Synthesis of the molecular hybrid inspired by Largazole and Psammaplin A

Article abstract of DOI:10.1016/j.tet.2017.12.025

One important class of HDAC (histone deacetylation enzymes) inhibitors is the sulfur-containing marine natural products with structural diversity. Inspired by two structurally distinguishing examples, Largazole and Psammaplin A, which possess macrocyclic depsipeptide and simple linear amide scaffold respectively, we designed one novel molecular hybrid by replacing the alkene moiety in Largazole with a semirigid amide bond. This hybrid compound has been synthesized from L-malic acid in 10 steps with an overall yield of 7%. The preliminary biological assays suggest that the replacement of trans olefin moiety with amide bond will lead to an unbeneficial effect on the inhibition against HDACs.

Full text of DOI:10.1016/j.tet.2017.12.025

Accepted Manuscript
Synthesis of the molecular hybrid inspired by Largazole and Psammaplin A
Xiaoling Yu, Bingbing Zhang, Guangsheng Shan, Yue Wu, Feng-Ling Yang, Xinsheng
Lei
PII:
S0040-4020(17)31285-1
10.1016/j.tet.2017.12.025
TET 29181
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 12 October 2017
Revised Date: 11 December 2017
Accepted Date: 12 December 2017
Please cite this article as: Yu X, Zhang B, Shan G, Wu Y, Yang F-L, Lei X, Synthesis of the molecular
hybrid inspired by Largazole and Psammaplin A, Tetrahedron (2018), doi: 10.1016/j.tet.2017.12.025.
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ACCEPTED MANUSCRIPT
Graphical Abstract
Synthesis of the molecular hybrid inspired
by Largazole and Psammaplin A
Leave this area blank for abstract info.
Xiaoling Yu a, b , Bingbing Zhang a , Guangsheng Shan a , Yue Wu a , Feng-Ling Yang b , and Xinsheng Lei a*
a School of Pharmacy, Fudan University, 826 Zhangheng Road, Pudong Zone, Shanghai 201203, China
b College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou, Henan Province
450001, China.

1
ACCEPTED MANUSCRIPT
Tetrahedron
journal homepage: www.elsevier.com
Synthesis of the molecular hybrid inspired by Largazole and Psammaplin A
Xiaoling Yu a, b, Bingbing Zhang a, Guangsheng Shan a, Yue Wu a, Feng-Ling Yang b, and Xinsheng Lei a,
a School of Pharmacy, Fudan University, 826 Zhangheng Road, Pudong Zone, Shanghai 201203, China.
b College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou, Henan Province 450001, China.
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
One important class of HDAC (histone deacetylation enzymes) inhibitors is the sulfur-
containing marine natural products with structural diversity. Inspired by two structurally
distinguishing examples, Largazole and Psammaplin A, which possess macrocyclic depsipeptide
and simple linear amide scaffold respectively, we designed one novel molecular hybrid by
replacing the alkene moiety in Largazole with a semirigid amide bond. This hybrid compound
has been synthesized from L-malic acid in 10 steps with an overall yield of 7%. The preliminary
biological assays suggest that the replacement of trans olefin moiety with amide bond will lead
to an unbeneficial effect on the inhibition against HDACs.
Available online
Keywords:
Marine natural product
Largazole
Psammaplin A
Hybrid
2009 Elsevier Ltd. All rights reserved
.
HDAC inhibitors
———
Corresponding author. Tel.: +0-086-21-51980128; fax: +0-086-21-51980128; e-mail: leixs@fudan.edu.cn

2
Tetrahedron
ACCEPTED MANUSCRIPT
effects.2 So far, at least 18 HDAC isoforms have been identified
1. Introduction
and categorized into the following classes: Zn2+-dependent Class
I (HDAC1, 2, 3, 8), Class II (class IIa: HDAC4, 5, 7, 9; class IIb:
HDAC6, 10) and Class IV (HDAC11).3 The importance of
individual HDAC isoforms in cell function and cancer biology is
not fully understood although several pan-selective HDAC
inhibitors, such as SAHA, Belinostat, Panobinostat, Chidamide
and Romidepsin (FK228), have been approved for the treatment
of cutaneous T-cell lymphoma.4 At present, significant efforts are
devoted to explore the class- and even isoform-specific HDAC
The regulation of histone-tailed lysine acetylation by histone
acetyltransferases (HATs) and histone deacetylases (HDACs)
exerts an important epigenetic regulatory mechanism in the
biochemistry of life process.1 The deacetylated histone can form
transcriptionally inactive condensed forms of chromatin, which
can result in silencing of tumor suppressor genes and promoting
cancer initiation and/or progression. Therefore, inhibition of
histone deacetylases will revert hyperacetylation and
transcriptional repression of targeted genes and exert antitumor
inhibitors
in
cancer
therapy.5
Figure 1 The representative HDAC inhibitors and the molecular hybrid based on Largazole and Psammaplin A.
Natural products are widely used as an important source in
drug discovery, and their pivotal importance has been particularly
supported by some antitumor drugs. Largazole was recently
discovered in the realm of natural HDAC inhibitors by Leusch’s
group from a marine cyanobacterium of the genus Symploca.6
The preliminary biological assays revealed Largazole is a potent
and highly selective Class I HDAC inhibitor6, and its superior
HDAC activities and anticancer properties attracted several
research groups in the community of organic and medicinal
chemistry.7-10 As a 16-membered macrocyclic depsipeptide
natural product, Largazole possesses a dense combination of
unusual structural features including its 3-hydroxy-7-
mercaptohept-4-enoic acid unit. Notably, this side chain unit is
commonly shared in several other natural HDAC inhibitors,
structural similarity between the side chain units in Largazole and
Psammaplin A indicates that these units may play a similar role
in the inhibition of HDACs. As a result, we decided to replace
the alkene moiety in Largazole with the semirigid amide moiety.
Herein, we report our work on synthesis of this novel molecular
hybrid 1 based on Largazole and Psammaplin A (Figure 1).
2. Results and discussion
including
Romidepsin
(FK228),11
FR901375,12
and
Spiruchostatins (Figure 1).13 To our interest, this unit appears to
mimic the side chain of the extremely potent HDAC inhibitor
Psammaplin A, a member of another family of natural products
isolated from several marine sponges (Figure 1).14 All these
natural products can be activated in vitro and in vivo through
either reductive cleavage of a disulfide bond or hydrolysis of
thioester to expose the free sulfhydryl residue, which coordinates
with Zn2+ cation of the HDACs, exerting inhibitory effects
against HDACs (Figure 1).
Figure 2 Retrosynthetic analysis of the target compound.
Amide bond is considered as a good isostere for trans olefin
The convergent synthetic approach to the hybrid was adopted
as shown in Figure 2. The target compound could be assembled
based on the steric demand, bond length and angle15. The

3
from three units, L-Fmoc-Valine, the thiazole-thiazoline
fragment (12), and the side chain fragment 16. Fragment 16
could be prepared from commercially available L-malic acid 13
and amine 21 via amide formation. The linear depsipeptides were
then subjected to deprotection at the N- and C-terminus followed
by macrolactamization at the upper or lower amide bond, and
thiol deprotection to afford the macrolactam thiol, and further
acylation would complete the synthesis of the hybrid (1).
As shown in Scheme 1, methyl α-methylcysteine HCl 5 was
ACCEPTED MANUSCRIPT
readily accessible with the requisite (R)-stereochemistry via the
Pattenden modification of the Seebach protocol on L-cysteine
methyl ester.17-18 After that, the thiazole-thiazoline fragment 11a
was prepared via several steps from Glycine amide HCl
according to Xie’s modified protocol16, and the overall yield was
in line with that described. Then 11a could be hydrolyzed to the
free acid (12a) or deprotected to the free amine (12b).
To accomplish the synthesis of hybrid 1, thiazole-thiazoline
fragment (11a) was prepared according to the known method.16-18
Scheme 1 Preparation of the advanced intermediate 12a and 12b.
With the required fragments in hand, we proceeded to
construct the modified depsipeptide 1. Initially, the key
macrolactamization was chosen at the upper amide (Scheme 2).
Thus, L-malic acid 13 was condensed in anhydrous DMSO at rt
with hexafluoroacetone, prepared from CF3COCF3ꢀ3H2O in the
presence of concentrated sulfuric acid,19 giving the masked acid
14a in 86% yield. In order to obtain the desired amide with the
naked acid group of 14a and the amine 12b, tremendous effort
was demonstrated that the selective condensation is difficult,
generally giving the undesired amide (Scheme 2). The ester
group of 14a was very reactive due to the two CF3 groups, thus
amidation took place there to form the undesired product.
Considering that there might be somewhat trouble in the selective
hydrolysis of the methyl ester group in the precursor of 20
(Scheme 2) in the late stage, we gave up the macrolactamization
at the upper amide.
Subsequently, TFA-mediated removal of the t-Boc and 2-
(trimethylsilyl)ethoxy groups was achieved in DCM at 0 C – rt
o
for overnight. After examination of several macrocyclization
conditions, compound 20 was obtained in 77% yield over two
steps, using HATU/HOBT/DIPEA for the condensation in
anhydrous DCM solution (the concentration of cyclization
precursor at about 0.001 M).
Our alternative approach was to conduct the key
macrolactamization at the lower amide (Scheme 3). 13 was
condensed with Cl3CCH(OH)2 in the presence of concentrated
sulfuric acid,20 giving the more stable masked acid 14b in 95%
yield, which was further esterified with TMSCH2CH2OH (TSE-
OH) or MeOH to give 15a and 15b. Then, 15b was smoothly
demasked in ethanol with amine 21, which was prepared from
Ph3COH and HSCH2CH2OH in CF3CO2H at rt for 3 h (86%),21 to
give the amide 16b in 83% yield. Upon the activation of Fmoc-L-
valine with 2,4,6-Cl3-C6H2COCl in the presence of DMAP and
DIEPA in CH2Cl2 (DCM)23, the condensation with 16b afforded
intermediate 17b in 66% yield. Removal of Fmoc group was
performed with Et2NH in CH3CN at rt for 3 h, giving the crude
18b, which was subjected to EDCI/HOBT-mediated coupling
with the acid 12a to give 19b in 77% yield.
Scheme 2 Attempt to synthesize hybrid 1 via the key
macrolactamization at the upper amide.

4
Tetrahedron
ACCEPTED MANUSCRIPT
Scheme 3 Synthesis of hybrid 1 and its thiol.
Noteworthily, we also attempted to execute the key
Finally, deprotection of Trt group in 20, using (i-Pr)3SiH
(TIPS) and CF3CO2H in anhydrous DCM solution, proceeded
smoothly to afford the free thiol (hybrid 1 thiol) in 70% yield,
and the subsequent acetylation with acetyl chloride under the
standard condition led to 1 in 63% yield. The structures of 20 and
1 were further determined by 1H, 13C NMR, COSY, HMQC and
HMBC spectra.
macrolactamization with 19a. Unfortunately, the selective
hydrolysis of the methyl ester in 19a was found to be difficult
under various conditions, which generally resulted in the
elimination reaction to give the corresponding fumaramide in a
significant yield.
Table 1. HDACs inhibitory activities and antiproliferative activities of Largazole, Hybrid 1 and their free thiols.
O
O
S
S
N
N
NH
O
NH
O
N
S
N
S
O
O
O
O
H
N
RS
N
H
RS
N
H
O
Largazole
: R= C7H15CO
Largazole thiol: R= H
Hybrid 1
: R= C7H15CO
Hybrid 1 thiol
: R= H
Compound
IC50 (nM)a
IC50 (µM)b
HCT116
0.184±0.014
ND
HDAC1
228
HDAC2
1520
0.9
HDAC3
HDAC8
120
HDAC6
A549
MDA-MB-231
1.37±0.416
NDc
Laragzole
Largazole thiol
Hybrid 1
2002
0.7
16140
102
0.467±0.104
ND
0.4
42
> 10000
1777
14
> 10000
2076
19
> 10000
2146
18
> 10000
> 10000
388
> 10000
> 10000
12
> 10
> 10
> 10
Hybrid 1 thiol
SAHA
ND
ND
ND
d
a SAHA was used as a positive control. Values are means of three experiments, and standard error of the IC50 was generally less than 10%.
b Laragzole was used as a positive control.
c Not detected at 10 µM.
d Not tested.
To examine the influence on the biological activities through
the replacement of the trans alkene in Largazole with the
semirigid amide bond, Laragzole and its free thiol were prepared
according to the known method.16 Then, we evaluated their
biological activities according to the known method22, including
HDACs inhibition (class I HDACs: HDAC1−3, 8; and a
representative of Class II HDACs: HDAC6) and cytotoxicities
against A549 (human lung carcinoma), HCT116 (human
colorectal carcinoma), and MDA-MB-231 (human breast
carcinoma) cell lines. The results are summarized in Table 1.
Compared with SAHA, Largazole thiol is a class I HDAC
inhibitor with highly potency (HDAC1-3: IC50 0.4-07 nM,
HDAC8: IC50 42 nM) and with a good selectivity over class II
HDAC (HDAC6: IC50 102 nM, selectivity of HDAC6/1: 250).
Largazole is a weaker HDAC inhibitor than its free thiol and
SAHA, and can be considered as a prodrug of the thiol to

5
ameliorate cell permeability. In our cell assay, Largazole exhibits
obvious antiproliferative activities against A549 (IC50 0.47 µM),
4.2.
2-(triethylsilyl)ethyl
2-((2R/S,4S)-5-oxo-2-
ACCEPTED MANUSCRIPT
(trichloromethyl)-1,3-dioxolan-4-yl)acetate (15b
)
HCT116 (0.18 µM) and MDA-MB-231 (1.37 µM), but the thiol
had no activities due to its poor cell permeability. These results
are consistent with those reported.6,10
To a stirring solution of 14b (1.06 g, 4.0 mmol), 2-
(trimethylsilyl)ethanol (0.29 mL, 2.0 mmol) and DMAP
(0.25 g, 0.2 mmol) in DCM, (50 mL), a solution of EDCI
(0.767 g, 4.0 mmol) in DCM (10 mL) was added dropwise
Although the synthetic hybrid 1 is structurally different from
Largazole only on the replacement of trans olefin moiety with
amide moiety, we unfortunately found Hybrid 1 free thiol did not
display obvious inhibitory effect on HDACs, and furthermore,
Hybrid 1 had no obvious antiproliferative activities against A549,
HCT116 and MDA-MB-231. One possible reason is that the
semirigid amide bond results in the off-range orientation of the
free sulfhydryl residue compared with the trans double bond,
which can’t coordinates well with Zn2+ cation of the HDACs.
The molecular modeling studies to account for the reduced
binding affinity of this analogue are under way.
o
at 0 C, and then continued to stir for overnight at rt. The
reaction solution was evaporated under reduce pressure, and
the residue was then dissolved into ethyl acetate (EA, 200
mL). The organic phase was washed with H2O (3 × 50 mL),
brine (50 mL) and dried with Na2SO4. After filtration and
evaporation, the residue was chromatographed on silica gel
with petroleum ether (PE)/EA (30/1) as the eluent, yielding
colourless oil (0.602 g, 83%). [α]20 26.5 (c 0.63, CHCl3),
D
1H NMR (400 MHz, CDCl3): δ 5.91 (m, 1H, C
HCCl3), 4.70-
4.90 (m, 1H, OC
H), 4.20-4.30 (m, 2H, OCH2), 3.04-2.95 (m,
2H, CH2CO), 1.08-1.00 (m, 2H, CH2SiMe3), 0.05 (s, 9H,
CH2SiMe3), 13C NMR (150 MHz, CDCl3): δ 170.3, 169.0,
105.1, 97.8, 71.5, 64.1, 35.8, 17.3, -1.6, ESI-MS m/z 385.2,
387.1 [M+Na]+, HRMS-ESI m/z [M+Na]+, calcd for
C11H17Cl3O5SiNa 384.9803, found 384.9801.
3. Conclusions
In conclusion, inspired by the similarity between the side
chains of two natural HDAC inhibitors Largazole and
Psammaplin A, we designed a new analogue by replacing the
trans olefin in Largazole with semirigid amide bond. The
synthesis of this analogue was accomplished in 10 steps with an
overall yield of 7%. The preliminary biological assays suggest
that the trans olefin moiety in this family of the sulfur-containing
despeptides, especially in Largazole, is very important structural
unit in the inhibition against HDACs, and its amide type isostere
will lead to an unbeneficial effect.
4.3. S-Tritylaminoethanethiol (21
)
Aminoethanethiol (2.32 g, 30 mmol) was dissolved in 50
mL of TFA. Triphenylcarbinol (7.81 g, 30 mmol) was added
to the solution portionwise, under stirring at room
temperature, until the solution became clear. The reaction
mixture, a dense deeply red liquid, was stirred for 3 h and
then distilled to remove the solvent under reduce pressure.
The residue was dissolved in a mixture of DCM and 3.0 M
NaOH aqueous solution, and then the organic layer was
isolated, washed with water and brine, and dried with
MgSO4. After filtration and evaporation, the residue was
chromatographed on silica gel with DCM/MeOH (15/1) as
the eluent, yielding yellowish solid (8.21 g, 86%). Mp 91-93
oC,21 1H NMR (400 MHz, CDCl3): δ 7.18-7.47 (m, 15H,
4. Experimental section
The chemicals and reagents were purchased from Acros, Alfa
Aesar, and National Chemical Reagent Group Co. Ltd., P. R.
China, and used without further purification. Anhydrous solvents
(THF, MeOH, DMF, DCM, and CH3CN) used in the reactions
were dried and freshly distilled before use. All the reactions were
carried out under Ar atmosphere, otherwise stated else. The
progress of the reactions was monitored by TLC (silica-coated
glass plates) and visualized under UV light, and by using iodine
or phosphomolybdic acid. Melting points were measured on a
SGW X-4 microscopy melting point apparatus without
ArH), 2.58 (t, J = 6.4 Hz, 2H, NCH2CH2), 2.32 (t, J = 6.4 Hz,
2H, SCH2CH2), 1.08 (brs, 2H, NH2), ESI-MS m/z 320.2
[M+H]+.
4.4.
(S)-2-(trimethylsilyl)ethyl
3-hydroxy-4-oxo-4-((2-
)
1
correction. H NMR and 13C NMR spectra were recorded either
(tritylthio)ethyl)amino)butanoate (16b
on a 400 MHz Varian Instrument at 25 °C or 600 MHz Bruke
Instrument at 25 °C, using TMS as an internal standard,
respectively. Multiplicity is tabulated as s for singlet, d for
doublet, dd for doublet of doublet, t for triplet, and m for
multiplet. HRMS spectra were recorded on Finnigan-Mat-95
mass spectrometer, equipped with ESI source. Experimental
procedures for the preparation of compound 11a and Largazole
are prepared as the reported literature.16
A solution of 21 (1.75 g, 5.47 mmol), 15b (2.38 g, 6.56
mmol) in anhydrous EtOH (50 mL) was stirred for overnight
at rt and then distilled to remove the solvent under reduce
presure. The residue was chromatographed on silica gel with
DCM/MeOH (70/1) as the eluent, yielding a foamly solid
1
(2.06 g, 90%). [α]20 -16.4 (c 0.50, CHCl3), H NMR (400
D
MHz, CDCl3): 7.50-7.30 (m, 15H, Ar
1H, N ), 4.35 (m, 1H, OC CH2), 4.18 (t,
OCH2CH2SiMe3), 3.86 (d, = 4.0 Hz, 1H, O
NHCH2CH2S), 2.87 (dd, = 17.2, 3.6 Hz, 1H, OCHCH2),
2.61 (dd, = 17.2, 8.8 Hz, 1H, OCHCH2), 2.37 (t, = 6.4 Hz,
2H, NHCH2CH2S), 0.97 (t, 8.4 Hz, 2H,
H
), 6.91 (t,
= 8.4 Hz, 2H,
), 3.08 (m, 2H,
J = 5.6 Hz,
H
H
J
H
4.1. Chloralides of L-Malic Acid (14b
)
J
J
A mixture of chloral hydrate (18.8 g, 114 mmol), L-malic
acid (13.5 g, 100 mmol) and concentrated H2SO4 (25 mL)
was stirred for 3 h at rt. The reaction mixture was then
poured into iced H2O (250 mL) with stirring. The collected
precipitate was washed with H2O (2 × 50 mL) and dissolved
in EtOAc (EA, 250 mL). The organic layer was washed with
sat. NaCl (2 × 50 mL) solution, dried with anhydrous
J
J
J
=
OCH2CH2SiMe3), 0.09 (s, 9H, OCH2CH2SiMe3), 13C NMR
(150 MHz, CDCl3): δ 173.3, 172.0, 144.9, 129.9, 128.3,
127.1, 68.8, 67.1, 63.8, 38.5, 38.1, 32.2, 17.6, -1.2, ESI-MS
m/z 536.7 [M+H]+, HRMS-ESI m/z [M+Na]+, calcd for
C30H37NO4SSiNa 558.2105, found 558.2108.
Na2SO4 and evaporated to give chloralide of L-malic acid
o
(
14b, 24.6 g, 93%) as a white solid. Mp 130-132 C.20 1H
4.5.
butoxycarbonyl)amino)-3-methylbutanoyl)oxy)-4-oxo-4-((2-
(tritylthio)ethyl)amino)butanoate (17b
(S)-2-(trimethylsilyl)ethyl
3-(((S)-2-((tert-
NMR (400 MHz, CDCl3): δ 3.0-3.2 (m, 2H, CH2CO), 4.80-
5.00 (m, 1H, OC
H), 5.8-6.0 (m, 1H, CHCCl3), ESI-MS m/z
)
285.2 [M+Na]+.

6
Bioorganic & Medicinal Chemistry
To a stirring solution of Fmoc-L-Valine (2.79 g,
4.8.
(S)-2-(trimethylsilyl)ethyl
3-(((S)-2-((R)-2'-(((tert-
butoxycarbonyl)amino)methyl)-4-methyl-4,5-dihydro-[2,4'-
bithiazole]-4-carboxamido)-3-methylbutanoyl)oxy)-4-oxo-4-
ACCEPTED MANUSCRIPT
8.21mmol), DIPEA (2.72 mL, 1.65 mmol) in DCM (20 mL),
2,4,6-Cl3-C6H2COCl (1.72 mL, 11.0 mmol) was added
dropwise at 0 OC, and then continued to stir for 1.5 h. Then a
((2-(tritylthio)ethyl)amino)butanoate (19b
)
solution of 16b (2.89 g, 5.48 mmol) and DMAP (0.736 g
，
A solution of EDCI (0.461 g, 2.4 mmol) in DCM (10 mL)
was added dropwise to the stirring solution of 18b (1.53 g,
2.4 mmol), 12a (0.715 g, 2.0 mmol) and HOBT (0.325 g, 2.4
mmol) in DCM (20 mL) over 30 min at rt, and then
continued to stir for 3 h. After removal of the solvent under
reduce pressure, the residue was dissolved in EA (200 mL),
the organic phase was washed with a sat. NaHCO3 solution
(3 × 50 mL), brine (50 mL), and dried with Na2SO4. After
filtration and evaporation, the residue was chromatographed
on silica gel with EA as the eluent, yielding a white solid
(1.50 g, 77%). Mp 63-65 oC, [α]20D -45.6 (c 0.24, CHCl3), 1H
NMR (600 MHz, CDCl3) δ 7.90 (s, 1H, ArHH), 7.46-7.15 (m,
6.02 mmol) in DCM (10 mL) was added dropwise to the
stirring reaction solution, and stirred for another 3 h at rt.
After evaporation under reduce precursor, and the residue
was then dissolved into DCM (200 mL). The organic phase
was washed with H2O (3 × 50 mL), brine (50 mL) and dried
with Na2SO4. Upon filtration and evaporation, the residue
was chromatographed on silica gel with PE/EA (5/1) as the
eluent, yielding a foamy solid (3.08 g, 66%). [α]20 -7.10 (c
D
1
0.55, CHCl3), H NMR (400 MHz, CDCl3): δ 7.91 (d,
J =
8.0 Hz, 2H, Ar
Ar , N CO2CH2CH), 6.82 (brs, 1H, N
OC CH2), 5.28 (brs, 1H, C NHFmoc), 4.48-4.59 (m, 2H,
NHCO2CH2CH), 4.36-4.39 (m, 1H, NHCO2CH2C ), 4.27-
4.32 (m, 2H, OCH2CH2SiMe3), 3.14-3.19 (m, 2H,
NHCH2CH2S), 3.19 (dd, = 17.2, 4.0 Hz, 1H, OCHCH2),
2.90 (dd, = 17.2, 8.0 Hz, 1H, OCHCH2), 2.49-2.55 (m, 2H,
NHCH2CH2S), 2.33-2.42 (m, 1H, C Me2), 0.96 (t, = 6.8
Hz, 2H, OCH2CH2SiMe3), 0.90-0.80 (m, 6H, CH e2), 0.02
H
), 7.69 (m, 2H, Ar
H
), 7.30-7.56 (m, 18H,
H
H
H), 5.63-5.65 (m, 1H,
H
H
16H, Ar
5.45 (dd,
CH2N Boc), 4.64 (d,
= 6.0, 4.8 Hz, 1H, Me2CHC
H2CH2SiMe3), 3.67 (d, = 12.0 Hz, 1H, CqCH2S), 3.30-
= 12.0 Hz, 1H,
= 16.8, 3.6 Hz, 1H, CH2CO), 2.93 (m,
1H, CH2NHCO), 2.77 (dd, = 16.8, 9.0 Hz, 1H, CH2CO),
2.50 (m, 1H, NHCH2CH2S), 2.40 (m, 1H, NHCH2CH2S)
2.27 (m, 1H, Me2C CH), 1.52 (s, 3H, CH3), 1.48 (s, 9H,
Me3), 0.98 (m, 2H, CH2CH2SiMe3), 0.93 (d, = 7.2 Hz, 3H,
Me2CHCH), 0.92 (d, = 7.2 Hz, 3H, Me2CHCH), 0.03 (s,
H
, CHN
= 9.0, 3.6 Hz, 1H, CO2C
= 5.4 Hz, 2H, CH2NHBoc), 4.24 (dd,
NHCO), 4.16 (m, 2H,
H
CO), 7.09 (t,
J
= 5.4 Hz, 1H, CH2N
HCO),
H
J
HCH2), 5.30 (brs, 1H,
H
J
J
J
H
J
C
J
H
J
3.20 (m, 1H, CH2NHCO), 3.07 (d,
CqCH2S), 3.00 (dd,
J
M
J
(s, 9H, CMe3), 13C NMR (150 MHz, CDCl3): δ 170.8, 170.0,
168.3, 157.2, 144.9, 144.0, 143.9, 141.7, 129.7, 128.3, 128.1,
127.5, 127.1, 125.3, 120.4, 120.4, 71.0, 67.4, 67.1, 63.7,
59.7, 47.5, 38.6, 37.0, 31.8, 30.9, 19.5, 17.6, 17.6, -1.2, ESI-
MS m/z 879.4 [M+Na]+, HRMS-ESI m/z [M+Na]+, calcd for
C50H56N2O7SSiNa 879.3470, found 379.3471.
J
,
H
C
J
J
9H, SiMe3). 13C NMR (150 MHz, CDCl3) δ 175.5, 169.5,
169.0, 168.8, 167.6, 162.9, 155.0, 147.9, 144.1, 129.0, 127.3,
126.1, 121.1, 84.2, 82.4, 79.9, 70.2, 66.1, 62.7, 57.7, 41.8,
40.1, 37.9, 36.4, 30.9, 29.3, 27.7, 24.2, 18.6, 17.1, 16.7, -2.1.
ESI-MS m/z 996.4 [M+Na]+, HRMS-ESI m/z [M+Na]+,
calcd for C49H63N5O8S3SiNa 996.3500, found 996.3512.
4.6.
(S)-2-(trimethylsilyl)ethyl
3-(((S)-2-amino-3-
methylbutanoyl)oxy)-4-oxo-4-((2-
(tritylthio)ethyl)amino)butanoate (18b
)
A solution of 17b (1.33 g, 1.55 mmol), Et2NH (0.8 mL,
7.8 mmol) in anhydrous CH3CN (20 mL) was stirred for 3 h
at rt and then distilled to remove the solvent under reduce
presure. The residue was chromatographed on silica gel with
PE/EA (1/1) as the eluent, yielding a colorless oil (0.660 g,
4.9. (5R,8S,11S)-8-isopropyl-5-methyl-6,9,13-trioxo-N-(2-
(tritylthio)ethyl)-10-oxa-3,17-dithia-7,14,19,20-
tetraazatricyclo[14.2.1.12,5]icosa-1(18),2(20),16(19)-
1
triene-11-carboxamide (20
)
68%). H NMR (400 MHz, CDCl3): 7.19-7.39 (m, 15H,
Ar
4.8 Hz, 1H, OC
3.29 (d, = 5.2 Hz, 1H, C
1H, OCHCH2), 2.81 (dd,
2.38 (t,
H
), 6.52 (t,
J
H
= 5.2 Hz, 1H, N
CH2), 4.10-4.20 (m, 2H, OCH2CH2SiMe3),
H), 5.46-5.49 (dd, J = 7.2,
Trifluoroacetic acid (3.0 mL) was added dropwise to the
stirring solution of 19b (0.293 g, 0.3 mmol) in DCM (15 mL)
J
HNH2), 2.91 (dd,
J = 16.8, 4.8 Hz,
o
over 30 min at 0 C, and then warmed to rt and stirred for
J
= 16.8, 4.8 Hz, 1H, OCHCH2),
overnight. After removal of the solvent under reduce
pressure, the residue was dissolved in toluene (Tol, 50 mL)
and evaporated in vacuo, and then cycled again to remove
CF3CO2H. The residue was subsequently treated with ether
(50mL), and resulted a white precipitate. After filtration, the
white crude product was dissolved in acetonitrile (300 mL,
about 1.0 mM solution), and to the solution was added
dropwise a solution of HATU (0.229 g, 0.6 mmol), HOBT
(0.082g, 0.6 mmol) in acetonitrile (10 mL) at rt over 30 min.
Sixteen hours later, the reaction mixture was evaporated
under reduce pressure, the residue was dissolved in EA (200
mL), the organic phase was washed with a sat. NaHCO3
solution (3 × 50 mL), brine (50 mL), and dried with Na2SO4.
After filtration and evaporation, the residue was
chromatographed on silica gel with EA as the eluent,
yielding a white foamy solid (0.144 g, 64%). [α]20D -0.256 (c
J
= 6.4 Hz, 2H, NHCH2CH2S), 1.97-2.06 (m, 1H,
C
H
Me2), 0.96 (t, = 6.4 Hz,, 2H, OCH2CH2SiMe3), 0.93 (d,
J
J
= 6.4 Hz, 3H, CHMe2), 0.86 (d, J = 6.4 Hz, 3H, CHMe2),
0.02 (s, 9H, CMe3), ESI-MS m/z 635.3 [M+H]+.
4.7. (R)-2'-(((tert-butoxycarbonyl)amino)methyl)-4-methyl-
4,5-dihydro-[2,4'-bithiazole]-4-carboxylic acid (12a
)
A LiOH aqueous solution (2.0 M, 1.70 mL, 5.2 mmol)
was added dropwise to a stirring solution of 11a (0.983 g,
2.6 mmol) in THF/H2O (4/1, 25 mL) at rt, and then
continued to stir for 2 h. After quenched with a sat. KHSO4
aqueous solution (30 mL), the mixture was extracted with
EA (3 × 50 mL), the combined organic phase was washed
with brine (3 × 50 mL), and dried with MgSO4. After
filtration and evaporation, the residue was chromatographed
on silica gel with EA as the eluent, yielding a white solid
(0.790 g, 85%). Mp 75-77 oC.16 [α]20D -52.6 (c 0.60, CHCl3),
1H NMR (400 MHz, CDCl3): δ 9.77 (s, 1H, CO2H), 7.89 (s,
1
0.39, CHCl3), H NMR (400 MHz, CDCl3): δ 7.72 (s, 1H,
ArH
H
), 7.57 (t,
J
= 6.0 Hz, 1H, SCH2CH2N
CO), 7.16-7.30 (m, 15H, Ar
= 6.0 Hz, 1H, ArHCH2N CO), 5.46 (m, 1H, CO2C
H
CO), 7.41 (d,
J
= 6.0 Hz, 1H, CHN
H
H), 6.54 (t,
1H, Ar
H
), 5.38 (brs, 1H, N
H), 4.65 (d, J = 6.4 Hz, 2H,
J
H
HCH2),
C
H2NHBoc), 3.87 (d,
J
= 11.2 Hz, 1H, CH2S), 3.13 (d, J =
5.00 (dd,
8.0, 4.0 Hz, 1H, Me2CHC
J
= 18, 8.0 Hz, 1H, ArHCH2NHCO), 4.53 (dd,
NHCO), 4.07 (dd, = 18, 4.0
Hz, 1H, ArHCH2NHCO), 3.92 (d, = 12 Hz, 1H, CqCH2S),
J =
11.2 Hz, 1H, CH2S), 1.57 (s, 3H, CH3), 1.47 (s, 9H, CMe3),
H
J
ESI-MS m/z 380.1 [M+Na]+.
J

7
3.27 (d,
NHCH2CH2S, CH2CO), 2.72 (dd, 1H,
H2CO), 2.36 (t, = 8.0 Hz, 2H, NHCH2CH2S), 2.01-2.13
(m, 1H, Me2C CH), 1.69 (s, 3H, CqCH3), 0.84 (d, = 6.0
J
= 12 Hz, 1H, CqCH2S), 3.27-3.06-3.17 (m, 3H,
33.4, 32.0, 30.1, 29.2, 28.4, 25.9, 25.0, 22.9, 19.5, 17.6,
ACCEPTED MANUSCRIPT
J
= 16, 8.0 Hz, 1H,
14.4. HRMS-ESI m/z: [M+H]+, calcd for C28H42N5O6S3
C
J
640.2292, found 640.2293.
H
J
Hz, 3H, Me2CH), 0.70 (d,
J
= 6.0 Hz, 3H, Me2CH), 13C
Acknowledgments
NMR (150 MHz, CDCl3) δ HRMS-ESI m/z [M+Na]+, calcd
for C39H41N5NaO5S3 778.2162, found 778.2168.
We thank National Natural Foundation of China (No.
21472024 and No. 21242008) and National High-tech R&D
Program of China (2013AA092903) for the financial support. We
are grateful to Dr. Han-Qing Dong and Dr. Jie-Fei Cheng for
their helpful discussion.
4.10.
(5R,8S,11S)-8-isopropyl-N-(2-mercaptoethyl)-5-
methyl-6,9,13-trioxo-10-oxa-3,17-dithia-7,14,19,20-
tetraazatricyclo[14.2.1.12,5]icosa-1(18),2(20),16(19)-
triene-11-carboxamide (hybrid
1 thiol)
References and notes
To a solution of 20 (0.144 g, 0.2 mmol) in DCM (20 mL)
at 0 C was added triisopropylsilane (0.082 mL, 0.4 mmol)
o
1.
2.
3.
Delcuve, G. P.; Khan, D. H.; Davie, J. R Clin. Epigenetics 2012,
4, 5.
Manal, M.; Chandrasekar, M. J. N.; Jeyapal Gomathi Priya, J. G.;
Nanjan, M. J. Bioorg. Chem. 2016, 67, 18.
followed by trifluoroacetic acid (0.2 mL, 2.69 mmol). After
being stirred for 1.5 h at rt, the reaction mixture was
concentrated in vacuo. The crude product was purified by
flash chromatography on silica gel with ethyl acetate to
obtain the free thiol (0.078 g, 70%), as a white solid. Mp
a) De Ruijter, A. J.; Van Gennip, A. H.; Caron, H. N.; Kemp, S.;
Van Kuilenburg, A. B. Biochem. J. 2003, 370, 737; b) Johnstone,
R. W. Nat. Rev. Drug Discovery 2002, 1, 287; c) Emanuele, S.;
Lauricella,,M; Tesoriere, G. Int. J. Oncol. 2008, 33, 637.
Pan, D.-S.; Yang, Q.-J.; Fu, X.; Shan, S.; Zhu, J.-Z.; Zhang, K; Li,
Z.-B.; Ning, Z.-Q.; Lu, X.-P. MedChemComm 2014, 5, 1789.
a) Roche, J.; Bertrand, P. Eur. J. Med. Chem. 2016, 121, 451; b)
Balasubramanian, S.; Verner, E.; Buggy, J. J. Cancer Lett. 2009,
280, 211; c) Tabackman, A. A.; Frankson, R.; Marsan, E. S.; Perry,
K.; Cole, K. E. J. Struct. Biol. 2016, 195, 373.
a) Taori, K.; Paul, V. J.; Luesch, H. J. Am. Chem. Soc. 2008, 130,
1806; b) Ying,Y.; Taori,K.; Kim, H.; Hong, J.; Hendrik Luesch, H.
J. Am. Chem. Soc. 2008, 130, 8455; c) Bowers, A.; West, N.;
Taunton, J.; Schreiber, S. L.; Bradner, J. E.; Williams, R. M. J.
Am. Chem. Soc. 2008, 130, 11219.
o
1
128-129 C, [α]20 -6.06 (c 0.132, CHCl3), H NMR (600
D
MHz, CDCl3) δ 7.77 (s, 1H, ArHH), 7.75 (brs, 1H,
4.
5.
SCH2CH2N
(brs, 1H, ArHCH2N
(dd, = 17.4, 7.8 Hz, 1H, ArHCH2NHCO), 4.57 (dd,
4.2 Hz, 1H, Me2CHC NHCO), 4.32 (dd, = 17.4, 4.8 Hz,
1H, ArHCH2NHCO), 3.91 (d, = 11.4, 1H, CqCH2S), 3.52-
3.40 (m, 3H, NHCH2CH2S, CqCH2S), 3.25 (dd, = 14.4, 4.2
Hz, 1H, CH2CO), 2.82 (dd, = 14.4, 5.4 Hz, 1H, CH2CO),
2.64-2.45 (m, 2H, NHCH2CH2S), 2.10-2.14 (m, 1H,
Me2C CH), 1.81 (s, 3H, CqCH3), 0.85 (d, = 6.6 Hz, 3H,
Me2CH), 0.74 (d,
= 6.6 Hz, 3H, Me2CH). 13C NMR (150
H
CO), 7.25 (d,
J
= 9.6 Hz, 1H, CHN
H
CO), 6.75
CH2), 5.16
= 9.0,
HCO), 5.57 (m, 1H, CO2C
H
J
J
H
J
J
6.
7.
J
J
H
J
Maolanon, A. R.; Kristensen, H. M. E; Leman, L. J.; Ghadiri, M.
R.; Olsen, C. A. ChemBioChem. 2016, 17, 1, and references
therein.
Reddy, D. N.; Ballante, F.; Chuang. T.; Pirolli, A.; Marrocco, B;
Marshall, G. R. J. Med. Chem. 2016, 59, 1613.
J
MHz, CDCl3) δ 172.7, 169.5, 168.1, 147.1, 124.5, 84.4, 84.2,
71.8, 60.4, 58.3, 43.1, 42.6, 41.4, 38.4, 33.0, 32.0, 29.7, 29.4,
24.7, 24.4,19.2, 17.2, 14.2. HRMS-ESI m/z: [M+ H]+, calcd
for C20H28N5O5S3 514.1247, found 514.1245.
8.
9.
Yao, Y.; Tu, Z; Liao, C; Wang, Z; Li, S; Yao, H; Li, Z; Jiang, S. J.
Med. Chem. 2015, 58, 7672.
10.
11.
12.
Clausen, D. J.; Smith, W. B.; Haines, B. E.; Wiest, O.; Bradner, J.
E.; Williams, R. M. Bioorg. Med. Chem. 2015, 23, 5061.
Ueda, H.; Nakajima, H.; Hori, Y.; Fujita, T.; Nishimura, M.; Goto,
T.; Okuhara, M. J. Antibiot. 1994, 47, 301.
a) Shigematsu, N.; Ueda, H.; Takase, S.; Tanaka, H. J. Antibiot.
1994, 47, 311; b) Ueda, H.; Manda, T.; Matsumoto, S.;
Mukumoto, S.; Nishigaki, F.; Kawamura, I.; Shimomura, K. J.
Antibiot. 1994, 47, 315.
4.11. S-(2-((5R,8S,11S)-8-isopropyl-5-methyl-6,9,13-trioxo-
10-oxa-3,17-dithia-7,14,19,20-
tetraazatricyclo[14.2.1.12,5]icosa-1(18),2(20),16(19)-
triene-11-carboxamido)ethyl) octanethioate (hybrid 1
)
To a stirred solution of the free thiol (n0.029 g, 0.057
o
mmol) in DCM (10 mL) at 0 C were added Et3N (16 µL,
13.
14.
Masuoka, Y.; Nagai, A.; Shin-ya, K.; Furihata, K.; Nagai, K.;
Suzuki, K.; Hayakawa, Y.; Seto, H. Tetrahedron Lett. 2001, 42,
411.
Baud, M. G. J.; Leiser, T.; Haus, P.; Samlal, S.; Wong, A. C.;
Wood, R. J.; Petrucci, V.; Gunaratnam, M.; Hughes, S. M.;
Buluwela, L.; Turlais, F.; Neidle, S.; Meyer-Almes, F.-J.; White,
A. J. P.; Fuchter, M. J. J. Med. Chem. 2012, 55, 1731.
Hann, M. M.; Sammes, P. G.; Kennewell, P. D.; Taylor, J. B. J.
Chem. Soc., Perkin Trans. I 1982, 307.
Xiao, Q.; Wang, L.-P.; Jiao, X.-Z; Liu, X.-Y.; Wu, Q.; Xie,P. J.
Asian Nat. Prod. Res. 2010, 12, 940.
Jeanguenat, A.; Seebach, D. J. Chem. Soc., Perkin Trans. I 1991,
2291.
Mulqueen, G. C.; Pattenden, D.; Whiting, D. A. Tetrahedron
1993, 49, 5359.
Burger, K.; Windeisen, E.; Pires, R. J. Org. Chem. 1995, 60, 7641.
Rankin, G. O.; Sun, H.; McCain, B.; Hubbard, J. L.; Chai, M.
Synthesis, 2002, 2165.
Maltese, M. J. Org. Chem. 2001, 66, 7615.
Chen, F.; Gao, A.-H.; Li, J.; Nan, F.-J. ChemMedChem 2009, 4,
1269.
0.114 mmol) and octanoyl chloride (0.068 mL, 0.339 mmol).
Then the mixture was warmed to rt and stirred for 2 h, the
reaction was quenched with methanol (10 mL) and the
mixture was concentrated in vacuo. Purification of the crude
product by column chromatography on silica gel with EA as
solvent gave hybrid
[α]20D -2.61 (c 0.537, CHCl3), 1H NMR (600 MHz, CDCl3) δ
7.76 (s, 1H, ArHH), 7.66 (brs, 1H, SCH2CH2N CO), 7.24
(d, = 9.0 Hz, 1H, CHN CO), 6.62 (brs, 1H,
ArHCH2N CO), 5.54 (brs, 1H, CO2C CH2), 5.15 (d,
13.8 Hz, 1H, ArHCH2NHCO), 4.58-4.60 (dd, = 9.0, 4.2
Hz,1H, Me2CHC NHCO), 4.32 (d, = 13.8 Hz, 1H,
ArHCH2NHCO), 3.94 (d, = 10.8 Hz, 1H, CqCH2S), 3.40-
3.41 (m, 2H, NHCH2CH2S), 3.36 (d, = 10.8 Hz, 1H,
CqCH2S), 3.24 (d, = 10.2 Hz, 1H, CH2CO), 2.90-2.97 (m,
2H, NHCH2CH2S), 2.79 (d, = 10.2 Hz, 1H, CH2CO), 2.50
(t, = 7.8 Hz, 2H, C6H13CH2CO), 2.13-2.14 (m, 1H,
Me2C CH), 1.83 (s, 3H, CqCH3), 1.62 (m, 2H,
CH3CH2C4H8CH2CO), 1.26-1.33 (m, 8H,
CH3CH2C4H8CH2CO), 0.88 (t, 6.0 Hz, 3H,
H3C6H13CO), 0.86 (d, = 6.0 Hz, 3H, Me2CH), 0.74(d,
1 (0.023 g, 63%, as a white foamy solid.
15.
16.
17.
18.
H
J
H
H
H
J =
J
H
J
J
19.
20.
J
J
21.
22.
J
J
H
23.
Bhansali,P.; Hanigan, C. L.; Casero Jr., R. A.; Tillekeratne, L. M.
V. J. Med. Chem. 2011, 54, 7453.
J
=
C
J
J =
13
6.0 Hz, 3H, Me2CH). C NMR (150 MHz, CDCl3) δ 173.2,
169.8, 168.6, 168.0, 72.2, 58.6, 44.5, 43.5, 41.9, 40.0, 38.7,

8
Bioorganic & Medicinal Chemistry
ACCEPTED MANUSCRIPT
Supplementary Material
Supplementary data (Synthetic procedures for preparation of
known compounds and NMR spectra for the important
compounds) associated with this article can be found.