seen in Scheme 1, naphthol 6 was easily ob-
tained from 5-bromoveratraldehyde 5.12 Sequential
manipulation involving protection, reduction, and fur-
ther methylation afforded the benzyl methyl ether 7.
Oxidation under DDQ conditions provided the benzyl
ether 8. Subsequent introduction of a methylamino
group gave 9 in 89% yield.13
d
OMe
MeHN
9
Scheme 1. Reagents and conditions: (a) LiAlH4, THF, rt, 61%; (b)
MeI, NaH, DMF, rt, 65%; (c) DDQ, t-BuOH, H2O–CH2Cl2, rt, 84%;
(d) MeNH2, THF, 0°C, 89%.
3. Synthesis of the 1,3-diol analogues 14, 15, and 16
Oxirane 10 was obtained in good yield from 6 by Me3SI
(Scheme 2). Isomerization and the Horner–Wadsworth–
Emmons reaction provided the a,b-unsaturated ester 11
in 57% yield, which was exposed to DIBAL to afford an
OBn OMe
a - d
e, f
O
6
MeO
OMe
10
OBn OMe
OBn OMe
g, h
O
CO Et
2
MeO
OH
MeO
OMe
OMe
11
12
OBn OMe
OH OMe
O
O
O
O
i, j
MeO
MeO
OMe
OMe
13
14
k
O
OMe
O
O
OMe
l
O
O
O
O
MeHN
MeO
O
15
16
Scheme 2. Reagents and conditions: (a) MeI, K2CO3, DMF, rt, 92%; (b) LiAlH4, THF, rt, 80%; (c) SO3ÆPy, TEA, DMSO, CH2Cl2, rt, 100%; (d)
Me3SI, NaH, DMSO, THF, 0°C, quant.; (e) ZnBr2, PhH, reflux; (f) (EtO)2P(O)CH2CO2Et, NaH, THF, À78°C, 57% in two steps; (g) DIBAL, THF,
À78°C, 88%; (h) L-(+)-DIPT, Ti(Oi-Pr)4, TBHP, MS4A, CH2Cl2, À20°C, 85%, 95% ee; (i) Red-AlÒ, THF, À78°C; (j) 2,2-dimethoxypropane, CSA,
0°C, 71% (13) and 25% (14) in two steps; (k) DDQ, H2O-1,4-dioxane, rt, 89%; (l) MeNH2, MeOH, 0°C, 38%.

A. Shimbashi et al. / Bioorg. Med. Chem. Lett. 15 (2005) 61–65
63
allyl alcohol. Sharpless asymmetric epoxidation of the
allyl alcohol was carried out with L-(+)-DIPT to give
the epoxy alcohol 12 in 85% yield (95% ee). Reductive
cleavage of the epoxide ring with Red-AlÒ, and acetal-
protection of the 1,3-diol gave 13 and 14. Immediate
oxidation of 13 afforded quinone 15 in 63% yield from
12. Finally, introduction of a methylamino group pro-
vided the methylamine 16 in 38% yield.
K2CO3 in MeOH afforded 21. Introduction of a methyl-
amino group and deacetylation provided 22 in 74% yield
from 20.
5. Synthesis of the naphthoquinone derivatives 23–28
To obtain 23–28, the synthetic route to 19–22 was
slightly modified (Scheme 4). Upon using the same
method as described in Scheme 3, deacetylation was
unsuccessful. Accordingly, the order of oxidation and
desilylation was reversed. Thus, the silyl ether 17 was re-
acted with the corresponding aldehyde, and oxidized to
naphthoquinones, which were successfully converted
into the allyl alcohols, 23, 25, and 27. Treatment of
23, 25, and 27 with MeNH2 gave the corresponding
methylamines 24, 26, and 28.
4. Synthesis of the naphthalene 18 and the naphthoqui-
nones 19, 21, 22
Compound 6 was converted into the silyl ether 17,
according to the previous synthesis of the natural prod-
uct 112 (Scheme 3). A lithiated derivative of 17 was re-
acted with valeraldehyde and successive desilylation,
afforded the allyl alcohol 18 in 80% yield. Compound
18 was oxidized under DDQ conditions to give the
naphthoquinone 19 in 79% yield. Naphthoquinones 21
and 22 were obtained from the allyl alcohol 18, as fol-
lows. Selective acetylation, oxidation to the quinone,
and selective demethylation at the C-6 position gave
20 in 39% yield from 18. Compound 20 was reacted with
6. Biological activity
The naphthalene and naphthoquinone derivatives were
submitted to assessment for in vitro inhibition of
Cdc25A phosphatase using 4-nitrophenyl phosphatase
OBn OMe
Br
ref. 12
a, b
6
MeO
OTBDMS
OMe
17
OBn OMe OH
O
O
OMe OH
c
MeO
OH
MeO
OH
OMe
18
19
d - f
O
OH OH
O
O
OH OH
g
MeO
OH
MeO
OAc
O
21
20
h, i
O
O
OH OH
MeHN
OH
22
Scheme 3. Reagents and conditions: (a) valeraldehyde, n-BuLi, THF, À78°C; (b) TBAF, THF, 0°C, 80% in two steps; (c) DDQ, t-BuOH, H2O–
CH2Cl2, rt, 79%; (d) Ac2O, Py, CH2Cl2, rt, 85%; (e) DDQ, t-BuOH, H2O–CH2Cl2, 0°C, 87%; (f) BBr3, CH2Cl2, À78°C, 53%; (g) K2CO3, MeOH, rt,
quant.; (h) MeNH2, THF, 0°C, 88%; (i) K2CO3, MeOH, rt, 84%.

64
A. Shimbashi et al. / Bioorg. Med. Chem. Lett. 15 (2005) 61–65
OBn OMe
O
OMe OH
a - c
Br
R
MeO
OTBS
MeO
d
OH
OMe
O
17
23, 25, 27
O
O
OMe OH
R
MeHN
OH
24, 26, 28
a
c yield
d yield
aldehyde
R
propionaldehyde
hexaldehyde
61%
Et
24%
47%
16%
23
25
27
24
26
28
n-pentyl
Ph
54%
75%
benzaldehyde
Scheme 4. Reagents and conditions: (a) aldehyde, n-BuLi, THF, À78°C; (b) DDQ, t-BuOH, H2O–CH2Cl2, 0°C; (c) TBAF, AcOH, THF, rt;
(d) MeNH2, THF, 0°C.
(pNPP) as a substrate. As shown in Table 1, the low
activities of naphthalenes 14 and 18 indicated the naph-
thoquinone structure has a positive effect on inhibition
of the Cdc25A phosphatase. The naphthoquinones pos-
sessing OMe groups at the C-2 position (8, 15, 19, 21,
23, 25, and 27) showed stronger Cdc25A inhibitory
activity than the methylamino derivatives (9, 16, 22,
24, 26, and 28) as well as ( )-1. In preceding investiga-
tions of Cdc25A phosphatase inhibition, hydrophilic
subunits, such as a carboxylic acid, were reported to
serve as a surrogate phosphatase. A hydrophobic sub-
structure, such as a long side chain, might occupy a
hydrophobic binding pocket, when the molecule is
bound to Cdc25A.14 However, our compounds without
a carboxylic acid and a long alkyl side chain (8, 15),
showed effective Cdc25A phosphatase-inhibitory activ-
ity.15 Furthermore, length of alkyl side chain (19, 23,
25), and conversion of alkyl side chain into phenyl
group (27) provided no serious effect on the inhibitory
activity. In addition, protection at C-6 as a methyl ether
exhibited no serious activity difference (19, 21), which
indicated little contribution of the hydrogen bond be-
tween the C-4 carbonyl oxygen and the C-6 phenol
group.
In conclusion, the naphthalene and naphthoquinone
derivatives were synthesized and their inhibitory activity
against Cdc25A phosphatase was evaluated. It was de-
tected that the naphthoquinone core with a methoxy
group, instead of a methylamino group as in natural
pyranonaphthoquinones 1–4, is an important factor to
show high activity of Cdc25A phosphatase inhibition,
and the long alkyl side chain might not be required to
inhibit the phosphatase, contrary to the previously re-
ported data.
Table 1. Inhibitory activity against Cdc25A protein phosphatase
(pNPP)a
Compound
IC50 (lg/mL)
Compound
IC50 (lg/mL)
8
0.4
>10
>10
0.4
22
23
>10
1.86
>10
1.6
9
14
15
16
18
19
21
24
25
>10
>10
1.77
1.51
0.005
26
27
>10
2.74
>10
>10
Further investigation of the structure–activity relation-
ship and selectivity for Cdc25A is in progress.
28
( )-1
b
Na3VO4
Acknowledgements
a Cdc25A assay in vitro: The activity of the GST-cdc25A was measured
in a 96-well microtiter plate using pNPP (sigma) as a substrate.
Approximately 30lg of purified GST-cdc25A was preincubated at
37°C for 15min in reaction buffer containing of 50mM Tris–HCl
(pH7.5) and various concentrations of inhibitors in 1% MeOH, and
then reaction was started by adding of 5mM pNPP and incubated at
37°C for 60min. Absorbance at 405nm was measured.
b Positive control.
This work was supported by Grant-in-Aid for the 21st
Century COE program ꢀKeio Life Conjugated Chemis-
try,ꢁ as well as Scientific Research C from the Ministry
of Education, Culture, Sports, Science, and Technology,
Japan. A.S. was financially supported by the same COE
program.

A. Shimbashi et al. / Bioorg. Med. Chem. Lett. 15 (2005) 61–65
65
10. (a) Juan, C.-C.; Wu, F.-Y. H. Biochem. Biophys. Res.
Commun. 1993, 190, 907–911; (b) Ham, S. W.; Park, H. J.;
Lim, D. H. Bioorg. Chem. 1997, 25, 33–36; (c) Nishikawa,
Y.; Carr, B. J.; Wang, M.; Kar, S.; Finn, F.; Dowd, P.;
Zheng, Z. Z.; Kerns, J.; Naganathan, S. J. Biol. Chem.
1995, 270, 28304–28310; (d) Ni, R.; Nishikawa, Y.;
Carr, B. I. J. Biol. Chem. 1998, 273, 9906–9911;
(e) Ham, S. W.; Park, J.; Lee, S.-J.; Kim, W.; Kang, K.;
Choi, K. H. Bioorg. Med. Chem. Lett. 1998, 8, 2507–
2510.
References and notes
1. (a) Strausfeld, U.; Labbe, J. C.; Fesquet, D.; Cavadore, J.
C.; Picard, A.; Sadhu, K.; Russell, P.; Doree, M. Nature
1999, 351, 242–245; (b) Gautier, J.; Solomon, M. J.;
Booher, R. N.; Bazan, J. F.; Kirschner, M. W. Cell 1999,
67, 197–211; (c) Millar, J. B. A.; McGowan, C. H.;
Lenaers, G.; Jones, R.; Russell, P. EMBO J. 1991, 10,
4301–4309; (d) Nilsso, I.; Hoffmann, I. Prog. Cell Cycle
Res. 2000, 4, 107–114.
11. Kulanthaivel, P.; Perun, T. J., Jr.; Belvo, M. D.; Strobel,
R. J.; Paul, D. C.; Williams, D. C. J. Antibiot. 1999, 52,
256–262, The activity of 1:15.5% inhibition at 10lM
concentration.
12. (a) Shimbashi, A.; Ishikawa, Y.; Nishiyama, S. Tetrahe-
dron Lett. 2004, 45, 939–941; (b) Shimbashi, A.; Tsuchiya,
A.; Imoto, M.; Nishiyama, S. Bull. Chem. Soc. Jpn. 2004,
77, 1925–1930; (c) Iinuma, M.; Tanaka, T.; Matsuura, S.
Chem. Pharm. Bull. 1984, 32, 2296–2300.
2. (a) Russell, P.; Nurse, P. Cell 1987, 49, 559–567; (b)
Parker, L.; Atheron-Fessler, S.; Piwnica-Worms, H. Proc.
Natl. Acad. Sci. U.S.A. 1992, 89, 2917–2921.
3. (a) Sadhu, K.; Reed, S. I.; Richardson, H.; Russell, P.
Proc. Natl. Acad. Sci. U.S.A. 1990, 87, 5130–5143; (b)
Galaktionov, K.; Beach, D. Cell 1991, 67, 1181–1194; (c)
Jinno, S.; Suto, K.; Nagata, A.; Igarashi, M.; Kanaoka,
Y.; Nojima, H.; Okayama, H. EMBO J. 1994, 13, 1549–
1556.
13. Tao, X. L.; Cheng, J.-F.; Nishiyama, S.; Yamamura, S.
Tetrahedron 1994, 50, 2017–2028.
4. (a) Ref. 3c; (b) Hoffmann, I.; Draetta, G.; Karsenti, E.
EMBO J. 1994, 13, 4302–4310.
14. (a) Baratte, B.; Meijier, L.; Galaktionov, K.; Beach, D.
Anticancer Res. 1992, 12, 873–880; (b) Moran, E. J.;
Sarshar, S.; Cargill, J. F.; Shahbaz, M. M.; Lio, A.; Mjalli,
A. M. M.; Armstrong, R. W. J. Am. Chem. Soc. 1995, 117,
10787–10788; (c) Cao, X.; Moran, E. J.; Siev, D.; Lio, A.;
Ohashi, C.; Mjalli, A. M. M. Bioorg. Med. Chem. Lett.
1995, 5, 2953–2958; (d) Rice, R. L.; Rusnak, J. M.;
Yokokawa, F.; Yokokawa, S.; Messner, D.; Boynton, A.
L.; Wipf, P.; Lazo, J. S. Biochemistry 1997, 36, 15965–
15974; (e) Bergnes, G.; Gilliam, C. L.; Boisclair, M. D.;
Blanchard, J. L.; Blake, K. V.; Epstein, D. M.; Pal, K.
Bioorg. Med. Chem. Lett. 1999, 332, 19–24; (f) Wipf, P.;
Aslan, D. C.; Luci, D. K.; Southwick, E. C. Biotech.
Bioeng. 2000, 71, 58–70.
15. Other Cdc25A inhibitors: Indolyldihydroxyquinone ana-
logue was a potent deactivator (IC50 = 1.0lM, Ref. 9).
Dysidiolide derivative effectively arrested cells in G1/S
phase (IC50 = 13lM, Ref. 7). Glucolipsin A analogue
(IC50 = 1.6lM, Ref. 8) and menadione derivative
(IC50 = 3.8lM, Ref. 10) showed good selectivity for
Cdc25A. The IC50 values (lM) of 8 and 15 were 1.5 and
1.2, and their effective selectivity against Cdc25A was
determined by comparison with PTP1B (IC50 = 20 lM for
8, 13lM for 15).
5. (a) Galaktionov, K.; Lee, A. K.; Eckstein, J.; Draetta, G.;
Meckler, J.; Loda, M.; Beach, D. Science 1995, 269, 1575–
1577; (b) Galaktionov, K.; Chen, X.; Beach, D. Nature
1996, 382, 511–517; (c) Gasparotto, D.; Maestro, R.;
Piccinin, S.; Vukosavljievic, T.; Barzan, L.; Sulfaro, S.;
Boiocchi, M. Cancer Res. 1997, 57, 2366–2368; (d) Wu, W.
G.; Fan, Y. H.; Kemp, B. L.; Walsh, G.; Mao, L. Cancer
Res. 1998, 58, 4082–4085; (e) Dixon, D.; Moyana, T.;
King, M. J. Exp. Cell Res. 1998, 240, 236–243; (f) Cangi,
M. G.; Cukor, B.; Soung, P.; Signoretti, S.; Moreira, G.;
Ranashinge, M.; Cady, B.; Pagano, M.; Loda, M. J. Clin.
Invest. 2000, 106, 753–761.
6. (a) Draetta, G.; Eckstein, J. Biochem. Biophys. Acta 1997,
1332, M53; (b) Carnero, A. Br. J. Cancer 2002, 87, 129–
133; (c) Lyon, M. A.; Ducruet, A. P.; Wipf, P.; Lazo, J. S.
Nat. Rev. Drug Discovery 2002, 1, 961–976.
7. Study on structure–activity relationship of dysidiolide:
Takahashi, M.; Dodo, K.; Sugimoto, Y.; Aoyagi, Y.;
Yamada, Y.; Hashimoto, Y.; Shirai, R. Bioorg. Med.
Chem. Lett. 2000, 10, 2571–2574.
8. Furstner, A.; Ruiz-Caro, J.; Prinz, H.; Waldmann, H. J.
¨
Org. Chem. 2004, 69, 459–467.
9. Sohn, J.; Kiburz, B.; Li, Z.; Deng, L.; Safi, A.; Pirrung, M.
C.; Rudolph, J. J. Med. Chem. 2003, 46, 2580–2588.