Design, synthesis and preliminary activity assay of 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid derivatives as novel Histone deacetylases (HDACs) inhibitors

Article abstract of DOI:10.1016/j.bmc.2010.01.060

Histone deacetylases (HDACs) are enzymes involved in tumor genesis and development. Herein, we report a novel series of 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid derivatives as HDACs inhibitors. The preliminary biological screening showed that most

Full text of DOI:10.1016/j.bmc.2010.01.060

Bioorganic & Medicinal Chemistry 18 (2010) 1761–1772
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Design, synthesis and preliminary activity assay of
1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid derivatives as novel
Histone deacetylases (HDACs) inhibitors
Yingjie Zhang a, Jinhong Feng a, Chunxi Liu b, Lei Zhang a, Jie Jiao a, Hao Fang a, Li Su a, Xiaopan Zhang a,
Jian Zhang a, Minyong Li a, Binghe Wang c, Wenfang Xu a,
*
a Department of Medicinal Chemistry, School of Pharmacy, Shandong University, Ji’nan, Shandong, 250012, PR China
b Department of Pharmaceutics, School of Pharmacy, Shandong University, Ji’nan, Shandong, 250012, PR China
c Department of Chemistry, Georgia State University, Atlanta, 30303, USA
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 enzymes involved in tumor genesis and development. Herein, we
report a novel series of 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid derivatives as HDACs inhibitors.
The preliminary biological screening showed that most of our compounds exhibited potent inhibitory
Received 2 January 2010
Revised 27 January 2010
Accepted 28 January 2010
Available online 6 February 2010
activity against HDACs. Within this series, five compounds, 13a (IC50 = 0.58 0.10
1.00 0.16 M), 8l (IC50 = 1.06 0.14 M), 7i (IC50 = 1.17 0.19 M) and 7a (IC50 = 1.29 0.15
sessed better HDACs inhibitory activity than Vorinostat (IC50 = 1.48 0.20 M). So these five compounds
l
M), 7d (IC50
=
l
l
l
l
M) pos-
l
Keywords:
could be used as novel lead compounds for further design of HDACs inhibitors. The anti-proliferative
activities of a few compounds and the structure–activity relationships are also briefly discussed.
Ó 2010 Elsevier Ltd. All rights reserved.
1,2,3,4-Tetrahydroisoquinoline-3-carboxylic
acid derivatives
Histone deacetylases
Inhibitor
Synthesis
1. Introduction
cause many diseases, of which cancer is the most dreadful. It has
been shown that global hypoacetylation of histone H4 is a common
HDACs belong to a family of enzymes that remove the acetyl
group from the -amide groups of lysine residues located on the
hallmark of cancer, and changes in H4 acetylation happen early in
the tumorigenesis.8 Many experiments revealed that overexpres-
sion of class I and II HDACs, especially class I isozymes, is associ-
ated with many kinds of tumors.9–16
Up to now, many kinds of HDAC inhibitors (HDACi) have been
studied for their therapeutic effects on cancers,17 including
hydroxamates, cyclic peptides, aliphatic acids and benzamides.18
Suberoylanilide hydroxamic acid (SAHA, Vorinostat) (Fig. 1), the
first HDACi approved by the FDA in 2006 for treatment of cutane-
ous T-cell lymphoma, validated HDACi as a strategy for cancer
therapy.19,20
Figure 1 also showed the common pharmacophore of HDACi,
which includes three parts, the Zn2+ binding group, linker and sur-
face recognition domain. In the design of HDACi, much attention
has been paid to the ‘linker’ optimization. One effective strategy
is replacing the flexile carbochain linker with conformationally re-
stricted structures. Four clinical trial HDAC inhibitors containing ri-
gid aromatic rings or heterocyclic rings in the ‘linker’ are
represented in Figure 1. Encouraged by these successful com-
pounds, we focused our attention on searching for other rigid ac-
tive structures as the ‘linker’ of HDACi. Based on our literatures
investigation, we found that Tic, short for 1,2,3,4-tetrahydroiso-
e
nucleosomal histones. The hydrolysis of the acetyl group results in
the consequence that the positive charge density on the N-termini
of nucleosomal histones increases, which strengthens the interac-
tion with the negatively charged DNA and blocks the access of tran-
scription factors.1–3 In this way, HDACs involve in the remodeling of
chromatin and down-regulate many genes expression. In addition,
deacetylation of nucleosomal histones plays roles in the other gen-
ome function, including chromatin assembly, DNA repair and
recombination.4,5 Recently, more and more non-histone proteins,
such as transcription factors, cytoskeletal proteins, molecular chap-
erones and nuclear import factors, have been revealed as the HDACs
substrates.6 HDACs are divided into four distinct classes. Class I, II
and IV HDACs are zinc-dependent metalloproteinase, whereas class
III HDACs (sirtuins) have a unique mechanism of deacetylation
dependent on the NAD+ cofactor.7
Because of the multiple functions of HDACs in gene expression
and other important cell processes, dysregulation of HDACs will
* Corresponding author.
E-mail address: wfxu@yahoo.cn (W. Xu).
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.01.060

1762
Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
O
H
N
SAHA
HO
N
H
(Vorinostat)
O
Zinc ion
Binding
Group
Surface
Recognition
Domain
Linker
H
N
N
H
N
N
H
HO
N
N
O
H
N
HO
HO
O
Panobinostat (LBH-589) (Phase III)22
SB-939 (Phase I)21
O
O
S
NEt2
N
H
O
N
N
N
H
N
H
N
O
N
HO
O
O
ITF-2357 (Phase II)23
R306465 (Phase I)24
Figure 1. The structures of SAHA and four clinical trial HDACi containing rigid aromatic ring or heterocyclic ring in the ‘linker’. The three parts of HDACi pharmacophore are
indicated in three different colors, respectively.21–24
quinoline-3-carboxylic acid, is a kind of non-protein
a
-amino acids
should be noted that the structure of our designed Tic derivative
is quite consistent with the common pharmacophore of HDACi.
with distinct geometrical conformation and biological activity. The
structure of Tic has been used in many compounds designs (Fig. 2).
The most successful example was substituting the Tic residue for
the proline residue of enalapril. This modification led to a new ap-
proved angiotensin converting enzyme inhibitor (ACEi), quinapril
(Fig. 3).32 However, there is little research on the structure of Tic
in HDACi design. In this paper, we introduce the rigid Tic fragment
to the ‘linker’ of HDACi and design a novel HDACi scaffold, which is
shown in Figure 4a. The chirality of our target structure is deter-
mined by the chirality of HDACs peptide substrate (Fig. 4b). It
2. Chemistry
The target compounds were synthesized following the proce-
dures described in Schemes 1 and 2. The starting material 3, 5-diio-
do-L-tyrosine, compound 1 was converted to compound 2 by
Pictet–Spengler cyclization according to the literature.33 The com-
pound 2 was protected by Boc, followed by catalytic hydrogenoly-
sis to give the key intermediate 4. The compound 4 was coupled
R2
O
X
R1
NH
R2
R10 R7 R6
O
N
m
n
O
N
R9
O
R1
N
O
O
R
5 R4
R3
N
N
R8
Y
n
NH2
O
α vβ 3 integrins antagonist25
Prolyl endopeptidases
inhibitor 26
OH
SH
δ - Opioid receptor
H
antagonist or agnoist27, 28
N
R3
NH2
O
O
O
S
O
R2
N
N
H
NH
R4
R3
N
R1
O
N
O
O
O
R4
N
S
R1
OH
O
O
R5
R2
Farnesyl transferase
inhibitor29
κ - Opioid receptor antagonist30
MMP-8 inhibitor31
Figure 2. Compounds containing the fragment of 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid (Tic). The structure of the Tic fragment is indicated in red.25–31

Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
1763
3. Result and discussion
Considering the available X-ray crystal structure of human
HDAC8 and the high homology of all HDACs isoforms, all the target
compounds were tested for their inhibitory ability against HDAC8,
with Vorinostat (SAHA) being used as positive control.
From the results listed in Table 1, it could be found that most of
our target compounds showed comparable inhibitory activity with
O
N
O
N
N
H
N
H
O
O
COOH
O
O
COOH
enalapril32
quinapril (an approved ACEi)32
Vorinostat, and compounds 13a (IC50 = 0.58 0.10
1.00 0.16 M), 8l (IC50 = 1.06 0.14 M), 7i (IC50 = 1.17 0.19
M) and 7a (IC50 = 1.29 0.15 M) were even more potent than
Vorinostat (IC50 = 1.48 0.20 M). In the 7 series compounds,
lM), 7d (IC50 =
l
l
Figure 3. Replacing the proline residue of enalapril led to a new approved drug
quinapril.
l
l
l
although two more potent compounds, 7d and 7i were obtained
by modification of the R1 phenyl group of 7a, most modification
of the R1 phenyl group of 7a seemed detrimental to the activity,
such as, inserting methylene between the phenyl group and amide
group (7b and 7c), introducing substituent groups to the phenyl
group (7e–7h, 7j–7m) and replacing the phenyl group with ali-
phatic groups (7n–7p). However, this trend was converse in the
8 series compounds. Compared with compound 8a, other com-
pounds were more potent except 8p. Generally, compounds with
aromatic R1 groups (7a–7m, 8a–8m) were more potent than com-
pounds with aliphatic R1 groups (7n–7p, 8n–8p).
with a few kinds of aromatic or aliphatic amines by classical DCC/
HOBT method to yield 5a–5p, and then converted to the corre-
sponding 6a–6p. The ester groups of 6a–6p were treated with
NH2OK in methanol to get the compounds 7a–7p. Finally the com-
pounds 8a–8p were obtained by deprotecting the Boc groups of
7a–7p.
The compounds 13a–13c were synthesized according to the
methods described in Scheme 2. The hydantoin structures of com-
pounds 13b and 13c could be supported by 1H NMR, HRMS and
many literatures.34–36 We suppose that the cyclization of 13b
and 13c might result from the nucleophilic attack of the amide N
atom to the carbonyl C atom of Cbz group. However, the exact
mechanism of hydantoin formation and the reason why 13a do
not form a hydantoin structure need deep investigation.
Significantly, almost all 7 series compounds except 7j and 7l
possessed lower IC50 value than that of their corresponding 8 series
compounds. For example, the IC50 of 7a was 1.29 0.15
lM, much
lower than that of 8a (8.21 1.68 M). The most notable exception
l
was compound 8l, which was much more potent than 7l. We pre-
O
O
*
R1
H
N
N
C-terminus
*
H
H
N
N
N
H
HO
O
O
HN
N-terminus
O
O
Surface
Recognition
Domain
Zinc ion
Binding
Group
Linker
a. (S)-Tic derivatives as HDACi
b. Peptide Substrate of HDACs
Figure 4. (a) The novel HDACi scaffold containing Tic fragment in the ‘linker’. (b) The structure of the HDACs peptide substrate.
O
O
O
O
I
I
I
c
OH
a
OH
OH
NH.HCL
b
OH
Boc
N
NH2
N
HO
Boc
HO
HO
HO
I
I
I
1
2
3
4
O
O
O
R1
R1
R1
d
e
N
N
f
N
H
N
H
H
H
N
O
N
N
HO
O
O
Boc
Boc
HO
Boc
O
O
5a-5p
6a-6p
7a-7p
O
R1
g
N
H
H
NH.HCL
N
HO
O
O
8a-8p
Scheme 1. Reagents and conditions: (a) (CH2O)n, 37% HCl, CH3OCH2CH2OCH3, 72–75 °C, 18 h; (b) (Boc)2O, 1 N NaOH, THF; (c) 10% Pd–C, H2, Et3N, MeOH; (d) R1NH2, DCC,
HOBT, anhydrous THF; (e) BrCH2COOHCH3, K2CO3, anhydrous DMF; (f) NH2OK, CH3OH; (g) HCl, anhydrous EtOAc.

1764
Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
O
O
O
O
I
R2
b
c
OH
OH
NH.HCl
a
OH
Cbz
N
H
NH
N
N
HO
HO
HO
HO
R2
Cbz
I
2
9
10
11a-11c
O
N
H
Cbz
13a, R2:
H
N
N
N
O
HO
O
O
R2
d
O
O
N
e
H
O
O
N
O
Cbz
13b, R2:
13c, R2:
O
N
R2
H
N
12a-12c
HO
O
Scheme 2. Reagents and conditions: (a) 10% Pd–C, H2, Et3N, MeOH, H2O; (b) Cbz–Cl, 1 M NaOH; (c) R2NH2, DCC, HOBT, THF; (d) BrCH2COOCH, K2CO3, DMF; (e) NH2OK,
CH3OH.
sume that the potent activity of 8l might result from higher affinity
between the large fused aromatic naphthalene group of 8l and the
residues of HDAC8 protein. While in compound 7l, the Boc group
with large volume probably restricts the large naphthalene group’s
free rotation and sufficient interaction with HDAC8 protein, which
leads to the decreased activity. In spite of this, the tertiary butyl-
oxycarbonyl group (Boc group) located on the secondary amine
atom seemed beneficial to the activity of our compounds. In order
to investigate the relationship between the inhibitory activity and
the groups located on the secondary amine atom, the benzyloxy-
carbonyl group (Cbz group) was chose to protect the secondary
amine atom as the surrogate of Boc group. As a result, other three
compounds were achieved (Scheme 2). Interestingly, under basic
condition, Cbz groups of compounds 13b and 13c were eliminated
to form the hydantoin structures, which could be supported by 1H
NMR, HRMS and many literatures.34–36 The inhibitory activities of
these three compounds are listed in Table 2. The rigid compounds
13b and 13c were almost inactive, while 13a (IC50 = 0.58 0.10
docking study was carried out via Sybyl/FlexX module. The molec-
ular structures of 13a and 7d were generated with Sybyl/Sketch
module and optimized using Powell’s method with the Tripos force
field with convergence criterion set at 0.05 kcal/(Å mol), and as-
signed with Gasteiger–HÜckel method. The residues in a radius
of 8.0 Å around TSA in the co-crystal structure of HDAC8 and TSA
(PDB entry: 1T64) were selected as the active site. Other docking
parameters implied in the program were kept default. The docking
results showed that the carbonyl and hydroxyl oxygens of
hydroxamate could chelate with the zinc ion in HDACs. Both the
methoxyphenyl group of 7d and the phenethyl group of 13a could
form p–p interaction with Tyrosine 100 (Y100). This could explain
why compounds with aromatic R1 groups (7a–7m, 8a–8m) were
more potent than compounds with aliphatic R1 groups (7n–7p,
8n–8p). Because the Cbz group of 13a was longer than the Boc
group of 7d, the Cbz group could insert into pocket X to form stron-
ger hydrophobic interaction. Although the docking results were
consistent with our inference and revealed the reason why com-
pound 13a was more active than 7d, the exact binding mode of
our compounds with HDAC8 should be verified from further
X-ray co-crystal studies. In order to find more potent compounds
having stronger affinity with pocket X and to validate the docking
results, derivatives with diverse N-substituted groups are under
research in our lab.
l
M) exhibited much lower IC50 value compared with its corre-
sponding Boc-protecting compound 7c (IC50 = 2.67 0.15 M). It
was remarkable that compound 13a was even more potent than
7d (IC50 = 1.00 0.16 M), the most potent compound of all our 7
l
l
and 8 series compounds. The result that 13a possessed the most
potent inhibitory activity among all our target compounds further
confirmed our hypothesis that the substituent on the secondary
amine atom was very crucial to the activity and perhaps more
important than the R1 group of these derivatives.
4. Conclusions
In summary, we developed a novel series of 1,2,3,4-tetrahydro-
isoquinoline-3-carboxylic acid derivatives as potent HDACs inhib-
itors. Among them, compounds 13a, 7d, 8l, 7i and 7a exhibited
better anti-HDACs and anti-proliferative activity than Vorinostat.
Therefore, using these compounds as leads, efforts are currently
underway in our laboratoty to search new 1,2,3,4-trtrahydroiso-
quinoline-3-carboxylic acid derivatives as more potent HDACs
inhibitors.
Encouraged by the potent anti-HDACs activities of compounds
7d, 7i, 8l and 13a, we then evaluated their anti-proliferative activ-
ities against HCT116, SKOV3 and HL60 cell lines using MTT assay.
As can be seen in Figure 5, for each compound, the order of its IC50
values against these three tumor cell lines was HCT116 <
SKOV3 < HL60, which was due to the HDACs expression levels in
these three cell lines (HCT116 > SKOV3 > HL60). These results indi-
rectly provided proof that the observed anti-proliferative effect on
the cells was through inhibition of HDACs. The activities against all
the three cell lines of compounds 7d, 7i and 8l were better than
that of SAHA. Unexpectedly, the anti-proliferative activity of 13a
was not as excellent as its anti-HDACs enzyme activity. We sup-
pose that the easiness of benzyl ester hydrolysis in 13a might
result in its decreased cell activity.
5. Experimental section
5.1. Chemistry: general procedures
Unless specified otherwise, all starting materials, reagents and
solvents were commercially available. All reactions except those
in aqueous media were carried out by standard techniques for
the exclusion of moisture. All reactions were monitored by thin-
In order to determine the interaction between our compounds
and HDACs, compounds 13a and 7d were docked into the active
site of HDAC8 (PDB entry: 1T64) using SYBYL 7.3 (Fig. 6). Molecular

Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
1765
Table 1
The structures and inhibitory activities of compounds 7a–7p, 8a–8p against HDAC8
O
O
R1
R1
N
N
H
H
H
H
N
NH.HCl
N
N
HO
O
HO
O
Boc
O
O
7a-p
8a-p
a
Compounds
IC50 of HDAC8 (
lM)
R1
7a
8a
1.29 0.15
8.21 1.68
7b
8b
3.41 0.21
5.10 0.49
7c
8c
2.67 0.15
4.07 0.61
7d
8d
1.00 0.16
5.57 0.44
O
7e
8e
1.65 0.18
3.82 0.29
7f
8f
4.00 0.38
4.13 0.51
7g
8g
1.77 0.25
3.62 0.37
7h
8h
2.56 0.33
3.23 0.25
F
7i
8i
1.17 0.19
3.20 0.41
Cl
7j
8j
3.78 0.66
3.39 0.0.39
7k
8k
1.55 0.24
3.34 0.23
F
Cl
7l
8l
4.25 0.27
1.06 0.14
7m
8m
1.98 0.23
2.21 0.19
7n
8n
3.02 0.25
3.54 0.67
7o
8o
4.42 0.42
5.77 1.09
7p
8p
4.58 0.57
12.17 1.87
O
H
N
OH
Vorinostat
1.48 0.20
N
H
O
a
Data represent mean values of at least three independent experiments and standard deviations are calculated based on the results of these experiments.

1766
Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
Table 2
The structures and inhibitory activities of compounds 13a–13c against HDAC8
a
Compounds
Structures
IC50 of HDAC8 (
l
M)
O
N
H
O
13a
0.58 0.10
H
N
N
N
N
HO
O
O
O
O
O
O
H
N
N
13b
>50
HO
O
O
O
H
N
N
13c
>50
HO
O
O
O
H
N
OH
Vorinostat
1.48 0.20
N
H
O
a
Data represent mean values of at least three independent experiments and standard deviations are calculated based on the results of these experiments.
4.5
>3.78
4
3.5
3
HCT116
SKOV3
HL60
>2.12
2.5
2
>2.0
>2.0
1.66
1.5
1
0.98
0.96
0.75
0.68
0.79
0.49
0.42
0.37
0.5
0
0.17
0.048
7d
7i
8l
13a
SAHA
Compounds
Figure 5. Anti-proliferative activities of the indicated compounds against three
tumor cell lines. The columns represent the mean values of five independent
experiments.
layer chromatography on 0.25 mm silica gel plates (60GF-254) and
visualized with UV light, chloride ferric or iodine vapor. Proton
NMR spectrums were determined on a Brucker DRX spectrometer
(600 MHz), d in parts per million and J in Hertz, using TMS as an
internal standard. Measurements were made in DMSO-d6 solu-
tions. ESI-MS were determined on an API 4000 spectrometer.
HRMS spectrums were conducted by Shandong Analysis and Test
Center. Melting points were determined on an electrothermal
melting point apparatus and were uncorrected.
Figure 6. The docking modes of compounds 13a, 7d with HDAC8. The protein is
represented by molecular surface. Zinc ion is indicated. Y100 stands for the tyrosine
100 residue. The yellow arc indicates the pocket X of the protein. 13a is depicted in
purple and 7d in white. (atom types: polar H, sky-blue; N, dark blue; O, red).
d6) d 3.07 (dd, J = 16.8 Hz, 10.8 Hz, 1H), 3.22 (dd, J = 16.8 Hz,
4.8 Hz, 1H), 4.02 (d, J = 16.2 Hz, 1H), 4.15 (d, J = 16.2 Hz, 1H), 4.32
(dd, J = 4.8 Hz, 10.8 Hz, 1H), 7.73 (s, 1H), 9.68 (s, 1H), 10.00 (br s,
2H), 14.17 (br s, 1H).
5.1.1. (S)-7-Hydroxy-6,8-diiodo-Tic hydrochloride (2)33
A suspension of 3,5-diiodo-L-tyrosine (1, 30.0 g, 69.3 mmol) in
concentrated HCl (250 mL), 1,2-dimethoxyethane (20 mL) and
paraformaldehyde (7.8 g, 260.0 mmol) was stirred vigorously and
slowly heated to 72 °C. After 0.5 h, concentrated HCl (50 mL), 1,2-
dimethoxyethane (10 mL) and paraformaldehyde (5.2 g, 173.3
mmol) were added and stirring was continued for 18 h at 72–
75 °C. The suspension was then cooled in an ice bath and filtered.
The filter cake was washed thoroughly with 1,2-dimethoxyethane
and dried under vacuum to get 19.41 g of compound 2 as a white
powder. Yield: 58%, ESI-MS m/z: 446.2 [M+H]+; 1H NMR (DMSO-
5.1.2. (S)-2-(tert-Butoxycarbonyl)-7-hydroxy-6,8-diiodo-Tic
(3)33
To a solution of compound 2 (4.81 g, 10.0 mmol) in 22 mL of 1 N
NaOH, was added a solution of (Boc)2O (2.40 g, 11.0 mmol) in 5 mL
THF. The solution was kept between pH 9 and 11 by addition of 1 N
NaOH. After stirring the mixture at room temperature for 6 h, the
THF was evaporated in vacuum with the residues being adjusted

Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
1767
to pH 4–5 with 1 N aqueous citric acid. Then the mixture was ex-
tracted with EtOAc (3 Â 15 mL). The extractions were combined,
washed with brine (3 Â 10 mL), dried over Mg2SO4 and evaporated
to give 4.99 g of crude product compound 3 as a light yellow pow-
der. This product was used for the following reaction without fur-
ther purification. 1H NMR (DMSO-d6) d 1.34 + 1.40 (s, 9H, cis/trans),
2.87–3.00 (m, 2H), 4.13–4.41 (m, 2H), 4.61–4.75 (m, 1H), 7.57 (s,
1H), 9.41 (br s, 1H), 12.71 (br s, 1H).
5.1.4.7. (S)-tert -Butyl 3-(4-fluorophenylcarbamoyl)-7-hydroxy-
3,4-dihydroisoquinoline-2(1H)-carboxylate (5h). Compound 5h
was synthesized following the procedure described in Section
5.1.4.
5.1.4.8. (S)-tert-Butyl 3-(3-chlorophenylcarbamoyl)-7-hydroxy-
3,4-dihydroisoquinoline-2(1H)-carboxylate (5i). Compound 5i
was synthesized following the procedure described in Section
5.1.4.
5.1.3. (S)-2-(tert-Butoxycarbonyl)-7-hydroxy-Tic (4)33
To a solution of compound 3 (2.73 g, 5.0 mmol) in 30 mL of
anhydrous methanol, were added triethylamine (1.11 g,
11.0 mmol) and 10% Pd/C (0.23 g). After stirring the mixture under
hydrogen atmosphere for 5 h, the solution was filtered on Celite,
the filtrate concentrated and acidified with 1 N aqueous citric acid
until pH 4–5 then extracted with EtOAc (3 Â 20 mL). The organic
layers were combined, washed with brine (3 Â 20 mL), dried over
Mg2SO4 and evaporated to give 1.10 g of crude product compound
4 as a light yellow powder. This product was used for the following
5.1.4.9. (S)-tert -Butyl 3-(2,4-dimethylphenylcarbamoyl)-7-hyd-
roxy-3,4-dihydroisoquinoline-2(1H)-carboxylate (5j). Compound
5j was synthesized following the procedure described in Section
5.1.4.
5.1.4.10. (S)-tert -Butyl 3-(3-chloro-4-fluorophenylcarbamoyl)-
7-hydroxy-3,4-dihydroisoquinoline-2(1H)-carboxylate
(5k). Compound 5k was synthesized following the procedure de-
scribed in Section 5.1.4.
reaction without further purification. 1H NMR (DMSO-d6)
d
1.39 + 1.45 (s, 9H, cis/trans), 2.92–3.04 (m, 2H), 4.26–4.51 (m,
2H), 4.57–4.82 (m, 1H), 6.52 (s, 1H), 6.57 (d, J = 8.4 Hz, 1H), 6.97
(d, J = 8.4 Hz, 1H), 9.28 (s, 1H), 12.60 (s, 1H).
5.1.4.11. (S)-tert -Butyl 7-hydroxy-3-(naphthalen-1-ylcarba-
moyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (5l). Com-
pound 5l was synthesized following the procedure described in
Section 5.1.4.
5.1.4. (S)-tert-Butyl 7-hydroxy-3-(phenylcarbamoyl)-3,4-
dihydroisoquinoline-2(1H)-carboxylate (5a)
5.1.4.12. (S)-tert -Butyl 3-(biphenyl-4-ylcarbamoyl)-7-hydroxy-
3,4-dihydroisoquinoline-2(1H)-carboxylate (5m). Compound
5m was synthesized following the procedure described in Section
5.1.4.
At 0 °C, to a solution of compound 4 (2.93 g, 10.0 mmol) and
HOBT (1.49 g, 11.0 mmol) in 40 mL of anhydrous THF, was added
a solution of DCC (2.27 g, 11.0 mmol) in 10 mL of anhydrous THF,
and after 30 min, the aniline (1.02 g, 11.0 mmol). After stirring
the mixture at room temperature over night, THF was evaporated
with the residues being taken up in EtOAc (40 mL) and freezed in
refrigerator. Then DCU was filtered off, the filtrate washed with
saturated Na2CO3 (3 Â 10 mL), 1 N HCl (3 Â 10 mL), brine
(3 Â 10 mL) and dried over Mg2SO4. The solvent was evaporated
to give 2.55 g of crude product compound 5a as a yellow powder.
This product was used for the following reaction without further
purification. ESI-MS m/z: 369.5 [M+H]+; 1H NMR (DMSO-d6) d
1.30 + 1.45 (s, 9H, cis/trans), 2.85–3.11 (m, 2H), 4.27–4.52 (m,
2H), 4.55–4.74 (m, 1H), 6.57–6.64 (m, 2H), 6.96–7.04 (m, 2H),
7.25–7.30 (m, 2H), 7.49–7.56 (m, 2H), 9.29 (s, 1H), 9.96 (s, 1H).
5.1.4.13. (S)-tert
-Butyl 7-hydroxy-3-(pentylcarbamoyl)-3,4-
dihydroisoquinoline-2(1H)-carboxylate (5n). Compound 5n
was synthesized following the procedure described in Section
5.1.4.
5.1.4.14. (S)-tert-Butyl 3-(hexylcarbamoyl)-7-hydroxy-3,4-dihy-
droisoquinoline-2(1H)-carboxylate (5o). Compound 5o was
synthesized following the procedure described in Section 5.1.4.
5.1.4.15. (S)-tert-Butyl 3-(tert -butylcarbamoyl)-7-hydroxy-3,4-
dihydroisoquinoline-2(1H)-carboxylate (5p). Compound 5p
was synthesized following the procedure described in Section
5.1.4.
5.1.4.1. (S)-tert-Butyl 3-(benzylcarbamoyl)-7-hydroxy-3, 4-dihy-
droisoquinoline-2(1H)-carboxylate (5b). Compound 5b was
synthesized following the procedure described above.
5.1.5. (S)-tert-Butyl 7-(2-methoxy-2-oxoethoxy)-3-
(phenylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6a)
5.1.4.2. (S)-tert -Butyl 7-hydroxy-3-(phenethylcarbamoyl)-3,4-
dihydroisoquinoline-2(1H)-carboxylate (5c). Compound 5c was
synthesized following the procedure described in Section 5.1.4.
A mixture of 5a (2.55 g, 6.93 mmol), K2CO3 (1.91 g, 13.86 mmol)
and methyl bromoacetate (2.12 g, 13.86 mmol) in 40 mL of anhy-
drous DMF was stirred at room temperature for 3 h. The mixture
was poured to 300 mL of H2O and extracted with EtOAc
(3 Â 40 mL). The organic layers were combined, washed with brine
(3 Â 20 mL), dried over Mg2SO4 and evaporated under vacuum. The
residues were purified by flash column chromatography (petro-
leum/EtOAc 3:1) to give 1.16 g of desired compound 6a as colorless
oil. Yield: 38%, ESI-MS m/z: 441.6 [M+H]+; 1H NMR (DMSO-d6) d
1.33 + 1.47 (s, 9H, cis/trans), 2.93–3.16 (m, 2H), 3.70 (s, 3H),
4.34–4.58 (m, 2H), 4.62–4.88 (m, 1H), 4.78 (s, 2H), 6.72–6.90 (m,
2H), 7.02–7.16 (m, 2H), 7.26–7.31 (m, 2H), 7.49–7.56 (m, 2H),
10.00 (s, 1H).
5.1.4.3. (S)-tert -Butyl 7-hydroxy-3-(4-methoxyphenylcarba-
moyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (5d). Com-
pound 5d was synthesized following the procedure described in
Section 5.1.4.
5.1.4.4. (S)-tert-Butyl 7-hydroxy-3-(p-tolylcarbamoyl)-3,4-dihy-
droisoquinoline-2(1H)-carboxylate (5e). Compound 5e was syn-
thesized following the procedure described in Section 5.1.4.
5.1.4.5. (S)-tert-Butyl 7-hydroxy-3-(o-tolylcarbamoyl)-3,4-dihy-
droisoquinoline-2(1H)-carboxylate (5f). Compound 5f was syn-
thesized following the procedure described in Section 5.1.4.
Compounds 6b–p were synthesized following the procedure
described above.
5.1.4.6. (S)-tert-Butyl 7-hydroxy-3-(m-tolylcarbamoyl)-3,4-dihy-
droisoquinoline-2(1H)-carboxylate (5g). Compound 5g was
synthesized following the procedure described in Section 5.1.4.
5.1.5.1. (S)-tert -Butyl 3-(benzylcarbamoyl)-7-(2-methoxy-2-
oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6b). Yield: 38%.

1768
Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
5.1.5.2. (S)-tert -Butyl 7-(2-methoxy-2-oxoethoxy)-3-(pheneth-
ylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6c). Yield: 40%.
the residues being recrystallized with EtOAc/EtOH to give 0.23 g
of title compound 7a as a white powder. Yield: 26%, mp: 179–
181 °C. IR (KBr) 3329, 3260, 2976, 2924, 2855, 1699, 1663,
1601 cmÀ1 1H NMR (DMSO-d6) d 1.30 + 1.46 (s, 9H, cis/trans),
;
5.1.5.3. (S)-tert -Butyl 7-(2-methoxy-2-oxoethoxy)-3-(4-meth-
oxyphenylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxyl-
ate (6d). Yield: 36%.
2.89–3.19 (m, 2H), 4.34–4.65 (m, 2H), 4.44 (s, 2H), 4.57–4.80 (m,
1H), 6.70–6.88 (m, 2H), 7.01–7.17 (m, 2H), 7.24–7.32 (m, 2H),
7.49–7.56 (m, 2H), 8.97 (s, 1H), 10.01 (s, 1H), 10.81 (s, 1H); HRMS
(AP-ESI) m/z calcd for C23H28N3O6 [M+H]+ 442.1978, found:
442.1980.
5.1.5.4. (S)-tert-Butyl 7-(2-methoxy-2-oxoethoxy)-3-(p-tolylcar-
bamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6e). Yield: 33%.
Compounds 7b–p were synthesized following the procedure
described above.
5.1.5.5. (S)-tert-Butyl 7-(2-methoxy-2-oxoethoxy)-3-(o-tolylcar-
bamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6f). Yield: 40%.
5.1.6.1. (S)-tert -Butyl 3-(benzylcarbamoyl)-7-(2-(hydroxyami-
no)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(7b). Yield: 22%, mp: 153–156 °C. IR (KBr) 3299, 2976, 2928,
2868, 1670 cmÀ1 1H NMR (DMSO-d6) d 1.33 + 1.47 (s, 9H, cis/
;
5.1.5.6. (S)-tert -Butyl 7-(2-methoxy-2-oxoethoxy)-3-(m-tolylc-
trans), 2.95–3.11 (m, 2H), 4.03–4.20 (m, 2H), 4.36–4.53 (m, 2H),
4.40 (s, 2H), 4.56–4.76 (m, 1H), 6.80–6.89 (m, 2H), 6.99–7.11 (m,
2H), 7.15–7.20 (m, 2H), 7.23–7.34 (m, 2H), 8.33 (s, 1H), 9.02 (s,
1H), 10.76 (s, 1H); HRMS (AP-ESI) m/z calcd for C24H30N3O6
[M+H]+ 456.2135, found: 456.2137.
arbamoyl)-3,
4-dihydroisoquinoline-2(1H)-carboxylate
(6g). Yield: 39%.
5.1.5.7. (S)-tert -Butyl 3-(4-fluorophenylcarbamoyl)-7-(2-meth-
oxy-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6h). Yield: 34%.
5.1.6.2. (S)-tert -Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-
(phenethylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carbox-
ylate (7c). Yield: 29%, mp: 155–157 °C. IR (KBr) 3315, 3200, 2978,
5.1.5.8. (S)-tert-Butyl 3-(3-chlorophenylcarbamoyl)-7-(2-meth-
oxy-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6i). Yield: 31%.
2931, 2864, 1664 cmÀ1 1H NMR (DMSO-d6) d 1.35 + 1.45 (s, 9H,
;
cis/trans), 2.54–2.64 (m, 2H), 2.83–2.95 (m, 2H), 3.13–3.27 (m,
2H), 4.28–4.57 (m, 2H), 4.44 (s, 2H), 4.52–4.76 (m, 1H), 6.76–
6.81 (m, 2H), 7.02–7.19 (m, 3H), 7.24–7.28 (m, 2H), 7.92 (s, 1H),
8.97 (s, 1H), 10.81 (s, 1H); HRMS (AP-ESI) m/z calcd for C25H31N3O6-
Na [M+Na]+ 492.2111, found: 492.2129.
5.1.5.9. (S)-tert-Butyl 3-(2,4-dimethylphenylcarbamoyl)-7-(2-
methoxy-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carbox-
ylate (6j). Yield: 48%.
5.1.5.10. (S)-tert-butyl 3-(3-chloro-4-fluorophenylcarbamoyl)-
5.1.6.3. (S)-tert -Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-(4-
methoxyphenylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-car-
boxylate (7d). Yield: 25%, mp: 167–169 °C. IR (KBr) 3268, 3150,
7-(2-methoxy-2-oxoethoxy)-3,
4-dihydroisoquinoline-2(1H)-
carboxylate (6k). Yield: 37%.
2978, 2933, 2915, 2854, 2836, 1702, 1671, 1613 cmÀ1 1H NMR
;
5.1.5.11. (S)-tert-Butyl 7-(2-methoxy-2-oxoethoxy)-3-(naphtha-
len-1-ylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6l). Yield: 29%.
(DMSO-d6) d 1.30 + 1.46 (s, 9H, cis/trans), 2.89–3.17 (m, 2H), 3.72
(s, 3H), 4.32–4.65 (m, 2H), 4.44 (s, 2H), 4.60–4.80 (m, 1H), 6.75–
6.90 (m, 4H), 7.06–7.24 (m, 1H), 7.38–7.47 (m, 2H), 8.98 (s, 1H),
9.86 (s, 1H), 10.82 (s, 1H); HRMS (AP-ESI) m/z calcd for
C24H30N3O7 [M+H]+ 472.2084, found: 472.2090.
5.1.5.12. (S)-tert-Butyl 3-(biphenyl-4-ylcarbamoyl)-7-(2-meth-
oxy-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6m). Yield: 31%.
5.1.6.4. (S)-tert-Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-(p -
tolylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(7e). Yield: 27%, mp: 164–166 °C. IR (KBr) 3267, 3138, 2980, 2916,
5.1.5.13. (S)-tert -Butyl 7-(2-methoxy-2-oxoethoxy)-3-(pentylc-
arbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6n). Yield: 39%.
2859, 1700, 1664, 1610 cmÀ1 1H NMR (DMSO-d6) d 1.30 + 1.45 (s,
;
9H, cis/trans), 2.23 (s, 3H), 2.87–3.17 (m, 2H), 4.32–4.65 (m, 2H),
4.44 (s, 2H), 4.56–4.79 (m, 1H), 6.74–6.88 (m, 2H), 7.05–7.17 (m,
3H), 7.35–7.46 (m, 2H), 8.97 (s, 1H), 9.91 (s, 1H), 10.81 (s, 1H);
HRMS (AP-ESI) m/z calcd for C24H30N3O6 [M+H]+ 456.2135, found:
456.2138.
5.1.5.14. (S)-tert-Butyl 3-(hexylcarbamoyl)-7-(2-methoxy-2-oxo-
ethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate (6o). Yield:
41%.
5.1.5.15. (S)-tert -Butyl 3-(tert -butylcarbamoyl)-7-(2-methoxy-
2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(6p). Yield: 38%.
5.1.6.5. (S)-tert-Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-(o -
tolylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(7f). Yield: 26%, mp: 166–168 °C. IR (KBr) 3252, 2975, 2911, 2859,
1694, 1660, 1616 cmÀ1 1H NMR (DMSO-d6) d 1.39 + 1.47 (s, 9H,
;
5.1.6. (S)-tert-Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-
(phenylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(7a)
To a solution of compound 6a (0.88 g, 2.0 mmol) in 10 mL of
anhydrous methanol, was added a solution of NH2OK (0.14 g,
6 mmol) in 3.5 mL of anhydrous methanol. The mixture was stirred
for 0.5 h and the solvent was evaporated under vacuum. The resi-
dues were acidified with 2 N HCl until pH 3–4 then extracted with
EtOAc (3 Â 10 mL). The organic layers were combined, washed
with brine (3 Â 10 mL), dried over Mg2SO4 and evaporated with
cis/trans), 1.96 + 2.05 (s, 3H, cis/trans), 3.01–3.20 (m, 2H), 4.43 (s,
2H), 4.45–4.55 (m, 2H), 4.60–4.86 (m, 1H), 6.78–6.87 (m, 2H),
7.05–7.19 (m, 5H), 8.97 (s, 1H), 9.28 (s, 1H), 10.81 (s, 1H); HRMS
(AP-ESI) m/z calcd for C24H30N3O6 [M+H]+ 456.2135, found:
456.2131.
5.1.6.6. (S)-tert-Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-(m-
tolylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(7g). Yield: 33%, mp: 183–184 °C. IR (KBr) 3332, 3150, 2976, 2928,
2865, 1696, 1664, 1612 cmÀ1 1H NMR (DMSO-d6) d 1.30 + 1.46 (s,
;

Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
1769
9H, cis/trans), 2.30 (s, 3H), 2.91–3.15 (m, 2H), 4.35–4.63 (m, 2H),
4.41 (s, 2H), 4.56–4.82 (m, 1H), 6.75–6.88 (m, 3H), 7.09–7.18 (m,
2H), 7.25–7.29 (m, 2H), 8.98 (s, 1H), 9.93 (s, 1H), 10.82 (s, 1H);
HRMS (AP-ESI) m/z calcd for C24H29N3O6Na [M+Na]+ 478.1954,
found: 478.1971.
(s, 1H), 10.12 (s, 1H), 10.82 (s, 1H); HRMS (AP-ESI) m/z calcd for
C29H32N3O6 [M+H]+ 518.2291, found: 518.2294.
5.1.6.13. (S)-tert -Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-
(pentylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(7n). Yield: 25%, mp: 159–161 °C. IR (KBr) 3363, 3235, 2962,
2931, 2861, 1693, 1669, 1647 cmÀ1 1H NMR (DMSO-d6) d 0.79–
;
5.1.6.7. (S)-tert -Butyl 3-(4-fluorophenylcarbamoyl)-7-(2-(hyd-
roxyamino)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-car-
boxylate (7h). Yield: 28%, mp: 167–169 °C. IR (KBr) 3261, 3164,
0.86 (m, 3H), 1.08–1.14 (m, 2H), 1.17–1.23 (m, 2H), 1.30–1.36
(m, 2H), 1.39 + 1.46 (s, 9H, cis/trans), 2.95–3.05 (m, 4H), 4.31–
4.65 (m, 2H), 4.48 (s, 2H), 4.60–4.87 (m, 1H), 6.74 (s, 1H), 6.81
(d, J = 8.4 Hz, 1H), 7.04 (d, J = 8.4 Hz, 1H), 7.90 (s, 1H), 9.07 (s,
1H), 10.69 (s, 1H); HRMS (AP-ESI) m/z calcd for C22H34N3O6
[M+H]+ 435.2369, found: 435.2364.
2979, 2933, 2917, 2856, 1700, 1663, 1619 cmÀ1
;
1H NMR
(DMSO-d6) d 1.30 + 1.46 (s, 9H, cis/trans), 2.91–3.16 (m, 2H),
4.35–4.63 (m, 2H), 4.44 (s, 2H), 4.50–4.77 (m, 1H), 6.76–6.88 (m,
2H), 7.10–7.20 (m, 3H), 7.51–7.58 (m, 2H), 9.07 (s, 1H), 10.64 (s,
1H), 10.76 (s, 1H); HRMS (AP-ESI) m/z calcd for C23H27FN3O6
[M+H]+ 482.1703, found: 482.1733.
5.1.6.14. (S)-tert -Butyl 3-(hexylcarbamoyl)-7-(2-(hydroxyami-
no)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(7o). Yield: 26%, mp: 105–107 °C. IR (KBr) 3311, 3180, 2957,
5.1.6.8. (S)-tert-Butyl 3-(3-chlorophenylcarbamoyl)-7-(2-(hyd-
roxyamino)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-car-
boxylate (7i). Yield: 23%, mp: 182–184 °C. IR (KBr) 3314, 3188,
2931, 2858, 1690, 1661, 1639 cmÀ1 1H NMR (DMSO-d6) d 0.81–
;
0.85 (m, 3H), 1.07–1.23 (m, 8H), 1.36 + 1.45 (s, 9H, cis/trans),
2.86–3.01 (m, 4H), 4.31–4.56 (m, 2H), 4.14 (s, 2H), 4.44–4.75 (m,
1H), 6.74 (s, 1H), 6.81 (d, J = 8.4 Hz, 1H), 7.05 (d, J = 8.4 Hz, 1H),
7.75 (s, 1H), 8.97 (s, 1H), 10.81 (s, 1H); HRMS (AP-ESI) m/z calcd
for C23H36N3O6 [M+H]+ 450.2604, found: 450.2610.
3114, 2977, 2932, 2918, 1699, 1664, 1620 cmÀ1
;
1H NMR
(DMSO-d6) d 1.30 + 1.46 (s, 9H, cis/trans), 2.91–3.17 (m, 2H),
4.37–4.57 (m, 2H), 4.44 (s, 2H), 4.60–4.78 (m, 1H), 6.75–6.89 (m,
2H), 7.08–7.17 (m, 2H), 7.30–7.35 (m, 1H), 7.37–7.45 (m, 1H),
7.71–7.76 (m, 1H), 8.97 (s, 1H), 10.23 (s, 1H), 10.82 (s, 1H); HRMS
(AP-ESI) m/z calcd for C23H26ClN3O6Na [M+Na]+ 498.1408, found:
498.1431.
5.1.6.15. (S)-tert-Butyl 3-(tert-butylcarbamoyl)-7-(2-(hydroxya-
mino)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxyl-
ate (7p). Yield: 28%, mp: 176–178 °C. IR (KBr) 3307, 3239, 2978,
; d
2920, 2854, 2697, 1675, 1640 cmÀ1 1H NMR (DMSO-d6)
5.1.6.9. (S)-tert-Butyl 3-(2, 4-dimethylphenylcarbamoyl)-7-(2-
(hydroxyamino)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-
carboxylate (7j). Yield: 28%, mp: 154–156 °C. IR (KBr) 3247,
1.14 + 1.21 (s, 9H, cis/trans), 1.36 + 1.44 (s, 9H, cis/trans), 2.74–
3.03 (m, 2H), 4.25–4.57 (m, 2H), 4.45 (s, 2H), 4.19–4.76 (m, 1H),
6.77 (s, 1H), 6.80 (d, J = 8.4 Hz, 1H), 7.08 (d, J = 8.4 Hz, 1H), 7.31
(s, 1H), 8.96 (s, 1H), 10.80 (s, 1H); HRMS (AP-ESI) m/z calcd for
C21H31N3O6Na [M+Na]+ 444.2111, found: 444.2119.
3150, 2977, 2919, 2861, 1696, 1666, 1618 cmÀ1
;
1H NMR
(DMSO-d6) d 1.39 + 1.47 (s, 9H, cis/trans), 1.91 + 2.00 (s, 3H, cis/
trans), 2.21 (s, 3H), 3.00–3.18 (m, 2H), 4.41–4.54 (m, 2H), 4.44 (s,
2H), 4.57–4.84 (m, 1H), 6.77–6.86 (m, 2H), 6.89–7.04 (m, 3H),
7.11–7.16 (m, 1H), 8.97 (s, 1H), 9.21 (s, 1H), 10.82 (s, 1H); HRMS
(AP-ESI) m/z calcd for C25H31N3O6Na [M+Na]+ 492.2111, found:
492.2118.
5.1.7. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-phenyl-1,2,3,4-
tetrahydroisoquinoline-3-carboxamide hydrochloride (8a)
To a solution of compound 7a (0.15 g, 0.34 mmol) in 5 mL of dry
EtOAc, was added a solution of EtOAc (10 mL) saturated by dry HCl
gas. The reaction solution was stirred at room temperature for 8 h
when the precipitation appeared. The suspension was filtered with
the filter being washed with ether to give 0.12 g of desired com-
pound 8a as a white powder. Yield: 91%, mp: 187–189 °C. IR
5.1.6.10. (S)-tert -Butyl 3-(3-chloro-4-fluorophenylcarbamoyl)-
7-(2-(hydroxyamino)-2-oxoethoxy)-3,4-dihydroisoquinoline-
2(1H)-carboxylate (7k). Yield: 40%, mp: 163–165 °C. IR (KBr)
3326, 3145, 2979, 2930, 2852, 1694, 1675, 1628 cmÀ1 1H NMR
;
(KBr) 3192, 3134, 3056, 2981, 2808, 2606, 2489, 1677, 1606 cm
(DMSO-d6) d 1.30 + 1.46 (s, 9H, cis/trans), 2.91–3.17 (m, 2H),
4.37–4.62 (m, 2H), 4.44 (s, 2H), 4.54–4.77 (m, 1H), 6.76–6.89 (m,
2H), 7.10–7.16 (m, 1H), 7.32–7.46 (m, 2H), 7.82–7.87 (m, 1H),
8.97 (s, 1H), 10.25 (s, 1H), 10.82 (s, 1H); HRMS (AP-ESI) m/z calcd
for C23H25ClFN3O6Na [M+Na]+ 516.1314, found: 516.1310.
À1
;
1H NMR (DMSO-d6) d 2.98–3.37 (m, 2H), 4.28–4.30 (m, 2H),
4.32–4.37 (m, 1H), 4.45 (s, 2H), 6.89–6.92 (m, 2H), 7.12–7.23 (m,
2H), 7.36–7.41 (m, 2H), 7.65–7.67 (m, 2H), 9.00 (br s, 1H), 9.60
(br s, 1H), 9.81 (br s, 1H), 10.77 (s, 1H), 10.87 (s, 1H); HRMS (AP-
ESI) m/z calcd for C18H20N3O4 [M+H]+ 342.1454, found: 342.1459.
Compounds 8b–p were synthesized following the procedure
described above.
5.1.6.11. (S)-tert -Butyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-
(naphthalen-1-ylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-
carboxylate (7l). Yield: 35%, mp: 142–144 °C. IR (KBr) 3263,
5.1.7.1. (S)-N-Benzyl-7-(2-(hydroxyamino)-2-oxoethoxy)-1,2,3,
4-tetrahydroisoquinoline-3-carboxamide hydrochloride
(8b). Yield: 94%, mp: 175–177 °C. IR (KBr) 3216, 3063, 3033,
2976, 2930, 2868, 1746, 1677, 1627 cmÀ1 1H NMR (DMSO-d6) d
;
1.41 + 1.49 (s, 9H, cis/trans), 3.11–3.28 (m, 2H), 4.41–4.64 (m,
2H), 4.45 (s, 2H), 4.67–4.97 (m, 1H), 6.66–6.91 (m, 2H), 7.18–
7.23 (m, 1H), 7.38–7.53 (m, 3H), 7.59–7.52 (m, 3H), 8.05–8.13
(m, 1H), 8.98 (s, 1H), 9.91 (s, 1H), 10.82 (s, 1H); HRMS (AP-ESI)
m/z calcd for C27H30N3O6 [M+H]+ 492.2135, found: 492.2131.
2926, 2851, 2812, 2604, 2485, 1672, 1617 cmÀ1
;
1H NMR
(DMSO-d6) d 2.91–3.28 (m, 2H), 4.15–4.27 (m, 2H), 4.31–4.41 (m,
3H), 4.45 (s, 2H), 6.87–6.90 (m, 2H), 7.17–7.20 (m, 1H), 7.28–7.39
(m, 5H), 9.90 (br s, 1H), 9.22 (s, 1H), 9.53 (br s, 1H), 9.74 (br s,
1H), 10.87 (s, 1H); HRMS (AP-ESI) m/z calcd for C19H22N3O4
[M+H]+ 356.1610, found: 356.1680.
5.1.6.12. (S)-tert-Butyl 3-(biphenyl-4-ylcarbamoyl)-7-(2-(hydrox-
yamino)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxyl-
ate (7m). Yield: 29%, mp: 144–146 °C. IR (KBr) 3428, 3255, 2976,
5.1.7.2. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-phenethyl-1,2,
3,4-tetrahydroisoquinoline-3-carboxamide hydrochloride
(8c). Yield: 92%, mp: 167–169 °C. IR (KBr) 3222, 3062, 3028,
; d
2923, 2860, 1701, 1662, 1594 cmÀ1 1H NMR (DMSO-d6)
1.32 + 1.47 (s, 9H, cis/trans), 2.93–3.19 (m, 2H), 4.37–4.65 (m, 2H),
4.44 (s, 2H), 4.55–4.83 (m, 1H), 6.76–6.90 (m, 2H), 7.11–7.18 (m,
1H), 7.31–7.33 (m, 1H), 7.42–7.45 (m, 2H), 7.60–7.67 (m, 6H), 8.97
2933, 2802, 2604, 2484, 1669, 1615 cmÀ1 1H NMR (DMSO-d6) d
;
2.75–2.80 (m, 2H), 2.79–3.16 (m, 2H), 3.38–3.49 (m, 2H), 4.06–

1770
Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
4.30 (m, 3H), 4.44 (s, 2H), 6.84 (s, 1H), 6.88 (d, J = 8.4 Hz, 1H), 7.14 (d,
J = 8.4 Hz, 1H), 7.22–7.26 (m, 3H), 7.31–7.34 (m, 2H), 8.70 (s, 1H),
8.99 (br s, 1H), 9.42 (br s, 1H), 9.58 (br s, 1H), 10.86 (s, 1H); HRMS
(AP-ESI) m/z calcd for C21H26N3O3 [M+H]+ 370.1767, found:
370.1778.
5.1.7.9. (S)-N-(2,4-Dimethylphenyl)-7-(2-(hydroxyamino)-2-oxo-
ethoxy)-1,2,3,4-tetrahydroisoquinoline-3-carboxamide hydro-
chloride (8j). Yield: 88%, mp: 163–165 °C. IR (KBr) 3443, 3190,
3022, 2975, 2921, 2601, 2484, 1673, 1616 cmÀ1 1H NMR (DMSO-
d6) d 2.19 (s, 3H), 2.27 (s, 3H), 2.99–3.42 (m, 2H), 4.22–4.25 (m, 2H),
4.30–4.33 (m, 1H), 4.45 (s, 2H), 6.85 (s, 1H), 6.88–6.89 (m, 1H),
7.02–7.03 (m, 1H), 7.08 (s, 1H), 7.20–7.33 (m, 2H), 8.99 (s, 1H), 9.60
(br s, 1H), 9.71 (br s, 1H), 9.98 (s, 1H), 10.87 (s, 1H); HRMS (AP-ESI)
m/z calcd for C20H24N3O4 [M+H]+ 370.1767, found: 370.1770.
;
5.1.7.3. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-(4-methoxy-
phenyl)-1,2,3,4-tetrahydroisoquinoline-3-carboxamide hydro-
chloride (8d). Yield: 94%, mp: 176–178 °C. IR (KBr) 3195, 3055,
3006, 2934, 2836, 2606, 2487, 1675, 1614 cmÀ1
;
1H NMR
(DMSO-d6) d 2.99–3.42 (m, 2H), 3.74 (s, 3H), 4.28–4.30 (m, 2H),
4.36–3.38 (m, 1H), 4.46 (s, 2H), 6.89–6.91 (m, 2H), 6.94–6.96 (m,
2H), 7.20–7.21 (m, 1H), 7.57–7.59 (m, 2H), 8.99 (br s, 1H), 9.60
(br s, 1H), 9.83 (br s, 1H), 10.79 (s, 1H), 10.89 (s, 1H); HRMS (AP-
ESI) m/z calcd for C19H22N3O5 [M+H]+ 372.1559, found: 372.1563.
5.1.7.10. (S)-N-(3-Chloro-4-fluorophenyl)-7-(2-(hydroxyamino)-
2-oxoethoxy)-1,2,3,4-tetrahydroisoquinoline-3-carboxamide
hydrochloride (8k). Yield: 90%, mp: 173–175 °C. IR (KBr) 3196,
3135, 3047, 2980, 2935, 2607, 2486, 1681, 1615 cmÀ1 1H NMR
;
(DMSO-d6) d 3.00–3.47 (m, 2H), 4.27–4.40 (m, 3H), 4.46 (s, 2H),
6.85 (s, 1H), 6.88–6.89 (m, 1H), 7.17–7.20 (m, 1H), 7.46–7.47 (m,
1H), 7.64–7.66 (m, 1H), 7.99–8.00 (m, 1H), 9.00 (br s, 1H), 9.67 (br
s, 1H), 9.91 (br s, 1H), 10.90 (s, 1H), 11.50 (s, 1H); HRMS (AP-ESI)
m/z calcd for C18H18ClFN3O4 [M+H]+ 394.0970, found: 394.0987.
5.1.7.4. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-p-tolyl-1,2,3,
4-tetrahydroisoquinoline-3-carboxamide hydrochloride (8e).
Yield: 90%, mp: 182–184 °C. IR (KBr) 3188, 3123, 3038, 2922,
2806, 2604, 2483, 1677, 1612 cmÀ1 1H NMR (DMSO-d6) d 2.78
;
5.1.7.11. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-(naphthalen-
-1-yl)-1,2,3,4-tetrahydroisoquinoline-3-carboxamide hydrochlo-
ride (8l). Yield: 85%, mp: 230–232 °C. IR (KBr) 3422, 3176, 3050,
(s, 3H), 2.99–3.44 (m, 2H), 4.28–4.38 (m, 3H), 4.46 (s, 2H), 6.85
(s, 1H), 6.89 (d, J = 8.4 Hz, 1H), 7.15 (d, J = 8.4 Hz, 1H), 7.18 (d,
J = 7.8 Hz, 2H), 7.56 (d, J = 7.8 Hz, 2H), 9.00 (br s, 1H), 9.62 (br s,
1H), 9.89 (br s, 1H), 10.82 (s, 1H), 10.90 (s, 1H); HRMS (AP-ESI)
m/z calcd for C19H22N3O4 [M+H]+ 356.1610, found: 356.1661.
3013, 2981, 2611, 2489, 1738, 1672 cmÀ1 1H NMR (DMSO-d6) d
;
3.15–3.59 (m, 2H), 4.15–4.35 (m, 3H), 4.47 (s, 2H), 6.91 (s, 1H),
6.94 (d, J = 8.4 Hz, 1H), 7.29 (d, J = 8.4 Hz, 1H), 7.56–7.62 (m, 3H),
7.68–7.71 (m, 1H), 7.87–7.89 (m, 1H), 7.99–8.01 (m, 1H), 8.14–
8.16 (m, 1H), 9.01 (br s, 1H), 9.66 (br s, 1H), 9.80 (br s, 1H), 10.78
(s, 1H), 10.89 (s, 1H); HRMS (AP-ESI) m/z calcd for C22H22N3O4
[M+H]+ 392.1610, found: 392.1629.
5.1.7.5. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-o-tolyl-1,2,3,
4-tetrahydroisoquinoline-3-carboxamide hydrochloride (8f).
Yield: 96%, mp: 177–179 °C. IR (KBr) 3187, 3025, 2918, 2800,
2739, 2606, 2489, 1675, 1615 cmÀ1 1H NMR (DMSO-d6) d 2.25
;
(s, 3H), 3.05–3.45 (m, 2H), 4.28–4.40 (m, 3H), 4.46 (s, 2H), 6.83–
6.92 (m, 2H), 7.17–7.37 (m, 5H), 9.00 (br s, 1H), 9.59 (br s, 1H),
9.78 (br s, 1H), 10.20 (s, 1H), 10.88 (s, 1H); HRMS (AP-ESI) m/z calcd
for C19H22N3O4 [M+H]+ 356.1610, found: 356.1652.
5.1.7.12. (S)-N-(Biphenyl-4-yl)-7-(2-(hydroxyamino)-2-oxoeth-
oxy)-1,2,3,4-tetrahydroisoquinoline-3-carboxamide hydrochlo-
ride (8m). Yield: 87%, mp: 178–180 °C. IR (KBr) 3240, 3179, 3104,
3032, 2977, 2601, 2486, 1677, 1607 cmÀ1 1H NMR (DMSO-d6) d
;
3.02–3.47 (m, 2H), 4.32–4.42 (m, 3H), 4.46 (s, 2H), 6.90 (s, 1H),
6.91 (d, J = 8.4 Hz, 1H), 7.22 (d, J = 8.4 Hz, 1H), 7.34–7.37 (m, 1H),
7.46 (d, J = 7.8 Hz, 2H), 7.67 (d, J = 7.8 Hz, 2H), 7.71 (d, J = 8.4 Hz,
2H), 7.78 (d, J = 8.4 Hz, 2H), 9.00 (br s, 1H), 9.64 (br s, 1H), 9.82
(br s, 1H), 10.89 (s, 1H), 11.04 (s, 1H); HRMS (AP-ESI) m/z calcd
for C24H24N3O4 [M+H]+ 418.1767, found: 418.1759.
5.1.7.6. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-m-tolyl-1,2,3,
4-tetrahydroisoquinoline-3-carboxamide hydrochloride (8g).
Yield: 89%, mp: 173–175 °C. IR (KBr) 3235, 3150, 3028, 2920,
2723, 2598, 2477, 1679, 1617 cmÀ1 1H NMR (DMSO-d6) d 2.31
;
(s, 3H), 2.99–3.42 (m, 2H), 4.29–4.32 (m, 2H), 4.37–4.39 (m, 1H),
4.45 (s, 2H), 6.87 (s, 1H), 6.91 (d, J = 8.4 Hz, 1H), 6.96–6.97 (m,
1H), 7.21 (d, J = 8.4 Hz, 1H), 7.25–7.27 (m, 1H), 7.45–7.46 (m,
2H), 8.99 (s, 1H), 9.60 (br s, 1H), 9.71 (br s, 1H), 10.71 (s, 1H),
10.87 (s, 1H); HRMS (AP-ESI) m/z calcd for C19H22N3O4 [M+H]+
356.1610, found: 356.1668
5.1.7.13. (S)-7-(2-(Hydroxyamino)-2-oxoethoxy)-N-pentyl-1,2,3,
4-tetrahydroisoquinoline-3-carboxamide hydrochloride (8n).
Yield: 91%, mp: 168–170 °C. IR (KBr) 3234, 3076, 2955, 2930,
2861, 2598, 2482, 1670, 1610 cmÀ1 1H NMR (DMSO-d6) d 0.88
;
(t, J = 6.6 Hz, 3H), 1.24–1.30 (m, 4H), 1.47 (q, J = 6.6 Hz, 2H),
2.87–3.25 (m, 2H), 3.16 (t, J = 6.0 Hz, 2H), 4.09–4.32 (m, 3H), 4.44
(s, 2H), 6.81 (s, 1H), 6.77 (d, J = 8.4 Hz, 1H), 7.13 (d, J = 8.4 Hz,
1H), 8.69 (s, 1H), 9.00 (br s, 1H), 9.44 (br s, 1H), 9.68 (br s, 1H),
10.88 (s, 1H); HRMS (AP-ESI) m/z calcd for C17H26N3O4 [M+H]+
336.1923, found: 336.1932.
5.1.7.7. (S)-N-(4-Fluorophenyl)-7-(2-(hydroxyamino)-2-oxoeth-
oxy)-1,2,3,4-tetrahydroisoquinoline-3-carboxamide hydrochlo-
ride (8h). Yield: 91%, mp: 171–173 °C. IR (KBr) 3209, 3152, 3059,
2809, 2605, 2487, 1679, 1620 cmÀ1 1H NMR (DMSO-d6) d 3.00–
;
3.43 (m, 2H), 4.29–4.32 (m, 2H), 4.36–4.39 (m, 1H), 4.45 (s, 2H),
6.89 (s, 1H), 6.90–6.92 (m, 1H), 7.20–7.29 (m, 3H), 7.68–7.70 (m,
2H), 9.04 (s, 1H), 9.63 (br s, 1H), 9.74 (br s, 1H), 10.83 (s, 1H),
10.97 (s, 1H); HRMS (AP-ESI) m/z calcd for C18H19FN3O4 [M+H]+
360.1360, found: 360.1377.
5.1.7.14. (S)-N-hexyl-7-(2-(hydroxyamino)-2-oxoethoxy)-1,2,3,
4-tetrahydroisoquinoline-3-carboxamide hydrochloride (8o).
Yield: 95%, mp: 173–175 °C. IR (KBr) 3230, 3074, 2955, 2930,
2857, 2601, 2484, 1669, 1618 cmÀ1 1H NMR (DMSO-d6) d 0.88
;
(t, J = 6.6 Hz, 3H), 1.29–1.43 (m, 6H), 1.46 (q, J = 6.6 Hz, 2H),
2.87–3.25 (m, 2H), 3.13–3.19 (m, 2H), 4.07–4.32 (m, 3H), 4.44 (s,
2H), 6.86 (s, 1H), 6.88 (d, J = 8.4 Hz, 1H), 7.17 (d, J = 8.4 Hz, 1H),
8.66 (s, 1H), 9.00 (br s, 1H), 9.45 (br s, 1H), 9.67 (br s, 1H), 10.89
(s, 1H); HRMS (AP-ESI) m/z calcd for C18H28N3O4 [M+H]+
350.2080, found: 350.2075.
5.1.7.8. (S)-N-(3-chlorophenyl)-7-(2-(hydroxyamino)-2-oxoeth-
oxy)-1,2,3,4-tetrahydroisoquinoline-3-carboxamide hydrochlo-
ride (8i). Yield: 92%, mp: 193–195 °C. IR (KBr) 3234, 3182, 3057,
2984, 2930, 2604, 2485, 1677, 1618 cmÀ1 1H NMR (DMSO-d6) d
;
3.00–3.47 (m, 2H), 4.27–4.40 (m, 3H), 4.46 (s, 2H), 6.89 (s, 1H),
6.90–6.91 (m, 1H), 7.19–7.22 (m, 2H), 7.40–7.43 (m, 1H), 7.59–
7.61 (m, 1H), 7.88 (s, 1H), 9.00 (br s, 1H), 9.67 (br s, 1H), 9.90 (br
s, 1H), 10.90 (s, 1H), 11.36 (s, 1H); HRMS (AP-ESI) m/z calcd for
C18H19ClN3O4 [M+H]+ 376.1064, found: 376.1078.
5.1.7.15. (S)-N-tert -Butyl-7-(2-(hydroxyamino)-2-oxoethoxy)-
1,2,3,4-tetrahydroisoquinoline-3-carboxamide hydrochloride
(8p). Yield: 93%, mp: 179–181 °C. IR (KBr) 3466, 3234, 3069,

Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
1771
2973, 2932, 2615, 2491, 1671, 1618 cmÀ1
;
1H NMR (DMSO-d6) d
Compounds 13a–c were synthesized according to the same pro-
cedure as compound 7a.
1.33 (s, 9H), 2.83–3.25 (m, 2H), 3.97–3.99 (m, 1H), 4.21 (d,
J = 15.6 Hz, 1H), 4.30 (d, J = 15.6 Hz, 1H), 4.44 (s, 2H), 6.85 (s, 1H),
6.87 (d, J = 8.4 Hz, 1H), 7.18 (d, J = 8.4 Hz, 1H), 8.25 (s, 1H), 8.99
(s, 1H), 9.36 (br s, 1H), 9.62 (br s, 1H), 10.88 (s, 1H); HRMS (AP-
ESI) m/z calcd for C16H24N3O4 [M+H]+ 322.1767, found: 322.1756.
5.1.12. (S)-Benzyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-
(phenethylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-
carboxylate (13a)
Yield: 23%, mp: 151–153 °C. IR (KBr) 3264, 3233, 3027, 2929,
5.1.8. (S)-7-Hydroxy-Tic (9)33
2857, 1671, 1649 cmÀ1 1H NMR (DMSO-d6) d 2.53–2.57 (m, 2H),
;
A solution of compound 2 (4.45 g, 9.24 mmol) in EtOH (150 mL)
and H2O (50 mL) containing triethylamine (3.08 g, 30.5 mmol) was
hydrogenated in the presence of 10% Pd/C (0.55 g) for 4 h. The cat-
alyst was filtered off with the solution being evaporated until crys-
tals started to appear. The pH was adjusted to 6 with 2 N HCl. After
cooling overnight in the refrigerator, the crystals were filtered,
washed with cold water and dried to give 1.23 g of desired com-
pound 9. This product was used for the following reaction without
further purification. Mp: 253–255 °C. 1H NMR (DMSO-d6) d 2.79
(dd, J = 16.8 Hz, 10.8 Hz, 1H), 3.03 (dd, J = 16.8 Hz, 4.8 Hz, 1H),
3.32 (br s, 1H), 3.47 (dd, J = 4.8 Hz, 10.8 Hz, 1H), 4.04–4.09 (m,
2H), 6.57 (s, 1H), 6.64 (d, J = 8.4 Hz, 1H), 6.99 (d, J = 8.4 Hz, 1H),
8.92 (br s, 1H), 9.47 (br s, 1H).
2.93–3.01 (m, 2H), 3.15–3.18 (m, 2H), 4.41 (s, 2H), 4.51–4.72 (m,
3H), 5.06–5.19 (m, 2H), 6.78–6.83 (m, 2H), 7.03–7.08 (m, 2H),
7.16–7.44 (m, 9H), 8.01 (s, 1H), 8.96 (s, 1H), 10.81 (s, 1H); HRMS
(AP-ESI) m/z calcd for C28H30N3O6 [M+H]+ 504.2135, found:
504.2125.
5.1.12.1. (S)-Benzyl 7-(2-(hydroxyamino)-2-oxoethoxy)-3-(phe-
nylcarbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(13b). Yield: 25%, mp: 145–147 °C. IR (KBr) 3050, 2933, 1768,
1709, 1617 cmÀ1 1H NMR (DMSO-d6) d 2.98–3.18 (m, 2H), 4.40–
;
4.52 (m, 2H), 4.45 (s, 2H), 4.87–4.90 (m, 1H), 6.86–6.93 (m, 2H),
7.22–7.23 (m, 1H), 7.40–7.51 (m, 5H), 8.97 (s, 1H), 10.83 (s, 1H);
HRMS (AP-ESI) m/z calcd for C19H18N3O5 [M+H]+ 368.1246, found:
368.1249.
5.1.9. (S)-2-(Benzyloxycarbonyl)-7-hydroxy-Tic (10)37
At 0 °C, to a solution of compound 9 (1.23 g, 6.38 mmol) in 7 mL
of 1 N NaOH, was added Cbz–Cl (1.20 g, 7.01 mmol) dropwise. The
solution was kept between pH 9–11 by addition of 1 N NaOH. After
stirring the mixture for 3 h, the solution was adjusted to pH 4–5
with 1 N HCl. Then the mixture was extracted with EtOAc
(3 Â 10 mL). The extractions were combined, washed with brine
(3 Â 10 mL), dried over Mg2SO4 and evaporated to give 2.00 g of
crude product compound 10 as light yellow oil. This product was
used for the following reaction without further purification. 1H
NMR (DMSO-d6) d 3.00–3.07 (m, 2H), 4.38–4.65 (m, 2H), 4.84–
4.85 (m, 1H), 5.10–5.19 (m, 2H), 6.59–6.63 (m, 2H), 6.98–7.00
(m, 1H), 7.30–7.43 (m, 5H), 9.33 (br s, 1H), 12.65 (br s, 1H).
Compounds 11a–c were synthesized according to the same pro-
cedure as compound 5a.
5.1.12.2. (S)-Benzyl 3-(biphenyl-4-ylcarbamoyl)-7-(2-(hydroxya-
mino)-2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxyl-
ate (13c). Yield: 20%, mp: 187–189 °C. IR (KBr) 3466, 1772, 1718,
1614 cmÀ1; 1H NMR (DMSO-d6) d 3.01–3.19 (m, 2H), 4.42–4.45 (m,
2H), 4.67 (s, 2H), 4.89–4.92 (m, 1H), 6.82–6.92 (m, 2H), 7.21–7.24
(m, 1H), 7.39–7.41 (m, 1H), 7.49–7.52 (m, 4H), 7.71–7.80 (m, 4H),
8.97 (s, 1H), 10.83 (s, 1H); HRMS (AP-ESI) m/z calcd for C25H22N3O5
[M+H]+ 444.1553, found: 444.1559.
5.2. Biological materials and methods
5.2.1. In vitro HDAC8 assay
IC50 values against HDAC8 were determined in 15 mM Tris–HCl,
pH 8.0, at 37 °C, using Boc-Lys (acetyl)-AMC as substrate and
HDAC8 expressed in Escherichia coli as the enzyme. The HDAC8
was expressed and purified essentially as the protocol described
in the supporting materials and methods of Ref. 38. The compound
samples and the positive control drug were diluted to various con-
centrations. Firstly, on the 96-well plate, HDAC8 was incubated at
37 °C with compound sample for 5 min, and then the substrate was
added. The HDAC8 reaction was allowed to proceed for 30 min,
during which Boc-Lys (acetyl)-AMC could be hydrolyzed to Boc-
Lys-AMC. Secondly, trypsin solution was added to transform Boc-
Lys-AMC to AMC (4-amino-7-methylcoumarin) for another
20 min. Finally, fluorescence intensity was measured using a
microplate reader (ex: 390 nm and em: 460 nm). The inhibition ra-
tios were calculated from the fluorescence intensity readings of
inhibited wells relative to those of control wells, and the IC50 val-
ues were determined using a regression analysis of the concentra-
tion/inhibition data.
5.1.10. (S)-Benzyl 7-hydroxy-3-(phenethylcarbamoyl)-3,4-
dihydroisoquinoline-2(1H)-carboxylate (11a)
Yield: 67%, ESI-MS m/z: 431.2 [M+H]+; 1H NMR (DMSO-d6) d
3.04–3.22 (m, 2H), 4.65–4.90 (m, 3H), 5.04–5.18 (m, 2H), 6.75–
6.79 (m, 1H), 6.89–6.94 (m, 1H), 7.02–7.06 (m, 1H), 7.11–7.54
(m, 10H), 9.33 (br s, 1H), 10.06 (s, 1H).
5.1.10.1. (S)-Benzyl 7-hydroxy-3-(phenylcarbamoyl)-3,4-dihy-
droisoquinoline-2(1H)-carboxylate (11b). Yield: 69%.
5.1.10.2. (S)-Benzyl 3-(biphenyl-4-ylcarbamoyl)-7-hydroxy-3,4-
dihydroisoquinoline-2(1H)-carboxylate (11c). Yield: 71%.
Compounds 12a–c were synthesized according to the same pro-
cedure as compound 6a.
5.1.11. (S)-Benzyl 7-(2-methoxy-2-oxoethoxy)-3-(phenethylc-
arbamoyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (12a)
Yield: 37%, ESI-MS m/z: 503.3 [M+H]+; 1H NMR (DMSO-d6) d
3.03–3.21 (m, 2H), 3.69 (s, 3H), 4.64–4.89 (m, 3H), 4.76 (s, 2H),
5.04–5.18 (m, 2H), 6.74–6.78 (m, 1H), 6.87–6.92 (m, 1H), 7.01–
7.05 (m, 1H), 7.10–7.53 (m, 10H), 10.06 (s, 1H).
5.2.2. MTT Assay
HCT116, SKOV3 and HL60 cells were respectively grown in
RPMI1640 medium containing 10% FBS at 37 °C in 5% CO2 humid-
ified incubator. Cell proliferation was determined by the MTT (3-
[4,5-dimethyl-2-thiazolyl]-2,5-diphenyl-2H-tetrazolium bromide)
assay. Briefly, cells were plated in a 96-well plate at 10,000 cells
per well, cultured for 4 h in complete growth medium, then treated
5.1.11.1. (S)-Benzyl 7-(2-methoxy-2-oxoethoxy)-3-(phenylcarba-
moyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (12b). Yield:
39%.
with 2000, 400, 80, 16, 3.2, 0.64
MTT solution was added to each well. After further incubation for
4 h, formazan formed from MTT was extracted by adding 200
lg/mL of compounds for 48 h. 0.5%
5.1.11.2. (S)-Benzyl 3-(biphenyl-4-ylcarbamoyl)-7-(2-methoxy-
2-oxoethoxy)-3,4-dihydroisoquinoline-2(1H)-carboxylate
(12c). Yield: 40%.
lL
DMSO and mixing for 15 min. Optical density was read with an ELI-
SA reader at 570 nm.

1772
Y. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1761–1772
17. Butler, L. M.; Agus, D. B.; Scher, H. I.; Higgins, B.; Rose, A.; Cordon-Cardo, C.;
Acknowledgements
Thaler, H. T.; Rifkind, R. A.; Marks, P. A.; Richon, V. M. Cancer Res. 2000, 60, 5165.
18. Garber, K. Nat. Biotechnol. 2007, 25, 17.
This work was supported by National Nature Science Founda-
tion of China (Grant Nos. 30772654 and 36072541) and the Ph.D.
Programs Foundation of Ministry of Education of P. R. China (No.
20060422029) and National High Technology Research and Devel-
opment Program of China (863 project; Grant No. 2007AA02Z314).
19. Grant, S.; Easley, C.; Kirkpatrick, P. Nat. Rev. Drug Disc. 2007, 6, 21.
20. Marks, P. A.; Breslow, R. Nat. Biotechnol. 2007, 25, 84.
21. Sangthongpitag, K.; Wu, X.; Khng, H.; Goh, S.; Ng, P.; Hu, C.; Bonday, Z.; Wang,
H.; Wood, J.; Entzeroth, M. Eur. J. Cancer Suppl. 2006, 4, 52.
22. Fiskus, W.; Pranpat, M.; Bali, P.; Balasis, M.; Kumaraswamy, S.; Boyapalle, S.;
Rocha, K.; Wu, J.; Giles, F.; Manley, P. W.; Atadja, P.; Bhalla, K. Blood 2006, 108,
645.
23. Armeanu, S.; Pathil, A.; Venturelli, S.; Mascagni, P.; Weiss, T. S.; Gottlicher, M.;
Gregor, M.; Lauer, U. M.; Bitzer, M. J. Hepatol. 2005, 42, 210.
24. Arts, J.; Angibaud, P.; Marien, A.; Floren, W.; Janssens, B.; King, P.; van Dun, J.;
Janssen, L.; Geerts, T.; Tuman, R. W.; Johnson, D. L.; Andries, L.; Jung, M.;
Janicot, M.; van Emelen, K. Br. J. Cancer 2007, 97, 1344.
References and notes
1. Christian, A. H.; Stuart, L. S. Curr. Opin. Chem. Biol. 1997, 1, 300.
2. Kouzarides, T. Curr. Opin. Genet. Dev. 1999, 9, 40.
3. Wolffe, A. P. Science (Washington) 1996, 272, 371.
4. Polo, S. E.; Almouzni, G. Cancer Lett. 2005, 220, 1.
25. Wang, A.; U’Prichard, D. C.; Marder, V. J. US Patent 2002/0061885 A1.
26. Vendeville, S.; Goossens, F.; Debreu-Fontaine, M. A.; Landry, V.; Davioud-
Charvet, E.; Grellier, P.; Scharpe, S.; Sergheraert, G. Bioorg. Med. Chem. 2002, 10,
1719.
27. Balboni, G.; Salvadori, S.; Guerrini, R.; Negri, L.; Giannini, E.; Jinsmaa, Y.; Bryant,
S. D.; Lazarus, L. H. J. Med. Chem. 2002, 45, 5556.
28. Balboni, G.; Guerrini, R.; Salvadori, S.; Bianchi, C.; Rizzi, D.; Bryant, S. D.;
Lazarus, L. H. J. Med. Chem. 2002, 45, 713.
29. Kotha, S.; Sreenivasachary, N. Eur. J. Org. Chem. 2001, 17, 3375.
30. Cueva, J. P.; Cai, T. B.; Mascarella, S. W.; Thomas, J. B.; Navarro, H. A.; Carroll, F.
I. J. Med. Chem. 2009, 52, 7463.
5. Vidanes, G. M.; Bonilla, C. Y.; Toczyski, D. P. Cell 2005, 121, 973.
6. Glozak, M. A.; Sengupta, N.; Zhang, X. H.; Seto, E. Gene 2005, 363, 15.
7. Yang, X.; Seto, E. Nat. Rev. Mol. Cell Biol. 2008, 9, 206.
8. Fraga, M. F.; Ballestar, E.; Villar-Garea, A.; Boix-Chornet, M.; Espada, J.; Schotta,
G.; Bonaldi, T.; Haydon, C.; Ropero, S.; Petrie, K.; Iyer, N. G.; Perez-Rosado, A.;
Calvo, E.; Lopez, J. A.; Cano, A.; Calasanz, M. J.; Colomer, D.; Piris, M. A.; Ahn, N.;
Imhof, A.; Caldas, C.; Jenuwein, T.; Esteller, M. Nat. Genet. 2005, 37, 391.
9. Halkidou, K.; Gaughan, L.; Cook, S.; Leung, H. Y.; Neal, D. E.; Robson, C. N.
Prostate 2004, 59, 177.
10. Choi, J. H.; Kwon, H. J.; Yoon, B.-I.; Kim, J. H.; Han, S. U.; Joo, H. J.; Kim, D.-Y. Jpn.
J. Cancer Res. 2001, 92, 1300.
11. Wilson, A. J.; Byun, D.-S.; Popova, N.; Murray, L. B.; L’Italien, K.; Sowa, Y.;
Arango, D.; Velcich, A.; Augenlicht, L. H.; Mariadason, J. M. J. Biol. Chem. 2006,
281, 13548.
12. Zhang, Z. H.; Yamashita, H.; Toyama, T.; Sugiura, H.; Ando, Y.; Mita, K.;
Hamaguchi, M.; Hara, Y.; Kobayashi, S.; Iwase, H. Breast Cancer Res. Treat. 2005,
94, 11.
31. Matter, H.; Schudok, M.; Schwab, W.; Thorwart, W.; Barbier, D.; Billen, G.;
Haase, B.; Neises, B.; Weithmann, K. U.; Wollmann, T. Bioorg. Med. Chem. 2002,
10, 3529.
32. Klutchko, S.; Blankley, C. J.; Fleming, R. W.; Hinkley, J. M.; Werner, A. E.; Nordin,
I.; Holmes, A.; Hoefle, M. L.; Cohen, D. M.; Essenburg, A. D.; Kaplan, H. R. J. Med.
Chem. 1986, 29, 1953.
33. Verschueren, K.; Toth, G.; Tourwé, D.; Lebl, M.; Van Binst, G.; Hruby, V.
Synthesis 1992, 5, 458.
13. Zhu, P.; Martin, E.; Mengwasser, J.; Schlag, P.; Janssen, K.-P.; Gottlicher, M.
Cancer Cell. 2004, 5, 455.
34. Bodanszky, M.; Scheehan, T. J.; Ohdetti, A. M.; Lande, S. J. Am. Chem. Soc. 1963,
85, 991.
14. Huang, B. H.; Laban, M.; Leung, C. H.-W.; Lee, L.; Lee, C. K.; Salto-Tellez, M.;
Raju, G. C.; Hooi, S. C. Cell Death Differ. 2005, 12, 395.
35. Maclaren, A. J. Aust. J. Chem. 1958, 11, 360.
36. Dekker, A. C.; Taylor, P. S.; Rruton, S. J. J. Biol. Chem. 1949, 180, 155.
37. Faul, M. M.; Winneroski, L. L.; York, J. S.; Reinhard, M. R.; Hoying, R. C.; Gritton,
W. H.; Dominianni, S. J. Heterocycles 2001, 55, 689.
38. Vannini, A.; Volpari, C.; Filocamo, G.; Caroli Casavola, E.; 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.
15. Song, J.; Noh, J. H.; Lee, J. H.; Eun, J. W.; Ahn, Y. M.; Kim, S. Y.; Lee, S. H.; Park, W.
S.; Yoo, N. J.; Lee, J. Y.; Nam, S. W. APMIS 2005, 113, 264.
16. Zhang, Z. H.; Yamashita, H.; Toyama, T.; Sugiura, H.; Omoto, Y.; Ando, Y.;
Mita, K.; Hamaguchi, M.; Hayashi, S.-I.; Iwase, H. Clin. Cancer Res. 2004, 10,
6962.