Design and synthesis of a novel series of cyclohexyloxy-pyridyl derivatives as inhibitors of diacylglycerol acyl transferase 1

Article abstract of DOI:10.1039/c2md20187a

A novel series of potent diacylglycerol acyl transferase 1 inhibitors was developed from the clinical candidate AZD3988. Replacement of the phenyl cyclohexyl-ethanoate side chain with substituted oxy-linked side chains to introduce changes in shape and polarity, reduce lipophilicity and mask the hydrogen bond donors with internal hydrogen bond acceptors led to improvements in solubility, unbound clearance and excellent selectivity over the related enzyme acyl-coenzyme A:cholesterol acyltransferase 1. A comparison of the small molecule crystal structures of compound 4 and compound 28 is described. Compounds in this series have good ADMET properties and provide an exposure-dependent decrease in circulating plasma triglyceride levels in a rat oral lipid tolerance test. The Royal Society of Chemistry 2013.

Full text of DOI:10.1039/c2md20187a

MedChemComm
View Article Online
View Journal | View Issue
CONCISE ARTICLE
Design and synthesis of a novel series of cyclohexyloxy-
pyridyl derivatives as inhibitors of diacylglycerol acyl
transferase 1†
Cite this: Med. Chem. Commun., 2013,
4, 151
a
Alleyn T. Plowright, Peter Barton,b Stuart Bennett,b Alan M. Birch,b Susan Birtles,b
*
Linda K. Buckett,b Roger J. Butlin,b Robert D. M. Davies,b Anne Ertan,c
Pablo Morentin Gutierrez,b Paul D. Kemmitt,b Andrew G. Leach,b Per H. Svensson,cd
Andrew V. Turnbullb and Michael J. Waringb
A novel series of potent diacylglycerol acyl transferase 1 inhibitors was developed from the clinical
candidate AZD3988. Replacement of the phenyl cyclohexyl-ethanoate side chain with substituted oxy-
linked side chains to introduce changes in shape and polarity, reduce lipophilicity and mask the
hydrogen bond donors with internal hydrogen bond acceptors led to improvements in solubility,
unbound clearance and excellent selectivity over the related enzyme acyl-coenzyme A:cholesterol
acyltransferase 1. A comparison of the small molecule crystal structures of compound 4 and compound
28 is described. Compounds in this series have good ADMET properties and provide an exposure-
dependent decrease in circulating plasma triglyceride levels in a rat oral lipid tolerance test.
Received 6th July 2012
Accepted 10th August 2012
DOI: 10.1039/c2md20187a
www.rsc.org/medchemcomm
inhibitors,5–7 including a series of oxadiazole amides leading to
the clinical candidate AZD3988 (1) (Fig. 1).6 Compound 1
Introduction
The rate of developing obesity is alarming considering the
connection between a body mass index above 30 and diseases
such as diabetes, hypertension, arterial cardiovascular disease,
stroke, and some forms of cancer. Hence, there is strong
demand for drugs capable of treating both obesity and type 2
diabetes. Inhibition of diacylglycerol acyl transferase-1 (DGAT-
1), a membrane bound enzyme found in the endoplasmic
reticulum, is of increasing interest as a mechanism for thera-
peutic treatment of diabetes, obesity and other diseases which
together constitute metabolic syndrome.1–3 DGAT-1 decient
(Dgat-1-) mice are viable and resistant to weight gain when fed a
high-fat diet and show increased insulin and leptin sensitivity.4
A number of small molecule inhibitors of DGAT-1 have been
reported and the progress of some compounds into clinical
development supports the potential for therapeutic use.
Recently we have reported different series of DGAT-1
contains a phenyl cyclohexyl-ethanoate side chain, a fragment
which appears to be a privileged structure as it is contained
within several DGAT-1 inhibitors,8–10 including LCQ-908 (2) and
PF-4620110 (3) which have both progressed to the clinic.8,11
As part of the ongoing research to develop the oxadiazole
amide series, variations in this fragment were sought to provide
structural variation and optimise the compounds further.
Despite 1 displaying many positive criteria a potential area of
improvement was in the physicochemical properties (thermo-
dynamic solubility of crystalline 1 ¼ 2 mM).12 Whilst 1 appeared
to have good selectivity over its closest structural homologue,
acyl-coenzyme A:cholesterol acyltransferase 1 (ACAT-1),13 with
an ꢀ40 fold in vitro enzyme selectivity, it was found to cause
aCardiovascular and Gastrointestinal Innovative Medicines Unit, AstraZeneca R&D,
¨
Pepparedsleden 1, Molndal, 43183, Sweden. E-mail: alleyn.plowright@astrazeneca.
com; Tel: +46 (0)31 7761572
bCardiovascular and Gastrointestinal Innovative Medicines Unit, AstraZeneca R&D,
Mereside, Alderley Park, Cheshire, SK10 4TG, UK
c
¨
¨
Medicines Evaluation, AstraZeneca R&D, 151 85 Sodertalje, Sweden
dDepartment of Applied Physical Chemistry, Royal Institute of Technology, SE-10044,
Stockholm, Sweden
† Electronic supplementary information (ESI) available: Synthetic details for the
preparation of compounds and crystallographic data. CCDC 886319 and
886320. For ESI and crystallographic data in CIF or other electronic format, see
DOI: 10.1039/c2md20187a
Fig. 1 Some DGAT-1 inhibitors.
This journal is ª The Royal Society of Chemistry 2013
Med. Chem. Commun., 2013, 4, 151–158 | 151

View Article Online
MedChemComm
Concise Article
adrenal toxicities in dog toxicology studies which may impact isothiocyanate and then cyclised using 1-ethyl-3-(3-di-methyl-
clinical utility.14 Hence greater selectivity towards ACAT-1 was aminopropyl)carbodiimide hydrochloride to give the 1,3,4-oxa-
sought. Aqueous solubility is driven by a number of factors diazole ring system which nally underwent ester hydrolysis to
including lipophilicity and solid-state interactions in the crystal give the desired acid derivatives 9.
lattice. Strategies to explore both of these aspects were devised
The oxy-linked pyridyl derivatives were synthesized in one of
and herein our continued investigations to improve compound the two ways (Scheme 2). Firstly, treating the anion of 5 with the
properties to provide an alternative series of acidic DGAT-1 substituted 2-uoro-5-bromopyridine in an analogous fashion
inhibitors are reported.
to the rst step in Scheme 1 gave the 2-alkoxy-5-bromopyridine
11. Buchwald coupling of 11 with benzophenone imine under
palladium catalysis followed by hydrogenation of the imine
gave the required pyridylamine 12. Alternatively, performing a
Mitsunobu reaction between the cyclohexanol 5 and the
substituted 2-hydroxypyridine 13 using diisopropylazodi-
carboxylate and triphenylphosphine gave the intermediate 14
which was hydrogenated as before to give the required pyr-
idylamine 12. In the case of compound 28, where a uoro-
substituent was required ortho to the pyridyl nitrogen, the
Mitsunobu reaction was performed using 6-chloro-5-nitro-
pyridin-2-ol (i.e. R2 ¼ 6-Cl in 13) to give 14. The 6-chloro group
was then converted to the 6-uoro group using tetrabuty-
lammonium cyanide and hexauorobenzene in DMSO, which
upon hydrogenation gave the pyridylamine intermediate 12
with the required substitution pattern. The pyridylamines 12
were transformed into the nal oxadiazole acids 9 using
essentially the same chemistry as described in steps (d)–(g) in
Scheme 1.
Results and discussion
Chemistry
The synthesis of the key oxy-linked phenyl and pyridyl deriva-
tives is described in Schemes 1 and 2. Treatment of a mixture of
cis- and trans-ethyl-4-hydroxycyclohexane carboxylate 5 with
sodium hydride gave the alkoxide anion which underwent a
nucleophilic aromatic substitution reaction with 1-uoro-4-
nitrobenzene 6. This provided a mixture of cis- and trans-
intermediates, which were separated by ash column chroma-
tography to provide the pure cis-cyclohexyl product, which was
hydrogenated to give the substituted aniline 7. Acylation of 7
using methyl oxalylchloride gave the oxalate ester which was
displaced with hydrazine monohydrate to give 8. The mono-
hydrazide 8 was reacted with the required substituted aryl
Table 1 Variations in the oxy-linked side chain and aniline substitution
hDGAT- hACAT-
Sol.b
a
a
1 IC50
1 IC50
R1
R2
(mM)
(mM)
Log D (mM) LLE
Scheme 1 Synthesis of oxy-linked phenyl derivatives. Reagents and conditions:
(a) NaH (60% in oil), DMF, 0 ꢁC to rt; (b) 6, DMF, 110 ꢁC; (c) Pd (10% on C), H2,
EtOH : THF (1.5 : 1); (d) methyl oxalylchloride, pyridine, CH2Cl2; (e) N2H4$H2O,
EtOH; (f) (i) R3C6H4NCS, DMA, (ii) EDC, 60 ꢁC; and (g) LiOH, THF : H2O : MeOH
(1 : 1 : 2), 35 ꢁC.
1
—
—
0.002
0.14
0.08
3.0
2.3
2
6
5.6
4.6
15
3,4-diF
—
16
17
18
19
3,4-diF
4-F
0.004
0.005
0.018
1.1
—
3
—
1.8
1.2
1.4
1.9
5
7.1
6.3
6.2
4-CN
14.3
0.45
—
43
2,4,5-triF 0.007
2,4,5-triF 0.008
Scheme 2 Synthesis of oxy-pyridyl derivatives. Reagents and conditions: (a) NaH
(60% in oil), DMF, 0 ꢁC to rt; (b) 10, DMF, 100 ꢁC; (c) if R1 ¼ COOH, then HCl,
MeOH; (d) benzophenone imine, PdOAc2, Cs2CO3, (S)-(ꢂ)-2,20-bis(diphenylphos-
phino)-1,10-binaphthyl, THF, 60 ꢁC; (e) Pd (10% on C), H2, ethyl acetate; (f) 13,
20
1.3
1.9
27
6.2
diisopropylazodicarboxylate, PPh3, THF, 180 ꢁC, microwave; (g) for 28 where R2
¼
a
6-Cl, Bu4NCN, DMSO, hexafluorobenzene, 0 ꢁC to rt; and (h) Pd (10% on C), H2,
All IC50 values reported are mean values from $2 experiments.
b
Measured by incubation of crystalline materials for 24 h at pH 7.4.12
ethyl acetate.
152 | Med. Chem. Commun., 2013, 4, 151–158
This journal is ª The Royal Society of Chemistry 2013

View Article Online
Concise Article
MedChemComm
energy is minimized when these two dipole moments oppose
one another as is seen in the observed conformation. The
second conformational feature is that the N–H bond of the
group linking the oxadiazole to the phenyl ring has the ortho-
uoro substituent towards it rather than the unsubstituted
ortho position. Again, the dipole moment of the N–H bond is
opposed to that of the C–F bond in this conformation whereas
the alternative conformation would present a C–H bond with a
reinforcing dipole which would be higher in energy. These
conformational preferences are conrmed by quantum
mechanical calculations (RHF/6-31G*) on a fragment of 4 with
the cyclohexyl-ethanoic acid group removed, which shows that
the lowest energy conformation is the one found in the crystal
structure.18,19 Rotation by 180ꢁ about the oxadiazole–amide
bond costs 5.0 kcal molꢂ1. Rotation about the oxadiazole–N
bond by the same angle costs 2.1 kcal molꢂ1. No minimum
energy structure could be found for rotation about the N-phenyl
bond without also rotating about the oxadiazole–N bond.
The intermolecular interactions revealed by the crystal
structure include (i) the amide NH interacting with the carbonyl
of an adjacent amide; (ii) the aniline NH interacting with one of
the oxadiazole nitrogens and (iii) the carboxylate residue in 4
Compound proling
During the discovery of 1 it had been found that the carboxylic
acid functionality was important for combining potent DGAT-1
inhibition in vitro with good oral activity in vivo and hence,
for this study the SAR focused on acidic derivatives. The lip-
ophilicity of 1 is not high (log D ¼ 3, Table 1),12 but as the
permeability of 1, as measured in MDCK cells (Papp A-B 12 ꢃ
10ꢂ6 cm sꢂ1), is high there is a scope for lipophilicity to be
reduced to improve compound characteristics including phys-
icochemical properties.15 In parallel, crystals of the closely
related derivative 4 (Fig. 1), itself being a potent DGAT-1
inhibitor (IC50 ¼ 3 nM), suitable for X-ray structural determi-
nation were grown to understand the interactions existing in
the solid state within this series (Fig. 2).16 A view of the X-ray
structure in Fig. 2a shows the plane of molecules that are
arranged adjacent to one another and reveals two interesting
conformational features and a number of intermolecular
interactions. The rst conformational feature is that the amide
carbonyl is oriented away from the oxadiazole nitrogens.
¨
Bostrom et al. have shown that the 1,3,4-oxadiazole has a large
associated dipole moment.17 The amide carbonyl also has a
large associated dipole moment and the overall conformational
Fig. 2 Two views of the crystal structure of the acetic acid co-complex of compound 4. Panel (a) shows a view from above the plane of adjacent molecules and panel
(b) shows how these planes stack.
This journal is ª The Royal Society of Chemistry 2013
Med. Chem. Commun., 2013, 4, 151–158 | 153

View Article Online
MedChemComm
Concise Article
forming a tight dimer with the acetic acid also present in the 0.01%) and high intrinsic metabolic clearance in rat hepato-
structure. The quantum mechanical calculations can also be cytes (Clint ¼ 22 mL minꢂ1 mgꢂ1) so was not pursued. Intro-
used to investigate the strengths of intermolecular interactions. duction of an oxy-link between the phenyl and cyclohexyl rings
In particular, Kenny has shown that electrostatic potential of 1 to provide the cis-cyclohexyl derivative 16 proved more
calculations correlate closely with observed values of log Ka and benecial and excellent inhibition of hDGAT-1 was achieved.
log Kb which are experimental measures of a molecule’s Compound 16 also showed excellent selectivity against hACAT-1
hydrogen bonding capacity.20,21 These calculations reveal that (280 fold).
the same features that drive the conformational preference for
Combining the cis-cyclohexyloxy side chain with alternative
the molecule also inuence its ability to interact with other aniline substitution on the oxadiazole ring was explored
molecules. The conformation of the same model compound extensively and a large variety of substitution was tolerated with
described above that has uorine oriented towards the NH has a a similar SAR to what was observed in the initial series.6 As
computed log Ka for the NH of 2.02 while the same compound increasing molecular weight and lipophilicity would be detri-
lacking the ortho uoro has a log Ka of 2.24. Hence, any through- mental to physicochemical properties the focus was primarily
bond polarizing effect of uorine is counteracted by a through- on small variations of aniline (Table 1). Replacement of the
space masking of the dipole of the N–H bond by that of the C–F meta-uoro substituent of 16 with a hydrogen atom to give the 4-
bond. The alternative view of the crystal structure in Fig. 2b uoro derivative 17 gave excellent inhibition of hDGAT-1 with a
shows how the molecules stack on top of one another and
reveals that the various layers alternate the orientation of the
molecule and that the molecules are slightly twisted with
Table 2 Variations in the phenyl cyclohexyloxy acid side chains
respect to one another indicating that there are no signicant
stacking interactions between the layers.
Hence, the initial design strategies to improve physico-
chemical properties were focused on reducing the overall lip-
ophilicity and disrupting the intricate framework of intra- and
hDGAT- hACAT-
1 IC50
1 IC50
Sol.b
Log D (mM) LLE
a
b
intermolecular hydrogen bonding observed in the crystal lattice
of 4 by (i) introducing changes in shape and polarity into the
side chain to reduce the efficiency of crystal packing; (ii)
reducing the overall lipophilicity of the phenyl cyclohexyl-
ethanoate side chain, whilst maintaining the high ligand lip-
ophilicity efficiency (LLE) of 1; and (iii) masking the hydrogen
bond donors which are required for DGAT-1 inhibition in this
series with internal hydrogen bond acceptors to break up the
hydrogen bonding network in the crystal lattice to increase
solubility. Compounds were tested in vitro for inhibition
towards recombinant human DGAT-1 (hDGAT-1).5 To ensure
selectivity, inhibition of human ACAT-1 (hACAT-1) and human
DGAT-2, which although serving the same function is only
distantly related to DGAT-1 structurally,22 was monitored to nd
opportunities for enhancing selectivity. Solubility was
measured on crystalline materials.12
Initial attempts to introduce changes in the shape and
increase the polarity in the side chain were made through
introduction of an oxygen atom. The phenyl-trans-cyclohexyl
unit in 1 is linear and relatively at and therefore packs well in
the lattice. The introduction of an oxygen linker completely
changes the shape to the one that packs less efficiently. This can
be exaggerated further by changing from a trans-conguration
in which both substituents can be placed in equatorial posi-
tions to a cis-conguration in which one of the substituents
must become axial. Axially substituted cyclohexanes have a
shape that tessellates less well and so might reduce the effi-
ciency of packing in the lattice. Initially this was attempted on
the bis-phenyl side chain substituted with a carboxylic acid in
the ortho-, meta- or para-position. Whilst good hDGAT-1 inhi-
bition was observed with both the meta- (15) and para-deriva-
tives (data not shown) the solubility of both remained poor.
R1
R2
(mM)
(mM)
21
22
3,4-diF
0.005
6.5
1.9
1.9
49 6.5
69 6.5
2,4,5-triF 0.003
2,4,5-triF 0.015
2,4,5-triF 0.008
0.31
23
24
25
26
27
28
6.5
0.86
5.6
1.5
4.8
1.1
1.8
2.3
1.4
2.2
1.9
1.8
13 6.0
11 5.8
23 6.8
3,4-diF
3,4-diF
3,4-diF
0.005
0.007
0.003
4
6.0
15 6.6
2,4,5-triF 0.004
2,4,5-triF 0.006
2200 6.6
29
1.3
1.6
240 6.6
a
All IC50 values reported are mean values from $2 experiments.
b
Measured by incubation of crystalline materials for 24 h at pH 7.4.12
In addition, 15 showed high rat plasma protein binding (Fu
<
154 | Med. Chem. Commun., 2013, 4, 151–158
This journal is ª The Royal Society of Chemistry 2013

View Article Online
Concise Article
MedChemComm
corresponding reduction in lipophilicity and hence improve- to a reduction of log D to 1.9 and still provided good perme-
ment in LLE. However, the solubility was still poor. Substitution ability as measured in MDCK cells. Additional uoro-substitu-
on aniline with a 4-CN group (18) led to reduced potency versus tion in the side chain, for example, 22, also gave high
hDGAT-1 so was not pursued further.
permeability.23 This was hypothesised as both hydrogen bond
Alternatively, in an attempt to disrupt the intermolecular donors could now be masked through intramolecular hydrogen
hydrogen bond between the hydrogen bond donor of the aniline bonds. However, introduction of an additional heteroatom into
and an oxadiazole nitrogen acceptor in a similar fashion to that the side chain such as the pyridyl derivative 21 gave a signicant
shown with 4 (Fig. 2), a uoro substituent was added to the reduction in permeability, despite the compound having the
aniline ring ortho to the anilino-NH to give the 2,4,5-triuoro same log D as 19 and 22. Further reduction of the lipophilicity of
derivative 19. Excellent inhibition of hDGAT-1 was achieved, the the side chain led to a signicant reduction in permeability as
fraction unbound in rat plasma protein binding showed a 10- exemplied with 25 (MDCK Papp A-B 1.1 ꢃ 10ꢂ6 cm sꢂ1). The
fold increase (0.4%) compared with 1 and solubility was clearance of the compounds in human hepatocytes was low in
increased to 43 mM. The corresponding trans-derivative, 20, had most cases. However, differences emerged in in vivo DMPK
the same potency against hDGAT-1 but interestingly was 3-fold experiments. The oxy-linked phenyl derivatives 19 and 22 had
less potent than the cis-derivative against hACAT-1.
excellent DMPK parameters in rat including low clearance and
Attempts were made to reduce the lipophilicity further high bioavailability. The clearance in the dog was signicantly
(Table 2). These included reducing the size of the cyclohexyl higher, particularly aer normalizing for plasma protein
ring to the cyclobutyl, which led to a 10-fold reduction in binding to give the unbound clearance. This was in contrast to
potency against hDGAT-1 (data not shown), and introducing an the pyridyl derivative 21, which had signicantly higher clear-
additional heteroatom into the side chain by replacing the ance in the rat mainly due to increased renal clearance. In the
phenyl ring with a pyridyl ring as exemplied with 21. dog the clearance, in particular the unbound clearance, was
Compound 21 maintained good potency and achieved excellent signicantly lower. In summary, the cis-uorophenyl derivatives
selectivity versus hACAT-1 as well as improved solubility. The 19 and 22 offered improvements in solubility and permeability
nal strategy to improve solubility by disrupting the hydrogen and an excellent rat DMPK prole, whereas the pyridyl deriva-
bonding framework was to replicate the effect of the uorine tive 21 offered advantages in selectivity against hACAT-1 and
atom adjacent to the aniline NH and also mask the amide lower unbound clearance in the dog.
hydrogen bond donor with an ortho-uoro substituent. This
To achieve a combination of these properties two strategies
hypothesis was tested by synthesising the ortho-uoro derivative were undertaken. Firstly, slightly increasing the lipophilicity of
22. Compound 22 gave excellent inhibition of hDGAT-1, high the pyridyl derivative 21 to increase permeability and reduce
LLE and gratifyingly a further small, but consistent, increase in renal clearance gave 26 (Table 2). Both excellent potency
solubility. Interestingly, 22 also had good potency against towards hDGAT-1 and selectivity over hACAT-1 were achieved as
hACAT-1. This was similar to the des-uoro derivative 19 sug- well as signicantly reducing in vivo clearance in the rat.
gesting that the 2-uoro substituent in the aniline ring was However, despite a moderate increase in permeability as
good for hACAT-1 activity. As observed with previous examples, measured in MDCK cells, solubility was also decreased so the
the corresponding trans-derivative 23 had signicantly reduced optimal prole was not achieved. The second strategy involved
potency towards hACAT-1. Introduction of a uoro group into replacing the benzene ring with pyridine in the side chain of 22
the meta-position relative to the amide (24) gave no benecial to bring the benets of the pyridyl but with minimal impact on
effect on either hDGAT-1 potency or solubility.
overall lipophilicity. This led to derivatives 27, 28 and 29. All
Further proling revealed that as the lipophilicity was three compounds had excellent potency against hDGAT-1,
reduced over one log unit compared to the starting point 1 then improved selectivity against hACAT-1 compared with 22 and
the impact on both permeability and clearance would be of maintained very low metabolic clearance in human hepato-
paramount importance (Table 3). Introduction of an oxy-link in cytes. Moreover, good improvements in both rat DMPK
combination with the 2,4,5-triuoroanilino moiety, as in 19, led parameters as well as a reduction in unbound clearance in the
dog were achieved for 28. In addition, 28 brought together all
strategies for improving solubility in this series by introducing
shape changes and reducing lipophilicity and attempting to
Table 3 DMPK parameters of leading derivatives
break the interactions in the crystal lattice through introducing
heteroatoms and masking the hydrogen bond donors. Gratify-
ingly 28 achieved excellent solubility balanced with good
permeability, which both contribute to the high bioavailability
observed in rat and dog DMPK experiments.
To assess the true impact on the interactions existing in the
solid state, crystals of 28 were grown which were suitable for
X-ray structural determination (Fig. 3).24 The X-ray structure
reveals that compared to the structure of 4, the void density has
increased from 10.1% of the unit cell to 12.7% of the unit cell,25
commensurate with a decreased packing efficiency. As before,
MDCK
Hu Hep
Rat Cl (Clu)
Dog Cl (Clu)
F% kgꢂ1
Papp A-B
Clint (mL
(mL minꢂ1 Rat (mL minꢂ1 Dog
10ꢂ6 (cm sꢂ1
)
minꢂ1 mgꢂ1
)
kgꢂ1
)
)
F%
19 11
<2
<2
<2
<2
<2
<2
1.1 (470)
7.4 (740)
0.4 (200)
0.7 (230)
3.6 (600)
1.8 (300)
115 11 (920)
33 4.7 (130)
85 6.5 (1700)
82 2.3 (100)
78 5.1 (300)
100 4.6 (200)
23
60
76
35
84
61
21 2.0
22 33a
26 5.1
27 16
28 15
a
Measured using Caco-2 cells.
This journal is ª The Royal Society of Chemistry 2013
Med. Chem. Commun., 2013, 4, 151–158 | 155

View Article Online
MedChemComm
Concise Article
Fig. 3 Two views of the crystal structure of compound 28. Panel (a) shows a view from above the plane of adjacent molecules and panel (b) shows how these planes
stack.
the ortho-uoro substituent masks the aniline NH but inter-
estingly the amide NH is not masked. Instead, uorine is
twisted away from the NH which ensures that while the NH is a
better donor, the pyridyl ring is twisted with respect to the
amide and so adopts a shape that packs less well in the solid
state. The consequence of all these changes is a much more
complex structure in which the plane of adjacent molecules
(Fig. 3a) is held together by van der Waals interactions,
including some p–p stacking. These planes are in turn held
together in a cross-hatched fashion because of the L shape of
the molecule which arises because of the axial positioning of
the oxy-pyridyl group. Each molecule is involved in a dimeric
pairing (Fig. 3b, the light blue and pink molecules) and these
dimers then present hydrogen bonding groups that interact
with the acids of adjacent molecules. These are difficult to
compare to the acid dimers in 4 but the data suggest that this
network of interactions contributes less to the lattice energy.
In general, compounds in this series showed no signicant
activity against human DGAT-2 (e.g. 26; IC50 ¼ 28 mM), the hERG
Fig. 4 PK/PD analysis showing the relationship between plasma triglycerides
(TAG) and free compound levels in plasma for 26 in a rat oral lipid tolerance test. A
direct response (Emax) model was used to fit the PK/PD data (see ESI†).
156 | Med. Chem. Commun., 2013, 4, 151–158
This journal is ª The Royal Society of Chemistry 2013

View Article Online
Concise Article
MedChemComm
encoded potassium channel (e.g. 26; IC50 > 30 mM), or the cyto- 3 D. Matsuda and H. Tomoda, Curr. Opin. Invest. Drugs, 2007,
chrome P450 enzyme isoforms CYP1A2, CYP2C9, CYP2C19,
CYP2D6 and CYP3A4 (e.g. 26; all IC50s > 10 mM). Compounds in
this series were also tested to conrm in vivo inhibition of DGAT-1
8, 836.
4 H. C. Chen and R. V. Farese, Arterioscler., Thromb., Vasc. Biol.,
2005, 25, 482.
using a rat oral lipid tolerance test.5 Treatment with 26 resulted in 5 A. M. Birch, S. Birtles, L. K. Buckett, P. D. Kemmitt,
an exposure-dependent decrease in circulating plasma triglyc-
eride levels. Fig. 4 shows the relationship between plasma
G. J. Smith, T. J. D. Smith, A. V. Turnbull and
S. J. Y. Wang, J. Med. Chem., 2009, 52, 1558.
triglycerides and free compound levels in plasma for 26 in this 6 W. McCoull, M. S. Addie, A. M. Birch, S. Birtles, L. K. Buckett,
model. PK-PD analysis indicates that the in vivo generated free
IC50 (0.2 nM) is in good agreement with the in vitro IC50 in rat
microsomes (0.9 nM) and a functional cell assay measuring
triglyceride synthesis in the HuTu80 human cell line (4 nM).
R. J. Butlin, S. S. Bowker, S. Boyd, S. Chapman,
R. D. M. Davies, C. S. Donald, C. P. Green, C. Jenner,
P. D. Kemmitt, A. G. Leach, G. C. Moody, P. Morentin
Gutierrez, N. J. Newcombe, T. Nowak, M. J. Packer,
A. T. Plowright, J. Revill, P. Schoeld, C. Sheldon, S. Stokes,
A. V. Turnbull, S. J. Y. Wang, D. P. Whalley and
J. M. Wood, Bioorg. Med. Chem. Lett., 2012, 22, 3873.
7 M. J. Waring, A. M. Birch, S. Birtles, L. K. Buckett, R. J. Butlin,
L. Campbell, P. Morentin Gutierrez, P. D. Kemmitt,
A. G. Leach, P. A. MacFaul, C. O’Donnell and
A. V. Turnbull, Med. Chem. Commun., submitted for
publication.
Conclusion
In summary, a novel series of hDGAT-1 inhibitors is described.
Design hypotheses were devised to introduce changes in shape
and polarity into the side chain, reduce lipophilicity and mask
the hydrogen bond donors with internal hydrogen bond
acceptors to improve the solubility and unbound clearance
prole of the clinical candidate 1. These changes led to the 8 R. L. Dow, J. Li, M. P. Pence, E. M. Gibbs, J. L. LaPerle,
replacement of the phenyl cyclohexyl-ethanoate side chain, a
fragment present in a number of hDGAT-1 inhibitors, with
substituted oxy-linked side chains. Combining the best features
of two sub-series, including balancing lipophilicity and
permeability, provided potent inhibitors of hDGAT-1 and dis-
played increased solubility and excellent ADMET parameters,
whilst maintaining good properties such as LLE. Simulta-
neously, greater selectivity against the related enzyme hACAT-1
J. Litcheld, D. W. Piotrowski, M. J. Munchhof,
T. B. Manion, W. J. Zavadoski, G. S. Walker,
R. K. McPherson, S. Tapley, E. Sugarman, A. Guzman-Perez
and P. DaSilva-Jardine, ACS Med. Chem. Lett., 2011, 2, 407.
9 R. L. Dow, M. Andrews, G. E. Aspnes, G. Balan, E. M. Gibbs,
A. Guzman-Perez, K. Karki, J. L. LaPerle, J. Li, J. Litcheld,
M. J. Munchhof, C. Perreault and L. Patel, Bioorg. Med.
Chem. Lett., 2011, 21, 6122.
was achieved. The comparison of the small molecule X-ray 10 V. S. C. Yeh, D. W. A. Beno, S. Brodjian, M. E. Brune,
structures of 4 and 28 shows marked differences in the solid
state. Compound 28 adopts a different shape due to the axial
positioning of the cis-oxy-pyridyl group. The planes of the
molecules are held together by van der Waals interactions,
including p–p stacking, and each molecule is involved in a
dimeric pairing leading to an overall decreased packing effi-
ciency of 28. Finally, inhibition of DGAT-1 in vitro translates to
S. C. Cullen, B. D. Dayton, M. K. Dhaon, H. D. Falls,
J. Gao, N. Grihalde, P. Hajduk, T. M. Hansen, A. S. Judd,
A. J. King, R. C. Klix, K. J. Larson, Y. Y. Lau, K. C. Marsh,
S. W. Mittelstadt, D. Plata, M. J. Rozema, J. A. Segreti,
E. J. Stoner, M. J. Voorbach, X. Wang, X. Xin, G. Zhao,
C. A. Collins, B. F. Cox, R. M. Reilly, P. R. Kym and
A. J. Souers, J. Med. Chem., 2012, 55, 1751.
the in vivo setting as compound 26 is shown to decrease circu- 11 http://clinicaltrials.gov/ct2/show/NCT01146522?term¼LCQ-
lating plasma triglyceride levels in an exposure-dependent
manner in a rat oral lipid tolerance test.
908&rank¼1.
12 log D, plasma-protein binding and solubility measurements
were made as described in: D. Buttar, N. Colclough,
S. Gerhardt, P. A. MacFaul, S. D. Phillips, A. T. Plowright,
P. Whittamore, K. Tam, K. Maskos, S. Steinbacher and
H. Steuber, Bioorg. Med. Chem., 2010, 18, 7486.
Acknowledgements
The research was sponsored by AstraZeneca. All authors were
employees of AstraZeneca at the time of the research. The authors 13 S. Cases, S. J. Smith, Y. W. Zheng, H. M. Myers, T.-Y. Chang,
thank Scott Boyd, Craig Donald, Kristin Goldberg and Adele Lloyd
for the synthesis and purication of test compounds, Usha
B.-L. Li, C. C. Y. Chang, Y. Urano and Y. Am., J. Endocrinol.
Metab., 2009, 297, E1–E9.
Chauhan for the testing of compounds in biological assays and 14 E. Floetmann, L. K. Buckett, A. V. Turnbull, C. Hammond,
colleagues in Physical Chemistry, DMPK and safety assessment
for the provision of secondary ADMET data.
C. Hallberg, A. Birch, D. Lees and H. B. Jones, manuscript
in preparation.
15 M. J. Waring, Bioorg. Med. Chem. Lett., 2009, 19, 2844.
16 X-ray data for compound 4: molecular formula
¼
Notes and references
C
23H21F3N4O4, C2H4O2, formula weight ¼ 534.491, crystal
˚
ꢀ
1 V. A. Zammit, L. K. Buckett, A. V. Turnbull, H. Wure and
A. Proven, Pharmacol. Ther., 2008, 118, 295.
2 B. K. Hubbard, I. Enyedy, T. A. Gilmore and M. Serrano-Wu,
Expert Opin. Ther. Pat., 2007, 17, 1331.
system ¼ triclinic, space group ¼ P1, a ¼ 5.1885(5) A, b ¼
˚
˚
11.4587(10) A, c ¼ 21.469(2) A, a ¼ 75.373(5), b ¼
3
˚
86.058(5), g ¼ 83.124(4), V ¼ 1225.2(2) A , T ¼ 200 K, Z ¼
2, Dc ¼ 1.449 Mg mꢂ3, (Mo-Ka) l ¼ 0.71073 A, m ¼ 0.12
˚
This journal is ª The Royal Society of Chemistry 2013
Med. Chem. Commun., 2013, 4, 151–158 | 157

View Article Online
MedChemComm
mmꢂ1
Concise Article
,
5552 reections measured, 3628 independent 22 K. D. Lardizabal, J. T. Mai, N. W. Wagner, A. Wyrick,
reections, 1988 observed reections [I> 2.0
R1_obs ¼ 0.053, goodness of t ¼ 0.92.
s
(I)],
T. Voelker and D. J. Hawkins, J. Biol. Chem., 2001, 276, 38862.
23 D. Dalvit and A. Vulpetti, ChemMedChem, 2012, 7, 262.
¨
´
17 J. Bostrom, A. Hogner, A. Llinas, E. Wellner and 24 X-ray data for compound 28: molecular formula
¼
A. T. Plowright, J. Med. Chem., 2012, 55, 1817.
C
21H17F4N5O5, formula weight ¼ 495.40, crystal system ¼
˚
18 P. C. Hariharan and J. A. Pople, Theor. Chim. Acta, 1973, 28, 213.
19 M. J. Frisch, G. W. Trucks, H. B. Schlegel, G. E. Scuseria,
M. A. Robb, J. R. Cheeseman, G. Scalmani, V. Barone,
B. Mennucci, G. A. Petersson, H. Nakatsuji, M. Caricato,
X. Li, H. P. Hratchian, A. F. Izmaylov, J. Bloino and
G. Zheng, et al., Gaussian 09, Revision A.1, Gaussian, Inc.,
Wallingford CT, 2009.
monoclinic, space group ¼ C2/c, a ¼ 29.7773(14) A, b ¼
˚
˚
6.3200(3) A, c ¼ 23.2587(11) A, b ¼ 100.671(3), V ¼
4301.4(4) A , T ¼ 200 K, Z ¼ 8, Dc ¼ 1.530 Mg mꢂ3, (Mo-
3
˚
Ka) l ¼ 0.71073 A, m ¼ 0.13 mmꢂ1, 55003 reections
˚
measured, 4007 independent reections, 2557 observed
reections [I> 2.0 s (I)], R1_obs ¼ 0.043, goodness of t ¼
1.02.
˚
20 P. W. Kenny, J. Chem. Soc., Perkin Trans. 2, 1994, 199.
21 P. W. Kenny, J. Chem. Inf. Model., 2009, 49, 1234.
25 Computed in mercury with a probe radius of 0.3 A on a grid
˚
of 0.2 A spacing on the solvent accessible surface.
158 | Med. Chem. Commun., 2013, 4, 151–158
This journal is ª The Royal Society of Chemistry 2013