Tetronic Acid Library
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 20 3221
NaOH (1.0 mL), and the mixture was stirred at 23 °C for 69
h. The mixture was acidified with 1 N HCl to pH 1-2 and
extracted with AcOEt. The organic layer was washed with
brine, dried (Na2SO4), and concentrated. The residue was
purified by silica gel column chromatography (CHCl3/MeOH,
10:1) and then treated with 0.5 N HCl to give 34 (14 mg, 100%)
as a colorless solid.
Refer en ces
(1) (a) Fischer, E. H.; Charbonneau, H.; Tonks, N. K. Protein
Tyrosine Phosphatases: A Diverse Family of Intracellular and
Transmembrane Enzymes. Science 1991, 253, 401-406. (b)
Stone, R. L.; Dixon, J . E. Protein-tyrosine Phosphatases. J . Biol.
Chem. 1994, 269, 31323-31326. (c) Sun, H.; Tonks, N. K. The
coordinated action of protein tyrosine phosphatases and kinases
in cell signaling. Trends Biochem. Sci. 1994, 19, 480-485 and
references therein.
(2) (a) Burke, T. R., J r.; Zhang, Z.-Y. Protein-Tyrosine Phos-
phatases: Structure, Mechanism, and Inhibitor Discovery.
Biopolymer 1998, 47, 225-241. (b) Widlanski, T. S.; Myers, J .
K.; Stec, B.; Holtz, K. M.; Kantrowitz, E. R. The road less
travelled: taming phosphatases. Chem. Biol. 1997, 4, 489-492
and references therein.
(3) Last year, non-peptide-like reversible inhibitors of PTP1B were
reported. (a) Andersen, H. S.; Iversen, L. F.; J eppesen, C. B.;
Branner, S.; Norris, K.; Rasmussen, H. B.; Møller, K. B.; Møllere,
N. P. H. 2-(Oxalylamino)-benzoic Acid Is a General, Competitive
Inhibitor of Protein-tyrosine Phosphatases. J . Biol. Chem. 2000,
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Benzothiophene Biphenyls as Inhibitors of Protein Tyrosine
Phosphatase 1B with Antihyperglycemic Properties. J . Med.
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Syn th esis of (R)-3-Hexadecan oyl-5-m eth yltetr on ic Acid
(35). To a solution of 6a (400 mg, 1.08 mmol) and methyl (R)-
lactate (0.10 mL, 1.08 mmol) in THF (7.0 mL) was added silver
trifluoroacetate (240 mg, 1.08 mmol), and the mixture was
stirred overnight at 23 °C while shielded from light. After
addition of silver trifluoroacetate (100 mg, 0.45 mmol), the
mixture was stirred for 2 days at 23 °C. The mixture was
quenched by the addition of water, extracted with CH2Cl2,
dried (Na2SO4), and concentrated. The residue was purified
by silica gel column chromatography (hexane/AcOEt, 10:1) to
give methyl (R)-2-(3-oxo-octadecanoyl)oxypropionate (369 mg,
89%, a 2:1 mixture of keto and enol forms) as a colorless oil.
To a solution of this ester (224 mg, 0.58 mmol) in THF (1.0
mL) was added TBAF (1 M THF solution, 0.87 mL, 0.87 mmol),
and the mixture was stirred at 23 °C for 1 day. After addition
of TBAF (0.87 mL, 0.87 mmol), the mixture was further stirred
overnight at 23 °C. The reaction was quenched by the addition
of 1 N aqueous HCl (100 mL), and the mixture was then
extracted with CH2Cl2. The combined organic layers were
washed with brine, dried (Na2SO4), and concentrated. The
residue was purified by silica gel column chromatography
(CHCl3/MeOH, 30:1) and thin layer silica gel chromatography
and then was treated with 0.5 N aqueous HCl to give 35 (166
mg, 81%) as a pale-yellow solid.
Syn th esis of 3-Hexa d eca n oyltetr on ic Acid (36). Similar
to the synthesis of 35, compound 36 was synthesized from 6a
and methyl glycolate (24%, pale-yellow solid; see Supporting
Information).
VHR Assa ys. VHR was overexpressed in E. coli and
purified. Assays were performed using pNPP as a substrate
at 37 °C in 50 mM succinate, 1 mM EDTA, 150 mM NaCl, pH
6.0, in the presence or absence of the compound. Immediately
after addition of purified VHR (0.6 µM) and pNPP (10 mM),
the reaction mixture (200 µL) was incubated at 37 °C for 3
min. The phosphatase reaction was then terminated by the
addition of 1 N NaOH (1 mL). The phosphatase activities were
measured as absorbance changes at 405 nm.
Cd c25B Assa ys. Cdc25B (aa 377-565) was overexpressed
as His6-tag-fusion protein in E. coli and purified. Assays were
performed using OMFP as a substrate at 37 °C in 100 mM
Tris, 1 mM DTT, 40 mM NaCl, 20% glycerol, pH 8.2, in the
presence or absence of the compound. Immediately after the
addition of purified cdc25B (0.04 µM) and OMFP (200 µM),
the reaction mixture (700 µL) was incubated at 37 °C for 20
min. The phosphatase reaction was then terminated by the
addition of 1 M MES (pH 5.5)-6 M guanidine (150 µL). The
phosphatase activities were measured as absorbance changes
at 477 nm.
Molecu la r Mod elin g. The initial structure generated by
docking 3-acetyltetronic acid anion to VHR manually was
optimized by molecular mechanics calculation using the
computer program Hyper-Chem (Hypercube Inc., Canada).
Further analysis of the 3D structure of the VHR-RK682 (1)
complex using molecular dynamics simulation will be pub-
lished elsewhere.
(4) Horiguchi, T.; Nishi, K.; Hakoda, S.; Tanida, S.; Nagata, A.;
Okayama, H. Dnacin A1 and dnacin B1 are antitumor antibiotics
that inhibit cdc25B phosphatase activity. Biochem. Pharmacol.
1994, 48, 2139-2141.
(5) (a) Gunasekera, S. P.; McCathy, P. J .; Kelly-Borges, M.; Lob-
kovsky, E.; Clardy, J . Dysidiolide: A Novel Protein Phosphatase
Inhibitor from the Caribbean Sponge Dysidea etheria de Lauben-
gels. J . Am. Chem. Soc. 1996, 118, 8759-8760. (b) Blanchard,
J . L.; Epstein, D. M.; Boisclair, M. D.; Rudolph, J .; Pal, K.
Dysidiolide and Related γ-Hydroxy Butenolide Compounds as
Inhibitors of the Protein Tyrosine Phosphatase, CDC25. Bioorg.
Med. Chem. Lett. 1999, 9, 2537-2538. (c) Takahashi, M.; Dodo,
K.; Sugimoto, Y.; Aoyagi, Y.; Yamada, Y.; Hashimoto, Y.; Shirai,
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Structure-Activity Relationship. Bioorg. Med. Chem. Lett. 2000,
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Messner, D. J .; Boynton, A. L.; Wipf, P.; Lazo, J . S. A Targeted
Library of Small-Molecule, Tyrosine, and Dual-Specificity Phos-
phatase Inhibitors Derived from Rational Core Design and
Random Side Chain Variation. Biochemistry 1997, 15965-
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Menadione as an Inhibitor of the cdc25 Phosphatase. Bioorg.
Chem. 1997, 25, 33-36. (c) Peng, H.; Zalkow, L. H.; Abraham,
R. T.; Powis, G. Novel CDC25A Phosphatase Inhibitors from
Pyrolysis of 3-R-Azido-B-homo-6-oxa-4-cholesten-7-one on Silica
Gel. J . Med. Chem. 1998, 41, 4677-4680. (d) Bergnes, G.;
Gilliam, C. L.; Boisclair, M. D.; Blanchard, J . L.; Blake, K. V.;
Epstein, D. M.; Pal, K. Generation of an Ugi Library of
Phosphate Mimic-Containing Compounds and Identification of
Novel Dual Specific Phosphatase Inhibitors. Bioorg. Med. Chem.
Lett. 1999, 9, 2849-2854. (e) El-Subbagh, H. I.; Abadi, A. H.;
Al-Khawad, I. E.; Al-Rashood, K. A. Synthesis and Antitumor
Activity of Some New Substituted Quinolin-4-one and 1,7-
Naphthyridin-4-one Analogs. Arch. Pharm. Pharm. Med. Chem.
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Lett. 2000, 10, 615-617. (g) Wipf, P.; Aslan, D. C.; Luci, D. K.;
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A.; Mjalli, A. M. M.; Armstrong, R. W. Radio Frequency Tag
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(10) (a) Ishibashi, T.; Bottaro, D. P.; Chan, A.; Miki, T.; Aaronson,
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(b) Aroca, P.; Bottaro, D. P.; Ishibashi, T.; Aaronson, S. A.;
Ack n ow led gm en t . We thank Professor Hiroto
Okayama (University of Tokyo) for the generous gift of
cdc25 cDNA. This work was supported in part by The
Hayashi Memorial Foundation for Female Natural
Scientists, The Kato Memorial Bioscience Foundation,
and The Mochida Memorial Foundation for Medical and
Pharmaceutical Research.
Su p p or t in g In for m a t ion Ava ila b le: Spectral data for
compounds 2-36. This material is available free of charge via
the Internet at http://pubs.acs.org.