Design and synthesis of non-hydroxamate histone deacetylase inhibitors: Identification of a selective histone acetylating agent

Article abstract of DOI:10.1016/j.bmc.2005.04.002

A series of suberoylanilide hydroxamic acid (SAHA)-based non-hydroxamates was designed, synthesized, and evaluated for their histone deacetylase (HDAC) inhibitory activity. Among these, methyl sulfoxide 15 inhibited HDACs in enzyme assays and caused hyperacetylation of histone H4 while not inducing the accumulation of acetylated α-tubulin in HCT116 cells.

Full text of DOI:10.1016/j.bmc.2005.04.002

Bioorganic & Medicinal Chemistry 13 (2005) 4332–4342
Design and synthesis of non-hydroxamate histone deacetylase
inhibitors: identification of a selective histone acetylating agent
Takayoshi Suzuki,* Azusa Matsuura, Akiyasu Kouketsu, Shinya Hisakawa,
Hidehiko Nakagawa and Naoki Miyata*
Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabe-dori, Mizuho-ku, Nagoya, Aichi 467-8603, Japan
Received 17 March 2005; revised 4 April 2005; accepted 4 April 2005
Available online 4 May 2005
Abstract—A series of suberoylanilide hydroxamic acid (SAHA)-based non-hydroxamates was designed, synthesized, and evaluated
for their histone deacetylase (HDAC) inhibitory activity. Among these, methyl sulfoxide 15 inhibited HDACs in enzyme assays and
caused hyperacetylation of histone H4 while not inducing the accumulation of acetylated a-tubulin in HCT116 cells.
ꢀ 2005 Elsevier Ltd. All rights reserved.
1. Introduction
SAHA, which are thought to chelate the zinc ion in
the active site in a bidentate fashion through its CO
and OH groups.7 However, hydroxamic acids occasion-
ally have produced problems such as poor pharmacoki-
netics and severe toxicity.8 In addition, although
isozyme-selective HDAC inhibitors are considered to
be useful not only as tools for probing the biology of
the enzyme but also as drugs with low toxicity, many
of the known hydroxamate HDAC inhibitors do not
distinguish well among the HDAC isozymes.9 There-
fore, it is desirable to find non-hydroxamate HDAC
inhibitors to improve the pharmacokinetics, toxicity,
and isozyme selectivity of hydroxamates. To date, some
types of non-peptide non-hydroxamate HDAC inhibi-
tors have been reported.10 However, many of these have
either reduced potency or metabolic disadvantages.
Thus, there remains a need to develop non-hydroxamate
HDAC inhibitors. Here, we report the design, synthesis,
enzyme inhibition, and in-cell selective histone deacetyl-
ase inhibition of SAHA-based non-hydroxamates.
The dynamic homeostasis of the nuclear acetylation of
histones is regulated by the opposing activity of the
enzymes, histone acetyl transferases (HATs) and histone
deacetylases (HDACs). The reversible acetylation of his-
tone lysine residues by HATs and HDACs is one of the
several possible regulatory mechanisms of gene expres-
sion.1 When HDACs are inhibited, histone hyperacetyl-
ation occurs. The disruption of the chromatin structure
by histone hyperacetylation leads to the transcriptional
activation of genes2 associated with some disease states
such as cancer3 and inflammation.4 Indeed, HDAC
inhibitors such as trichostatin A (TSA) and suberoylan-
ilide hydroxamic acid (SAHA) (Fig. 1) have potent
anti-cancer effects and some of them are currently in
phase I/II clinical trials.5
Previously reported HDAC inhibitors are mostly
hydroxamic acid derivatives,6 typified by TSA and
Figure 1. Structures of TSA and SAHA.
Keywords: Histone deacetylase inhibitor; Non-hydroxamate.
*
Corresponding authors. Tel./fax: +81 52 836 3407; e-mail addresses: suzuki@phar.nagoya-cu.ac.jp; miyata-n@phar.nagoya-cu.ac.jp
0968-0896/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2005.04.002

T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
4333
2. Chemistry
The compounds prepared for this study are shown in Ta-
ble 1. The routes used for synthesis of the compounds are
shown in Schemes 1–4. Compounds 1 and 6 were pre-
pared as shown in Scheme 1, starting with dicarboxylic
acids 23. The condensation of dicarboxylic acids 23 with
an equivalent amount of aniline gave mono-anilides 24.
The carboxylic acid 24b was converted to Weinreb amide
25 in the presence of EDCI and HOBt. Compound 25
was allowed to react with lithium aluminum hydride at
0 ꢁC to give aldehyde 26. Reductive amination of 26 with
mono-Boc ethylenediamine, Boc protection of the resul-
tant amine, chromatographic purification, and removal
of the two Boc groups afforded ethylenediamine 1. Carb-
oxylic acid 24a was converted to acylsulfonamide 6 by
acylation of methanesulfonamide in the presence of
CDI and DBU in DMF.
Amine 27 served as a key common intermediate for the
synthesis of compounds 2–5, 8–14, 17, and 18 (Scheme
2). Carboxylic acid 24b was converted to amine 27 by
Curtius rearrangement of carboxylic acid 24b, treatment
of the resulting isocyanates with benzyl alcohol, and re-
moval of the Z group by hydrogenolysis. Amine 27
underwent a reaction with diketene to give acetoacet-
amide 2. An appropriate carboxylic acid was reacted
with amine 27 in the presence of EDCI and HOBt in
DMF to give compounds 3, 12, 13, 17, and 18. Hydroly-
sis of ester 3 provided carboxylic acid 4, and treatment of
3 with 25% aqueous ammonia produced amide 5. Amine
27 was converted to diethyl carbamoylphosphonate 8
Scheme 2. Reagents and conditions: (a) diphenylphosphoryl azide,
Et3N, benzene, reflux; (b) BnOH, reflux; (c) H2, 5% Pd–C, MeOH; (d)
diketene, rt (for 2); (e) RCOOH, EDCI, HOBt, DMF, rt (for 3, 12, 13,
17, and 18); (f) EtSCOP(O)(OEt)2, rt (for 8); (g) (i) bromoacetyl
chloride, Et3N, THF, rt; (ii) P(OEt)3, tetrabutyl ammonium iodide,
reflux (for 10); (h) (i) thiazolidine-2,3-dicarboxylic acid 3-tert-butyl
ester, EDCI, HOBt, DMF, rt; (ii) 4NHClÆAcOEt, AcOEt, rt (for 11);
(i) tetrazole-5-acetic acid, bis(2-oxo-3-oxazolidinyl)phosphinic chlo-
ride, 0 ꢁC to rt (for 14); (j) 2 N aq NaOH, MeOH, rt; (k) 25% aq NH3,
MeOH, rt; (l) TMSBr, MeCN, rt; (m) chloromethylsulfonyl chloride,
Et3N, 0 ꢁC to rt.
Scheme 3. Reagents and conditions: (a) Na2SO3, EtOH, H2O, reflux;
(b) SOCl2, DMF, toluene, reflux; (c) 25% aq NH3, 4-(dimethyl-
amino)pyridine, pyridine, CH2Cl2, rt; (d) 2 N aq NaOH, EtOH, rt; (e)
aniline, EDCI, HOBt, DMF, rt; (f) AcOH, 1,10-carbonyldiimidazole,
DBU, DMF, rt.
upon reacting with S-ethyl diethylphosphonothiolfor-
mate, and following ester group removal with trimethyl-
silyl bromide gave carbamoylphosphonic acid 9.
Acylation of amine 27 with bromoacetyl chloride fol-
lowed by an Arbazov reaction provided diethyl phospho-
nate 10. Coupling between amine 27 and N-Boc
Scheme 1. Reagents and conditions: (a) aniline, 180 ꢁC; (b) N,O-
dimethylhydroxylamine hydrochloride, Et3N, EDCI, HOBt, DMF, rt;
(c) LiAlH4, THF, 0 ꢁC; (d) N-(2-aminoethyl)carbamic acid tert-butyl
ester, NaBH(OAc)3, CH2Cl2, AcOH, rt; (e) (Boc)2O, Et3N, rt; (f) TFA,
CH2Cl2, rt; (g) MsNH2, 1,10-carbonyldiimidazole, DBU, DMF, rt.

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T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
3. Results and discussion
3.1. Enzyme assays
The compounds synthesized in this study were tested
with an in vitro assay using a HeLa nuclear extract rich
in HDAC activity. The results are summarized in Table
1. SAHA was used as a positive control, which inhibited
100% of the HDAC activity at 100 lM and had an IC50
of 0.28 lM.
Table 1. HDAC enzyme inhibition data for compounds 1–22a
Compound
R
n
% inhibition at 100 lM
1
–NHCH2CH2NH2
–NHCOCH2COMe
–NHCOCH2COOEt
–NHCOCH2COOH
–NHCOCH2CONH2
–CONHSO2Me
6
5
5
5
5
6
6
5
5
5
12 1.2
12 1.3
2
3
0
1.4
Scheme 4. Reagents and conditions: (a) LiOHÆH2O, EtOH, THF,
H2O, rt; (b) (COCl)2, DMF, CH2Cl2, rt; (c) aniline, Et3N, CH2Cl2, rt;
(d) 15% aq NaSMe, EtOH, rt; (e) 1 equiv of m-chloroperoxybenzoic
acid, CH2Cl2, rt; (f) AcSK, EtOH, rt; (g) 2 N aq NaOH, EtOH, rt; (h)
5-bromovaleric acid ethyl ester, NaOEt, EtOH, rt; (i) 2 equiv of m-
chloroperoxybenzoic acid, CH2Cl2, rt.
4
4.1 0.56
1.1 2.2
21 3.0
2.5 1.9
5
6
7
–SO2NHAc
8
–NHCOP(O)(OEt)2
–NHCOP(O)(OH)2
–NHCOCH2P(O)(OEt)2
0 1.6
5.8 1.3
9
10
0
0
0.36
11
12
5
5
1.5
thiazolidine-2-carboxylic acid in the presence of EDCI
and HOBt and subsequent deprotection of the Boc group
under acidic conditions produced 2-thiazolidinecarboxa-
mide 11. Amine 27 was reacted with tetrazole-5-acetic
acid in the presence of bis(2-oxo-3-oxazolidinyl)phosphi-
nic chloride to give tetrazole-5-acetamide 14. Chloro-
methyl sulfonamide 19 was obtained by the coupling of
amine 27 with chlorosulfonyl chloride.
21 4.6
2.0 1.1
17 1.7
13
14
5
5
The preparation of sulfonamide derivatives 7 and 16 was
achieved via sulfonyl chloride 29 (Scheme 3). Bromide
28 was converted to sulfonyl chloride 29 by a reaction
with Na2SO3 and by subsequent treatment with thionyl
chloride. Treatment of the sulfonyl chloride 29 with 25%
aqueous ammonia and subsequent hydrolysis yielded
sulfonamide 30. The condensation of carboxylic acid
30 with aniline gave compound 16. Sulfonamide 16
was acetylated to the reversed acylsulfonamide 7 in the
same procedure employed in the preparation of 6.
15
16
17
18
19
20
21
22
–S(O)Me
–SO2NH2
6
6
5
5
5
6
6
6
62 2.0
10 0.97
11 3.5
3.7 2.3
–NHCOCH@CH2
–NHCOC„CH
–NHSO2CH2Cl
–SO2(CH2)4COOH
–S(O)(CH2)4COOH
–SO2Me
0
0.56
6.1 3.3
12 6.1
33 2.0b
a Values are means standard deviations of a minimum of three sep-
arate experiments.
Compounds 15, 20, and 21 were synthesized from ester
28 as depicted in Scheme 4. Acid chloride 31 was pre-
pared from ester 28 by hydrolysis of the ethyl ester
and subsequent reaction with oxalyl chloride. The ami-
no group of aniline was acylated with the acid chloride
31 to give the amide 32. Sulfide 33 was obtained by
the alkylation of methylmercaptan with bromide 32.
Oxidation of 33 with 1 equiv of MCPBA gave the sulf-
oxide 15. Bromide 32 was also treated with potassium
thioacetate to give the thioester, after which deacetyla-
tion of the thioacetate under alkaline conditions gave
thiol 34. Thiol 34 was alkylated with ethyl 5-bromoval-
erate, oxidized with 1 or 2 equivalent amounts of
MCPBA, and then hydrolyzed to give compounds 20
and 21.
b Data taken from the literature (Ref. 10i).
The co-crystal structure of an archaebacterial HDAC
homologue (HDAC-like protein, HDLP) with SAHA
revealed that the hydroxamic acid group coordinates
the zinc ion in the active site through its CO and OH
groups.7 Based on these data, we designed compounds
1–14 which were expected to chelate zinc ion in the ac-
tive site in a bidentate fashion. Notably, an acylsulfon-
amide moiety has been reported to bind zinc ion in a
bidentate fashion through its CO and SO2 groups,11
and carbamoylphosphonic acids are known to be matrix

T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
4335
metalloproteinase inhibitors,12 another form of zinc-
dependent metalloenzyme. However, these compounds
were found to be totally inactive, although acylsulfon-
amide 6 and 2-thiophenecarboxamide 12 showed a cer-
tain level of inhibitory activity against HDACs. The
reason that compounds 1–14 were weakly active is un-
clear, but it is probably because they lack interaction
with amino acid residues such as Tyr and His in the ac-
tive site of HDACs.7
Figure 2. Western blot analysis of histone hyperacetylation and a-
tubulin acetylation induction in HCT 116 cells produced by compound
15 and by reference compound SAHA.
Next, compounds with methyl sulfoxide (15) and sulfon-
amide13 (16), which are expected to coordinate zinc ions
monodentately, were evaluated for anti-HDAC activity.
Interestingly, although the inhibitory ability of mono-
dentate zinc-binding groups (ZBGs) was thought to be
less than that of bidentate ZBGs, methyl sulfoxide 15
was more potent than compounds 1–14 in inhibiting
68% of the HDAC activity at 100 lM, with an IC50 of
48 lM.
Western blot analysis was performed (Fig. 2). HCT116
human colon cancer cells were treated with SAHA or
compound 15 for 8 h at 37 ꢁC. As reported previously,
5 lM of SAHA caused the accumulation of both acetyl-
ated histone H4 and acetylated a-tubulin. On the other
hand, 100 lM of compound 15 did not induce a-tubulin
acetylation although the same level of histone H4 hyper-
acetylation as with SAHA was observed. These results
suggested that compound 15 is inactive toward TDAC
(HDAC6) in cells.
The crystal structure of the HDLP/SAHA complex
made it clear that there are nucleophilic amino acids
such as His in the active center of HDACs.7 Compounds
bearing acrylamide (17), 2-propynamide (18), and chlo-
romethanesulfonamide (19) could form covalent bonds
with nucleophilic amino acids of the enzyme and func-
tion as irreversible HDAC inhibitors. However, these
compounds were only weakly active against HDACs.
4. Conclusion
We have designed and prepared a series of SAHA-based
non-hydroxamate compounds as (i) analogues bearing
functional groups expected to chelate zinc ion biden-
tately (compounds 1–14) or monodentately (compounds
15 and 16), (ii) irreversible inhibition-oriented com-
According to the data of the crystal structure of the
˚
HDLP/SAHA complex, there is a 14 A long internal
cavity adjacent to the active site.7,14 We designed the
cavity-targeting compounds 20 and 21 modeled after
sulfoxide 15 and sulfone 22. Sulfone 22 has been re-
ported to inhibit HDACs.10i The carboxylic acid of 20
and 21 could interact with Arg 27 (HDLP numbering),
˚
pounds (compounds 17–19), and (iii) 14 A internal cav-
ity-targeting compounds (compounds 20 and 21), and
evaluated their inhibitory effect on HDACs. Among
them, methyl sulfoxide 15 inhibited HDACs in enzyme
assays and showed great HDAC/TDAC selectivity in
cellular assays.
˚
a component in the construction of the 14 A internal
cavity, and these compounds were anticipated to be
more potent HDAC inhibitors. However, contrary to
expectation, compounds 20 and 21 did not show strong
activity as compared with their parent compounds 15
and 22. At the time this work was ongoing, the X-ray
structure of human HDAC8 was published.15 According
to this work, there is no such large internal cavity adja-
cent to the active site. This may be the reason why com-
pounds 20 and 21 did not exhibit inhibitory activity
against human HDACs.
In conclusion, we have identified a novel lead compound,
from which potent HDAC isozyme-selective inhibitors
can be developed. The findings of this study should also
pave the way for the development of new medicines with-
out side effects caused by interference with microtubule
dynamics associated with HDAC6. Currently, detailed
structure–activity relationship studies of methyl sulfox-
ide-based HDAC inhibitors are under way.
3.2. Cellular assays
5. Experimental section
5.1. Chemistry
HDACs are also responsible for the deacetylation of
non-histone proteins.1c Notably, HDAC6, one of the
isozymes of HDAC, has recently been reported to be a
a-tubulin deacetylase (TDAC).16 The reversible acetyla-
tion of a-tubulin is involved in microtubule stability.
Therefore, compounds with high HDAC/TDAC selec-
tivity are desirable for biological studies and medicinal
use. However, many hydroxamate HDAC inhibitors
such as TSA and SAHA cause the accumulation of acet-
ylated histones as well as acetylated a-tubulin, which
indicates that they do not discriminate between HDAC6
and the other isozymes.9 To investigate the selectivity of
sulfoxide 15, the most active compound in this study,
Melting points were determined using a Yanagimoto mi-
cro melting point apparatus or a Buchi 545 melting
¨
point apparatus and were left uncorrected. Proton nu-
clear magnetic resonance spectra (1H NMR) were re-
corded on a JEOL JNM-LA400 or JEOL JNM-LA500
spectrometer in solvent as indicated. Chemical shifts
(d) were reported in parts per million relative to the
internal standard tetramethylsilane. Elemental analysis
was performed with a Yanaco CHN CORDER NT-5
analyzer, and all values were within 0.4% of the

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T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
calculated values. High-resolution mass spectra
(HRMS) were recorded on a JEOL JMS-SX102A mass
spectrometer. GC–MS analyses were performed on a
Shimadzu GCMS-QP2010. Reagents and solvents were
purchased from Aldrich, Tokyo Kasei Kogyo, Wako
Pure Chemical Industries, and Kanto Kagaku and used
without purification. Flash column chromatography
was performed using Silica Gel 60 (particle size 0.046–
0.063 mm) supplied by Merck.
2.37 (2H, t, J = 7.3 Hz), 1.76 (2H, quintet, J = 7.3 Hz),
1.68 (2H, quintet, J = 7.4 Hz), 1.42 (2H, quintet,
J = 7.3 Hz).
5.1.4. 7-(2-Aminoethylamino)heptanoic acid phenylamide
ditrifluoroacedic acid salt (1Æ2TFA). To a solution of 26
(240 mg, 1.09 mmol) obtained above in CH2Cl2 (3 mL)
and AcOH (1 mL) was added N-(2-aminoethyl)car-
bamic acid tert-butyl ester (190 mg, 1.19 mmol) at room
temperature. The solution was stirred for 15 min, fol-
lowed by the addition of sodium triacetoxy borohydride
(475 mg, 2.24 mmol). The solution was stirred overnight
at room temperature. To the solution were added trieth-
ylamine (12 mL) and di-tert-butyl dicarbonate (300 mg,
1.37 mmol), and the mixture was stirred at room tem-
perature for 5 h. The reaction mixture was poured into
water and extracted with AcOEt. The AcOEt layer
was separated, washed with water, saturated aqueous
NaHCO3 and brine, and was dried over Na2SO4. Filtra-
tion and concentration in vacuo and purification by sil-
ica gel flash chromatography (n-hexane/AcOEt = 5:2)
gave 120 mg of colorless oil. To a solution of the oil in
CH2Cl2 (2 mL) was added trifluoroacetic acid (3 mL),
and the mixture was stirred at room temperature for
1 h. The solution was concentrated in vacuo to give a
solid. The residue was triturated in ether to give
112 mg (21%) of 1Æ2TFA as a crude solid. The solid
was recrystallized from AcOEt and collected by filtra-
tion to give 103 mg of 1Æ2TFA as colorless crystals: mp
113–114 ꢁC; 1H NMR (DMSO-d6 400 MHz, d; ppm)
9.87 (1H, s), 7.58 (2H, d, J = 7.8 Hz), 7.28 (2H, t,
J = 7.6 Hz), 7.02 (1H, t, J = 7.1 Hz), 3.20–3.05 (4H,
m), 2.95 (2H, t, J = 8.1 Hz), 2.31 (2H, t, J = 7.1 Hz),
1.66–1.50 (4H, m), 1.42–1.26 (4H, m); Anal. Calcd for
C15H25N3OÆ2TFA: C, 46.44; H, 5.54; N, 8.55. Found:
C, 46.25; H, 5.44; N, 8.27.
5.1.1. 6-Phenylcarbamoylhexanoic acid (24b). A mixture
of aniline (5.80 g, 62.3 mmol) and pimeric acid (23b,
10.0 g, 62.4 mmol) was stirred at 180 ꢁC for 1 h. After
cooling, the mixture was diluted with AcOEt–THF
and the slurry was filtered. The filtrate was washed with
saturated aqueous NaHCO3 and the aqueous layer was
acidified with concentrated HCl. The precipitated crys-
tals were collected by filtration to give 7.11 g (49%) of
1
24b as a white solid: H NMR (DMSO-d6 400 MHz,
d; ppm) 11.97 (1H, broad s), 9.83 (1H, s), 7.58 (2H, d,
J = 7.8 Hz), 7.27 (2H, t, J = 7.9 Hz), 7.01 (1H, t,
J = 7.4 Hz), 2.67 (2H, t, J = 7.4 Hz), 2.21 (2H, t,
J = 7.3 Hz), 1.62–1.49 (4H, m), 1.34–1.27 (2H, m).
5.1.2. Heptanedioic acid methoxymethylamide phenyl-
amide (25). To a solution of 24b (5.7 g, 24.2 mmol) ob-
tained
above
and
N,O-dimethylhydroxylamine
hydrochloride (4.80 g, 49.2 mmol) in DMF (70 mL)
were added 1-ethyl-3-(3-dimethylaminopropyl)carbodi-
imide hydrochloride (6.00 g, 31.3 mmol), 1-hydroxy-
1H-benzotriazole monohydrate (4.80 g, 31.3 mmol),
and triethylamine (7.50 mL, 54.0 mmol) and the mixture
was stirred overnight at room temperature. The reaction
mixture was poured into water and extracted with
AcOEt. The AcOEt layer was separated, washed with
water, 2 N aqueous HCl, saturated aqueous NaHCO3
and brine, and was dried over Na2SO4. Filtration and
concentration in vacuo and separation by silica gel flash
chromatography (n-hexane/AcOEt = 8:1) gave 6.34 g
5.1.5. Phenylcarbamoylheptanoic acid (24a). Compound
24a was prepared from aniline and suberic acid in 51%
yield by using the same procedure described for 24b:
1H NMR (DMSO-d6 400 MHz, d; ppm) 11.98 (1H,
broad s), 9.84 (1H, s), 7.58 (2H, d, J = 7.8 Hz), 7.28
(2H, t, J = 7.8 Hz), 7.01 (1H, t, J = 7.3 Hz), 2.29 (2H,
t, J = 7.4 Hz), 2.20 (2H, t, J = 7.3 Hz), 1.58 (2H, t,
J = 7.2 Hz), 1.50 (2H, t, J = 7.1 Hz), 1.29 (4H, m).
1
(94%) of 25 as a colorless crystal: H NMR (CDCl3,
400 MHz, d; ppm) 7.54 (2H, d, J = 7.8 Hz), 7.41 (1H,
broad s), 7.31 (2H, t, J = 7.6 Hz), 7.09 (1H, t,
J = 7.6 Hz), 3.67 (3H, s), 3.18 (3H, s), 2.45 (2H, t,
J = 7.3 Hz), 2.39 (2H, t, J = 7.3 Hz), 1.77 (2H, quintet,
J = 7.3 Hz), 1.70 (2H, quintet, J = 7.6 Hz), 1.44 (2H,
quintet, J = 7.8 Hz).
5.1.6. 8-Methanesulfonylamino-8-oxooctanoic acid phen-
ylamide (6). To a solution of 24a (500 mg, 2.01 mmol) in
DMF (5 mL) was added 1,1-carbonyldiimidazole
(360 mg, 2.22 mmol). After 1 h, methanesulfonamide
(210 mg, 2.21 mmol) and 1,8-diazabicyclo[5,4,0]-7-unde-
cene (330 lL, 2.21 mmol) were added to the reaction
mixture. After overnight stirring, the mixture was
poured into 2 N aqueous HCl and the precipitated crys-
tals were collected by filtration to give 595 mg (91%) of 6
as a crude solid. The solid was recrystallized from
AcOEt and collected by filtration to give 511 mg of 6
as colorless crystals: mp 145–146 ꢁC; 1H NMR
(DMSO-d6 400 MHz, d; ppm) 11.64 (1H, broad s),
9.84 (1H, broad s), 7.58 (2H, d, J = 8.0 Hz), 7.28 (2H,
t, J = 7.8 Hz), 7.01 (1H, t, J = 7.1 Hz), 3.22 (3H, s),
2.29 (2H, t, J = 7.3 Hz), 2.26 (2H, t, J = 7.3 Hz),
1.58 (2H, quintet, J = 7.4 Hz), 1.52 (2H, quintet,
5.1.3. 7-Oxoheptanoic acid phenylamide (26). To a sus-
pension of lithium aluminum hydride (300 mg,
7.91 mmol) in THF (30 mL) was added a solution of
25 (2.50 g, 8.98 mmol) obtained above in THF (20 mL)
dropwise with cooling by an ice-water bath. The reac-
tion mixture was stirred at 0 ꢁC for 30 min. To the mix-
ture were added water (0.3 mL), 15% aqueous NaOH
(0.3 mL), and water (0.9 mL) in a sequential order and
the slurry was filtered. After the solid was washed with
THF (10 mL), the combined filtrates were concentrated
in vacuo. The residue was purified by silica gel flash
chromatography (n-hexane/AcOEt = 1:1) to give 1.42 g
(72%) of 26 as a white solid: 1H NMR (CDCl3,
400 MHz, d; ppm) 9.77 (1H, s), 7.52 (2H, d,
J = 8.1 Hz), 7.32 (2H, t, J = 7.6 Hz), 7.25 (1H, broad
s), 7.10 (1H, t, J = 7.3 Hz), 2.47 (2H, t, J = 7.1 Hz),

T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
4337
J = 7.3 Hz), 1.36–1.22 (4H, m); MS (EI) m/z: 326 (M+);
Anal. Calcd for C15H22N2O4S: C, 55.19; H, 6.79; N,
8.58. Found: C, 55.00; H, 6.72; N, 8.73.
5.1.9. N-(5-Phenylcarbamoylpentyl)malonamic acid ethyl
ester (3). Mp 107–110 ꢁC; 1H NMR (CDCl3, 500
MHz, d; ppm) 7.52 (2H, d, J = 8.2 Hz), 7.32 (2H, t,
J = 7.9 Hz), 7.28 (1H, broad s), 7.18 (1H, broad s),
7.10 (1H, t, J = 7.3 Hz), 4.19 (2H, q, J = 7.1 Hz), 3.31
(2H, q, J = 6.6 Hz), 3.29 (2H, s), 2.37 (2H, t,
J = 7.5 Hz), 1.77 (2H, quintet, J = 7.5 Hz), 1.59 (2H,
quintet, J = 7.3 Hz), 1.43 (2H, quintet, J = 7.5 Hz),
1.28 (3H, t, J = 7.2 Hz); MS (EI) m/z: 320 (M+); Anal.
Calcd for C17H24N2O4: C, 63.73; H, 7.55; N, 8.74.
Found: C, 63.82; H, 7.65; N, 8.47.
5.1.7. 6-Aminohexanoic acid phenylamide (27). To a sus-
pension of 24b (1.11 g, 4.73 mmol) and triethylamine
(699 mg, 6.90 mmol) in benzene (3 mL) was added
diphenylphosphoryl azide (1.83 g, 6.64 mmol) and the
mixture was heated at reflux temperature for 1 h. Next,
benzyl alcohol (1.20 mL, 11.6 mmol) was added and the
reaction mixture was stirred at reflux temperature for
24 h. It was then concentrated in vacuo and the residue
was dissolved in AcOEt. The AcOEt solution was
washed with 0.4 N aqueous HCl, water, saturated aque-
ous NaHCO3, and brine, and was dried over Na2SO4.
Filtration and concentration in vacuo, and purification
by recrystallization from CHCl3–n-hexane gave 1.01 g
(63%) of (6-phenylcarbamoylpentyl)carbamic acid
5.1.10. Thiophene-2-carboxylic acid (5-phenylcarbamoyl-
1
pentyl)amide (12). Mp 139–141 ꢁC; H NMR (CDCl3,
500 MHz, d; ppm) 7.53 (2H, d, J = 7.6 Hz), 7.52 (1H,
d, J = 4.9 Hz), 7.45 (1H, d, J = 4.9 Hz), 7.35 (1H, broad
s), 7.31 (2H, t, J = 7.9 Hz), 7.10 (1H, t, J = 7.3 Hz), 7.04
(1H, t, J = 4.9 Hz), 6.27 (1H, broad s), 3.47 (2H, q,
J = 6.6 Hz), 2.40 (2H, t, J = 7.2 Hz), 1.80 (2H, quintet,
J = 7.4 Hz), 1.67 (2H, quintet, J = 7.0 Hz), 1.48 (2H,
quintet, J = 7.7 Hz); MS (EI) m/z: 316 (M+); HRMS
Calcd for C17H20N2O2S 316.125. Found 316.125.
1
benzyl ester as a colorless needle: H NMR (DMSO-d6
400 MHz, d; ppm) 9.81 (1H, s), 7.57 (2H, d,
J = 7.8 Hz), 7.37–7.22 (8H, m), 7.00 (1H, t, J =
7.4 Hz), 4.99 (2H, s), 2.99 (2H, q, J = 6.5 Hz), 2.28
(2H, t, J = 7.4 Hz), 1.58 (2H, quintet, J = 7.6 Hz), 1.43
(2H, quintet, J = 7.1 Hz), 1.32 (2H, quintet,
J = 7.8 Hz); MS (EI) m/z: 340 (M+).
5.1.11. Thiazole-2-carboxylic acid (5-phenylcarbamoyl-
1
pentyl)amide (13). Mp 130–133 ꢁC; H NMR (DMSO-
d6, 500 MHz, d; ppm) 9.84 (1H, broad s), 8.85 (1H,
broad s), 8.02 (1H, d, J = 3.4 Hz), 7.99 (1H, d,
J = 3.1 Hz), 7.57 (2H, d, J = 7.9 Hz), 7.27 (2H, t,
J = 7.9 Hz), 7.01 (1H, t, J = 7.0 Hz), 3.26 (2H, q,
J = 6.7 Hz), 2.30 (2H, t, J = 7.5 Hz), 1.59 (2H, quintet,
J = 7.6 Hz), 1.56 (2H, quintet, J = 7.9 Hz), 1.32 (2H,
quintet, J = 7.5 Hz); MS (EI) m/z: 317 (M+); HRMS
Calcd for C16H19N3O2 S 317.120. Found 317.119.
A solution of (6-phenylcarbamoylpentyl)carbamic acid
benzyl ester(1.00 g, 2.95 mmol) obtained above in
MeOH (50 mL) was stirred under H2 (atmospheric pres-
sure) in the presence of 5% Pd/C (106 mg) at room tem-
perature for 7 h. The catalyst was removed by filtration
through Celite, and the filtrate was concentrated in
vacuo. The residue was purified by silica gel flash chro-
matography (CHCl3/MeOH/iPrNH2 = 19:1:1) to give
584 mg (96%) of 27 as a white solid: 1H NMR
(DMSO-d6 400 MHz, d; ppm) 9.83 (1H, s), 7.58 (2H,
d, J = 7.6 Hz), 7.27 (2H, t, J = 7.9 Hz), 7.01 (1H, t,
J = 7.3 Hz), 2.55 (2H, m), 2.29 (2H, t, J = 7.4 Hz),
1.59 (2H, quintet, J = 7.4 Hz), 1.37–1.30 (4H, m).
5.1.12. N-(5-Phenylcarbamoylpentyl)acrylamide (17). Mp
1
166–168 ꢁC; H NMR (DMSO-d6, 500 MHz, d; ppm)
9.84 (1H, s), 8.07 (1H, s), 7.58 (2H, d, J = 7.9 Hz),
7.27 (2H, t, J = 7.9 Hz), 7.01 (1H, t, J = 7.5 Hz), 6.20
(1H, dd, J = 10.1, 17.1 Hz), 6.05 (1H, dd, J = 2.2,
17.1 Hz), 5.55 (1H, dd, J = 2.2, 10.1 Hz), 3.12 (2H, q,
J = 5.8 Hz), 2.29 (2H, t, J = 7.5 Hz), 1.59 (2H, quintet,
J = 7.5 Hz), 1.45 (2H, quintet, J = 7.9 Hz), 1.30 (2H,
quintet, J = 7.3 Hz); MS (EI) m/z: 260 (M+); Anal.
Calcd for C15H20N2O2: C, 69.20; H, 7.74; N, 10.76.
Found: C, 68.97; H, 7.76; N, 10.64.
5.1.8. 6-(3-Oxobutyrylamino)hexanoic acid phenylamide
(2). To a solution of 27 (107 mg, 0.52 mmol) obtained
above in MeOH (3 mL) was added diketene (0.3 mL,
3.89 mmol) at 0 ꢁC and the resulting mixture was stirred
at room temperature for 9 h. The solution was concen-
trated in vacuo to give a crude solid. The solid was puri-
fied by silica gel flash column chromatography (AcOEt
only to AcOEt/MeOH = 9:1) to give 108 mg (72%) of 2
as a yellow solid. The solid (108 mg) was recrysta-
llized from n-hexane–AcOEt–MeOH and collected by
filtration to give 69 mg of 2 as brown crystals: mp 126–
5.1.13. Propynoic acid (5-phenylcarbamoylpentyl)amide
1
(18). Mp 143–145 ꢁC; H NMR (DMSO-d6, 500 MHz,
d; ppm) 9.84 (1H, s), 8.70 (1H, s), 7.58 (2H, d,
J = 7.9 Hz), 7.28 (2H, t, J = 7.9 Hz), 7.01 (1H, t,
J = 7.3 Hz), 4.09 (1H, s), 3.07 (2H, q, J = 6.7 Hz), 2.29
(2H, t, J = 7.3 Hz), 1.58 (2H, quintet, J = 7.3 Hz), 1.43
(2H, quintet, J = 7.6 Hz), 1.28 (2H, quintet,
J = 7.3 Hz); MS (EI) m/z: 258 (M+); HRMS Calcd for
C15H18N2O2: 258.137. Found: 258.137.
1
128 ꢁC; H NMR (DMSO-d6, 500 MHz, d; ppm) 9.85
(1H, s), 8.02 (1H, s), 7.56 (2H, d, J = 8.2 Hz), 7.28 (2H,
t, J = 7.9 Hz), 7.01 (1H, t, J = 7.3 Hz), 3.27 (2H, s),
3.05 (2H, q, J = 6.5 Hz), 2.29 (2H, t, J = 7.5 Hz), 2.12
(3H, s), 1.58 (2H, quintet, J = 7.3 Hz), 1.43 (2H, quintet,
J = 7.5 Hz), 1.30 (2H, quintet, J = 7.8 Hz); MS (EI) m/z:
290 (M+); Anal. Calcd for C16H22N2O3: C, 66.18; H,
7.64; N, 9.65. Found: C, 66.04; H, 7.40; N, 9.48.
5.1.14. N-(5-Phenylcarbamoylpentyl)malonamic acid (4).
To a solution of 3 (134 mg, 0.42 mmol) in MeOH (3 mL)
was added 2 N aqueous NaOH (0.5 mL, 1.00 mmol), and
the mixture was stirred at room temperature for 30 min.
The reaction mixture was concentrated and dissolved in
AcOEt. The solution was washed with 2 N aqueous HCl
and brine, and was dried over Na2SO4. Filtration and
Compounds 3, 12, 13, 17, and 18 were prepared from
amine 27 and an appropriate carboxylic acid using the
procedure described for 25.

4338
T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
concentration in vacuo gave a crude solid. The solid was
recrystallized from n-hexane–AcOEt and collected by fil-
tration to give 54 mg (43%) as colorless crystals: mp 128–
130 ꢁC; H NMR (DMSO-d6, 500 MHz, d; ppm) 9.84
(1H, s), 8.32 (1H, s), 8.02 (1H, s), 7.58 (2H, d,
J = 7.6 Hz), 7.28 (2H, t, J = 7.9 Hz), 7.01 (1H, t,
J = 7.5 Hz), 3.10 (2H, s), 3.05 (2H, q, J = 6.4 Hz), 2.29
(2H, t, J = 7.5 Hz), 1.59 (2H, quintet, J = 7.5 Hz), 1.43
(2H, quintet, J = 7.3 Hz), 1.30 (2H, quintet,
J = 7.6 Hz); Anal. Calcd for C15H20N2O4: C, 61.63; H,
6.90; N, 9.58. Found: C, 61.67; H, 6.95; N, 9.47.
(5 mL) was stirred overnight at room temperature. After
the addition of MeOH (1 mL), the solution was concen-
trated in vacuo and the residue was triturated in AcOEt–
MeOH–n-hexane to give 57 mg (43%) of 9 as a crude so-
lid. The solid was recrystallized from AcOEt–MeOH
and collected by filtration to give 53 mg of 9 as colorless
crystals: mp 154–156 ꢁC; 1H NMR (DMSO-d6,
500 MHz, d; ppm) 9.85 (1H, s), 8.35 (1H, broad s),
7.58 (2H, d, J = 8.2 Hz), 7.28 (2H, t, J = 8.1 Hz), 7.01
(1H, t, J = 7.3 Hz), 3.10 (2H, q, J = 6.7 Hz), 2.29 (2H,
t, J = 7.6 Hz), 1.59 (2H, quintet, J = 7.6 Hz), 1.46 (2H,
quintet, J = 7.6 Hz), 1.27 (2H, quintet, J = 7.6 Hz);
Anal. Calcd for C13H19N2O5P: C, 49.68; H, 6.09; N,
8.91. Found: C, 49.51; H, 6.06; N, 8.82.
1
5.1.15. N-(5-Phenylcarbamoylpentyl)acrylamide (5). To a
solution of 3 (61 mg, 0.19 mmol) in MeOH (1.5 mL) was
added 25% aqueous NH3 (1.5 mL), and the mixture was
stirred overnight at room temperature. The reaction
mixture was concentrated in vacuo and the residue
was triturated in AcOEt to give 16 mg (28%) of 5 as a
crude solid. The solid was recrystallized from n-hex-
ane–AcOEt–MeOH and collected by filtration to give
13 mg of 5 as colorless crystals: mp 166–168 ꢁC; 1H
NMR (DMSO-d6, 500 MHz, d; ppm) 9.84 (1H, s), 7.97
(1H, t, J = 5.3 Hz), 7.58 (2H, d, J = 7.9 Hz), 7.41 (1H,
broad s), 7.28 (2H, t, J = 7.9 Hz), 7.01 (1H, t,
J = 7.31 Hz), 6.99 (1H, broad s), 3.05 (2H, q,
J = 6.5 Hz), 2.97 (2H, s), 2.28 (2H, t, J = 7.6 Hz), 1.58
(2H, quintet, J = 7.6 Hz), 1.43 (2H, quintet,
J = 7.2 Hz), 1.30 (2H, quintet, J = 7.5 Hz); MS (EI) m/z:
291 (M+); Anal. Calcd for C15H21N3O3: C, 61.84; H,
7.27; N, 14.42. Found: C, 61.68; H, 7.25; N, 14.17.
5.1.18. [(5-Phenylcarbamoylpentylcarbamoyl)methyl]-phos-
phonic acid diethyl ester (10). A mixture of 27 (618 mg,
3.00 mmol), Et3N (1.0 mL, 7.20 mmol), and bromoace-
tyl chloride (1.0 mL, 6.5 mmol) in THF (20 mL) was
stirred at room temperature for 1 h. The reaction mix-
ture was diluted with CHCl3. The solution was washed
with saturated aqueous NaHCO3, water, and brine,
and was dried over Na2SO4. Filtration and concentra-
tion in vacuo, and purification by silica gel flash column
chromatography gave 693 mg (71%) of a 1:1 mixture of
bromoacetyl 6-anilino-6-oxohexanamide and chloroace-
tyl 6-anilino-6-oxohexanamide as a light yellow solid:
1H NMR (CDCl3, 400 MHz, d; ppm) 7.52 (2H, d,
J = 8.1 Hz), 7.32 (2H, t, J = 7.8 Hz), 7.10 (1H, t,
J = 7.3 Hz), 6.64, 6.57 (1H, broad s), 4.03, 3.86 (2H,
s), 3.33 (2H, m), 2.38 (2H, t, J = 7.4 Hz), 1.78 (2H, m),
1.61 (2H, m), 1.43 (2H, m).
5.1.16. (5-Phenylcarbamoylpentylcarbamoyl)phosphonic
acid diethyl ester (8). Under Ar atmosphere, to a solution
of ethyl chloroformate (1.14 g, 9.12 mmol) in dry toluene
(50 mL) was slowly added triethylphosphite (2.82 g,
17.0 mmol) at 0 ꢁC and the resulting mixture was stirred
at room temperature for 4 h. The solution was concen-
trated and purified by silica gel flash column chromato-
graphy (AcOEt/n-hexane = 1:2) to give 1.25 g (61%) of
S-ethyl diethylphosphonothiolformate as a crude oil:
1H NMR (CDCl3, 500 MHz, d; ppm) 4.25 (4H, m),
3.01 (2H, q, J = 8.5 Hz), 1.37 (6H, t, J = 5.5 Hz), 1.28
(3H, t, J = 9.3 Hz); MS (EI) m/z 226 (M+).
To a solution of the solid (164 mg, 0.54 mmol) obtained
above in P(OEt)3 (852 mg, 5.13 mmol) was added tetra-
butylammonium iodide (81.9 mg, 0.22 mmol). The mix-
ture was stirred overnight at reflux temperature. After
cooling, the reaction mixture was evaporated in vacuo
to remove triethylphosphite. Purification by silica gel
flash column chromatography (AcOEt only to AcOEt/
MeOH = 9:1) gave 147 mg of 10 as a colorless oil
1
(74%): H NMR (CDCl3, 500 MHz, d; ppm) 7.94 (1H,
broad s), 7.57 (2H, d, J = 7.9 Hz), 7.30 (2H, t,
J = 7.9 Hz), 7.08 (1H, t, J = 7.3 Hz), 6.89 (1H, broad
s), 4.13 (4H, m), 3.27 (2H, q, J = 6.5 Hz), 2.84 (2H, d,
J = 20.7 Hz), 2.35 (2H, t, J = 7.3 Hz), 1.75 (2H, quintet,
J = 7.5 Hz), 1.56 (2H, quintet, J = 7.3 Hz), 1.42 (2H,
quintet, J = 7.5 Hz), 1.33 (6H, t, J = 7.0 Hz); MS (EI)
m/z: 384 (M+); HRMS Calcd for C18H29N2O5P
384.181, Found 384.182.
To a solution of S-ethyl diethylphosphonothiolformate
(679 mg, 3.00 mmol) obtained above in MeCN (5 mL)
was added 27 (215 mg, 1.04 mmol), and the solution
was stirred overnight at room temperature. The solution
was concentrated and purified by silica gel flash column
chromatography (AcOEt/n-hexane = 1/2 to AcOEt
1
only) to give 208 mg (54%) of 8 as a colorless oil: H
NMR (CDCl3, 500 MHz, d; ppm) 7.65 (1H, broad s),
7.55 (2H, d, J = 7.9 Hz), 7.30 (2H, t, J = 7.9 Hz), 7.22
(1H, broad s), 7.09 (1H, t, J = 7.5 Hz), 4.22 (4H, m),
3.34 (2H, q, J = 6.6 Hz), 2.36 (2H, t, J = 7.3 Hz), 1.75
(2H, quintet, J = 7.6 Hz), 1.59 (2H, quintet, J =
7.5 Hz), 1.41 (2H, quintet, J = 7.6 Hz), 1.36 (6H, t,
J = 7.2 Hz); MS (EI) m/z: 370 (M+); HRMS Calcd for
C17H27N2O5P 370.166. Found 370.165.
5.1.19. Thiazolidine-2-carboxylic acid (5-phenylcarba-
moylpentyl)amide hydrochloric acid salt (11ÆHCl). To a
solution of thiazolidine-2-carboxylic acid (704 mg,
5.29 mmol) in MeOH (10 mL) was added Boc2O
(1.97 g, 9.04 mmol) and Et3N (0.5 mL), and the mixture
was stirred overnight at room temperature. The solution
was concentrated, dissolved with AcOEt and the AcOEt
solution was extracted with saturated aqueous NaHCO3
and the aqueous layer was neutralized with 2 N HCl.
The aqueous layer was extracted with ether and the or-
ganic layer was washed with brine, and dried over
Na2SO4. Filtration and concentration in vacuo, and
5.1.17. (5-Phenylcarbamoylpentylcarbamoyl)phosphonic
acid (9). A solution of 8 (156 mg, 0.42 mmol) and
bromotrimethylsilane (614 mg, 4.01 mmol) in MeCN

T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
4339
purification by silica gel flash column chromatography
(n-hexane/AcOEt = 2:1) gave 966 mg of thiazolidine-
2,3-dicarboxylic acid 3-tert-butyl ester (78%) as a crude
solid: H NMR (CDCl3, 500 MHz, d; ppm) 11.09 (1H,
broad s), 5.33, 5.15 (1H, s), 3.99, 3.83 (2H, m), 3.24,
3.01 (2H, m), 1.48, 1.45 (9H, s).
water bath. The solution was stirred at room temperature
for 30 min. The reaction mixture was poured into water
and extracted with CHCl3. The CHCl3 layer was sepa-
rated, washed with water, 1 N aqueous HCl and brine,
and was dried over Na2SO4. Filtration and concentration
in vacuo, and purification by silica gel flash column chro-
matography (n-hexane/AcOEt = 2:1 to 1:1) gave 54 mg
(23%) of 19 as a yellow solid. The solid was recrystallized
from n-hexane–CHCl3 and collected by filtration to give
1
2-(5-Phenylcarbamoylpentylcarbamoyl)thiazolidine-3-
carboxylic acid tert-butyl ester was prepared from amine
27 and thiazolidine-2,3-dicarboxylic acid 3-tert-butyl es-
ter obtained above in 79% yield using the procedure de-
scribed for 25. 1H NMR (DMSO-d6, 500 MHz, d; ppm)
9.85 (1H, s), 7.98 (1H, broad s), 7.58 (2H, d, J = 7.9 Hz),
7.28 (2H, t, J = 7.9 Hz), 7.01 (1H, t, J = 7.5 Hz), 5.14,
5.02 (1H, s), 3.75, 3.67 (2H, m), 3.06 (2H, m), 2.98
(2H, m), 2.29 (2H, t, J = 7.7 Hz), 1.58 (2H, quintet,
J = 7.5 Hz), 1.40 (2H, m), 1.34 (9H, s), 1.32 (2H, quin-
tet, J = 7.5 Hz); MS (EI) m/z: 421 (M+).
1
46 mg of 19 as colorless crystals: mp 122–124 ꢁC; H
NMR (CDCl3, 500 MHz, d; ppm) 7.51 (2H, d,
J = 7.9 Hz), 7.32 (2H, t, J = 7.9 Hz), 7.15 (1H, broad s),
7.11 (1H, t, J = 7.3 Hz), 4.76 (1H, broad s), 4.49 (2H,
s), 3.23 (2H, q, J = 6.7 Hz), 2.39 (2H, t, J = 7.2 Hz),
1.77 (2H, quintet, J = 7.5 Hz), 1.65 (2H, quintet, J =
7.2 Hz), 1.48 (2H, quintet, J = 7.7 Hz); MS (EI) m/z:
318 (M+); Anal. Calcd for C13H19ClN2O3S: C, 48.97;
H, 6.01; N, 8.79. Found: C, 48.88; H, 5.85; N, 8.71.
To a solution of 2-(5-phenylcarbamoylpentylcarbamoyl)-
thiazolidine-3-carboxylic acid tert-butyl ester (180 mg,
0.43 mmol) in AcOEt (5 mL) was added 4 N HClÆAcOEt
(1.0 mL, 4.00 mmol), and the mixture was stirred
overnight at room temperature. The solution was con-
centrated in vacuo and triturated in AcOEt–MeOH–n-
hexane to give 112 mg (81%) of 11ÆHCl as a crude solid.
The solid was recrystallized from AcOEt–MeOH and
collected by filtration to give 102 mg as colorless crys-
5.1.22. 7-Chlorosulfonylheptanoic acid ethyl ester (29).
To an aqueous solution (7 mL) of anhydrous sodium
sulfite (2.03 g, 16.1 mmol) was added a solution of 7-
bromoheptanoic acid ethyl ester (28, 2.0 g, 8.43 mmol)
in EtOH (5 mL) and the solution was boiled under re-
flux with stirring for 2 h. The solution was evaporated
to dryness and the solid was dried in vacuo at 60 ꢁC.
This white solid was placed in a flask, toluene (30 mL)
was added followed by a catalytic amount of DMF,
and then thionyl chloride (6.2 mL, 85.0 mmol) was
added dropwise. The mixture was boiled under reflux
with stirring for 5 h, diluted with AcOEt, washed with
aqueous saturated cold water, and brine, and dried over
MgSO4. Filtration and concentration in vacuo, and
purification by silica gel flash chromatography (n-hex-
ane/AcOEt = 4:1) gave 2.02 g (93%) of 29: 1H NMR
(CDCl3, 400 MHz, d; ppm) 4.13 (2H, q, J = 7.1 Hz),
3.66 (2H, t, J = 7.8 Hz), 2.31 (2H, t, J = 7.3 Hz), 2.06
(2H, quintet, J = 7.8 Hz), 1.66 (2H, quintet,
J = 7.3 Hz), 1.53 (2H, quintet, J = 7.8 Hz), 1.41 (2H,
quintet, J = 7.1 Hz), 1.26 (2H, quintet, J = 7.1 Hz).
1
tals: mp 144–147 ꢁC; H NMR (DMSO-d6, 500 MHz,
d; ppm) 9.90 (1H, s), 8.56 (1H, broad s), 7.59 (2H, d,
J = 7.6 Hz), 7.28 (2H, t, J = 7.9 Hz), 7.01 (1H, t,
J = 7.5 Hz), 5.17 (1H, s), 3.60 (1H, m), 3.45 (1H, m),
3.11 (4H, m), 2.30 (2H, t, J = 7.1 Hz), 1.59 (2H, quintet,
J = 7.5 Hz), 1.47 (2H, quintet, J = 7.0 Hz), 1.31 (2H,
quintet, J = 7.5 Hz); MS (EI) m/z: 321 (M+); HRMS
Calcd for C16H23N3O2S 321.151. Found 321.151.
5.1.20. 6-(2-1H-Tetrazol-5-ylacetylamino)hexanoic acid
phenylamide (14). A solution of 1H-tetrazole-5-acetic
acid (76.8 mg, 0.60 mmol) and 27 (105 mg, 0.51 mmol)
in CH2Cl2 (1 mL) was cooled to 0 ꢁC and bis(2-oxo-3-
oxazolidinyl)phosphinic chloride (206 mg, 0.79 mmol)
was added. The mixture was stirred overnight at room
temperature and concentrated in vacuo. To the residue
was added saturated aqueous NaCl and the precipitate
was collected to give 160 mg (99%) of 14 as a pink solid.
The solid was recrystallized from MeOH and collected
by filtration to give 14 mg of 14 as pink crystals: mp
5.1.23. 7-Sulfamoylheptanoic acid phenylamide (16). To a
mixture of 25% aqueous NH3 (10 mL), a catalytic
amount of 4-(dimethylamino)pyridine, and THF
(20 mL) was added a solution of 29 (1.33 g, 5.18 mmol)
obtained above in THF (10 mL), and the mixture was
stirred at room temperature for 1 h. The reaction mix-
ture was poured into water and extracted with AcOEt.
The AcOEt layer was separated, washed with water, satu-
rated aqueous NaHCO3 and brine, and was dried over
Na2SO4. Filtration and concentration in vacuo and
purification by silica gel flash chromatography (n-hex-
ane/AcOEt = 1:1) gave 1.11 g (91%) of 7-sulfamoylhep-
tanoic acid ethyl ester as a crude oil.
1
174–176 ꢁC; H NMR (DMSO-d6, 500 MHz, d; ppm)
9.88 (1H, s), 8.34 (1H, broad s), 7.58 (2H, d, J =
7.6 Hz), 7.28 (2H, t, J = 7.9 Hz), 7.01 (1H, t,
J = 7.5 Hz), 3.86 (2H, s), 3.09 (2H, q, J = 6.5 Hz), 2.30
(2H, t, J = 7.5 Hz), 1.59 (2H, quintet, J = 7.1 Hz), 1.45
(2H, quintet, J = 7.5 Hz), 1.31 (2H, quintet, J =
7.8 Hz); Anal. Calcd for C15H20N6O2Æ1/4H2O: C, 56.15;
H, 6.44; N, 26.19. Found: C, 56.07; H, 6.25; N, 26.30.
Compound 16 was prepared from 7-sulfamoylheptanoic
acid ethyl ester obtained above in 64% yield using the
1
procedure described for 4 and 25: mp 163–164 ꢁC; H
5.1.21. 6-Chloromethanesulfonylaminohexanoic acid phen-
ylamide (19). To a solution of 27 (153 mg, 0.74 mmol)
and triethylamine (0.40 mL, 2.87 mmol) in CHCl3
(3 mL) was added chloromethylsulfonyl chloride
(311 mg, 2.09 mmol) dropwise with cooling by an ice-
NMR (DMSO-d6 400 MHz, d; ppm) 9.85 (1H, broad
s), 7.58 (2H, d, J = 8.1 Hz), 7.28 (2H, t, J = 7.6 Hz),
7.01 (1H, t, J = 7.1 Hz), 6.72 (2H, broad s), 2.95 (2H,
t, J = 7.8 Hz), 2.30 (2H, t, J = 7.6 Hz), 1.69 (2H, quintet,
J = 8.0 Hz), 1.59 (2H, quintet, J = 7.6 Hz), 1.40 (2H,

4340
T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
quintet, J = 7.8 Hz), 1.32 (2H, quintet, J = 7.1 Hz); MS
(EI) m/z: 284 (M+); Anal. Calcd for C13H20N2O3S: C,
54.91; H, 7.09; N, 9.85. C, 54.83; H, 7.18; N, 9.66.
(2H, quintet, J = 7.3 Hz), 1.42 (4H, m); MS (EI) m/z:
251 (M+).
5.1.27. 7-Methanesulfinylheptanoic acid phenylamide
(15). To a solution of 33 (80 mg, 0.32 mmol) obtained
above in CH2Cl2 (3 mL) was added 3-chloroperoxyben-
zoic acid (65%, 85 mg, 0.32 mmol). The mixture was
stirred at room temperature for 30 min. To the reaction
mixture was added saturated aqueous NaHCO3 and satu-
rated aqueous Na2S2O3 and the mixture was stirred at
room temperature for 1 h. The reaction mixture was
poured into water and extracted with CHCl3. The
CHCl3 layer was separated, washed with water and
brine, and was dried over Na2SO4. Filtration and con-
centration in vacuo and purification by silica gel flash
chromatography (AcOEt/MeOH = 10:1) gave 66 mg
(77%) of 15 as a crude solid. The solid was recrystallized
from n-hexane–AcOEt and collected by filtration to give
5.1.24. 7-Acetylsulfamoylheptanoic acid phenylamide (7).
Compound 7 was prepared from 16 obtained above and
acetic acid in 61% yield using the procedure described
for 6: mp 174–176 ꢁC; H NMR (DMSO-d6 400 MHz,
d; ppm) 11.60 (1H, broad s), 9.85 (1H, broad s), 7.58
(2H, d, J = 7.6 Hz), 7.28 (2H, t, J = 7.6 Hz), 7.01 (1H,
t, J = 7.3 Hz), 3.34 (2H, t, J = 7.8 Hz), 2.29 (2H, t,
J = 7.3 Hz), 1.65 (2H, quintet, J = 7.8 Hz), 1.57 (2H,
quintet, J = 7.3 Hz), 1.40 (2H, quintet, J = 7.6 Hz),
1.30 (2H, quintet, J = 7.6 Hz); Anal. Calcd for
C15H22N2O4SÆ1/10H2O: C, 54.89; H, 6.82; N, 8.54.
Found: C, 55.03; H, 6.83; N, 8.16.
1
5.1.25. 7-Bromoheptanoic acid phenylamide (32). 7-
Bromoheptanoic acid was prepared from 28 in 99%
yield using the procedure described for 4. In this case,
1
50 mg of 15 as colorless crystals: mp 121–122 ꢁC; H
NMR (CDCl3, 400 MHz, d; ppm) 7.65 (1H, broad s),
7.54 (2H, d, J = 7.8 Hz), 7.31 (2H, t, J = 7.6 Hz), 7.09
(1H, t, J = 7.3 Hz), 2.80–2.60 (2H, m), 2.56 (3H, s),
2.36 (2H, t, J = 7.4 Hz), 1.90–1.65 (4H, m), 1.65–1.35
(4H, m); MS (EI) m/z: 267 (M+); Anal. Calcd for
C14H21NO2S: C, 62.89; H, 7.92; N, 5.35. Found: C,
62.64; H, 7.89; N, 5.35.
1
LiOH was used instead of NaOH: H NMR (CDCl3,
400 MHz, d; ppm) 3.41 (2H, t, J = 6.8 Hz), 2.37 (2H, t,
J = 7.3 Hz), 1.87 (2H, quintet, J = 6.8 Hz), 1.66 (2H,
quintet, J = 7.6 Hz), 1.54–1.32 (4H, m).
To a suspension of 7-bromoheptanoic acid (2.64 g,
12.6 mmol) obtained above in CH2Cl2 (30 mL) were
added oxalyl chloride (1.65 mL, 18.9 mmol) and a cata-
lytic amount of DMF. The mixture was stirred at room
temperature for 2 h. The solvent was removed by evapo-
ration in vacuo to give acid chloride 31. To a solution of
aniline (3.50 g, 37.6 mmol) and triethylamine (5.30 mL,
38.1 mmol) in CH2Cl2 (40 mL) was added a solution
of 31 obtained above in CH2Cl2 (10 mL) dropwise cool-
ing in an ice-water bath. The mixture was stirred at
room temperature for 1 h. It was diluted with AcOEt
and washed with aqueous saturated NaHCO3, water
and brine, before being dried over MgSO4. Filtration
and concentration in vacuo, and purification by silica
gel flash chromatography (n-hexane/AcOEt = 3:1) gave
3.13 g (87%) of 32: 1H NMR (CDCl3, 400 MHz,
d; ppm) 7.51 (2H, d, J = 8.1 Hz), 7.32 (2H, t, J = 7.6 Hz),
7.15 (1H, broad s), 7.10 (1H, t, J = 7.6 Hz), 3.41 (2H, t,
J = 6.8 Hz), 2.36 (2H, t, J = 7.3 Hz), 1.87 (2H, quintet,
J = 7.1 Hz), 1.75 (2H, quintet, J = 7.3 Hz), 1.49 (2H, quin-
tet, J = 7.6 Hz), 1.41 (2H, quintet, J = 6.8 Hz).
5.1.28. 5-(6-Phenylcarbamoylhexylsulfanyl)pentanoic acid
ethyl ester (35). To a solution of a mixture of 32 (2.80 g,
9.85 mmol) in EtOH (30 mL) was added potassium thio-
acetate (1.70 g, 14.9 mmol). The mixture was stirred at
reflux temperature for 7 h. The reaction mixture was
poured into water and extracted with AcOEt. The
AcOEt layer was separated, washed with water, satu-
rated aqueous NaHCO3 and brine, and was dried over
Na2SO4. Filtration and concentration in vacuo and
purification by silica gel flash chromatography (n-hex-
ane/AcOEt = 5:2) gave 2.70 g (98%) of thioacetic acid
1
S-(6-phenylcarbamoylhexyl) ester as a crude solid: H
NMR (CDCl3, 400 MHz, d; ppm) 7.51 (2H, d,
J = 8.0 Hz), 7.32 (2H, t, J = 7.3 Hz), 7.22 (1H, broad
s), 7.10 (1H, t, J = 7.3 Hz), 2.86 (2H, t, J = 7.1 Hz),
2.35 (2H, t, J = 7.3 Hz), 2.32 (3H, s), 1.73 (2H, quintet,
J = 7.1 Hz), 1.59 (2H, quintet, J = 7.1 Hz), 1.40 (4H, m);
MS (EI) m/z: 279 (M+).
Compound 34 was prepared from 32 obtained above in
87% yield using the procedure described for 4: 1H NMR
(CDCl3, 400 MHz, d; ppm) 7.51 (2H, d, J = 8.0 Hz),
7.32 (2H, t, J = 7.6 Hz), 7.12 (1H, broad s), 7.10 (1H,
t, J = 7.1 Hz), 2.53 (2H, q, J = 7.3 Hz), 2.36 (2H, t,
J = 7.6 Hz), 1.74 (2H, quintet, J = 7.1 Hz), 1.63 (2H,
quintet, J = 7.1 Hz), 1.42 (4H, m), 1.33 (1H, t,
J = 7.8 Hz); MS (EI) m/z: 237 (M+).
5.1.26. 7-Methylsulfanylheptanoic acid phenylamide (33).
To a solution of 32 (300 mg, 1.06 mmol) in EtOH
(10 mL) was added methylmercaptan sodium salt (15%
in water, 1.50 g, 3.21 mmol) and the solution was stirred
at room temperature for 5 h. The reaction mixture was
diluted with AcOEt and washed with water and brine,
and was dried over MgSO4. Filtration and concentration
in vacuo, and purification by silica gel flash chromato-
graphy (n-hexane/AcOEt = 2:1) gave 262 mg (99%) of
33 as a crude solid. The solid was recrystallized from n-
hexane–AcOEt and collected by filtration to give
To a solution of 34 (1.0 g, 4.21 mmol) obtained above
and 5-bromovaleric acid ethyl ester (1.0 mL, 6.32 mmol)
in EtOH (15 mL) was added sodium ethoxide in EtOH
(20%, 1.60 g, 4.70 mmol) with cooling by an ice-water
bath. The solution was stirred overnight at room tem-
perature. The reaction mixture was poured into ice
water and extracted with AcOEt. The AcOEt layer
was separated, washed with water and brine, and was
1
217 mg of 33 as a colorless crystal: H NMR (CDCl3,
400 MHz, d; ppm) 7.51 (2H, d, J = 8.0 Hz), 7.32 (2H, t,
J = 7.8 Hz), 7.16 (1H, broad s), 7.10 (1H, t, J =
7.6 Hz), 2.49 (2H, t, J = 7.1 Hz), 2.36 (2H, t, J = 7.3
Hz), 2.09 (3H, s), 1.74 (2H, quintet, J = 7.3 Hz), 1.61

T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
4341
dried over Na2SO4. Filtration and concentration in va-
cuo and purification by silica gel flash chromatography
(n-hexane/AcOEt = 2:1) gave 1.39 g (91%) of 35 as a
(1H, t, J = 7.3 Hz), 3.08 (2H, t, J = 7.3 Hz), 3.05 (2H,
t, J = 7.3 Hz), 2.30 (2H, t, J = 7.3 Hz), 2.26 (2H, t,
J = 7.3 Hz), 1.73–1.52 (8H, m), 1.41 (2H, quintet,
J = 7.4 Hz), 1.33 (2H, quintet, J = 7.3 Hz); MS (EI)
m/z: 369 (M+); Anal. Calcd for C18H27NO5S: C, 58.51;
H, 7.37; N, 3.79. Found: C, 58.43; H, 7.42; N, 3.52.
1
white solid: H NMR (CDCl3, 500 MHz, d; ppm) 7.52
(2H, d, J = 7.9 Hz), 7.32 (2H, t, J = 7.6 Hz), 7.20 (1H,
broad s), 7.10 (1H, t, J = 7.4 Hz), 4.13 (2H, q,
J = 7.2 Hz), 2.53–2.48 (4H, m), 2.36 (2H, t,
J = 7.4 Hz), 2.32 (2H, t, J = 7.3 Hz), 1.76–1.70 (4H,
m), 1.63–1.57 (4H, m), 1.50–1.35 (4H, m), 1.25 (3H, t,
J = 7.1 Hz); MS (EI) m/z: 365 (M+).
5.2. Biology
5.2.1. HDAC inhibitory activity assay. The assay of
HDAC activity was performed using an HDAC fluores-
cent activity assay/drug discovery kit (AK-500, BIO-
MOL Research Laboratories). HeLa nuclear extracts
(0.5 lL/well) were incubated at 37 ꢁC with 25 lM of
Fluor de LysTM substrate and various concentrations of
samples. Reactions were stopped after 30 min by adding
Fluor de LysTM Developer with trichostatin A, which
stops further deacetylation. Then, 15 min after addition
of this developer, the fluorescence of the wells was mea-
sured on a fluorometric reader with excitation set at
360 nm and emission detection set at 460 nm, and the
% inhibition was calculated from the fluorescence read-
ings of inhibited wells relative to those of control wells.
The concentration of compound which results in 50%
inhibition was determined by plotting the log[Inh] versus
the logit function of the % inhibition. IC50 values of
SAHA and compound 15 are determined using a regres-
sion analysis of the concentration/inhibition data.
5.1.29. 5-(6-Phenylcarbamoylhexane-1-sulfinyl)pentanoic
acid (20). 5-(6-Phenylcarbamoylhexane-1-sulfinyl)penta-
noic acid ethyl ester was prepared from 35 obtained
above in 81% yield using the procedure described for
1
15: H NMR (CDCl3, 500 MHz, d; ppm) 7.53 (2H, d,
J = 7.9 Hz), 7.48 (1H, broad s), 7.31 (2H, t,
J = 7.7 Hz), 7.09 (1H, t, J = 7.3 Hz), 4.13 (2H, q,
J = 7.0 Hz), 2.69 (2H, quintet, J = 7.1 Hz), 2.64 (2H,
quintet, J = 6.7 Hz), 2.38–2.34 (4H, m), 1.83–1.75 (8H,
m), 1.60–1.40 (4H, m), 1.26 (3H, t, J = 7.0 Hz); MS
(EI) m/z: 381 (M+).
Compound 20 was prepared from 5-(6-phenylcarba-
moylhexane-1-sulfinyl)pentanoic acid ethyl ester ob-
tained above in 98% yield using the procedure
described for 4: mp 122–123 ꢁC; 1H NMR (CDCl3,
500 MHz, d; ppm) 7.85 (1H, broad s), 7.54 (2H, d,
J = 7.9 Hz), 7.30 (2H, t, J = 7.7 Hz), 7.09 (1H, t, J =
7.4 Hz), 2.78 (2H, m), 2.67 (2H, m), 2.39 (2H, t,
J = 6.5 Hz), 2.36 (2H, t, J = 7.6 Hz), 1.88–1.66 (8H,
m), 1.58–1.35 (4H, m); MS (EI) m/z: 353 (M+); Anal.
Calcd for C18H27NO4SÆ1/5MeOH: C, 60.74; H, 7.79;
N, 3.89. Found: C, 60.96; H, 7.73; N, 3.52.
5.2.2. Western blot analysis. HCT116 human colon can-
cer cells were purchased from American Type Culture
Collection (ATCC, Manassas, VA, U.S.A.) and cultured
in McCoy5A culture medium containing penicillin and
streptomycin, supplemented with fetal bovine serum as
described in the ATCC instructions. HCT-116 cells
(5 · 105) treated for 8 h with samples at the indicated
concentrations in 10% FBS-supplemented McCoyꢀs 5A
medium were collected and sonicated. Protein concen-
trations of the lysates were determined by using a Brad-
ford protein assay kit (Bio-Rad Laboratories);
equivalent amounts of proteins from each lysate were re-
solved in 15% SDS-polyacrylamide gel and then trans-
ferred onto nitrocellulose membranes (Bio-Rad
Laboratories). After blocking with Tris-buffered saline
(TBS) containing 3% skimmed milk for 30 min, the
transblotted membrane was incubated with hyperacetyl-
ated histone H4 antibody (Upstate Biotechnology)
(1:2000 dilution) or acetylated a-tubulin antibody (SIG-
MA) (1:2000 dilution) in TBS containing 3% skimmed
milk at 4 ꢁC overnight. After probed with the primary
antibody, the membrane was washed twice with water,
followed by incubation with goat anti-rabbit or anti-
mouse IgG-horseradish peroxidase conjugates (diluted
1:10000 or 1:5000) for 1.5 h at room temperature and
washed twice with water. The immunoblots were visual-
ized by enhanced chemiluminescence.
5.1.30. 5-(6-Phenylcarbamoylhexane-1-sulfonyl)pentanoic
acid (21). To a solution of 35 (600 mg, 1.64 mmol) in
CH2Cl2 (10 mL) was added 3-chloroperoxybenzoic acid
(65%, 960 mg, 3.62 mmol). The mixture was stirred
overnight at room temperature. To the reaction mixture
was added saturated aqueous NaHCO3 and saturated
aqueous Na2S2O3 and the mixture was stirred at room
temperature for 1 h. The reaction mixture was poured
into water and extracted with CHCl3. The CHCl3 layer
was separated, washed with water and brine, and was
dried over Na2SO4. Filtration and concentration in va-
cuo and purification by silica gel flash chromatography
(n-hexane/AcOEt = 1:3) gave 650 mg (99%) of 5-(6-phen-
ylcarbamoylhexane-1-sulfonyl)pentanoic acid ethyl ester
1
as a crude solid: H NMR (CDCl3, 500 MHz, d; ppm)
7.52 (2H, d, J = 8.0 Hz), 7.32 (2H, t, J = 7.6 Hz), 7.28
(1H, broad s), 7.10 (1H, t, J = 7.6 Hz), 4.14 (2H, q,
J = 7.0 Hz), 2.98–2.23 (4H, m), 2.38–2.34 (4H,
m),1.95–1.83 (4H, m), 1.81–1.73 (4H, m), 1.51 (2H,
quintet, J = 7.7 Hz), 1.45 (2H, quintet, J = 7 Hz), 1.26
(3H, t, J = 7.3 Hz); MS (EI) m/z: 397 (M+).
Compound 21 was prepared from 5-(6-phenylcarba-
moylhexane-1-sulfonyl)pentanoic acid ethyl ester ob-
tained above in 94% yield using the procedure
described for 4: mp 153–154 ꢁC; H NMR (DMSO-d6
500 MHz, d; ppm) 12.1 (1H, broad s), 9.85 (1H, s),
7.58 (2H, d, J = 8.6 Hz), 7.28 (2H, t, J = 7.7 Hz), 7.01
Acknowledgments
1
This work was supported in part by a grant from the
Health Sciences Foundation from the Ministry of
Health, Labor and Welfare of Japan.

4342
T. Suzuki et al. / Bioorg. Med. Chem. 13 (2005) 4332–4342
2002, 12, 3443; (c) Wada, C. K.; Frey, R. R.; Ji, Z.; Curtin,
References and notes
M. L.; Garland, R. B.; Holms, J. H.; Li, J.; Pease, L. J.;
Guo, J.; Glaser, K. B.; Marcotte, P. A.; Richardson, P. L.;
Murphy, S. S.; Bouska, J. J.; Tapang, P.; Magoc, T. J.;
Albert, D. H.; Davidsen, S. K.; Michaelides, M. R. Bioorg.
Med. Chem. Lett. 2003, 13, 3331; (d) Vasudevan, A.; Ji, Z.;
Frey, R. R.; Wada, C. K.; Steinman, D.; Heyman, H. R.;
Guo, Y.; Curtin, M. L.; Guo, J.; Li, J.; Pease, L.; Glaser,
K. B.; Marcotte, P. A.; Bouska, J. J.; Davidsen, S. K.;
Michaelides, M. R. Bioorg. Med. Chem. Lett. 2003, 13,
3909; (e) Suzuki, T.; Nagano, Y.; Matsuura, A.; Kohara,
A.; Ninomiya, S.; Kohda, K.; Miyata, N. Bioorg. Med.
Chem. Lett. 2003, 13, 4321; (f) Wu, T. Y. H.; Hassig, C.;
Wu, Y.; Ding, S.; Schultz, P. G. Bioorg. Med. Chem. Lett.
2004, 14, 449; (g) Vaisburg, A.; Bernstein, N.; Frechette,
S.; Allan, M.; Abou-Khalil, E.; Leit, S.; Moradei, O.;
Bouchain, G.; Wang, J.; Woo, S. H.; Fournel, M.; Yan, P.
T.; Trachy-Bourget, M.-C.; Kalita, A.; Beaulieu, C.; Li,
Z.; MacLeod, A. R.; Besterman, J. M.; Delorme, D.
Bioorg. Med. Chem. Lett. 2004, 14, 283; (h) Suzuki, T.;
Kouketsu, A.; Matsuura, A.; Kohara, A.; Ninomiya, S.;
Kohda, K.; Miyata, N. Bioorg. Med. Chem. Lett. 2004, 14,
3313; (i) Suzuki, T.; Matsuura, A.; Kouketsu, A.; Nakag-
awa, H.; Miyata, N. Bioorg. Med. Chem. Lett. 2005, 15,
331; (j) Chen, B.; Petukhov, P. A.; Jung, M.; Velena, A.;
Eliseeva, E.; Dritschilo, A.; Kozikowski, A. P. Bioorg.
Med. Chem. Lett. 2005, 15, 1389; (k) Suzuki, T.; Nagano,
Y.; Kouketsu, A.; Matsuura, A.; Maruyama, S.; Kuro-
taki, M.; Nakagawa, H.; Miyata, N. J. Med. Chem. 2005,
48, 1019.
1. (a) Grozinger, C. M.; Schreiber, S. L. Chem. Biol. 2002, 9,
3; (b) Kouzarides, T. Curr. Opin. Genet. Dev. 1999, 9, 40;
(c) Hassig, C. A.; Schreiber, S. L. Curr. Opin. Chem. Biol.
1997, 1, 300.
2. Taunton, J.; Hassig, C. A.; Schreiber, S. L. Science 1996,
272, 408.
3. Sambucetti, L. C.; Fischer, D. D.; Zabludoff, S.; Kwon, P.
O.; Chamberlin, H.; Trogani, N.; Xu, H.; Cohen, D. J.
Biol. Chem. 1999, 274, 34940.
4. Leoni, F.; Zaliani, A.; Bertolini, G.; Porro, G.; Pagani, P.;
`
Pozzi, P.; Dona, G.; Fossati, G.; Sozzani, S.; Azam, T.;
Bufler, P.; Fantuzzi, G.; Goncharov, I.; Kim, S. H.;
Pomerantz, B. J.; Reznikov, L. L.; Siegmund, B.; Dina-
rello, C. A.; Mascagni, P. Proc. Natl. Acad. Sci. U.S.A.
2002, 99, 2995.
5. (a) Yoshida, M.; Horinouchi, S.; Beppu, T. BioEssays
1995, 17, 423; (b) Richon, V. M.; Webb, Y.; Merger, R.;
Sheppard, T.; Jursic, B.; Ngo, L.; Civoli, F.; Breslow, R.;
Rifkind, R. A.; Marks, P. A. Proc. Natl. Acad. Sci. U.S.A.
1996, 93, 5705.
6. For a review, see: (a) Miller, T. A.; Witter, D. J.;
Belvedere, S. J. Med. Chem. 2003, 46, 5097; For a review,
see: (b) Yoshida, M.; Matsuyama, A.; Komatsu, Y.;
Nishino, N. Curr. Med. Chem. 2003, 10, 2351; For a
review, see: (c) Miller, T. A. Expert Opin. Ther. Pat. 2004,
14, 791.
7. Finnin, M. S.; Donigian, J. R.; Cohen, A.; Richon, V. M.;
Rifkind, R. A.; Marks, P. A.; Breslow, R.; Pavletich, N. P.
Nature 1999, 401, 188.
11. Mastrolorenzo, A.; Supuran, C. T. Met.-Based Drugs.
2000, 7, 49.
8. (a) Mulder, G. J.; Meerman, J. H. Environ. Health Perspect.
1983, 49, 27; (b) Vassiliou, S.; Mucha, A.; Cuniasse, P.;
Georgiadis, D.; Lucet-Levannier, K.; Beau, F.; Kannan,
R.; Murphy, G.; Knaeuper, V.; Rio, M. C.; Basset, P.;
Yiotakis, A.; Dive, V. J. Med. Chem. 1999, 42, 2610.
9. (a) Matsuyama, A.; Yoshimatsu, Y.; Shimazu, T.;
Sumida, Y.; Osada, H.; Komatsu, Y.; Nishino, N.;
Khochbin, S.; Horinouchi, S.; Yoshida, M. EMBO J.
2002, 21, 6820; (b) Haggarty, S. J.; Koeller, K. M.; Wong,
J. C.; Grozinger, C. M.; Schreiber, S. L. Proc. Natl. Acad.
Sci. U.S.A. 2003, 100, 4389; (c) Wong, J. C.; Hong, R.;
Schreiber, S. L. J. Am. Chem. Soc. 2003, 125, 5586; (d)
Glaser, K. B.; Li, J.; Pease, L. J.; Staver, M. J.; Marcotte,
P. A.; Guo, J.; Frey, R. R.; Garland, R. B.; Heyman, H.
R.; Wada, C. K.; Vasudevan, A.; Michaelides, M. R.;
Davidsen, S. K.; Curtin, M. L. Biochem. Biophys. Res.
Commun. 2004, 325, 683.
12. Breuer, E.; Salomon, C. J.; Katz, Y.; Chen, W.; Lu, S.;
Ro¨schenthaler, G. V.; Hadar, R.; Reich, R. J. Med. Chem.
2004, 47, 2826.
13. Gao, J.; Cheng, X.; Chen, R.; Sigal, G. B.; Bruce, J. E.;
Schwartz, B. L.; Hofstadler, S. A.; Anderson, G. A.;
Smith, R. D.; Whitesides, G. M. J. Med. Chem. 1996, 39,
1949.
14. Wang, D. F.; Wiest, O.; Helquist, P.; Lan-Hargest, H. Y.;
Wiech, N. L. J. Med. Chem. 2004, 47, 3409.
15. (a) Somoza, J. R.; Skene, R. J.; Katz, B. A.; Mol, C.; Ho,
J. D.; Jennings, A. J.; Luong, C.; Arvai, A.; Buggy, J. J.;
Chi, E.; Tang, J.; Sang, B.-C.; Verner, E.; Wynands, R.;
Leahy, E. M.; Dougan, D. R.; Snell, G.; Navre, M.;
Knuth, M. W.; Swanson, R. V.; McRee, D. E.; Tari, L. W.
Structure 2004, 12, 1325; (b) Vannini, A.; Volpari, C.;
Filocamo, G.; Casavola, E. C.; Brunetti, M.; Renzoni, D.;
Chakravarty, P.; Paolini, C.; Francesco, R. D.; Gallinari,
10. (a) Suzuki, T.; Ando, T.; Tsuchiya, K.; Fukazawa, N.;
Saito, A.; Mariko, Y.; Yamashita, T.; Nakanishi, O. J.
Med. Chem. 1999, 42, 3001; (b) Frey, R. R.; Wada, C. K.;
Garland, R. B.; Curtin, M. L.; Michaelides, M. R.; Li, J.;
Pease, L. J.; Glaser, K. B.; Marcotte, P. A.; Bouska, J. J.;
Murphy, S. S.; Davidsen, S. K. Bioorg. Med. Chem. Lett.
P.; Steinkuhler, C.; Marco, S. D. Proc. Natl. Acad. Sci.
¨
U.S.A. 2004, 101, 15064.
16. Hubbert, C.; Guardiola, A.; Shao, R.; Kawaguchi, Y.; Ito,
A.; Nixon, A.; Yoshida, M.; Wang, X.; Yao, T. Nature
2002, 417, 455.