Design, synthesis, and biological evaluation of novel dual inhibitors of secretory phospholipase A2 and sphingomyelin synthase

Article abstract of DOI:10.1002/cjoc.201300079

A novel series of eight SMS and sPLA2 dual inhibitors containing indole and α-amino cyanide fragments of different length and substitution position was synthesized and evaluated by three different in vitro assays. Biological evaluation showed that all compounds provided inhibitory effects against SMS (about 50% inhibition at 100 μmol/L) and sPLA2 (14-32 μmol/L). All the compounds had the SMS activity better than the positive control compound D609 in SMS2 homogenate, with compounds 5b and 5e ideal for liver homogenate and SMS2 high expression cell homogenate, respectively. The secretory phospholipase A2 (sPLA2) and sphingomyelin (SMS) are the key enzymes to atherosclerosis. In this paper, we combined the single sPLA2 and SMS inhibitors with a carbon chain to get a series of dual inhibitors. The biological evaluation showed these compounds had the moderate inhibition against both enzymes. Copyright

Full text of DOI:10.1002/cjoc.201300079

FULL PAPER
DOI: 10.1002/cjoc.201300079
Design, Synthesis, and Biological Evaluation of Novel Dual
Inhibitors of Secretory Phospholipase A2 and
Sphingomyelin Synthase
Xing Gao, Haojun Gong, Peng Men, Lu Zhou,* and Deyong Ye*
Department of Medicinal Chemistry, School of Pharmacy, Fudan University, Shanghai 201203, China
A novel series of eight SMS and sPLA2 dual inhibitors containing indole and α-amino cyanide fragments of
different length and substitution position was synthesized and evaluated by three different in vitro assays. Biological
evaluation showed that all compounds provided inhibitory effects against SMS (about 50% inhibition at 100 μmol/L)
and sPLA2 (14－32 μmol/L). All the compounds had the SMS activity better than the positive control compound
D609 in SMS2 homogenate, with compounds 5b and 5e ideal for liver homogenate and SMS2 high expression cell
homogenate, respectively.
Keywords atherosclerosis, secretory phospholipase A2 (sPLA2), sphingomyelin synthase (SMS), multi-target
drug design, dual inhibitors
Introduction
ploys ceramide and phosphatidylcholine (PC) as sub-
strates to generate SM and diacylglycerol (DAG).[11]
There are two main isoforms of the enzyme, SMS1 and
SMS2.[12] SMS activity links directly to cell functions
which may have impact on disease development.[13]
Some evidence indicates that high plasma SM levels are
closely related to the development of AS. The low-den-
sity lipoprotein (LDL) extracted from human athero-
sclerotic lesions is much richer in SM than the LDL
from plasma,[14,15] indicating that the distribution and
synthesis of SM increase in these lesions. Native lipo-
protein SM also promotes the SM aggregation in arter-
ies.[16] Further clinical research confirmed these results,
pointing out that plasma SM levels are a risk factor for
AS.[17] Accordingly SMS has begun to attract more and
more attention as a potential anti-AS target. However,
the crystal structure of SMS has not been resolved and
few associated inhibitors have been reported.
Secretory phospholipase A2 (sPLA2), part of the
group II non-pancreatic family of phospholipase A2, is a
14-kDa, calcium-dependent enzyme synthesized by
many different mammalian tissues including vascular
smooth muscle cells,[18] platelets,[19] neutrophils,[20] liver
cells,[21] spleen[22], and a number of other mesenchymal
tissues. It hydrolyzes phospholipids at the sn-2 position
releasing some precursors of important mediators of
inflammation and inflammatory response.[23,24] Based on
the inflammatory nature of AS, Menschikowski et al.
first reported the expression of sPLA2 in atherosclerotic
plaques and the possible role of sPLA2 in the process of
Atherosclerosis (AS)—the principal cause of myo-
cardial infarction, coronary artery disease, stroke, and
gangrene of the extremities—is a chronic inflammatory
disease related to dysbolism of lipids and responsible
for most mortality in westernized societies and several
developing countries.[1,2] Many cellular and molecular
level experiments indicate AS is not only related to lipid
dysbolism and inflammation,[3] but also to some other
pathological changes such as oxidative stress,[4] immu-
nomodulation,[5] thrombosis,[6] diabetes[7] and metabolic
syndrome.[8] Nevertheless, these results cannot explain
the whole pathogenic process of AS independently. Tra-
ditional anti-AS drugs are mainly lipid-lowering drugs,
including statins and fibrates. Although they play im-
portant roles in lowering LDL-cholesterol levels by in-
hibiting a single target, the incidence of AS remains
high. This shows that LDL-cholesterol level is not the
only indicator of AS. Since AS is a multi-factor &
multi-target disease, single target therapy has certain
limitations and multi-target treatment may be more
suitable for this complex disease.
Sphingomyelin (SM) is a major component of cellu-
lar membranes in a wide range of organisms from pro-
tozoa to mammals. It plays a role in the formation of
lipid rafts, specialized areas in cellular membranes with
important functions in intracellular signal transduction[9]
and membrane trafficking.[10] Sphingomyelin synthase
(SMS) is the last enzyme for SM biosynthesis. It em-
*
E-mail: zhoulu@fudan.edu.cn (L. Zhou); dyye@shmu.edu.cn (D. Ye); Tel.: 0086-021-51980117; Fax: 0086-021-51980125
Received January 29, 2013; accepted March 1, 2013; published online April 3, 2013.
Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/cjoc.201300079 or from the author.
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Novel Dual Inhibitors of Secretory Phospholipase A2 and Sphingomyelin Synthase
atherogenesis.[25,26] The findings of Bobryshev et al.[27]
is also consistent with these conclusions. sPLA2 can also
directly induce the aggregation and fusion of LDL par-
ticles.[28] For both reasons, sPLA2 has been identified as
an important risk factor of cardiovascular disease and a
predictor of AS.
the amide NH group bound to the residue His 48. Sub-
stition of the 1-benzyl group at the 2-position retained
or improved the activity,[39] allowing the flexibility to
introduce lipophilic groups.
D609 (Figure 1B), the only reported small-molecular
compound as SMS inhibitor, was not suitable to use in
our dual inhibitors because of its instability and low
activity (IC50 ＝375 μnol/L).[40,41] Instead we chose
Dy105 (IC50＝12 μmol/L, Figure 1C), a compound de-
veloped in our lab. The structure of α-amino cyanide in
molecule Dy105 was crucial to provide SMS inhibitory
activity. So we retained this group and designed a linker
to attach to the sPLA2 pharmacophore to form a series
of dual inhibitors (Figure 1). Compounds 5a－5f had
the linker containing carbon atoms between 2 to 5
(Scheme 1). Because the molecular weights of 5a－5f
were greater than 500, the simplified structures (com-
pounds 8a, 8b) with lower molecular weight were de-
signed as better drug models (Scheme 2).
The reported research mainly focused on the selec-
tive inhibitors against the single enzymes sPLA2 or
SMS. The first small-molecule sPLA2 inhibitors were
reported in 1980s, quickly followed by many more.[29,30]
But the research of SMS inhibitors was few.[31] Multi-
target drugs developed at selected molecular targets in-
volved in the pathogenic pathway have received more
attention in recent years, especially in complex diseases
like cancer, AIDs, and atherosclerosis,[32,33] and recent
research indicated that SMS and PLA2 had interaction in
membrane binding.[34] Generally, there are two ways to
design multitarget inhibitors－fusion and linking. Wei
and coworkers designed multitarget anti-inflammatory
inhibitors by combining molecular docking with com-
mon pharmacophore matching.[35] Cashman and co-
workers synthesized dual inhibitors for depression ther-
apy by linking a novel selective serotonin reuptake in-
hibitor (SSRI) to a PDE4 inhibitor.[36] Both works
showed advantage over simple additive effects of co-
administered agents in whole blood assays or animal
experiments. Here we used a linking strategy to link a
known sPLA2 inhibitor to newly developed SMS in-
hibitor by varying the linker length and position.
Experimental
Chemistry
The general method of synthesis for the compounds
5a－5f is depicted in Scheme 1. Indomethacin 1 was
used as starting material to synthesize compound 2 by a
one-pot method.[42] Benzylation of compound 2 with
substituted benzyl chlorides gave compounds 3a－3d.
The etherification and Strecker reaction subsequently
aforded target compounds 5a－5f. The synthesis route
of compounds 8a, 8b, as shown in Scheme 2, was simi-
lar to that of compounds 5a－5f.
Bach et al.[37,38] first reported a series of indole-3-
acetamides and related compounds as potential sPLA2
inhibitors (Figure 1A). X-ray crystallographic studies of
the crystalline complex between inhibitors and enzymes
showed that the indoyl group of the inhibitor was lo-
cated in the active site of the enzyme, while the amide
carbonyl group with the calcium in the active site and
Biological evaluation
Materials The reagents 1,2-dimyristoyl-sn-
glycero-3-phosphocholine (DMPC) and 6-((N-(7-
nitrobenz-2-oxa-1,3-diazol-4-yl)amio)hexanoyl)-
sphingosine (C6-NBD-Ceramide) were purchased from
Santa Cruz (USA), N-(N-(7-nitro-2,1,3-benzoxadiazol-
4-yl)-epsilon-aminohexanoyl)sphingosylphosphoryl
choline (C6-NBD-SM) was obtained from Sigma-
Aldrich (USA). ICR rats were provided by Sino-British
S1PPR/BK Lab Animal Ltd and were maintained at
standard laboratory diet and water. High expression
SMS2 enzyme was supplied by Institutes of Biomedical
Sciences, Fudan University and the homogenate was
prepared with the appropriate concentration. The sPLA2
enzyme was supplied by College of Chemistry and Mo-
lecular Engineering, Peking University.
O
NH2
O
CN
N
N
O
SHK
H
O
C
S
B
A
O
NH2
O
N
Enzyme-based assay of SMS in high expression
SMS2 homogenate[43]
NC
NH
Linker
O
280 µL of enzyme-test buffer containing 10 mmol/L
4-(2-hydroxyethyl) piperazine-1-ethanesulfonic acid
(HEPES) (pH 7.4), 3 mmol/L MnCl2 and 0.3% bovine
serum albumin (BSA) in water (Milli-Q), 4 µL of SMS2
homogenates and 10 µL of DMSO with various concen-
trations of test compounds was mixed and preincubated
O
sPLA2 inhibitor
SMS inhibitor
Figure 1 (A) sPLA2 inhibitor indole-3-acetamide; (B) the re-
ported SMS inhibitor D609; (C) SMS inhibitor Dy105; below:
the designed SMS and sPLA2 dual inhibitors.
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Scheme 1 Synthesis route of dual inhibitors 5
O
O
OH
NH2
O
O
Cl
n Cl
O
NH2
O
a
N
O
N
b
O
N
H
O
n Cl
2
Cl
3
1
O
O
NH2
CHO
NH2
NH2
OH
O
O
NC
N
CHO
N
NH
d
c
A
A
O
n O
O
nO
4
5
Reagents and conditions: (a) ClCOOC2H5, NMM, NH3H2O, NaOH, THF, r.t.; (b) NaH, DMF, r.t.; (c) K2CO3, MeCN, reflux; (d)TMSCN, I2, MeCN, r.t.
Scheme 2 Synthesis route of dual inhibitors 8
O
O
O
CHO
OH
NH2
NH2
NH2
O
O
OHC
O
e
N
A
N
H
f
N
O
Cl
7
6
2
O
NH2
NH2
H
N
NC
O
N
g
A
O
8
Reagents and conditions: (e) Cl(CH2)4Cl, NaH, DMF, r.t.; (f) K2CO3, MeCN, reflux; (g) TMSCN, I2, MeCN, r.t.
at 37 ℃ for 30 min. Then 6 µL of DMSO containing
2.32 mmol/L C6-NBD-Ceramide and 40 mmol/L
DMPC was added to initiate the enzymatic reactions
and incubated at 37 ℃ for an additional 2 h. The reac-
tions were terminated by the addition of 600 µL of
ethanol. The mixture was then centrifuged and the su-
pernatant was gathered for HPLC analysis. Extracts (20
µL) from the supernatant were analyzed for the forma-
tion of C6-NBD-SM by HPLC using the reversed-phase
C18 column and an isocratic elution with methanol/
water/trifluoroacetic acid [88∶12∶0.1 (V∶V∶V)].
The flow rate of the mobile phase was 1.0 mL/min. De-
tection was accomplished with a fluorescence spectro-
photometer (λexcited＝475 nm, λemssion＝525 nm). The
retention time of C6-NBD-SM and C6-NBD-ceramide
was 13.6 min and 14.8 min respectively. A comparison
of the formation of C6-NBD-SM in the test group with
the control group yielded the inhibition rate. The per-
centage inhibition rate could be calculated by the equa-
tion: inhibition%＝1-S/S0, where S is the peak area of
C6-NBD-SM in test group and S0 is the peak area of
C6-NBD-SM in control group.
Enzyme-based assay of SMS in liver homogenate
The ICR rats livers were dissected, weighed and then
homogenized with homogenate buffer (pH 7.4) con-
taining 0.25 mol/L sucrose, 50 mmol/L Tris-HCl and 1
mmol/L EDTA. The homogenates were subjected to
centrifugation and the supernatant was used for bio-
logical assays. In this assay system, 244 µL of enzyme-
test buffer, 40 µL of liver homogenates (contained 2 mg
of total proteins) and 10 µL of DMSO with various
concentrations of test compounds were mixed and pre-
incubated at 37 ℃ for 30 min. Then 6 µL of DMSO
containing 2.32 mmol/L of C6-NBD-ceramide and 40
mmol/L (DMPC) were added and incubated at 37 ℃
for an additional 2 h. The following operations includ-
ing the termination of the enzyme catalytic reactions,
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Novel Dual Inhibitors of Secretory Phospholipase A2 and Sphingomyelin Synthase
extraction and determination of the products were the
same as shown above.
excitation at 377 nm and emission at 470 nm. Fluores-
cent signals were monitored by a kinetics modeling
program. Reported IC50 values, determined by plotting
concentration-velocity curves, are the mean of at least
three separate experiments. The initial velocities of the
change in fluorescence intensity with various inhibitor
concentrations were used to calculate the IC50 values
using the following equation: v0/v＝1＋[I]/IC50, where
v0 is the velocity in control group, v is the velocity in
test group, and [I] is the concentrations of inhibitors.
[44]
Enzyme-based assay of sPLA2
The synthesized gene of hnp-sPLA2 was ligated to a
pET21a vector and transformed into Escherichia coli
cells (BL21(DE3) strain). The purification included a
refolding process.[34] The assay was carried out in a
96-well plate utilizing a multiwell fluorometer. All so-
lutions were prepared with high purity water. The buffer
used in all the experiments was 50 mmol/L Tris-HCl
(pH 8.0), 100 mmol/L NaCl, 10 μmol/L ANS, 10
mmol/L CaCl2. The substrate was 2 mmol/L DMPC-
deoxycholic acid (DCA) buffer solution, which was
sonicated for 1 min to form a micelle suspension. All
the samples were dissolved in DMSO. A 200 μL reac-
tion solution contained 15 nmol/L sPLA2, 20 μL of sub-
strate, 0.1 mg/mL BSA, and 10 μL of inhibitor solution.
The hydrolysis reaction was monitored with an “interfa-
cial probe”, called ANS. The reaction was monitored by
Results and Discussion
In vitro activities of compounds 5a－5f and 8a, 8b
against SMS and sPLA2 were evaluated (Table 1). SMS
activity was evaluated at the compound concentration of
100 μmol/L. From Table 1, we found compounds 5d
and 5b had the best activities in liver homogenate and
SMS2 cell homogenate, respectively. Some compounds
had the SMS2 activity significantly better than liver
Table 1 In vitro inhibitory activity against SMS and sPLA2
Inhibition% in liver
homogenatea
Inhibition% in SMS2
homogenatea
IC50 in sPLA2/
Structure
(μmol•L-1)
O
NH2
O
N
36%
29%
38%
48%
51%
42%
31.7
H
N
NC
O
O
O
5a
NH2
O
H
N
20.0
NC
N
O
O
O
5b
NH2
O
N
18.3
H
NC
N
O
O
5c
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Continued
Inhibition% in liver
homogenatea
Inhibition% in SMS2
homogenatea
IC50 in sPLA2/
(μmol•L-1)
Structure
O
NH2
O
N
O
H
43%
41%
14.1
NC
N
O
5d
O
NH2
O
N
O
48%
43%
15.7
NH
CN
O
5e
O
NH2
O
N
O
40%
40%
14.7
O
NH
CN
5f
O
NH2
HN
O
26%
46%
27.3
NC
N
O
8a
O
H
NC
N
NH2
O
29%
NDb
ND
49%
22%
78%
29.5
ND
ND
N
O
8b
O
SHK
S
D609
CN
N
H
O
Dy105
a The compound concentration was 100 μmol/L. b no data.
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Novel Dual Inhibitors of Secretory Phospholipase A2 and Sphingomyelin Synthase
Acknowledgement
homogenate activity (compounds 5a, 5b, 8a, 8b). Most
compounds were better than D609 (not shown in Table
1, about 25%), although the activity was lower than lead
compound Dy105. In the sPLA2 evaluation system, all
the compounds showed inhibitory activities in the mi-
cromolar level, in which compound 5d had the best ac-
tivity, and most compounds had the better activity than
diphenyl compounds (8a, 8b).
The work was funded by the National Natural Sci-
ence Foundation of China (Nos. 30973641, 20902013),
Specialized Research Fund for the Doctoral Program of
Higher Education (No. 20090071110054), Chinese
Ministry of Education, the open grant of the State Key
Laboratory of Bio-organic and Natural Products Chem-
istry, CAS, and open grant of Institute of Bioscience,
Fudan University. The enzymes were supplied by
Yan-hui Xu group, Institutes of Biomedical Sciences,
Fudan University and Lu-hua Lai group, College of
Chemistry and Molecular Engineering, Peking Univer-
sity.
All the compounds remained active against sPLA2
with different intensities based on the chain length and
substituted position in the target compounds. When the
linker contained five carbon atoms (compounds 5d－5f),
the compounds offered better inhibitory activity against
sPLA2. The ortho-substituted 5d and 8a were slightly
suprior to the meta- and para-substituted 5e－5f and 8b.
The results showed that SMS inhibitor fragment was not
fitted for sPLA2 pocket and longer linker length would
weaken the counter effect. Although sPLA2 and SMS
shared the same fragment of substrate, phosphotidyl
choline, we could assume that unknown substrate bind-
ing model of SMS was different from that of sPLA2.
Most of the compounds showed moderate activity
against SMS in liver homogenate and SMS2 expressed
insect cell homogenate. Compounds 5d and 5b had the
best activities in liver homogenate and SMS2 cell ho-
mogenate, respectively. Comparison of 5d－5f and 8a,
8b, showed that the different substituted position of
α-amino cyanide impacted the SMS activities. The
meta-substituted 5e provided better inhibition than the
ortho- and para-substituted 5d and 5f, both in liver ho-
mogenate and SMS2 cell homogenate. The meta-sub-
stituted 8b was also slightly more active than the ortho-
substituted 8a. Compounds 5b and 8a, 8b gave the best
results in these two systems. Since there are two main
SMS isoforms in liver homogenate (SMS1 and SMS2),
the higher inhibition of SMS2 over SMS1 in liver ho-
mogenate showed that the compounds might have high
selectivity against SMS2. This hypothesis needs further
confirmation due to the absence of high expression
SMS1 cells. Based on the present results, we could con-
clude that meta-substitute and shorter linker length (3
carbons for 5c) would enhance the selectivity against
SMS2.
References
[1] Ross, R. Nature 1993, 362, 801.
[2] Ross, R. N. Engl. J. Med. 1999, 340, 115.
[3] Libby, P. Nature 2002, 420, 868.
[4] Alexander, R. W. Hypertension 1995, 25, 155.
[5] Nicoletti, A.; Caligiuri, G.; Hansson, G. K. J. Intern. Med. 2000, 247,
397.
[6] Prandoni, P.; Bilora, F.; Marchiori, A.; Bernardi, E.; Petrobelli, F.;
Lensing, A. W. A.; Prins, M. H.; Girolami, A. N. Engl. J. Med. 2003,
348, 1435.
[7] Beckman, J. A.; Creager, M. A.; Libby, P. JAMA-J. Am. Med. Assoc.
2002, 287, 2570.
[8] Grundy, S. M. Circulation 2002, 105, 2696.
[9] Simons, K.; Toomre, D. Nat. Rev. Mol. Cell. Biol. 2000, 1, 31.
[10] Holthuis, J. C. M.; Pomorski, T.; Raggers, R. J.; Sprong, H.; Van
Meer, G. Physiol. Rev. 2001, 81, 1689.
[11] Voelker, D. R.; Kennedy, E. P. Biochemistry 1982, 21, 2753.
[12] Kidani, Y.; Ohshima, K.; Sakai, H.; Kohno, T.; Baba, A.; Hattori, M.
Biochem. Biophys. Res. Commun. 2012, 417, 1014.
[13] Zhao, Y. R.; Dong, J. B.; Li, Y.; Wu, M. P. Life Sci. 2012, 90, 867.
[14] Guyton, J. R.; Klemp, K. F. Arterioscler. Thromb. Vasc. Biol. 1996,
16, 4.
[15] Schissel, S. L.; Jiang, X. C.; Tweedie-Hardman, J.; Jeong, T. S.;
Camejo, E. H.; Najib, J.; Rapp, J. H.; Williams, K. J.; Tabas, I. J.
Biol. Chem. 1998, 273, 2738.
[16] Jiang, X. C.; Paultre, F.; Pearson, T. A.; Reed, R. G.; Francis, C. K.;
Lin, M.; Berglund, L.; Tall, A. R. Arterioscler. Thromb.Vasc. Biol.
2000, 20, 2614.
[17] Schlitt, A.; Blankenberg, S.; Yan, D.; von Gizycki, H.; Buerke, M.;
Werdan, K.; Bickel, C.; Lackner, K. J.; Meyer, J.; Rupprecht, H. J.;
Jiang, X. C. Nutr. Metab. 2006, 3, 5.
[18] Nakano, T.; Arita, H. FEBS Lett. 1990, 273, 23.
[19] Kramer, R. M.; Hession, C.; Johansen, B.; Hayes, G.; McGray, P.;
Chow, E. P.; Tizard, R.; Pepinsky, R. B. J. Biol. Chem. 1989, 264,
5768.
[20] Forst, S.; Weiss, J.; Elsbach, P.; Maraganore, J. M.; Reardon, I.;
Heinrikson, R. L. Biochemistry 1986, 25, 8381.
[21] Aarsman, A. J.; de Jong, J. G.; Arnoldussen, E.; Neys, F. W.; van
Wassenaar, P. D.; Van den Bosch, H. J. Biol. Chem. 1989, 264,
10008.
[22] Ono, T.; Tojo, H.; Kuramitsu, S.; Kagamiyama, H.; Okamoto, M. J.
Biol. Chem. 1988, 263, 5732.
[23] Murakami, M.; Taketomi, Y.; Miki, Y.; Sato, H.; Hirabayashi, T.;
Yamamoto, K. Prog. Lipid Res. 2011, 50, 152.
[24] Dennis, E. A.; Cao, J.; Hsu, Y. H.; Magrioti, V.; Kokotos, G. Chem.
Rev. 2011, 111, 6130.
[25] Menschikowski, M.; Kasper, M.; Lattke, P.; Schiering, A.; Schiefer,
S.; Stockinger, H.; Jaross, W. Atherosclerosis 1995, 118, 173.
[26] Schiering, A.; Menschikowski, M.; Mueller, E.; Jaross, W. Athero-
Conclusions
In summary, we have designed and synthesized a se-
ries of novel SMS and sPLA2 dual inhibitors with mod-
erate inhibitory activity using a linking strategy. Some
compounds showed better SMS inhibitory activities
than D609 and possessed selectivity against SMS2 iso-
form. Among these compounds, 5b and 5e were ideal in
liver homogenate and SMS2 cell homogenate, respec-
tively. The activity against sPLA2 was only slightly
decreased compared to the leading compound. Further
studies will focus on developing more dual inhibitors
and testing by in vivo evaluations.
Chin. J. Chem. 2013, 31, 1164—1170
© 2013 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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Gao et al.
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sclerosis 1999, 144, 73.
[37] Dillard, R. D.; Bach, N. J.; Draheim, S. E.; Berry, D. R.; Carlson, D.
G.; Chirgadze, N. Y.; Clawson, D. K.; Hartley, L. W.; Johnson, L.
M.; Jones, N. D.; McKinney, E. R.; Mihelich, E. D.; Olkowski, J. L.;
Schevitz, R. W.; Smith, A. C. J. Med. Chem. 1996, 39, 5119.
[38] Dillard, R. D.; Bach, N. J.; Draheim, S. E.; Berry, D. R.; Carlson, D.
G.; Chirgadze, N. Y.; Clawson, D. K.; Hartley, L. W.; Johnson, L.
M.; Jones, N. D.; McKinney, E. R.; Mihelich, E. D.; Olkowski, J. L.;
Schevitz, R. W.; Smith, A. C. J. Med. Chem. 1996, 39, 5137.
[39] Zhou, L.; Fang, C.; Wei, P.; Liu, S. Y.; Liu, Y.; Lai, L. H. J. Med.
Chem. 2008, 51, 3360.
[27] Bobryshev, Y. V.; Crozier, J. A.; Lord, R. S. A.; Tran, D.; Jamal, O.
S.; Pärsson, H. N.; Scott, K. F. Atherosclerosis 1996, 127, 283.
[28] Hakala, J. K.; Oorni, K.; Pentikainen, M. O.; Hurt-Camejo, E.;
Kovanen, P. T. Arterioscler. Thromb. Vasc. Biol. 2001, 21, 1053.
[29] Magrioti, V.; Kokotos, G. Expert Opin. Ther. Pat. 2013, 23, 333.
[30] Mouchlis, V. D.; Barbayianni, E.; Mavromoustakos, T. M.; Kokotos,
G. Curr. Med. Chem. 2011, 18, 2566.
[31] Delgado, A.; Casas, J.; Llebaria, A.; Abad, J. L.; Fabrias, G. Bio-
chim. Biophys. Acta-Biomembranes 2006, 1758, 1957.
[32] Morphy, R.; Rankovic, Z. Drug Discov. Today 2007, 12, 156.
[33] Hopkins, A. L. Nat. Chem. Biol. 2008, 4, 682.
[40] Amtmann, E. Drugs Exp. Clin. Res. 1996, 22, 287.
[41] Gonzalez-Roura, A.; Casas, J.; Llebaria, A. Lipids 2002, 37, 401.
[42] Liu, Y.; Han, X. F.; Huang, C. K.; Hao, X.; Lai, L. H. Bioorg. Med.
Chem. Lett. 2005, 15, 4540.
[34] Chiou, Y. L.; Lin, S. R.; Chang, L. S. Chem. Phys. Lipids 2011, 164,
378.
[35] Wei, D. G.; Jiang, X. L.; Zhou, L.; Chen, J.; Chen, Z.; He, C.; Yang,
K.; Liu, Y.; Pei, J. F.; Lai, L. H. J. Med. Chem. 2008, 51, 7882.
[36] Cashman, J. R.; Voelker, T.; Zhang, H.; O'Donnell, J. M. J. Med.
Chem. 2009, 52, 1530.
[43] Deng, X. D.; Sun, H.; Gao, X.; Gong, H. J.; Lu, W. B.; Chu, Y.;
Zhou, L.; Ye, D. Y. Anal. Lett. 2012, 45, 1581.
[44] Huang, C. K.; Zhou, L.; Liu, Y.; Lai, L. H. Anal. Biochem. 2006,
351, 11.
(Lu, Y.)
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