Novel N-hydroxyfurylacrylamide-based histone deacetylase (HDAC) inhibitors with branched CAP group (Part 2)

Article abstract of DOI:10.1016/j.bmc.2013.06.009

Histone deacetylases (HDACs) are significant enzymes involved in tumor genesis and development. Herein, we report a series of novel N-hydroxyfurylacryl-amide-based HDAC inhibitors, which are marked by introducing branched hydrophobic groups as the capping group. The inhibitory activity of the synthesized compounds against HDACs and several tumor cell lines are firstly determined. Fifteen compounds with promising activities are selected for further evaluation of target selectivity profile against recombinant human HDAC1, HDAC4 and HDAC6. Compounds 10a, 10b, 10d and 16a exhibit outstanding selectivity against HDAC6. Analysis of HDAC4 X-ray structure and HDAC1, HDAC6 homology model indicates that these enzyme differ significantly in the rim near the surface of the active site. Although TSA has been known as a pan-HDAC inhibitor, it exhibits outstanding selectivity for HDAC6 over HDAC4. For further physicochemical properties study, six compounds are chosen for determination of their physicochemical properties including log D_7.4 and aqueous solubility. The results suggest that compounds with a smaller framework and with hydrophilicgroups are likely to have better aqueous solubility.

Full text of DOI:10.1016/j.bmc.2013.06.009

Bioorganic & Medicinal Chemistry 21 (2013) 5339–5354
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Novel N-hydroxyfurylacrylamide-based histone deacetylase (HDAC)
inhibitors with branched CAP group (Part 2)
Taotao Feng a,b, Hai Wang a,b, Hong Su a,b, Hui Lu a,b, Liqin Yu a,b, Xiaojin Zhang a,b,d, Haopeng Sun a,b,c,
,
⇑
Qidong You a,b,
⇑
a Jiangsu Key Laboratory of Drug Design and Optimization, China Pharmaceutical University, Nanjing 210009, China
b State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China
c Department of Medicinal Chemistry, School of Pharmacy, China Pharmaceutical University, Nanjing 210009, China
d Department of Organic Chemistry, School of Science, China Pharmaceutical University, Nanjing 210009, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Histone deacetylases (HDACs) are significant enzymes involved in tumor genesis and development.
Herein, we report a series of novel N-hydroxyfurylacryl-amide-based HDAC inhibitors, which are
marked by introducing branched hydrophobic groups as the capping group. The inhibitory activity of
the synthesized compounds against HDACs and several tumor cell lines are firstly determined. Fifteen
compounds with promising activities are selected for further evaluation of target selectivity profile
against recombinant human HDAC1, HDAC4 and HDAC6. Compounds 10a, 10b, 10d and 16a exhibit
outstanding selectivity against HDAC6. Analysis of HDAC4 X-ray structure and HDAC1, HDAC6 homol-
ogy model indicates that these enzyme differ significantly in the rim near the surface of the active site.
Although TSA has been known as a pan-HDAC inhibitor, it exhibits outstanding selectivity for HDAC6
over HDAC4. For further physicochemical properties study, six compounds are chosen for determina-
tion of their physicochemical properties including logD7.4 and aqueous solubility. The results suggest
that compounds with a smaller framework and with hydrophilicgroups are likely to have better aque-
ous solubility.
Received 15 April 2013
Revised 5 June 2013
Accepted 6 June 2013
Available online 15 June 2013
Keywords:
HDAC inhibitors
N-hydroxyfurylacrylamide
HDAC6 selectivity
Anti-tumor
Homology modelling
Molecular docking
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
acetylation of lysine residues, resulting in condensation of chromo-
somal DNA and gene transcriptional repression.7 In addition, many
Epigenetics refers to the heritable changes in gene expression or
phenotype that are stable between cell division, and sometimes
between generations, but do not involve changes in the underlying
DNA sequence of the organism.1,2 Epigenetic alterations, including
DNA methylation, histone modifications, nucleosome positioning,
and small noncoding RNAs (miRNA, siRNA), play an important role
in cancer progression.3–5 The histone proteins are widely described
as essential players in the control of gene expression via modifica-
tion through chemical reactions such as acetylation, phosphoryla-
tion, and methylation. The amino termini of histones extend from
the nucleosomal core and are modified by histone acetyltransfer-
ases (HATs) and histone deacetylases (HDACs) during the cell cy-
non-histone proteins have been identified to substrate of HDACs.
HDAC6 participates in the deacetylation of nonhistone proteins,
such as a-tubulin and HSP90, as well as regulating important bio-
logical processes including microtubule stability and function, and
molecular chaperon activity.8,9 Abnormality of histone deacetyla-
tion has been observed in human tumors, and inhibition of HDAC
has become a novel and validated therapeutic strategy against can-
cers.10 Therefore, HDACs are widely recognized as promising tar-
gets for therapeutic intervention in cancer.
Till now, eighteen HDACs have been reported and they can be
divided into four distant classes. Class I (HDAC1–3 and 8), class II
(HDAC4–7, 9 and 10) and class IV (HDAC11) HDACs are all Zn2+
-
cle.6 HATs transfer acetyl groups to the
e
-amino group of lysine
dependent metalloproteases, while class III (SIRT1–7) HDACs are
side chains of the histone protein, leading to local expansion of
chromatin and increased accessibility of regulatory proteins to
DNA, whereas HDACs oppose the effects of HATs and reverse the
NAD+-dependent.11,12
Currently, a great number of HDAC inhibitors, including SAHA,
ITF2357, MS-275, LAQ824, TSA, Tubacin, MGCD0103, FK228, are re-
ported as antitumor agents (Fig. 1). Among them, Vorinostat
(SAHA)13 and romidepsin (FK228)14 have been approved by US
FDA for the treatment of relapsed cutaneous T-cell lymphoma
(CTCL).15,16 Most HDAC inhibitors, as exemplified by Vorinostat,
can be described by the cap, linker, and zinc-binding group (ZBG).17
⇑
Corresponding authors. Tel./fax: +86 25 83271216 (H.S.); tel./fax: +86 25
83271351 (Q.Y.).
E-mail addresses: sunhaopeng@163.com (H. Sun), youqidong@gmail.com,
youqd@cpu.edu.cn (Q. You).
0968-0896/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2013.06.009

5340
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
linker
cap
ZBG
O
O
H
N
OH
N
H
O
N
H
NH2
H
N
O
N
Vorinostat(SAHA)
MS275
O
OH
N
O
O
O
OH
N
H
OH
N
H
N
N
H
LAQ824
TSA
Figure 1. The structures of known HDAC inhibitors.
Half of these inhibitors have the hydroxamic moiety, a typical
zinc-binding group. However, hydroxamic acid usually lacks selec-
tivity due to its strong zinc-binding ability, therefore, how to im-
prove the target selectivity of the HDACs inhibitors has been a
main problem that attracts the attentions of medicinal chemists
all over the world. Sequence aligenment of HDACs indicates that
the principal difference among the individual subtype exists in
the loops forming shallow grooves around the rim of enzyme pock-
et.18,19 Close examination of corresponding crystal structures fur-
ther indicates that these enzyme differ significantly in the rim
near the surface of the active site,20,21 which suggests that capping
group of HDAC inhibitors is quite important to the target selectiv-
ity. As a result, rational modification of capping group is a promis-
ing strategy for the improvement of selectivity of the HDAC
inhibitors.22 Several groups has reported HDAC selective inhibitors
with modified capping group (e.g., compounds 1, 2, 3 and 4,
Fig. 2).23–29 According to the structure–activity relationship (SAR)
data of these reported inhibitors, although the elements account
for selectivity are less understood,30,31 it seemed that the size of
capping group is the key point for governing enzyme selectivity.
Our laboratory has focused on the discovery of selective HDAC
inhibitors for a long time. In the previous work, we have designed
and synthesized a class of novel N-hydroxybenzamide-based
HDAC inhibitors with branched capping group, which exhibit
outstanding in vitro antitumor activities, and some of them
showed selectivity against HDAC1.30 With the aim to gain further
knowledge about the structural requirements for the selectivity
of the inhibitors, herein we report the design and synthesis of no-
vel N-hydroxyfurylacrylamide compounds with branched capping
group. Compared to the previouly reported compounds,30 the N-
hydroxyfurylacryl-amide was utilized as the zinc binding group.
The steric benzyl, 4-phenylbenzyl and homopiperonyl were taken
as the hydrophobic group so as to accommodate the binding pock-
et of HDACs. Another side chain at the amine N (represented by R
group in Schemes 1–3) in order to explore the interaction with the
shallow grooves near the rim of HDAC catalytic pockets.
The HDACs enzyme inhibitory activity as well as the anti-prolif-
erative effects of the synthesized compounds were initially
determined. These compounds displayed good HDAC inhibitory
activity and many of them had promising anti-proliferative activity
in several tumor cell lines. For further evaluation of our com-
pounds, fifteen compounds were retained for a target selectivity
profile study. Human HDAC1, HDAC4 and HDAC6 were used as
representative of class I, class IIa and class IIb HDACs, respectively.
To elucidate the potential binding mode of the inhibitors, docking
studies were performed using HDAC1, HDAC6 homology models
and HDAC4 X-ray structure. Through SAR study we found that
compounds with branched capping groups tend to selectively
inhibit HDAC6. Additionally, compounds with good selective
profiles on HDAC6 generally showed decreased antiproliferative
O
HN
S
O
O
O
N
N
N
H
HN
O
N
O
Ph
O
NH HN
O
O
Ph
2
1
O
O
NH
O
N
H
HN
O
NH2
H
N
O
S
NH
O
O
3
4
Figure 2. The structures of selective HDAC inhibitors with cyclic peptide or branched capping group.

T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
5341
activities against cancer cells, which was in accordance with previ-
ously reported results that HDAC6-selective compounds show re-
duced antiproliferative properties.32,33 Finally, Compound 10a,
10b, 10d, 10g, 10h and 13f that show excellent selectivity and
acceptable anti-proliferative activities were further evaluated for
their physicochemical properties including logD7.4 and intrinsic
solubility. Compounds 10b and 10d that show promising HDAC6
selective profile, drug-like properties which makes them ideal lead
compounds for the development of agents treating HDAC6-rele-
vant disease.
(10a–10d) displayed good potency against the enzymes. Replacing
alkyl substitutes with longer alkyl substitutes (10e–10g) resulted
in decreased inhibitory activity, which is consistent with our previ-
ous result that the longer alkyl substitutes might have weaker
affinity.30 We further examined analogs with phenoxyalkyl substi-
tutes to investigate the effects of different substitutions on the
inhibition of HDACs. Modification of the R alkyl group of 10a,
10b, 10c seemed detrimental to the activity. In detail, incorporat-
ing phenoxy to the ethyl (10a and 10i) and replacing the butyl with
aromatic groups (10d and 10l) can cause the decrease in the inhib-
itory effects. Compound 10h bearing 2-hydroxy-ethyl showed
noticeable HDACs inhibitory activity, indicating that the hydroxy-
ethyl unit may interact with the hydration shell of aspartic acidglu-
tamic acid in the rim of the HDAC binding channel.34
2. Result and discussion
2.1. Chemistry
Inorder to investgate the effect of capping group upon HDAC
activity, we synthesized compounds 13a–13l and 16a–16k. When
introduced a hydrophobic benzene ring into capping group, 13a–
13l displayed moderate HDAC inhibitory activity. This might be
attributed to unfavorable steric effect of 4-phenylbenzyl substitu-
tion. In this series, compound 13f inhibited the HDAC activity with
The synthetic route of compounds 10a–10n was shown in
Scheme 1. In detail, furfural 5 was treated with triethyl phospho-
nacetate in absolute ethanol containing anhydrous potassium car-
bonate to give (E)-ethyl-3-(furan-2-yl)acrylate 6. (E)-Ethyl-3-(5-
formylfuran-2-yl)acrylate
7 was prepared through Vilsmeier–
an IC50 = 0.089 lM, which is consistant with the above mentioned
Hacck reaction. Benzylamine was reacted with 7 to give Schiff’s
base, which was further reduced with NaBH4 to provide intermedi-
ate 8. Compound 8 was alkylated with various alkyl bromides to
generate 9a–9n, which was subsequently reacted with hydroxyl-
amine to give target analogs 10a–10n.
For the synthesis of compounds 13a–13l, 4-phenylbenzylamine
was treated with compound 7 to give Schiff’s base, which was re-
duced with NaBH4 to provide intermediate 11. Compound 11 was
alkylated with various alkyl bromides to generate 12a–12l, which
was subsequently reacted with hydroxylamine to give desired ana-
logs 13a–13l.
The compounds 16a–16k were synthesized according to the
methods described in Scheme 1. Homopiperonylamine was treated
with compound 7 to give Schiff’s base, which was reduced with
NaBH4 to provide intermediate 14. Compound 14 was alkylated
with various alkyl bromides to generate 15a–15k, which was sub-
sequently reacted with hydroxylamine to give desired analogs
16a–16k.
hypothesis that the 2-hydroxy-ethyl substitution may interact
with the hydration shell of aspartic acid and glutamic acid in the
rim of the HDAC binding channel.34
Compound 16a–16k, having two methylene groups between the
basic N and the homopiperonyl, resulted in the decreased inhibi-
tory activity. In comparison with the activity variation of 13a–
13l, the decreased inhibitory activity was owing to the hydropho-
bic homopiperonyl and the longer carbon chain. Interestingly,
compound 16g bearing 2-hydroxy-ethyl substitution also dis-
played maximum HDAC inhibitory activity with an IC50 = 0.40
in this series.
lM
Anti-proliferative activities of all compounds were evaluated on
four tumor cell lines including PC3 (prostatecancer), HCT116 (co-
lon cancer), A549 (lung cancer) and HepG2 (liver cancer). Anti-pro-
liferative cytotoxic activity was measured by MTT [3-(4,5-
dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide] as-
say. The obtained EC50 values of the types of tumer cell lines com-
pared with SAHA are summarized in Table 1. In series 10a–10n,
although analogs 10a–10d displayed good HDAC inhibitory activ-
ity, they just showed moderate growth inhibitory activity to four
human tumor cell lines. For examples, analogs 10b and 10d exhib-
ited moderate inhibitory against four cell lines with EC50 values
2.2. Biological evaluation
The HDACs enzymes inhibitory activities of these compounds
were determined in vitro using the commercially available Drug
Discovery Kit AK500 and the results are summarized in Table 1.
For compounds 10a–10n, analogs bearing short alkyl substitutes
ranging from 16.72 to 25.03 lM and 14.45 to 24.30 lM,
respectively. This might be ascribed to enzyme isoform selectivity
O
a
R1
b
c
O
O
O
OHC
N
H
CHO
O
O
O
O
O
O
O
8
11
5
6
7
14
O
O
R1
N
R1
O
d
O
e
OH
N
R
N
H
R
9a-9n
10a-10n
12a-12l
13a-13l
16a-16k
15a-15k
Scheme 1. Synthesis of N-hydroxyfurylacrylamides 10a-10n, 13a-13l, 16a-16k. Reagents and conditions (a) (C2H5)2P(O)CH2CO2C2H5, K2CO3, EtOH, 82%; (b) POCl3, DMF,
82.5%; (c) (1) R1NH2, CH3OH, reflux, 3 h; (2) CH3OH, NaBH4, 43.7%-76.8%; (d) K2CO3, KI, CH3CN, RX, 35 °C, 36.1%-96.1%; (e) NH2OH, KOH, CH3OH, rt, 36.2–46.0%.

5342
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
Table 1
HDACs inhibitory activities and Anti-proliferative activities of indicated compounds
Compd
R
R1
HDACs IC50
(
lM)
PC3 EC50
(l
M)
HCT116 EC50
(lM)
A549 EC50
(l
M)
HepG2 EC50 (lM)
10a
10b
10c
10d
10e
10f
10g
10h
10i
Ethyl
Propyl
Iso-propyl
Butyl
Iso-butyl
Hexyl
Octyl
2-Hydroxy-ethyl
2-Phenoxyethyl
2-(p-Tolyloxy)ethyl
3-Phenoxypropyl
3-(p-Tolyloxy)propyl
4-Phenoxybutyl
4-(p-Tolyloxy)butyl
Ethyl
Iso-propyl
Amyl
Octyl
Benzyl
2-Hydroxy-ethyl
2-Phenoxyethyl
2-(p-Tolyloxy)ethyl
3-Phenoxypropyl
3-(p-Tolyloxy)propyl
4-Phenoxybutyl
4-(p-Tolyloxy)butyl
Ethyl
Propyl
Butyl
Iso-butyl
Amyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
Benzyl
4-Phenylbenzyl
4-Phenylbenzyl
4-Phenylbenzyl
4-phenylbenzyl
4-Phenylbenzyl
4-Phenylbenzyl
4-Phenylbenzyl
4-Phenylbenzyl
4-Phenylbenzyl
4-phenylbenzyl
4-Phenylbenzyl
4-Phenylbenzyl
Homopiperonyl
Homopiperonyl
homopiperonyl
Homopiperonyl
Homopiperonyl
Homopiperonyl
Homopiperonyl
Homopiperonyl
Homopiperonyl
Homopiperonyl
Homopiperonyl
—
0.13
0.17
0.18
0.54
3.17
2.65
2.55
0.17
>2.55
1.87
1.48
2.54
2.12
1.31
1.25
1.61
>2.39
>2.17
>2.55
0.089
1.21
1.71
>2.07
1.66
>2.02
>1.96
2.31
1.59
>2.69
>2.69
3.49
2.28
0.40
104.33
25.03
26.91
20.73
18.84
23.79
42.32
>138.12
>127.55
51.72
44.85
42.17
35.86
15.69
67.87
24.15
13.42
36.83
14.31
15.91
15.90
18.82
25.22
18.06
23.61
37.65
66.48
7.57
6.37
24.90
22.68
18.32
26.31
8.96
38.37
23.25
34.52
14.45
20.46
11.04
6.59
97.18
52.32
52.59
25.76
7.95
10.62
9.82
8.40
15.59
14.71
28.80
69.82
8.36
26.45
23.03
16.60
11.49
22.36
15.57
84.68
50.61
78.23
39.34
72.51
35.39
87.92
37.39
27.04
16.29
46.83
8.30
>166.67
16.72
67.52
24.30
27.68
21.80
3.96
128.12
44.72
12.96
14.95
5.95
14.74
16.29
12.85
15.51
19.71
36.93
66.61
22.56
18.10
19.40
12.74
11.65
32.90
15.92
81.66
28.72
54.33
38.01
65.34
73.20
104.94
68.26
66.71
109.53
97.69
12.20
0.47
12.07
20.08
14.83
7.24
6.02
6.43
5.62
1.84
2.74
8.23
10.43
9.01
4.02
9.53
6.93
2.05
2.22
2.76
2.76
65.96
15.87
16.99
17.06
74.79
91.40
69.50
30.55
28.47
37.24
35.88
0.57
10j
10k
10l
10m
10n
13a
13b
13c
13d
13e
13f
13g
13h
13i
13j
13k
13l
16a
16b
16c
16d
16e
16f
16g
16h
16i
33.20
16.14
37.15
34.51
92.50
23.72
18.67
18.80
15.54
>166.67
Benzyl
2-Hydroxy-ethyl
2-Phenoxyethyl
2-(p-Tolyloxy)ethyl
3-Phenoxypropyl
4-Phenoxybutyl
—
2.36
0.84
>2.22
1.57
0.053
16j
16k
SAHA
and cellular membrane permeability. Compared to other cell lines,
most of the compounds showed potent inhibitory effect on
HCT116, the most potent analogs 10g exhibited very strong growth
inhibition with EC50 of 0.47 lM. The data indicate that these new
analogs are more sensitive on the HCT116 cells.
ring to the end of alkyl resulted in decreased inhibitory activity
against HDAC6 (10a vs 10j, 13a vs 13h, 16a vs 16i). The isoform
selectivity analysis shows that capping group of HDAC inhibitors
is quite important to selectivity. Tubacin, the HDAC6 selective
inhibitor with bulk capping group displays seven fold selectivity
for HDAC6 over HDAC136 and has been used widely in cell biology
studies.37,38 As a comparision, compound 10b, the most selective
inhibitor, displayed 60-fold selectivity for HDAC6 over HDAC4
and 20-fold selectivity for HDAC6 over HDAC1. Therefore, the data
indicate that isoform selective inhibitors can be generated with
bulk branched capping group. This is different from the N-
hydroxybenzamide-based HDAC inhibitors, which displayed
HDAC1 selectivity. Compounds in this study are much more conju-
gated, compared to N-hydroxybenzamide group, by inserting eth-
ylene link between the C2-furan position and the carbonyl group
of the hydroxamic acid. We speculated that the conjugated struc-
ture may contribute to the selectivity for directing the cap group of
the inhibitor to interact with the rim of the catalytic pocket of the
HDAC enzymes. On the other hand, Mai et al. have disclosed a ser-
ies of aroyl-pyrrolyl-hydroxy-amides (APHAs).39,40 Some APHAs
showed distinct selectivity against HD1-B and HD1-A, two maize
deacetylases that are homologous of mammalian class I and class
IIa HDACs, respectively. The most selective APHAs showed no
inhibitory activity against HDAC1 but was able to inhibit HDAC4.
Though APHAs and our compounds have the similarly conjugated
structure, the branched cap group of our compounds are more
flexible than APHPs. This branched cap group may contribute to
the selectivity for comfortably occupying the shallow groove at
the rim of the catalytic pocket of the HDAC enzymes. To elucidate
Compound series 13a–13l showed preferable growth inhibitory
activity to series 10a–10n. All compound in this series exhibited
more potent growth inhibitory activity relative to SAHA on PC3 cell
line. Compound 13a were comparable to SAHA on A549 and HepG2
cell line. The compound series 16a–16k which have moderate
HDAC inhibitory activity showed poor growth inhibitory activity
to four human tumor cell lines, while some compounds were active
on the PC-3 cell lines.
We further selected 15 compounds with acceptable activities
on both enzyme inhibition and anti-proliferation to evaluate the
possible class specificity (Table 2). Human HDAC1, HDAC4 and
HDAC6 were taken as markers for class I, class IIa and class IIb
HDACs, respectively.35 These compounds exhibited unexpected
potent HDAC6 inhibition with IC50 of ranging from 0.18 to
4.65
lM, but displayed poor inhibitory activity against HDAC1
and HDAC4 with IC50 of 2.37–31.38
l
M and 3.96–68.11 M,
l
respectively. Therefore, these compounds are highly selective
and potent inhibitors of HDAC6, especially compounds 10a, 10b,
10d and 16a. Analogs bearing different capping group (benzyl, 4-
phenylbenzyl and homopiperonyl) showed comparable HDAC6
inhibitory activity (compound 10a, 13a and 16a). For compounds
bearing alkyl substitutes (compound 10a, 10d, 10e and 10h),
the order of their IC50 values against HDAC6 was
n-butyl < ethyl < hydroxyethyl < isobutyl. Incorporating benzene

T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
5343
Table 2
HDAC1, HDAC4 and HDAC6 inhibitory activities, log D7.4 and intrinsic solubility of indicated compounds
Compd
HDAC1 IC50
(l
M)
HDAC4 IC50
(l
M)
HDAC6 IC50
(lM)
HDAC1/HDAC6
HDAC4/HDAC6
logD, pH 7.4
Intrinsic solubility (mM)
10a
10b
10c
10d
10e
10g
10h
10i
10j
10k
10l
10n
13a
13f
13h
16a
16i
5.44
4.95
2.37
4.69
20.62
ND
12.71
20.23
31.38
6.48
9.66
4.73
3.42
ND
10.73
10.63
3.96
7.26
12.32
ND
11.25
50.56
68.11
12.29
13.22
12.64
5.721
ND
0.25
0.18
0.34
0.23
1.10
ND
0.43
1.93
3.13
0.79
0.93
0.55
0.83
ND
21.44
28.15
6.94
20.01
18.68
42.26
60.50
11.63
30.94
11.16
1.95
2.54
ND
2.97
ND
5.28
1.62
ND
ND
ND
ND
ND
ND
1.80
ND
ND
ND
3.00
2.80
ND
2.30
ND
1.90
4.00
ND
ND
ND
ND
ND
ND
3.11
ND
ND
ND
29.68
10.48
10.03
8.21
10.41
8.63
26.27
26.18
21.77
15.59
14.24
23.06
6.87
4.11
4.09
7.88
17.80
0.31
0.0049
9.07
1.14
0.48
4.65
>100
0.0016
3.59
16.57
3.83
7.95
41.93
11.29
19.95
52.45
>100
0.085
MS-275
TSA
ND
ND
ND
ND
3.06
53.12
ND = not determined.
the selectivity of compound 10b for HDAC6 over HDAC1 and
HDAC4, we constructed the HDAC1 and HDAC6 homology model.
The homology model was constructed by Discovery Studio 3.0
using the Homology Modeling protocol as the method described
in the literature.41 Analysis of HDAC4 X-ray structure and HDAC1,
HDAC6 homology modeled catalytic pockets reveals that, while
the active site is highly conserved, the shallow grooves S of the
catalytic channel rim differ greatly among the three isozymes.
Two regions, named A and B, represent boundaries of shallow
grooves S rim (Fig. 3). Region A corresponds to Leu262 in HDAC1,
Leu299 in HDAC4 and Leu274 in HDAC6; Region B corresponds to
Phe196 in HDAC1, Phe227 in HDAC4 and, Phe205 in HDAC6. Com-
pared to Phe196 in HDAC1 and Phe227 in HDAC4, Phe205 in
HDAC6 orients in an opposite direction. Variation at the B region
produces a significantly wider groove rim in the homology model
of HDAC6. The structural manifestation of this variation is that the
measured A–B distance is 8.75 Å in HDAC1 and 8.66 Å in HDAC4,
compared to10.98 Å in HDAC6. In order to understand the binding
mode of the HDAC6 selective inhibitor, compound 10b was docked
into the active site of HDAC1, HDAC6 homology model and HDAC4
X-ray structure using CDOCKER module inbuilt in Discovery Studio
3.0. Data show that compound 10b with bulk branched capping
group is large enough to comfortably occupy the groove S in
HDAC6, but not HDAC1 and HDAC4. In the mode of the
10b–HDAC6 complex, the carbonyl and hydroxyl oxygens of
hydroxamate could chelate with the zinc ion, zinc ion was also
coordinated to two aspartic acids (Asp267, Asp174) and two histi-
dine (His136, His135). The furan ring was bounded in the tubular
hydrophobic pocket (Fig. 4).The ‘cap’ of 10b was found to interact
with the rim of the HDAC binding channel and the hydrophobic
Figure 3. Top-down view of active site for HDAC1, HDAC4 and HDAC6. Distances between boundaries of the grooves S rim are shown with arrows. Letters A–B denote two
boundary areas surrounding the grooves rim.

5344
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
Figure 4. The docking modes of compounds 10b with HDAC4 X-ray structure and HDAC1, HDAC6 homology models. The protein is represented by molecular surface. Letters
A–B denote two boundary areas surrounding the grooves S. Compound 10b and zinc ion is indicated green.
surface. The aromatic benzene ring of 10b showed favorable
p
–
p
and two histidine (His159, His158). However, the carbonyl and hy-
droxyl oxygens of hydroxamate did not chelate with the zinc ion.
The N-hydroxyfurylacrylamide group had little direct contact with
the enzyme. Weak van der Waals interactions of the hydrophobic
propyl group with Leu299 could be observed. The furan group of
stacking interaction with Phe204. Favorable van der Waals interac-
tions of the aromatic benzene ring with Pro273 and Leu274 were
observed. Favorable van der Waals interactions of the hydrophobic
propyl group of 10b with Pro206 and Met207 were also observed
at the rim of groove S. The furan ring could form hydrophobic
interaction with Leu274. Therefore, the bulk branched capping
group gives the extra stabilization energy by occupying the shal-
low groove S to form stronger hydrophobic interaction. These
observations can explain the high binding energy of 10b with
HDAC6 (–CDOCKER energy 42.88 kcal/mol). In the mode of the
10b–HDAC1 complex, 10b completely inserted into the long inter-
nal cavity, forming an unsuitable molecular conformation and
unpleasant binding energy (–CDOCKER energy À35.33 kcal/mol).
Compound 10b was surrounded by the hydrophobic residue
Ala127, Gly128, Gly129 and Leu130. Arg25 could form hydrogen
bonds with the hydroxamic acid of the inhibitors. The zinc ion
was stably coordinated to two aspartic acids (Asp255, Asp167)
and two histidine (His132, His131). However, the carbonyl and hy-
droxyl oxygens of hydroxamate did not chelate with the zinc ion.
Additionally, it did not interact with the rim of the HDAC binding
channel and the hydrophobic surface. As for HDAC4, 10b also bin-
ded with an unrational conformation, which showed an reverse
orientation compared to that in the 10b–HDAC6 complex. The zinc
ion was stably coordinated to two aspartic acids (Asp196, Asp290)
10b could form p–p interaction with Phe227. In addition, the shal-
low groove S of HDAC4 was also not occupied by 10b, which was
ascribed to the decreased binding energy (19.29 kcal/mol). Taken
together, the comparison of the binding conformations and the
interaction energies in the three complexes has confirmed the tar-
get selective profile of our compound.
Interestingly, The HDAC4 IC50/HDAC6 IC50 value of TSA is 53,
although TSA has been known as a pan-HDAC inhibitor. To better
understand the selectivity of TSA for HDAC6 over HDAC4, TSA
were docked into HDAC4 X-ray structure and HDAC6 homology
model. Analysis of HDAC4 binding mode showed that the
hydroxamic acid interacted with the zinc ion and polar residue
His198 (Fig. 5). Phe168 and Phe227 sandwiched the hydrophobic
alkyl chain of the inhibitors. The methyl group near the benzene
ring could form hydrophobic interaction with Pro298 and Leu299.
The cap group of TSA was outside the peptide binding groove and
did not make any interactions. Additional, the shallow groove S of
HDAC4 was not occupied by TSA. The analysis of the binding
mode obtained for the HDAC6 homology model showed that
His136 could form hydrogen bonds with the hydroxamic acid of

T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
5345
Figure 5. The docking modes of compounds TSA with HDAC4 X-ray structure and HDAC6 homology models. The protein is represented by molecular surface. Letters A-B
denote two boundary areas surrounding the grooves S. Compound TSA is indicated magenta. Zinc ion is indicated green.
the inhibitors. Phe145 and Phe204 in HDAC6 sandwiched the al-
kyl chain linker between their hydrophobic portions. The aro-
matic cap group of TSA could form hydrophobic interaction
with Pro273 and Leu274. In addition, the shallow groove S of
HDAC6 was also not occupied by TSA. Docking analysis showed
that there was no significant difference in TSA–HDAC binding
mode between HDAC4 and HDAC6 at the entrance of the binding
pocket. Also, the binding mode of TSA was diferent from com-
pound 10b. Then, we analysis the chelation geometry of the cen-
tral zinc cation to explore this selectivity of TSA (Fig. 6). The
measured distance between carbonyl and hydroxyl oxygens of
hydroxamate and zinc ion is 2.18 Å, 4.19 Å in HDAC4 and
2.70 Å, 2.55 Å in HDAC6. HDAC6 ligand–protein complexes shows
the expected bidentate chelation geometry of the central zinc cat-
ion. However, the observed geometry in the HDAC4 structures is
more in line with the weaker monodentate binding mode of the
central zinc cation. Therefore, the comparison of the chelation
mode of hydroxamate with the zinc ion in the two complexes
has confirmed the target selective profile of TSA.
structures of HDAC4 (PDB ID: 2VQM) and HDAC7 (PDB ID: 3C0Z
and 3C10) bound to hydroxamate inhibitors. None of the ligand–
protein complexes showed the expected bidentate chelation geom-
etry of the central zinc cation, as observed in the structures of li-
gand-bound human HDAC8 (PDB ID: 1T64 and 1T69) and
bacterial homologs (PDB ID: 1ZZ1). The observed geometry in the
published HDAC4 and HDAC7 structures, however, was more in
line with the weaker monodentate binding mode of the neutral
form of the hydroxamic acid.42 This could also help us to under-
stand the selectivity of TSA in our study.
In an effort to identify potential drug like compounds, six com-
pounds were chosen for determination of their physicohemical
properties including logD7.4 and aqueous solubility. The partition
coefficient (logD) at pH 7.4 and aqueous solubility were deter-
mined according to the method of Avdeef and Tsinman43 on a
Gemini Profiler instrument (pION) by the ‘goldstandard’ Av-
deefÀBucher potentiometric titration method.44
As shown in Table 2, all six compounds displayed good aqueous
solubility. Generally, the introduction of hydrophilic groups re-
sulted in a significant increase in solubility. For instance, compar-
ing the solubility of 10h and 10a, showed that the former
compound were more soluble than the latter compound. Introduc-
tion of hydrophobic aliphatic chain with bulk volume led to poor
On the other hand, Bradner et al. described that hydroxamates
such as vorinostat, belinostat, and panobinostat showed much re-
duced potency against class IIa enzymes including HDAC4. They
interpreted this observation based on the available crystal
Figure 6. Examination of Zinc chelation by TSA. Compound TSA is indicated magenta. Zinc ion is indicated green.

5346
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
solubility. For examples, compound 10g with octyl group displayed
reduced aqueous solubility compared to the compound 10a with
ethyl substituent. Furthermore, it was shown that compound 10h
with a benzyl as capping group were more soluble than compound
13f with a 4-phenylbenzyl substituent. There was significant neg-
ative correlation between the logD7.4 and aqueous solubility of the
six tested compounds. These results suggest that compounds with
a smaller framework and with hydrophilic groups are likely to have
better aqueous solubility.
HDAC6-relevant disease such as autoimmune disorders, neurode-
generative disorders.
4. Experimental
4.1. General
Melting points were determined on a Mel-TEMP II melting point
apparatus and are uncorrected. Infrared (IR) spectra (KBr) were re-
corded on a Nicolet Impact 410 instrument (KBr pellet). 1H NMR
spectra were recorded with a Bruker Avance 300 MHz spectrome-
ter at 300 K, using TMS as an internal standard. MS spectra were
recorded on a Shimadzu GC–MS 2010 (EI) or a Mariner Mass Spec-
trum (ESI), or a LC/MSD TOF HR-MS Spectrum. All compounds were
routinely checked by TLC and 1H NMR. TLCs and preparative thin-
layer chromatography were performed on silica gel GF/UV 254, and
the chromatograms were performed on silica gel (200–300 mesh)
visualized under UV light at 254 and 365 nm. All solvents were re-
agent grade and, when necessary, were purified and dried by stan-
dards methods. Concentration of solutions after reactions and
extractions involved the use of a rotary evaporator operating at a
reduced pressure of ca. 20 Torr. Organic solutions were dried over
anhydrous sodium sulfate. Analytical results are within (0.40% of
the theoretical values).
3. Conclusions
We have synthesized and evaluated a series of hydroxamic
acids with branched capping group. Although the HDACs inhibi-
tory activity of most compounds is weaker than SAHA, com-
pounds 13f exhibited comparable HDACs inhibitory activity
with SAHA. Interestingly, compounds (10h, 13f, 16g) with
hydroxyethyl unit all displayed maximum HDACs inhibitory
activity. Docking study on LAQ824 carried out on homology mod-
eling of HDAC1 revealed that the hydroxyethyl unit was found to
interact with the hydration shell of aspartic acid and glutamic
acid in the rim of the HDAC binding channel.34 Thus, the hydroxy-
ethyl group is crucial in enzyme inhibitory activity. We also eval-
uated anti-proliferative activities of all compounds on four tumor
cell lines using PC3, HCT116, A549 and HepG2. These compounds
displayed good HDAC inhibitory activity, and many of them had
promising anti-proliferative activity in several tumor cell lines.
We next tested the isoform selectivity of fifteen selected com-
pounds against human HDAC1, HDAC4 and HDAC6. Compounds
10a, 10b, 10d and 16a demonstrated outstanding selectivity
against HDAC6. Furthermore, previously reported works indicated
that single-treatment of HDAC6 selective inhibitors did not show
anti-proliferative activity in general.32,33 On the other hand,
although compounds 10a, 10b and 10d demonstrated outstand-
ing selectivity against HDAC6, their HDAC1 inhibitory activity
was not vanished. The two reasons could explain why compounds
with good selective profiles on HDAC6 showed decreased, but not
lost, antiproliferative activities on tumor cells. Homology model
and docking analysis showed that compound 10b with bulk
branched capping group was large enough to comfortably occupy
the groove S in HDAC6, but not HDAC1 and HDAC4. These results
emphasized the importance of cap group for the target selectivity
and provide guidance for the design of HDAC6 selectivity inhibi-
tors. In an effort to identify potential drug like compounds, six
compounds that show excellent selectivity and acceptable anti-
proliferative activities were chosen for determination of their
physicohemical properties including logD7.4 and aqueous solubil-
ity. Compound 10b and 10d displayed good aqueous solubility
with an intrinsic solubility of 2.80 mM and 2.30 mM. The results
suggest that compounds with a smaller framework and with
hydrophilic groups are likely to have better aqueous solubility.
Creating isoform selectivity HDAC inhibitors has been challeng-
ing.45 However, selective inhibitors would provide powerful
chemical tools to elucidate the individual functions of the HDAC
isoforms.37 In addition, isoform selective inhibitors have the po-
tential to improve therapeutic outcomes in the clinic compared
to broad spectrum inhibitors. Unlike other histone deacetylase,
inhibition of HDAC6 does not appear to be associated with any
serious toxicity, making it an excellent drug target.44 Despite
the involvement of HDAC6 in disease states, there are relativity
few examples of HDAC6 selective inhibitors.46–50 Therefore, com-
pounds 10b and 10d that show promising HDAC6 selective pro-
file, drug-like properties which makes them ideal lead
compounds for the development of agents treating other
4.1.1. (E)-Ethyl 3-(furan-2-yl)acrylate (6)
Furfural (5, 0.1 mol, 8.28 mL) in absolute ethanol (10 mL) was
added in one portion to a mixture of triethyl 2-phosphonobutyrate
(0.12 mol, 23.8 mL) and anhydrous potassium carbonate (0.3 mol,
41.4 g) in absolute ethanol (100 mL). After being stirred at 70 °C
for 2 h, the reaction mixture was cooled to room temperature, fil-
trated and washed with ethanol (2 Â 20 mL). The filtrate was con-
centrated and distilled under reduced pressure to give pure
yellowish solid (13.6 g) in 82% yield; mp 24–26 °C [lit.51 24 °C].
4.1.2. (E)-Ethyl 3-(5-formylfuran-2-yl)acrylate (7)
POCl3 (0.13 mol, 11.9 mL) was added to a cooled (0 °C) mixture
of (E)-ethyl 3-(furan-2-yl)acrylate (0.12 mol, 19.6 g) and DMF
(0.13 mol, 10.0 mL) over a period of 20 min. After stirring at
60 °C for 2 h, the mixture was cooled to room temperature and
poured onto crushed ice (200 g). The pH of the mixture was ad-
justed to 7 with 10% NaOH. The resulted precipitate was filtered
and purified to give brown solid (19.2 g) in 82.5% yield by crystal-
lization with a mixture of ethanol and water; mp 73.4–74 °C [lit.52
74 °C].
4.1.3. General procedure for the synthesis of compounds 8, 11,
14
4.1.3.1. (E)-Ethyl 3-{5-[(benzylamino)methyl]furan-2-yl}acry-
late (8).
A solution of 7 (9.7 g, 0.05 mol) and benzylamine
(5.5 mL, 0.05 mol) in methanol (50 mL) was refluxed for 3 h. After
cooling to room temperature, NaBH4 (1.9 g, 0.05 mol) was added
slowly in three batches to the mixture and stirred for 30 min.
The solution was evaporated to dryness and treated with water
(50 mL). The mixture was extracted with ethyl acetate
(2 Â 30 mL). The combined organic solution was washed with
brine (10 mL) and the solution was treated with hydrochloric acid
to give hydrochloride of the product. The precipitated was filtered
and washed with ethyl acetate. The filter cake was moved into a
beaker, 2 N NaOH was added into the beaker. Stir until the solid
dissolve. The resulted mixture was extracted with ethyl acetate
(50 mL Â 2). The combined organic phase was washed with satu-
rated brine (10 mL), dried and evaporated to dryness to furnish
the product (9.1 g). Yield: 63.8%; oil. MS (EI) m/z 285 (M).

T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
5347
4.1.3.2. (E)-Ethyl 3-{5-[(biphenyl-4-ylmethylamino)methyl]-
furan-2-yl}acrylate (11). Compound 11 (15.25 g, 76.8% yield)
was synthesized from 7 (10.7 g, 0.055 mol) and 4-biphenylethyl-
amine (10.1 g, 0.055 mol) according to the procedure used to syn-
thesize 8; mp 63–65 °C; MS (EI) m/z 361 (M).
was synthesized from 8 (1.0 g, 3.5 mmol) and 2-bromoethanol
(0.77 mL, 10.5 mmol) according to the procedure used to synthe-
size 9a; oil; MS (EI) m/z 329 (M+).
4.1.4.9. (E)-Ethyl 3-{5-{[benzyl(2-phenoxyethyl)amino]methyl}-
furan-2-yl}acrylate (9i). Compound 9i (0.91 g, 74.8% yield) was
synthesized from 8 (1.0 g, 3.5 mmol) and (2-bromoethoxy)benzene
(0.72 g, 3.6 mmol) according to the procedure used to synthesize
9a; oil; MS (EI) m/z 405 (M+).
4.1.3.3.
(E)-Ethyl
3-{5-{[2-(benzo[d][1,3]dioxol-5-yl)ethyl-
amino]methyl}furan-2-yl}acrylate (14). Compound 14 (6.0 g,
43.7% yield) was synthesized from 7 (8.54 g, 0.044 mol) and homo-
piperonylamine (6.61 g, 0.04 mol) according to the procedure used
to synthesize 8; oil; MS (EI) m/z 343 (M+).
4.1.4.10. (E)-Ethyl 3-{5-{benzyl[2-(p-tolyloxy)ethyl]amino}-
methyl}furan-2-ylacrylate (9j). Compound 9j (0.79 g, 62.9%
yield) was synthesized from 8 (0.85 g, 3 mmol) and 1-(2-bromo-
ethoxy)-4-methylbenzene (0.77 g, 3.6 mmol) according to the pro-
cedure used to synthesize 9a; oil; MS (EI) m/z 419 (M+).
4.1.4. General procedure for the synthesis of compounds 9a–9p,
12a–12p, 15a–15p
4.1.4.1. (E)-Ethyl 3-{5-{[benzyl(ethyl)amino]methyl}furan-2-
yl}acrylate (9a). Bromoethane (0.45 mL, 6 mmol) was dropped
to the mixture of 8 (0.86 g, 3 mmol), K2CO3 (0.83 g, 6 mmol) and
KI (catalytic amount) in CH3CN. The mixture was stirred at 35 °C
for 8 h. The reaction mixture was filtered. The filtrate was concen-
trated and purified by chromatography on a silica gel column, elut-
ing with a mixture of ethyl acetate and petroleum ether to give the
desired product (0.60 g). Yield: 64.1%, oil. 1H NMR (DMSO-d6,
300 MHz, d ppm) 0.99 (t, 3H, J = 6.6, CH2CH3), 1.06 (t, 3H, J = 6.0,
COOCH2CH3), 2.46 (q, 2H, J = 6.6, CH2CH3), 3.57 (s, 2H, CH2), 3.61
(s, 2H, CH2), 3.98 (q, J = 6.0, 2H, COOCH2CH3), 6.24 (d, 1H, J = 15.6,
vinyl-H), 6.39 (s, 1H, furan-H), 6.69 (s, 1H, furan-H), 7.24 (d, 1H,
J = 15.0, vinyl-H), 7.27–7.33 (m, 5H, Ar-H); MS (EI) m/z 313 (M+).
4.1.4.11.
(E)-Ethyl
3-{5-[benzyl(3-phenoxypropyl)amino]-
methyl}furan-2-ylacrylate (9k). Compound 9k (1.09 g, 86.6%
yield) was synthesized from 8 (0.85 g, 3 mmol) and (3-bromoprop-
oxy)benzene (0.77 g, 3.6 mmol) according to the procedure used to
synthesize 9a; oil; MS (EI) m/z 419 (M+).
4.1.4.12. (E)-Ethyl 3-{5-{{benzyl[3-(p-tolyloxy)propyl]amino}-
methyl}furan-2-yl}acrylate (9l). Compound 9l (1.01 g, 77.9%
yield) was synthesized from 8 (0.85 g, 3 mmol) and 1-(3-bromo-
propoxy)-4-methylbenzene (0.82 g, 3.6 mmol) according to the pro-
cedure used to synthesize 9a; oil; MS (EI) m/z 433 (M+).
4.1.4.2. (E)-Ethyl 3-{5-{[benzyl(propyl)amino]methyl}furan-2-
yl}acrylate (9b). Compound 9b (1.10 g, 96.1% yield) was synthe-
sized from 8 (1.0 g, 3.5 mmol) and 1-bromopropane (0.96 mL,
10.5 mmol) according to the procedure used to synthesize 9a;
oil; MS (EI) m/z 327 (M+).
4.1.4.13. (E)-Ethyl 3-{5-{[benzyl(4-phenoxybutyl)amino]methyl}-
furan-2-yl}acrylate (9m). Compound 9m (0.90 g, 59.4% yield)
was synthesized from 8 (1.0 g, 3.5 mmol) and (4-bromobutoxy)ben-
zene (0.86 g, 3.6 mmol) according to the procedure used to synthe-
size 9a; oil; MS (EI) m/z 433 (M+).
4.1.4.3. (E)-Ethyl 3-{5-{[benzyl(isopropyl)amino]methyl}furan-
2-yl}acrylate (9c). Compound 9c (0.41 g, 36.1% yield) was syn-
thesized from 8 (1.0 g, 3.5 mmol) and 2-bromopropane (0.96 mL,
10.5 mmol) according to the procedure used to synthesize 9a;
oil; MS (EI) m/z 327 (M+).
4.1.4.14. (E)-Ethyl 3-{5-{{benzyl[4-(p-tolyloxy)butyl]amino}-
methyl}furan-2-yl}acrylate (9n). Compound 9n (1.26 g, 80.5%
yield) was synthesized from 8 (1.0 g, 3.5 mmol) and 1-(4-bromo-
butoxy)-4-methylbenzene(0.87 g, 3.6 mmol) according to the pro-
cedure used to synthesize 9a; oil; MS (EI) m/z 447 (M+).
4.1.4.4. (E)-Ethyl 3-{5-{[benzyl(butyl)amino]methyl}furan-2-
yl}acrylate (9d). Compound 9d (0.93 g, 77.9% yield) was synthe-
sized from 8 (1.0 g, 3.5 mmol) and 1-bromobutane (1.13 mL,
10.5 mmol) according to the procedure used to synthesize 9a;
oil; MS (EI) m/z 341 (M+).
4.1.4.15. (E)-Ethyl 3-{5-{[(biphenyl-4-ylmethyl)(ethyl)amino]-
methyl}furan-2-yl}acrylate (12a). Compound 12a (1.26 g,
92.2% yield) was synthesized from 11 (1.26 g, 3.5 mmol) and 1-
bromoethane (0.78 mL, 10.5 mmol) according to the procedure
used to synthesize 9a; oil; MS (EI) m/z 389 (M+).
4.1.4.5. (E)-Ethyl 3-{5-{[benzyl(isobutyl)amino]methyl}furan-2-
yl}acrylate (9e). Compound 9e (0.83 g, 69.2% yield) was synthe-
sized from 8 (1.0 g, 3.5 mmol) and 1-bromo-2-methylpropane
(1.13 mL, 10.5 mmol) according to the procedure used to synthe-
size 9a; oil; MS (EI) m/z 341 (M+).
4.1.4.16.
(E)-Ethyl
3-{5-{[(biphenyl-4-ylmethyl)(isopro-
pyl)amino]methyl}furan-2-yl}acrylate (12b). Compound 12b
(0.51 g, 36.1% yield) was synthesized from 11 (1.26 g, 3.5 mmol)
and 2-bromopropane (0.96 mL, 10.5 mmol) according to the proce-
dure used to synthesize 9a; oil; MS (EI) m/z 403 (M+).
4.1.4.6. (E)-Ethyl 3-{5-{[benzyl(hexyl)amino]methyl}furan-2-
yl}acrylate (9f). Compound 9f (1.10 g, 85.2% yield) was synthe-
4.1.4.17. (E)-Ethyl 3-{5-{[(biphenyl-4-ylmethyl)(pentyl)amino]-
methyl}furan-2-yl}acrylatee (12c). Compound 12c (1.36 g,
90.1% yield) was synthesized from 11 (1.26 g, 3.5 mmol) and bro-
mopentane (0.87 mL, 7 mmol) according to the procedure used to
synthesize 9a; oil; MS (EI) m/z 431 (M+).
sized from
8 (1.0 g, 3.5 mmol) and bromohexane (0.99 mL,
7 mmol) according to the procedure used to synthesize 9a; oil;
MS (EI) m/z 369 (M+).
4.1.4.7.
(E)-Ethyl-3-{5-{[benzyl(octyl)amino]methyl}furan-2-
4.1.4.18. (E)-Ethyl 3-{5-{[(biphenyl-4-ylmethyl)(octyl)amino]-
methyl}furan-2-yl}acrylate (12d). Compound 12d (1.08 g,
65.5% yield) was synthesized from 11 (1.26 g, 3.5 mmol) and
bromooctane (1.22 mL, 7 mmol) according to the procedure used
to synthesize 9a; oil; MS (EI) m/z 473 (M+).
yl}acrylate (9g). Compound 9g (0.91 g, 65.5% yield) was synthe-
sized from 8 (1.0 g, 3.5 mmol) and bromooctane (1.22 mL, 7 mmol)
according to the procedure used to synthesize 9a; oil; MS (EI) m/z
397 (M+).
4.1.4.8. (E)-Ethyl 3-{5-{[benzyl(2-hydroxyethyl)amino]methyl}-
furan-2-yl}acrylate (9h). Compound 9h (1.08 g, 93.8% yield)
4.1.4.19. (E)-Ethyl 3-{5-{[benzyl(biphenyl-4-ylmethyl)amino]-
methyl}furan-2-yl}acrylate (12e). Compound 12e (1.14 g,

5348
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
72.4% yield) was synthesized from 11 (1.26 g, 3.5 mmol) and ben-
zylbromide (0.5 mL, 4.2 mmol) according to the procedure used to
synthesize 9a; oil; MS (EI) m/z 451 (M+).
4.1.4.30. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl]
(isobutyl)amino}methyl}furan-2-yl}acrylate (15d). Compound
15d (0.42 g, 42.1% yield) was synthesized from 14 (0.86 g,
2.5 mmol) and 1-bromo-2-methylpropane (1.1 mL, 10 mmol)
according to the procedure used to synthesize 9a; oil; MS (EI) m/
z 399 (M+).
4.1.4.20. (E)-Ethyl 3-{5-{[(biphenyl-4-ylmethyl)(2-hydroxy-
ethyl)amino]methyl}furan-2-yl}acrylate (12f). Compound 12f
(1.33 g, 93.8% yield) was synthesized from 11 (1.26 g, 3.5 mmol)
and 2-bromoethanol (0.77 mL, 10.5 mmol) according to the proce-
dure used to synthesize 9a; oil; MS (EI) m/z 405 (M+).
4.1.4.31. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl]
(pentyl)amino}methyl}furan-2-yl}acrylate (15e). Compound
15e (0.46 g, 58.6% yield) was synthesized from 14 (0.65 g,
1.9 mmol) and bromopentane (1.0 mL, 8 mmol) according to the
procedure used to synthesize 9a; oil; MS (EI) m/z 413 (M+).
4.1.4.21. (E)-Ethyl 3-{5-{[(biphenyl-4-ylmethyl)(2-phenoxyeth-
yl)amino]methyl}furan-2-yl}acrylate (12g). Compound 12g
(1.08 g, 74.8% yield) was synthesized from 11 (1.26 g, 3 mmol)
and (2-bromoethoxy)benzene (0.72 g, 3.6 mmol) according to the
procedure used to synthesize 9a; oil; MS (EI) m/z 481 (M+).
4.1.4.32. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl](be-
nyl)amino}methyl}furan-2-yl}acrylate (15f). Compound 15f
(0.94 g, 72.4% yield) was synthesized from 14 (1.0 g, 3 mmol) and
benzylbromide (0.43 mL, 3.6 mmol) according to the procedure
used to synthesize 9a; oil; MS (EI) m/z 433 (M+).
4.1.4.22.
(E)-Ethyl
3-{5-{(biphenyl-4-ylmethyl)[2-(p-tolyl-
oxy)ethyl]amino}methyl}furan-2-ylacrylate (12h). Compound
12h (0.93 g, 62.9% yield) was synthesized from 11 (1.26 g, 3 mmol)
and 1-(2-bromoethoxy)-4-methylbenzene (0.77 g, 3.6 mmol)
according to the procedure used to synthesize 9a; oil; MS (EI) m/
z 495 (M+).
4.1.4.33. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl](2-
hydroxyethyl)amino}methyl}furan-2-yl}acrylate (15g). Com-
pound 15g (0.56 g, 47.9% yield) was synthesized from 14 (1.0 g,
3 mmol) and 2-bromoethanol (0.64 mL, 9 mmol) according to the
procedure used to synthesize 9a; oil; MS(EI) m/z 387 (M+).
4.1.4.23. (E)-Ethyl 3-{5-[(biphenyl-4-ylmethyl)(3-phenoxypro-
pyl)amino]methyl}furan-2-ylacrylate (12i). Compound 12i
(1.29 g, 86.6% yield) was synthesized from 11 (1.26 g, 3 mmol)
and (3-bromopropoxy)benzene (0.77 g, 3.6 mmol) according to
the procedure used to synthesize 9a; oil; MS (EI) m/z 495 (M+).
4.1.4.34. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl](2-
phenoxyethyl)amino}methyl}furan-2-yl}acrylate (15h). Com-
pound 15h (0.96 g, 69.1% yield) was synthesized from 14 (1.0 g,
3 mmol) and (2-bromoethoxy)benzene (0.91 g, 4.5 mmol) accord-
ing to the procedure used to synthesize 9a; oil; MS (EI) m/z 463
(M+).
4.1.4.24.
(E)-Ethyl
3-{5-{{(biphenyl-4-ylmethyl)[3-(p-tolyl-
(12j). Com-
oxy)propyl]amino}methyl}furan-2-yl}acrylate
pound 12j (1.19 g, 77.9% yield) was synthesized from 11 (1.26 g,
3 mmol) and 1-(3-bromopropoxy)-4-methylbenzene (0.82 g,
3.6 mmol) according to the procedure used to synthesize 9a; oil;
MS (EI) m/z 509 (M+).
4.1.4.35. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl][2-
(p-tolyloxy]ethyl}amino}methyl}furan-2-yl}acrylate
(15i). Compound 15i (0.78 g, 53.6% yield) was synthesized from
14 (1.0 g, 3 mmol) and 1-(2-bromoethoxy)-4-methylbenzene
(0.77 g, 3.6 mmol) according to the procedure used to synthesize
9a; oil; MS (EI) m/z 477 (M+).
4.1.4.25. (E)-Ethyl 3-{5-{[(biphenyl-4-ylmethyl)(4-phenoxybu-
tyl)amino]methyl}furan-2-yl}acrylate (12k). Compound 12k
(1.06 g, 59.4% yield) was synthesized from 11 (1.26 g, 3.5 mmol)
and (4-bromobutoxy)benzene (0.86 g, 3.6 mmol) according to the
procedure used to synthesize 9a; oil; MS (EI) m/z 509 (M+).
4.1.4.36. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl](3-
phenoxypropyl)amino}methyl}furan-2-yl}acrylate
(15j). Compound 15j (1.28 g, 89.4% yield) was synthesized from
14 (1.0 g, 3 mmol) and (3-bromopropoxy)benzene (0.97 g,
4.5 mmol) according to the procedure used to synthesize 9a; oil;
MS (EI) m/z 477 (M+).
4.1.4.26.
(E)-Ethyl
3-{5-{{(biphenyl-4-ylmethyl)[4-(p-tolyl-
oxy)butyl]amino}methyl}furan-2-yl}acrylate (12l). Compound
12l (1.47 g, 80.5% yield) was synthesized from 11 (1.26 g,
3.5 mmol) and 1-(4-bromobutoxy)-4-methylbenzene (0.87 g,
3.6 mmol) according to the procedure used to synthesize 9a; oil;
MS (EI) m/z 523 (M+).
4.1.4.37. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl](4-
phenoxybutyl)amino}methyl}furan-2-yl}acrylate (15k). Com-
pound 15k (1.0 g, 67.9% yield) was synthesized from 14 (1.0 g,
3 mmol) and (4-bromobutoxy)benzene (1.0 g, 4.5 mmol) according
to the procedure used to synthesize 9a; oil; MS (EI) m/z 491 (M+).
4.1.4.27. (E)-Ethyl 3-{5-{[(2-(benzo[d][1,3]dioxol-5-yl)ethy-
l](ethyl)amino}methyl}furan-2-ylacrylate
(15a). Compound
15a (0.52 g, 46.4% yield) was synthesized from 14 (1.0 g, 3 mmol)
and 1-bromoethane (0.67 mL, 9 mmol) according to the procedure
used to synthesize 9a; oil; MS (EI) m/z 371 (M+).
4.1.5. General procedure for the synthesis of compounds 10a–
10p, 13a–13p, 16a–16p
4.1.5.1.
hydroxyacrylamide (10a).
(E)-3-{5-{[Benzyl(ethyl)amino]methyl}furan-2-yl}-N-
Hydroxylamine hydrochloride
4.1.4.28. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl]-
(propyl)amino}methyl}furan-2-yl}acrylate (15b). Compound
15b (0.60 g, 62.3% yield) was synthesized from 14 (0.86 g,
2.5 mmol) and 1-bromopropane (0.91 mL, 10 mmol) according to
the procedure used to synthesize 9a; oil; MS(EI) m/z 385 (M+).
(1.28 g, 18.4 mmol) in methanol (3.2 mL) was mixed with KOH
(1.03 g, 18.4 mmol) at 40 °C in methanol (6.0 mL), after stirring for
5 min the mixture was cooled to 0 °C for 30 min. The precipitate
was filtered. Compound 9a (0.60 g, 2.0 mmol) was then added to
the filtrate followed by addition of KOH (0.06 g, 1.0 mmol). The mix-
ture was stirred at room temperature for 1 h. The solution was added
to stirring cold water (20 mL), and the pH was adjusted to 7 by add-
ing 2 N HCl. The mixture was extracted with ethyl acetate
(15 mL Â 2). The combined organic solution was washed with satu-
rated brine (5 mL), dried and evaporated to dryness. The residue
4.1.4.29. (E)-Ethyl 3-{5-{{[2-(benzo[d][1,3]dioxol-5-yl)ethyl]-
(butyl)amino}methyl}furan-2-yl}acrylate
(15c). Compound
15c (0.58 g, 58.1% yield) was synthesized from 14 (0.86 g,
2.5 mmol) and 1-bromobutane (0.81 mL, 7.5 mmol) according to
the procedure used to synthesize 9a; oil; MS (EI) m/z 399 (M+).

T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
5349
solid was purified by recrystallization. Yield 43.3% (0.26 g); oil. 1H
NMR (DMSO-d6, 300 MHz, d ppm) 0.99 (t, 3H, J = 6.6, CH2CH3), 2.46
(q, 2H, J = 6.6, CH2CH3), 3.57 (s, 2H,CH2), 3.61 (s, 2H, CH2), 6.24 (d,
1H, J = 15.6, vinyl-H), 6.39 (s, 1H, furan-H), 6.69 (s, 1H, furan-H),
7.24 (d, 1H, J = 15.0, vinyl-H), 7.27–7.33 (m, 5H, Ar-H), 9.0 (s, 1H,
CONHOH), 10.0 (s, 1H, CONHOH); MS (ESI) m/z 299 (MÀH); HRMS
4.1.5.6. (E)-3-{5-{[Benzyl(hexyl)amino]methyl}furan-2-yl}-N-
hydroxyacrylamide (10f). Compound 10f (0.32 g, 45.1% yield)
was synthesized from 9f (0.71 g, 2.0 mmol) according to the proce-
dure used to synthesize 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d
ppm): 0.82 (t, 3H, J = 6.9, CH3), 1.13–1.24 (m, 6H, CH2 Â 3), 1.40–
1.46 (m, 2H, CH2), 2.37 (t, 2H, J = 7.1, CH2), 3.58 (s, 2H, CH2), 3.60
(s, 2H, CH2), 6.21 (d, 1H, J = 15.6, vinyl-H), 6.39 (d, 1H, J = 3.0, fur-
an-H), 6.71 (d, 1H, J = 3.0, furan-H), 7.23 (d, 1H, J = 15.9, vinyl-H),
7.23–7.35 (m, 5H, Ar-H), 9.03 (s, 1H, CONHOH), 10.77 (s, 1H, CON-
HOH); MS (ESI) m/z 357 (M+H); HRMS calcd for C21H29N2O3
calcd for C17H21N2O3 [MH]+: 301.1547; found: 301.1544; IR (KBr)
m
(cmÀ1) = 3416, 3224, 2968, 1621. HPLC purity: 99. 0% (tR = 4.54 min,
80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
[MH]+: 357.2173; found: 357.218; IR (KBr)
m
(cmÀ1) = 3260, 3024,
4.1.5.2. (E)-3-{5-{[Benzyl(propyl)amino]methyl}furan-2-yl}-N-
hydroxyacrylamide (10b). Compound 10b (0.25 g, 39.3% yield)
was synthesized from 9b (0.62 g, 2.0 mmol) according to the pro-
cedure used to synthesize 10a; oil. 1H NMR (DMSO-d6, 300 MHz,
d ppm): 0.82 (t, 3H, J = 7.1, CH3), 1.47 (m, 2H, CH2CH3), 2.36 (t,
2H, J = 6.9, CH2CH2CH3), 3.58 (s, 2H, CH2), 3.61 (s, 2H, CH2), 6.20
(d, 2H, J = 15.3, vinyl-H), 6.39 (s, 1H, furan-H), 6.70 (s, 1H, furan-
H), 7.23 (d, 2H, J = 15.6, vinyl-H), 7.25–7.34 (m, 5H, Ar-H), 8.99
(s, 1H, CONHOH), 10.75 (s, 1H, CONHOH); MS (ESI) m/z 313
(MÀH); HRMS calcd for C18H23N2O3 [MH]+: 315.1703; found:
2861, 1580. HPLC purity: 98. 6% (tR = 5.65 min, 80% MeCN/20%
H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.7.
(E)-3-{5-{[Benzyl(octyl)amino]methyl}furan-2-yl}-N-
hydroxyacrylamide (10g). Compound 10g (0.32 g, 41.2% yield)
was synthesized from 9g (0.76 g, 2.0 mmol) according to the proce-
dure used to synthesize 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d
ppm): 0.90 (t, 3H, J = 6.6, CH3), 1.26 (m, 10H, CH2 Â 5), 1.50 (m,
2H,CH2), 2.43 (t, 2H, J = 6.7,CH2), 3.64 (s, 2H, CH2), 3.67 (s, 2H,
CH2), 6.27 (d, 1H, J = 15.6, vinyl-H), 6.45 (d,1H, J = 3.0, furan-H),
6.76 (d, 1H, J = 3.0, furan-H), 7.28 (d, 1H, J = 15.6, vinyl-H), 7.31–
7.41 (m, 5H, Ar-H), 9.08 (s, 1H, CONHOH), 10.08 (s, 1H, CONHOH);
MS (ESI) m/z 383 (MÀH); HRMS calcd for C23H33N2O3 [MH]+:
315.1700; IR (KBr)
m
(cmÀ1) = 3430, 3231, 2975, 1665, 1456. HPLC
purity: 98. 7% (tR = 5.44 min, 80% MeCN/20% H2O/0.1% TFA),
0.5 mL/min in 20 min.
385.2486; found: 385.2475; IR (KBr)
m
(cmÀ1) = 3416, 3253, 2940,
1620, 1534. HPLC purity: 98. 1% (tR = 6.34 min, 80% MeCN/20%
H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.3. (E)-3-{5-{[Benzyl(isopropyl)amino]methyl}furan-2-yl}-
N-hydroxyacrylamide (10c). Compound 10c (0.12 g, 37.5%
yield) was synthesized from 9c (0.31 g, 1.0 mmol) according to
the procedure used to synthesize 10a; oil. 1H NMR (DMSO-d6;
300 MHz, d ppm): 1.01 (s, 3H, CH3), 1.03 (s, 3H, CH3), 2.91 (m,
1H, CH), 3.58 (s, 2H, CH2), 3.60 (s, 2H, CH2), 6.20 (d, 1H, J = 15.6, vi-
nyl-H), 6.34 (d, 1H, J = 2.7, furan-H), 6.59 (s, 1H, furan-H), 7.11 (d,
1H, J = 15.0, vinyl-H), 7.18–7.36 (m, 5H, Ar-H); MS (ESI) m/z 313
(MÀH); HRMS calcd for C18H23N2O3 [MH]+: 315.1703; found:
4.1.5.8. (E)-3-{5-{[Benzyl(2-hydroxyethyl)amino]methyl}furan-
2-yl}-N-hydroxyacrylamide (10h). Compound 10h (0.24 g, 37.5%
yield) was synthesized from 9h (0.63 g, 2.0 mmol) according to the
procedure used to synthesize 10a; oil. 1H NMR (DMSO-d6, 300 MHz,
d ppm): 2.53 (t, 2H, J = 6.0, CH2), 3.51 (t, 2H, J = 6.0, OCH2), 3.63 (s,
2H, CH2),3.67 (s, 2H, CH2), 4.41 (t, 1H, J = 5.1, OH), 6.21 (d, 1H,
J = 15.6, vinyl-H), 6.42 (s, 1H, furan-H), 6.72 (s,1H, furan-H), 7.25 (d,
1H, J = 15.6, vinyl-H), 7.2–7.35 (m, 5H, Ar-H), 9.01 (s, 1H, CONHOH),
10.75 (s, 1H, CONHOH); MS (ESI) m/z 315 (MÀH); HRMS calcd for
315.1712; IR (KBr)
m
(cmÀ1) = 3438, 3267, 2954, 1616, 1495. HPLC
purity: 99. 1% (tR = 4.86 min, 80% MeCN/20% H2O/0.1% TFA),
0.5 mL/min in 20 min.
C
17H21N2O4 [MH]+: 317.1496; found: 317.1486; IR (KBr)
m
(cmÀ1) = 3217, 2925, 2847, 1740. HPLC purity: 98.3% (tR = 3.17 min,
4.1.5.4.
(E)-3-{5-{[Benzyl(butyl)amino]methyl}furan-2-yl}-N-
80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
hydroxyacrylamide (10d). Compound 10d (0.28 g, 43.4% yield)
was synthesized from 9d (0.65 g, 2.0 mmol) according to the pro-
cedure used to synthesize 10a; oil. 1H NMR (DMSO-d6, 300 MHz,
d ppm): 0.81 (t, 3H, J = 7.2, CH3), 1.25 (m, 2H, CH2), 1.45 (m, 2H,
CH2), 2.38 (t, 2H, J = 7.1, CH2), 3.58 (s, 2H, CH2), 3.61 (s, 2H, CH2),
6.21 (d, 1H, J = 15.3, vinyl-H), 6.39 (d, 1H, J = 3.3, furan-H), 6.70
(d, 1H, J = 3.0, furan-H), 7.23 (d, 1H, J = 15.6, vinyl-H), 7.23–7.34
(m, 5H, Ar-H), 9.02 (s, 1H, CONHOH), 10.76 (s, 1H, CONHOH); MS
4.1.5.9. (E)-3-{5-{[Benzyl(2-phenoxyethyl)amino]methyl}furan-
2-yl}-N-hydroxyacrylamide (10i). Compound 10i (0.34 g, 43.1%
yield) was synthesized from 9i (0.80 g, 2.0 mmol) according to the
procedure used to synthesize 10a; oil. 1H NMR (DMSO-d6,
300 MHz, d ppm): 2.84 (t, 2H, J = 5.6, NCH2), 3.72 (s, 2H, CH2),
3.76 (s, 2H, CH2), 4.06 (t, 2H, J = 5.4, CH2O), 6.24 (d, 1H, J = 15.6, vi-
nyl-H), 6.45 (d, 1H, J = 2.1, furan-H), 6.72 (d, 1H, J = 1.8, furan-H),
6.88–6.93 (m, 3H, Ar-H), 7.27 (d, 1H, J = 15.6, vinyl-H), 7.22–7.39
(m, 7H, Ar-H), 9.04 (s, 1H, CONHOH), 10.76 (s, 1H, CONHOH); MS
(ESI) m/z 327 (MÀH); HRMS calcd for
C
19H25N2O3 [MH]+:
329.1860; found: 329.1866; IR (KBr)
m
(cmÀ1) = 3416, 3246, 2968,
1659, 1449. HPLC purity: 98. 0% (tR = 6.78 min, 80% MeCN/20%
H2O/0.1% TFA), 0.5 mL/min in 20 min.
(ESI) m/z 393 (M+H); HRMS calcd for
C
23H25N2O4 [MH]+:
393.1809; found: 393.1816; IR (KBr)
m
(cmÀ1) = 3032, 2918, 1722,
1641. HPLC purity: 98. 9% (tR = 4.48 min, 80% MeCN/20% H2O/
4.1.5.5. (E)-3-{5-{[Benzyl(isobutyl)amino]methyl}furan-2-yl}-
N-hydroxyacrylamide (10e). Compound 10e (0.30 g, 45.1%
yield) was synthesized from 9e (0.65 g, 2.0 mmol) according to
the procedure used to synthesize 10a; oil. 1H NMR (DMSO-d6,
300 MHz, d ppm): 0.81 (s, 3H, CH3), 0.84 (s, 3H, CH3), 1.81 (m,
1H, CH), 2.15 (d, 2H, J = 7.5, CH2CH), 3.57 (s, 2H, CH2), 3.59 (s,
2H, CH2), 6.20 (d, 1H, J = 15.3, vinyl-H), 6.38 (d, 1H, J = 3.3, furan-
H), 6.70 (d, 1H, J = 3.0, furan-H), 7.23 (d, 1H, J = 15.3, vinyl-H),
7.22–7.35 (m, 5H, Ar-H), 8.98 (s, 1H, CONHOH), 10.76 (s, 1H, CON-
HOH); MS (ESI) m/z 327 (MÀH); HRMS calcd for C19H25N2O3
0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.10. (E)-3-{5-{{Benzyl[2-(p-tolyloxy)ethyl]amino}methyl}-
furan-2-yl}-N-hydroxyacrylamide
(10j). Compound
10j
(0.35 g, 43.6% yield) was synthesized from 9j (0.81 g, 2.0 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR
(DMSO-d6, 300 MHz, d ppm): 2.20 (s, 3H, CH3), 2.81 (t, 2H, J = 6.8,
NCH2), 3.70 (s, 2H, CH2), 3.74 (s, 2H, CH2), 4.00 (t, 2H, J = 6.8,
CH2O), 6.21 (d, 1H, J = 15.6, vinyl-H), 6.43 (d, 1H, J = 3.0, furan-H),
6.69 (d, 1H, J = 2.7, furan-H), 6.76 (d, 2H, J = 8.4, Ar-H), 7.03 (d,
2H, J = 8.1, Ar-H), 7.22 (d, 1H, J = 15.3, vinyl-H), 7.21–7.37 (m, 5H,
Ar-H); MS (ESI) m/z 405 (MÀH); HRMS calcd for C24H27N2O4
[MH]+: 329.1860; found: 329.1863; IR (KBr)
m
(cmÀ1) = 3473,
3437, 3238, 2953, 1658. HPLC purity: 98. 7% (tR = 6.57 min, 80%
MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
[MH]+: 407.1965; found: 407.1969; IR (KBr)
m
(cmÀ1) = 3181,

5350
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
3039, 2875, 1609. HPLC purity: 98. 4% (tR = 6.42 min, 80% MeCN/
20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
(DMSO-d6, 300 MHz, d ppm): 1.12 (t, 3H, J = 7.1, CH2CH3), 2.56 (q,
2H, J = 7.1, CH2CH3), 3.63 (s, 2H, CH2), 3.73 (s, 2H, CH2), 6.29 (d, 2H,
J = 15.6, vinyl-H), 6.48 (d, 1H, J = 3.3, furan-H), 6.77 (d, 1H, J = 3.0, fur-
an-H), 7.29 (d, 2H, J = 15.6, vinyl-H), 7.39–7.77 (m, 9H, Ar-H), 9.06 (s,
1H, CONHOH), 10.8(s, 1H, CONHOH); MS (ESI)m/z 375 (MÀH); HRMS
calcd for C23H25N2O3 [MH]+: 377.1860; found: 377.1861; IR (KBr)
4.1.5.11.
(E)-3-{5-{[Benzyl(3-phenoxypropyl)amino]methyl}-
(10k). Compound 10k
furan-2-yl}-N-hydroxyacrylamide
(0.36 g, 43.8% yield) was synthesized from 9k (0.81 g, 2.0 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR
(DMSO-d6, 300 MHz, d ppm): 1.89 (m, 2H, CH2), 2.58 (t, 2H, J = 6.7,
CH2), 3.63 (s, 2H, CH2), 3.65 (s, 2H, CH2), 3.96 (t, 2H, J = 6.3,CH2),
6.14 (d, 1H, J = 15.6, vinyl-H), 6.41 (d, 1H, J = 3.0, furan-H), 6.69 (d,
1H, J = 3.0, furan-H), 6.77–6.93 (m, 3H, Ar-H), 7.28 (d, 1H, J = 15.6, vi-
nyl-H), 7.20–7.35 (m, 7H, Ar-H), 9.03 (s, 1H, CONHOH), 10.73 (s, 1H,
CONHOH); MS (ESI) m/z 407 (M+H); HRMS calcd for C24H27N2O4
m
(cmÀ1) = 3440, 2968, 2918, 2839, 1630. HPLC purity: 97. 3%
(tR = 4.76 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.16.
(E)-3-{5-{[(Biphenyl-4-ylmethyl)(isopropyl)amino]-
methyl}furan-2-yl}-N-hydroxyacrylamide (13b). Compound
13b (0.15 g, 38.3% yield) was synthesized from 12b (0.39 g,
1.0 mmol) according to the procedure used to synthesize 10a; oil.
1H NMR (DMSO-d6, 300 MHz, d ppm): 1.03 (s, 3H, CH3), 1.05 (s,
3H, CH3), 2.93 (m, 1H, CH), 3.62 (s, 2H, CH2), 3.65 (s, 2H, CH2),
6.08 (d, 1H, J = 15.6, vinyl-H), 6.41 (d, 1H, J = 2.7, furan-H), 6.71
(d, 1H, J = 2.7, furan-H), 7.24(d, 1H, J = 15.6, vinyl-H), 7.33–
7.68(m, 9H, Ar-H), 9.01(s, 1H, CONHOH), 10.8(s, 1H, CONHOH);
MS (ESI) m/z 389 (MÀH); HRMS calcd for C24H27N2O3 [MH]+:
[MH]+: 407.1965; found: 407.1971; IR (KBr)
m
(cmÀ1) = 3452, 2085,
1655, 1634. HPLC purity: 97. 9% (tR = 5.91 min, 80% MeCN/20%
H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.12. (E)-3-{5-{{Benzyl[3-(p-tolyloxy)propyl]amino}methyl}-
furan-2-yl}-N-hydroxyacrylamide (10l). Compound 10l (0.28 g,
43.6% yield) was synthesized from 9l (0.63 g, 1.5 mmol) according
to the procedure used to synthesize 10a; oil. 1H NMR (DMSO-d6,
300 MHz, d ppm): 1.86 (t, 2H, J = 6.5, CH2), 2.01 (s, 3H, CH3), 2.61
(m, 2H, CH2),3.62 (s, 2H, CH2), 3.64 (s, 2H, CH2), 3.90 (t, 2H,
J = 6.9, CH2O), 6.22 (d, 1H, J = 15.6, vinyl-H), 6.35 (d, 1H, J = 3.0, fur-
an-H), 6.69 (d, 1H, J = 3.0, furan-H), 6.74 (d, 2H, J = 8.4, Ar-H), 7.05
(d, 2H, J = 8.4, Ar-H), 7.24 (d, 1H, J = 15.0, vinyl-H), 7.12–7.35 (m,
5H, Ar-H), 9.08 (s, 1H, CONHOH), 10.72 (s, 1H,CONHOH); MS
391.2016; found: 391.2013; IR (KBr)
m
(cmÀ1) = 3423, 2968, 1655,
1630. HPLC purity: 99. 1% (tR = 5.13 min, 80% MeCN/20% H2O/
0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.17. (E)-3-{5-{[(Biphenyl-4-ylmethyl)(pentyl)amino]-
methyl}furan-2-yl}-N-hydroxyacrylamide
(13c). Compound
13c (0.34 g, 40.4% yield) was synthesized from 12c (0.84 g, 2.0 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR
(DMSO-d6, 300 MHz, d ppm): 0.85 (s, 3H, CH3), 1.23 (m, 4H,
CH2 Â 2), 1.48 (m, 2H, CH2), 2.41 (q, 2H, J = 4.5, CH2), 3.63 (s, 2H,
CH2), 3.64 (s, 2H, CH2), 6.23 (d, 1H, J = 15.6, vinyl-H), 6.41 (d, 1H,
J = 3.0, furan-H), 6.71 (d, 1H, J = 3.0, furan-H), 7.25 (d, 1H, J = 15.6, vi-
nyl-H), 7.33–7.49 (m, 5H, Ar-H), 7.65–7.68 (m, 4H, Ar-H), 9.0 (s, 1H,
CONHOH), 10.8 (s, 1H, CONHOH); MS (ESI) m/z 419 (M+H); HRMS
(ESI) m/z 421 (M+H); HRMS calcd for
C
25H29N2O4 [MH]+:
421.2122; found: 421.2119; IR (KBr)
m
(cmÀ1) = 3573, 3423, 1730,
1570. HPLC purity: 98. 5% (tR = 6.84 min, 80% MeCN/20% H2O/
0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.13.
(E)-3-{5-{[Benzyl(4-phenoxybutyl)amino]methyl}-
furan-2-yl}-N-hydroxyacrylamide (10m). Compound 10m
(0.25 g, 40.2% yield) was synthesized from 9m (0.63 g, 1.5 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR
(DMSO-d6, 300 MHz, d ppm): 1.62–1.73 (m, 4H, CH2 Â 2), 2.48 (t,
2H, J = 6.4, CH2), 3.60 (s, 2H, CH2), 3.62 (s, 2H, CH2), 3.88 (t, 2H,
J = 6.2, CH2O), 6.22 (d, 1H, J = 15.6, vinyl-H), 6.40 (d, 1H, J = 3.0, fur-
an-H, 6.70 (d, 1H, J = 3.0, furan-H), 6.88 (d, 1H, J = 15.6, vinyl-H),
6.78–6.90 (m, 3H, Ar-H), 6.99–7.59 (m, 7H, Ar-H), 9.04 (s, 1H, CON-
HOH), 10.77 (s, 1H, CONHOH); MS (ESI) m/z 421 (M+H); HRMS
calcd for C25H29N2O4 [MH]+: 421.2122; found: 421.2120; IR (KBr)
calcd for C26H31N2O3 [MH]+: 419.2329; found: 419.2335; IR (KBr)
m
(cmÀ1) = 3200, 3039, 2928, 1641. HPLC purity: 98. 1% (tR = 4.26 min,
80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.18. (E)-3-{5-{[(Biphenyl-4-ylmethyl)(octyl)amino]methyl}-
furan-2-yl}-N-hydroxyacrylamide
(13d). Compound
13d
(0.37 g, 40.1% yield) was synthesized from 12d (0.92 g, 2.0 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR
(DMSO-d6, 300 MHz, d ppm): 0.82 (t, 3H, J = 6.4, CH3), 1.19 (m,
10H, CH2 Â 5), 1.46 (m, 2H,CH2), 2.41 (t, 2H, J = 6.7, CH2), 3.63 (s,
2H, CH2), 3.65 (s, 2H, CH2), 6.25 (d, 1H, J = 15.3, vinyl-H), 6.41 (d,
1H, J = 2.7, furan-H), 6.71 (d, 1H, J = 2.7, furan-H), 7.26 (d, 1H,
J = 15.6, vinyl-H), 7.33–7.48 (m, 5H, Ar-H), 7.65–7.67 (m, 4H, Ar-
H), 9.0 (s, 1H, CONHOH), 10.8 (s, 1H, CONHOH); MS (ESI) m/z 461
(M+H); HRMS calcd for C29H37N2O3 [MH]+: 461.2799; found:
m
(cmÀ1) = 3566, 3437, 1698, 1552. HPLC purity: 98. 8%
(tR = 5.09 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in
20 min.
4.1.5.14. (E)-3-{5-{{Benzyl[4-(p-tolyloxy)butyl]amino}methyl}-
furan-2-yl}-N-hydroxyacrylamide
(10n). Compound
10n
(0.28 g, 43.3% yield) was synthesized from 9n (0.65 g, 1.5 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR
(DMSO-d6, 300 MHz, d ppm): 1.61–1.68 (m, 4H, CH2 Â 2), 2.21 (s,
3H, CH3), 2.44 (t, 2H, J = 6.4, CH2), 3.59 (s, 2H, CH2), 3.62 (s, 2H,
CH2), 3.84 (t, 2H, J = 5.9, CH2O), 6.22 (d, 1H, J = 15.6, vinyl-H),
6.39 (d, 1H, J = 3.0, furan-H), 6.68 (d, 1H, J = 2.7, furan-H), 6.75 (d,
2H, J = 8.4, Ar-H), 7.05 (d, 2H, J = 8.4, Ar-H), 7.26 (d, 1H, J = 15.6, vi-
nyl-H), 7.19–7.34 (m, 5H, Ar-H), 9.04 (s, 1H, CONHOH), 10.77 (s,1H,
CONHOH); MS (ESI) m/z 435 (M+H); HRMS calcd for C26H31N2O4
461.2812; IR (KBr)
m
(cmÀ1) = 3025, 2925, 2854, 1637, 1505. HPLC
purity: 98. 2% (tR = 5.49 min, 80% MeCN/20% H2O/0.1% TFA),
0.5 mL/min in 20 min.
4.1.5.19. (E)-3-{5-{[Benzyl(biphenyl-4-ylmethyl)amino]methyl}-
furan-2-yl}-N-hydroxyacrylamide
(13e). Compound
13e
(0.33 g, 37.7% yield) was synthesized from 12e (0.88 g, 2.0 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR
(DMSO-d6, 300 MHz, d ppm): 3.60 (s, 6H, CH2 Â 3), 6.26 (d, 1H,
J = 15.6, vinyl-H), 6.38 (d, 1H, J = 3.0, furan-H), 6.58 (d, 1H, J = 3.0,
furan-H), 7.21 (d, 1H, J = 15.6, vinyl-H), 7.34–7.45 (m, 10H, Ar-H),
7.64–7.67 (m, 4H, Ar-H); MS (ESI) m/z 437 (MÀH); HRMS calcd
[MH]+: 435.2278; found: 435.2276; IR (KBr)
m
(cmÀ1) = 3480,
3423, 2925, 1634. HPLC purity: 97. 6% (tR = 7.74 min, 80% MeCN/
20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
for C28H27N2O3 [MH]+: 439.2016; found: 439.2022; IR (KBr)
m
4.1.5.15. (E)-3-{5-{[(Biphenyl-4-ylmethyl)(ethyl)amino]methyl}-
(cmÀ1) = 3027, 2925, 2871, 1598. HPLC purity: 98. 0%
(tR = 4.81 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in
20 min.
furan-2-yl}-N-hydroxyacrylamide
(13a). Compound
13a
(0.33 g, 44.3% yield) was synthesized from 12a (0.75 g, 2.0 mmol)
according to the procedure used to synthesize 10a; oil. 1H NMR

T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
5351
4.1.5.20.
(E)-3-{5-{[(Biphenyl-4-ylmethyl)(2-hydroxyethyl)-
12j (0.74 g, 1.5 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 1.90 (m, 2H,
CH2), 2.21 (s, 3H, CH3), 2.58 (t, 2H, J = 6.3, NCH2), 3.66 (s, 2H,
CH2), 3.68 (s, 2H, CH2), 3.94 (t, 2H, J = 5.8, CH2O), 6.24 (d, 1H,
J = 15.6, vinyl-H), 6.44 (s, 1H, furan-H), 6.72 (s, 1H, furan-H), 6.74
(d, 2H, J = 8.4, Ar-H), 7.03 (d, 2H, J = 8.1, Ar-H), 7.24 (d, 1H,
J = 15.6, vinyl-H), 7.33–7.49 (m, 5H, Ar-H), 7.59 (d, 2H, J = 8.1, Ar-
H), 7.64 (d, 2H, J = 7.5, Ar-H), 9.03 (s, 1H, CONHOH), 10.77 (s, 1H,
CONHOH); MS (ESI) m/z 497 (M+H); HRMS calcd for C31H33N2O4
amino]methyl}furan-2-yl}-N-hydroxyacrylamide (13f). Com-
pound 13f (0.28 g, 36.2% yield) was synthesized from 12f (0.80 g,
2.0 mmol) according to the procedure used to synthesize 10a; oil.
1H NMR (DMSO-d6, 300 MHz, d ppm): 2.57 (t, 2H, J = 6.3, NCH2),
3.53 (t, 2H, J = 5.1, CH2OH), 3.68 (s, 2H, CH2), 3.71 (s, 2H, CH2), 4.43
(t, 1H, J = 4.8, CH2OH), 6.24 (d, 1H, J = 15.6, vinyl-H), 6.44 (d, 1H,
J = 2.4, furan-H), 6.72 (s, 1H), 7.24 (d, 1H, J = 15.6, vinyl-H), 7.33–
7.38 (m, 1H, Ar-H), 7.44–7.49 (m, 4H, Ar-H), 7.64 (d, 1H, J = 8.4, Ar-
H), 7.66 (d, 1H, J = 7.8, Ar-H), 9.03 (s, 1H, CONHOH), 10.77 (s, 1H,
CONHOH); MS (ESI) m/z 393 (M+H); HRMS calcd for C23H25N2O4
[MH]+:497.2435; found: 497.2444; IR (KBr)
m
(cmÀ1) = 3388,
3196, 2918, 1614, 1580. HPLC purity: 97. 6% (tR = 4.94 min, 80%
MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
[MH]+: 393.1809; found: 393.1816; IR (KBr)
m
(cmÀ1) = 3409, 2932,
1630, 1559. HPLC purity: 99. 3% (tR = 3.94 min, 80% MeCN/20%
H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.25.
(E)-3-{5-{[(Biphenyl-4-ylmethyl)(4-phenoxybutyl)-
amino]methyl}furan-2-yl}-N-hydroxyacrylamide (13k). Com-
pound 13k (0.32 g, 43.5% yield) was synthesized from 12k
(0.74 g, 1.5 mmol) according to the procedure used to synthesize
10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 1.66 (m, 2H, CH2),
1.73 (m, 2H, CH2), 2.50 (t, 2H, J = 6.1, NCH2), 3.64 (s, 2H, CH2),
3.67 (s, 2H, CH2), 3.89 (t, 2H, J = 6.0, CH2O), 6.24 (d, 1H, J = 15.6, vi-
nyl-H), 6.43 (d, 1H, J = 3.0, furan-H), 6.71 (d, 1H, J = 3.0, furan-H),
6.85–6.89 (m, 3H, Ar-H), 7.23 (d, 1H, J = 15.6, vinyl-H), 7.24–7.26
(m, 2H, Ar-H), 7.35–7.38 (m, 1H, Ar-H), 7.42–7.49 (m, 3H, Ar-H),
7.62 (d, 2H, J = 7.8, Ar-H), 7.65 (d, 2H, J = 7.5, Ar-H); MS (ESI) m/z
497 (M+H); HRMS calcd for C31H33N2O4 [MH]+:497.2435; found:
4.1.5.21.
(E)-3-{5-{[(Biphenyl-4-ylmethyl)(2-phenoxyethyl)-
amino]methyl}furan-2-yl}-N-hydroxyacrylamide (13g). Com-
pound 13g (0.31 g, 43.9% yield) was synthesized from 12g (0.70 g,
1.5 mmol) according to the procedure used to synthesize 10a; oil.
1H NMR (DMSO-d6, 300 MHz, d ppm): 2.87 (t, 2H, J = 5.3, NCH2),
3.77 (s, 2H, CH2), 3.80 (s, 2H, CH2), 4.10 (t, 2H, J = 5.4, CH2O), 6.25
(d, 1H, J = 15.6, vinyl-H), 6.49 (s, 1H, furan-H), 6.73 (s, 1H, furan-
H), 6.89–6.92 (m, 3H, Ar-H), 7.26 (d, 1H, J = 15.0, vinyl-H), 7.24–
7.29 (m, 2H, Ar-H), 7.33–7.38 (m, 1H, Ar-H), 7.45 (d, 2H, J = 7.9,Ar-
H), 7.47 (d, 2H, J = 8.1, Ar-H), 7.64 (d, 2H, J = 8.1, Ar-H), 7.66 (d, 2H,
J = 7.8, Ar-H), 9.04 (s,1H, CONHOH), 10.77 (s, 1H, CONHOH); MS
(ESI) m/z 469 (M+H); HRMS calcd for C29H29N2O4 [MH]+:469.2122;
497.2446; IR (KBr)
purity: 98.7% (tR = 4.57 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/
min in 20 min.
m
(cmÀ1) = 3024, 2939, 2861, 1599, 1495. HPLC
found: 469.2128; IR (KBr)
m
(cmÀ1) = 3200, 2918, 1602, 1495. HPLC
purity: 98. 9% (tR = 4.73 min, 80% MeCN/20% H2O/0.1% TFA),
0.5 mL/min in 20 min.
4.1.5.26.
(E)-3-{5-{{[Biphenyl-4-ylmethyl][4-(p-tolyloxy)
butyl]amino}methyl}furan-2-yl}-N-hydroxyacrylamide
4.1.5.22. (E)-3-{5-{{(Biphenyl-4-ylmethyl)[2-(p-tolyloxy)ethyl]-
amino}methyl}furan-2-yl}-N-hydroxyacrylamide (13h). Com-
pound 13h (0.33 g, 46.0% yield) was synthesized from 12h (0.72 g,
1.5 mmol) according to the procedure used to synthesize 10a; oil.
1H NMR (DMSO-d6, 300 MHz, d ppm): 2.12 (s, 3H, CH3), 2.85 (t, 2H,
J = 5.2, NCH2), 3.76 (s, 2H, CH2), 3.79 (s, 2H, CH2), 4.06 (t, 2H,
J = 5.1, CH2O), 6.25 (d, 1H, J = 15.6, vinyl-H), 6.48 (s, 1H, furan-H),
6.73 (s, 1H, furan-H), 6.79 (d, 2H, J = 7.8, Ar-H), 7.05 (d, 2H, J = 8.2,
Ar-H), 7.25 (d, 1H, J = 15.6, vinyl-H), 7.33–7.38 (m, 1H, Ar-H),
7.46–7.47 (m, 4H, Ar-H), 7.63 (d, 2H, J = 8.1, Ar-H), 7.65 (d, 2H,
J = 7.9, Ar-H), 9.03 (s, 1H, CONHOH), 10.77 (s, 1H, CONHOH); MS
(13l). Compound 13l (0.31 g, 41.2% yield) was synthesized from
12l (0.76 g, 1.5 mmol) according to the procedure used to synthesize
10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 1.65–1.70 (m, 4H,
CH2 Â 2), 2.20 (s, 3H, CH3), 3.34 (m, 2H, CH2), 3.63 (t, 2H, J = 6.6,
CH2), 3.66 (s, 2H, CH2), 3.84 (t, 2H, J = 6.1, CH2O), 6.25 (d, 1H,
J = 15.6, vinyl-H), 6.42 (s, 1H, furan-H), 6.73 (s, 1H, furan-H), 6.74
(d, 2H, J = 7.8, Ar-H), 7.02 (d, 2H, J = 8.0, Ar-H), 7.25 (d, 1H, J = 15.0,
vinyl-H), 7.35–7.45 (m, 5H, Ar-H), 7.61 (d, 1H, J = 7.8, Ar-H), 7.64
(d, 1H, J = 7.8, Ar-H), 9.04 (s, 1H, CONHOH), 10.77 (s, 1H, CONHOH);
MS (ESI) m/z 511 (M+H); HRMS calcd for
C32H35N2O4
[MH]+:511.2591; found: 511.2594; IR (KBr) (cmÀ1) = 3438, 3260,
m
(ESI) m/z 481 (MÀH); HRMS calcd for
C
30H31N2O4
2954, 2769, 1630. HPLC purity: 99. 0% (tR = 7.53 min, 80% MeCN/
20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
[MH]+:483.2278; found: 483.2289; IR (KBr) (cmÀ1) = 3210, 3032,
m
2918, 1612. HPLC purity: 99. 6% (tR = 6.41 min, 80% MeCN/20%
H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.27. (E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](ethyl)a-
mino}methyl}furan-2-yl}-N-hydroxyacrylamide (16a). Com-
pound 16a (0.14 g, 40.6% yield) was synthesized from 15a
(0.36 g, 1.0 mmol) according to the procedure used to synthesize
10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 1.0 (t, 3H, J = 6.4,
CH3), 2.51 (q, 2H, J = 6.4, NCH2), 2.52–2.64 (m, 4H,CH2 Â 2), 3.68
(s, 2H, CH2), 5.94 (s, 2H, OCH2O), 6.20 (d, 1H, J = 15.4, vinyl-H),
6.37 (d, 1H, J = 3.0, furan-H), 6.62–6.65 (m, 1H, Ar-H), 6.68 (d, 1H,
J = 3.0, furan-H), 6.76–6.79 (m, 2H, Ar-H), 7.21 (d, 1H, J = 15.5, vi-
nyl-H), 8.96 (s, 1H, CONHOH), 10.67 (s, 1H, CONHOH); MS (ESI)
m/z 357 (MÀH); HRMS calcd for C19H23N2O5 [MH]+:359.1601;
4.1.5.23.
(E)-3-{5-{[(Biphenyl-4-ylmethyl)(3-phenoxypropyl)-
amino]methyl}furan-2-yl}-N-hydroxyacrylamide (13i). Com-
pound 13i (0.30 g, 41.6% yield) was synthesized from 12i (0.72 g,
1.5 mmol) according to the procedure used to synthesize 10a; oil.
1H NMR (DMSO-d6, 300 MHz, d ppm): 1.93 (t, 2H, J = 5.7, NCH2),
2.61 (m, 2H, CH2), 3.67 (s, 2H, CH2), 3.69 (s, 2H, CH2), 3.99 (t, 2H,
J = 6.0, CH2O), 6.24 (d, 1H, J = 15.6, vinyl-H), 6.44 (d, 1H, J = 2.1, fur-
an-H), 6.70 (s, 1H, furan-H), 6.88 (d, 1H, J = 15.3, vinyl-H), 6.84–
6.90 (m, 2H, Ar-H), 7.21–7.27 (m, 3H, Ar-H), 7.33–7.37 (m, 1H, Ar-
H), 7.41–7.48 (m, 4H, Ar-H), 7.59 (d, 2H, J = 8.1, Ar-H), 7.64 (d, 2H,
J = 8.1, Ar-H), 9.01 (s, 1H, CONHOH), 10.75 (s, 1H, CONHOH); MS
(ESI) m/z 483 (M+H); HRMS calcd for C30H31N2O4 [MH]+:483.2278;
found: 359.1609; IR (KBr) m
(cmÀ1) = 3200, 2932, 1609, 1488. HPLC
purity: 98. 1% (tR = 4.68 min, 80% MeCN/20% H2O/0.1% TFA),
0.5 mL/min in 20 min.
found: 483.2280; IR (KBr)
m
(cmÀ1) = 3025, 2939, 1602, 1580. HPLC
purity: 99. 2% (tR = 5.14 min, 80% MeCN/20% H2O/0.1% TFA),
4.1.5.28.
(E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](pro-
0.5 mL/min in 20 min.
pyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(16b). Compound 16b (0.15 g, 42.0% yield) was synthesized from
15b (0.37 g, 1.0 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 0.82 (t, 3H,
J = 6.1, CH3), 1.42 (m, 2H, CH2), 2.42 (t, 2H, J = 6.4, NCH2), 2.50–
4.1.5.24.
(E)-3-{5-{{(Biphenyl-4-ylmethyl)[3-(p-tolyloxy)pro-
pyl]amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(13j). Compound 13j (0.31 g, 42.0% yield) was synthesized from

5352
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
2.60 (m, 4H, CH2 Â 2), 3.68 (s, 2H, CH2), 5.94 (s, 2H, OCH2O), 6.18
(d, 1H, J = 15.6, vinyl-H), 6.38 (s, 1H, furan-H), 6.63–6.79 (m, 4H,
Ar-H), 7.21 (d, 1H, J = 15.6, vinyl-H), 9.0 (s, 1H, CONHOH), 10.7 (s,
1H, CONHOH); MS (ESI) m/z 373 (M+H); HRMS calcd for
1630, 1488. HPLC purity: 99. 2% (tR = 6.52 min, 80% MeCN/20%
H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.33.
(E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](2-
C
20H25N2O5 [MH]+:373.1758; found: 373.1770; IR (KBr)
m
hydroxyethyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(16g). Compound 16g (0.14 g, 38.5% yield) was synthesized from
15g (0.37 g, 1.0 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 2.51 (t, 2H,
J = 6.0, CH2), 2.57–2.62 (m, 4H, CH2 Â 2), 3.46 (t, 2H, J = 5.1, CH2),
3.73 (s, 2H, CH2), 4.35 (t, 1H, J = 4.8, CH2OH), 5.94 (s, 2H, OCH2O),
6.19 (d, 1H, J = 15.3, vinyl-H), 6.4 (d, 1H, J = 3.0, furan-H), 6.62–
6.65 (m, 1H, Ar-H), 6.69 (d, 1H, J = 3.0, furan-H), 6.77–6.78 (m, 2H,
Ar-H), 7.21 (d, 1H, J = 15.6, vinyl-H), 9.0 (s, 1H, CONHOH), 10.7 (s,
1H, CONHOH); MS (ESI) m/z 375 (M+H); HRMS calcd for
(cmÀ1) = 3209, 2968, 2868, 1439. HPLC purity: 98. 5%
(tR = 5.66 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in
20 min.
4.1.5.29. (E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](butyl)a-
mino}methyl}furan-2-yl}-N-hydroxyacrylamide (16c). Com-
pound 16c (0.15 g, 39.4% yield) was synthesized from 15c (0.38 g,
1.0 mmol) according to the procedure used to synthesize 10a; oil.
1H NMR (DMSO-d6, 300 MHz, d ppm): 0.84 (t, 3H, J = 7.2, CH3),
1.22 (m, 2H, CH2), 1.27 (m, 2H, CH2), 2.44 (t, 2H, J = 7.1, NCH2),
2.55–2.64 (m, 4H, CH2 Â 2), 3.67 (s, 2H, CH2), 5.94 (s, 2H, OCH2O),
6.19 (d,1H, J = 15.6, vinyl-H), 6.37 (d, 1H, J = 3.3, furan-H), 6.62–
6.66 (m, 1H, Ar-H), 6.69 (d, 1H, J = 3.0, furan-H), 6.77–6.79 (m,
2H, Ar-H), 7.21 (d, 1H, J = 15.6, vinyl-H), 9.0 (s, 1H, CONHOH),
10.7 (s,1H, CONHOH); MS (ESI) m/z 387 (M+H); HRMS calcd for
C
19H23N2O6 [MH]+:375.1551; found: 375.1563; IR (KBr)
m
(cmÀ1) = 3224, 2889, 1726, 1662. HPLC purity: 98. 8% (tR = 4.29 min,
80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.34. (E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](2-phen-
oxyethyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(16h). Compound 16h (0.17 g, 39.1% yield) was synthesized from
15h (0.45 g, 1.0 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 2.68 (m, 4H,
CH2 Â 2), 2.91 (m, 2H, CH2), 3.82 (s, 2H, CH2), 4.1 (m, 2H, CH2),
5.94 (s, 2H, OCH2O), 6.23 (d, 1H, J = 15.6, vinyl-H), 6.42 (d, 1H,
J = 3.0, furan-H), 6.65 (d, 1H, J = 3.0, furan-H), 6.68–6.92 (m, 6H,
Ar-H), 7.30 (d, 1H, J = 15.3, vinyl-H), 7.09–7.53 (m, 2H, Ar-H); MS
C
21H27N2O5 [MH]+:387.1914; found: 387.1926; IR (KBr)
m
(cmÀ1) = 3180, 2954, 2875, 1637, 1577. HPLC purity: 98. 8%
(tR = 6.19 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in
20 min.
4.1.5.30.
(E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](isobu-
tyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(ESI) m/z 449 (MÀH); HRMS calcd for
C
25H27N2O6
(16d). Compound 16d (0.15 g, 39.2% yield) was synthesized from
15d (0.38 g, 1.0 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 0.86 (s, 3H,
CH3), 0.88 (s, 3H, CH3), 1.77 (m, 1H, CH), 2.26 (d, 2H, J = 7.5,
NCH2), 2.62–2.67 (m, 4H, CH2 Â 2), 3.73 (s, 2H, CH2), 6.0 (s, 2H,
OCH2O), 6.25 (d, 1H, J = 15.6, vinyl-H), 6.43 (d, 1H, J = 3.0, furan-
H), 6.68 (d, 1H, J = 3.0, furan-H), 6.71 (m, 3H, Ar-H), 7.26 (d, 1H,
J = 15.6, vinyl-H); MS (ESI) m/z 385 (MÀH); HRMS calcd for
[MH]+:451.1864; found: 451.1880; IR (KBr)
m
(cmÀ1) = 3200,
2968, 2861, 1744, 1609. HPLC purity: 97. 5% (tR = 4.36 min, 80%
MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.35. (E)-3-{5-{{ [2-(Benzo[d][1,3]dioxol-5-yl)ethyl](2-(p-tol-
yloxy)ethyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(16i). Compound 16i (0.21 g, 45.3% yield) was synthesized from
15i (0.46 g, 1.0 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 2.28 (s, 3H,
CH3), 2.73–2.74 (m, 4H, CH2 Â 2), 2.95 (t, 2H, J = 5.2, CH2), 3.87
(s, 2H, CH2), 4.04 (t, 2H, J = 5.1, CH2), 5.99 (s, 2H, OCH2O), 6.27 (d,
1H, J = 15.6, vinyl-H), 6.48 (d, 1H, J = 3.0, furan-H), 6.70–6.85 (m,
5H, Ar-H), 6.76 (d, 1H, J = 3.0, furan-H), 6.86 (d, 2H, J = 8.4, Ar-H),
7.13 (d, 2H, J = 8.4, Ar-H), 7.27 (d, 1H, J = 15.6, vinyl-H); MS (ESI)
m/z 465 (M+H); HRMS calcd for C26H29N2O6 [MH]+:465.2020;
C
21H27N2O5 [MH]+:387.1914; found: 387.1923; IR (KBr)
m
(cmÀ1) = 3293, 2952, 2927, 1730. HPLC purity: 98. 0%
(tR = 6.71 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in
20 min.
4.1.5.31.
(E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](pen-
tyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
found: 465.2034; IR (KBr)
m
(cmÀ1) = 3025, 2918, 1612, 1510. HPLC
(16e). Compound 16e (0.16 g, 41.5% yield) was synthesized from
15e (0.40 g, 1.0 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 0.84 (t, 3H,
J = 6.4, CH3), 1.21–1.63 (m, 6H, CH2 Â 3), 2.43 (t, 2H, J = 6.7,
NCH2), 2.50–2.60 (m, 4H, CH2 Â 2), 3.68 (s, 2H, CH2), 5.94 (s, 2H,
OCH2O), 6.09 (d, 1H, J = 15.3, vinyl-H), 6.37 (s, 1H, furan-H),
6.64–6.80 (m, 4H, 3Ar-H, 1 furan-H), 7.23 (d, 1H, J = 15.6, vinyl-
purity: 98. 3% (tR = 7.16 min, 80% MeCN/20% H2O/0.1% TFA),
0.5 mL/min in 20 min.
4.1.5.36. (E)-3-{5-{{ [2-(Benzo[d][1,3]dioxol-5-yl)ethyl](3-phen-
oxypropyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(16j). Compound 16j (0.17 g, 37.5% yield) was synthesized from
15j (0.46 g, 1.0 mmol) according to the procedure used to synthe-
size 10a; oil. 1H NMR (DMSO-d6, 300 MH, d ppm): 0.91 (m, 2H,
CH2), 1.87 (t, 2H, J = 5.8, CH2), 2.56–2.74 (m, 4H, CH2 Â 2), 3.88
(s, 2H, CH2), 3.96 (t, 2H, J = 6.0, CH2), 5.95 (s, 2H, OCH2O), 6.26 (d,
1H, J = 15.3, vinyl-H),6.43 (d, 1H, J = 3.0, furan-H), 6.73 (d, 1H,
J = 2.4, furan-H), 6.80–7.25 (m, 6H, Ar-H), 7.30 (d, 1H, J = 15.3, vi-
nyl-H), 7.25–7.4 (m, 2H, Ar-H); MS (ESI) m/z 463 (MÀH); HRMS
calcd for C26H29N2O6 [MH]+:465.2020; found: 465.2024; IR (KBr)
H); MS (ESI) m/z 401 (M+H); HRMS calcd for
C22H29N2O5
[MH]+:401.2071; found: 401.2080; IR (KBr) (cmÀ1) = 3200,
m
2968, 2861, 1744, 1609. HPLC purity: 97. 6% (tR = 6.26 min, 80%
MeCN/20% H2O/0.1% TFA), 0.5 mL/min in 20 min.
4.1.5.32. (E)-3-5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](benzyl)a-
mino}methyl}furan-2-yl}-N-hydroxyacrylamide (16f). Com-
pound 16f (0.18 g, 44.2% yield) was synthesized from 15f (0.40 g,
1.0 mmol) according to the procedure used to synthesize 10a; oil.
1H NMR (DMSO-d6, 300 MHz, d ppm): 2.59–2.67 (m, 4H, CH2 Â 2),
3.66 (s, 2H, CH2), 3.68 (s, 2H, CH2), 5.94 (s, 2H, OCH2O), 6.23 (d, 1H,
J = 15.6, vinyl-H), 6.41 (d, 1H, J = 3.0, furan-H), 5.59–6.61 (m, 1H,
Ar-H), 6.70 (d, 1H, J = 3.0, furan-H), 6.76–6.85 (m, 2H, Ar-H), 7.21–
7.42 (m, 5H, Ar-H), 9.04 (s, 1H, CONHOH), 10.77 (s, 1H, CONHOH);
m
(cmÀ1) = 2932, 1601, 1491, 1442. HPLC purity: 98. 8%
(tR = 5.73 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in
20 min.
4.1.5.37. (E)-3-{5-{{[2-(Benzo[d][1,3]dioxol-5-yl)ethyl](4-phe-
noxybutyl)amino}methyl}furan-2-yl}-N-hydroxyacrylamide
(16k). Compound 16k (0.18 g, 37.2% yield) was synthesized from
15k (0.48 g, 1.0 mmol) according to the procedure used to synthe-
MS (ESI) m/z 405 (MÀH); HRMS calcd for
C24H25N2O5
[MH]+:421.1758; found: 421.1767; IR (KBr) (cmÀ1) = 3032, 2940,
m

T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
5353
size 10a; oil. 1H NMR (DMSO-d6, 300 MHz, d ppm): 1.64 (m, 2H,
CH2), 1.74 (m, 2H, CH2), 2.56 (t, 2H, J = 6.2, CH2), 2.58–2.67 (m,
4H, CH2 Â 2), 3.76 (s, 2H, CH2), 3.97 (t, 2H, J = 6.3, CH2), 5.99 (s,
2H, OCH2O), 6.26 (d, 1H, J = 15.6, vinyl-H), 6.44 (d, 1H, J = 3.3, fur-
an-H), 6.72–6.74 (m, 1H, Ar-H), 6.74 (d, 1H, J = 3.0, furan-H), 6.82–
6.85 (m, 2H, Ar-H), 6.94–6.99 (m, 3H, Ar-H), 7.25–7.35 (m, 2H, Ar-
H), 7.32 (d, 1H, J = 15.6,vinyl-H); MS (ESI) m/z 479 (M+H); HRMS
calcd for C27H31N2O6 [MH]+:479.2177; found: 479.2171; IR (KBr)
2VQM) was used for the structural model of HDAC4. Water mole-
cules were removed from three structure. Then, CDOCKER was
used to perform the docking of 10b to the catalytic site of HDAC1,
HDAC4 and HDAC6. All default parameters were used in the dock-
ing process. CHARMm-based molecular dynamics (1000 steps)
were used to generate random ligand 10b conformations, and the
position of any ligand 10b was optimized in the binding site using
rigid body rotation followed by simulated annealing at 700 K. Final
energy minimization was set as the full potential mode. The final
binding conformation of 10b was determined on the basis of en-
ergy, other parameters were set as default. Docking study of ligand
TSA was carried out as described above.
m
(cmÀ1) = 2960, 2875, 1602, 1488. HPLC purity: 99. 2%
(tR = 6.93 min, 80% MeCN/20% H2O/0.1% TFA), 0.5 mL/min in
20 min.
4.2. Bioactivity test
Acknowledgements
4.2.1. General
4.2.1.1. Ligands and materials. SAHA (kind gift of Merck) was
This work was supported by the project 81230078 (key pro-
gram), 81202463 (youth foundation) and 91129732 of National
Natural Science Foundation of China, Program of State Key Labora-
tory of Natural Medicines, China Pharmaceutical University (No.
JKGQ201103), 2012AA020301 of 863 program, 2010ZX09401-401
and 2009ZX09501-003 of the National Major Science and Technol-
ogy Project of China (Innovation and Development of New Drugs).
dissolved in DMSO and used at 5
lM. MS-275 (kind gift from
Schering AG) was dissolved in ethanol and used at 5
l
M. All other
compounds were dissolved in DMSO and used at the indicated
concentrations.
4.2.1.2. Fluorimetric HDACs assays. The HDAC Fluorescent
Activity Assay is based on the Fluor deLys Substrate and Developer
combination (BML-AK500-0001, New York, USA) and has been car-
ried out according to supplier’s instructions. First, the Fluor deLys
Substrate, which comprises an acetylated lysine side chain, has
been incubated with HeLa nuclear extractin presence or absence
of the inhibitors. Deacetylation of the substrate sensitizes the sub-
strate so that, in the second step, treatment with the Developer
produces a fluorophore. Fluorescence has been quantified with a
TECAN inphinite M200 station.
References and notes
1. Arrowsmith, C. H.; Bountra, C.; Fish, P. V.; Lee, K.; Schapira, M. Nat. Rev. Drug
Disc. 2012, 11, 384.
2. Dhanak, D. ACS Med. Chem. Lett. 2012, 3, 521.
3. Brait, M.; Sidransky, D. Toxicol. Mech. Method 2011, 21, 275.
4. McCaughan, J. A.; McKnight, A. J.; Courtney, A. E.; Maxwell, A. P. Transplantation
2012, 94, 1.
5. Dawson, M. A.; Kouzarides, T. Cell 2012, 150, 12.
6. Grunstein, M. Nature 1997, 389, 349.
7. Gregoretti, I.; Lee, Y. M.; Goodson, H. V. J. Mol. Biol. 2004, 338, 17.
8. Hubbert, C.; Guardiola, A.; Shao, R.; Kawaguchi, Y.; Ito, A.; Nixon, A.; Yoshida,
M.; Wang, X. F.; Yao, T. P. Nature 2002, 417, 455.
9. Kovacs, J. J.; Murphy, P. J.; Gaillard, S.; Zhao, X.; Wu, J. T.; Nicchitta, C. V.;
Yoshida, M.; Toft, D. O.; Pratt, W. B.; Yao, T. P. Mol. Cell. 2005, 18, 601.
10. Marks, P. A.; Rifkind, R. A.; Richon, V. M.; Breslow, R. Clin. Cancer Res. 2001, 7,
759.
11. Vannini, A.; Volpari, C.; Gallinari, P.; Jones, P.; Mattu, M.; Carfi, A.; De
Francesco, R.; Steinkuhler, C.; Di Marco, C. EMBO Rep. 2007, 8, 879.
12. Bertrand, P. Eur. J. Med. Chem. 2010, 45, 2095.
4.2.1.3. Cellular proliferation assay. All compounds were dis-
solved in DMSO with the stock concentration of 10 mg/mL, and di-
luted with medium freshly before drug administration. Cell lines
were seeded into 96-well flat bottom plates at density of 4000
cells/well. Twenty-four hours after seeding, each compounddilution
was added in duplicate and incubation continued at 37 °C in a
humidified atmosphere containing 5% CO2. After 48 h, add 20 lL
13. FDA approval and new drug application (NDA) documents for Zolinza (SAHA,
MTT at 5 mg/mL in PBS (filter sterilized, light protected, and stored
at 4 °C) per well, and after 4 h of incubation at 37 °C, the fluorescence
was measured at 570 nm using Thermo Multiskan Spectrum.
vorinostat).
http://www.accessdata.fda.gov/drugsatfda_docs/label/2009/
021991s004lbl.pdf
14. FDA approval and new drug application (NDA) documents for Istodax (FK228,
romidepsin).
022393lbl.pdf
www.accessdata.fda.gov/drugsatfda_docs/label/2009/
4.2.1.4. HDAC1, HDAC4 and HDAC6 enzyme inhibition
assays. HDAC1, HDAC4 and HDAC6 enzyme inhibition assays
were performed by the Reaction Biology Corporation, Malvern,
PA, using the Reaction Biology HDAC Spectrum platform.
(www.reactionbiology.com) The HDAC1, HDAC 4, HDAC 6 assays
used isolated recombinant human protein. Substrate for HDAC1,
HDAC4 and HDAC6 assays is a fluorogenic peptide from p53 resi-
dues 379–382 (RHKKAc). Compounds were dissolved in DMSO
and tested in 10-dose IC50 mode with 3-fold serial dilution starting
15. Zain, J.; Kaminetzky, D.; O’Connor, O. A. Expert Rev. Hematol. 2010, 3, 187.
16. Mai, A.; Altucci, L. Int. J. Biochem. Cell. B. 2009, 41, 199.
17. Sternson, S. M.; Wong, J. C.; Grozinger, C. M.; Schreiber, S. L. Org. Lett. 2001, 3,
4239.
18. Nielsen, T. K.; Hildmann, C.; Dickmanns, A.; Schwienhorst, A.; Ficner, R. J. Mol.
Biol. 2005, 354, 107.
19. 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.
20. 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.
21. Vannini, A.; Volpari, C.; Filocamo, G.; Casavola, E. C.; Brunetti, M.; Renzoni, D.;
Chakravarty, P.; Paolini, C.; De Francesco, R.; Gallinari, P.; Steinkuhler, C.; Di
Marco, S. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 15064.
22. Su, H.; Altucci, L.; You, Q. Mol. Cancer. Ther. 2008, 7, 1007.
23. Furumai, R.; Komatsu, Y.; Nishino, N.; Khochbin, S.; Yoshida, M.; Horinouchi, S.
Proc. Natl. Acad. Sci. U.S.A. 2001, 98, 87.
at 100
10-dose IC50 with 3-fold serial dilution starting at 10
Compound MS275 was tested in a 10-dose IC50 with 3-fold serial
dilution starting at 100 M. IC50 values were extracted by curve-
lM. Control Compound Trichostatin A (TSA) was tested in a
lM. Control
l
fitting the dose/response slopes.
24. Hildmann, C.; Wegener, D.; Riester, D.; Hempel, R.; Schober, A.; Merana, J.;
Giurato, L.; Guccione, S.; Nielsen, T. K.; Ficner, R.; Schwienhorst, A. J. Biotechnol.
2006, 124, 258.
4.C3o. mputational docking
25. Siliphaivanh, P.; Harrington, P.; Witter, D. J.; Otte, K.; Tempest, P.; Kattar, S.;
Kral, A. M.; Fleming, J. C.; Deshmukh, S. V.; Harsch, A.; Secrist, P. J.; Miller, T. A.
Bioorg. Med. Chem. Lett. 2007, 17, 4619.
26. Jones, P.; Altamura, S.; Chakravarty, P. K.; Cecchetti, O.; De Francesco, R.;
Gallinari, P.; Ingenito, R.; Meinke, P. T.; Petrocchi, A.; Rowley, M.; Scarpelli, R.;
Serafini, S.; Steinkuhler, C. Bioorg. Med. Chem. Lett. 2006, 16, 5948.
27. Heltweg, B.; Dequiedt, F.; Marshall, B. L.; Brauch, C.; Yoshida, M.; Nishino, N.;
Verdin, E.; Jung, M. J. Med. Chem. 2004, 47, 5235.
All computational work was done using Discovery Studio 3.0
package (Accelrys Inc.) Compound 10b was built using the ‘Sketch
and Edit Molecule’ module. The HDAC1 and HDAC6 homology
model was constructed by Discovery Studio 3.0 using the Homol-
ogy Modeling protocol as the method described in the literature.41
The crystal structure deposited in the Protein Data Bank (PDB ID

5354
T. Feng et al. / Bioorg. Med. Chem. 21 (2013) 5339–5354
28. Suzuki, T.; Kouketsu, A.; Itoh, Y.; Hisakawa, S.; Maeda, S.; Yoshida, M.;
Nakagawa, H.; Miyata, N. J. Med. Chem. 2006, 49, 4809.
38. Namdar, M.; Perez, G.; Ngo, L.; Marks, P. A. Proc. Natl. Acad. Sci. U.S.A. 2003,
2010, 107.
29. Itoh, Y.; Suzuki, T.; Kouketsu, A.; Suzuki, N.; Maeda, S.; Yoshida, M.; Nakagawa,
H.; Miyata, N. J. Med. Chem. 2007, 50, 5425.
39. Mai, A.; Massa, S.; Pezzi, R.; Rotili, D.; Loidl, P.; Brosch, G. J. Med. Chem. 2003, 46,
4826.
30. Su, H.; Yu, L.; Nebbioso, A.; Carafa, V.; Chen, Y.; Altucci, L.; You, Q. Bioorg. Med.
Chem. Lett. 2009, 19, 6284.
31. Butler, K. V.; Kalin, G.; Brochier, C.; Vistoli, G.; Langley, B.; Kozikowski, A. P. J.
Am. Chem. Soc. 2010, 132, 10842.
32. Schäfer, S.; Saunders, L.; Eliseeva, E.; Velena, A.; Jung, M.; Schwienhorst, A.;
Strasser, A.; Dickmanns, A.; Ficner, R.; Schlimme, S.; Sippl, W.; Verdin, E.; Jung,
M. Bioorg. Med. Chem. 2011, 2008, 16.
33. Schäfer, S.; Saunders, L.; Schlimme, S.; Valkov, V.; Wagner, J. M.; Kratz, F.; Sippl,
W.; Verdin, E.; Jung, M. ChemMedChem 2009, 4, 283.
34. Remiszewski, S. W.; Sambucetti, L. C.; Bair, K. W.; Bontempo, J.; Cesarz, D.;
Chandramouli, N.; Chen, R.; Cheung, M.; Cornell-Kennon, S.; Dean, K.;
Diamantidis, G.; France, D.; Green, M. A.; Howell, K. L.; Kashi, R.; Kwon, P.;
Lassota, P.; Martin, M. S.; Mou, Y.; Perez, L. B.; Sharma, S.; Smith, T.; Sorensen,
E.; Taplin, F.; Trogani, N.; Versace, R.; Walker, H.; Weltchek-Engler, S.; Wood,
A.; Wu, A.; Atadja, P. J. Med. Chem. 2003, 46, 4609.
35. HDAC1,HDAC4,HDAC6 inhibition assays were performed by the Reaction
Biology Corporation,Malvern,PA, using the Reaction Biology HDAC Spectrum
platform. www.reactionbiology.com
40. Mai, A.; Massa, S.; Pezzi, R.; Simeoni, S.; Rotili, D.; Nebbioso, A.; Scognamiglio,
A.; Altucci, L.; Loidl, P.; Brosch, G. J. Med. Chem. 2005, 48, 3344.
41. Lu, H.; Chen, Y.; Yang, B.; You, Q. Acta Pharm. Sin. B. 2011, 1, 240.
42. Bradner, J. E.; West, N.; Grachan, M. L.; Greenberg, E. F.; Haggarty, S. J.;
Warnow, T.; Mazitschek, R. Nat. Chem. Biol. 2010, 3, 238.
43. Avdeef, A.; Tsinman, O. Pharm. Res. 2008, 25, 2613.
44. Avdeef, A.; Bucher, J. J. Anal. Chem. 1987, 50, 2137.
45. Bieliauskas, A. V.; Pflum, M. K. H. Chem. Soc. Rev. 2008, 37, 1402.
46. Witt, O.; Deubzer, H. E.; Milde, T.; Oehme, I. Cancer Lett. 2009, 277, 8.
47. Ontoria, J. M.; Altamura, S.; Di Marco, A.; Ferrigno, F.; Laufer, R.; Muraglia, E.;
Palumbi, M. C.; Rowley, M.; Scarpelli, R.; Schultz-Fademrecht, C.; Serafini, S.;
Steinkühler, C.; Jones, P. J. Med. Chem. 2009, 52, 6782.
48. Schafer, S.; Saunders, L.; Schlimme, S.; Valkov, V.; Wagner, J. M.; Kratz, F.; Sippl,
W.; Verdin, E.; Jung, M. ChemMedChem 2009, 4, 283.
49. Auzzas, L.; Larsson, A.; Matera, R.; Baraldi, A.; Deschênes-Simard, B.; Giannini,
G.; Cabri, W.; Battistuzzi, G.; Gallo, G.; Ciacci, A.; Vesci, L.; Pisano, C.; Hanessian,
S. J. Med. Chem. 2010, 53, 8387.
50. Choi, S. E.; Weerasinghe, S. V. W.; Pflum, M. K. H. Bioorg. Med. Chem. Lett. 2011,
22, 6139.
51. Haru, O.; Haruo, S. Synthesis 1972, 138.
36. Estiu, G.; Greenberg, E.; Harrison, C. B.; Kwiatkowski, N. P.; Mazitschek, R.;
Bradner, J. E.; Wiest, O. J. J. Med. Chem. 2008, 51, 2898.
37. Hideshima, T.; Bradner, J. E.; Wong, J.; Chauhan, D.; Richardson, P.; Schreiber, S.
L.; Anderson, K. C. Proc. Natl. Acad. Sci. U.S.A. 2005, 102, 8567.
52. Brochet, C.; Syssa, J. L.; Mouloungui, Z.; Delmas, M.; Gaset, A. Synth. Commun.
1991, 21, 1735.