Ezetimibe analogs with a reorganized azetidinone ring: Design, synthesis, and evaluation of cholesterol absorption inhibitions

Article abstract of DOI:10.1016/j.bmcl.2006.09.078

The underlying principle of drug design in this paper is that the maximum retention of the functional groups that exist in the marketed drug would provide a higher probability for comparable safety while the conformational changes in the newly created ana

Full text of DOI:10.1016/j.bmcl.2006.09.078

Bioorganic & Medicinal Chemistry Letters 17 (2007) 101–104
Ezetimibe analogs with a reorganized azetidinone ring: Design,
synthesis, and evaluation of cholesterol absorption inhibitions
Xianxiu Xu,a,b Renzhong Fu,a Jin Chen,a Shengwu Chenc and Xu Baia,b,
*
aThe Center for Combinatorial Chemistry and Drug Discovery, Jilin University, 75 Haiwai Street, Changchun, Jilin 130012, PR China
bChangchun Discovery Sciences, Ltd, 75 Haiwai Street, Changchun, Jilin 130012, PR China
cJilin Natural Pharmatech Co., Ltd, Changchun, Jilin 130012, PR China
Received 31 May 2006; revised 23 September 2006; accepted 28 September 2006
Available online 30 September 2006
Abstract—The underlying principle of drug design in this paper is that the maximum retention of the functional groups that exist in
the marketed drug would provide a higher probability for comparable safety while the conformational changes in the newly created
analogs should not constitute a significant structural variation to adversely affect biological activity. Four individual isomers of
backbone re-organized ezetimibe analogs were designed and synthesized. Their effects on the cholesterol levels in rat serum were
evaluated by a high-cholesterol and high-fat diet feeding experiment. All the new analogs showed significant effect in lowering
the levels of total cholesterol in serum.
Ó 2006 Elsevier Ltd. All rights reserved.
Atherosclerotic coronary heart disease (CHD) remains a
major concern in healthcare due to its high morbidity
and mortality.1 Lowering the level of cholesterol in blood
has been shown to be an effective way to treat and pre-
vent CHD.2 There are two recognized sources of choles-
terol in the serum: biosynthesis in the liver and
absorption of dietary cholesterol in the small intestine.3,4
Statins have been prescribed as the predominant class of
cholesterol-lowering agents since 1980s.5 They inhibit
3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA)
reductase, the catalyst of the rate-limiting step of choles-
terol biosynthesis in liver.6 Recently, ezetimibe (Fig. 1,
1), a blocker of intestinal sources of cholesterol, has
become an increasingly important choice for reducing
serum cholesterol level. It inhibits the absorption of
dietary or recycled cholesterol in the intestine and can
be used either alone or in combination with a statin.
Thus far, ezetimibe is the only marketed example of this
new class of anti-cholesterolemia drugs.7–9
molecular target of ezetimibe have been disclosed. Evi-
dence suggests that the compound interacts with the
Niemann Pick C1-like 1 (NPC1L1) protein which plays
a critical role in cholesterol transporting through the
intestine wall.10 Despite the absence of a molecular tar-
get the SAR studies on azetidinone structures have been
thoroughly investigated. Very recently, it has been dis-
closed that the azetidinone backbone possibly plays
the role of a structural scaffold to appropriately position
the required ring substituents since the mesylated azeti-
dine analog (Fig. 1, 2) showed comparable bioactivity
by an in vitro assay.11 This study could further encour-
OMs
OH
OH
OH
F
N
N
F
O
F
F
1
2
Ezetimibe was discovered through in vivo screening uti-
lizing an animal-feeding model to identify active com-
pounds. Recently, many reports on the potential
OH
F
OH
F
N
O
Keywords: Backbone re-organized ezetimibe analogs; Azetidin-2-one;
Cholesterol absorption inhibition.
3
*
Figure 1. The structures of ezetimibe 1, its azetidine analog 2, and re-
organized analogs 3.
Corresponding author. Tel.: +86 431 5188955; fax: +86 431
5188900; e-mail: xbai@jlu.edu.cn
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.09.078

102
X. Xu et al. / Bioorg. Med. Chem. Lett. 17 (2007) 101–104
Cl
OBn
OBn
O
F
MeO
R
R
O
4
N
N
b
+
O
H2N
CHO
F
F
N
a
8a
, R= (S)-1-[4'-MeO
-Ph-(CH3)CH]
8b,
R = (S)-1-[4'-MeO
-Ph-(CH )CH]
+
3
c
c
OBn
OMe
5
9a
9b
, R = H
, R = H
OBn
6
7
Scheme 1. Synthesis of trans-azetidinone fragments. Reagents and conditions: (a) toulene, reflux; (b) Et3N, toluene, 47% for 8a, 26% for 8b in two
steps; (c) CAN, 53% for 9a, 55% for 9b.
age the search for new cholesterol absorption inhibitors
away from the traditional azetidinone structures. We
have been interested in discovering compounds that
inhibit cholesterol absorption by exploring ezetimibe
analogs in which the substituents are regiochemically
rearranged on the azetidinone scaffold (Fig. 1, 3). This
paper discloses results of the design, synthesis, and in vi-
vo evaluation of cholesterol absorption inhibition.
tion, both 3S and 3R forms often demonstrate compara-
ble activity without consistent preference. These SAR
results, in particular the last point regarding the stereo-
chemistry at 3-position prompted us to consider swap-
ping the substituents between the N-position and the
3-position. The underlying principle is that the maxi-
mum retention of the functional groups that exist in
the marketed drug would provide a higher probability
for comparable safety while the conformational changes
in the newly created backbone should not constitute a
significant structural variation to adversely affect biolog-
ical activity. As a result, the re-organized ezetimibe ana-
log 3 (Fig. 1) was devised.
As reported in the literature, the pharmacophore of
ezetimibe has the following requirements: (1) azetidi-
none as the required backbone; (2) N-phenyl is required,
but could tolerate a range of substituents; (3) the phenyl
tethered with three-carbon chain at 3-position is neces-
sary for activity; (4) p-hydroxyl or p-methoxyphenyl
at 4-position is a necessity; (5) unlike the chiral center
at C-4 which shows a clear preference for S configura-
Four individual isomers of 3 with trans configuration of
the azetidinone backbone were prepared according to
Schemes 1–3. Scheme 1 depicts the synthesis of trans-
azetidinone fragments in optically active forms. Com-
mercially available S-1-(4-methoxyphenyl) ethanamine
5 was treated with 4-(benzyloxy)benzaldehyde 6 in
refluxing toluene to give imine 7. Reactions of acid chlo-
ride 4 and imine 7 using Et3N as the base in refluxing
toluene led to diastereoisomers 8a and 8b with a ratio
of 2 to 1 in 73% combined yield over two steps. Diaste-
reoisomers 8a and 8b were readily separated by flash
chromatography. The oxidative removal of the chiral
benzylamine group by ceric ammonium nitrate (CAN)
gave more than 50% of the desired products 9.
Ph
H
OH
Ph
O
O
X
N
11a
B
F
Cl
.
BH3 SMe2
85%
F
12a, X = Cl
13a, X = I
10
NaI
95%
Ph
H
OH
X
Ph
O
N
11b
B
F
10
12b
.
, X = Cl
BH3 SMe2
87%
The optically active alcohol fragments 13a and 13b were
prepared by a modified literature procedure12 as depict-
ed in Scheme 2. Both optically pure enantiomers of
CBS catalyst 11 were obtained by Corey’s procedure.13
NaI
93%
13b, X = I
Scheme 2. Synthesis of alcohol fragments.
OR
OH
OBn
F
N
OH
I
KOH
O
HN
DMSO/THF
90%
+
F
O
F
14a, R = Bn
3a, R = H
Pd/C, H2
90%
F
9a
13a
OH
F
OH
F
OH
OH
OH
OH
F
F
N
O
N
F
N
O
O
3c
3d
3b
F
Scheme 3. Synthesis of final targets.

Table 1. Physical properties of backbone re-organized ezetimibe analogs
3a
3b
3c
3d
1H NMR (300 MHz, CDCl3)
1.85–1.92 (m, 2H), 2.93–3.02 (m, 1H),
3.70–3.80 (m, 1H), 4.14 (d, 1H,
J = 2.1 Hz), 4.41 (d, 1H, J = 2.1 Hz),
4.69–4.73 (m, 1H), 6.89 (d, 2H,
J = 8.4 Hz), 6.97–7.07 (m, 4H), 7.18–
7.29 (m, 6H)
1.83–1.92 (m, 2H), 2.76 (d, 1H, OH,
J = 3.9 Hz), 3.16–3.25 (m, 1H), 3.45–
3.54 (m, 1H), 4.14 (d, 1H,
J = 2.1 Hz), 4.43 (d, 1H, J = 2.1 Hz),
4.74–4.79 (m, 1H), 5.43 (s, 1H, OH),
6.88 (d, 2H, J = 8.4 Hz), 6.98–7.07
(m, 4H), 7.21–7.31 (m, 6H)
37.86, 38.10, 63.18, 63.59, 70.14,
115.39 (d, J = 20.63 Hz), 116.21 (d,
J = 18.3 Hz), 116.33, 128.25 (d,
J = 8.03 Hz), 128.30, 128.74, 130.08
(d, J = 8.18 Hz), 132.51 (d,
J = 2.25 Hz), 142.40 (d, J = 2.25 Hz),
158.34, 161.79 (d, J = 241.5 Hz),
162.10 (d, J = 241.5 Hz), 168.18.
150–151.3
1.83–1.91 (m, 2H), 3.18–3.25 (m, 1H),
3.46–3.53 (m, 1H), 4.14 (d, 1H,
J = 2.1 Hz), 4.43 (d, 1H, J = 2.1 Hz),
4.75–4.79 (m, 1H), 6.87 (d, 2 H,
J = 8.1 Hz), 6.97–7.07 (m, 4H), 7.20–
7.33 (m, 6H)
1.86–1.93 (m, 2H), 2.94–3.02 (m, 1H),
3.09 (d, 1H, OH, J = 3.6 Hz), 3.71–
3.80 (m, 1H), 4.15 (d, 1H,
J = 2.1 Hz), 4.41 (d, 1H, J = 2.1 Hz),
4.69–4.75 (m, 1H), 5.69 (s, 1H, OH),
6.90 (d, 2H, J = 8.7 Hz), 6.98–7.08
(m, 4H), 7.19–7.30 (m, 6H)
13C NMR (75 MHz, DMSO-d6)
37.78, 37.86, 62.85, 63.72, 70.09,
115.41 (d, J = 21.23 Hz), 116.22 (d,
J = 20.03 Hz), 116.38, 128.21 (d,
J = 8.03 Hz), 128.22, 128.70, 130.08
(d, J = 8.03 Hz), 132.53 (d,
J = 2.85 Hz), 142.49 (d, J = 2.33 Hz),
158.34, 161.80 (d, J = 240.98 Hz),
162.11 (d, J = 241.58 Hz), 168.13
65.8–67.3
37.87, 38.10, 63.18, 63.59, 70.14,
115.40 (d, J = 20.63 Hz), 116.24 (d,
J = 21.23 Hz), 116.33, 128.25 (d,
J = 7.5 Hz), 128.30, 128.75, 130.09
(d, J = 8.03 Hz), 132.55 (d,
J = 2.85 Hz), 142.39 (d, J = 2.33 Hz),
158.34, 161.80 (d, J = 240.98 Hz),
162.10 (d, J = 242.10 Hz), 168.17
149–150.8
37.78, 37.85, 62.84, 63.71, 70.07,
115.40 (d, J = 20.55 Hz), 116.20 (d,
J = 19.5 Hz), 116.33, 128.21 (d,
J = 6.9 Hz), 128.25, 128.68, 130.08
(d, J = 8.03 Hz), 132.51, 142.48,
158.32, 161.79 (d, J = 240.38 Hz),
162.10 (d, J = 241.5 Hz), 168.10
Mp (°C)
67.6–68.5
392.2
3.037
MS [M+Hꢀ18]+
RT (min)
392.2
3.042
392.2
3.021
392.2
3.028
20
½aꢁD (c = 1, MeOH)
+90.2°
+128.2°
ꢀ127.4°
ꢀ90.2°

104
X. Xu et al. / Bioorg. Med. Chem. Lett. 17 (2007) 101–104
Table 2. Results of in vivo experiments16
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Entry
Animal groupa
TC or
Changes (%)b
LDL-cho or
Changes (%)b
1
2
3
4
5
6
7
High-cholesterol diet
Normal diet
ezetimibe
5.40 2.25
ꢀ62***,c
ꢀ51***
ꢀ26*
2.93 1.54
ꢀ59***
ꢀ47**
ꢀ22
3a
3b
3c
3d
ꢀ17
ꢀ5
8. (a) Clader, J. W. J. Med. Chem. 2004, 47, 1; (b) Burnett,
D. A. Curr. Med. Chem. 2004, 11, 1873; Clader, J. W.
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H. R., Jr. Bioorg. Med. Chem. 1998, 6, 1429.
ꢀ25*
ꢀ29*
ꢀ18
ꢀ35
***P < 0.001; **P < 0.01; *P < 0.05.
a 8–18 rats per group.
b Percentile was calculated by comparing to the one in animals fed by
high-cholesterol diets.
c Compared to high-cholesterol diet.
10. (a) Altmann, S. W.; Davis, H. R., Jr.; Zhu, L.-J.; Yao, X.;
Hoos, L. M.; Tetzloff, G.; Iyer, S. P. N.; Maguire, M.;
Golovko, A.; Zeng, M.; Wang, L.; Murgolo, N.; Grazi-
ano, M. P. Science 2004, 303, 1201; (b) Calvo, M. G.;
Lisnock, J.; Bull, H. G.; Hawes, B. E.; Burnett, D. A.;
Braun, M. P.; Crona, J. H.; Davis, H. R.; Dean, D. C.;
Detmers, P. A.; Graziano, M. P.; Hughes, M.; MacIntyre,
D. E.; Ogawa, A.; O’Neill, K. A.; Iyer, S. P.; Shevell, D.
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102, 8132.
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E. M. J. Med. Chem. 2005, 48, 6035; (b) Ritter, T.;
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icant effect in lowering the total cholesterol and LDL (3b
is the exception) in serum. These SAR trends may pro-
vide insights into the further design of novel cholesterol
absorption inhibitors.
In conclusion, four individual isomers of backbone re-
organized ezetimibe analogs were designed and synthe-
sized. Their effects on the cholesterol levels in rat serum
were evaluated by a high-cholesterol and high-fat diet-
feeding experiment. All of the new compounds showed
significant effects in lowering the levels of total cholester-
ol in serum. This information could be valuable in
designing new cholesterol absorption inhibitors.
12. Corey, E. J.; Reichard, G. A. Tetrahedron Lett. 1989, 30,
5207.
Acknowledgments
13. Xavier, L. C.; Mohan, J. J.; Mathre, D. J.; Thompson, A.
S.; Carroll, J. D.; Corley, E. G.; Desmond, R. Org. Syn.
Cv 9, 676.
14. CCDC 620888 contains the supplementary crystallograph-
ic data for 8b. These data can be obtained free of charge
from The Cambridge Crystallographic Data Center via
www.ccdc.cam.ac.uk/data_request/cif.
The authors wish to express gratitude to Mr. Guangdi
Yang, Ms. Ling Ye, and Dr. Mao Li for conducting
X-ray analysis of products 3a and 8b. This work was
partially supported by a Grant (No. 2005132) from the
Bureau of Science and Technology of Changchun City.
15. CCDC 620889 contains the supplementary crystallograph-
ic data for 3a. These data can be obtained free of charge
from The Cambridge Crystallographic Data Center via
www.ccdc.cam.ac.uk/data_request/cif.
References and notes
16. Cholesterol-feeding experiment: Male Wistar rats with
weight of 150–180 g were purchased from the Center of
Research Animals, the College of Basic Medicines, Jilin
University. The high-fat and high-cholesterol diet was
made from 87.8% regular feeds, 2% cholesterol, 10% hog
fat, and 0.2% propylthiouracil. The animals were raised
for 1 week, divided into individual groups according to
their levels of TC and LDL-cholesterol in serum, and then
fed for 2 weeks on regular diet, high-fat and high-
cholesterol diet, high-fat and high-cholesterol diet with
ezetimibe or individual drugs, respectively. The drugs were
made to a mixture with 0.5% sodium carboxy methyl
cellulose(40 mg/100 ml), administered at a single dosage of
4 mg/kg/day by intragastric perfusion. After fasted for
16 h, blood samples were collected by cutting the tails, and
the serum was isolated and measured for the level of total
cholesterol and LDL-cholesterol. The data were calculated
by statistical methods.
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