Design, synthesis and biological evaluation of novel indole derivatives as potential HDAC/BRD4 dual inhibitors and anti-leukemia agents

Article abstract of DOI:10.1016/j.bioorg.2018.12.011

HDAC inhibitors and BRD4 inhibitors were considered to be potent anti-cancer agents. Recent studies have demonstrated that HDAC and BRD4 participate in the regulation of some signal paths like PI3K-AKT. In this work, a series of indole derivatives that co

Full text of DOI:10.1016/j.bioorg.2018.12.011

BioorganicChemistry84(2019)410–417
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Design, synthesis and biological evaluation of novel indole derivatives as
potential HDAC/BRD4 dual inhibitors and anti-leukemia agents
Gaoliang Cheng, Zhi Wang, Jinyu Yang, Yu Bao, Qihao Xu, Linxiang Zhao, Dan Liu⁎
Key Laboratory of Structure-Based Drugs Design & Discovery of Ministry of Education, Shenyang Pharmaceutical University, Shenyang 110016, China
A R T I C L E I N F O
A B S T R A C T
Keywords:
HDAC inhibitors and BRD4 inhibitors were considered to be potent anti-cancer agents. Recent studies have
demonstrated that HDAC and BRD4 participate in the regulation of some signal paths like PI3K-AKT. In this
work, a series of indole derivatives that combine the inhibitory activities of BRD4 and HDAC into one molecule
were designed and synthesized through the structure-based design method. Most compounds showed potent
HDAC inhibitory activity and moderate BRD4 inhibitory activity. In vitro anti-proliferation activities of the
synthesized compounds were also evaluated. Among them, 19f was the most potent inhibitor against HDAC3
with IC50 value of 5 nM and BRD4 inhibition rate of 88% at 10 μM. It was confirmed that 19f could up-regulate
the expression of Ac-H3 and reduce the expression of c-Myc by western blot analysis. These results indicated that
19f was a potent dual HDAC/BRD4 inhibitor and deserved further investigation.
HDAC and BRD4 dual inhibitor
Indole derivatives
c-Myc
Ac-H3
Anti-proliferative activity
1. Introduction
of CTCL and peripheral T-cell lymphoma (PTCL) [10]. Belinostat (2)
was approved in 2014 by the FDA to treat PTCL [11]. Panobinostat (3)
In the past few years, epigenetic mechanisms have emerged to be
relevant to a wide range of diseases including cancers, diabetes, cardiac
diseases, and neurological disorders [1–4]. Lysine post-translational
modification (PTM), one kind of well-studied epigenetic mechanisms,
plays an extensive role in cell signaling, such as phosphorylation, me-
thylation, acetylation and ubiquitination [5]. Acetylation level of the
lysine in the N-terminal tails of histone is a vital epigenetic mark related
to genes transcription [6]. Over 24,000 lysine acetylations were found
in human cells which indicate that lysine acetylation plays many im-
portant roles in cell signal transduction [7]. Acetylation of the lysine is
regulated by histone acetyltransferases (HATs) and histone deacetylases
(HDACs) which also regarded as “writers” and “erasers”. HDACs cata-
lyze the remove of acetyl to make the chromatin structure constrained,
thereby prevent the transcription. The 18 isoforms of HDACs are
grouped into four classes: class I (HDACs 1, 2, 3 and 8), class II (HDACs
4, 5, 6, 7, 9 and 10), class III (SIRT1-7) and class IV (HDAC 11) [8].
As four drugs have been approved by FDA for the treatment of
lymphoma and multiple myeloma in the past eleven years, HDAC in-
hibitors become a promising therapy for the treatment of cancers.
Vorinostat (1), also known as SAHA, was approved in 2006 by FDA as
the first-in-class HDAC inhibitor for the treatment of cutaneous T-cell
lymphoma (CTCL) [9]. Romidepsin, the only one ratified natural pro-
duct as HDAC inhibitor, was approved in 2009 by FDA for the treatment
was approved in 2015 by FDA as an orphan drug for the treatment of
multiple myeloma (MM) [12].
Bromodomain and extra terminal (BET), another important epige-
netic modulator, also participates in the regulation of genes transcrip-
tion [13]. The BET family proteins can recognize acetylated lysine re-
sidues in histones H3 and H4 and thereby mediate signaling
transduction [14]. The BET family is composed of bromodomain-con-
taining protein 2 (BRD2), BRD3, BRD4, and bromodomain testis spe-
cific protein (BRDT). All the subtypes contain two N-terminal bromo-
domains (BD1 and BD2) and an extra C-terminal domain (ET) [15].
BRD4 was identified in 1988 as a component of the mammalian med-
iator complex; a coactivator plays an essential role in the regulation of
transcription by RNA polymerase II (RNA Pol II) in eukaryotes [16].
BRD4 plays an important role in cellular processes, for instance, tran-
scription, cell proliferation, differentiation and apoptosis [17]. In ad-
dition, BRD4 can enhance expression of many oncogenes, such as c-
Myc, Pim1 and Bcl2. Thus, BRD4 came to be a promising therapeutic
target for cancers. Many BRD4 inhibitors have been reported in the past
few years, including RVX-208, (+)-JQ1 (4), CPI-0610 (5), I-BET151
(6), and some of them are in clinical trials for the treatment of acute
myelocytic leukemia (AML), MM and small cell lung cancer (SCLC)
[18,19] (see Fig. 1).
Recent studies have indicated that HDAC and BRD4 are associated
⁎ Corresponding author.
E-mail address: sammyld@163.com (D. Liu).
https://doi.org/10.1016/j.bioorg.2018.12.011
Received 14 September 2018; Received in revised form 3 December 2018; Accepted 10 December 2018
Availableonline10December2018
0045-2068/©2018ElsevierInc.Allrightsreserved.

G. Cheng et al.
BioorganicChemistry84(2019)410–417
Fig. 1. Structures of some reported HDAC inhibitors and BRD4 inhibitors.
with similar biological phenotypes related to cancer and combination of
the HDAC inhibitor LBH589 and BET inhibitor I-BET151 synergistically
induces apoptosis of melanoma cells [20,21]. Moreover, combination of
Panobinostat and (+)-JQ1 synergistically down-regulates the expres-
sion of N-Myc and Bcl2 [22]. In addition, combination of Mocetinostat
and (+)-JQ1 synergistically inhibits the Ras/MAPK signal pathway
[23]. All these conclusions promote us to develop dual HDAC/BRD4
inhibitors.
consist of a hydrogen bond acceptor group as a mimic of Kac such as 3,
5-dimethylisoxazole, which interact with the conserved asparagine
(Asn140), and a hydrophobic group coupled with the hydrogen bond
acceptor group via a parent nucleus, which occupies the groove of WPF
shelf [24–26] (Fig. 2A). It is notable that the ZA channel, a hydrophobic
tunnel connects BRD4 active pocket and solvent area, is not fully oc-
cupied. Coincidently, the linker group of HDAC inhibitor is probably
favorable to the ZA channel. Based on these findings, we designed and
synthesized a series of compounds as candidate dual HDAC/BRD4 in-
hibitors. Compound 7 was taken as basic skeleton firstly. The reported
BRD4 inhibitors were analysed and compound P-0014 [27] was chosen
because of the similar structure to compound 7. Then the active group
of BRD4 inhibitor P-0014 was introduced to basic skeleton (compound
7) to obtain the novel molecule (Fig. 2C). We hope the new molecule
keeping both HDAC3 inhibitory activity and BRD4 inhibitory power.
the critical segments of both were retained and hybridized to make
the new compounds’ dual inhibitory activities.
We found the compound (7) bearing indole skeleton as HDAC in-
hibitor, IC50(HDAC1) = 0.58 μM, IC50(HDAC3) = 0.06 μM. Docking
study of compound 7 and HDAC3 demonstrated that the hydroxamic
group chelates the zinc ion at the bottom of the HDAC active site and
the carbon chain can occupy the hydrophobic tunnel of the HDAC ac-
tive site. In addition, indole skeleton and the substituent at 3-position
can form hydrophobic interactions with the amino acid residues at the
entrance of the HDAC active site (Fig. 2B). On the other hand, the SAR
of BRD4 inhibitors have been carefully studied. Many BRD4 inhibitors
Fig. 2. (A) Binding mode of P-0014 to BRD4 (PDB ID: 5UOO); (B) Binding mode of compound 7 to HDAC3 (PDB ID: 4A69); (C) Schematic diagram of construct novel
dual BRD4/HDAC inhibitors.
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BioorganicChemistry84(2019)410–417
Scheme 1. Synthesis of target compounds 13a and 13b. Reagents and conditions: (a) NaNO2, SnCl2, HCl (12 mol/L), H2O, 0 °C, 2 h; (b) phenylacetaldehyde, EtOH,
reflux, 5 h; (c) 3,5-Dimethylisoxazole-4-boronic acid pinacol ester, NaHCO3, Pd (dppf) Cl2, 1,2-dimethoxyethane, H2O, reflux, 3 h. (d) Cs2CO3, MeCN, reflux, 4 h; (e)
NH2OH (50 wt% in water), NaOH (2 mol/L), MeOH, r.t., 2 h.
2. Results and discussion
length with n = 1 (19b) made the optimal value, with IC50 values of
0.122 µM, 0.179 µM, 0.118 µM and 0.023 µM against HDACs, HDAC1,
HDAC2 and HDAC3, respectively. For the R group on the phenyl, 3-
position substituted compounds demonstrated the superior HDAC in-
hibitory activity than 2-position and 4-position substituted compounds
(exemplified by 19j, 19l and 19n). In addition, compounds with an
electrophilic R group showed more potent activity than the compounds
with electron-donating R group (exemplified by 19l and 19o).
All the synthesized compounds were evaluated the inhibition effects
for BRD4 at 100 µM, 10 µM and 1 µM, with (+)-JQ1 as positive control.
As shown in Table 1, most of them could inactive BRD4 over 50% ration
at 10 µM. Compound 19f with para-F substitution on phenyl showed the
most powerful inhibitory activity against BRD4 (88%) at 10 µM. Di-
versity length of the chains on 1-positon of indole were tolerable to
BRD4 inhibition due to the long ZA channel in BRD4. All these results
suggested that the designed compounds exhibited HDAC/BRD4 dual
inhibitory activity.
2.1. Chemistry
The synthetic route to target compounds 13a and 13b was illu-
strated as Scheme 1. 4-Bromoaniline was treated with sodium nitrite
and stannous chloride in hydrochloric acid at 0 °C to give p-bromo-
phenylhydrazine hydrochloride (9), which was reacted with phenyla-
cetaldehyde to obtain compound 10. Intermediate 10 was coupled with
3, 5-Dimethylisoxazole-4-boronic acid pinacol ester and the product
was alkylated to give intermediate 12a and 12b. The ester groups of
intermediate 12a and 12b were converted into corresponding hydro-
xamic acid of compounds 13a and 13b through ammonolysis.
Compounds 19a-19o were synthesized from 5-bromine indole by the
route displayed in Scheme 2. 5-bromine indole was acylated at 3-po-
sition to give intermediates 15a-15h, which were hydrogenated by li-
thium aluminum hydride in the next step. Then the final products were
synthesized from intermediates 17a-17h through coupled reaction, al-
kylation reaction and ammonolysis reaction successively.
2.3. Cell growth inhibition assay
The anti-proliferative effects of novel compounds were tested
against THP-1 (human acute monocytic leukemia) cell line with
Vorinostat and (+)-JQ1 as positive controls. The results of the anti-
proliferation assay of synthesized compounds were summarized in
Table 2. All the compounds showed medium potency against THP-1 cell
lines. Compounds 19d and 19o exhibited greatest anti-proliferative
activities, with GI50 value of 8.79 µM and 7.82 µM respectively.
2.2. In vitro HDAC and BRD4 inhibitory activity
To explore the biological activity, the HDAC inhibitory activities of
novel compounds against HeLa nuclear extracts and recombinant
human HDAC1, 2, 3, 6 enzymes were investigated, using Vorinostat as
positive control. As shown in Table 1, all the compounds manifested
inhibitory activities against class I HDACs at submicromolar con-
centration. However, all of them exhibited weaker activities against
HDAC6 with the IC50 values more than 1 µM. The data indicated that
these compounds were selective class I HDACs inhibitors. Ulteriorly,
compound 19f showed selective HDAC3 inhibitory activity with IC50 of
5 nM. QSAR studies suggested that the length of the chain at 1-position
of indole had a significant influence on HDAC inhibitory activity. The
length with 6 carbons was more favorable for HDAC inhibitory activity
than 5 carbons (exemplified by 19c and 19d). The length of the short
chain at 3- position of indole also had influence on the activity. Chain
2.4. Intra-cellular target validation
We then explored whether these compounds could inhibit HDAC
and BRD4 in cellular condition. The levels of c-Myc, acetylated histone
H3 (Ac-H3) and α-tubulin were determined by western blot assay in
THP-1 cells treated with compound 19f at different concentrations for
24 h. In THP-1 cell line, compound 19f promoted acetylation of histone
H3 but not α-tubulin, which proved that compound 19f is a selective
Scheme 2. Synthesis of target compounds 19a-19o. Reagents and conditions: (a) appropriate acyl chloride, AlCl3, CH2Cl2, r.t., 4 h; (b) LiAlH4, THF, r.t., 2 h; (c) 3,5-
Dimethylisoxazole-4-boronic acid pinacol ester, NaHCO3, Pd (dppf) Cl2, 1,2-dimethoxyethane, H2O, reflux, 3 h. (d) Cs2CO3, MeCN, reflux, 4 h; (e) NH2OH (50 wt% in
water), NaOH (2 mol/L), MeOH, r.t., 2 h.
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BioorganicChemistry84(2019)410–417
Table 1
The HDAC and BRD4 inhibitory activities of compounds 13a, 13b and 19a-19o.a
Compd.
R
n
m
IC50 (μM)
BRD4 Inhibition Rate (%)
HDACs
HDAC1
HDAC2
HDAC3
HDAC6
@ 100 μM
@ 10 μM
@ 1 μM
13a
H
5
6
5
6
5
6
5
6
5
6
5
6
5
6
5
6
6
0
0
1
1
2
2
1
1
2
2
1
1
1
1
1
1
1
0.223
0.183
0.160
0.122
0.307
0.172
0.286
0.291
0.207
0.030
0.369
0.149
0.244
0.094
0.332
0.186
0.083
0.076
0.031
0.013
0.021
0.013
0.019
0.021
0.034
0.016
0.018
0.008
0.032
0.009
0.033
0.006
0.041
0.023
0.011
0.002
0.260
0.287
0.189
0.179
0.247
0.240
0.341
0.181
0.259
0.079
0.314
0.220
0.222
0.253
0.650
0.431
0.031
0.103
0.022
0.016
0.011
0.011
0.021
0.017
0.023
0.011
0.031
0.007
0.046
0.031
0.019
0.012
0.053
0.033
0.005
0.008
0.154
0.180
0.110
0.118
0.538
0.139
0.517
0.298
0.946
0.487
> 1
0.023
0.009
0.012
0.019
0.064
0.016
0.073
0.033
0.086
0.034
0.120
0.202
0.089
0.023
0.076
0.028
0.039
0.005
0.017
0.064
0.062
0.050
0.062
0.024
0.072
0.018
0.013
0.027
0.010
0.012
0.007
0.009
0.010
0.005
0.007
0.002
0.004
0.007
0.012
0.009
0.013
0.006
0.011
0.007
0.003
0.006
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
> 1
0.039
86
91
96
96
98
94
96
95
97
95
95
97
94
96
97
95
92
66
76
64
65
49
64
76
88
52
50
59
55
71
69
63
76
41
21
34
16
31
39
24
19
28
9
13b
H
19a
H
19b
H
19c
H
19d
H
19e
4-F
19f
4-F
19g
4-F
19h
4-F
16
18
14
15
14
18
13
12
19i
2-CH3
2-CH3
3-CH3
3-CH3
4-CH3
4-CH3
4-CF3
19j
> 1
19k
0.568
0.127
0.956
0.438
0.079
0.055
0.067
0.013
0.071
0.038
0.010
0.007
19l
19m
19n
19o
Vorinostat
(+)-JQ1
0.002
97
96
95
a
Each value was reproduced in three experiments.
using the Discovery Studio 3.0 software package. Compound 19f was
selected for docking evaluation as representative example and the
predicted binding modes were shown in Fig. 4 below.
Table 2
The anti-proliferative activities of indicated compounds and positive controls
against THP-1 cell lines.a
Compd.
As illustrated by Fig. 4A and B, compound 19f bind to HDAC3 by
embedding the flexible chain into the hydrophobic tunnel and an-
choring the terminal hydroxamic acid group at the bottom of the
pocket. Besides the coordination with zinc ion, extra hydrogen bonds
were found between the eNH of the hydroxamic acid group and HIS134
as well as HIS135. In addition, the indole ring of compound 19f showed
hydrophobic and van der Waals interactions with residues PHE199 and
PHE200 of the receptor protein. Furthermore, the phenyl group formed
a hydrophobic interaction with the residue LEU266. All these ob-
servations may explain the strong enzymatic inhibitory activity of the
synthesized compounds.
GI50(μM, THP-1)
Compd.
GI50(μM, THP-1)
13a
13b
19a
19b
19c
19d
19e
19f
11.29
12.43
12.45
12.13
10.06
8.79
1.22
1.63
2.01
1.39
0.97
19i
14.82
13.93
24.21
23.34
16.00
12.96
7.82
0.78
1.23
2.35
1.87
2.01
1.42
19j
19k
19l
19m
19n
0.65
21.26
15.5
2.54
19o
0.54
1.23
Vorinostat
(+)-JQ1
0.51
0.04
0.02
19g
19h
12.98
14.11
1.35
2.23
0.13
a
Each value was reproduced in three experiments.
The possible binding mode of compound 19f on BRD4 was exhibited
in Fig. 4C and D. The 3, 5-dimethylisoxazole group functioned as a Kac
mimic and interacted with the residue ASN140 through a hydrogen
bond. Moreover, a hydrogen bond was observed between the oxygen
atom of the 3, 5-dimethylisoxazole group and the vicinal hydrone. In
the solvent-exposed area, the hydroxamic acid group formed two hy-
drogen bonds with the residue PRO86. Apart from hydrogen bonding,
the indole ring showed hydrophobic interactions with residues TRP81
and PRO82. Additionally, a π-π interaction was found between the
phenyl group and the residue TRP81. In general, the binding modes
pointed us the key interactions between synthesized compounds and
the two targets which will guide us in the future design.
3. Conclusion
We reported herein a series of indole derivatives as HDAC/BRD4
dual inhibition agents. The specific structure-activity relationship of
these compounds was summarized in detail simultaneously. The enzy-
matic assay revealed that the synthesized compounds exhibited an ex-
cellent potency of inhibiting HDAC and a good potency of inhibiting
BRD4. Compound 19f demonstrated the supreme HDAC inhibitory ac-
tivity with the IC50 value of 5 nM and BRD4 inhibitory activity with the
inhibition rate of 88% at the concentration of 10 µM. All compounds
showed preferable anti-proliferation activity against human acute
monocytic leukemia THP-1 cell lines with the GI50 value of 7.82 µM to
24.21 µM. Compound 19f as the representative compound was used for
further mechanistic studies. The results indicated that the tumor cell
growth inhibitory effects were correlated with the decreased protein
Fig. 3. Validation of HDAC and BRD4 inhibition in THP-1 cells after 24 h of
treatment with compound 19f at 2.5, 5, 10, 20 μM.
class I HDACs inhibitor. This finding is in accord with the enzyme-based
assay. Moreover, the level of c-Myc was efficiently decreased in a dose-
dependent manner (see Fig. 3).
2.5. Docking study
A docking analysis was carried out for further illustrating the pos-
sible binding modes of the synthesized compounds on HDAC and BRD4
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G. Cheng et al.
BioorganicChemistry84(2019)410–417
Fig. 4. Possible binding modes of 19f to HDAC3 (A and B), BRD4 (C and D).
levels of c-Myc and the increased protein levels of Ac-H3. Molecular
docking analysis showed that the hydroxamic acid group could form
hydrogen bonds with the residues of not only HDAC but BRD4. The
indole ring and the phenyl group are also essential for the key hydro-
phobic interactions between the compound and the targets. A novel
class of HDAC/BRD4 dual inhibitors bearing indole scaffold were pro-
vided, and deserved further research.
13.2 mmol) in ethyl alcohol (50 mL) was added hyacinthin (1.5 g,
12.5 mmol) at N2 atmosphere and the reaction was heated to reflux for
3 h. The mixture solution was concentrated to dryness, added water
(100 mL) and extracted with ethyl acetate (80 mL × 3). The organic
layers were combined, washed with brine, dried with Na2SO4 and
evaporated. Finally, the resulting residue was purified by column
chromatography on silica gel as indicated to give 10 as white solid.
To a solution of 10 (0.6 g, 2.2 mmol) in glycol dimethyl ether and
water mixture (30 mL, GDE: H2O = 10:1) were added 3, 5-
Dimethylisoxazole-4-boronic acid pinacol ester (0.54 g, 2.4 mmol),
NaHCO3 (0.56 g, 6.6 mmol) and Pd(dppf)Cl2 (0.1 g, 0.1 mmol) succes-
sively at N2 atmosphere and the reaction was warmed to 80 °C for 4 h.
The mixture solution was added water (100 mL) and extracted with
ethyl acetate (80 mL × 3). The organic layers were combined, washed
with brine, dried with Na2SO4 and evaporated. Finally, the resulting
residue was purified by column chromatography on silica gel as in-
dicated to give 11 as white solid.
4. Experimental
4.1. Chemistry
The melting points were determined on an electrically heated X-4
digital visual melting point apparatus and were uncorrected. Mass
spectra (MS) were determined on a Finnigan MAT/USA spectrometer
(LC–MS). 1H NMR and 13C NMR spectrum were recorded on Bruker AV-
400 or ARX 600 spectrometers with tetramethylsilane (TMS) used as
the internal standard. Chemical shifts were reported in ppm (δ).
High–resolution mass spectra were obtained on Bruker micro TOF–Q in
the ESI mode (HR–ESI–MS). All reactions were performed with com-
mercially available reagents and they were used without further pur-
ification. All reactions were monitored by thin-layer chromatography
(TLC) carried on fluorescent precoated plates GF254 (Qindao Haiyang
Chemical, China) and detection of the components was made by short
UV light. Column chromatography was performed with silica gel 60
(200–300 mesh).
To a solution of 11 (0.5 g, 1.75 mmol) in acetonitrile (30 mL) were
added Cs2CO3 (2.5 g, 5 mmol) and methyl 7-bromoheptanoate (0.44 g,
2 mmol) or methyl 6-bromohexanoate (0.42 g, 2 mmol). The reaction
was warm to 70 °C for 2 h. The mixture solution was added water
(50 mL) and extracted with ethyl acetate (30 mL × 3). The organic
layers were combined, washed with brine, dried with Na2SO4 and
evaporated. Finally, the resulting residue was purified by column
chromatography on silica gel as indicated to give 12a or 12b as white
solid.
To a solution of 12a or 12b (0.5 mmol) in methanol (15 mL) were
added NaOH (2 mol/L, 4 mL) and NH2OH (50 wt% in water, 3 mL)
dropwise successively at 0 °C. The reaction was warmed to room tem-
perature for 3 h. The mixture solution was concentrated to dryness, and
the obtained solid was dissolved in water (10 mL). The resulting solu-
tion was adjusted to neutral with 1 mol/L aqueous solution of HCl and
extracted with CH2Cl2 (10 mL × 3). The organic layers were combined,
washed with brine, dried with Na2SO4 and evaporated. Finally, the
4.1.1. General procedure for the synthesis of 13a-13b
Material 8 (3 g, 17.4 mmol) was added to NaNO2 aqueous solution
(1 mol/L, 50 mL) and the mixture was stirred for 30 min at 0 °C. Then
the mixture was warmed to room temperature for 90 min. A solution of
SnCl2 (12 g, 52.2 mmol) in concentrated hydrochloric acid (10 mL) was
added dropwise to the mixture for 2 h at 0 °C. The resulting precipitate
was filtered to give 9 as white solid. To a solution of 9 (2.9 g,
414

G. Cheng et al.
BioorganicChemistry84(2019)410–417
resulting residue was purified by column chromatography on silica gel
4.1.2.2. 7-(3-benzyl-5-(3,
5-dimethylisoxazol-4-yl)-1H-indol-1-yl)-N-
as indicated to give 13a or 13b as white solid.
hydroxyheptanamide (19b). Yield: 23.6%. Mp: 57.8–58.3 °C. ESI-MS:
m/z, 444.0 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s, 1H),
8.64 (s, 1H), 7.49 (d, J = 8.4 Hz, 1H), 7.34 (s, 1H), 7.31 (d, J = 7.5 Hz,
2H), 7.26 (s, 1H), 7.25 (d, J = 7.8 Hz, 2H), 7.14 (t, J = 7.3 Hz, 1H),
7.07 (d, J = 10.0 Hz, 1H), 4.12 (t, J = 7.1 Hz, 2H), 4.04 (s, 2H), 2.31 (s,
3H), 2.14 (s, 3H), 1.92 (t, J = 7.4 Hz, 2H), 1.74 (dd, J = 13.5, 6.7 Hz,
2H), 1.49–1.43 (m, 2H), 1.28–1.24 (m, 4H). 13C NMR (101 MHz,
DMSO) δ: 169.46, 164.59, 158.84, 142.43, 135.57, 128.79, 128.53,
128.13, 126.99, 126.10, 122.47, 119.95, 119.66, 117.51, 114.14,
110.36, 45.65, 36.45, 32.59, 30.14, 28.59, 26.39, 25.39, 11.67,
10.95. HRMS (ESI+) m/z calcd for C27H31N3O3 [M−H]− 444.2365
found: 444.2222.
4.1.1.1. 6-(5-(3,
5-dimethylisoxazol-4-yl)-3-phenyl-1H-indol-1-yl)-N-
hydroxyhexanamide (13a). Yield: 28.5%. Mp: 88.5–89.6 °C. ESI-MS:
m/z, 416.2 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s, 1H),
8.65 (s, 1H), 7.80 (s, 1H), 7.77 (d, J = 0.8 Hz, 1H), 7.68 (d, J = 7.1 Hz,
2H), 7.64 (d, J = 8.4 Hz, 1H), 7.43 (t, J = 7.8 Hz, 2H), 7.23 (t,
J = 7.3 Hz, 1H), 7.18 (dd, J = 8.5, 1.3 Hz, 1H), 4.23 (t, J = 7.3 Hz,
2H), 2.40 (s, 3H), 2.22 (s, 3H), 1.95 (t, J = 7.4 Hz, 2H), 1.82 (dd,
J = 15.0, 7.5 Hz, 2H), 1.56 (dd, J = 15.0, 7.5 Hz, 2H), 1.30 (dd,
J = 15.0, 7.9 Hz, 2H). 13C NMR (151 MHz, DMSO) δ: 69.40, 164.83,
158.83, 136.22, 135.54, 129.29, 127.93, 126.94, 125.91, 123.10,
121.55, 120.17, 117.42, 115.49, 111.17, 45.94, 32.51, 29.87, 26.27,
25.10, 11.67, 10.95. HRMS (ESI+) m/z calcd for C25H27N3O3 [M−H]−
416.2052 found: 416.1970.
4.1.2.3. 6-(5-(3, 5-dimethylisoxazol-4-yl)-3-phenethyl-1H-indol-1-yl)-N-
hydroxyhexanamide (19c). Yield: 30.8%. Mp: 66.6–67.1 °C. ESI-MS:
m/z, 444.3 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s, 1H),
8.65 (s, 1H), 7.72 (t, J = 7.5 Hz, 1H), 7.49 (s, 1H), 7.27 (s, 2H), 7.26 (s,
2H), 7.19 (s, 1H), 7.17 (dd, J = 8.6, 4.4 Hz, 1H), 7.08 (d, J = 9.7 Hz,
1H), 4.10 (t, J = 7.0 Hz, 2H), 3.01–2.96 (m, 2H), 2.97–2.92 (m, 2H),
2.39 (s, 3H), 2.22 (s, 3H), 1.94 (d, J = 8.6 Hz, 2H), 1.72 (dt, J = 14.7,
7.2 Hz, 2H), 1.51 (d, J = 8.6 Hz, 2H), 1.39–1.31 (m, 2H). 13C NMR
(101 MHz, DMSO) δ: 169.37, 165.04, 158.62, 143.29, 141.61, 134.02,
130.15, 129.68, 128.93, 128.71, 128.65, 126.29, 124.95, 123.68,
116.39, 109.08, 60.96, 44.68, 32.60, 31.71, 26.28, 25.15, 11.69,
10.85. HRMS (ESI+) m/z calcd for C27H31N3O3 [M−H]− 444.2365
found: 444.2279.
4.1.1.2. 7-(5-(3,
5-dimethylisoxazol-4-yl)-3-phenyl-1H-indol-1-yl)-N-
hydroxyheptanamide (13b). Yield: 32.1%. Mp: 72.1–72.9 °C. ESI-MS:
m/z, 430.1 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.31 (s, 1H),
8.64 (d, J = 1.6 Hz, 1H), 7.80 (s, 1H), 7.78 (d, J = 1.1 Hz, 1H), 7.67 (d,
J = 7.1 Hz, 2H), 7.64 (d, J = 8.5 Hz, 1H), 7.43 (t, J = 7.7 Hz, 2H), 7.23
(t, J = 7.4 Hz, 1H), 7.18 (dd, J = 8.4, 1.5 Hz, 1H), 4.24 (t, J = 7.2 Hz,
2H), 2.40 (s, 3H), 2.23 (s, 3H), 1.93 (t, J = 7.4 Hz, 2H), 1.81 (dd,
J = 13.6, 6.7 Hz, 2H), 1.51–1.44 (m, 2H), 1.33–1.27 (m, 4H). 13C NMR
(101 MHz, DMSO) δ: 169.44, 164.82, 158.82, 136.24, 135.55, 129.28,
127.94, 126.91, 125.90, 123.10, 121.55, 120.19, 117.43, 115.49,
111.15, 46.03, 32.57, 30.04, 28.57, 26.43, 25.39, 11.67, 10.95.
HRMS (ESI+) m/z calcd for C26H29N3O3 [M−H]− 430.2209 found:
430.2125.
4.1.2.4. 7-(5-(3, 5-dimethylisoxazol-4-yl)-3-phenethyl-1H-indol-1-yl)-N-
hydroxyheptanamide (19d). Yield: 22.1%. Mp: 70.2–71.0 °C. ESI-MS:
m/z, 458.1 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s, 1H),
8.65 (s, 1H), 7.50 (d, J = 1.4 Hz, 1H), 7.48 (d, J = 8.5 Hz, 1H), 7.26 (s,
2H), 7.25 (s, 2H), 7.18 (s, 1H), 7.17–7.15 (m, 1H), 7.08 (dd, J = 8.4,
1.6 Hz, 1H), 4.10 (t, J = 7.0 Hz, 2H), 3.01–2.97 (m, 2H), 2.97–2.93 (m,
2H), 2.39 (s, 3H), 2.22 (s, 3H), 1.92 (t, J = 7.4 Hz, 2H), 1.74–1.68 (m,
2H), 1.49–1.43 (m, 2H), 1.27–1.21 (m, 4H). 13C NMR (101 MHz,
DMSO) δ: 169.48, 164.54, 158.72, 141.97, 134.98, 129.87, 129.64,
129.55, 129.16, 128.95, 128.87, 128.80, 128.58, 127.92, 117.34,
114.00, 110.48, 48.98, 32.57, 30.38, 28.61, 26.41, 25.37, 11.57,
10.83. HRMS (ESI+) m/z calcd for C28H33N3O3 [M−H]− 458.2522
found: 458.2187.
4.1.2. General procedure for the synthesis of 19a-19o
To a mixture of AlCl3 (1.5 g, 11.6 mmol) and dry CH2Cl2 (30 mL)
was added appropriate acyl chloride (8.5 mmol) dropwise at 0 °C. Then
a solution of 5-bromine indole (1.5 g, 7.6 mmol) in dry CH2Cl2 (20 mL)
was added dropwise to the mixture for 3 h. The mixture solution was
added water (50 mL) and extracted with CH2Cl2 (30 mL × 3). The or-
ganic layers were combined, washed with brine, dried with Na2SO4 and
evaporated. Finally, the resulting residue was purified by column
chromatography on silica gel as indicated to give 15a-15h as light pink
solid.
To a solution of 15a-15h (4.05 mmol) in dry THF (50 mL) were
added LiAlH4 (0.46 g, 12 mmol) and boron fluoride ethyl ether (3 mL,
24 mmol) successively at room temperature for 1 h. The reaction was
quenched by 2 mol/L aqueous solution of HCl and extracted with ethyl
acetate (10 mL × 3). The organic layers were combined, washed with
brine, dried with Na2SO4 and evaporated. Finally, the resulting residue
was purified by column chromatography on silica gel as indicated to
give 16a-16h as white solid.
4.1.2.5. 6-(5-(3, 5-dimethylisoxazol-4-yl)-3-(4-fluorobenzyl)-1H-indol-1-
yl)-N-hydroxyhexanamide (19e). Yield: 18.8%. Mp: 55.6–57.1 °C. ESI-
MS: m/z, 448.0 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s,
1H), 8.66 (s, 1H), 7.49 (d, J = 8.4 Hz, 1H), 7.34 (t, J = 2.3 Hz, 2H),
7.32 (t, J = 2.8 Hz, 1H), 7.27 (s, 1H), 7.08 (dd, J = 6.7, 2.0 Hz, 2H),
7.06 (d, J = 1.7 Hz, 1H), 4.14–4.10 (m, 2H), 4.03 (s, 2H), 2.32 (s, 3H),
2.15 (s, 3H), 1.93 (t, J = 7.3 Hz, 2H), 1.77–1.70 (m, 2H), 1.53 (p,
J = 7.5 Hz, 2H), 1.28–1.20 (m, 3H). 13C NMR (101 MHz, DMSO) δ:
169.35, 164.55, 158.71, 135.74, 130.58, 130.53, 127.83, 127.59,
122.47, 120.06, 119.84, 117.34, 115.29, 115.15, 113.91, 110.53,
45.65, 32.53, 30.40, 29.99, 26.30, 25.11, 11.55, 10.83. HRMS (ESI+)
m/z calcd for C26H28FN3O3 [M−H]− 448.2115 found: 448.2089.
Compounds 19a-19o were synthesized from 16a to 16 h by the
synthetic method from 10 to 13a as white solid.
4.1.2.1. 6-(3-benzyl-5-(3,
5-dimethylisoxazol-4-yl)-1H-indol-1-yl)-N-
hydroxyhexanamide (19a). Yield: 24.9%. Mp: 59.5–60.2 °C. ESI-MS:
m/z, 430.2 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s, 1H),
4.1.2.6. 7-(5-(3,5-dimethylisoxazol-4-yl)-3-(4-fluorobenzyl)-1H-indol-1-
8.65 (dd, J = 7.9, 1.7 Hz, 1H), 7.49 (d, J = 8.5 Hz, 1H), 7.34 (d,
J = 1.2 Hz, 1H), 7.31 (d, J = 7.1 Hz, 2H), 7.27 (s, 1H), 7.25 (t,
J = 6.3 Hz, 2H), 7.14 (t, J = 7.3 Hz, 1H), 7.07 (dd, J = 8.4, 1.6 Hz,
1H), 4.12 (t, J = 7.1 Hz, 2H), 4.04 (s, 2H), 2.31 (s, 3H), 2.14 (s, 3H),
1.94 (t, J = 7.3 Hz, 2H), 1.74 (dt, J = 14.9, 7.3 Hz, 2H), 1.53 (dt,
J = 15.1, 7.5 Hz, 2H), 1.29–1.24 (m, 2H). 13C NMR (101 MHz, DMSO)
δ: 169.40, 164.54, 158.72, 141.96, 134.98, 129.87, 129.55, 128.88,
128.59, 126.77, 126.05, 122.41, 119.90, 117.35, 114.00, 110.49,
48.98, 36.45, 32.50, 30.00, 28.94, 26.29, 25.11, 11.56, 10.84. HRMS
(ESI+) m/z calcd for C26H29N3O3 [M−H]− 430.2209 found: 430.2078.
yl)-N-hydroxyheptanamide (19f). Yield: 19.3%. Mp: 53.9–54.9 °C. ESI-
MS: m/z, 462.2 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s,
1H), 8.65 (s, 1H), 7.49 (d, J = 8.4 Hz, 1H), 7.34 (d, J = 5.9 Hz, 2H),
7.32 (d, J = 5.7 Hz, 1H), 7.26 (s, 1H), 7.09–7.08 (m, 1H), 7.07 (d,
J = 2.4 Hz, 1H), 7.07–7.05 (m, 1H), 4.14–4.10 (m, 2H), 4.03 (s, 2H),
2.32 (s, 3H), 2.15 (s, 3H), 1.92 (t, J = 7.4 Hz, 2H), 1.73 (dt, J = 13.0,
6.3 Hz, 2H), 1.50–1.43 (m, 2H), 1.30–1.24 (m, 4H). 13C NMR
(101 MHz, DMSO) δ: 169.46, 164.55, 158.71, 135.75, 130.57,
130.52, 127.82, 127.59, 122.46, 120.05, 119.84, 117.34, 115.28,
115.14, 113.92, 110.52, 45.70, 32.56, 30.39, 30.12, 28.54, 26.41,
415

G. Cheng et al.
BioorganicChemistry84(2019)410–417
25.41, 11.56, 10.83. HRMS (ESI+) m/z calcd for C27H30FN3O3
1.74 (dt, J = 14.7, 7.3 Hz, 2H), 1.52 (dd, J = 14.8, 7.4 Hz, 2H),
1.28–1.24 (m, 2H). 13C NMR (101 MHz, DMSO) δ: 169.35, 164.52,
158.70, 141.88, 137.54, 135.70, 129.52, 128.48, 127.95, 127.47,
127.28, 126.69, 126.01, 122.37, 119.94, 117.35, 114.08, 110.48,
45.64, 32.53, 31.28, 30.00, 26.30, 25.12, 21.38, 11.56, 10.84. HRMS
(ESI+) m/z calcd for C27H31N3O3 [M−H]− 445.2365 found: 444.2287.
[M−H]− 462.2271 found: 462.2182.
4.1.2.7. 6-(5-(3, 5-dimethylisoxazol-4-yl)-3-(4-fluorophenethyl)-1H-indol-
1-yl)-N-hydroxyhexanamide (19g). Yield: 22.5%. Mp: 61.0–61.3 °C. ESI-
MS: m/z, 462.2 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32 (s,
1H), 8.66 (s, 1H), 7.49 (d, J = 2.1 Hz, 1H), 7.48 (d, J = 4.3 Hz, 1H),
7.28 (dd, J = 10.3, 3.9 Hz, 2H), 7.17 (s, 1H), 7.08 (d, J = 1.6 Hz, 1H),
7.06 (d, J = 8.9 Hz, 2H), 4.10 (t, J = 7.0 Hz, 2H), 2.99–2.95 (m, 2H),
2.95–2.91 (m, 2H), 2.39 (s, 3H), 2.22 (s, 3H), 1.93 (t, J = 7.4 Hz, 2H),
1.75–1.69 (m, 2H), 1.55–1.49 (m, 2H), 1.24–1.20 (m, 2H). 13C NMR
(101 MHz, DMSO) δ: 169.35, 164.59, 158.84, 135.57, 130.54, 130.49,
128.12, 127.01, 122.49, 119.97, 119.67, 117.51, 115.21, 115.08,
113.99, 110.38, 45.58, 35.60, 32.52, 30.00, 27.16, 26.26, 25.12,
11.65, 10.93. HRMS (ESI+) m/z calcd for C27H30FN3O3 [M−H]−
462.2271 found: 462.2193.
4.1.2.12. 7-(5-(3, 5-dimethylisoxazol-4-yl)-3-(3-methylbenzyl)-1H-indol-
1-yl)-N-hydroxyheptanamide (19l). Yield: 23.7%. Mp: 59.0–59.6 °C.
ESI-MS: m/z, 458.4 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32
(s, 1H), 8.64 (s, 1H), 7.48 (d, J = 8.4 Hz, 1H), 7.37 (s, 1H), 7.26 (s, 1H),
7.14 (t, J = 7.4 Hz, 2H), 7.10 (d, J = 7.6 Hz, 1H), 7.07 (dd, J = 8.4,
1.1 Hz, 1H), 6.96 (d, J = 7.3 Hz, 1H), 4.12 (t, J = 6.9 Hz, 2H), 3.99 (s,
2H), 2.33 (s, 3H), 2.23 (s, 3H), 2.16 (s, 3H), 1.92 (t, J = 7.4 Hz, 2H),
1.74 (dd, J = 13.0, 6.7 Hz, 2H), 1.46 (dd, J = 13.9, 6.8 Hz, 2H), 1.26
(d, J = 2.6 Hz, 4H). 13C NMR (101 MHz, DMSO) δ: 169.44, 164.52,
158.71, 141.90, 137.54, 135.69, 129.51, 128.47, 127.95, 127.50,
127.28, 126.69, 126.00, 122.37, 119.94, 117.35, 114.08, 110.47,
45.68, 32.57, 31.26, 30.13, 28.57, 26.43, 25.42, 21.38, 11.56, 10.85.
HRMS (ESI+) m/z calcd for C28H33N3O3 [M−H]− 459.2522 found:
458.2470.
4.1.2.8. 7-(5-(3, 5-dimethylisoxazol-4-yl)-3-(4-fluorophenethyl)-1H-indol-
1-yl)-N-hydroxyheptanamide (19h). Yield: 27.1%. Mp: 78.5–79.1 °C.
ESI-MS: m/z, 476.2 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32
(s, 1H), 8.64 (s, 1H), 7.48 (s, 1H), 7.48 (d, J = 5.2 Hz, 1H), 7.27 (dd,
J = 8.6, 5.7 Hz, 2H), 7.17 (s, 1H), 7.08 (d, J = 2.3 Hz, 1H), 7.06 (dd,
J = 9.2, 2.3 Hz, 2H), 4.10 (t, J = 7.0 Hz, 2H), 2.99–2.95 (m, 2H),
2.95–2.92 (m, 2H), 2.39 (s, 3H), 2.22 (s, 3H), 1.92 (t, J = 7.4 Hz, 2H),
1.74–1.68 (m, 2H), 1.49–1.43 (m, 2H), 1.23 (dd, J = 17.4, 10.3 Hz,
4H). 13C NMR (101 MHz, DMSO) δ: 169.45, 164.59, 158.84, 135.57,
130.53, 130.48, 128.12, 127.04, 122.48, 119.96, 119.67, 117.51,
115.20, 115.06, 113.95, 110.36, 45.64, 35.59, 32.57, 30.13, 28.57,
27.12, 26.38, 25.37, 11.65, 10.93. HRMS (ESI+) m/z calcd for
4.1.2.13. 6-(5-(3, 5-dimethylisoxazol-4-yl)-3-(4-methylbenzyl)-1H-indol-
1-yl)-N-hydroxyhexanamide (19m). Yield: 24.3%. Mp: 61.3–62.2 °C.
ESI-MS: m/z, 444.3 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32
(s, 1H), 8.66 (s, 1H), 7.48 (d, J = 8.4 Hz, 1H), 7.32 (s, 1H), 7.23 (s, 1H),
7.18 (d, J = 7.6 Hz, 2H), 7.06 (d, J = 7.8 Hz, 3H), 4.11 (s, 3H), 3.99 (s,
2H), 2.32 (s, 3H), 2.23 (s, 3H), 2.15 (s, 3H), 1.93 (s, 2H), 1.73 (s, 2H),
1.52 (s, 2H), 1.25 (s, 3H). 13C NMR (101 MHz, DMSO) δ: 169.35,
164.52, 158.71, 145.64, 138.83, 135.72, 134.93, 130.11, 129.98,
129.14, 128.79, 122.38, 119.92, 117.36, 114.25, 110.46, 45.63,
32.53, 30.92, 30.01, 26.30, 25.11, 20.95, 11.55, 10.83. HRMS (ESI+)
m/z calcd for C27H31N3O3 [M−H]− 444.2365 found: 444.2281.
C
28H33FN3O3 [M−H]− 476.2428 found: 476.2341.
4.1.2.9. 6-(5-(3, 5-dimethylisoxazol-4-yl)-3-(2-methylbenzyl)-1H-indol-1-
yl)-N-hydroxyhexanamide (19i). Yield: 32.2%. Mp: 72.1–73.1 °C. ESI-
MS: m/z, 444.3 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.31 (s,
1H), 8.65 (s, 1H), 7.49 (d, J = 8.4 Hz, 1H), 7.34 (d, J = 1.2 Hz, 1H),
7.25–7.22 (m, 1H), 7.15–7.12 (m, 1H), 7.11–7.09 (m, 2H), 7.09–7.06
(m, 2H), 4.11 (t, J = 7.0 Hz, 2H), 4.03 (s, 2H), 2.33 (s, 3H), 2.29 (s,
3H), 2.16 (s, 3H), 1.92 (t, J = 7.4 Hz, 2H), 1.75–1.69 (m, 2H),
1.54–1.48 (m, 2H), 1.23 (d, J = 3.3 Hz, 2H). 13C NMR (101 MHz,
DMSO) δ: 169.34, 164.53, 158.71, 139.57, 136.35, 135.68, 130.40,
129.59, 128.09, 127.82, 126.35, 126.11, 122.39, 119.97, 119.91,
117.37, 112.76, 110.50, 45.63, 32.52, 30.01, 29.22, 26.28, 25.11,
19.55, 11.60, 10.88. HRMS (ESI+) m/z calcd for C27H31N3O3 [M−H]−
444.2365 found: 444.2221.
4.1.2.14. 7-(5-(3, 5-dimethylisoxazol-4-yl)-3-(4-methylbenzyl)-1H-indol-
1-yl)-N-hydroxyheptanamide (19n). Yield: 21.1%. Mp: 65.5–66.0 °C.
ESI-MS: m/z, 458.0 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32
(s, 1H), 8.64 (d, J = 1.1 Hz, 1H), 7.48 (d, J = 8.4 Hz, 1H), 7.32 (d,
J = 1.0 Hz, 1H), 7.23 (s, 1H), 7.18 (d, J = 7.9 Hz, 2H), 7.06 (d,
J = 7.6 Hz, 3H), 4.11 (t, J = 7.2 Hz, 2H), 3.99 (s, 2H), 2.32 (s, 3H),
2.23 (s, 3H), 2.15 (s, 3H), 1.92 (t, J = 7.3 Hz, 2H), 1.73 (dd, J = 13.5,
6.7 Hz, 2H), 1.49–1.42 (m, 2H), 1.29–1.24 (m, 4H). 13C NMR
(101 MHz, DMSO) δ: 169.46, 164.52, 158.71, 145.64, 138.84,
135.72, 134.93, 130.11, 129.97, 129.13, 128.78, 122.37, 119.92,
117.36, 114.26, 110.45, 45.68, 32.57, 30.91, 30.13, 28.55, 26.41,
25.41, 20.95, 11.55, 10.83. HRMS (ESI+) m/z calcd for C28H33N3O3
[M−H]−. 458.2522 found: 458.2446.
4.1.2.10. 7-(5-(3, 5-dimethylisoxazol-4-yl)-3-(2-methylbenzyl)-1H-indol-
1-yl)-N-hydroxyheptanamide (19j). Yield: 26.5%. Mp: 77.4–78.5 °C.
ESI-MS: m/z, 458.3 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.31
(s, 1H), 8.64 (s, 1H), 7.49 (d, J = 8.4 Hz, 1H), 7.35 (d, J = 1.2 Hz, 1H),
7.25–7.21 (m, 1H), 7.15–7.12 (m, 1H), 7.11–7.09 (m, 2H), 7.09–7.06
(m, 2H), 4.11 (t, J = 7.0 Hz, 2H), 4.03 (s, 2H), 2.33 (s, 3H), 2.28 (s,
3H), 2.16 (s, 3H), 1.91 (t, J = 7.4 Hz, 2H), 1.74–1.68 (m, 2H),
1.48–1.40 (m, 2H), 1.23 (s, 4H). 13C NMR (101 MHz, DMSO) δ:
169.44, 164.53, 158.71, 139.58, 136.34, 135.68, 130.41, 129.59,
128.09, 127.85, 126.36, 126.11, 122.39, 119.97, 119.92, 117.38,
112.76, 110.49, 45.67, 32.57, 30.12, 29.22, 28.55, 26.40, 25.42,
4.1.2.15. 7-(5-(3, 5-dimethylisoxazol-4-yl)-3-(3-(trifluoromethyl)benzyl)-
1H-indol-1-yl)-N-hydroxyheptanamide
(19o). Yield:
18.9%.
Mp:
57.2–58.1 °C. ESI-MS: m/z, 512.1 [M−H]−
.
1H NMR (600 MHz,
DMSO‑d6) δ: 10.32 (s, 1H), 8.65 (s, 1H), 7.68 (s, 1H), 7.63 (d,
J = 6.9 Hz, 1H), 7.51 (s, 2H), 7.49 (s, 1H), 7.39 (s, 1H), 7.35 (s, 1H),
7.08 (dd, J = 8.4, 1.5 Hz, 1H), 4.15 (s, 2H), 4.13 (t, J = 7.1 Hz, 2H),
2.31 (s, 3H), 2.13 (s, 3H), 1.91 (d, J = 7.7 Hz, 2H), 1.76–1.71 (m, 2H),
1.51–1.47 (m, 2H), 1.25 (d, J = 7.0 Hz, 4H). 13C NMR (101 MHz,
DMSO) δ: 169.41, 164.54, 158.69, 143.68, 135.67, 133.07, 129.62,
127.83, 127.76, 125.18, 122.86, 122.54, 120.18, 119.77, 117.31,
113.20, 110.60, 45.72, 35.49, 32.79, 30.09, 28.88, 28.04, 27.61,
25.31, 19.03, 11.49, 10.77. HRMS (ESI+) m/z calcd for
19.54, 11.61, 10.88. HRMS (ESI+) m/z calcd for
C28H33N3O3
[M−H]− 458.2522 found: 458.2438.
4.1.2.11. 6-(5-(3, 5-dimethylisoxazol-4-yl)-3-(3-methylbenzyl)-1H-indol-
1-yl)-N-hydroxyhexanamide (19k). Yield: 25.2%. Mp: 69.3–70.7 °C.
C
28H30F3N3O3 [M−H]− 512.2239 found: 512.2143.
ESI-MS: m/z, 444.2 [M−H]−
.
1H NMR (600 MHz, DMSO‑d6) δ: 10.32
(s, 1H), 8.65 (s, 1H), 7.48 (d, J = 8.4 Hz, 1H), 7.36 (s, 1H), 7.26 (s, 1H),
7.13 (d, J = 7.2 Hz, 2H), 7.10 (d, J = 7.8 Hz, 1H), 7.07 (dd, J = 8.4,
1.5 Hz, 1H), 6.96 (d, J = 7.5 Hz, 1H), 4.12 (t, J = 7.1 Hz, 2H), 3.99 (s,
2H), 2.32 (s, 3H), 2.24 (s, 3H), 2.15 (s, 3H), 1.93 (t, J = 7.4 Hz, 2H),
4.2. Biological assays
4.2.1. HDAC inhibition fluorescence assay
In this HDACs assay, HeLa nuclear extracts (Enzo Life Sciences,
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G. Cheng et al.
BioorganicChemistry84(2019)410–417
USA) were used as a source of histone deacetylase. The recombinant
human HDAC1, 2, 3 and 6 were purchased from BPS Bioscience (USA).
All reactions were performed in the black half area 96–well micro-
plates. A serial dilution of the inhibitors (5 µL/well) and enzymes (5 µL/
well) were pre-incubated in HDAC buffer (10 µL/well) at 25 °C for
15 min, and then fluorogenic substrate (5 µL/well) Boc–Lys(Ac)–AMC
was added. After incubation at 37 °C for 60 min, the mixture was
stopped by the addition of developer (25 µL/well) for 10 min.
Fluorescence intensity was measured using the Thermo Scientific
Varioskan Flash Station at excitation and emission wavelengths of 355
(or 360 for the HeLa NuEx) and 460 nm, respectively. The IC50 values
were extracted by curve fitting the dose/response slopes.
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Acknowledgment
This work was supported by the National Nature Science
Foundation of China (No. 81473086).
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