Identification of Reversible Small Molecule Inhibitors of Endothelial Lipase (EL) That Demonstrate HDL-C Increase in Vivo

Article abstract of DOI:10.1021/acs.jmedchem.9b01831

Endothelial lipase (EL) hydrolyzes phospholipids in high-density lipoprotein (HDL) resulting in reduction in plasma HDL levels. Studies with murine transgenic, KO, or loss-of-function variants strongly suggest that inhibition of EL will lead to sustained plasma high-density lipoprotein cholesterol (HDL-C) increase and, potentially, a reduced cardiovascular disease (CVD) risk. Herein, we describe the discovery of a series of oxadiazole ketones, which upon optimization, led to the identification of compound 12. Compound 12 was evaluated in a mouse pharmacodynamics (PD) model and demonstrated a 56% increase in plasma HDL-C. In a mouse reverse cholesterol transport study, compound 12 stimulated cholesterol efflux by 53% demonstrating HDL-C functionality.

Full text of DOI:10.1021/acs.jmedchem.9b01831

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Article
Identification of Reversible Small Molecule Inhibitors of
Endothelial Lipase (EL) which Demonstrate HDL-C Increase In Vivo
George O. Tora, Soong-Hoon Kim, Zulan Pi, James A. Johnson, Ji Jiang, Monique S. Phillips, John L.
Lloyd, Lynn Abell, Hao Lu, Gregory Locke, Leonard Adam, David Taylor, Xiaohong Yin, Kamelia Behnia, Lei
Zhao, Richard Yang, Michael Basso, Christian Caporuscio, Alice ye Chen, Eddy Liu, Todd G. Kirchgessner,
Joelle M. Onorato, Carol Ryan, Sarah C. Traeger, David A. Gordon, Ruth R. Wexler, and Heather J. Finlay
J. Med. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jmedchem.9b01831 • Publication Date (Web): 28 Jan 2020
Downloaded from pubs.acs.org on February 2, 2020
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Identification of Reversible Small Molecule Inhibitors of
Endothelial Lipase (EL) which Demonstrate HDL-C
Increase In Vivo
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,†
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George Tora,* Soong-Hoon Kim, Zulan Pi, James A. Johnson, Ji Jiang, Monique Phillips,
†
∞
ƾ
£
‡
‡
John Lloyd, Lynn M. Abell, Hao Lu, Gregory Locke, Leonard P. Adam, David S. Taylor,
‡
§
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Xiaohong Yin, Kamelia Behnia, Lei Zhao, Richard Yang, Michael Basso, Christian
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‡
‡
‡
§
Caporuscio, Alice Ye Chen, Eddie Liu, Todd Kirshgessner, Joelle M. Onorato, Carol
‡
†
†
Ryan, Sarah C. Traeger, ℇ David Gordon,‡ Ruth R. Wexler and Heather J. Finlay.
Departments of Discovery †Chemistry, ‡Biology, ±Pharmaceutics, £Mechanistic Biochemistry,
§Pharmaceutical Candidate Optimization, ℇDiscovery Analytical Sciences, Bristol-Myers Squibb,
Research and Development, P.O. Box 5400, Princeton, New Jersey 08543-5400, United States
KEYWORDS Endothelial Lipase, HDL, Hepatic Lipase
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ABSTRACT: Endothelial lipase (EL) hydrolyzes phospholipids in HDL resulting in reduction in
plasma HDL levels. Studies with murine transgenic, KO or loss-of-function variants strongly
suggest that inhibition of EL will lead to sustained plasma HDL-C increase and, potentially, a
reduced CVD risk. Herein, we describe the discovery of a series of oxadiazole ketones, which
upon optimization, led to the identification of Compound 12. Compound 12 was evaluated in a
mouse PD model and demonstrated a 56% increase in plasma HDL-C. In a mouse reverse
cholesterol transport study, Compound 12 stimulated cholesterol efflux by 53% demonstrating
HDL-C functionality.
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INTRODUCTION
Cardiovascular disease (CVD) is the number one cause of mortality globally. An estimated
17.9 million people died from CVD in 2016, representing 31% of all global deaths. Of these,
an estimated 85% are due to heart attack and stroke.1 Coronary heart disease is caused by
buildup of plaque in arteries resulting in narrowing of the small blood vessels that supply
blood and oxygen to the heart. Every year approximately 735,000 Americans have a heart
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attack.2 In 2016, the total estimated cost, (health care costs, medications and loss in
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productivity) due to heart disease in the US alone was $555B, with costs expected to reach
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Elevated levels of low density lipoprotein cholesterol (LDL-C) are associated with increased
atherosclerotic plaque formation and coronary artery disease (CAD)4. Reduction of LDL-C
has become the cornerstone of treatment to reduce the risk of CV events. Current drug
therapy reduces CAD events by 25-30%5 and there is a further need to reduce events and
decrease mortality associated with CAD. It has been shown that high density lipoprotein
cholesterol (HDL-C) is an independent risk factor for CAD events and plays a central role in
reverse cholesterol transport (RCT). RCT is a process where HDL-C cholesterol is removed
from the periphery (e.g., atherosclerotic plaques in blood vessels) to the liver for
elimination.6,7 HDL-C may also serve to reduce blood vessel injury through its antioxidant and
anti-inflammatory functions.8 Recently a phase III study of the cholesterol ester transfer
protein (CETP) inhibitor, anacetrapib, did not demonstrate benefit on mortality despite robust
increase in HDL-C.9 It is unclear why elevation of HDL-C through inhibition of CETP was not
beneficial, however, evaluation of EL as an alternative mechanism to raise HDL-C could
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provide further insight and generate therapeutic agents that may demonstrate cardiovascular
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benefit.10,11
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Endothelial lipase is a member of the triglyceride lipase family which includes hepatic
lipase, (HL) lipoprotein lipase, (LPL) and pancreatic lipase, (PL).12 The triglyceride lipase
gene family plays a central role in dietary fat absorption. Endothelial lipase is the most
recently discovered member of the triglyceride lipase gene family, with 44% structural
homology to human LPL and 41% homology to human HL 13,14 Unlike LPL and HL, EL is
secreted from vascular endothelial cells and hepatocytes and is believed to function at the
site of synthesis to influence plasma HDL-C through phospholipase activity.15 EL
preferentially hydrolyses the SN-1 (PLA1) ester bond of phosphatidylcholine (PC) present in
HDL-C particles, releasing Lyso-PC and ultimately reducing plasma HDL-C and the number
of HDL particles that participate in RCT. Studies with murine transgenic, KO or loss-of-
function variants demonstrate higher levels of plasma HDL-C. Therefore, 16,17,18 selective
inhibition of EL should increase HDL-C and provide a potential benefit in the treatment of
cardiovascular disease. Despite the central role that EL plays in HDL-C metabolism, only a
few EL inhibitors have been reported.19,20 The first disclosed small molecule EL inhibitor
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(XEN445, 1, Figure 1) increased HDL-C in WT mice at a plasma concentration of 9.9 µM at
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16 h post the last dose.21 In our hands, this compound did not demonstrate inhibition of EL in
serum or plasma based assays. A subsequent disclosure of the irreversible thiocarbamate EL
inhibitor (2) was reported to increase HDL-C at higher doses of 30 and 100 mg/kg22
We have previously disclosed our efforts to identify reversible EL inhibitors with oral
bioavailability23,26-27 (4 and 5, Figure 1). Those compounds, however, did not demonstrate
elevation in HDL-C in our mouse pharmacodynamic model despite achieving target plasma
exposures. Therefore, identification of a selective, reversible EL inhibitor to demonstrate
pharmacological proof of concept remained the focus of our effort.
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Me
O
OH
N
F
O
O
OH
Cl
Cl
N
O
S
N
O
H
N
N
H
Me
F3C
3
2
EL IC50 = 0.15 M
EL IC50 = 0.68 M
1
EL IC50 = 0.24 M
O
O
O
N
S
NH HN
N
S
N
N
NH2
H
O
N
S
O
O
O
S
O
S
Me
O
F
N
O
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4
EL IC50 = 0.03 M
HL IC50 = 26 M
EL IC50 = 0.02 M
HL IC50 = 0.05 M
Figure 1. Recently disclosed EL inhibitors
RESULTS
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Compound 3 was the first compound advanced to the mouse PD model, (based on potency
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in a high throughput in vitro vesicle based assay) and demonstrated no effect on HDL-C. This
led to the development of a human serum (hSerum) and a mouse plasma assay (mPlasma) as
a means to evaluate EL inhibitors in a more biologically relevant environment23 Subsequent
optimization for the in vitro mPlasma potency led to the identification of the EL/HL sulfonated
benzothiazole-oxadiazole inhibitor 4.26 SAR was further developed concurrent with improving
the PK profile resulting in compound 5 which had good hSerum potency (EL IC50 0.12 µM) with
acceptable selectivity verses HL (88 fold).27 Despite compound 3 exhibiting good mouse p.o.
PK, with a Ctrough concentration of >10 fold over mPlasma IC50 of 0.12 µM during the course of
a 4 day PK/PD study, an increase in HDL-C was, again, not observed.
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During the course of our studies to synthesize analogs of 5, we found that non sulfonated
amides like 6 spontaneously oxidized to the keto-amides 7 (Figure 2). This oxidation could be
accomplished chemically in both the amide and oxadiazole series, 8 (Figure 2). We sought to
establish the stability and reversibility with respect to enzyme activity of these ketones while at
the same time maintaining EL activity.
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Amides
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4
O
R
R
O
R
R
N
'air'
N
S
N
N
S
Sponteneous [O]
O
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5
Oxadiazole
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S
Air/Oxidant
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Figure 2. Spontaneous oxidation of amides to keto-amides
Compounds 12-16 were prepared by the general method described in Scheme 1.26 The
methylene bridging the heterocycles was regio-selectively oxidized and the sulfamide
deprotected in one pot to give ketones 12-16. Initially a H2O2/AcOH mixture was used to effect
this oxidation, but yields were low and the conditions were not tolerant to diverse functional
groups. Subsequently, we discovered that using a hypervalent iodine species,
[bis(trifluoroacetoxy)iodo]benzene (PIFA) to effect this oxidation resulted in high yields and was
more general for selective oxidation in this series.
Table 1. Selectivity versus HL SAR
O
O
N
S
N
N
NH2
H
O
N
S
O
R
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EL
HL
EL
EL
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2
3
Compound
R
(µM)a
(µM)a
(µM) a
(µM) a
4
5
human serum
mouse plasma
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9
12
13
0.005
<0.01
0.002
0.01
0.1
0.06
<0.2
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<0.2
F
F
14
15
0.03
0.02
0.04
0.4
ndb
N
O
<0.01
0.005
0.70
O
N
H
O
16
0.02
0.03
0.8
ndb
HN
aInhibition was run at 3 concentrations to calculate IC50 values, bnot determined.
Br
O
b
H
Br
a
O
N
N
N
O
Br
N
O
OH
O
O
N
S
N
NH2
S
O
N
S
OEt
NH
H
O
10b
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10a
OH
HO
B
Br
e
d
+
c
Br
N
N
N
H
NHBoc
O
N
N
N
N
S
S
NH2
S
O
O
O
10d
10c
f
N
N
N
N
H
N
NHBoc
O
N
N
H
NH2
O
S
S
N
S
S
O
O
O
O
O
11
12
Scheme 1. Reagents and conditions: (a) Hydrazine, EtOH, rt, 94%; (b) T3P, DIEA, Dioxane,
67%; (c) TFA/ CH2Cl2, 100%. (d) N-(tert-butoxycarbonyl)-N-[4-(dimethylazaniumylidene)-1,4-
dihydropyridin-1-ylsulfonyl]azanide (N-TDDSA), Pyridine, CH2Cl2, 74%; (e) Xphos 2nd Gen.,
K3PO4, THF, 11-62%; (f) PIFA, CH2Cl2/H2O then TFA, 14-50%
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The inhibitory activity of compounds for EL and HL was evaluated using a fluorogenic
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substrate assay. In this assay the lipid hydrolysis was measured in HDL in the presence of
overexpressed endogenous human EL from conditioned media obtained from HT-1080 cells.
The HL assay was run in the same manner with HL partially purified from conditioned media
obtained from COS7 cells. (Table 1). (see supporting information).23 EL inhibition was also
measured in the presence of hSerum and mPlasma. Test compounds were dissolved in DMSO in a
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96 well plate and combined with a pre-mixed solution of D31-POPC-HDL/human serum and EL. The
plate was incubated at 37 °C for 2 h then quenched with 100% ethanol containing D3-heptadecanoic acid
(internal standard). The plate was sealed, centrifuged and transferred to a PCR plate for LC/MS analysis.
Ketones 12 to 16 showed improved potency in both the mPlasma and hSerum assays relative
to compound 2, but we did not observe any improvement in selectivity for EL versus HL. Given
the excellent EL mouse plasma potency, Compound 12 was chosen as a bench mark compound
to evaluate in vivo.
We were concerned that the ketone could make a covalent irreversible bond with the enzyme
and, as such, compound 12 was evaluated for reversibility. Compound 12 was pre-incubated
with the enzyme and then dialyzed. The activity of the enzyme was checked at 24h to determine
whether enzyme activity was restored. It was found that at 3 uM, 90% of activity was restored
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at 24h and at 9 uM, 64% of activity was restored at 24h, consistent with a competitive mode of
inhibition.
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Compound 12 was advanced to pharmacokinetic studies to determine exposure (Table 2).
The target concentration was >5 fold over the mouse plasma IC50 at Ctrough plasma level.
Compound 12 was dosed at 50 mg/kg by p.o., s.c. and i.p. administration (Table 2). When dosed
i.p., the 24 h plasma exposure was 9.9 fold over the mouse plasma IC50. Having achieved target
exposures, we advanced compound 12 to the mouse pharmacodynamic model to evaluate HDL-
C plasma levels.
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Table 2. Mouse pharmacokinetic propertiesa for compound 12
C24
(µM)
AUC0-24h
(µM.h)
Dosing
method
Dose
(mg/kg)
Cmax(µM)
POb
SCc
IPd
50
50
50
10
2.9
19
0.25
0.19
0.50
48
21
170
b
aData from fasted, male, C57BL/6 mice (n = 6). Vehicle: EtOH/PEG400/H2O (10/70/20),
d
cVehicle: EtOH/PEG400/H2O (10/60/30), Vehicle: 2% PVP-K/0.15%SLS/97.85% H2O, dosed
as a nano suspension
Compound 12 was dosed at 50 mg/kg, i.p. QD in wild type mice (n=6) for 2 days. Mice were
fasted 5 hours prior to bleed on day minus 1 and the first dose administered 18 h post bleed
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with the final dose administered 24 h after the first dose. 1 h post the second dose, the mice
were fasted for 5 h and blood samples collected at 30 h post initial dose (6 h post final dose). A
56% elevation in plasma HDL-C concentration was observed at 30 hours (6 hours post last
dose). The plasma concentration was 520 ± 0.32 nM consistent with previous PK studies (Figure
4).
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Figure 4: HDL-C elevation with compound 12 i.p., QD adminstration at 30 hours post first dose,
(6 hours post second dose).
It was encouraging to observe robust, and exposure-dependent, elevation of HDL-C in this
model. To demonstrate that the HDL-C elevation was due to on-target EL inhibition, a PD study
(50 mg/kg, i.p, BID) in HL KO and EL KO mice was undertaken (Figure 5). Mice were fasted 5
hours prior to baseline bleed on day minus 1. Three doses were administered with the final dose
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administered at 24 h after the first dose. 1 h post third dose, the mice were fasted for 5 h and
blood samples collected at 30 h post initial dose (6 h post final dose). In the HL KO PD study, a
32% elevation of HDL was observed at 30 hours, at a plasma concentration of 11.0 ± 7.4 µM.
In the EL knockout PD study no elevation of HDL was observed at a plasma concentration of
8.5 ± 1.1 µM. These studies showed that the HDL-C elevation observed in wild type mice was
due to EL inhibition and not due to HL inhibition. Sustained HDL-C increase by compound 12 in
WT and KO HL mouse is consistent with EL inhibition as the mechanism for HDL-C elevation.
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Figure 5: HL and EL knock out PD study
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Unlike humans, wild type mice have a relatively high level of HDL-C concentration and low
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levels of LDL-C. In the dual transgenic hCETP/apoB-100 mouse, the lipid profile more closely
resembles that of humans. Compound 12 elevated HDL-C by 36% when dosed at 50 mg/kg,
i.p., BID in the dual transgenic mice and did not affect the LDL-C level.
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It has also been shown that HDL-C plays a central role in RCT, a multi-step process resulting
in the net movement of cholesterol from peripheral tissues back to the liver.7 Cellular cholesterol
efflux is mediated by HDL-C, acting in conjunction with the enzyme lecithin:cholesterol
acyltransferase (LCAT). In a RCT WT mouse model,24-25 mice (n=3) were dosed with compound
12 at 50 mg/kg, i.p, QD, followed by i.v administration of 3H-labelled cholesterol on day 3 (Figure
6). The labelled particles were rapidly taken up by macrophages in the reticuloendothelium
system (RES). The nadir of this initial clearance phase was observed 15-30 minutes post
injection (Figure 6). The reappearance of label after 30 minutes represents the efflux of
cholesterol out of the RES macrophages onto circulating HDL. Therefore, the macrophage
specific efflux was due to the formulation of 3H-cholesterol tracer (albumin complexed
nanoparticles) and the manner in which these particles are distributed following i.v
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adminstration. The net movement of H-labelled cholesterol back to the plasma compartment
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was measured over 2.5 hours. In this experiment, compound 12 stimulated cholesterol efflux by
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53% demonstrating HDL-C functionality.
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37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Figure 6. Compound 12 stimulates Macrophage Cholesterol Efflux. Initial cholesterol efflux rates
determined by linear regression and two-tailed p values for differences in slopes between the
treatment groups calculated (Graphpad Prism, 5.01)
As the physiological effect of HL inhibition is not well understood, we continued our search for
a compound with an improved selectivity profile with respect to HL and an improved oral PK
profile. We chose to substitute the sulfamide with other functional groups in an effort to improve
HL selectivity and ADME properties.
Compound 21 was generated as shown in Scheme 2 from sulfur isocyanatidic chloride, which
was acylated with 2-chloroethanol and condensed with compound 17 to afford intermediate
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Journal of Medicinal Chemistry
oxazolidine 18. Compound 18, a versatile intermediate, was converted to various
alkylsulfamides, for example, compound 19 in 69% yield. XPhos 2nd Gen mediated Suzuki
coupling followed by selective oxidation afforded 21 in 24-50% yields. Compounds 22-24 were
synthesized by the general route shown in Scheme 2.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Scheme 2.
O
O
N
O
NH2
N
S
N
HO
N
N
O
S
N
b
O
N
H
a
NH2
O
N
O
N
S
O
O
Cl
Cl
S
Br
Br
18
10c
O
O
N
S
N
N
O
N
O
H
H
N
S
O
N
S
c, d
N
N
H
N
H
O
N
O
21
S
19
Br
Scheme 2. Reagents and conditions: (a) TEA, CH2Cl2, rt, 73%; (b) TEA, ACN, 95ºC, 69%;
(c) Xphos 2nd. Gen., K3PO4, THF, 22-68%; (d) PIFA, CH2Cl2/H2O then TFA, 5-50%.
Scheme 3.
N
O
Boc
b, c
O
NHNH2
N
S
a
O
O
S
+
N
N
O
Boc
N
S
N
HO
H2N
NH2
22a
22b
O
O
N
S
N
O
S
O
H2N
N
H2N
N
O
O
d
N
N
N
N
S
S
O
22
22c
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Scheme 3. Reagents and conditions: (a) T3P, DIEA, Dioxane, 19%; (b) TFA/DCM (c) 1,4-
dioxane, 100 °C, 53%; (d)H2O2/AcOH, 21%
1
2
3
4
5
6
7
8
9
Scheme 4.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
b
a
N
H
N
N
NH2
N
N
N
N
N
H
N
N
NH2
NH2
S
N
S
S
O
O
O
O
O
O
25b
25
25a
Scheme 4. Reagents and conditions: (a) KOCN,AcOH/H2O, rt, 31%; (b) H2O2, AcOH, 23%.
Substitution of the terminal nitrogen with a methyl resulted in compound 21 which had EL
hSerum potency of 140 nM, but no selectivity against HL. This primary sulfamide was the most
potent compound in this series and we focused on further optimization of 12. Substitution of the
internal nitrogen with a methyl group resulted in a loss of EL hSerum potency (22). There was
no improvement in potency by substituting the α-position with a methyl. Both the S and R methyl
substitutions (23 and 24) resulted in 30-100 fold loss in potency as compared to 12. The
corresponding urea analogue, compound 25, was also prepared (Scheme 3) but was less potent
in the EL hSerum assay and showed no improvement in HL selectivity. Having realized no
selectivity improvement as a result of SAR on the sulfamide, we sought to keep the primary
sulfamide constant and explore SAR at the C5 and C6 positions.
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Journal of Medicinal Chemistry
1
2
3
Table 3. Sulfamide SAR
4
5
6
7
N
R
N
8
9
O
N
S
O
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
EL
HL
EL
EL
Compound
R
(µM) b
(µM)b
(µM)b
(µM)b
human
serum
mouse
plasma
nde
O
S
O
Me
21
0.02
0.02
0.01
0.1
N
N
H
H
O
O
S
N
NH2
22a
0.003
7
1
Me
Me
O
O
S
N
NH2
23a
24a
25
0.09
0.2
0.06
0.03
2
10
1
0.2
H
Me
O
O
S
N
H
NH2
3
O
N
H
NH2
0.03
>0.01
1
bInhibition was run at 3 concentrations to calculate IC50 values.
Previously disclosed SAR26 on benzothiazole–linked sulfones showed that selectivity could be
improved by incorporating polar aryl substituents at C-6. However, the strategy to replace the
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Page 18 of 32
phenyl group with 4-fluoro pyridyl, (14) in the ketone series was unsuccessful. Subsequently,
we sought to modify the electronics of the benzothiazole core to improve selectivity by
incorporating fluorine at C5 and C6. Combining polarity at C6 with fluorine substitution at C5,
gave compound 28 (Table 4), which maintained EL activity with a modest increase in selectivity
versus HL, (9 fold). We synthesized additional C6 substituted compounds with C5 fluorine
substitution (Table 4) and observed an increase in selectivity over HL while maintaining the EL
hSerum potency as shown by compound 29 and 30, (EL hSerum potency of 0.06 uM and 0.02
uM, EL selectivity 38 and 41 fold respectively). Less polar amide, compound 31 had reduced
EL selectivity (8 fold). Similar SAR was observed by reversing the substitution of polar
substituent at C5 and fluorine substitution at C6 (32 and 33). To further evaluate our compounds
and prioritize for mouse PK, compounds were tested in HL KO mPlasma assays. Compounds
which had better HL KO mPlasma activity together with improved EL selectivity were progressed
to mouse PK. Compound 32 (EL hSerum potency of 0.04 uM, EL selectivity of >32, HL KO
mPlasma IC50 of 0.2 µM and excellent human, rat and mouse liver microsome stability) was
selected in the C-6 Fluoro series. In the C5 fluoro series, compound 31 (EL hSerum potency of
0.04 µM, EL selectivity of >9, HL KO mPlasma IC50 of 0.2 µM and >80% liver microsome stability
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
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57
58
59
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in human, rat and mouse) was advanced to rat PK studies with 32 (Table 4). Both compounds
showed good exposure on i.p. QD dosing (Table 5): compound 31 (Cmax = 7.1 µM, C24 = 0.26
µM, AUC = 58.0 µM*h at 50 mg/kg) and compound 32 (Cmax = 21.0 µM, C24 = 0.46 µM, AUC =
36.0 µM*h at 50 mg/kg). Hence, both compounds 31 and 32 were advanced to a mouse PD
study.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Table 4: C5 and C6 Fluoro effect on selectivity versus HL
O
O
N
S
N
N
NH2
5
6
H
R2
R1
O
N
S
O
Compound
R1
R2
EL
HL
EL
HL KO
Met Stab
H/Mb
(µM)b
(µM)b
(µM)
IC50 (µM)a
human
serum
14
28
H
F
0.03
0.02
0.04
nd
nd
78/68
--/66
F
F
N
N
0.001
0.009
0.02
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1
2
3
4
5
6
7
8
29
30
F
F
0.0005
0.0009
0.02
0.04
0.06
0.02
0.2
nd
81/--
HN
O
O
80/-
N
O
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
F
31
32
33
F
<0.0005
<0.0005
<0.0005
0.004
0.02
0.05
0.04
0.2
0.2
0.2
0.2
87/--
H
F
N
F
O
F
96/100
98/100
HN
O
F
F
0.004
H
F
N
F
O
aInhibition was run at 3 concentrations to calculate IC50 values. Percentage of
compound (0.5 uM) remaining after 10 minute incubation in human, rat and mouse liver
microsomes.
b
Table 5. Mouse i.p. PK propertiesa
Cmin
(µM)
AUC0-24h
(µM.h)
Dose
(mg/kg)
Cmax(µM)
Compound
31b
32b
50c
50c
7.1
21
0.25
0.46
58
36
adata from fasted, male, C57BL/6 mice (n = 6), bVehicle: 2% PVP-K/0.15% SLS/97.85%
c
H2O. Dosed as a nano suspension
Wild-type C57BL/6 mice (n = 6) were dosed with compounds 31 and 32 at 50 mg/kg i.p.
QD. Separately, a set of mice (n = 3) were dosed as per protocol and utilized to determine
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1
2
3
4
5
6
7
8
9
plasma exposure at 24 h. This satellite group showed a lower than projected exposure
(target coverage: 5X mouse IC50 at Ctrough) at 24 h (32:C24h = 96 nM, 4.8X lower and 31:
C24h = 36 nM, 7.2X lower). The lipid profile was measured using FPLC analysis of blood
samples taken at 30h (6h post final dose) and showed a 38% and 37% increase in HDL-C
concentrations for compounds 31 and 32 respectively.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
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40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Figure 5: HDL-C elevation with compound 31 and 32 following i.p. QD administration.
DISCUSSION AND CONCLUSION
In summary, we have identified small molecule, reversible EL inhibitors which
demonstrated a significant elevation of HDL-C in a WT-mouse PD model. We have also
shown that dual EL-HL inhibitor, 12, raised HDL-C in WT and HL KO mice but had no
effect on EL KO mice demonstrating that HDL-C increase is due to EL inhibition.
Compound 12 demonstrated increased reverse cholesterol transport in mouse in addition
to causing a sustained and exposure related plasma HDL-C elevation in mice, thus
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1
2
3
4
5
6
7
8
9
demonstrating functionality of the elevated HDL. In addition, we have identified
compounds with potency for EL and increased selectivity versus HL, which demonstrated
increased HDL-C in the mouse PD model (compounds 31 and 32).
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
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EXPERIMENTAL SECTION
Chemistry: General Methods. All solvents and reagents were used as obtained. Reactions
involving air or moisture sensitive reagents were carried out under argon atmosphere. Microwave
®
reactions were performed using Biotage Initiator reactors. NMR spectra were recorded in a
®
deuterated solvent with a Bruker or JEOL Fourier transform spectrometers operating at
1
13
®
®
frequencies as follows: H NMR: 400 MHz (Bruker or JEOL ) or 500 MHz (JEOL ). C NMR:
100 MHz or 125 MHz (Bruker). Spectra data are reported in the format: chemical shift
(multiplicity, coupling constants, number of hydrogens). Chemical shifts are specified in ppm
downfield of a tetramethylsilane internal standard ( units, tetramethylsilane = 0 ppm) and/or
referenced to solvent peaks, which in 1H NMR spectra appear at 2.49 ppm for CD2HSOCD3, 3.30
ppm for CD2HOD, and 7.24 ppm for CHCl3, and which in 13C NMR spectra appear at 39.7 ppm
for CD3SOCD3. All 13C NMR spectra were proton decoupled. All coupling constants (J) are
reported in Hertz.
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1
2
3
4
LCMS chromatograms were obtained on a Shimadzu HPLC system running Discovery VP
5
6
®
software, coupled with a Waters Micromass ZQ mass spectrometer running MassLynx version
7
8
9
3.5 software and using the following methods. For each method the LC column was maintained at
room temperature and UV detection was set to 220 nm. Analytical purity was >95% unless stated
otherwise.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
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41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Method B: A linear gradient using solvent A (10% methanol, 90% water, 0.1% of TFA) and
solvent B (90% methanol, 10% water, 0.1% of TFA); 0-100% of solvent B over 2 min and then
100% of solvent B over 1 min. Column: PHENOMENEX® Luna 5 μm C18 (2.0 x 30 mm). Flow
rate was 1 mL/min.
Method M: A linear gradient using of Solvent A (0.05% TFA, 100% water) and Solvent B
(0.05% TFA, 100% ACN); 2 to 98% B over 1 min, with 0.5 min hold time at 98% B. Column:
Waters BEH C18 (2.1 x 50 mm). Flow rate: 0.8 mL/min.
2-(6-Bromobenzo[d]thiazol-2-yl)acetohydrazide (10a): Ethyl 2-(6-bromobenzo[d]thiazol-2-
yl)acetate (3.0 g, 10.0mmol) was combined with hydrazine (3.14 mL, 100 mmol) in methanol
(60 mL) and stirred at room temperature. After 16 h, a precipitate formed that was collected by
filtration and washed with ethyl acetate (2X) to afford compound 7 (2.7 g, 94%) as a white solid.
LC/MS m/z = 288.0 [M+H], RT = 0.68 min (Method M). MS (ES). 1H NMR (500 MHz, DMSO-
d6) δ 9.47 (br s, 1H), 8.37 (d, J=2.2 Hz, 1H), 7.89 (d, J=8.8 Hz, 1H), 7.64 (dd, J=8.7, 2.1 Hz,
1H), 4.36 (br d, J=3.9 Hz, 2H), 3.99 (s, 2H)
tert-Butyl
((5-((6-bromobenzo[d]thiazol-2-yl)methyl)-1,3,4-oxadiazol-2-
yl)methyl)carbamate
(10b): Compound 10a (400 mg, 1.40 mmol) and 2-((tert-
butoxycarbonyl)amino)acetic acid (294 mg, 1.68 mmol) were dissolved in dioxane (10 mL) and
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2
3
4
5
6
7
8
9
treated with 1-propanephosphonic acid cyclic anhydride (2.08 mL, 3.49 mmol) (50% in ethyl
acetate) and DIEA (0.610 mL, 3.49 mmol). The reaction mixture was heated at 70°C. After 1 h,
the reaction was allowed to cool to rt and treated with additional 1-propanephosphonic acid cyclic
anhydride (2.08 mL, 3.49 mmol) (50% in ethyl acetate) and DIEA (0.610 mL, 3.49 mmol). The
reaction mixture was stirred at 105 °C for 16 h. After allowing to cool to rt, the reaction was
concentrated under reduced pressure and the residue was purified on ISCO (0-100% Hex/EtOAc)
to afford compound 10b (400 mg, 67%). LC/MS m/z = 427.1 (M+H). RT = 1.91 min (Method B).
1H NMR (500 MHz, CHLOROFORM-d) δ 7.83 (dd, J=1.7, 0.8 Hz, 1H), 7.41 (d, J=1.7 Hz, 1H),
7.39 (d, J=1.9 Hz, 1H), 4.76 (s, 2H), 4.59 (d, J=5.0 Hz, 2H), 1.47 (s, 9H)
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(5-((6-Bromobenzo[d]thiazol-2-yl)methyl)-1,3,4-oxadiazol-2-yl)methanamine
(10c):
Compound 10b (300 mg, 0.677 mmol) was combined with DCM (4.0 mL) and treated with TFA
(4.0 mL) at room temperarure for 30 min. The solvents were evaporated under reduced pressure
and azeotroped with toluene (3X). The resulting white solid was dissolved in DCM and washed
with phosphate buffer (1.5 M), brine, dried over sodium sulfate, filtered and concentrated under
reduced pressure to afford compound 10c (260 mg, quant.) which was used as is in the next step.
LC/MS m/z = 327.0 (M+H). RT = 1.34 min (Method B).
tert-Butyl
(N-((5-((6-bromobenzo[d]thiazol-2-yl)methyl)-1,3,4-oxadiazol-2-
yl)methyl)sulfamoyl)carbamate (10d): To a solution of compound 10c (220 mg, 0.640 mmol) in
DCM (10 mL) and pyridine (0.520 mL, 6.40 mmol) was added (tert-butoxycarbonyl)((4-
(dimethyliminio)pyridin-1(4H)-yl)sulfonyl)amide (203 mg, 0.673 mmol). After 16 h, the reaction
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1
2
3
4
5
6
7
8
9
mixture was evaporated under reduced pressure to dryness, dissolved in EtOAc and poured into
satd NH4Cl. The aqueous portion was extracted with EtOAc (3X) and the combined organic
extracts were washed with brine, dried over sodium sulfate, filtered and concentrated under
reduced pressure. The residue was purified on ISCO (0 to 100% EtOAc/DCM) to give compound
10d (246 mg, 74% ) as a white solid. LC/MS m/z = 505.9 (M+H). RT = 1.82 min (Method B). 1H
NMR (400 MHz, CHLOROFORM-d) δ 7.95 (d, J=2.0 Hz, 1H), 7.80 (d, J=8.5 Hz, 1H), 7.54 (dd,
J=8.8, 2.0 Hz, 1H), 4.66 (s, 2H), 4.47 (s, 2H), 1.30 (s, 9H)
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
tert-Butyl
(N-((5-((6-phenylbenzo[d]thiazol-2-yl)methyl)-1,3,4-oxadiazol-2-
yl)methyl)sulfamoyl)carbamate (11): To a solution of Compound 10d (110 mg, 0.210 mmol)
and phenylboronic acid (38.5 mg, 0.320 mmol) in dioxane (2 mL) and water (0.5 mL) was added
potassium phophate (112 mg, 0.530 mmol), and 2nd Gen. Xphos precatalyst (16.57 mg, 0.020
mmol). The mixture was degassed by purging reaction mixture with argon and heated at 90 °C for
2 h. After allowing to cool to room temperature, the reaction mixture was filtered and concentrated
under reduced pressure. The residue was purified on ISCO (0-15% DCM/Methanol) to afford
compound 11 (52 mg, 48%) as a light brown solid. LC/MS m/z = 501.9 [M+H], RT = 1.96 min
(Method B).
N-((5-((6-Phenyl-1,3-benzothiazol-2-yl)carbonyl)-1,3,4-oxadiazol-2-yl)methyl)sulfamide
(12). To a solution of compound 11 (117 mg, 0.290 mmol) in CH2Cl2 (5 mL), TFA (0.1 mL) and
water (26 µL, 1.5 mmol) was added [bis(trifluoroacetoxy)iodo]benzene (376 mg, 0.870 mmol) at
room tempeartue. The mixture mixture was stirred for 10 min and concentrated under reduced
pressure. The residue was purified on ISCO (0-10% DCM/Methanol) to afford compound 12 (21
mg, 18%) as a yellow solid. LC/MS m/z = 416.3 (M+H). RT = 1.81 min (Method B). HRMS m/z
1
+
(ESI) calcd for C17H13N5O4S2 ([M+H] ) 416.0482, found 416.0467. H NMR (500MHz,
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5
6
ACETONITRILE-d3) δ 8.44 (d, J=1.3 Hz, 1H), 8.37 (d, J=8.5 Hz, 1H), 8.00 (dd, J=8.7, 1.7 Hz,
1H), 7.80 - 7.78 (m, 2H), 7.56 - 7.53 (m, 2H), 7.46 (m, 1H), 6.01 (br t, J=5.5 Hz, 1H), 5.60 - 5.34
7
8
9
13
(m, 2H), 4.61 (s, 2H). C NMR (126MHz, ACETONITRILE-d3) δ 172.1, 167.6, 163.5, 162.0,
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
154.2, 143.2, 140.9, 139.6, 130.6 (s, 2C), 129.8, 128.9, 128.8 (s, 2C), 127.5, 122.1, 39.2.
Compounds 31 and 32 were prepared from 10d using the methods and conditions
described for 12
4-(5-Fluoro-2-{5-[(sulfamoylamino)methyl]-1,3,4-oxadiazole-2-carbonyl}-1,3-
benzothiazol-6-yl)-N-(2,2,2-trifluoroethyl)benzamide (31). 26% yield. LCMS (method M)
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retention time = 0.75 min, (M+H) = 559.0. HRMS m/z (ESI) calcd for C20H14F4N6O5S2 ([M-H]
) 557.0319, found 557.0332. 1H NMR (500 MHz, Acetone-d6) δ 8.51 (d, J=7.3 Hz, 1H), 8.44 (t,
J=6.3 Hz, 1H), 8.19 (d, J=11.0 Hz, 1H), 8.11 (d, J=8.5 Hz, 2H), 7.84 (dd, J=8.2, 1.6 Hz, 2H), 6.85
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(t, J=5.8 Hz, 1H), 6.27 (s, 2H), 4.75 (d, J=6.0 Hz, 2H), 4.23 (qd, J=9.5, 6.6 Hz, 2H). C NMR
(126 MHz, Acetone-d6) δ 171.3, 167.6, 167.5, 165.4, 161.6, 160.5 (d, J=247.0 Hz, 1C), 154.8 (d,
J=12.7 Hz, 1C), 139.2, 134.8 (d, J=1.8 Hz, 1C), 134.7, 131.5 (d, J=17.3 Hz, 2C), 130.4 (d, J=2.7
Hz, 1C), 128.8 (s, 2C), 125.4 (d, J=4.5 Hz, 1C), 125.9 (q, J=278.8 Hz, 1C), 112.7 (d, J=25.4 Hz,
1C), 41.5 (q, J=34.5 Hz, 1C), 39.0. 19F NMR (471 MHz, Acetone-d6) δ -72.70 (t, J=9.5 Hz, 3F),
-117.94 (t, J=8.7 Hz, 1F).
N-({5-[6-Fluoro-5-(1-oxo-2,3-dihydro-1H-isoindol-5-yl)-1,3-benzothiazole-2-carbonyl]-
1,3,4-oxadiazol-2-yl}methyl)aminosulfonamide (32). 34% yield. LCMS (method E) retention
+
-
time = 0.73 min, (M+H) = 489.0. HRMS m/z (ESI) calcd for C27H26N6O6ClS3 ([M-H] )
487.0289, found 487.0302. 1H NMR (500 MHz, DMF-d7) δ 8.57 - 8.56 (m, 1H), 8.56 - 8.55 (m,
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1H), 8.46 (d, J=10.1 Hz, 1H), 7.98 (s, 1H), 7.92 - 7.89 (m, 1H), 7.89 - 7.86 (m, 1H), 7.71 (br t,
J=6.1 Hz, 1H), 6.98 (s, 2H), 4.75 (d, J=6.0 Hz, 2H), 4.60 (s, 2H). 13C NMR (126 MHz, DMF-d7)
δ 170.4, 170.0, 167.3, 161.1, 159.4 (d, J=250.7 Hz, 1C), 150.9, 145.3, 138.5 (br d, J=12.7 Hz,
1C), 138.1, 133.3, 132.1, 130.5 (br d, J=17.3 Hz, 1C), 129.4 (br d, J=2.7 Hz, 1C), 127.9 (br d,
J=4.5 Hz, 1C), 125.1 (br d, J=2.7 Hz, 1C), 123.4, 110.3 (br d, J=30.0 Hz, 1C), 45.6, 38.2. 19F
NMR (471 MHz, DMF-d7) δ -114.74 (br t, J=8.7 Hz, 1F). Orthogonal HPLC purity: Method O1:
RT = 5.02 min, 96%; Method O2: RT = 5.86 min, 99%.
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ASSOCIATED CONTENT
Supporting Information.
The supporting information is available free of charge at
Experimental details on chemistry general methods; experimental section; biological activity;
revesibility assays and reversibility study for compound 12; standard deviations of EL IC50, HL
IC50 and EL hSerum IC50 for compounds 12, 31 and 32; pharmacokinetics, single-dose mouse
pharmacokinetics and mouse PD studies; Molecular formula strings(CSV)
AUTHOR INFORMATION
Corresponding Author
E-mail: george.tora@bms.com.
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Present Address
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(K.B) Glaxosmithkline PRR DPU, 1250 S Collegeville Rd, Collegeville, PA 19426, United States.
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(L.A.) Agios Pharmaceuticals, 88 Sidney Street, Cambridge MA 02139, United States.
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(R.Y.) Sollers Institute, 55 Parsonage Road, Edison NJ 08837, United States.
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(H.L.) EMD Serono Research and Development, 45 Middlesex Turnpike, Billerica, MS 01821, United States.
Author Contributions
All authors have given approval to the final version of the manuscript.
Notes
The authors were employed by Bristol-Myers Squibb at the time of this work and some authors
may own stock.
ACKNOWLEDGMENT
The authors would like to thank Dr. Arvind Mathur, Rick Rampulla and colleagues at
Department of Discovery Synthesis at Biocon Bristol-Myers Squibb RD Center for the
preparation of intermediates.
ABBREVIATIONS
HDL, high density lipoprotein; HDL-C, high density lipoprotein cholesterol; KO, knock-
out; SAR, structure activity relationship, CVD, cardiovascular disease; EL, endothelial
lipase; HL, hepatic lipase; LPL, lipoprotein lipase; PK, pharmokinetic; PD,
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pharmacodynamic; FPLC, fast protein liquid chromatography; N-TDDSA, N-(tert-
butoxycarbonyl)-N-[4-(dimethylazaniumylidene)-1,4-dihydropyridin-1-ylsulfonyl]azanide
BID, dosing twice daily, QD, dosing once daily; H2O2, Hydrogen peroxide; AcOH, acetic
acid; PIFA, [bis(trifluoroacetoxy)iodo]benzene.
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REFERENCES
1. World Health Organization. Fact sheet. Cardiovascular diseases (CVDs). 17 May 2017.
2. Mozaffarian D, Benjamin, E. J.; Go, A. S.; Arnett, D, K.; Blaha, M. J.; Cushman, M.; de Ferranti, S.; Després, J.
P.; Fullerton, H. J.; Howard, V. J.; Huffman, M. D.; Judd, S. E.; Kissela, B. M.; Lackland, D. T.; Lichtman, J.
H.; Lisabeth, L. D.; Liu, S.; Mackey, R. H.; Matchar, D. B.; McGuire, D. K.; Mohler, E. R 3rd.; Moy, C.
S.; Muntner, P.; Mussolino, M. E.; Nasir, K.; Neumar, R. W.; Nichol, G.; Palaniappan, L.; Pandey, D. K.; Reeves,
M. J.; Rodriguez, C. J.; Sorlie, P. D.; Stein, J.; Towfighi, A.; Turan, T. N.; Virani, S. S.; Willey, J. Z.; Woo, D.;
Yeh, R. W.; Turner, M. B.. Heart disease and stroke statistics—2015 update: a report from the American Heart
Association. Circulation. 2015;131:e29-322.
3. American Heart Association 1/17DS11775, 2017
4. (a) Martinez, L. O.; Jacquet, S.; Tercé, F.; Collet, X.; Perret, B.; Barbaras, R. New insight on the molecular
mechanisms of high-density lipoprotein cellular interactions. Cell. Mol. Life Sci. 2004, 61 (18), 2343-2360; (b)
Gordon, D. J.; Probstfield, J. L.; Garrison, R. J.; Neaton, J. D.; Castelli, W. P.; Knoke, J. D.; Jacobs, D. R., Jr.;
Bangdiwala, S.; Tyroler, H. A. High-density lipoprotein cholesterol and cardiovascular disease. Four prospective
American studies. Circulation 1989, 79 (1), 8-15
5. Khera, A. V.; Cuchel, M.; Llera-Moya, M.; Rodriques, A.; Burke, M.; Jafri, K.; French, B. C.; Phillips, J. A.;
Mucksavage, M. L.; Wilensky, R. L.; Mohler, E. R.; Rothblat, G. H.; Rader, D. Cholesterol efflux capacity, high-
density lipoprotein function, and atherosclerosis. J. N. Engl. J. Med. 2011, 364, 127-135.
6. Gordon, T.; Castelli, W. P.; Hjortland, M. C.; Kannel, W. B.; Dawber, T. R. High density lipoprotein as a
protective factor against coronary heart disease. The Framingham study. J. Am. Med. 1977, 707-714.
7. Annema, W.; Tietge, U. J. Role of hepatic lipase and endothelial lipase in high-density lipoprotein-mediated
reverse cholesterol transport. Curr. Atheroscler. Rep 2011, 13, 257-265.
8. Soran, H.; Hama, S.; Yadav, R.; Durrington, P. N. HDL functionality. Curr. Opin. Lipidol. 2012, 23 (4), 353-366.
9. Filippatos, T. D.; Kei, A.; Elisaf, M. S. Anacetrapib, a New CETP Inhibitor: The New Tool for the Management
of Dislypidemias? Diseases 2017, 5, 21.
10. Hafiane, A.; Genest, J. HDL, Atherosclerosis, and Emerging Therapies. Cholesterol 2013, 2013, 1−18.
11. DeSantis, P.; Coleman, T.; Schiekofer, S.; Nawroth, P. P.; Schlimmer, P.; Schneider, J. G. Mini-Rev. Med. Chem.
2008, 8, 619-627.
12. Jaye, M.; Lynch, K. J.; Krawiec, J.; Marchadier, D.; Maugeais, C.; Doan, K.; South, V.; Amin, D.; Perrone, M.;
Rader, D. J. A novel endothelial-derived lipase that modulates HDL metabolism. Nat. Genet. 1999, 21 (4), 424-
428.
13. Hirata, K.; Dichek, H. L.; Cioffi, J. A.; Choi, S. Y.; Leeper, N. J.; Quintana, L.; Kronmal, G. S.; Cooper, A. D.;
Quertermous, T. Cloning of a unique lipase from endothelial cells extends the lipase gene family. J. Biol. Chem.
1999, 274, 14170-14175.
14. Jaye, M.; Lynch, K. J.; Krawiec, J.; Marchadier, D.; Maugeais, C.; Doan, K.; South, V.; Amin, D.; Perrone, M.;
Rader, D. A novel endothelial-derived lipase that modulates HDL metabolism. J. Nat. Genet. 1999, 21, 424-428.
15. Choi, S. Y.; Hirata, K.; Leo, G. C.; Ishida, T.; Quertermous, T.; Cooper, A. D. Endothelial lipase: a new lipase on
the block. J. Lipid Res. 2002, 43, 1763-1769.
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