Structure-activity relationship studies of a series of novel δ-lactam-based histone deacetylase inhibitors

Article abstract of DOI:10.1021/jm0613828

We synthesized a series of δ-lactam-based HDAC inhibitors that were identified with various degrees of anti-inflammatory and cell growth inhibitory activities. Compounds possessing significant HDAC inhibitory activity exhibited both anti-inflammatory and cell growth inhibitory activities as well as significant tumor growth inhibition in the in vivo tumor xenograft experiments. Besides, these compounds demonstrated antiinflammatory properties in vitro via suppression of the production of the proinflammatory cytokine TNF-α and nitric oxide by LPS-stimulated RAW264.7 cells.

Full text of DOI:10.1021/jm0613828

J. Med. Chem. 2007, 50, 2737-2741
2737
Structure-Activity Relationship Studies of a Series of Novel δ-Lactam-Based Histone
Deacetylase Inhibitors
Hwan Mook Kim,† Dong-Kyu Ryu,† Yongseok Choi,‡ Bum Woo Park,† Kiho Lee,† Sang Bae Han,† Chang-Woo Lee,†
Moo-Rim Kang,† Jong Soon Kang,† Shanthaveerappa K. Boovanahalli,† Song-Kyu Park,*,† Jung Whan Han,§ Tae-Gyu Chun,|,
Hee-Yoon Lee,| Ky-Youb Nam,# Eun Hyun Choi, and Gyoonhee Han*,
BioeValuation Center, KRIBB, Daejeon 305-806, Korea, School of Life Sciences and Biotechnology, Korea UniVersity, Seoul 136-701 Korea,
College of Pharmacy, Sung Kyun Kwan UniVersity, Suwon, Gyeonggi-do 440-746, Korea, Department of Chemistry, KAIST, Daejeon 305-701,
Korea, Research Institute Bioinformatics and Molecular Design (BMD), Yonsei Engineering Research Complex, Seodaemun-gu, Seoul 120-740,
Korea, and Department of Biotechnology, Yonsei UniVersity, Seodaemun-gu, Seoul 120-740, Korea
ReceiVed December 1, 2006
We synthesized a series of δ-lactam-based HDAC inhibitors that were identified with various degrees of
anti-inflammatory and cell growth inhibitory activities. Compounds possessing significant HDAC inhibitory
activity exhibited both anti-inflammatory and cell growth inhibitory activities as well as significant tumor
growth inhibition in the in ViVo tumor xenograft experiments. Besides, these compounds demonstrated anti-
inflammatory properties in Vitro via suppression of the production of the proinflammatory cytokine TNF-R
and nitric oxide by LPS-stimulated RAW264.7 cells.
Introduction
properties of SAHA have been demonstrated via in Vitro and
in ViVo suppression of proinflammatory cytokine tumor necrosis
factor (TNF-R) as well as inhibition of in Vitro production of
cytokine-induced nitric oxide (NO).5a Based on these findings,
we hypothesized that the dual anti-inflammatory and antipro-
liferative activity of HDAC inhibitors is in part attributable to
the potent HDAC inhibitory activity. These key findings
prompted us to investigate a series of potent HDAC inhibitors
for their potential to exhibit anti-inflammatory and antiprolif-
erative activities.
In our search for novel HDAC inhibitors, we recently
disclosed a series of δ-lactam-based HDAC inhibitors6 and we
identified a lead compound (3) in this series, which showed
significant HDAC inhibitory activity comparable to SAHA.
Based on this lead molecule, we performed a closely related
structural activity relationship (SAR) study and then generated
a new series of potent δ-lactam-based HDAC inhibitors, which
were evaluated for their anti-inflammatory and antiproliferative
activities. These studies were culminated in the identification
of a series of potent δ-lactam-based HDAC inhibitors, which
demonstrated significant inhibitions of HDAC activity as well
as anti-inflammatory and cell growth inhibitory activities.
Herein, we report our findings derived from a systematic
evaluation of correlation of HDAC inhibition to anti-inflam-
matory and cell growth inhibitory activities of a new series of
novel δ-lactam-based HDAC inhibitors.
The first connection between inflammation and cancer was
made much earlier by Rudolf Virchow when he discovered
leucocytes in neoplastic tissues and linked tumor with inflam-
mation.1 Since then, a substantial body of evidence has been
published to support the conclusion that chronic inflammation
represents a key risk factor for cancer and even as a means to
prognose/diagnose cancer at the onset of the disease.2 In
addition, recent data have expanded the concept that inflam-
mation is a critical component of tumor progression. The longer
the inflammation persists, the higher the risk of associated
carcinogenesis.3 Therefore, new strategies for the prevention
and treatment of tumors associated with inflammatory diseases
would open new therapeutic avenues.
Histone deacetylases (HDACsa) play an important role in gene
transcription. Many recent studies show that inhibition of
HDACs elicits anticancer effects in several types of tumor cells
by inhibition of cell growth and induction of cell differentiation
and this has led to the development of specific inhibitors for
cancer chemotherapy. In this context, several programs for the
development of HDAC inhibitors as an anticancer drug have
been initiated, and currently, several compounds are in both
preclinical development and clinical trials.4 HDAC inhibitors
such as suberoylanilide hydroxamic acid (SAHA, 1) and
trichostatin A (TSA, 2), the class of hydroxamic acid-containing
hybrid polar molecules, possess antitumor activity as well as
anti-inflammatory properties (Figure 1).5 The anti-inflammatory
Chemistry
The synthesis of δ-lactam analogues 13-15 and 21 was
accomplished according to the synthetic protocols as illustrated
in Schemes 1 and 2.6 As shown in Scheme 1, reaction of
4-bromo-1-butene with the appropriate amines 4 yielded the
corresponding alkylated secondary amines 5. EDC-mediated
coupling of these amines 5 with the appropriate acids 6 afforded
the respective intermediates 7-9 in good yields. Further
cyclization was achieved under mild conditions using catalytic
2-3 mol % of Grubb’s catalyst (I) in high yields. The resulting
ester derivatives 10-12 were then reacted with KONH2 to
obtain the desired hydroxamides 13-15. Following a similar
sequence of reactions, commercially available 2, 4-dimethoxy
* To whom correspondence should be addressed. Tel.: +82-42-860-
4689 (S.-K.P.); +82-2-2123-2882 (G.H.). Fax: +82-42-860-4605 (S.-K.P.);
+82-2-3362-7265 (G.H.). E-mail: spark123@kribb.re.kr (S.-K.P.);
gyoonhee@yonsei.ac.kr (G.H.).
† Bioevaluation Center, KRIBB.
‡ Korea University.
§ Sung Kyun Kwan University.
| Department of Chemistry, KAIST.
Yonsei University.
# Research Institute Bioinformatics and Molecular Design (BMD).
a Abbreviations: HDAC, histone deacetylase; SAHA, suberoylanilide
hydroxamic acid; TSA, trichostatin A; TNF-R, tumor necrosis factor R;
NO, nitric oxide; CAP, hydrophobic aromatic groups; DMB, dimethoxy-
benzyl; EDC, 1-ethyl-3-(3-dimethylaminopropyl)-dicarboximide hydrochlo-
ride.
10.1021/jm0613828 CCC: $37.00 © 2007 American Chemical Society
Published on Web 05/04/2007

2738 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 11
Brief Articles
Figure 1. Chemical structure of SAHA and TSA.
Scheme 1a
Table 1. In Vitro Inhibition of the Activity of HDAC Enzyme and the
Production of TNF-R and NO by SAHA and δ-Lactam Analogues
13-15 and 21a
IC50 (µM)
cmpd
13a
13b
13c
13d
14a
14b (3)
14c
14d
15b
15c
15d
21a
21b
21c
SAHA
m
n
R1
R2
HDACb
TNF-Rc
NOc
1
2
3
4
1
2
3
4
2
3
4
1
1
1
1
1
1
1
2
2
2
2
3
3
3
2
2
2
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
H
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
NAe
NA
NA
NA
NA
NA
NA
3.31
1.90
1.10
1.30
NA
NA
NA
NA
10.96
5.06
0.76
NA
NA
NA
NA
3.09
4.27
2.71
1.80
NA
6.57
6.74
NA
NA
0.67
0.35
0.27
0.13
NA
3.21
NA
NA
-CHdCH2
4.27
0.50
0.11
NA
5.20
1.88
DMPhd
a Values are the means of a minimum of three experiments. b HDAC
enzyme obtained from HeLa cell lysate. c For NO and TNF-R assays,
inhibitors were treated in RAW264.7 cells with the range between 0.1 µM
and 10 µM. IC50 values were calculated to evaluate their ability for
suppressing the production of NO and TNF-R. d DMPh ) 2,4-dimethoxy
phenyl. e NA ) IC50 values are higher than 10 µM.
a Reagents and conditions: (a) 4-bromo-1-butene, hunig’s base, reflux;
(b) EDCI, DMAP, CH2Cl2, rt; (c) Grubb’s catalyst (I), rt; (d) KONH2,
MeOH, 0 °C to rt.
Scheme 2a
analogues 20a,b. Subsequent reaction of 20a,b with KONH2
afforded the corresponding hydroxamides 21a,b (Scheme 2).
Results and Discussion
Compounds 13-15 and 21 were assayed for their HDAC
inhibitory activity and anti-inflammatory activity, and the results
are tabulated as IC50 values in Table 1. SAHA, a known HDAC
inhibitor and antitumor agent, was used as a reference compound
for comparison. The partially purified HDAC enzyme was
obtained from HeLa cell lysate for measuring enzyme inhibitory
activity. A potent proinflammatory cytokine TNF-R and a well-
known proinflammatory key mediator NO, which play an
important role in inflammation, were used as indicators to assess
the anti-inflammatory activity of HDAC inhibitors.
HDAC inhibitors SAHA (1) and TSA (2) and most of their
analogues have an aliphatic chain between the hydroxamic acid
(zinc binder) and the hydrophobic aromatic groups (CAP group).
The alkyl chain of SAHA inserts into the tubular hydrophobic
pocket, and the hydroxamic acid group serves as a bidentate
chelator of the catalytic Zn2+ ion bound in the enzyme active
site.7 The zinc binder and cap groups of the compound 3 were
attached through a δ-lactam ring, which mimic the hydrophobic
tail group and the aliphatic chain of SAHA, and therefore, this
compound also should bind to the hydrophobic cavity in a
similar fashion as SAHA. Compound 3 showed potent HDAC
inhibition, with an IC50 value of 0.35 µM, and was chosen as
a starting template for the generation of a new series of δ-lactam
analogues. Because we hypothesized that the placement of
an appropriate spacer between the zinc binder and cap group
a Reagents and conditions: (a) 4-bromo-1-butene, hunig’s base, reflux;
(b) 6b, EDCI, DMAP, CH2Cl2, rt; (c) Grubb’s catalyst (I), rt; (d) TFA,
triethylsilane, reflux; (e) NaHMDS, -78 °C, 0.5 h, then Me2SO4 for 20a,
allyl bromide for 20b, -78 °C to 0 °C; (f) KONH2, MeOH, 0 °C to rt.
benzylamine 16 furnished the required intermediate 18, which
was subsequently converted directly to the required hydroxamide
21c under the usual reaction conditions described for the
previous hydroxamides. Displacement of the 2,4-dimethoxy
benzyl moiety in compound 18 was achieved under reflux
conditions in trifluoroacetic acid to obtain intermediate 19 in
good yield, which underwent alkylation to provide alkylated

Brief Articles
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 11 2739
Table 2. Cell Growth Inhibitory Activities of Analogues 14a-d and
would modulate the HDAC inhibitory activity, our SAR was
focused on the following modifications: (i) the variation of the
chain length between the zinc binder and the cap group by
retaining the δ-lactam ring, (ii) altering the cap group with
different alkyl and aromatic groups, and (iii) replacement of
the hydroxamic acid group with an o-aminoanilide moiety.
21ca
growth inhibitionb (µM)
cell line
HCT-15
LOX-IMVI
PC-3
ACHN
MDA-MB-231 breast
NCI-H23 lung
tissue
colon
melanoma NA 5.84 3.34 NA 6.63
14a 14b 14c 14d 21c SAHA
NAc 8.16 4.99 5.37 7.98
0.82
1.39
0.89
0.73
0.66
0.92
prostate
kidney
NA 8.58 5.04 1.32 4.55
NA 7.76 1.17 2.50 9.58
4.26 1.62 0.50 1.50 2.67
NA NA 3.79 5.14 NA
To assess the effect of the chain length, we prepared various
hydroxamic acid derivatives with varying chain lengths between
the zinc binder and the cap group, which resulted in the
analogues 13-15 having 1-4 carbon units between the cap
group and the δ-lactam ring, in addition to 1-3 carbon units
between the δ-lactam ring and the hydroxamic acid moiety. We
first examined analogues 13a-d having 1-4 carbon units
between the cap group and the δ-lactam ring, in addition to a
methylene bridge between the δ-lactam ring and the zinc binder
group for their ability to inhibit HDAC enzyme, but all of these
analogues were found to be inactive. We then investigated the
effect of elongation of the alkyl linker by two carbon units
between the δ-lactam ring and the zinc binder group by
maintaining 1-4 carbon units between the cap group and the
δ-lactam ring. Interestingly, this modification, which is repre-
sented by analogues 14a-d, demonstrated appreciably high
inhibitory activity as good as SAHA’s. Encouraged by these
results, we further increased the length of the alkyl linker by
three methylene units between δ-lactam ring and zinc binder
group, but this led to the complete loss of potency, with the
exception of analogue 15c, which displayed moderate inhibitory
activity. From these results it is evident that the potency of
δ-lactam analogues is directly related to the alkyl linker between
the cap group and the zinc binder group, suggesting that the
presence of 3-4 carbon units between the cap group and the
δ-lactam ring, in addition to two carbon units between the
δ-lactam ring and the zinc binder group is optimal for potency.
Having established the optimal chain length for potency, we
were interested in exploring the effects of altering the cap group
and the zinc binder group to assess whether such modifications
could affect potency. Replacements of the phenyl cap group
with methyl, allyl, and dimethoxy phenyl was investigated by
synthesizing analogues 21a-c. However, only dimethoxy
phenyl derivative 21c showed significant potency, with an IC50
value of 0.5 µM, which emphasizes that a phenyl group at this
position is optimal for potency.
a Values are the means of a minimum of three experiments. b Growth
inhibition was measured by SRB (sulforhodamine B) assay. c NA ) not
active.
Table 3. Inhibition of the Growth of the MDA-MB-231 Human Breast
Xenograft by Compound 14d and SAHAa
i.p.b
cmpd
dose (mg/kg)
30
14d
SAHA
inhibition (%)
inhibition (%)
51c
39d
a Nude mice (six per group) were treated. b 14d and SAHA were
administered (i.p.) after the size of tumors reached to 50-60 mm3. c P <
0.001. d P < 0.01.
IMVI, PC-3, ACHN, MDA-MB-231, and NCI-H23 cells. Cell
growth inhibition of the tested analogues was measured by SRB
assay,8 and the GI50 values obtained for the types of tumor cell
lines are summarized in Table 2. High cell growth inhibitory
potency against the entire cell types was observed with the
reference drug SAHA. Analogues 14c,d, the most potent HDAC
inhibitors among the series, demonstrated significant antipro-
liferative activity on all of the cell types examined, with the
exception of the inactivity of 14d on the LOX-IMVI cell line.
Whereas compounds 14b and 21c were inactive on the NCI-
H23 cell line and displayed considerable activity on all of the
remaining cell lines, compound 14a exhibited a considerable
growth inhibitory activity only on MDA-MB-231 cells. Thus,
overall tumor cell growth inhibitory activities of these analogues
are related to their HDAC inhibitory activities.
The most promising inhibitor 14d was selected to be further
evaluated for in ViVo tumor growth inhibitory activity, and the
results are summarized in Table 3. In this model, compound
14d and SAHA were administered daily (i.p., 30 mg/kg) to nude
mice after the sizes of MDA-MB-231 human breast xenografts
reached to 50-60 mm3. Compound 14d and SAHA caused 51%
and 39% inhibition of tumor growth, respectively, compared
to the group without treatment of HDAC inhibitors. Thus, both
compound 14d and SAHA displayed antitumor growth activities,
and 14d seemed to be better than SAHA in terms of inhibiting
the growth of tumor.
To understand the binding mode of these inhibitors, we
examined a docking model of human homology HDAC-1 model
based on the crystal structure of a bacterial HDAC homologue
(HDLP, PDB code 1C3R) and docked compounds 14d within
the site using the program InsightII/Discover (Figure 2).9 The
complex binding model for the analogs were carried out with
the Discover program.10 The δ-lactam ring of 14d passes
through a narrow channel in the binding pocket, which is formed
by two aromatic side chains of Phe-142 and Phe-197. The
aliphatic chain between the hydroxamate and the δ-lactam ring
of 14d was bound in the tubular hydrophobic pocket, and the
hydroxamic acid moiety of 14d chelates to the catalytic Zn2+
ion bound in the active site. The docking data suggest that the
chain length between the zinc binder group and the δ-lactam
ring is quite important to form the flexible conformation in the
binding pocket and that the carbonyl in the δ-lactam ring forms
To examine whether there exists a correlation between HDAC
enzyme inhibition and anti-inflammatory activity, these ana-
logues were further assayed for their ability to inhibit the
production of TNF-R and NO by RAW264.7 cells. Interestingly,
analogues 14a-d and 21c, possessing significant HDAC
inhibitory activity, greatly inhibited the production of both
TNF-R and NO, while compounds 15c and 21b, with moderate
HDAC inhibitory activity, displayed reasonable inhibition of
both TNF-R and NO. On the other hand, compounds 13a-d,
15b, and 21a, which failed to inhibit HDAC enzyme, were also
found to be inactive to inhibit the production of TNF-R and
NO, with the exception of analogue 15d, which inhibited NO
production considerably. Thus, only compounds capable of
inhibiting HDAC enzyme were able to inhibit the production
of TNF-R and NO. These results strongly support our hypothesis
that there exists a correlation between HDAC inhibitory activity
and anti-inflammatory activity among this class of compounds.
Because it is well documented in the literature that HDAC
inhibitors display cell growth inhibitory activity, analogues
14a-d and 21c, possessing significant HDAC inhibitory activ-
ity, were further tested for their antiproliferative effect using a
panel of cancer cell lines, which consisted of HCT-15, LOX-

2740 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 11
Brief Articles
General Procedures for 7-9. Compound 6a, 6b, or 6c (1.2
equiv) was dissolved in methylene chloride and then 5a in
methylene chloride was added via cannular at room temperature
under an Ar atmosphere. EDC (1.3 equiv) and DMAP (0.3 equiv)
were added, and the mixture was stirred for 8 h at room temperature.
The mixture was diluted with ethyl acetate and washed with 5%
HCl, satd NaHCO3 solution, and brine, and the organic layer was
dried over anhydrous MgSO4. The desired compound (9-11) was
obtained by flash chromatography.
3-(Benzyl-but-3-enyl-carbamoyl)-but-3-enoic Acid Methyl
Ester (7a). Compound 5a (248 mg, 1.53 mmol) was used and 7a
(170 mg, 38%, oil) was obtained by flash chromatography (ethyl
1
acetate/hexanes, 1/1). H NMR (CDCl3) δ 7.29 (d, J ) 6.9 Hz,
2H), 7.22 (t, J ) 6.9 Hz, 3H), 5.69 (br t, 1H), 5.23 (s, 2H), 5.04 (s,
1H), 4.97 (d, J ) 9.3 Hz, 1H), 4.71 (d, J ) 16.9 Hz, 2H), 3.61 (s,
3H), 3.42 (s, 4H), 2.30 (q, J ) 7.2 Hz, 2H); ESI (m/z) 310.5
(MNa+).
General Procedures for 10-12. Compound 7, 8, or 9 was
dissolved in methylene chloride (0.01 M), and the mixture was
gassed by Ar bubbling for 30 min. After Grubb’s catalyst (Type I,
0.02 equiv) was added, the reaction mixture was stirred for 12 h in
the dark. The mixture was concentrated in vacuo, and the desired
compound (10-12) was obtained by flash chromatography.
(1-Benzyl-2-oxo-1,2,5,6-tetrahydro-pyridin-3-yl)-acetic Acid
Methyl Ester (10a). Compound 7a (77 mg, 0.26 mmol) was used,
and 10a (50 mg, 74%, oil) was obtained by flash chromatography
Figure 2. Docked orientations for compound 14d bound to the HDAC1
catalytic core.
the weak hydrogen bond (2.41 Å) with the side chain of His-
170. Furthermore, a longer chain length between the δ-lactam
and the phenyl group and the phenyl hydrophobic CAP group
give the extra stabilization energy by hydrophobic interaction
with Pro-21, Phe-142, and Leu-263.
1
(ethyl acetate/hexanes, 1/2). H NMR (CDCl3) δ 7.30-7.23 (m,
5H), 6.42 (t, J ) 4.2 Hz, 1H), 4.61 (s, 2H), 3.69 (s, 3H), 3.32 (t,
J ) 7.2 Hz, 4H), 2.32 (dd, J ) 6.9 Hz, 2H); 13C NMR (CDCl3) δ
171.96, 164.46, 137.34, 136.80, 129.34, 128.55, 127.90, 127.32,
51.91, 50.02, 44.68, 36.38, 23.92; ESI (m/z) 282.4 (MNa+).
General Procedures for 13-15. Compound 10, 11, or 12 was
dissolved in methanol (0.5 M solution) and then NH2OK (1.7 M
suspension in methanol, 5.0 equiv) was added at 0 °C. The mixture
was stirred for 20 h at room temperature. 10% HCl was added to
pH 2-3, and the mixture was concentrated in vacuo. White solid
was removed on filter with eluting of methanol/chloroform (1/9).
The desired compound (13-15) was obtained by flash chroma-
tography.
A series of novel δ-lactam-based HDAC inhibitors possessing
both anti-inflammatory and antiproliferative activities were
identified through a systematic exploration of SAR studies of a
previously reported δ-lactam analogue. The potencies of these
δ-lactam analogues are related to the chain length between the
CAP group and the zinc binder group, with 3-4 carbon units
between the cap group and the δ-lactam ring, in addition to
two carbon units optimal for potency between the δ-lactam ring
and the zinc binder group, which was also further confirmed
by docking studies. The anti-inflammatory properties of these
inhibitors were strongly dependent on their HDAC inhibitory
potency. Only compounds possessing significant HDAC inhibi-
tory activity demonstrated potent anti-inflammatory properties.
Among them, compounds 14c and 14d emerged as the most
promising HDAC inhibitors, which also demonstrated significant
cell growth inhibitory activity against a panel of cell lines.
Moreover, compound 14d showed the considerable extent of
tumor growth inhibition in the in vivo tumor xenograft experi-
ment. Besides, these analogues also exhibited anti-inflammatory
properties via the potent in vitro and in vivo inhibition of
cytokine TNF-R and in vitro inhibition of LPS-induced NO. In
summary, these results revealed a correlation between HDAC
inhibition and anti-inflammatory and cell growth inhibitory
activities of this class of δ-lactam analogues, and further
investigation is warranted to establish the mechanism of
inhibitory activity. This represents the first report aimed at the
identification of a series of HDAC inhibitors possessing anti-
inflammatory and antiproliferative properties.
2-(1-Benzyl-2-oxo-1,2,5,6-tetrahydro-pyridin-3-yl)-N-hydroxy-
acetamide (13a). Compound 10a (17 mg, 0.07 mmol) was used
and 13a (10 mg, 55%, oil) was obtained by flash chromatography
1
(methanol/chloroform, 1/19). H NMR (CDCl3) δ 7.34-7.22 (m,
5H), 6.58 (t, J ) 4.5 Hz, 1H), 4.60 (s, 2H) 3.39-3.30 (m, 3H)
3.20 (s, 2H), 2.39-2.30 (m, 2H); 13C NMR (CDCl3) δ 168.44,
165.53, 138.56, 136.73, 128.77, 128.02, 127.88, 127.68, 50.33,
44.81, 36.63, 23.86; ESI (m/z) 283.4 (MNa+); HRMS (m/z, MH+)
calcd for C14H17N2O3, 261.1234; found, 261.1226.
4-[But-3-enyl-(2,4-dimethoxy-benzyl)-carbamoyl]-pent-4-eno-
ic Acid Methyl Ester (17). The same procedures as the general
procedures for 7a are followed. But-3-enyl-(2,4-dimethoxy-benzyl)-
amine (500 mg, 2.26 mmol) was used and 17 (640 mg, 78%, oil)
was obtained by flash chromatography (ethyl acetate/hexanes, 1/1).
1H NMR (CDCl3) δ 6.96 (d, J ) 7.6 Hz, 1H), 6.44-6.42 (m, 2H),
5.72 (t, J ) 8.3 Hz, 1H,), 5.12-4.97 (m, 4H), 4.47 (s, 2H), 3.76
(s, 6H), 3.63 (s, 3H), 3.33 (br t, 1H), 2.62-2.51 (m, 4H), 2.26 (q,
J ) 6.8 Hz, 2H); ESI (m/z) 384.5 (MNa+).
3-[1-(2,4-Dimethoxy-benzyl)-2-oxo-1,2,5,6-tetrahydro-pyridin-
3-yl]-propionic Acid Methyl Ester (18). The same procedures as
the general procedures for 10a are followed. Compound 17 (640
mg, 1.77 mmol) was used and 18 (580 mg, 98%, oil) was obtained
Experimental Section
1
by flash chromatography (ethyl acetate/hexanes, 1/1). H NMR
General. All chemicals were obtained from commercial suppliers
(CDCl3) δ 7.19 (d, J ) 3.0 Hz, 1H), 6.45-6.91 (m, 2H), 6.27 (t,
J ) 4.3 Hz, 1H), 4.55 (s, 2H), 3.78 (s, 6H), 3.63 (s, 3H), 3.30 (t,
J ) 7.1 Hz, 2H,), 2.63-2.50 (m, 4H), 2.23 (d, J ) 6.7 Hz, 2H),
1.57 (s, 2H); 13C NMR (CDCl3) δ 173.6, 164.8, 160.2, 158.5, 134.2,
133.9, 130.4, 118.0, 104.1, 98.3, 55.2, 51.3, 45.0, 44.3, 33.3, 26.6,
23.9; ESI (m/z) 356.5 (MNa+).
3-(2-Oxo-1,2,5,6-tetrahydro-pyridin-3-yl)-propionic Acid Meth-
yl Ester (19). Compound 18 (490 mg, 1.47 mmol) was dissolved
in 3 mL of TFA and then triethylsilane was added. The reaction
and used without further purification. Flash column chromatography
1
was performed with silica (Merck EM9385, 230-400 mesh). H
and 13C NMR spectra were recorded at 300 and at 75 MHz,
respectively. Proton and carbon chemical shifts are expressed in
ppm relative to internal tetramethylsilane; coupling constants (J)
are expressed in Hertz. Cells were obtained from R&D Systems,
Minneapolis, MN. Products from all reactions were purified to a
minimum purity of 96%, as determined by reversed phase HPLC
(see Supporting Information).

Brief Articles
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 11 2741
mixture was refluxed for 20 min at 8 °C. The solvent was removed
in vacuo, and the residue was diluted in chloroform (50 mL). The
organic layer was washed with satd NaHCO3 solution (10 mL) and
brine (10 mL). The organic layer was dried over anhydrous MgSO4.
Compound 19 (200 mg, 74%, solid) was obtained by flash
Supporting Information Available: Experimental procedures
and characterization data of intermediates and final compounds,
including biological procedures. This material is available free of
charge via the Internet at http://pubs.acs.org.
1
chromatography (ethyl acetate/hexanes, 1/1). H NMR (CDCl3) δ
References
6.45 (s, 1H), 6.36 (t, J ) 4.2 Hz, 2H), 3.61 (s, 3H), 3.31 (dt, J )
7.5 Hz, 2H), 2.55-2.45 (m, 4H), 2.27 (dd, J ) 6.7, 5.8 Hz, 2H);
13C NMR (CDCl3) δ 173.41, 166.82, 136.09, 133.48, 51.35, 39.52,
33.06, 26.02, 24.01; ESI (m/z) 220.4 (MNa+).
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3-(1-Methyl-2-oxo-1,2,5,6-tetrahydro-pyridin-3-yl)-propion-
ic Acid Methyl Ester (20a). To a solution of 19 (100 mg, 0.55
mmol) in THF (1.1 mL) was added NaHMDS (0.66 mL, 1.0 M in
THF, 1.2 equiv) dropwise at -78 °C. After stirring at -78 °C for
30 min, dimethyl sulfate was added dropwise. The reaction mixture
was stirred at -78 °C for 15 min and at 0 °C for 2 h. The reaction
was quenched by satd NH4Cl solution. The organic layer was
extracted by ethyl acetate (30 mL × 3), and the combined organic
layer was washed with satd NH4Cl solution (10 mL) and brine (10
mL) and dried over anhydrous MgSO4. Compound 20a (80 mg,
74%, oil) was obtained by flash chromatography (ethyl acetate/
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1
hexanes, 1/1). H NMR (CDCl3) δ 6.28 (t, J ) 4.2 Hz, 1H), 3.62
(s, 3H), 3.33 (t, J ) 7.2 Hz, 2H), 2.96 (d, J ) 3.2 Hz, 3H), 2.58-
2.47 (m, 4H), 2.33-2.27 (m, 2H); 13C NMR (CDCl3) δ 173.90,
165.53, 134.33, 134.20, 51.65, 47.80, 34.86, 33.53, 26.80, 23.93;
ESI (m/z) 220.4 (MNa+).
General Procedures for 21a-c. N-Hydroxy-3-(1-methyl-2-
oxo-1,2,5,6-tetrahydro-pyridin-3-yl)-propionamide (21a). The
same procedures are followed as for 13a. Compound 20a (40 mg,
0.20 mmol) was used and 21a (13.8 mg, 35%, oil) was obtained
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tiinflammatory properties via suppression of cytokines. Proc. Natl.
Acad. Sci. U.S.A. 2002, 99, 2995-3000. (b) Mishra, N.; Reilly, C.
M.; Brown, D. R.; Ruiz, P.; Gilkeson, G. S. Histone deacetylase
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Park, S.-K.; Han, J. W.; Lee, H. Y.; Lee, H. Y.; Han, G. Synthesis,
enzymatic inhibition, and cancer cell growth inhibition of novel delta-
lactam-based histone deacetylase (HDAC) inhibitors. Bioorg. Med.
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(10) (a) The computational complex model was solvated using a solvent
sphere of water extending 23.0 Å around the zinc ion, and only
residues within 5.0 Å of compound 14d were allowed to move during
the geometry optimizations using 500 steps of steepest decent and
3000 steps of conjugated gradient. The hydroxamic acid coordinates
of 14d were restrained using 10.0 kcal/mol harmonic forces, the zinc
ion VDW radius was taken from the work of Stote and Karplus. (b)
Stote, R. H.; Karplus, M. Zinc binding in proteins and solution: a
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1
by flash chromatography (methanol/chloroform, 1/19). H NMR
(CDCl3) δ 6.15 (t, J ) 4.3 Hz, 1H), 3.41 (t, J ) 7.2 Hz, 2H), 2.97
(s, 3H), 2.51 (t, J ) 7.5 Hz, 2H), 2.35 (m, 2H), 2.22 (t, J ) 7.5
Hz, 2H); 13C NMR (CDCl3) δ 170.56, 136.03, 47.72, 34.77, 31.68,
27.43, 23.61; ESI (m/z) 180.3 (M+ - H2O); HRMS (m/z; MH+)
calcd for C9H15N2O3, 199.1077; found, 199.1075.
3-(1-Allyl-2-oxo-1,2,5,6-tetrahydro-pyridin-3-yl)-N-hydroxy-
propionamide (21b). The same procedures are followed as for 13a.
Compound 20b (44 mg, 0.20 mmol) was used and 21b (20 mg,
45%, oil) was obtained by flash chromatography (methanol/
1
chloroform, 1/19). H NMR (CDCl3) δ 10.08 (br s, 1H), 8.76 (br
s, 1H), 6.38 (br t, 1H), 5.75-5.67 (m, 1H), 5.17 (d, J ) 5.4 Hz,
1H), 5.12 (s, 1H), 3.98 (d, J ) 5.4 Hz, 2H), 3.30 (t, J ) 7.0 Hz,
2H), 2.54 (s, 2H), 2.36-2.28 (m, 4H); 13C NMR (CDCl3) δ 170.17,
165.21, 136.15, 133.46, 132.91, 117.38, 49.25, 44.77, 32.72, 27.12,
23.74; ESI (m/z) 247.4 (MNa+); HRMS (m/z; MH+) calcd for
C11H17N2O3, 225.1234; found, 225.1232.
3-[1-(2,4-Dimethoxy-benzyl)-2-oxo-1,2,5,6-tetrahydro-pyridin-
3-yl]-N-hydroxy-propionamide (21c). The same procedures are
followed as for 13a. Compound 18 (46 mg, 0.14 mmol) was used
and 21c (32 mg, 73%, oil) was obtained by flash chromatography
(methanol/chloroform, 1/19). 1H NMR (CDCl3) δ 7.12 (d, J ) 9.0
Hz, 1H), 6.425-6.33 (m, 3H), 4.51 (s, 2H), 3.75 (s, 3H), 3.74 (s,
3H), 3.27 (t, J ) 6.9 Hz, 2H), 2.55 (m, 2H), 2.38 (m, 2H), 2.22
(m, 2H); 13C NMR (CDCl3) δ 170.1, 165.4, 160.2, 158.5, 135.8,
133.5, 130.4, 117.5, 104.2, 98.3, 55.3, 44.9, 44.6, 32.8, 27.1, 23.8;
ESI (m/z) 357.5 (MNa+); HRMS (m/z; MH+) calcd for C17H23N2O5,
335.1602; found, 335.1609.
Acknowledgment. This work was supported by a grant from
KRIBB Research Initiative program and Grant 0405-NS01-
0704-0001 of the Korean Health 21 R&D Project, Ministry of
Health and Welfare, the Republic of Korea.
JM0613828