Structure-Activity Relationship Explorations and Discovery of a Potent Antagonist for the Free Fatty Acid Receptor 2

Article abstract of DOI:10.1002/cmdc.202100356

Free fatty acid receptor 2 (FFA2) is a sensor for short-chain fatty acids that has been identified as an interesting potential drug target for treatment of metabolic and inflammatory diseases. Although several ligand series are known for the receptor, the

Full text of DOI:10.1002/cmdc.202100356

Accepted Article
Title: Structure-Activity Relationship Explorations and Discovery of a
Potent Antagonist for the Free Fatty Acid Receptor 2
Authors: Anders Højgaard Hansen, Henriette B. Christensen, Sunil
K. Pandey, Eugenia Sergeev, Alice Valentini, Julia Dunlop,
Domonkos Dedeo, Simone Fratta, Brian D. Hudson, Graeme
Milligan, Trond Ulven, and Elisabeth Rexen Ulven
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To be cited as: ChemMedChem 10.1002/cmdc.202100356
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01/2020

10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
Structure-Activity Relationship Explorations and Discovery of a
Potent Antagonist for the Free Fatty Acid Receptor 2
Anders Højgaard Hansen,[b]# Henriette B. Christensen,[b]# Sunil K. Pandey,[b] Eugenia Sergeev,[c] Alice
Valentini,[a] Julia Dunlop,[c] Domonkos Dedeo,[c] Simone Fratta,[a] Brian D. Hudson,[c] Graeme Milligan,[c]
Trond Ulven*[a],[b] and Elisabeth Rexen Ulven*[a]
[a]
A. Valentini, S. Fratta, Prof. T. Ulven, Prof. E. Rexen Ulven
Department of Drug Design and Pharmacology
University of Copenhagen
Universitetsparken 2, DK-2100 Copenhagen, Denmark
E-mail: tu@sund.ku.dk; eru@sund.ku.dk
[b]
[c]
Dr. A. Højgaard Hansen, H. B. Christensen, Dr. S. K. Panday, Prof. T. Ulven
Department of Physics, Chemistry and Pharmacy
University of Southern Denmark
Campusvej 55, DK-5230 Odense M, Denmark
Dr. E. Sergeev, J. Dunlop, D. Dedeo, Dr. B. D. Hudson, Prof. G. Milligan
Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology
College of Medical, Veterinary and Life Sciences, University of Glasgow
Glasgow G12 8QQ, Scotland, United Kingdom
Supporting information for this article is given via a link at the end of the document.
#These authors contributed equally
Abstract: Free fatty acid receptor 2 (FFA2) is a sensor for short-chain
fatty acids that has been identified as an interesting potential drug
target for treatment of metabolic and inflammatory diseases. Although
several ligand series are known for the receptor, there is still a need
for improved compounds. One of the most potent and frequently used
antagonists is the amide-substituted phenylbutanoic acid known as
CATPB. We here report the structure-activity relationship exploration
of this compound, leading to the identification of homologues with
increased potency. The preferred compound TUG-1958 was found,
besides improved potency, to have high solubility and favorable
pharmacokinetic properties.
reach endpoints for treatment of ulcerative colitis.[12] Reports also
indicate that FFA2 antagonism may counteract NSAID-induced
enteropathy and psoriasis.[13] Intriguingly, FFA2 was recently
found to be implicated in cell entry of the influenza A virus,
implying that FFA2 antagonists may have effects against
influenza A.[8a] Thus, the potential of FFA2 antagonists as
therapeutic agents is considered to be high in several areas, but
more research is required to investigate and realize this potential.
O
CF3
O
OH
N
N
O
HN
N
H
O
O
Cl
N
H
O
N
H
S
O
OH
N
Cl
1 (CATPB)
2 (BTI-A-404)
3 (GLPG0974)
Introduction
Figure 1. FFA2 antagonists.
The free fatty acid receptor 2 (FFA2) and the closely related FFA3
are G protein-coupled receptors that sense short-chain fatty acids
(SCFAs) such as acetate, propionate and butyrate,[1] and that
have been implicated in mediation of the physiological effects of
SCFA-producing gut microbiota.[2] FFA2 is relatively broadly
expressed in immune cells, intestine, adipose tissue and
pancreas,[3] and has been suggested as a target for treatment of
metabolic disorders, such as type 2 diabetes[4] and chronic kidney
disease,[5] inflammatory disorders, including inflammatory bowel
disease[6] and allergic asthma,[7] as well as infectious diseases.[8]
Antagonists were suggested for treatment of type 2 diabetes
based on results from FFA2 and FFA3 knockout mice.[9] The
observation that the FFA2 inverse agonist BTI-A-404 (2, Figure 1)
stimulates GLP-1 secretion in a cellular assay provides some
support for this,[10] although FFA2 agonists also have been
reported to stimulate GLP-1 secretion.[11] FFA2 antagonists are
also of interest for treatment of inflammatory bowel disease. As
the only compound in clinical trials to date, the antagonist
GLPG0974 (3, Figure 1), was found to inhibit migration and
activation of neutrophils in humans, even if this was insufficient to
The FFA2 antagonist CATPB (1, Figure 1) was initially disclosed
in a patent application from Euroscreen,[14] and subsequently
synthesized and characterized by us as an antagonist and inverse
agonist of FFA2.[15] Compared head-to-head, 1 and 3 were found
to be essentially equipotent as antagonists in both an inositol
phosphate assay and in a radioligand displacement binding
assay,[16] however, 1 exhibited considerably higher on- and off-
rates than 3.[16b] Partly because we were interested in better
understanding the basis for these differences, we conducted
structure-activity relationship investigations on 1. These
investigations led to the identification of homologues of 1 with an
extended carboxylic acid chain that have increased potency, as
presented herein. The preferred compound 37 (TUG-1958) was
found to have excellent solubility and favorable pharmacokinetic
properties.
1
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10.1002/cmdc.202100356
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CF3
CF3
Results and Discussion
(a), (b)
O
O
Compounds were initially synthesized as described by Brantis et
al.,[14] by condensation of arylacetic acid with malonate and
decarboxylation to form methyl 4-arylacetoacetate,[17] followed by
imine formation, catalytic hydrogenation under elevated pressure,
acylation of the resulting amine, and ester hydrolysis.[14] As we
observed some problems with the reproducibility of this protocol,
we explored alternative routes. A number of synthetic strategies
exist in the literature for producing β3-amino acid derivatives such
as 6 (Scheme 1). Besides the Arndt-Eistert homologation of α-
amino acids,[18] methods include enantioselective hydrogenation
of enamines[19] or ketones,[20] and Ellman’s sulfinamides.[21] The
latter was explored, as it offers a safe and convenient method to
access β-amino acid derived substrates without the use of either
toxic/explosive diazomethane or pressurized hydrogen.[22]
H2N
BocHN
OH
OMe
8
7
(c), (d)
CF3
CF3
(e)
BocHN
H2N
HCl
10
O
O
9
OEt
OEt
Scheme 2. Synthesis of γ4 -amino acid ethyl ester intermediates 10. Reagents
and conditions: (a) SOCl2, MeOH, 0 °C (5 min) → rt, 12 h, 85%. (b) Boc2O, Et3N,
DCM, rt, 22 h, 87%. (c) (i) DIBALH, -78 °C (30 min), toluene; (ii)
EtOC(O)CH2PPh3Br, KOtBu, DCM, -78 °C (1 h) → rt, 12 h, 59%. (d) Pd/C, H2,
EtOH, rt, 12 h, 86%. (e) HCl, dioxane, 0 °C (5 min) → rt, 99%.
Central intermediate 6 (Scheme 1) was synthesized by
reduction of 4-(trifluoromethyl)phenylacetic acid with LiAlH4 and
subsequent oxidation using 2-iodoxybenzoic acid (IBX)[23] to give
2-(4-(trifluoromethyl)phenyl)acetaldehyde, which was condensed
with (S)-(−)-2-methyl-2-propanesulfinamide using catalytic
CF3
CF3
O
(a), (b)
HCl
H2N
(
)
n
(
)
n
R
'
N
H
n = 1, R = Me (6)
n = 2, R = Et (10)
n = 0, R = Me (11)
O
OR
O
OH
pyridinium
p-toluenesulfonate
(PPTS)[24]
to
give
the
corresponding N-tert-butanesulfinyl imine 4 in 57% yield over two
steps. Next, diastereoselective addition of dimethyl malonate
(DMM) to neat 4 in presence of NaI[22a] produced 5 in moderate
yield. Treatment of 5 with refluxing aqueous acid caused amine
deprotection, ester hydrolysis, decarboxylation, and re-
esterification of the free β-amino acid to produce 6 in 91% yield
was achieved by addition of methanol to the refluxing reaction
mixture.
(c), (b)
CF3
O
O
S
R
N
H
O
OH
12
Scheme 3. Synthetic route for N-acylated and N-sulfonated amino acids.
Reagents and conditions: (a) (i) Carboxylic acid, EDC, HOBt, DIPEA, (DMAP),
DCM, 0 °C, 3.5 h; (ii) amine hydrochloride, 0 °C → rt, 8 h−2 d, 27−81%. (b) LiOH
(aq), THF, rt, 58−99%. (c) m-Tolylmethanesulfonyl chloride, Et3N, MeCN, rt (24
h) → 70 °C (48 h), 38%.
CF3
CF3
O
O
S
(a), (b), (c)
tBu
N
HO
4
(d)
CF3
CF3
The corresponding reverse amide of 1 was accessed as a
racemate by coupling of 13[26] with 3-chlorobenzylamine using a
O
S
(e)
COOMe
COOMe
H2N
tBu
N
bis(tetramethylene)fluoroformamidinium
hexafluorophosphate
H
COOMe
6
5
(BTFFH) promoted amide coupling[27] to give the amide tert-butyl
ester, which upon treatment with TFA in DCM produced the
desired carboxylic acid 14 (Scheme 4).
Scheme 1. Synthesis of β3-amino acid methyl ester intermediate 6. Reagents
and conditions: (a) LiAlH4, THF, rt, 91%. (b) IBX, AcOH, MeCN, rt → 70 °C, 3 h.
(c) (S)-(−)-2-Methyl-2-propanesulfinamide, PPTS, DCM, 12 h, rt, 67% (2 steps).
(d) DMM, NaI, neat, rt, 2 d, 40%. (e) (i) 6 M HCl, 110 °C, 3 h; (ii) 6 M HCl/MeOH,
110 °C, 12 h, 91%.
CF3
CF3
(a), (b)
H
N
HO
Cl
O
O
O
O
O
OH
13
14
g-Amino acid intermediate 10 was accessed in five steps from
the commercially available 7 (Scheme 2). Esterification of 7 using
thionyl chloride in MeOH followed by Boc protection in DCM
afforded 8, which was further reduced to the aldehyde with
DIBALH and in the same pot subjected to a Wittig reaction to give
the α,β unsaturated ethyl ester. Hydrogenation gave 9, and Boc-
deprotection afforded the desired (R)-γ4-amino acid ethyl ester
intermediate 10 in good yield (86%).[25]
Next, intermediate amines were coupled with carboxylic acids
to give N-acylated α, β3 and γ4-amino acid esters. Hydrolysis with
LiOH afforded the final carboxylic acids (Scheme 3). The
sulfonamide 12 was obtained in two steps starting from
intermediate 6 by reaction with m-tolylmethanesulfonyl chloride
followed by ester hydrolysis.
Scheme 4. Synthesis of 14. Reagents and conditions: (a) (i) BTFFH, DIPEA,
DCM, rt, 45 min; (ii) 3-chlorobenzylamine, 80 °C, 16 h, 43%. (b) TFA, DCM, rt,
2 h, 74%.
Compounds were tested in a [35S]GTPgS incorporation
assay,[26] well suited for Gi-coupled receptors, using membranes
of Flp-In T-Rex 293 cells induced to express human FFA2.[15]
CATPB (1) is an antagonist in this assay, capable of completely
inhibiting the effect of propionate with pIC50 = 6.65 ± 0.06 (Table
1).
2
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10.1002/cmdc.202100356
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the electron withdrawing chloro by the similarly sized electron
donating methyl (19) maintained potency, whereas larger meta-
substituents, including the electron withdrawing, hydrophobic
trifluoromethyl (20), the more polar, electron donating methoxy
(21), and the planar naphthyl (22), benzothiophene (23) and
indole (24) led to decreased potency, suggesting that increased
steric bulk is generally disfavored, whereas electron donating or
withdrawing properties seem less important.
Table 1. SAR exploration of the acetamide aryl group.
CF3
O
R
N
H
O
OH
cmpd
R
hFFA2
GTPγS[a]
pIC50
ClogP[b]
LLE[c]
Table 2. SAR exploration of the acetamide linker.
1
6.65 ± 0.06
6.07 ± 0.07
6.11 ± 0.11
6.53 ± 0.05
6.69 ± 0.21
6.12 ± 0.20
5.87 ± 0.13
5.90 ± 0.22
4.21
4.21
4.21
3.49
4.00
4.38
3.41
4.67
2.44
1.86
1.90
3.04
2.69
1.74
2.46
1.23
Cl
CF3
16
17
18
19
20
21
22
R
Cl
O
OH
Cl
cmpd
R
hFFA2
GTPγS[a]
pIC50
ClogP[b]
LLE[c]
O
25
Cl
5.58 ± 0.04
4.81 ± 0.05
~3.9
4.59
4.40
1.78
4.06
4.40
1.00
0.41
2.1
N
H
O
26
Cl
N
H
F3C
O
27
N
H
MeO
O
O
12
5.34 ± 0.08
5.21 ± 0.12
1.28
0.81
S
N
H
H
N
14 (rac)
Cl
23
24
6.11 ± 0.11
5.38 ± 0.12
4.52
3.48
1.59
1.90
O
S
[a] [35S]GTPγS-based binding assay measured with 0.3 mM propionate. Data (-
log M) is shown as mean ± SEM, n ³ 3. [b] ClogP values were calculated using
ChemBioDraw v. 16.0. [c] LLE values (LLE = pIC50−ClogP)
HN
Extension of the acetamide linker of 1 to the corresponding
propanamide 25 (Table 2) resulted in an order of magnitude loss
in potency, supporting the observation that increasing the size of
the phenylacetamide group is disfavored. Shortening to a m-
chlorobenzamide (26) led to >50-fold loss of potency, and
removal of the chlorophenyl to leave only the acetamide (27) gave
a >500-fold loss, indicating that the aromatic ring of the amide
side chain and its position is important to the overall potency of
this compound series. Conversion of the amide to a sulfonamide
(12) or reversing the amide (14) both resulted in significant loss of
potency. Overall, the SAR requirements of the phenylacetamide
fragment of 1 is relatively restricted with regard to both linker and
substitution pattern, with a small electron withdrawing or donating
substituents in the meta-position showing moderately increased
potency and all other modifications resulting in loss of activity.
Shifting attention to the carboxylic acid part, we found that
shortening by one methylene to give the phenylalanine derivative
28 (Table 3) led to a 2-fold decrease in potency. In contrast,
elongation by one methylene group (29) gave a compound that
was twice as potent as 1. The same trend was observed when the
methyl-substituted analogue 19 was shortened (30) or elongated
(31), confirming increased potency of the extended carboxylate
compounds.
[a] [35S]GTPγS-based binding assay measured with 0.3 mM propionate. Data
(-log M) is shown as mean ± SEM, n ³ 3. [b] ClogP values were calculated
using ChemBioDraw v. 16.0. [c] LLE values (LLE = pIC50−ClogP).
The methyl ester of 1 (intermediate 15) was also tested (pIC50
= 6.08 ± 0.03), confirming that the carboxylic acid is not critically
important for activity,[16b] as also observed for Galapagos’
azetidine amide FFA2 antagonist series,[6,28] but still resulted in a
reduction of potency by half a log unit, and considerably increased
lipophilicity as the ionizable group was lost.
The SAR exploration and optimization followed an iterative
analysis and design – synthesize – test cycle. We initially explored
the phenylacetamide part of 1, as the most accessible part with
intermediate 6 in hand. Isomers with the chloro-substituent in the
ortho- (16) or para-position (17) both showed a 3-fold decreased
potency relative to 1 (Table 1). In contrast, removal of the m-
chloro of 1 to give 18 only resulted in a minor loss of potency but
an increase in ligand lipophilicity efficiency (LLE),[29] as calculated
from ClogP values, due to the reduced lipophilicity. Thus, the m-
chloro of the phenylacetamide fragment at best contributes only
moderately to potency, whereas the o-chloro and the p-chloro
have negative effects on ligand activity.
Since
a
substituent in the meta-position of the
phenylacetamide appeared to be favored, we directed our
attention at exploring substituents in this position. Replacement of
3
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Table 3. SAR exploration of the linker to the carboxylate.
Table 4. SAR exploration of the extended carboxylate.
R2
CF3
O
O
R1
N
H
R
N
H
( )
n
OH
O
OH
O
cmpd
R1
R2
hFFA2
GTPγS[a]
pIC50
ClogP[b]
LLE[c]
cmpd
n
R
hFFA2
GTPγS[a]
pIC50
ClogP[b]
LLE[c]
32
33
34
28
29
30
31
0
2
0
2
Cl
Cl
6.32 ± 0.06
7.00 ± 0.21
6.22 ± 0.07
7.16 ± 0.06
4.10
4.52
3.39
4.31
2.22
2.48
2.83
2.85
CF3
CF3
7.04 ± 0.08
7.15 ± 0.10
4.69
3.73
2.35
3.42
F3C
MeO
Me
Me
CF3
CF3
CF3
6.68 ± 0.12
4.18 ± 0.13
6.58 ± 0.10
4.98
6.42
4.44
1.70
-2.24
2.14
[a] [35S]GTPγS-based binding assay measured with 0.3 mM propionate. Data
(-log M) is shown as mean ± SEM, n ³ 3. [b] ClogP values were calculated
using ChemBioDraw v. 16.0. [c] LLE values (LLE = pIC50−ClogP).
35
36
To investigate if the SAR observed for the CATPB analogues
was directly transferable to the elongated analogues, we
synthesized and tested the m-trifluoromethyl (32), m-methoxy
(33) and 1-naphthyl (34) analogues. Interestingly, 32 and 33
maintained potency similar to 29 and 31, in contrast to the more
than half a log unit reduced potency observed for 20 and 21. Thus,
SAR is not directly transferable from the original series to the
elongated carboxylic acid analogues. The naphthyl derivative 34
gave a moderate but significant decrease in potency, as also seen
in the original series (22, Table 1), whereas extension of the meta
substituent to pentyl derivative 35 resulted in a virtually inactive
37
38
39
CF3
CF3
7.41 ± 0.27
7.29 ± 0.18
3.95
4.02
3.46
3.27
F
F
F
F
CF3
7.16 ± 0.06
4.09
3.07
F
F
F
F
compound, demonstrating that
a restriction of the meta-
substituent size still is in operation. Introducing a methyl-branch
in 36, which was tested as a diastereomeric mixture, produced a
reasonably potent compound but would not represent an
improvement even if all activity was ascribed to only one of the
diastereomers.
40
41
42
CF3
CF3
H
6.97 ± 0.02
6.69 ± 0.03
5.55 ± 0.00
4.02
4.09
3.07
2.95
2.60
2.48
F
F
Since the elongated compound series showed higher
tolerance towards phenylacetamide substituents, we decided to
also explore m-fluoro derivative (37), as this substituent
represents only a moderately increased lipophilicity and protects
the phenyl against metabolism. To our delight, 37 resulted in the
most potent compound with a pIC50-value of 7.41 and also the
highest LLE-value. (It should be noted that LLE values are based
on calculated ClogP values. Estimated values of log D7.4 for
propanoic acids can be obtained by adding ~2.7.[30]) This result
prompted us to also explore the difluorinated analogues 38-41.
Compared to 37, an additional fluoro-substituent did not increase
potency for any of the compounds, and those with an ortho-fluoro
(40 and 41) showed distinctly reduced potency. Finally, probing
the importance of the para-trifluoromethyl at the other ring, we
found that its removal in 42 produced a compound with potency
reduced almost two orders of magnitude. Although the
simultaneous reduction in lipophilicity resulted in only one order
of magnitude reduced LLE compared to 37, it is clear that the
trifluoromethyl is important for the antagonist activity of the
compounds. Altogether, the elongated antagonist series shows
SAR features that are similar but not identical to the original
CATPB series, with a phenylacetamide more accommodating to
modifications and, notably, in general significantly increased
potency.
[a] [35S]GTPγS-based binding assay measured with 0.3 mM propionate. Data
(-log M) is shown as mean ± SEM, n ³ 3. [b] ClogP values were calculated
using ChemBioDraw v. 16.0. [c] LLE values (LLE = pIC50−ClogP).
We recently established a BRET binding assay based on
hFFA2 fused with nanoluciferase (NLuc) in the N-terminal and the
fluorescent antagonist tracer TUG-1609.[28] Selected compounds
were evaluated in this assay (Figure 2). The longer meta-chloro
substituted 29 showed the highest affinity in this assay, whereas
the shorter meta-chloro substituted 1 was second, both with pKi
values shifted ~0.35 relative to their GTPgS pIC50 value. On the
other hand, all fluoro-substituted compounds show pKi values
lower than their GTPgS pIC50 value and excellent correlation with
GTPgS values internally in the sub-series. Thus, the results
indicate that the chloro substituent conveys affinity that is not
translated to functional activity, whereas the binding of the fluoro-
substituted compounds is more effectively translated to
a
functional response. This may be explained by a higher ability of
the chloro-substituted compounds to effectively displace TUG-
1609. Further studies are required to elucidate the basis of this
substituent effect, but it may be related to subtle differences in the
binding site and/or differences in kinetic properties.
4
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Figure 2. Competition binding with TUG-1609 in a BRET assay with NLuc-hFFA2. Left: pKi values (-log M, mean ± SEM) and deviation from GTPgS pIC50 values.
Right: Plot of pKi against GTPgS pIC50 for chloro-substituted (blue) and fluoro-substituted (orange) compounds, trendlines through origo are shown.
Compound 1, 28, 29 and 37 were docked in a homology
model of hFFA2 based on the crystal structure of the closely
related hFFA1 (PBD: 5TZY). The carboxylate of 1 is embedded in
a polar network consisting of both hydrogen bonds and ionic
interactions with Arg180, Arg255, Tyr90 and Lys65 (Figure 3A).
Furthermore, there is hydrogen bond interaction between the
amide carbonyl and Ser86. The p-trifluoromethylphenyl is fitting
perfectly into the hydrophobic pocket between TM2 and TM3,
explaining why removal of the p-trifluoromethyl led to a significant
loss of potency. The phenylacetamide part of the ligand is fitted
into a pocket towards TM4 and the size and shape of the binding
cavity also explains why larger substituents are not favored and
that the meta-position seems to be best suited for substituents.
Shortening of the acid linker by one carbon (28, Figure 3B)
resulted in docking poses with inversion of the placements of the
two aromatic parts in the hydrophobic binding cavities and loss of
several of the polar interactions. On the other hand, elongating
the acid linker by one carbon (29, Figure 3C) kept the overall
binding pose observed with 1, but the extended space between
the acid and amide resulted in additional H-bond interactions
between the amide and Arg180 and NH-Ser86, and almost
perfect filling of hydrophobic pockets, providing a rationale for the
increased potency. The same pose was found for 37 (Figure 3D)
but with the m-fluoro filling another pocket than the m-chloro of 29.
Figure 3. Docking of A) 1, B) 28, C) 29 and D) 37 into a homology model of hFFA2 based on hFFA1 (PBD: 5TZY).
5
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39.52 ppm; 1H NMR (CD3OD): 3.31 ppm; 13C NMR (CD3OD): 49.00 ppm;
As the most potent functional antagonist with the highest LLE,
37 was selected for further investigations. Compound 37 was
completely selective over the related receptor FFA3 with no sign
of activity up to 100 µM. The compound exhibited excellent kinetic
solubility of 194 µM. Investigations of in vitro drug properties of 37
revealed high metabolic stability towards mouse liver microsomes,
with 95% remaining after 40 min incubation.
The pharmacokinetic properties of 37 was studied in mice,
revealing favorable properties with a half-life of almost two hours
and an oral bioavailability of 82% (Table 6).
High-resolution mass spectra (HRMS) were recorded on
a Bruker
micrOTOF-Q II (ESI) or QExactive Orbitrap mass spectrometer (MALDI).
Melting point ranges were determined on a Standford Research Systems
MPA100 using a ramp rate of 1 oC/min Specific optical rotation was
recorded on Anton Paar MCP 100 polarimeter. Purity was determined by
HPLC and confirmed by inspection of NMR spectra (1H and 13C NMR).
HPLC analysis was performed using a Dionex 120 or Gemini C18 column
(5 μm, 4.6x150 mm); flow: 1 mL/min; 10% MeCN in water (0-1 min), 10-
100% MeCN in water (1-10 min), 100% MeCN (11-15 min), with both
solvents containing 0.1% formic acid or 0.01% trifluoroacetic acid as
modifier; UV detection at 254 nm. All test compounds were of ≥95% purity.
Table 6. Pharmacokinetic properties in mice.[a]
(S)-3-(2-(3-Chlorophenyl)acetamido)-4-(4-(trifluoromethyl)phenyl)-
butanoic acid (1). Step 1: 2-(4-(Trifluoromethyl)phenyl)ethan-1-ol. LiAlH4
(3.76 g, 99.1 mmol) was added to a pre-dried round-bottom flask and the
flask was flushed with argon (x3). The flask was cooled to 0 °C and LiAlH4
was dissolved in dry THF (55 mL) and subsequently added 4-
(trifluoromethyl)phenylacetic acid (6.74 g, 33.0 mmol) dissolved in dry THF
(55 mL) (addition over 30 min). The reaction mixture was allowed to reach
rt and stirred for additional 3 h. At 0 °C, the reaction mixture was diluted
with wet THF (50 mL) and worked up according to the Feiser method, i.e.
dropwise addition of water (3.6 mL, 20 min), 15% NaOHaq (3.6 mL, 20 min),
and water (10.7 mL, 20 min). MgSO4 was added and the mixture stirred
for 15 min at rt, filtered through a glass filter funnel, washed with DCM (40
mL), and the filtrate concentrated in vacuo. The product was obtained after
flash chromatography (DCM) as a clear liquid (5.74 g, 91% yield): Rf = 0.19
(DCM); 1H NMR (400 MHz, CDCl3) δ 7.57 (d, J = 8.2 Hz, 2H), 7.35 (d, J =
8.1 Hz, 2H), 3.88 (t, J = 6.5 Hz, 2H), 2.92 (t, J = 6.5 Hz, 2H), 1.54 (s, 1H);
13C NMR (101 MHz, CDCl3) δ 143.0 (q, J = 1.3 Hz), 129.5 (2C), 129.0 (q,
J = 32.4 Hz), 125.6 (q, J = 3.8 Hz, 2C), 124.4 (q, J = 271.8 Hz), 63.4, 39.1;
EI-MS (70 eV) m/z 190 (M+). 1H NMR spectra in agreement with the
literature.[31]
parameters
37
Intravenous Administration (5 mg/kg)
112 min
t1/2
AUC0−∞
Vd
1620000 ng min mL−1
498 mL kg−1
CLtotal
3.09 mL min−1 kg−1
Oral Administration (10 mg/kg)
30 min
tmax
Cmax
AUC0−∞
F
8950 ng mL−1
2660000 ng min mL−1
82%
Step 2: 2-(4-(Trifluoromethyl)phenyl)acetaldehyde. To a solution of the
alcohol (5.07 g, 26.7 mmol) in MeCN (120 mL) were added AcOH (1.8 mL,
32.0 mmol) and IBX (10.2 g, 36.4 mmol) and the reaction stirred at rt for
30 min. The reaction was heated slowly to 70 °C and stirred for additional
3 h. NaHCO3 (4.0 g, 47.6 mmol) was added and the reaction stirred for 30
min, filtered through a plug of silica, washed with 20 mL DCM (x4) and the
filtrate concentrated in vacuo. The crude aldehyde was used in the next
step without further purification.
[a] Administered at 10 mg/kg po and 5 mg/kg iv in Balb/c mice.
Conclusion
In conclusion, our SAR studies of 1 resulted in the observation
that homologs with the carboxylic acid chain extended by one
methylene group have increased potency and the identification of
TUG-1958 (37) as the overall preferred compound. Further
studies found the compound to have excellent solubility and
pharmacokinetic properties in mice. Altogether, 37 represents a
significantly improved FFA2 antagonist.
Step
3:
(S,E)-2-Methyl-N-(2-(4-(trifluoromethyl)phenyl)ethylidene)-
propane-2-sulfinamide (4). In a pre-dried round-bottom flask under argon,
2-(4-(trifluoromethyl)phenyl)acetaldehyde (5.02 g), (S)-(−)-2-methyl-2-
propanesulfinamide (3.25 g, 26.8 mmol), and pyridinium p-
toluenesulfonate (278 mg, 1.11 mmol) were added dry DCM (53 mL) and
the reaction was stirred overnight under protection from light. The reaction
mixture was filtered through a pad of Celite, washed with 75 mL DCM (x4),
and the filtrate concentrated in vacuo.
4 was obtained after flash
chromatography (EtOAc/PE, 1:5) as a light yellow oil (4.45 g, 57% yield):
Rf = 0.11 (EtOAc/PE, 1:5); 1H NMR (400 MHz, CDCl3) δ 8.14 (t, J = 5.0 Hz,
1H), 7.60 (d, J = 8.2 Hz, 2H), 7.35 (d, J = 8.1 Hz, 2H), 3.91 – 3.87 (m, 2H),
1.18 (s, 9H); 13C NMR (101 MHz, CDCl3) δ 166.5, 139.1 (app s), 129.7
(2C), 129.7 (q, J = 32.6 Hz), 125.8 (q, J = 3.8 Hz, 2C), 124.2 (q, J = 271.9
Hz), 57.1, 42.3, 22.4 (3C); HRMS (ESI) calcd for C13H16F3NNaOS (M +
Na+) 314.0797, found 314.0796.
Experimental Section
Synthesis
All commercial starting materials and solvents were used without further
purification unless otherwise stated. THF was freshly distilled from
sodium/benzophenone. MeOH was freshly distilled from Mg. DCM was
freshly distilled and stored over 4Å sieves. TLC was performed on TLC
Silica gel 60 F254 plates and visualized at 254 nm or by staining with
ninhydrin, KMnO4 or FeCl3 stains. Petroleum ether (PE) refers to alkanes
with bp 60-80 °C. Water used in reactions was demineralized and water
used for freeze-drying was filtered Milli-Q. Purification by flash
chromatography was carried out using silica gel 60 (0.040-0.063 mm,
Merck). 1H and 13C NMR spectra were recorded at 400 and 101 MHz,
respectively, on a Bruker Avance III 400 at 300 K. Spectra were calibrated
relative to residual solvent peaks: 1H NMR (CDCl3): 7.26 ppm; 13C NMR
(CDCl3): 77.16 ppm; 1H NMR (DMSO-d6): 2.50 ppm; 13C NMR (DMSO-d6):
Step
4:
Dimethyl
2-((S)-1-(((S)-tert-butylsulfinyl)amino)-2-(4-
(trifluoromethyl)phenyl)ethyl)malonate (5). To a pre-dried microwave vial
were added 4 (4.45 g, 15.3 mmol), dimethyl malonate (2.9 mL, 30.6 mmol)
and NaI (2.29 g, 15.3 mmol). The vial was capped, flushed with argon (x3),
protected from light and the reaction was stirred for two days under argon.
The reaction mixture was diluted with water (75 mL), acidified with 1 M
HClaq (25 mL) and extracted with EtOAc (x3). The combined organic layers
were washed with brine, dried over Na2SO4, and concentrated in vacuo. 5
was obtained after flash chromatography (EtOAc/PE, 1:1) as a sticky oil
6
This article is protected by copyright. All rights reserved.

10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
(2.59 g, 40% yield): Rf = 0.14 (EtOAc/PE, 1:1); 1H NMR (400 MHz, CDCl3)
δ 7.54 (d, J = 8.0 Hz, 2H), 7.31 (d, J = 7.9 Hz, 2H), 4.55 (d, J = 9.4 Hz, 1H),
4.15 – 4.05 (m, 1H), 3.85 – 3.82 (m, 1H), 3.80 (s, 3H), 3.79 (s, 3H), 3.05 –
2.90 (m, 2H), 1.034 (s (shoulder), 3H), 1.029 (s, 6H); 13C NMR (101 MHz,
CDCl3) δ 168.6, 168.1, 142.1 (app s), 129.9 (2C), 129.2 (q, J = 32.4 Hz),
125.4 (q, J = 3.7 Hz, 2C), 124.2 (q, J = 271.8 Hz), 58.2, 56.2, 55.4, 53.0,
52.7, 40.1, 22.4 (3C); HRMS (ESI) calcd for C18H24F3NNaO5S (M + Na+)
446.1219, found 446.1235.
(S)-3-((m-Tolylmethyl)sulfonamido)-4-(4-(trifluoromethyl)-
phenyl)butanoic acid (12). Step 1: Under an inert atmosphere (argon), 6
(25.0 mg, 84 μmol) and Et3N (23 μL, 0.17 mmol) dissolved in dry MeCN
(60 μL) were added a 3 M solution of m-tolylmethanesulfonyl chloride (28
μL, 84 μmol) in dry MeCN and the reaction was stirred at rt over night. The
next day, the reaction was heated to 70 °C for 2 h, then diluted in DCM
and evaporated onto celite. The methyl ester of 12 of the title compound
was obtained after flash chromatography (EtOAc/PE, 1:3) as a sticky oil
(14 mg, 38% yield): Rf = 0.13 (EtOAc/PE, 1:3); 1H NMR (400 MHz, CDCl3)
δ 7.55 (d, J = 8.1 Hz, 2H), 7.25 – 7.15 (m, 5H), 7.11 (d, J = 7.5 Hz, 1H),
4.89 (d, J = 9.5 Hz, 1H), 4.17 – 4.06 (m, 2H), 3.81 – 3.72 (m, 1H), 3.70 (s,
3H), 2.97 (dd, J = 13.6, 5.9 Hz, 1H), 2.88 (dd, J = 13.6, 8.3 Hz, 1H), 2.44
Step 5: Methyl (S)-3-amino-4-(4-(trifluoromethyl)phenyl)butanoate
hydrochloride (6). To a round-bottomed flask was added 5 (725 mg, 1.71
mmol) and 6 M HClaq (6.8 mL), and the suspension was stirred at 110 °C
for 3 h to give a clear yellow solution. The reaction was cooled to rt, added
MeOH (27 mL) and stirred at 110 °C over night. The reaction mixture was
cooled to rt, added NaHCO3aq (sat) (20 mL), extracted with EtOAc (x3),
and dried over Na2SO4. The crude product was concentrated in vacuo, re-
dissolved in dioxane (2 mL), cooled to 0 °C, added HCl in dioxane (4 M,
0.6 mL) before concentration in vacuo to give a light yellow solid (465 mg,
91% yield): Rf (free amine) = 0.17 (EtOAc [5% MeOH]); 1H NMR (free
amine, 400 MHz, CDCl3) δ 7.57 (d, J = 7.9 Hz, 2H), 7.33 (d, J = 7.9 Hz,
2H), 3.69 (s, 3H), 3.55 – 3.46 (m, 1H), 2.84 (dd, J = 13.3, 5.4 Hz, 1H), 2.70
(dd, J = 13.2, 8.1 Hz, 1H), 2.50 (dd, J = 16.0, 4.0 Hz, 1H), 2.35 (dd, J =
16.0, 8.5 Hz, 1H), 2.20 (br s, 2H); 13C NMR (free amine, 101 MHz, CDCl3)
δ 172.6, 142.8 (app s), 129.7 (2C), 129.1 (q, J = 32.42 Hz), 125.6 (q, J =
3.7 Hz, 2C), 124.3 (q, J = 271.9 Hz), 51.7, 49.6, 43.7, 41.6; HRMS (ESI)
(dd, J = 16.8, 5.2 Hz, 1H), 2.35 (s, 3H), 2.30 (dd, J = 16.8, 5.0 Hz, 1H); 13
C
NMR (101 MHz, CDCl3) δ 171.8, 141.4 (app s), 138.7, 131.5, 129.9 (2C),
129.7, 129.6 (q, J = 32.6 Hz), 129.1, 128.8, 127.9, 125.8 (q, J = 3.7 Hz,
2C), 124.2 (q, J = 272.1 Hz), 60.5, 52.4, 52.1, 41.3, 37.6, 21.4; HRMS
(ESI) calcd for C20H22F3NNaO4S (M + Na+) 452.1114, found 452.1102.
Step 2: 12 was prepared as described for 1 using the methyl ester of 12
(9.5 mg, 22 μmol) and LiOHaq (0.5 M, 0.16 mL) and obtained as a white
sticky solid (9 mg, 98% yield): 1H NMR (400 MHz, CDCl3) δ 7.56 (d, J =
8.0 Hz, 2H), 7.27 – 7.15 (m, 5H), 7.11 (d, J = 7.4 Hz, 1H), 5.30 (d, J = 9.4
Hz, 1H), 4.18 – 4.01 (m, 2H), 3.79 – 3.67 (m, 1H), 3.01 – 2.88 (m, 2H),
2.51 (dd, J = 17.0, 5.2 Hz, 1H), 2.34 (s, 3H), 2.33 (dd, J = 16.9, 4.7 Hz,
1H); 13C NMR (101 MHz, CDCl3) δ 175.1, 141.4 (app s), 138.7, 131.6,
129.9 (2C), 129.8, 129.7 (q, J = 32.6 Hz), 129.0, 128.9, 127.9, 125.8 (q, J
= 3.7 Hz, 2C), 124.2 (q, J = 272.2 Hz), 60.6, 52.4, 41.1, 37.4, 21.4; HRMS
(ESI) calcd for C19H20F3NNaO4S (M + Na+) 438.0957, found 438.0958.
calcd for C12H15F3NO2 (M + H+) 262.1049, found 262.1053. [α]20 (free
D
amine) = -7° (c = 0.7, DCM).
HPLC: tR = 11.58 min, 95.9%. [α]20 = -9° (c = 0.2, DCM).
D
Step
6:
Methyl
(S)-3-(2-(3-chlorophenyl)acetamido)-4-(4-
(trifluoromethyl)phenyl)butanoate (15). Under inert atmosphere, a solution
of 2-(3-chlorophenyl)acetic acid (289 mg, 1.70 mmol), HOBt (284 mg, 1.85
mmol), DMAP (21 mg, 168 μmol), DIPEA (936 μL, 5.37 mmol) and EDC
hydrochloride (355 mg, 1.85 mmol) in dry DCM was stirred for 3.5 h at 0 °C.
A solution of cc (457 mg, 1.54 mmol) in dry DCM (1.5 mL) was added
dropwise at 0 °C, where after the reaction was brought to rt and stirred
overnight. The reaction mixture was diluted with DCM (25 mL) and added
sat NaHCO3aq (5 mL). The aqueous phase was washed with DCM (x3), the
combined organic layers were washed with brine, dried over Na2SO4,
concentrated in vacuo to give 15 after flash chromatography (EtOAc/PE,
1:1) as a white solid (375 mg, 59% yield): Rf = 0.16 (EtOAc/PE, 1:2); 1H
NMR (400 MHz, CDCl3) δ 7.51 (d, J = 8.1 Hz, 2H), 7.30 – 7.22 (m, 2H),
7.21 – 7.16 (m, 3H), 7.03 (dt, J = 6.8, 1.6 Hz, 1H), 6.07 (d, J = 8.5 Hz, 1H),
4.51 – 4.41 (m, 1H), 3.66 (s, 3H), 3.45 (s, 2H), 2.93 (dd, J = 13.7, 7.4 Hz,
1H), 2.84 (dd, J = 13.7, 7.2 Hz, 1H), 2.53 (dd, J = 16.2, 5.3 Hz, 1H), 2.47
(dd, J = 16.2, 5.3 Hz, 1H); 13C NMR (101 MHz, CDCl3) δ 171.9, 169.7,
141.6 (q, J = 1.4 Hz), 136.7, 134.8, 130.3, 129.6 (2C), 129.5, 129.2 (q, J =
32.3 Hz), 127.7, 127.5, 125.6 (q, J = 3.7 Hz, 2C), 124.3 (q, J = 272.0 Hz),
51.9, 47.4, 43.5, 39.6, 37.1. m.p.: 135.8-139.9 °C. HRMS (ESI) calcd for
C20H19ClF3NNaO3 (M + Na+) 436.0898, found 436.0883. HPLC: tR = 12.28
4-((3-Chlorobenzyl)amino)-4-oxo-3-(4-(trifluoromethyl)benzyl)-
butanoic acid (14). Step 1: tert-Butyl 4-((3-chlorobenzyl)amino)-4-oxo-3-
(4-(trifluoromethyl)benzyl)butanoate. To a pre-dried vial, 4-(tert-butoxy)-4-
oxo-2-(4-(trifluoromethyl)benzyl)butanoic acid (13)[27] (28 mg, 84 μmol)
and BTFFH (91 mg, 0.28 mmol) dissolved in dry DCM (200 μL) was added
DIPEA (66 μL, 0.38 mmol), and the reaction was stirred at rt under argon
for 45 min. Then (3-chlorophenyl)methanamine (15 μL, 0.12 mmol) was
added and the reaction was heated to 80 °C (capped system) until
consumption of starting material. The reaction mixture was filtered through
a plug of celite (EtOAc), the crude product was evaporated onto celite, and
the tert-butyl ester of 14 was obtained after flash chromatography
(EtOAc/PE, 1:10 à EtOAc/PE, 1:5) as a sticky oil (17 mg, 43% yield): Rf
= 0.08 (EtOAc/PE, 1:5); 1H NMR (400 MHz, CDCl3) δ 7.52 (d, J = 8.0 Hz,
2H), 7.28 (d, J = 8.0 Hz, 2H), 7.22 – 7.12 (m, 3H), 6.90 – 6.85 (m, 1H),
5.92 (t, J = 5.8 Hz, 1H), 4.35 – 4.22 (m, 2H), 3.04 (dd, J = 12.5, 8.5 Hz,
1H), 2.87 – 2.68 (m, 3H), 2.44 – 2.37 (m, 1H), 1.41 (s, 9H); 13C NMR (101
MHz, CDCl3) δ 173.4, 171.6, 143.3 (q, J = 1.1 Hz), 140.3, 134.6, 129.2 (q,
J = 32.5 Hz), 130.0, 129.5 (2C), 127.9, 127.7, 125.8, 125.6 (d, J = 3.8 Hz,
2C), 124.4 (q, J = 271.8 Hz), 81.5, 45.1, 43.1, 38.2, 38.1, 28.2 (3C); HRMS
(ESI) calcd for C23H25ClF3NNaO3 (M + Na+) 478.1367, found 478.1347.
min, 95.7%. [α]20 = -11° (c = 0.30, MeOH).
D
Step 7: To a solution of 15 (336 mg, 0.81 mmol) in THF (8 mL) cooled to
0 °C was added an aqueous solution of LiOH (0.5 M, 4.1 mL) and the
reaction was stirred at rt until consumption of starting material. The
reaction mixture was diluted with water and acidified with 1 M HClaq (until
pH 2-3). The aqueous phase was extracted with EtOAc (x3), the combined
organic layers were washed with brine, dried over Na2SO4 and
concentrated in vacuo to give 1 as a white solid (324 mg, 99% yield): 1H
NMR (400 MHz, CD3OD) δ 7.48 (d, J = 8.0 Hz, 2H), 7.30 (d, J = 8.0 Hz,
2H), 7.24 – 7.18 (m, 3H), 7.06 – 7.02 (m, 1H), 4.52 – 4.43 (m, 1H), 3.38 (s,
2H), 2.99 (dd, J = 13.7, 5.1 Hz, 1H), 2.83 (dd, J = 13.7, 9.0 Hz, 1H), 2.61
– 2.48 (m, 2H); 13C NMR (101 MHz, CD3OD) δ 174.5, 172.6, 144.0 (q, J =
1.5 Hz), 139.1, 135.2, 131.0 (2C), 130.9, 130.2, 129.8 (q, J = 32.1 Hz),
128.5, 128.0, 126.2 (q, J = 3.8 Hz, 2C), 125.8 (q, J = 271.0 Hz), 49.1, 43.5,
40.7, 39.7. m.p.: 159.6-160.4 °C. HRMS (ESI) calcd for C19H18ClF3NO3 (M
Step 2: In a vial, tert-butyl ester of 14 (15 mg, 33 μmol) dissolved in dry
DCM (200 μL) was added TFA (64 μL, 0.83 mmol) and the reaction was
stirred under argon at rt for 2 h. The reaction mixture was diluted with water
and extracted with EtOAc (x3), the combined organic layers were washed
with brine, dried over Na2SO4, and concentrated in vacuo. The crude
product was filtered through a plug of celite (EtOAc/PE, 1:2, 5 mL, and
then EtOAc, 10 mL), concentrated in vacuo, and freeze dried (water, 2 mL)
to give 14 as a sticky solid (10 mg, 74% yield); 1H NMR (400 MHz, DMSO-
d6) δ 12.18 (br s, 1H), 8.47 (t, J = 5.9 Hz, 1H), 7.61 (d, J = 8.1 Hz, 2H),
7.38 (d, J = 8.0 Hz, 2H), 7.28 – 7.21 (m, 2H), 7.21 – 7.17 (m, 1H), 7.03 –
6.98 (m, 1H), 4.29 – 4.12 (m, 2H), 3.02 – 2.94 (m, 1H), 2.89 (dd, J = 13.0,
8.1 Hz, 1H), 2.76 (dd, J = 13.0, 6.7 Hz, 1H), 2.56 (dd, J = 16.6, 8.8 Hz, 1H),
2.24 (dd, J = 16.6, 5.3 Hz, 1H); 13C NMR (101 MHz, DMSO-d6) δ 173.0,
172.8, 144.1 (app s), 142.0, 132.8, 129.8, 129.7 (2C), 126.9, 126.5, 125.6,
1
+ H+) 400.0922, found 400.0920. HPLC: tR = 11.36 min, 99.0%. [α]20 = -
125.0 (q, J = 3.9 Hz, 2C), 43.0, 41.4, 37.7, 35.8 (chemical shift values δCF,
D
J
and δCF, not observed); HRMS (ESI) calcd for C19H18ClF3NO3 (M + H+)
2
11° (c = 0.20, MeOH).
J
400.0922, found 400.0942.
7
This article is protected by copyright. All rights reserved.

10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
(S)-3-(2-(2-Chlorophenyl)acetamido)-4-(4-(trifluoromethyl)phenyl)-
butanoic acid (16). Step 1: The methyl ester of 16 was prepared as
described for 15 from 6 (30 mg, 0.11 mmol), 2-(2-chlorophenyl)acetic acid
(23 mg, 0.14 mmol), HOBt (19 mg, 0.14 mmol), DIPEA (100 μL, 0.57
mmol) and EDC hydrochloride (26 mg, 0.138 mmol) and obtained after
flash chromatography (EtOAc/PE, 7:3) as a white solid (35 mg, 73% yield):
Rf = 0.25 (EtOAc/PE, 7:3); 1H NMR (400 MHz, CDCl3) δ 7.47 (d, J = 8.0
Hz, 2H), 7.40 – 7.36 (m, 1H), 7.29 – 7.22 (m, 3H), 7.19 (d, J = 8.0 Hz, 2H),
6.09 (d, J = 8.7 Hz, 1H), 4.54 – 4.41 (m, 1H), 3.65 (s, 3H), 3.62 (d, J = 4.8
Hz, 2H), 2.94 (dd, J = 13.7, 7.4 Hz, 1H), 2.84 (dd, J = 13.7, 7.2 Hz, 1H),
2.53 (dd, J = 16.3, 5.4 Hz, 1H), 2.47 (dd, J = 16.2, 5.2 Hz, 1H); 13C NMR
(101 MHz, CDCl3) δ 171.8, 169.1, 141.5 (q, J = 1.2 Hz), 132.8, 131.6,
129.7, 129.5 (2C), 129 (q, J = 32.4 Hz), 129.0, 127.3, 124.1 (q, J = 271.9
Hz), 125.4 (q, J = 3.7 Hz, 2C), 51.8, 47.1, 41.6, 39.4, 36.9. HRMS (ESI)
calcd for C20H20ClF3NO3 (M + H+) 414.1078, found 414.1084.
129.2 (q, J = 32.5 Hz), 129.1 (2C), 127.5, 125.6 (q, J = 3.7 Hz, 2C), 124.3
(q, J = 271.9 Hz), 51.9, 47.3, 44.1, 39.6, 37.2; HRMS (ESI) calcd for
C20H20F3NNaO3 (M + Na+) 402.1287, found 402.1286.
Step 2: To a solution of the methyl ester of 18 (20 mg, 53 μmol) in THF
(0.5 mL) cooled to 0 °C was added an aqueous solution of LiOH (0.5 M,
0.26 mL), and the reaction was stirred at rt until consumption of starting
material. The reaction mixture was diluted with water and washed with
DCM (x1). The aqueous phase was acidified with 1 M HClaq, extracted with
EtOAc (x3), the combined organic layers were washed with brine, dried
over Na2SO4, and concentrated in vacuo to give 18 as a white solid (15
mg, 78% yield): 1H NMR (400 MHz, CD3OD) δ 7.48 (d, J = 8.0 Hz, 2H),
7.30 (d, J = 8.0 Hz, 2H), 7.27 – 7.17 (m, 3H), 7.14 – 7.08 (m, 2H), 4.54 –
4.42 (m, 1H), 3.39 (s, 2H), 2.99 (dd, J = 13.7, 5.2 Hz, 1H), 2.83 (dd, J =
13.6, 8.9 Hz, 1H), 2.60 – 2.47 (m, 2H); 13C NMR (101 MHz, CD3OD) δ
174.5, 173.4, 144.1 (app s), 136.8, 131.0 (2C), 130.0 (2C), 129.7 (q, J =
32.6 Hz), 129.5 (2C), 127.8, 126.1 (q, J = 3.8 Hz, 2C), 125.8 (q, J = 270.8
Hz), 49.0, 43.9, 40.7, 39.6. m.p.: 181.1-185.6 °C. HRMS (ESI) calcd for
C19H18F3NNaO3 (M + Na+) 388.1131, found 388.1112. HPLC: tR = 10.83
Step 2: 16 was prepared as described for 1 using methyl ester of 16 (35
mg, 85 µmol) and LiOHaq (0.5 M, 0.51 mL) and obtained as a white solid
(31 mg, 91% yield): 1H NMR (400 MHz, DMSO-d6) δ 8.11 (d, J = 8.4 Hz,
1H), 7.62 (d, J = 8.0 Hz, 2H), 7.41 (d, J = 7.9 Hz, 2H), 7.36 (dd, J = 7.8,
1.1 Hz, 1H), 7.27 – 7.12 (m, 3H), 4.30 (td, J = 8.1, 5.2 Hz, 1H), 3.47 (s,
2H), 2.91 (dd, J = 13.5, 5.1 Hz, 1H), 2.80 (dd, J = 13.5, 8.7 Hz, 1H), 2.43
(d, J = 6.7 Hz, 2H); 13C NMR (101 MHz, DMSO-d6) δ 172.3, 168.2, 143.5
(q, J = 1.2 Hz) 134.1, 133.5, 131.6, 130.0 (2C), 128.9, 128.3, 126.9 (q, J =
31.4 Hz), 126.8, 125.0 (q, J = 3.6 Hz, 2C), 124.2 (q, J = 273.0 Hz), 47.3,
39.8, 39.3, 39.0. m.p.: 194.9-198.9 °C. HRMS (ESI) calcd for
C19H17ClF3NNaO3 (M + Na+) 422.0741, found 422.0751. HPLC: tR = 11.32
min, 97.8%. [α]20 = -64° (c = 0.1, DCM).
D
(S)-3-(2-(m-Tolyl)acetamido)-4-(4-(trifluoromethyl)phenyl)butanoic
acid (19). Step 1: The methyl ester of 19 was prepared as described for
15 using 6 (228 mg, 0.77 mmol), 2-(m-tolyl)acetic acid (129 mg, 0.86
mmol), HOBt (143 mg, 0.94 mmol), DMAP (11 mg, 89 μmol), DIPEA (467
μL, 2.68 mmol) and EDC hydrochloride (179 mg, 0.92 mmol) and obtained
after flash chromatography (EtOAc/PE, 1:2) as a white solid (217 mg, 72%
yield): Rf = 0.16 (EtOAc/PE, 1:2); 1H NMR (400 MHz, CDCl3) δ 7.47 (d, J
= 8.0 Hz, 2H), 7.21 (t, J = 7.5 Hz, 1H), 7.15 (d, J = 8.0 Hz, 2H), 7.10 (d, J
= 7.6 Hz, 1H), 6.96 (s, 1H), 6.93 (d, J = 7.5 Hz, 1H), 6.00 (d, J = 8.5 Hz,
1H), 4.52 – 4.38 (m, 1H), 3.62 (s, 3H), 3.44 (s, 2H), 2.90 (dd, J = 13.7, 7.3
Hz, 1H), 2.82 (dd, J = 13.7, 7.0 Hz, 1H), 2.53 – 2.40 (m, 2H), 2.32 (s, 3H);
13C NMR (101 MHz, CDCl3) δ 171.8, 170.7, 141.6 (app s), 138.8, 134.6,
130.2, 129.7 (2C), 129.6, 129.3, 129.0, 128.6, 128.2, 126.4, 125.5 (q, J =
3.7 Hz, 2C), 124.3 (q, J = 271.9 Hz), 51.8, 47.3, 44.0, 39.5, 37.3, 21.4;
HRMS (ESI) calcd for C21H22F3NNaO3 (M + Na+) 416.1444, found
416.1442.
min, 98.0%. [α]20 = -1.8° (c = 0.11, MeOH).
D
(S)-3-(2-(4-Chlorophenyl)acetamido)-4-(4-(trifluoromethyl)phenyl)-
butanoic acid (17). Step 1: The methyl ester of 17 was prepared as
described for 15 using 6 (30 mg, 0.11 mmol), 2-(4-chlorophenyl)acetic acid
(23 mg, 0.14 mmol), HOBt (19 mg, 0.14 mmol), DIPEA (100 μL, 0.57
mmol) and EDC hydrochloride (26 mg, 0.14 mmol) and obtained after flash
chromatography (EtOAc/PE, 7:3) as a white solid (36 mg, 75% yield): Rf =
0.30 (EtOAc/PE, 7:3); 1H NMR (400 MHz, CDCl3) δ 7.50 (d, J = 8.0 Hz,
2H), 7.29 (d, J = 8.4 Hz, 2H), 7.19 (d, J = 7.9 Hz, 2H), 7.08 (d, J = 8.4 Hz,
2H), 6.03 (d, J = 8.6 Hz, 1H), 4.54 – 4.37 (m, 1H), 3.66 (s, 3H), 3.45 (s,
2H), 2.93 (dd, J = 13.7, 7.4 Hz, 1H), 2.83 (dd, J = 13.7, 7.3 Hz, 1H), 2.58
– 2.42 (m, 2H); 13C NMR (101 MHz, CDCl3) δ 171.9, 169.8, 141.4 (q, J =
1.2 Hz), 133.3, 133.1, 130.5 (2C), 129.5 (2C), 129.1 (q, J = 32.4 Hz), 129.0
(2C), 125.4 (q, J = 3.7 Hz, 2C), 124.2 (q, J = 272.0 Hz), 51.8, 47.2, 43.1,
39.4, 36.9; HRMS (ESI) calcd for C20H20ClF3NO3 (M + H+) 414.1078, found
414.1091.
Step 2: 19 was prepared as described for 18 using the methyl ester of 19
(124 mg, 0.32 mmol) and LiOHaq (0.5 M, 1.58 mL) and obtained as a white
solid (114 mg, 95% yield): 1H NMR (400 MHz, CD3OD) δ 7.46 (d, J = 8.0
Hz, 2H), 7.28 (d, J = 8.0 Hz, 2H), 7.13 (t, J = 7.6 Hz, 1H), 7.04 (d, J = 7.6
Hz, 1H), 6.99 (s, 1H), 6.94 – 6.89 (m, 1H), 4.50 – 4.41 (m, 1H), 3.35 (s,
2H), 2.98 (dd, J = 13.7, 5.2 Hz, 1H), 2.83 (dd, J = 13.7, 8.8 Hz, 1H), 2.59
– 2.47 (m, 2H), 2.29 (s, 3H); 13C NMR (101 MHz, CD3OD) δ 174.5, 173.5,
144.1 (app s), 139.2, 136.6, 131.0 (2C), 130.8, 129.7 (q, J = 32.2 Hz),
129.4, 128.5, 127.1, 126.1 (q, J = 3.8 Hz, 2C), 125.8 (q, J = 271.0 Hz),
49.0, 44.0, 40.6, 39.5, 21.4. m.p.: 167.3-168.7 °C. HRMS (ESI) calcd for
C20H21F3NO3 (M + H+) 402.1287, found 402.1301. HPLC: tR = 11.03 min,
Step 2: 17 was prepared as described for 1 using methyl ester of 17 (25
mg, 60 μmol) and LiOHaq (0.5 M, 0.36 mL) and obtained as a white solid
(22 mg, 91% yield): 1H NMR (400 MHz, DMSO-d6) δ 8.11 (d, J = 8.5 Hz,
1H), 7.56 (d, J = 8.0 Hz, 2H), 7.32 (d, J = 8.0 Hz, 2H), 7.27 (d, J = 8.4 Hz,
2H), 7.10 (d, J = 8.3 Hz, 2H), 4.32 – 4.16 (m, 1H), 3.29 (s, 2H), 2.88 (dd, J
= 13.5, 5.0 Hz, 1H), 2.75 (dd, J = 13.5, 8.7 Hz, 1H), 2.47 – 2.31 (m, 2H);
13C NMR (101 MHz, DMSO-d6) δ 172.2, 169.0, 143.4 (app s), 135.3, 131.0,
130.7 (2C), 130.0 (2C), 127.9 (2C), 124.8 (q, J = 3.3 Hz, 2C), 47.2, 41.6,
99.9%. [α]20 = +4° (c = 0.3, MeOH).
D
(S)-4-(4-(Trifluoromethyl)phenyl)-3-(2-(3-(trifluoromethyl)phenyl)-
acetamido)butanoic acid (20). Step 1: The methyl ester of 20 was
1
2
39.2, 39.0 (chemical shift values δCF, and δCF, not observed); HRMS
J
J
prepared as described for 15 using
6 (50 mg, 0.19 mmol), 2-(3-
(ESI) calcd for C19H17ClF3NNaO3 (M + Na+) 422.0741, found 422.0753.
(trifluoromethyl)phenyl)acetic acid (47 mg, 0.23 mmol), HOBt (34 mg, 0.23
mmol), DIPEA (167 μL, 0.96 mmol) and EDC hydrochloride (48 mg, 0.23
mmol) and obtained after flash chromatography (EtOAc/PE, 3:2) as a white
solid (54 mg, 63% yield): Rf = 0.15 (EtOAc/PE, 7:3); 1H NMR (400 MHz,
CDCl3) δ 7.55 (d, J = 7.8 Hz, 1H), 7.51 – 7.44 (m, 3H), 7.43 (d, J = 7.7 Hz,
1H), 7.34 (d, J = 7.7 Hz, 1H), 7.20 (d, J = 7.9 Hz, 2H), 6.19 (d, J = 8.5 Hz,
1H), 4.60 – 4.39 (m, 1H), 3.65 (s, 3H), 3.53 (s, 2H), 2.95 (dd, J = 13.7, 7.4
Hz, 1H), 2.85 (dd, J = 13.7, 7.3 Hz, 1H), 2.54 (dd, J = 16.3, 5.2 Hz, 1H),
2.47 (dd, J = 16.3, 5.2 Hz, 1H); 13C NMR (101 MHz, CDCl3) δ 171.9, 169.4,
141.4 (q, J = 1.1 Hz), 135.6, 132.6 (q, J = 1.1 Hz), 132.6 (q, J = 1.5 Hz),
131.1 (q, J = 32.2 Hz), 129.4 (2C), 129.3, 125.9 (q, J = 3.8 Hz, 2C), 125.4
(q, J = 3.5 Hz), 124.1 (q, J = 3.8 Hz), 124.1 (q, J = 271.5 Hz), 124.0 (q, J
HPLC: tR = 11.48 min, 98.9%.
(S)-3-(2-Phenylacetamido)-4-(4-(trifluoromethyl)phenyl)butanoic
acid (18). Step 1: The methyl ester of 18 was prepared as described for
15 using 6 (35 mg, 0.12 mmol), phenylacetic acid (18 mg, 0.13 mmol),
HOBt (22 mg, 0.14 mmol), DMAP (1.4 mg, 12 μmol), DIPEA (72 μL, 0.42
mmol) and EDC hydrochloride (28 mg, 0.14 mmol) and obtained after flash
chromatography (EtOAc/PE, 1:1) as a white solid (32 mg, 72% yield): Rf =
0.14 (EtOAc/PE, 1:2); 1H NMR (400 MHz, CDCl3) δ 7.48 (d, J = 8.0 Hz,
2H), 7.35 – 7.27 (m, 3H), 7.18 – 7.11 (m, 4H), 5.96 (d, J = 8.5 Hz, 1H),
4.50 – 4.41 (m, 1H), 3.63 (s, 3H), 3.49 (s, 2H), 2.90 (dd, J = 13.7, 7.3 Hz,
1H), 2.82 (dd, J = 13.7, 7.1 Hz, 1H), 2.53 – 2.41 (m, 2H); 13C NMR (101
MHz, CDCl3) δ 171.9, 170.5, 141.6 (app s), 134.8, 129.7 (2C), 129.4 (2C),
=
272.7 Hz), 51.8, 47.2, 43.4, 39.4, 36.8; HRMS (ESI) calcd for
C22H20F6NO3 (M + H+) 448.1342, found 448.1341.
8
This article is protected by copyright. All rights reserved.

10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
Step 2: 20 was prepared as described for 1 using the methyl ester of 20
(50 mg, 112 μmol) and LiOHaq (0.5 M, 0.90 mL) and obtained as a white
solid (43 mg, 89% yield): 1H NMR (400 MHz, DMSO-d6) δ 12.34 (br s, 1H),
8.19 (d, J = 8.4 Hz, 1H), 7.56 (d, J = 7.7 Hz, 1H), 7.51 (d, J = 8.1 Hz, 3H),
7.46 (d, J = 7.7 Hz, 1H), 7.39 (d, J = 7.6 Hz, 1H), 7.30 (d, J = 7.9 Hz, 2H),
4.30 – 4.20 (m, 1H), 3.42 (s, 2H), 2.89 (dd, J = 13.5, 4.8 Hz, 1H), 2.76 (dd,
J = 13.5, 8.8 Hz, 1H), 2.49 – 2.35 (m, 2H); 13C NMR (101 MHz, DMSO-d6)
δ 172.2, 168.9, 143.3 (q, J = 1.0 Hz), 137.7, 133.1 (q, J = 0.8 Hz), 129.9
(2C), 129.0, 128.9 (q, J = 31.4 Hz), 126.9 (q, J = 31.9 Hz), 125.5 (q, J =
3.2 Hz), 124.8 (q, J = 3.7 Hz, 2C), 124.4 (q, J = 271.5 Hz), 124.3 (q, J =
272.8 Hz), 123.8 (q, J = 3.7 Hz), 47.2, 41.9, 39.2, 38.9. m.p.: 164.5-
167.1 °C. HRMS (ESI) calcd for C20H17F6NNaO3 (M + Na+) 456.1005,
Hz, 2H), 7.47 (dt, J = 13.5, 7.4 Hz, 2H), 7.42 – 7.36 (m, 1H), 7.33 (d, J =
7.9 Hz, 2H), 7.28 (d, J = 7.0 Hz, 1H), 4.36 – 4.19 (m, 1H), 3.80 (s, 2H),
2.89 (dd, J = 13.4, 4.9 Hz, 1H), 2.80 (dd, J = 13.3, 8.7 Hz, 1H), 2.46 (dd, J
= 11.7, 1.9 Hz, 1H), 2.42 (dd, J = 11.7, 2.4 Hz, 1H); 13C NMR (101 MHz,
DMSO-d6) δ 172.3, 169.4, 143.5 (q, J = 0.9 Hz), 133.3, 132.6, 131.9, 130.0
(2C), 128.3, 127.6, 127.0, 126.9 (q, J = 31.4 Hz), 125.8, 125.5, 125.4,
124.9 (q, J = 3.4 Hz, 2C), 124.4 (q, J = 272.4 Hz), 124.2, 47.3, 39.8, 39.2,
38.9; HRMS (ESI) calcd for C23H20F3NNaO3 (M + Na+) 438.1287, found
438.1301. HPLC: tR = 10.52 min, 98.0%. [α]20 = -10.7° (c = 0.12, MeOH).
D
(S)-3-(2-(Benzo[b]thiophen-3-yl)acetamido)-4-(4-(trifluoromethyl)-
phenyl)butanoic acid (23). Step 1: The methyl ester of 23 was prepared
as described for 15 using 6 (35 mg, 0.12 mmol), 2-(benzo[b]thiophen-3-
yl)acetic acid (25 mg, 0.13 mmol), HOBt (22 mg, 0.14 mmol), DMAP (1.5
mg, 12 μmol), DIPEA (71 μL, 0.41 mmol) and EDC hydrochloride (28 mg,
0.15 mmol) and obtained after flash chromatography (EtOAc/PE, 1:3) as a
white solid (32 mg, 72% yield): Rf = 0.23 (EtOAc/PE, 1:3); 1H NMR (400
MHz, CDCl3) δ 7.92 – 7.87 (m, 1H), 7.64 – 7.60 (m, 1H), 7.42 – 7.33 (m,
4H), 7.24 (s, 1H), 7.02 (d, J = 8.0 Hz, 2H), 6.02 (d, J = 8.6 Hz, 1H), 4.52 –
4.39 (m, 1H), 3.75 (s, 2H), 3.53 (s, 3H), 2.83 (dd, J = 13.7, 7.4 Hz, 1H),
2.75 (dd, J = 13.7, 6.8 Hz, 1H), 2.48 – 2.33 (m, 2H); 13C NMR (101 MHz,
CDCl3) δ 171.7, 169.4 (d, J = 7.7 Hz), 141.4 (app s), 140.6, 138.4, 129.5
(2C), 129.3, 129.1 (q, J = 32.3 Hz), 125.5 (q, J = 3.7 Hz, 2C), 125.3, 125.0,
124.7, 124.3 (q, J = 272.0 Hz), 123.1, 121.8, 51.8, 47.1 (app d, J = 10.6
Hz), 39.5 (app d, J = 2.0 Hz), 37.2 (app d, J = 2.6 Hz), 37.0 (app d, J = 3.4
Hz); HRMS (ESI) calcd for C22H20F3NNaO3S (M + Na+) 458.1008, found
458.1022.
found 456.1025. HPLC: tR = 11.37 min, 99.3%. [α]20 = -2.6° (c = 0.11,
D
MeOH).
(S)-3-(2-(3-Methoxyphenyl)acetamido)-4-(4-(trifluoromethyl)phenyl)-
butanoic acid (21). Step 1: The methyl ester of 21 was prepared as
described for 15 using 6 (50 mg, 0.19 mmol), 2-(3-methoxyphenyl)acetic
acid (38 mg, 0.23 mmol), HOBt (34 mg, 0.23 mmol), DIPEA (167 μL, 0.96
mmol) and EDC hydrochloride (48 mg, 0.23 mmol) and obtained after flash
chromatography (EtOAc/PE, 3:2) as a white solid (58 mg, 74%): Rf = 0.15
(EtOAc/PE, 7:3); 1H NMR (400 MHz, CDCl3) δ 7.48 (d, J = 8.0 Hz, 2H),
7.25 (t, J = 7.8 Hz, 1H), 7.15 (d, J = 8.0 Hz, 2H), 6.84 (dd, J = 7.9, 2.7 Hz,
1H), 6.72 (d, J = 7.5 Hz, 1H), 6.71 (d, J = 2.2 Hz, 1H), 5.97 (d, J = 8.5 Hz,
1H), 4.58 – 4.28 (m, 1H), 3.79 (s, 3H), 3.64 (s, 3H), 3.47 (s, 2H), 2.90 (dd,
J = 13.7, 7.2 Hz, 1H), 2.82 (dd, J = 13.7, 7.0 Hz, 1H), 2.51 (dd, J = 16.0,
5.5 Hz, 1H), 2.45 (dd, J = 15.9, 5.4 Hz, 1H); 13C NMR (101 MHz, CDCl3) δ
171.7, 170.2, 160.0, 141.4 (q, J = 1.2 Hz), 136.0, 130.0, 129.5 (2C), 129.0
(q, J = 32.7 Hz), 125.4 (q, J = 3.7 Hz, 2C), 124.2 (q, J = 271.8 Hz), 121.5,
114.9, 112.8, 55.2, 51.7, 47.1, 44.0, 39.4, 37.1; HRMS (ESI) calcd for
C21H23F3NO4 (M + H+) 410.1574, found 410.1587.
Step 2: 23 was prepared as described for 1 using the methyl ester of 23
(30 mg, 69 μmol) and LiOHaq (0.5 M, 0.35 mL) and obtained as a white
solid (28 mg, 97% yield): 1H NMR (400 MHz, DMSO-d6) δ 12.33 (br s, 1H),
8.22 (d, J = 8.4 Hz, 1H), 7.97 – 7.92 (m, 1H), 7.69 – 7.65 (m, 1H), 7.54 (d,
J = 8.0 Hz, 2H), 7.38 – 7.31 (m, 4H), 7.30 (s, 1H), 4.34 – 4.23 (m, 1H),
3.59 (s, 2H), 2.90 (dd, J = 13.4, 4.8 Hz, 1H), 2.79 (dd, J = 13.5, 8.7 Hz,
1H), 2.49 – 2.38 (m, 2H); 13C NMR (101 MHz, DMSO-d6) δ 172.2, 168.5,
143.4 (app s), 139.3, 138.7, 130.5, 129.9 (2C), 126.9 (q, J = 32.1 Hz),
124.8 (q, J = 3.7 Hz, 2C), 124.4 (q, J = 271.9 Hz), 124.1, 124.0, 123.8,
122.7, 122.0, 47.3, 39.1, 38.9, 35.5; HRMS (ESI) calcd for
C21H18F3NNaO3S (M + Na+) 444.0852, found 444.0862. HPLC: tR = 11.56
Step 2: 21 was prepared as described for 1 using the methyl ester of 21
(50 mg, 127 μmol) and LiOHaq (0.5 M, 1.0 mL) and obtained as a white
solid (43 mg, 89% yield): 1H NMR (400 MHz, DMSO-d6) δ 12.29 (s, 1H),
8.07 (d, J = 8.3 Hz, 1H), 7.54 (d, J = 8.0 Hz, 2H), 7.31 (d, J = 7.9 Hz, 2H),
7.13 (t, J = 7.8 Hz, 1H), 6.76 (dd, J = 8.1, 2.6 Hz, 1H), 6.72 (t, J = 1.9 Hz,
1H), 6.67 (d, J = 7.5 Hz, 1H), 4.45 – 4.06 (m, 1H), 3.70 (s, 3H), 3.28 (d, J
= 3.5 Hz, 2H), 2.87 (dd, J = 13.4, 4.8 Hz, 1H), 2.77 (dd, J = 13.4, 8.5 Hz,
1H), 2.44 (dd, J = 15.6, 6.6 Hz, 1H), 2.37 (dd, J = 15.8, 7.1 Hz, 1H); 13C
NMR (101 MHz, DMSO-d6) δ 172.2, 169.4, 159.1, 143.4 (q, J = 1.0 Hz),
137.7, 130.0 (2C), 129.0, 126.9 (q, J = 31.7 Hz), 124.8 (q, J = 3.4 Hz, 2C),
124.4 (q, J = 272.4 Hz) 121.1, 114.7, 111.7, 54.9, 47.2, 42.5, 39.1, 38.8.
m.p.: 161.6-162.3 °C. HRMS (ESI) calcd for C20H20F3NNaO4 (M + Na+)
min, 99.6%. [α]20 = +0.5° (c = 0.2, DMSO).
D
(S)-3-(2-(1H-Indol-3-yl)acetamido)-4-(4-(trifluoromethyl)phenyl)-
butanoic acid (24). Step 1: The methyl ester of 24 was prepared as
described for 15 using 6 (50 mg, 0.19 mmol), 2-(1H-indol-3-yl)acetic acid
(40 mg, 0.23 mmol), HOBt (34 mg, 0.23 mmol), DIPEA (167 μL, 0.96
mmol) and EDC hydrochloride (48 mg, 0.23 mmol) and obtained after flash
chromatography (EtOAc/PE, 3:2) as a brown solid (65 mg, 81% yield): Rf
= 0.3 (EtOAc/PE, 3:2); 1H NMR (400 MHz, CDCl3) δ 8.17 (s, 1H), 7.47 –
7.40 (m, 2H), 7.37 (d, J = 8.0 Hz, 2H), 7.28 – 7.22 (m, 1H), 7.16 – 7.09 (m,
1H), 7.05 (d, J = 2.5 Hz, 1H), 7.03 (d, J = 8.0 Hz, 2H), 6.10 (d, J = 8.7 Hz,
1H), 4.66 – 4.29 (m, 1H), 3.67 (s, 2H), 3.51 (s, 3H), 2.84 (dd, J = 13.8, 7.1
Hz, 1H), 2.75 (dd, J = 13.8, 6.9 Hz, 1H), 2.44 (dd, J = 10.5, 3.6 Hz, 1H),
2.39 (dd, J = 10.4, 3.4 Hz, 1H); 13C NMR (101 MHz, CDCl3) δ 171.5, 170.9,
141.3 (app s), 136.3, 129.5 (2C), 125.3 (q, J = 3.8 Hz, 2C), 123.5, 122.7,
120.2, 118.6, 111.3, 108.9, 51.6, 46.9, 39.3, 37.2, 33.5 (chemical shift
418.1237, found 418.1256. HPLC: tR = 10.66 min, 99.7%. [α]20 = -3.1° (c
D
= 0.11, MeOH).
(S)-3-(2-(Naphthalen-1-yl)acetamido)-4-(4-(trifluoromethyl)phenyl)-
butanoic acid (22). Step 1: The methyl ester of 22 was prepared as
described for 15 using 6 (50 mg, 0.19 mmol), 2-(naphthalen-1-yl)acetic
acid (43 mg, 0.23 mmol), HOBt (34 mg, 0.23 mmol), DIPEA (167 μL, 0.96
mmol) and EDC hydrochloride (48 mg, 0.23 mmol) and obtained after flash
chromatography (EtOAc/PE, 3:2) as a white solid (56 mg, 68% yield): Rf =
0.30 (EtOAc/PE, 3:2); 1H NMR (400 MHz, CDCl3) δ 8.20 (s, 1H), 7.47 –
7.40 (m, 2H), 7.37 (d, J = 8.0 Hz, 2H), 7.27 – 7.22 (m, 2H), 7.15 – 7.10 (m,
1H), 7.05 (d, J = 2.5 Hz, 1H), 7.03 (d, J = 8.0 Hz, 2H), 6.10 (d, J = 8.7 Hz,
1H), 4.61 – 4.33 (m, 1H), 3.67 (s, 3H), 3.51 (s, 2H), 2.84 (dd, J = 13.7, 7.1
Hz, 1H), 2.75 (dd, J = 13.7, 6.8 Hz, 1H), 2.44 (dd, J = 10.3, 3.4 Hz, 1H),
2.39 (dd, J = 10.5, 5.7 Hz, 1H); 13C NMR (101 MHz, CDCl3) δ 171.5, 170.9,
141.3 (q, J = 0.7 Hz), 136.3, 129.5 (2C), 128.8 (q, J = 32.2 Hz), 126.9,
125.3 (q, J = 3.8 Hz, 2C), 124.1 (q, 272.3 Hz), 123.5, 122.7 (2C), 120.2,
118.6, 111.3, 108.9, 51.6, 46.9, 39.3, 37.2, 33.5; HRMS (ESI) calcd for
C24H23F3NO3 (M + H+) 430.1630, found 430.1623.
1
2
values δCF, J and δCF, J not observed); HRMS (ESI) calcd for C22H22F3N2O3
(M + H+) 419.1577, found 419.1577.
Step 2: 24 was prepared as described for 1 using the methyl ester of 24
(65 mg, 0.16 mmol) and LiOHaq (0.5 M, 1.24 mL) and obtained as a brown
solid (57 mg, 91% yield): 1H NMR (400 MHz, DMSO-d6) δ 12.25 (s, 1H),
10.83 (s, 1H), 7.93 (d, J = 8.2 Hz, 1H), 7.50 (d, J = 7.9 Hz, 2H), 7.39 (d, J
= 7.9 Hz, 1H), 7.30 (dd, J = 16.5, 8.0 Hz, 3H), 7.13 – 7.00 (m, 2H), 6.91 (t,
J = 7.4 Hz, 1H), 4.38 – 4.20 (m, 1H), 3.47 – 3.40 (s, 2H), 2.85 (dd, J = 13.5,
Step 2: 22 was prepared as described for 1 using the methyl ester of 22
(55 mg, 0.13 mmol) and LiOHaq (0.5 M, 1.0 mL) and obtained as a white
solid (47 mg, 88% yield): 1H NMR (400 MHz, DMSO-d6) δ 8.23 (d, J = 8.4
Hz, 1H), 7.90 (t, J = 7.3 Hz, 2H), 7.79 (d, J = 8.1 Hz, 1H), 7.54 (d, J = 7.9
5.2 Hz, 1H), 2.78 (dd, J = 13.5, 8.1 Hz, 1H), 2.40 (d, J = 6.7 Hz, 2H); 13
C
NMR (101 MHz, DMSO-d6) δ 172.3, 170.1, 143.4 (q, J = 0.9 Hz), 136.1,
130.0 (2C), 127.2, 126.8 (q, J = 32.3 Hz), 124.8 (q, J = 3.5 Hz, 2C), 124.2
(q, J = 272.5 Hz) 123.8, 120.9, 118.7, 118.2, 111.2, 108.7, 47.2, 39.1, 38.8,
9
This article is protected by copyright. All rights reserved.

10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
32.7; HRMS (ESI) calcd for C21H19F3N2NaO3 (M + Na+) 427.1240, found
Step 2: 27 was prepared as described for 1 using 27a (25 mg, 82 µmol)
and LiOHaq (0.5 M, 0.47 mL) and obtained as a white solid (21 mg, 88%
yield): 1H NMR (400 MHz, DMSO-d6) δ 7.87 (d, J = 8.3 Hz, 1H), 7.65 (d, J
= 7.9 Hz, 2H), 7.40 (d, J = 7.9 Hz, 2H), 4.79 – 3.95 (m, 1H), 2.86 (dd, J =
13.5, 5.5 Hz, 1H), 2.78 (dd, J = 13.5, 8.0 Hz, 1H), 2.43 – 2.30 (m, 2H), 1.72
(s, 3H); 13C NMR (101 MHz, DMSO-d6) δ 172.2, 168.5, 143.5 (app s),
129.9 (2C), 126.6 (q, J = 33.8 Hz), 124.8 (q, J = 3.7 Hz, 2C), 47.0, 39.1,
427.1259. HPLC: tR = 11.42 min, 97.2%. [α]20 = -1.0° (c = 0.10, MeOH).
D
(S)-3-(3-(3-Chlorophenyl)propanamido)-4-(4-(trifluoromethyl)phenyl)-
butanoic acid (25). Step 1: The methyl ester of 25 of the title compound
was prepared as described for 15 using 6 (35 mg, 0.12 mmol), 3-(3-
chlorophenyl)propanoic acid (24 mg, 0.13 mmol), HOBt (22 mg, 0.14
mmol), DMAP (1.6 mg, 13 μmol), DIPEA (71 μL, 0.41 mmol) and EDC
hydrochloride (27 mg, 0.14 mmol) and obtained after flash
chromatography (EtOAc/PE, 1:2) as a white solid (38 mg, 76% yield): Rf =
0.24 (EtOAc/PE, 1:1); 1H NMR (400 MHz, CD3OD) δ 7.54 (d, J = 8.1 Hz,
2H), 7.32 (d, J = 8.0 Hz, 2H), 7.24 – 7.14 (m, 3H), 7.07 (dt, J = 7.2, 1.4 Hz,
1H), 4.49 – 4.41 (m, 1H), 3.62 (s, 3H), 2.89 (dd, J = 13.7, 5.9 Hz, 1H), 2.85
– 2.77 (m, 3H), 2.51 (dd, J = 15.5, 6.0 Hz, 1H), 2.46 – 2.35 (m, 3H); 13C
NMR (101 MHz, CD3OD) δ 174.2, 173.0, 144.5, 144.0 (q, J = 1.2 Hz),
135.2, 131.0 (2C), 130.9, 129.8 (q, J = 32.3 Hz), 129.5, 127.9, 127.3, 126.2
(q, J = 3.9 Hz, 2C), 125.8 (q, J = 271.0 Hz), 52.2, 48.9, 40.8, 39.4, 38.3,
32.2; HRMS (ESI) calcd for C21H22ClF3NO3 (M + H+) 428.1235, found
428.1255.
1
38.6, 22.5 (chemical shift value δCF, J not observed). m.p.: 153.5-155.2 °C.
HRMS (ESI) calcd for C13H14F3NNaO3 (M + Na+) 312.0818, found
312.0829. HPLC: tR = 9.43 min, 99.6%. [α]20 = -3.9° (c = 0.10, MeOH).
D
(S)-2-(2-(3-Chlorophenyl)acetamido)-3-(4-(trifluoromethyl)phenyl)-
propanoic acid (28). Step 1: The methyl ester of 28 of the title compound
was prepared as described for 15 using 11 (41 mg, 0.14 mmol), 2-(3-
chlorophenyl)acetic acid (27 mg, 0.16 mmol), HOBt (26 mg, 0.17 mmol),
DMAP (1.8 mg, 14 μmol), DIPEA (88 μL, 0.50 mmol) and EDC
hydrochloride (32 mg, 0.17 mmol) and obtained after flash
chromatography (EtOAc/PE, 1:2) as a white solid (34 mg, 60% yield): Rf =
0.13 (EtOAc/PE, 1:2); 1H NMR (400 MHz, CD3OD) δ 7.51 (d, J = 8.0 Hz,
2H), 7.31 (d, J = 8.0 Hz, 2H), 7.25 – 7.18 (m, 3H), 7.09 – 7.04 (m, 1H),
4.74 (dd, J = 9.6, 5.1 Hz, 1H), 3.72 (s, 3H), 3.46 (s, 2H), 3.27 (dd, J = 14.0,
5.1 Hz, 1H), 3.03 (dd, J = 13.9, 9.6 Hz, 1H); 13C NMR (101 MHz, CD3OD)
δ 173.1, 173.0, 142.8 (q, J = 1.3 Hz), 138.8, 135.2, 130.91, 130.87 (2C),
130.2, 130.1 (q, J = 32.1 Hz), 128.5, 128.0, 126.3 (q, J = 3.8 Hz, 2C), 125.7
Step 2: 25 was prepared as described for 1 using the methyl ester of 25
(29 mg, 68 μmol) and LiOHaq (0.5 M, 0.34 mL) and obtained as a white
solid (27 mg, 95% yield): 1H NMR (400 MHz, CDCl3) δ 7.53 (d, J = 8.0 Hz,
2H), 7.24 (d, J = 8.0 Hz, 2H), 7.21 – 7.14 (m, 3H), 7.03 (dt, J = 6.4, 2.0 Hz,
1H), 6.14 (d, J = 8.7 Hz, 1H), 4.54 – 4.44 (m, 1H), 2.96 (dd, J = 13.7, 6.9
Hz, 1H), 2.91 – 2.81 (m, 3H), 2.57 – 2.37 (m, 4H); 13C NMR (101 MHz,
CDCl3) δ 175.7, 171.9, 142.6, 141.5 (app s), 134.4, 130.0, 129.7 (2C),
129.4 (q, J = 32.5 Hz), 128.6, 126.7, 126.7, 125.7 (q, J = 3.7 Hz, 2C), 124.3
(q, J = 272.1 Hz), 47.2, 39.7, 38.2, 36.9, 31.3; HRMS (ESI) calcd for
C20H19ClF3NNaO3 (M + Na+) 436.0898, found 436.0914. HPLC: tR = 11.63
(q,
J = 271.1 Hz), 54.7, 52.9, 43.0, 37.9; HRMS (ESI) calcd for
C19H17ClF3NNaO3 (M + Na+) 422.0741, found 422.0757.
Step 2: 28 was prepared as described for 18 using the methyl ester of 28
(25 mg, 63 μmol) and LiOHaq (0.5 M, 0.31 mL) and obtained as a white
solid (24 mg, 99% yield): 1H NMR (400 MHz, CD3OD) δ 7.50 (d, J = 8.0
Hz, 2H), 7.31 (d, J = 8.0 Hz, 2H), 7.25 – 7.18 (m, 3H), 7.09 – 7.05 (m, 1H),
4.72 (dd, J = 9.4, 4.8 Hz, 1H), 3.51 – 3.41 (m, 2H), 3.31 (dd, J = 13.8, 4.9
Hz, 1H), 3.03 (dd, J = 13.9, 9.5 Hz, 1H); 13C NMR (101 MHz, CD3OD) δ
174.2, 173.0, 143.1 (app s), 138.9, 135.2, 130.9 (2C), 130.2, 130.0 (q, J =
32.2 Hz), 128.5, 128.0, 127.1, 126.2 (q, J = 3.9 Hz, 2C), 125.8 (q, J = 271.1
Hz), 54.6, 43.1, 38.0. m.p.: 130.6-135.3 °C. HRMS (ESI) calcd for
C18H16ClF3NO3 (M + H+) 386.0765, found 386.0775. HPLC: tR = 11.66 min,
min, 98.5%. [α]20 = -14° (c = 0.2, DCM).
D
(S)-3-(3-Chlorobenzamido)-4-(4-(trifluoromethyl)phenyl)butanoic
acid (26) and (S)-3-acetamido-4-(4-(trifluoromethyl)phenyl)butanoic
acid (27). Step 1: In a pre-dried flask, 6 (74 mg, 0.283 mmol) and DMAP
(69 mg, 0.566 mmol) in dry DCM (1 mL) was added 3-chlorobenzoyl
chloride (253 µL, 0.20 mmol) and acetyl chloride (141 µL, 0.20 mmol) at
0 °C and the reaction was stirred for 20 h at rt. The contents were
concentrated and the methyl ester of 26 (26a) was obtained after flash
chromatography (EtOAc/PE, 3:2) as a white solid (29 mg, 26%) along with
the methyl ester of 27 (27a) as a thick syrup (27 mg, 31%): Rf = 0.4
(EtOAc/PE, 1:1) and Rf = 0.10 (EtOAc/PE, 1:1), respectively; 26a: 1H NMR
(400 MHz, CDCl3) δ 7.73 (t, J = 1.9 Hz, 1H), 7.62 – 7.54 (m, 3H), 7.50 –
7.45 (m, 1H), 7.41 – 7.32 (m, 3H), 7.10 (d, J = 8.7 Hz, 1H), 4.82 – 4.53 (m,
1H), 3.74 (s, 3H), 3.16 (dd, J = 13.6, 6.6 Hz, 1H), 2.97 (dd, J = 13.7, 8.1
Hz, 1H), 2.78 – 2.42 (m, 2H); 13C NMR (101 MHz, CDCl3) δ 172.5, 165.4,
141.6 (q, J = 1.1 Hz), 136.0, 134.9, 131.7, 129.9, 129.6 (2C), 129.2 (q, J =
32.6 Hz), 127.4, 125.6 (q, J = 3.5 Hz, 2C), 124.9, 124.5 (q, J = 272.0 Hz),
52.0, 47.7, 39.7, 36.3; HRMS (ESI) calcd for C19H18ClF3NO3 (M + H+)
400.0922, found 400.0932. 27a: 1H NMR (400 MHz, CDCl3) δ 7.55 (d, J =
7.9 Hz, 2H), 7.30 (d, J = 7.9 Hz, 2H), 6.22 (d, J = 9.4 Hz, 1H), 4.75 – 4.08
(m, 1H), 3.70 (s, 3H), 3.02 (dd, J = 13.7, 6.8 Hz, 1H), 2.87 (dd, J = 13.7,
7.9 Hz, 1H), 2.74 – 2.24 (m, 2H), 1.93 (s, 3H); 13C NMR (101 MHz, CDCl3)
δ 172.2, 169.5, 141.9 (app. s), 129.5 (2C), 129.1 (q, J = 32.5 Hz), 125.6
(q, J = 3.5 Hz, 2C), 124.5 (q, J = 272.0 Hz), 51.8, 47.0, 39.6, 36.6, 23.4.
98.9%. [α]20 = +15° (c = 0.2, MeOH).
D
(R)-4-(2-(3-Chlorophenyl)acetamido)-5-(4-(trifluoromethyl)phenyl)-
pentanoic
acid
(29).
Step
1:
Methyl
(S)-2-amino-3-(4-
(trifluoromethyl)phenyl)propanoate hydrochloride. At 0 °C, thionyl chloride
(1.1 mL, 15.0 mmol) was added dropwise to a round bottom flask
containing dry MeOH (7.5 mL) and the solution was stirred for 5 min. Then
(S)-2-amino-3-(4-(trifluoromethyl)phenyl)propanoic acid (7) (996 mg, 4.27
mmol) was added, the reaction was brought to rt and stirred under argon
until consumption of starting material. The reaction mixture was
concentrated, triturated with Et2O (x2), and dried in vacuo to afford a white
solid (1.03 g, 85% yield): Rf = 0.46 (DCM [1% MeOH]); 1H NMR (400 MHz,
DMSO-d6) δ 8.87 (s, 3H), 7.69 (d, J = 7.9 Hz, 2H), 7.51 (d, J = 7.8 Hz, 2H),
4.32 (t, J = 6.2 Hz, 1H), 3.67 (s, 3H), 3.33 (dd, J = 14.0, 5.5 Hz, 1H), 3.24
(dd, J = 13.8, 7.2 Hz, 1H); 13C NMR (101 MHz, DMSO-d6) δ 169.0, 139.8
(app s), 130.3 (2C), 127.8 (q, J = 31.6 Hz), 125.3 (q, J = 3.4 Hz, 2C), 124.2
(q, J = 274.6 Hz), 52.8, 52.6, 35.3; HRMS (ESI) calcd for C11H13F3NO2 (M
+ H+) 248.0893, found 248.0899. 1H NMR spectra in overall agreement
with literature.[32]
Step 2: 26 was prepared as described for 1 using 26a (18 mg, 45 μmol)
and LiOHaq (0.5 M, 0.27 mL) and obtained as a white solid (10 mg, 58%
yield): 1H NMR (400 MHz, DMSO-d6) δ 8.53 (d, J = 8.1 Hz, 1H), 7.77 (s,
1H), 7.70 (d, J = 7.7 Hz, 1H), 7.63 (d, J = 8.1 Hz, 2H), 7.59 (d, J = 8.0 Hz,
1H), 7.48 (t, J = 7.9 Hz, 1H), 7.44 (d, J = 8.0 Hz, 2H), 4.63 – 4.40 (m, 1H),
3.02 – 2.89 (m, 2H), 2.61 – 2.51 (m, 2H); 13C NMR (101 MHz, DMSO-d6)
δ 172.3, 164.3, 143.6 (q, J = 1.0 Hz), 136.5, 133.1, 131.0, 130.3, 130.0
(2C), 126.9, 125.9, 125.0 (q, J = 3.6 Hz, 2C), 48.0, 39.3, 38.8 (chemical
Step
2:
Methyl
(S)-2-((tert-butoxycarbonyl)amino)-3-(4-
(trifluoromethyl)phenyl)propanoate (8). At 0 °C, Et3N (0.88 mL, 6.45 mmol)
was added dropwise to a solution of 7 hydrochloride salt (915 mg, 3.23
mmol) dissolved in dry DCM (16.7 mL). Di-tert-butyl dicarbonate (993 mg,
4.55 mmol) was added and the reaction was stirred at rt under argon until
consumption of starting material. The reaction mixture was concentrated
and purified by flash chromatography (EtOAc/PE, 1:10) to give 8 as a white
solid (974 mg, 87% yield): Rf = 0.15 (EtOAc/PE, 1:8); 1H NMR (400 MHz,
CDCl3) δ 7.55 (d, J = 8.1 Hz, 2H), 7.26 (d, J = 8.5 Hz, 2H), 5.03 (d, J = 7.6
Hz, 1H), 4.62 (dd, J = 13.5, 6.3 Hz, 1H), 3.73 (s, 3H), 3.21 (dd, J = 13.7,
5.6 Hz, 1H), 3.08 (dd, J = 13.7, 6.2 Hz, 1H), 1.41 (s, 9H); 13C NMR (101
1
2
shift values δCF, and δCF, not observed); HRMS (ESI) calcd for
J
J
C18H15F3ClNNaO3 (M + Na+) 408.0585, found 408.0584. HPLC: tR = 11.39
min, 98.9%. [α]20 = -3.0° (c = 0.20, MeOH).
D
10
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10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
MHz, CDCl3) δ 171.9, 155.0, 140.3 (app s), 129.7 (2C), 129.4 (q, J = 32.6
Hz), 125.4 (q, J = 3.5 Hz, 2C), 124.2 (q, J = 272.0 Hz), 80.2, 54.2, 52.3,
38.3, 28.2 (3C); HRMS (ESI) calcd for C16H20F3NNaO4 (M + Na+) 370.1237,
found 370.1226. NMR is in overall agreement with literature.[33]
14.5; HRMS (ESI) calcd for C22H23ClF3NNaO3 (M + Na+) 464.1211, found
464.1226.
Step 7: 29 was prepared as described for 1 using the ethyl ester of 29 (15
mg, 34 μmol) and LiOHaq (0.5 M, 0.17 mL) and obtained as a white solid
(13 mg, 90% yield): 1H NMR (400 MHz, CD3OD) δ 7.46 (d, J = 8.0 Hz, 2H),
7.29 (d, J = 8.0 Hz, 2H), 7.24 – 7.17 (m, 3H), 7.05 – 7.00 (m, 1H), 4.20 –
4.10 (m, 1H), 3.38 (s, 2H), 2.94 (dd, J = 13.8, 5.1 Hz, 1H), 2.75 (dd, J =
13.8, 9.1 Hz, 1H), 2.38 – 2.29 (m, 2H), 1.98 – 1.88 (m, 1H), 1.79 – 1.67 (m,
1H); 13C NMR (101 MHz, CD3OD) δ 176.8, 173.0, 144.3 (app s), 139.1,
135.2, 131.0 (2C), 130.1, 129.6 (q, J = 32.3 Hz), 128.3, 128.0, 126.0 (q, J
= 3.8 Hz, 2C), 125.8 (q, J = 271.0 Hz), 51.4, 43.5, 41.7, 31.6, 31.1; HRMS
(ESI) calcd for C20H19ClF3NNaO3 (M + Na+) 436.0898, found 436.0907.
Step
3:
Ethyl
(S,E)-4-((tert-butoxycarbonyl)amino)-5-(4-
(trifluoromethyl)phenyl)pent-2-enoate. A solution of 8 (385 mg, 1.11 mmol)
in dry DCM (3.3 mL) under argon was cooled to -78 °C, added DIBALH in
toluene (1 M, 2.2 mL), and the reaction stirred for 30 min. Then, the
phosphonium
salt
((ethoxycarbonylmethyl)triphenylphosphonium
bromide) (952 mg, 2.22 mmol) and KOtBu (250 mg, 2.23 mmol)
suspended in dry DCM (2 mL) were added and the reaction was stirred for
1 h at -78 °C, after which the reaction was brought to rt and stirred
overnight. The reaction mixture was added a saturated solution of
potassium sodium tartrate (15 mL) and stirred for 30 min. The mixture was
extracted with EtOAc (x3), the combined organic phases were dried over
Na2SO4, concentrated in vacuo, and purified by flash chromatography
(EtOAc/PE, 1:15 à 1:6) to give a white solid (253 mg, 59% yield): Rf =
0.28 (EtOAc/PE, 1:4); 1H NMR (400 MHz, CDCl3) δ 7.56 (d, J = 8.0 Hz,
2H), 7.30 (d, J = 8.0 Hz, 2H), 6.89 (dd, J = 15.7, 5.1 Hz, 1H), 5.88 (dd, J =
15.7, 1.7 Hz, 1H), 4.65 (s, 1H), 4.50 (s, 1H), 4.19 (q, J = 7.1 Hz, 2H), 3.04
– 2.86 (m, 2H), 1.38 (s, 9H), 1.28 (t, J = 7.1 Hz, 3H); 13C NMR (101 MHz,
CDCl3) δ 166.1, 155.0, 146.8, 140.8 (app s), 129.9 (2C), 129.4 (q, J = 32.2
Hz), 125.7 – 125.6 (m), 125.6 – 122.9 (m), 121.8, 80.3, 60.7, 52.2, 40.9,
28.4 (3C), 14.3; ESI-HRMS calcd for C19H24F3NNaO4 (M + Na+) 410.1550,
found 410.1563.
HPLC: tR = 11.42 min, 97.8%. [α]20 = +25° (c = 0.1, MeOH).
D
(S)-2-(2-(m-Tolyl)acetamido)-3-(4-(trifluoromethyl)phenyl)propanoic
acid (30). Step 1: The methyl ester of 30 was prepared as described for
15 using 11 (71 mg, 0.25 mmol), 2-(m-tolyl)acetic acid (41 mg, 0.27 mmol),
HOBt (46 mg, 0.30 mmol), DMAP (3.2 mg, 26 μmol), DIPEA (153 μL, 0.88
mmol) and EDC hydrochloride (57 mg, 0.30 mmol) and obtained after flash
chromatography (EtOAc/PE, 1:3) as a white solid (58 mg, 62% yield): Rf =
0.16 (EtOAc/PE, 1:2); 1H NMR (400 MHz, CD3OD) δ 7.37 (d, J = 8.0 Hz,
2H), 7.16 (d, J = 8.0 Hz, 2H), 7.01 (t, J = 7.6 Hz, 1H), 6.92 (d, J = 7.6 Hz,
1H), 6.89 (s, 1H), 6.82 (d, J = 7.5 Hz, 1H), 4.61 (dd, J = 9.4, 5.1 Hz, 1H),
3.60 (s, 3H), 3.37 – 3.23 (m, 2H), 3.14 (dd, J = 14.0, 5.1 Hz, 1H), 2.91 (dd,
J = 13.9, 9.5 Hz, 1H), 2.16 (s, 3H); 13C NMR (101 MHz, CD3OD) δ 173.9,
173.0, 142.8 (q, J = 1.4 Hz), 139.2, 136.4, 130.9 (2C), 130.8, 130.0 (q, J =
32.2 Hz), 129.4, 128.6, 127.1, 126.2 (q, J = 3.8 Hz, 2C), 125.7 (q, J = 271.1
Hz), 54.6, 52.8, 43.5, 37.9, 21.4; HRMS (ESI) calcd for C20H20F3NNaO3 (M
+ Na+) 402.1287, found 402.1306.
Step
4:
Ethyl
(R)-4-((tert-butoxycarbonyl)amino)-5-(4-
(trifluoromethyl)phenyl)pentanoate (9). A vial containing the ethyl (S,E)-4-
((tert-butoxycarbonyl)amino)-5-(4-(trifluoromethyl)phenyl)pent-2-enoate
(210 mg, 0.54 mmol) dissolved in EtOH (3.5 mL) was flushed with argon
(x2) and added Pd/C (10%, 22 mg). The vial was flushed with hydrogen
gas (x3), and the reaction was stirred (H2, balloon) at rt over night. The
reaction mixture was filtered through a plug of celite (EtOAc), concentrated
in vacuo, and purified by flash chromatography (EtOAc/PE, 1:11 à 1:9) to
give 9 as a white solid (182 mg, 86% yield): Rf = 0.14 (EtOAc/PE, 1:10);
1H NMR (400 MHz, CDCl3) δ 7.53 (d, J = 8.0 Hz, 2H), 7.29 (d, J = 8.0 Hz,
2H), 4.41 (d, J = 8.7 Hz, 1H), 4.11 (q, J = 7.1 Hz, 2H), 3.91 – 3.78 (m, 1H),
2.90 – 2.76 (m, 2H), 2.43 – 2.28 (m, 2H), 1.91 – 1.79 (m, 1H), 1.69 – 1.57
(m, 1H), 1.36 (s, 9H), 1.23 (t, J = 7.1 Hz, 3H); 13C NMR (101 MHz, CDCl3)
δ 173.5, 155.5, 142.3 (app s), 129.9 (2C), 129.0 (q, J = 32.7 Hz), 125.4 (q,
J = 3.7 Hz, 2C), 124.4 (q, J = 271.8 Hz), 79.5, 60.7, 51.6, 41.9, 31.3, 29.4,
28.4 (3C), 14.3; HRMS (ESI) calcd for C19H26F3NNaO4 (M + Na+) 412.1706,
found 412.1709. [α]20D = -2° (c = 0.1, DCM).
Step 2: 30 was prepared as described for 18 using the methyl ester of 30
(52 mg, 0.14 mmol) and LiOHaq (0.5 M, 0.70 mL) and obtained as a white
solid (47 mg, 94% yield): 1H NMR (400 MHz, CD3OD) δ 7.46 (d, J = 8.1
Hz, 2H), 7.28 (d, J = 8.0 Hz, 2H), 7.11 (t, J = 7.5 Hz, 1H), 7.03 (d, J = 7.7
Hz, 1H), 6.99 (s, 1H), 6.92 (d, J = 7.5 Hz, 1H), 4.71 (dd, J = 9.2, 4.8 Hz,
1H), 3.49 – 3.37 (m, 2H), 3.28 (dd, J = 14.0, 4.8 Hz, 1H), 3.03 (dd, J = 13.9,
9.2 Hz, 1H), 2.27 (s, 3H); 13C NMR (101 MHz, CD3OD) δ 174.2, 173.9,
143.0 (app s), 139.2, 136.4, 130.9 (2C), 130.8, 129.9 (q, J = 32.2 Hz),
129.4, 128.6, 127.1, 126.1 (q, J = 3.8 Hz, 2C), 125.8 (q, J = 270.8 Hz),
54.5, 43.6, 38.0, 21.4. m.p.: 124.4-126.6 °C. HRMS (ESI) calcd for
C19H18F3NNaO3 (M + Na+) 388.1131, found 388.1139. HPLC: tR = 11.53
min, 96.2%. [α]20 = +10° (c = 0.2, MeOH).
D
(R)-4-(2-(m-Tolyl)acetamido)-5-(4-(trifluoromethyl)phenyl)pentanoic
acid (31). Step 1: The ethyl ester of 31 was prepared as described for 15
using (R)-10 (20 mg, 61 μmol), 2-(m-tolyl)acetic acid (10 mg, 68 μmol),
HOBt (11 mg, 74 μmol), DMAP (0.9 mg, 7 μmol), DIPEA (37 μL, 0.22
mmol) and EDC hydrochloride (14 mg, 74 μmol) and obtained after flash
chromatography (EtOAc/PE, 1:3 à 1:2) as a white solid (15 mg, 57%
yield): Rf = 0.16 (EtOAc/PE, 1:2); 1H NMR (400 MHz, CD3OD) δ 7.45 (d, J
= 8.0 Hz, 2H), 7.28 (d, J = 8.0 Hz, 2H), 7.12 (t, J = 7.6 Hz, 1H), 7.03 (d, J
= 7.6 Hz, 1H), 7.00 (s, 1H), 6.91 (d, J = 7.5 Hz, 1H), 4.17 – 4.06 (m, 1H),
4.09 (q, J = 7.1 Hz, 2H), 3.35 (s, 2H), 2.91 (dd, J = 13.8, 5.3 Hz, 1H), 2.76
(dd, J = 13.7, 8.9 Hz, 1H), 2.36 – 2.30 (m, 2H), 2.28 (s, 3H), 1.97 – 1.87
(m, 1H), 1.76 – 1.65 (m, 1H), 1.22 (t, J = 7.1 Hz, 3H); 13C NMR (101 MHz,
CD3OD) δ 174.9, 173.9, 144.3 (q, J = 1.4 Hz), 139.2, 136.8, 130.9 (2C),
130.7, 129.6 (q, J = 32.2 Hz), 129.4, 128.5, 127.0, 126.0 (q, J = 3.8 Hz,
2C), 125.8 (q, J = 270.9 Hz), 61.6, 51.2, 44.0, 41.7, 31.8, 30.9, 21.4, 14.5;
HRMS (ESI) calcd for C23H27F3NO3 (M + H+) 422.1938, found 422.1952.
Step 5: Ethyl (R)-4-amino-5-(4-(trifluoromethyl)phenyl)pentanoate
hydrochloride ((R)-10). In a vial, 9 (47.0 mg, 0.12 mmol) was cooled to 0 °C
and added HCl in dioxane (4 M, 2.4 mL), after which the reaction was
brought to rt and stirred under argon until complete consumption of starting
material as indicated by TLC. The reaction mixture was concentrated in
vacuo to give (R)-10 as a pale yellow solid (39 mg, 99% yield): Rf (free
amine) = 0.06 (EtOAc [5% MeOH]); HRMS (ESI) calcd for C14H19F3NO2
(M + H+) 290.1362, found 290.1376.
Step
6:
Ethyl
(R)-4-(2-(3-chlorophenyl)acetamido)-5-(4-
(trifluoromethyl)phenyl)pentanoate. The ethyl ester of 29 was prepared as
described for 15 using (R)-10 (17 mg, 52 μmol), 2-(3-chlorophenyl)acetic
acid (10 mg, 60 μmol), HOBt (10 mg, 63 μmol), DMAP (0.7 mg, 6 μmol),
DIPEA (32 μL, 0.18 mmol) and EDC hydrochloride (12 mg, 63 μmol) and
obtained after flash chromatography (EtOAc/PE, 1:2) as a white solid (17
mg, 75% yield): Rf = 0.17 (EtOAc/PE, 1:2); 1H NMR (400 MHz, CD3OD) δ
7.47 (d, J = 8.0 Hz, 2H), 7.29 (d, J = 8.0 Hz, 2H), 7.25 – 7.18 (m, 3H), 7.03
(m, 1H), 4.18 – 4.06 (m, 1H), 4.09 (q, J = 7.1 Hz, 2H), 3.38 (s, 2H), 2.92
(dd, J = 13.8, 5.2 Hz, 1H), 2.75 (dd, J = 13.8, 9.0 Hz, 1H), 2.38 – 2.31 (m,
Step 2: 31 was prepared as described for 1 using the ethyl ester of 31 (15
mg, 35 μmol) and LiOHaq (0.5 M, 0.17 mL) and obtained as a white solid
(11 mg, 82% yield): 1H NMR (400 MHz, CD3OD) δ 7.45 (d, J = 8.0 Hz, 2H),
7.28 (d, J = 8.0 Hz, 2H), 7.12 (t, J = 7.6 Hz, 1H), 7.03 (d, J = 7.6 Hz, 1H),
6.99 (s, 1H), 6.92 – 6.89 (m, 1H), 4.19 – 4.09 (m, 1H), 3.35 (s, 2H), 2.92
(dd, J = 13.7, 5.2 Hz, 1H), 2.76 (dd, J = 13.7, 8.9 Hz, 1H), 2.35 – 2.29 (m,
2H), 2.28 (s, 3H), 1.97 – 1.85 (m, 1H), 1.77 – 1.65 (m, 1H); 13C NMR (101
2H), 1.98 – 1.89 (m, 1H), 1.77 – 1.67 (m, 1H), 1.22 (t, J = 7.1 Hz, 3H); 13
C
NMR (101 MHz, CD3OD) δ 174.8, 172.9, 144.3 (q, J = 1.4 Hz), 139.2,
135.2, 130.9 (2C), 130.1, 129.6 (q, J = 32.2 Hz), 128.4, 128.0, 126.1 (q, J
= 3.8 Hz, 2C), 125.8 (q, J = 270.8 Hz), 61.6, 51.2, 43.5, 41.7, 31.8, 30.9,
11
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10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
MHz, CD3OD) δ 176.9, 173.9, 144.3 (q, J = 1.1 Hz), 139.2, 136.8, 130.9
(2C), 130.7, 129.5 (q, J = 32.2 Hz), 129.4, 128.5, 127.0, 126.0 (q, J = 3.8
Hz, 2C), 125.8 (q, J = 271.9 Hz), 51.3, 44.0, 41.6, 31.6, 31.0, 21.4. m.p.:
168.3-171.9 °C. HRMS (ESI) calcd for C21H23F3NO3 (M + H+) 394.1625,
+ Na+) 432.1393, found 432.1405. HPLC: tR = 10.92 min, 97.8%. [α]20
=
D
+29° (c = 0.2, MeOH).
(R)-4-(2-(Naphthalen-1-yl)acetamido)-5-(4-(trifluoromethyl)phenyl)-
pentanoic acid (34). Step 1: The ethyl ester of 34 was prepared as
described for 15 using (R)-10 (30 mg, 93 μmol), 2-(naphthalen-1-yl)acetic
acid (20 mg, 107 μmol), HOBt (17 mg, 112 μmol), DMAP (2 mg, 15 μmol),
DIPEA (60 μL, 0.34 mmol) and EDC hydrochloride (22 mg, 115 μmol) and
obtained after flash chromatography (DCM [1% MeOH] à DCM [2%
MeOH]) as a white solid (28 mg, 68% yield): Rf = 0.23 (DCM [2% MeOH]);
1H NMR (400 MHz, CDCl3) δ 7.93 – 7.81 (m, 3H), 7.56 – 7.47 (m, 2H),
7.43 (dd, J = 8.3, 6.9 Hz, 1H), 7.30 (dd, J = 7.0, 1.2 Hz, 1H), 7.25 (d, J =
7.6 Hz, 2H), 6.80 (d, J = 7.9 Hz, 2H), 5.14 (d, J = 8.9 Hz, 1H), 4.23 – 4.13
(m, 1H), 4.11 – 4.02 (m, 2H), 4.02 – 3.89 (m, 2H), 2.67 – 2.56 (m, 2H),
2.30 – 2.09 (m, 2H), 1.77 – 1.68 (m, 1H), 1.50 – 1.36 (m, 1H), 1.23 (t, J =
7.2 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 173.3, 170.6, 141.3 (q, J = 1.3
Hz), 134.2, 132.1, 131.1, 129.6 (q, J = 37.5 Hz), 129.5 (C2), 129.3, 129.0,
128.7, 128.4, 127.1, 126.5, 125.8, 125.3 (q, J = 3.8 Hz, 2C), 124.3 (q, J =
271.9 Hz), 123.8, 60.7, 49.7, 42.1, 40.7, 31.0, 29.1, 14.3; ESI-HRMS calcd
for C26H26F3NNaO3 (M + Na+) 480.1757, found 480.1779.
found 394.1642. HPLC: tR = 11.32 min, 95.9%. [α]20 = +8° (c = 0.1,
D
MeOH).
(R)-5-(4-(Trifluoromethyl)phenyl)-4-(2-(3-(trifluoromethyl)phenyl)-
acetamido)pentanoic acid (32). Step 1: The ethyl ester of 32 was
prepared as described for 15 using (R)-10 (20 mg, 62 μmol), 2-(3-
(trifluoromethyl)phenyl)acetic acid (15 mg, 72 μmol), HOBt (12 mg, 78
μmol), DMAP (2 mg, 16 μmol), DIPEA (40 μL, 0.22 mmol) and EDC
hydrochloride (15 mg, 78 μmol) and obtained after flash chromatography
(EtOAc/PE, 1:3) as a white solid (12 mg, 39% yield): Rf = 0.12 (EtOAc/PE
1:2); 1H NMR (400 MHz, CDCl3) δ 7.55 (d, J = 7.8 Hz, 1H), 7.49 – 7.40 (m,
4H), 7.34 (d, J = 7.7 Hz, 1H), 7.2 (d, J = 8.0 Hz, 2H), 5.52 (d, J = 8.4 Hz,
1H), 4.24 – 4.15 (m, 1H), 4.14 – 4.05 (m, 2H), 3.55 – 3.46 (m, 2H), 2.88 –
2.76 (m, 2H), 2.42 – 2.23 (m, 2H), 1.89 – 1.79 (m, 1H), 1.72 – 1.62 (m, 1H),
1.23 (t, J = 7.2 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 173.8, 169.8, 141.7
(app s), 135.8, 132.7, 131.4 (q, J = 32.5 Hz), 129.7 (2C), 129.5, 129.2 (q,
J = 32.5 Hz), 126.1 (q, J = 3.8 Hz), 125.4 (q, J = 3.8 Hz, 2C), 124.3 (q, J =
3.8 Hz), 124.3 (q, J = 272.0 Hz), 124.1 (q, J = 272.4 Hz), 60.9, 50.6, 43.7,
41.0, 31.1, 28.7, 14.3; ESI-HRMS calcd for C23H23F6NNaO3 (M + Na+)
498.1474, found 498.1465.
Step 2: 34 was prepared as described for 1 using the ethyl ester of 34 (21
mg, 46 μmol) and LiOHaq (0.5 M, 0.23 mL) and obtained after
recrystallization (MeOH) as a white solid (17 mg, 86% yield): 1H NMR (400
MHz, DMSO-d6) δ 12.05 (br s, 1H), 8.05 (d, J = 8.7 Hz, 1H), 7.91 (dd, J =
14.0, 8.2 Hz, 2H), 7.79 (d, J = 8.2 Hz, 1H), 7.56 – 7.31 (m, 6H), 7.28 (d, J
= 7.0 Hz, 1H), 4.04 – 3.94 (m, 1H), 3.89 – 3.76 (m, 2H), 2.84 (dd, J = 13.7,
5.2 Hz, 1H), 2.74 (dd, J = 13.5, 8.4 Hz, 1H), 2.37 – 2.14 (m, 2H), 1.84 –
1.71 (m, 1H), 1.65 – 1.54 (m, 1H); 13C NMR (101 MHz, DMSO-d6) δ 174.1,
169.6, 143.7 (app s), 133.2, 132.6, 131.9, 129.9 (2C), 128.2, 127.6, 126.9,
126.7 (q, J = 31.4 Hz), 125.7, 125.4, 125.3, 124.7 (q, J = 3.5 Hz, 2C), 124.8
(q, J = 271.0 Hz), 124.1, 49.3, 40.1, 32.5, 30.6, 30.4, 29.6; ESI-HRMS
calcd for C24H22F3NNaO3 (M + Na+) 452.1444, found 452.1459. HPLC: tR
= 11.12 min, 99.5%. [α]20D = +37° (c = 0.2, MeOH).
Step 2: 32 was prepared as described for 1 using the ethyl ester of 32 (11
mg, 22 μmol) and LiOHaq (0.5 M, 0.11 mL) and obtained as a white solid
(10 mg, 98% yield): 1H NMR (400 MHz, CD3OD) δ 7.52 (d, J = 5.8 Hz, 2H),
7.47 – 7.39 (m, 3H), 7.35 (d, J = 7.7 Hz, 1H), 7.29 (d, J = 8.1 Hz, 2H), 4.21
– 4.11 (m, 1H), 3.52 – 3.44 (m, 2H), 2.94 (dd, J = 13.7, 5.2 Hz, 1H), 2.76
(dd, J = 13.8, 9.1 Hz, 1H), 2.36 – 2.30 (m, 2H), 1.99 – 1.88 (m, 1H), 1.80
– 1.68 (m, 1H); 13C NMR (101 MHz, CD3OD) δ 176.8, 172.8, 144.3 (app
s), 138.3, 133.7, 131.8 (q, J = 32.0 Hz), 130.9 (2C), 130.2, 129.6 (q, J =
32.2 Hz), 126.8 (q, J = 3.9 Hz), 126.0 (q, J = 3.8 Hz, 2C), 125.8 (q, J =
270.9 Hz), 125.6 (q, J = 271.7 Hz), 124.6 (q, J = 3.9 Hz), 51.4, 43.5, 41.7,
31.6, 31.1; ESI-HRMS calcd for C21H19F6NNaO3 (M + Na+) 470.1161,
(R)-4-(2-(3-Pentylphenyl)acetamido)-5-(4-(trifluoromethyl)phenyl)-
pentanoic acid (35). Step 1: The ethyl ester of 35 was prepared as
described for 15 using (R)-10 (30 mg, 93 μmol), 2-(3-pentylphenyl)acetic
acid (21 mg, 102 μmol), HOBt (18 mg, 120 μmol), DMAP (2 mg, 18 μmol),
DIPEA (60 μL, 0.34 mmol) and EDC hydrochloride (25 mg, 128 μmol) and
obtained after flash chromatography (DCM [0.5% MeOH] à DCM [1%
MeOH]) as a white solid (32 mg, 71% yield): Rf = 0.16 (DCM [2% MeOH]);
1H NMR (400 MHz, CDCl3) δ 7.45 (d, J = 8.0 Hz, 2H), 7.23 (t, J = 7.6 Hz,
1H), 7.14 – 7.07 (m, 3H), 6.97 (s, 1H), 6.93 (d, J = 7.5 Hz, 1H), 5.26 (d, J
= 8.7 Hz, 1H), 4.23 – 4.14 (m, 1H), 4.14 – 4.05 (m, 2H), 3.51 – 3.42 (m,
2H), 2.77 (d, J = 6.4 Hz, 2H), 2.61 – 2.53 (m, 2H), 2.39 – 2.20 (m, 2H),
1.90 – 1.80 (m, 1H), 1.64 – 1.51 (m, 2H), 1.40 – 1.27 (m, 4H), 1.23 (t, J =
7.2 Hz, 3H), 0.88 (t, J = 6.9 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 173.4,
170.9, 144.1, 141.7 (q, J = 0.7 Hz), 134.7, 129.8 (2C), 129.6, 129.1, 129.0
(q, J = 33.9 Hz), 127.7, 126.7, 125.4 (q, J = 3.7 Hz, 2C), 124.4 (q, J =
271.9 Hz), 60.7, 49.9, 44.2, 40.9, 35.9, 31.7, 31.2, 31.1, 29.0, 22.6, 14.3,
14.1; ESI-HRMS calcd for C27H34F3NNaO3 (M + Na+) 500.2383, found
500.2392.
found 470.1138. HPLC: tR = 11.64 min, 98.2%. [α]20 = +13° (c = 0.2,
D
MeOH).
(R)-4-(2-(3-Methoxyphenyl)acetamido)-5-(4-(trifluoromethyl)phenyl)-
pentanoic acid (33). Step 1: The ethyl ester of 33 was prepared as
described for 15 using (R)-10 (21 mg, 63 μmol), 2-(3-
methoxyphenyl)acetic acid (12 mg, 75 μmol), HOBt (16 mg, 107 μmol),
DMAP (0.8 mg,
7 μmol), DIPEA (40 μL, 0.22 mmol) and EDC
hydrochloride (17 mg, 87 μmol) and obtained after flash chromatography
(EtOAc/PE, 1:3) as a white solid (7 mg, 27% yield): Rf = 0.10 (EtOAc/PE
1:2); 1H NMR (400 MHz, CDCl3) δ 7.46 (d, J = 7.2 Hz, 2H), 7.26 – 7.21 (m,
1H), 7.10 (d, J = 8.0 Hz, 2H), 6.87 – 6.82 (m, 1H), 6.73 – 6.68 (m, 2H),
5.33 – 5.27 (m, 1H), 4.23 – 4.13 (m, 1H), 4.10 (qd, J = 7.2, 1.0 Hz 2H),
3.79 (s, 3H), 3.46 (s, 2H), 2.83 – 2.72 (m, 2H), 2.39 – 2.22 (m, 2H), 1.81
(ddt, J = 11.6, 7.5, 4.3 Hz, 1H), 1.65 – 1.54 (m, 1H), 1.24 (t, J = 7.2 Hz,
3H); 13C NMR (101 MHz, CDCl3) δ 173.5, 170.6, 160.3, 141.7 (app s),
136.3, 130.3, 129.8 (2C), 129.0 (q, J = 32.3 Hz), 125.4 (q, J = 3.8 Hz, 2C),
124.4 (q, J = 272.0 Hz), 120.3, 115.2, 113.0, 60.8, 55.3, 50.0, 44.2, 40.9,
31.1, 29.0, 14.3; ESI-HRMS calcd for C23H26F3NNaO4 (M + Na+) 460.1706,
found 460.1410.
Step 2: 35 was prepared as described for 1 using the ethyl ester of 35
(22 mg, 47 μmol) and LiOHaq (0.5 M, 0.24 mL) and obtained as a white
solid (21 mg, >99% yield): 1H NMR (400 MHz, CDCl3) δ 7.45 (d, J = 8.0
Hz, 2H), 7.25 – 7.20 (m, 1H), 7.12 (d, J = 7.7 Hz, 1H), 7.07 (d, J = 8.0 Hz,
2H), 6.95 (s, 1H), 6.91 (d, J = 7.5 Hz, 1H), 5.32 (d, J = 9.0 Hz, 1H), 4.32 –
4.20 (m, 1H), 3.48 (s, 2H), 2.80 (dd, J = 13.9, 6.0 Hz, 1H), 2.72 (dd, J =
13.9, 6.9 Hz, 1H), 2.61 – 2.53 (m, 2H), 2.43 – 2.28 (m, 2H), 1.85 (ddt, J =
11.5, 7.5, 4.0 Hz, 1H), 1.65 – 1.52 (m, 3H), 1.38 – 1.27 (m, 4H), 0.88 (t, J
= 6.9 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 177.2, 171.8, 144.2, 141.4 (q,
J = 0.9 Hz), 134.4, 129.7 (2C), 129.6, 129.2, 129.1 (q, J = 32.5 Hz), 127.8,
126.7, 125.5 (q, J = 3.7 Hz, 2C), 124.3 (q, J = 272.0 Hz), 49.7, 43.9, 40.8,
35.9, 31.6, 31.2, 30.9, 29.4, 22.6, 14.1. m.p.: 128.4-130.9 °C. ESI-HRMS
calcd for C25H30F3NO3Na (M + Na+) 472.2070, found 472.2091. HPLC: tR
= 13.09 min, 95.0%; [α]20D = +30° (c = 0.2, MeOH).
Step 2: 33 was prepared as described for 1 using the ethyl ester of 33 (7
mg, 16 μmol) and LiOHaq (0.5 M, 80 μL) and obtained as a white solid (6
mg, 95% yield): 1H NMR (400 MHz, CD3OD) δ 7.45 (d, J = 8.0 Hz, 2H),
7.27 (d, J = 8.0 Hz, 2H), 7.18 – 7.12 (m, 1H), 6.81 – 6.75 (m, 2H), 6.74 –
6.67 (m, 2H), 4.18 – 4.08 (m, 1H), 3.76 (s, 3H), 2.92 (dd, J = 13.7, 5.2 Hz,
1H), 2.76 (dd, J = 13.7, 8.8 Hz, 1H), 2.41 – 2.23 (m, 2H), 1.97 – 1.86 (m,
1H), 1.78 – 1.66 (m, 1H); 13C NMR (101 MHz, CD3OD) δ 177.0, 173.7,
161.3, 144.3 (app s), 138.3, 131.0 (2C), 130.5, 129.6 (q, J = 32.2 Hz),
126.1 (q, J = 3.8 Hz, 2C), 126.0 (q, J = 270.6 Hz), 122.3, 115.8, 113.4,
55.6, 51.4, 44.1, 41.6, 31.7, 31.0; ESI-HRMS calcd for C21H22F3NNaO4 (M
12
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10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
(4R)-4-(2-Phenylpropanamido)-5-(4-(trifluoromethyl)phenyl)-
(R)-4-(2-(3,4-Difluorophenyl)acetamido)-5-(4-(trifluoromethyl)-
pentanoic acid (36). Step 1: The ethyl ester of 36 of the title compound
was prepared as described for 15 using (R)-10 (30 mg, 93 μmol), 2-
phenylpropanoic acid (20 μL, 111 μmol), HOBt (18 mg, 116 μmol), DMAP
(1 mg, 10 μmol), DIPEA (60 μL, 0.34 mmol) and EDC hydrochloride (23
mg, 121 μmol) and obtained after flash chromatography (DCM [1% MeOH]
à DCM [2% MeOH]) as a white solid (24 mg, 62% yield): Rf = 0.13 (DCM
[2% MeOH]); 1H NMR (400 MHz, CDCl3) δ 7.48 (d, J = 8.0 Hz, 2H), 7.38
(d, J = 8.0 Hz, 2H), 7.34 – 7.27 (m, 6H), 7.23 – 7.18 (m, 2H), 7.18 – 7.13
(m, 4H), 6.99 (d, J = 8.0 Hz, 2H), 5.31 (d, J = 8.5 Hz, 1H), 5.21 (d, J = 8.8
Hz, 1H), 4.26 – 3.95 (m, 6H), 3.51 – 3.39 (m, 2H), 2.86 – 2.64 (m, 4H),
2.40 – 2.20 (m, 3H), 2.18 – 2.07 (m, 1H), 1.86 – 1.73 (m, 2H), 1.63 – 1.52
(m, 2H), 1.46 (d, J = 7.2 Hz, 3H), 1.43 (d, J = 7.1 Hz, 3H), 1.25 (t, J = 7.2
Hz, 3H), 1.20 (t, J = 7.2 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 174.0,
173.7, 173.5, 173.4, 141.8, 141.5, 141.3, 141.0, 129.6 (2C), 129.6 – 128.2
(m) 129.0, 128.9, 127.5, 127.5, 127.3, 127.3, 125.5 – 125.1 (m), 125.9 –
122.7 (m), 60.7, 60.6, 50.1, 49.7, 47.4, 47.2, 41.0, 40.7, 31.1, 30.9, 29.0,
28.7, 18.3, 18.2, 14.2, 14.2; ESI-HRMS calcd for C23H26F3NNaO3 (M +
Na+) 444.1757, found 444.1770.
phenyl)pentanoic acid (38). Step 1: The ethyl ester of 38 of the title
compound was prepared as described for 15 using (R)-10 (20 mg, 62
μmol), 2-(3,4-difluorophenyl)acetic acid (14 mg, 81 μmol), HOBt (12 mg,
78 μmol), DMAP (0.8 mg, 7 μmol), DIPEA (40 μL, 0.23 mmol) and EDC
hydrochloride (18 mg, 92 μmol) and obtained after flash chromatography
(EtOAc/PE, 1:3) as a white solid (18 mg, 66% yield): Rf = 0.08 (EtOAc/PE,
1:2); 1H NMR (400 MHz, CDCl3) δ 7.50 (d, J = 8.0 Hz, 2H), 7.19 (d, J = 8.0
Hz, 2H), 7.12 – 7.05 (m, 1H), 6.97 (ddd, J = 10.8, 7.5, 2.1 Hz, 1H), 6.84
(ddd, J = 8.5, 4.1, 1.8 Hz, 1H), 5.49 (d, J = 8.5 Hz, 1H), 4.22 – 4.04 (m,
3H), 3.44 – 3.35 (m, 2H), 2.89 – 2.73 (m, 2H), 2.42 – 2.22 (m, 2H), 1.89 –
1.79 (m, 1H), 1.73 – 1.63 (m, 1H), 1.24 (t, J = 7.2 Hz, 3H); 13C NMR (101
MHz, CDCl3) δ 173.8, 169.9, 150.5 (dd, J = 249.3, 12.8 Hz), 149.9 (dd, J
= 248.5, 12.4 Hz), 141.8 (q, J = 1.1 Hz), 131.7 (dd, J = 5.9, 4.0 Hz), 129.7
(2C), 129.2 (q, J = 32.5 Hz), 125.5 (q, J = 3.8 Hz, 2C), 125.3 (dd, J = 6.2,
3.6 Hz), 125.6 – 122.9 (m), 118.1 (d, J = 57.7 Hz), 118.0 (d, J = 57.6 Hz),
60.9, 50.7, 43.0 (d, J = 0.8 Hz), 41.1, 31.1, 28.7, 14.3; ESI-HRMS calcd
for C22H22F5NNaO3 (M + Na+) 466.1412, found 466.1430.
Step 2: 38 was prepared as described for 1 using the ethyl ester of 38 (14
mg, 33 μmol) and LiOHaq (0.5 M, 0.17 mL) and obtained as a white solid
(12 mg, 90% yield): 1H NMR (400 MHz, CD3OD) δ 7.47 (d, J = 8.0 Hz, 2H),
7.31 (d, J = 8.0 Hz, 2H), 7.13 – 7.05 (m, 1H), 7.05 – 6.97 (m, 1H), 6.90 –
6.84 (m, 1H), 4.22 – 4.12 (m, 1H), 3.36 (s, 2H), 2.95 (dd, J = 13.8, 5.1 Hz,
1H), 2.74 (dd, J = 13.8, 9.3 Hz, 1H), 2.33 (t, J = 7.5 Hz, 2H), 2.00 – 1.88
(m, 1H), 1.79 – 1.68 (m, 1H); 13C NMR (101 MHz, CD3OD) δ 176.8, 172.8,
151.2 (dd, J = 246.5, 12.8 Hz), 150.6 (dd, J = 245.6, 12.6 Hz), 144.3 (q, J
= 1.2 Hz), 134.3 (dd, J = 6.1, 4.0 Hz), 130.9 (2C), 129.6 (q, J = 32.0 Hz),
126.4 (dd, J = 6.3, 3.6 Hz), 126.0 (q, J = 3.8 Hz, 2C), 125.8 (q, J = 271.0
Hz), 118.9 (d, J = 17.7 Hz), 118.0 (d, J = 17.3 Hz), 51.3, 42.9 (d, J = 0.9
Hz), 41.7, 31.6, 31.1; ESI-HRMS calcd for C20H18F5NNaO3 (M + Na+)
438.1099, found 438.1098. HPLC: tR = 11.12 min, 98.0%. [α]20D = +5° (c =
0.2, MeOH).
Step 2: 36 was prepared as described for 1 using the ethyl ester of 36 (11
mg, 26 μmol) and LiOHaq (0.5 M, 0.13 mL) and obtained as a white solid
(10 mg, 99% yield): 1H NMR (400 MHz, CDCl3) δ 7.48 (d, J = 7.9 Hz, 2H),
7.39 (d, J = 8.0 Hz, 2H), 7.35 – 7.27 (m, 6H), 7.22 – 7.16 (m, 2H), 7.15 –
7.09 (m, 4H), 6.99 (d, J = 8.0 Hz, 2H), 5.30 – 5.14 (m, 2H), 4.35 – 4.13 (m,
2H), 3.54 – 3.41 (m, 2H), 2.89 – 2.73 (m, 3H), 2.66 (dd, J = 13.9, 7.2 Hz,
1H), 2.44 – 2.15 (m, 4H), 1.94 – 1.76 (m, 2H), 1.68 – 1.51 (m, 2H), 1.47 (d,
J = 7.2 Hz, 3H), 1.42 (d, J = 7.2 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ
176.9, 176.8, 174.7, 174.4, 141.5, 141.2, 141.0, 140.7, 129.5 (2C), 129.1,
129.1, 129.3 – 128.7 (m), 127.6, 127.5, 127.5, 127.5, 125.5 – 125.2 (m),
125.6 – 122.7 (m), 49.8, 49.5, 47.3, 47.1, 40.9, 40.8, 30.9, 30.6, 29.5, 29.2,
18.1, 18.1; ESI-HRMS calcd for C21H22F3NNaO3 (M + Na+) 416.1444,
found 416.1464. HPLC: tR = 11.35 min, 95.1%. [α]20 = +88° (c = 0.2,
D
MeOH).
(R)-4-(2-(3,5-Difluorophenyl)acetamido)-5-(4-(trifluoromethyl)-
phenyl)pentanoic acid (39). Step 1: The ethyl ester of 39 was prepared
as described for 15 using (R)-10 (20 mg, 62 μmol), 2-(3,5-
difluorophenyl)acetic acid (13 mg, 74 μmol), HOBt (11 mg, 74 μmol),
DMAP (3 mg, 21 μmol), DIPEA (40 μL, 0.23 mmol) and EDC hydrochloride
(14 mg, 73 μmol) and obtained after flash chromatography (EtOAc/PE,
1:3) as a white solid (18 mg, 66% yield): Rf = 0.12 (EtOAc/PE, 1:2); 1H
NMR (400 MHz, CDCl3) δ 7.50 (d, J = 8.0 Hz, 2H), 7.19 (d, J = 8.0 Hz, 2H),
6.77 – 6.66 (m, 3H), 5.54 (d, J = 8.4 Hz, 1H), 4.22 – 4.15 (m, 1H), 4.15 –
4.05 (m, 2H), 3.47 – 3.37 (m, 2H), 2.89 – 2.78 (m, 2H), 2.39 (dt, J = 17.0,
7.1 Hz, 1H), 2.28 (dt, J = 17.0, 6.9 Hz, 1H), 1.89 – 1-79 (m, 1H), 1.74 –
1.64 (m, 1H), 1.24 (t, J = 7.1 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 173.9,
169.3, 163.3 (dd, J = 249.5, 12.9 Hz), 141.7 (q, J = 1.0 Hz), 138.4 (t, J =
9.4 Hz), 129.7 (2C), 129.2 (q, J = 32.6 Hz), 125.5 (q, J = 3.8 Hz, 2C), 124.3
(q, J = 272.0 Hz), 112.3 (dd, J = 18.4, 7.0 Hz, 2C), 103.0 (t, J = 25.1 Hz),
60.9, 50.7, 43.6 (t, J = 1.9 Hz), 41.0, 31.1, 28.6, 14.3; ESI-HRMS calcd for
C22H22F5NNaO3 (M + Na+) 466.1412, found 466.1416.
(R)-4-(2-(3-Fluorophenyl)acetamido)-5-(4-(trifluoromethyl)phenyl)-
pentanoic acid (37). Step 1: The ethyl ester of 37 was prepared as
described for 15 using (R)-10 (20 mg, 62 μmol), 2-(3-fluorophenyl)acetic
acid (11 mg, 73 μmol), HOBt (11 mg, 74 μmol), DMAP (2 mg, 15 μmol),
DIPEA (40 μL, 0.23 mmol) and EDC hydrochloride (15 mg, 78 μmol) and
obtained after flash chromatography (EtOAc/PE, 1:3) as a white solid (21
mg, 79% yield): Rf = 0.14 (EtOAc/PE, 1:2); 1H NMR (400 MHz, CDCl3) δ
7.48 (d, J = 8.0 Hz, 2H), 7.31 – 7.24 (m, 1H), 7.15 (d, J = 8.0 Hz, 2H), 7.02
– 6.96 (m, 1H), 6.90 (d, J = 7.6 Hz, 1H), 6.88 – 6.84 (m, 1H), 5.39 (d, J =
8.6 Hz, 1H), 4.23 – 4.14 (m, 1H), 4.14 – 4.05 (m, 2H), 3.51 – 3.41 (m, 2H),
2.80 (d, J = 6.6 Hz, 2H), 2.41 - 2.23 (m, 2H), 1.88 – 1.78 (m, 1H), 1.70 –
1.60 (m, 1H), 1.24 (t, J = 7.1 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 173.7,
170.0, 163.1 (d, J = 247.2 Hz), 141.7 (app s), 137.2 (d, J = 7.4 Hz), 130.6
(d, J = 8.4 Hz), 129.7 (2C), 129.1 (q, J = 32.5 Hz), 125.5 (q, J = 3.8 Hz,
2C), 125.0 (d, J = 3.0 Hz), 124.3 (q, J = 272.0 Hz), 116.4 (d, J = 21.5 Hz),
114.5 (d, J = 21.0 Hz), 60.9, 50.4, 43.7 (d, J = 1.6 Hz), 41.0, 31.1, 28.9,
14.3.; ESI-HRMS calcd for C22H23F4NNaO3 (M + Na+) 448.1506, found
448.1509.
Step 2: 39 was prepared as described for 1 using the ethyl ester of 39 (15
mg, 34 μmol) and LiOHaq (0.5 M, 0.17 mL) and obtained as a white solid
(13 mg, 91% yield): 1H NMR (400 MHz, CD3OD) δ 7.46 (d, J = 8.0 Hz, 2H),
7.31 (d, J = 8.1 Hz, 2H), 6.81 – 6.69 (m, 3H), 4.22 – 4.12 (m, 1H), 3.39 (s,
2H), 2.95 (dd, J = 13.8, 5.0 Hz, 1H), 2.74 (dd, J = 13.8, 9.3 Hz, 1H), 2.34
(t, J = 7.6 Hz, 2H), 1.99 – 1.89 (m, 1H), 1.79 – 1.69 (m, 1H); 13C NMR (101
MHz, CD3OD) δ 176.9, 172.3, 164.3 (dd, J = 247.0, 13.0 Hz), 144.3 (q, J
= 1.0 Hz), 141.1 (t, J = 9.8 Hz), 130.9 (2C), 129.7 (q, J = 32.3 Hz), 126.0
(q, J = 3.8 Hz, 2C), 125.8 (q, J = 271.0 Hz), 112.9 (dd, J = 18.6, 6.8 Hz,
2C), 103.0 (t, J = 25.7 Hz), 51.4, 43.4 (t, J = 1.8 Hz), 41.7, 31.7, 31.1; ESI-
HRMS calcd for C20H18F5NNaO3 (M + Na+) 438.1099, found 438.1087.
HPLC: tR = 11.20 min, 96.7%. [α]20D = +2° (c = 0.2, MeOH).
Step 2: 37 was prepared as described for 1 using the ethyl ester of 37 (15
mg, 35 μmol) and LiOHaq (0.5 M, 0.17 mL) and obtained as a white solid
(11 mg, 82% yield): 1H NMR (400 MHz, CD3OD) δ 7.46 (d, J = 8.0 Hz, 2H),
7.30 (d, J = 8.0 Hz, 2H), 7.27 – 7.20 (m, 1H), 6.97 – 6.85 (m, 3H), 4.21 –
4.11 (m, 1H), 3.40 (s, 2H), 2.94 (dd, J = 13.8, 5.1 Hz, 1H), 2.75 (dd, J =
13.8, 9.1 Hz, 1H), 2.38 – 2.29 (m, 2H), 1.99 – 1.87 (m, 1H), 1.78 – 1.68 (m,
1H); 13C NMR (101 MHz, CD3OD) δ 177.0, 173.0, 165.4, 163.0, 144.3 (app
s), 139.5 (d, J = 7.8 Hz), 131.1 (d, J = 8.4 Hz), 130.9 (2C), 129.6 (q, J =
32.2 Hz), 126.1 (q, J = 3.8 Hz, 2C), 125.8 (d, J = 2.9 Hz), 130.0 – 121.7
(m), 116.8 (d, J = 21.9 Hz), 114.5 (d, J = 21.3 Hz), 51.4, 43.6 (d, J = 1.8
Hz) 41.7, 31.7, 31.1; ESI-HRMS calcd for C20H19F4NNaO3 (M + Na+)
420.1193, found 420.1178. HPLC: tR = 11.03 min, 97.4%. [α]20D = +8° (c =
0.2, MeOH).
(R)-4-(2-(2,3-Difluorophenyl)acetamido)-5-(4-(trifluoromethyl)-
phenyl)pentanoic acid (40). Step 1: The ethyl ester of 40 was prepared
as described for 15 using (R)-10 (20 mg, 62 μmol), 2-(3,4-
13
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10.1002/cmdc.202100356
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difluorophenyl)acetic acid (12 mg, 68 μmol), HOBt (12 mg, 76 μmol),
DMAP (2 mg, 16 μmol), DIPEA (40 μL, 0.23 mmol) and EDC hydrochloride
(16 mg, 85 μmol) and obtained after flash chromatography (EtOAc/PE,
1:4) as a white solid (16 mg, 59% yield): Rf = 0.13 (EtOAc/PE, 1:2); 1H
NMR (400 MHz, CDCl3) δ 7.47 (d, J = 8.0 Hz, 2H), 7.20 (d, J = 8.0 Hz, 2H),
7.13 – 7.05 (m, 1H), 7.05 – 6.98 (m, 1H), 6.96 – 6.90 (m, 1H), 5.57 (d, J =
8.5 Hz, 1H), 4.23 – 4.14 (m, 1H), 4.11 (q, J = 7.1 Hz, 2H), 3.56 – 3.43 (m,
2H), 2.83 (d, J = 6.6 Hz, 2H), 2.43 – 2.25 (m, 2H), 1.90 – 1.80 (m, 1H),
1.73 – 1.62 (m, 1H), 1.24 (t, J = 7.1 Hz, 3H); 13C NMR (101 MHz, CDCl3)
δ 173.7, 169.0, 150.7 (dd, J = 248.7, 13.0 Hz), 149.0 (dd, J = 247.0, 12.9
Hz), 141.7 (q, J = 1.0 Hz), 129.6 (2C), 129.1 (q, J = 32.4 Hz), 126.2 (dd, J
= 3.3, 2.9 Hz), 125.5 (q, J = 3.8 Hz, 2C), 124.6 – 124.4 (m), 124.3 (q, J =
271.9 Hz), 116.6 (d, J = 17.1 Hz), 60.9, 50.6, 41.1, 36.8 (t, J = 2.4 Hz),
31.1, 28.8, 14.3; ESI-HRMS calcd for C22H22F5NNaO3 (M + Na+) 466.1412,
found 466.1415.
portion. The ice bath was removed, and reaction was stirred at rt. Followed
by TLC until the consumption of the starting material. Subsequently, the
residue was concentrated in vacuo and dried over high vacuum to give
430 mg of residue that was used directly in the next step. MS ES 222.1
[M+H]+.
Step 2: A solution of the amine hydrochloride product from the previous
step (430 mg, 1.7 mmol), 3-chlorophenylacetic acid (386 mg, 2.5 mmol)
and PyBOP (1.47 g, 2.8 mmol) in dry DMF (11 mL) was cooled to 0 °C was
slowly added DIPEA (1.2 mL, 6.7 mmol) and the reaction mixture was
stirred at rt overnight. The reaction mixture was diluted with EtOAc,
washed with 1 M HCl (x3) and brine (x3), dried over Na2SO4, concentrated
in vacuo and purified by semi-automated flash chromatography (EtOAc in
petroleum ether, 0-100%) to give 488 mg (88%) of ethyl (R)-4-(2-(3-
fluorophenyl)acetamido)-5-phenylpentanoate a white solid: Rf = 0.25
(EtOAc/heptane, 1:2); 1H NMR (400 MHz, CDCl3) δ 7.28 – 7.18 (m, 4H),
7.04 – 6.94 (m, 3H), 6.93 – 6.89 (m, 1H), 6.85 (dt, J = 9.6, 2.1 Hz, 1H),
5.37 (d, J = 8.8 Hz, 1H), 4.22 – 4.13 (m, 1H), 4.09 (qd, J = 7.2, 1.9 Hz, 2H),
3.44 (s, 2H), 2.78 – 2.66 (m, 2H), 2.38 – 2.21 (m, 2H), 1.87 – 1.77 (m, 1H),
1.68 – 1.55 (m, 1H), 1.22 (t, J = 7.1 Hz, 3H); 13C NMR (151 MHz, CDCl3)
δ 172.6, 168.8, 161.9 (d, J = 246.6 Hz), 136.2, 136.1 (d, J = 7.3 Hz), 129.4
(d, J = 8.3 Hz), 128.3, 127.44, 125.6, 124.0 (d, J = 2.9 Hz), 115.3 (d, J =
21.5 Hz), 113.3 (d, J = 21.0 Hz), 59.6, 49.2, 42.6 (d, J = 1.9 Hz), 39.9, 30.1,
27.8, 13.2.
Step 2: 40 was prepared as described for 1 using the ethyl ester of 40 (15
mg, 34 μmol) and LiOHaq (0.5 M, 0.17 mL) and obtained as a white solid
(13 mg, 95% yield): 1H NMR (400 MHz, CD3OD) δ 7.51 (d, J = 8.1 Hz, 2H),
7.36 (d, J = 8.0 Hz, 2H), 7.16 – 7.07 (m, 1H), 7.04 – 6.97 (m, 1H), 6.86 (dd,
J = 7.7, 6.3 Hz, 1H), 4.23 – 4.14 (m, 1H), 3.54 – 3.45 (m, 2H), 2.96 (dd, J
= 13.7, 5.3 Hz, 1H), 2.78 (dd, J = 13.7, 9.1 Hz, 1H), 2.43 – 2.29 (m, 2H),
2.00 – 1.89 (m, 1H), 1.81 – 1.69 (m, 1H); 13C NMR (101 MHz, CD3OD) δ
176.9, 171.8, 151.8 (dd, J = 246.1, 13.0 Hz), 150.2 (dd, J = 246.2, 12.9
Hz), 144.4 (q, J = 1.1 Hz), 130.9 (2C), 129.6 (q, J = 32.1 Hz), 127.4 – 127.3
(m), 126.6 – 126.4 (m), 126.1 (q, J = 3.8 Hz, 2C), 125.8 (q, J = 271.0 Hz),
125.2 (dd, J = 7.0, 4.8 Hz), 124.5, 121.8, 117.0 – 116.7 (m), 51.5, 41.8,
36.4 (t, J = 2.5 Hz), 31.7, 31.0; ESI-HRMS calcd for C20H18F5NNaO3 (M +
Step 3: To the ethyl ester from the previous step (200 mg, 0.6 mmol)
dissolved in THF/H2O (3:1, 12 mL) at 0 °C was added LiOH (118 mg, 2.8
mmol). The reaction was stirred at rt overnight, then acidified with 1 M
HClaq, and the aqueous phase was extracted with EtOAc (x3). The
combined organic layer was washed with brine, dried over Na2SO4 and
concentrated in vacuo. The residue was purified by preparative HPLC to
give 150 mg (81%) of 42 as a white solid: 1H NMR (400 MHz, CD3OD) δ
7.28 – 7.07 (m, 6H), 6.97 – 6.85 (m, 3H), 4.16 – 4.05 (m, 1H), 3.41 (s, 2H),
2.82 (dd, J = 13.7, 5.8 Hz, 1H), 2.70 (dd, J = 13.7, 8.3 Hz, 1H), 2.35 – 2.23
(m, 2H), 1.95 – 1.87 (m, 1H), 1.77 – 1.63 (m, 1H); 13C NMR (151 MHz,
CD3OD) δ 176.9, 173.0, 164.2 (d, J = 244.5 Hz), 139.6 (d, J = 6.5 Hz),
139.55, 131.1 (d, J = 8.3 Hz), 130.2 (2C), 129.3 (2C), 127.4, 125.9 (d, J =
2.9 Hz), 116.8 (d, J = 21.6 Hz), 114.5 (d, J = 21.0 Hz), 51.7, 43.5 (d, J =
1.8 Hz), 41.9, 31.7, 30.9. m.p.: 141.8-144.7s °C. MALDI-HRMS calcd for
C19H21FNO3 (M + H+) 330.1500, found 330.1499. HPLC: tR = 9.2 min, 96%.
[α]20D = +9.1° (c = 0.11, MeOH).
Na+) 438.1099, found 438.1117. HPLC: tR = 11.13 min, 99.0%. [α]20
=
D
+17° (c = 0.2, MeOH).
(R)-4-(2-(2,5-Difluorophenyl)acetamido)-5-(4-(trifluoromethyl)-
phenyl)pentanoic acid (41). Step 1: The ethyl ester of 41 was prepared
as described for 15 using (R)-10 (30 mg, 92 μmol), 2-(2,5-
difluorophenyl)acetic acid (18 mg, 105 μmol), HOBt (20 mg, 133 μmol),
DMAP (2 mg, 16 μmol), DIPEA (60 μL, 0.34 mmol) and EDC hydrochloride
(24 mg, 111 μmol) and obtained after flash chromatography (DCM [1%
MeOH] à DCM [2% MeOH]) as a white solid (28 mg, 70% yield): Rf = 0.29
(DCM [2% MeOH]); 1H NMR (400 MHz, CDCl3) δ 7.48 (d, J = 7.9 Hz, 2H),
7.20 (d, J = 7.9 Hz, 2H), 7.03 – 6.89 (m, 3H), 5.55 (d, J = 8.6 Hz, 1H), 4.23
– 4.07 (m, 3H), 3.51 – 3.38 (m, 2H), 2.90 – 2.77 (m, 2H), 2.44 – 2.23 (m,
2H), 1.90 – 1.80 (m, 1H), 1.73 – 1.62 (m, 1H), 1.24 (t, J = 7.1 Hz, 3H); 13
C
NMR (101 MHz, CDCl3) δ 173.7, 169.0, 158.8 (dd, J = 243.4, 2.2 Hz),
156.9 (dd, J = 241.1, 2.6 Hz), 141.8 – 141.7 (m), 129.7 (2C), 129.1 (q, J =
32.4 Hz), 125.5 (q, J = 3.7 Hz, 2C), 124.3 (q, J = 271.9 Hz), 123.7 (dd, J =
18.5, 8.1 Hz), 118.1 (dd, J = 24.3, 4.3 Hz), 116.6 (dd, J = 25.0, 8.8 Hz),
115.7 (dd, J = 24.0, 8.6 Hz), 60.9, 50.6, 41.0, 37.1 – 36.9 (m), 31.1, 28.8,
14.3; ESI-HRMS calcd for C22H22F5NNaO3 (M + Na+) 466.1412, found
466.1403.
Homology modeling
A homology model of hFFA2 was constructed using Modeller 9.17 based
on the crystal structure of hFFA1 (PDB code 5TZY).[34] The receptor was
prepared using Protein Preparation in Maestro (Schrödinger, LLC, version
12.7.161) and docked using the Induced-fit docking protocol with Glide
Extra Precision (XP) and default settings. Ligands 1, 28, 29 and 37 were
prepared by LigPrep using default settings and the OPLS4 forcefield.
Step 2: 41 was prepared as described for 1 using the ethyl ester of 41 (18
mg, 40 μmol) and LiOHaq (0.5 M, 0.20 mL) and obtained as a white solid
(16 mg, 95% yield): 1H NMR (400 MHz, DMSO-d6) δ 11.97 (s, 1H), 7.99
(d, J = 8.7 Hz, 1H), 7.58 (d, J = 8.0 Hz, 2H), 7.38 (d, J = 8.0 Hz, 2H), 7.20
– 7.12 (m, 1H), 7.12 – 7.04 (m, 1H), 7.00 – 6.93 (m, 1H), 4.04 – 3.91 (m,
1H), 3.45 – 3.37 (m, 2H), 2.85 (dd, J = 13.5, 5.2 Hz, 1H), 2.72 (dd, J = 13.5,
8.6 Hz, 1H), 2.35 – 2.15 (m, 2H), 1.81 – 1.71 (m, 1H), 1.64 – 1.53 (m, 1H);
13C NMR (101 MHz, DMSO-d6) δ 174.1, 168.1, 157.7 (dd, J = 239.2, 2.1
Hz), 156.6 (dd, J = 240.3, 2.0 Hz), 143.7 – 143.6 (m), 129.9 (2C), 126.8 (q,
J = 31.2 Hz), 125.2 (dd, J = 18.9, 8.6 Hz), 124.7 (q, J = 3.4 Hz, 2C), 124.4
(q, J = 271.8 Hz), 117.7 (dd, J = 24.4, 4.8 Hz), 116.1 (dd, J = 25.1, 9.0 Hz),
114.6 (dd, J = 24.0, 8.7 Hz), 49.3, 39.2, 35.1, 30.4, 29.5. m.p.: 160.3-
163.5 °C. ESI-HRMS calcd for C20H18F5NNaO3 (M + Na+) 438.1099, found
438.1114. HPLC: tR = 11.13 min, 97.0%. [α]20D = +38° (c = 0.2, MeOH).
In vitro pharmacology
Functional activity of the compounds was tested in
incorporation assay using membranes of Flp-In T-Rex 293 cells induced
to express human FFA2 as described by Hudson et al.[15,26]
a [
35S]GTPgS
Selected compounds were also evaluated in a BRET binding assay with
an NLuc-hFFA2 construct in Flp-In T-Rex 293 cells developed as
described by Christiansen et al.[35] and using the fluorescent antagonist
tracer TUG-1609 as described by Hansen et al.[28]
Microsomal stability
(R)-4-(2-(3-Fluorophenyl)acetamido)-5-phenylpentanoic acid (42)
Step 1: To ethyl (R)-4-((tert-butoxycarbonyl)amino)-5-phenylpentanoate
(535 mg, 1.7 mmol) at 0 °C, 4 M HCl in dioxane (20 ml) was added in one
The study was performed by Bienta (www.bienta.net). Mouse hepatic
microsomes were isolated from pooled, perfused livers of male BALB/c
mice (n = 50). Isolation was performed according to the standard
14
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10.1002/cmdc.202100356
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FULL PAPER
protocol.[36] The batches of microsomes were tested for quality control
using imipramine and propranolol as reference compounds. Microsomal
incubations were carried out in 96-well plates in 5 aliquots of 40 μL each
(one for each time point). Liver microsomal incubation medium comprised
PBS (100 mM, pH 7.4), MgCl2 (3.3 mM), NADPН (3 mM), glucose-6-
phosphate (5.3 mM), glucose-6-phosphate dehydrogenase (0.67
units/mL) with 0.42 mg of liver microsomal protein per ml. In the control
reactions the NADPH-cofactor system was substituted with PBS. 37 (2 μM,
final solvent concentration 1.6%) were incubated with microsomes at 37 °C,
shaking at 100 rpm. Each reaction was performed in duplicates. Five time
points over 40 minutes were analyzed. The reactions were stopped by
adding 12 volumes of 90% acetonitrile-water to incubation aliquots,
followed by protein sedimentation by centrifuging at 5500 rpm for 3
minutes. Supernatants were analyzed using HPLC-MS/MS on an API
3000 PE instrument.
[3]
a) I. Kimura, A. Ichimura, R. Ohue-Kitano, M. Igarashi, Physiol. Rev.
2020, 100, 171-210; b) G. Milligan, B. Shimpukade, T. Ulven, B. D.
Hudson, Chem. Rev. 2017, 117, 67-110.
[4]
[5]
J. Ghislain, V. Poitout, Nat. Rev. Endocrinol. 2021.
D. Mikami, M. Kobayashi, J. Uwada, T. Yazawa, K. Kamiyama, K.
Nishimori, Y. Nishikawa, S. Nishikawa, S. Yokoi, H. Kimura, I. Kimura,
T. Taniguchi, M. Iwano, Biochim. Biophys. Acta. Mol. Cell Biol. Lipids
2020, 1865, 158666.
M. Pizzonero, S. Dupont, M. Babel, S. Beaumont, N. Bienvenu, R.
Blanque, L. Cherel, T. Christophe, B. Crescenzi, E. De Lemos, P.
Delerive, P. Deprez, S. De Vos, F. Djata, S. Fletcher, S. Kopiejewski,
C. L'Ebraly, J. M. Lefrancois, S. Lavazais, M. Manioc, L. Nelles, L.
Oste, D. Polancec, V. Quenehen, F. Soulas, N. Triballeau, E. M. van
der Aar, N. Vandeghinste, E. Wakselman, R. Brys, L. Saniere, J. Med.
Chem. 2014, 57, 10044-10057.
[6]
[7]
[8]
J. H. Lee, D. S. Im, Biomol. Ther. 2021,
doi:10.4062/biomolther.2020.4176.
a) G. Wang, L. Jiang, J. Wang, J. Zhang, F. Kong, Q. Li, Y. Yan, S.
Huang, Y. Zhao, L. Liang, J. Li, N. Sun, Y. Hu, W. Shi, G. Deng, P.
Chen, L. Liu, X. Zeng, G. Tian, Z. Bu, H. Chen, C. Li, J. Virol. 2020, 94;
b) V. Sencio, A. Barthelemy, L. P. Tavares, M. G. Machado, D.
Soulard, C. Cuinat, C. M. Queiroz-Junior, M.-L. Noordine, S. Salomé-
Desnoulez, L. Deryuter, B. Foligné, C. Wahl, B. Frisch, A. T. Vieira, C.
Paget, G. Milligan, T. Ulven, I. Wolowczuk, C. Faveeuw, R. Le Goffic,
M. Thomas, S. Ferreira, M. M. Teixeira, F. Trottein, Cell reports 2020,
30, 2934-2947.e2936; c) I. Galvao, L. P. Tavares, R. O. Correa, J. L.
Fachi, V. M. Rocha, M. Rungue, C. C. Garcia, G. Cassali, C. M.
Ferreira, F. S. Martins, S. C. Oliveira, C. R. Mackay, M. M. Teixeira, M.
A. R. Vinolo, A. T. Vieira, Front. Immunol. 2018, 9, 142.
C. Tang, K. Ahmed, A. Gille, S. Lu, H. J. Grone, S. Tunaru, S.
Offermanns, Nat. Med. 2015, 21, 173-177.
In vivo pharmacokinetics
The study was performed by Bienta (www.bienta.net). Study design,
animal selection, handling and treatment were all in accordance with the
Enamine PK study protocols and Institutional Animal Care and Use
Guidelines. Animal treatment and plasma samples preparation were
conducted by the Animal Laboratory personnel at Enamine/Bienta. Male
BALB/c mice (9-11 weeks old, body weight: 22.1 to 29.6 g and average
body weight across all groups 24.9 g, SD = 1.51 g) were used in this study.
The animals were randomly assigned to the treatment groups before the
pharmacokinetic study; all animals were fasted for 4 h before dosing. Nine
time points (5, 15, 30, 60, 120, 240, 360, 480, and 1440 min) were set for
this pharmacokinetic study. Each of the time point treatment group
included 4 animals. There was also a control group of 2 animals.
Compound 37 in vehicle (DMSO – Cremophor EL – water w/5% mannitol,
1:1:8) was dosed at 10 mg/kg po and 5 mg/kg iv. Dosing volumes of
compounds or vehicle was 5 mL/kg. Mice were injected iv with 2,2,2-
tribromoethanol at the dose of 150 mg/kg prior to drawing the blood. Blood
collection was performed from the orbital sinus in microtainers containing
K2EDTA. Animals were sacrificed by cervical dislocation after the blood
samples collection. All samples were immediately processed, flash-frozen
and stored at -70°C until subsequent analysis. Before analysis, plasma
samples (50 μL) were added acetonitrile-methanol 1:1, v/v, 200 μL)
containing TUG-816 (cmp 1 in ref)[37] as internal standard (400 ng/mL).
After mixing by pipetting and centrifuging for 4 min at 6,000 rpm, 0.5 μL of
each supernatant was analyzed by HPLC-MS/MS on an API 3000 PE
instrument.
[9]
[10] B.-O. Park, S. H. Kim, G. Y. Kong, D. H. Kim, M. S. Kwon, S. U. Lee,
M.-O. Kim, S. Cho, S. Lee, H.-J. Lee, S.-B. Han, Y. S. Kwak, S. B. Lee,
S. Kim, Eur. J. Pharmacol. 2016, 771, 1-9.
[11] a) S. Forbes, S. Stafford, G. Coope, H. Heffron, K. Real, R. Newman,
R. Davenport, M. Barnes, J. Grosse, H. Cox, Diabetes 2015, 64, 3763-
3771; b) D. Bolognini, N. Barki, A. J. Butcher, B. D. Hudson, E.
Sergeev, C. Molloy, C. E. Moss, S. J. Bradley, C. Le Gouill, M. Bouvier,
A. B. Tobin, G. Milligan, Nat. Chem. Biol. 2019, 15, 489-498.
[12] S. Vermeire, V. Kojecký, V. Knoflícek, W. Reinisch, T. Van Kaem, F.
Namour, J. Beetens, F. Vanhoutte, J. Crohn's Colitis 2015, Suppl 1,
S39.
[13] a) Y. Akiba, K. Maruta, K. Narimatsu, H. Said, I. Kaji, A. Kuri, K. I.
Iwamoto, A. Kuwahara, J. D. Kaunitz, Am. J. Physiol. Gastrointest.
Liver Physiol. 2017, 313, G117-g128; b) Y. Lu, C. Fan, P. Li, Y. Lu, X.
Chang, K. Qi, Sci. Rep. 2016, 6, 37589.
[14] C. E. Brantis, F. Ooms, J. Bernard. Novel amino acid derivatives and
their use as GPR43 receptor modulators. WO2011092284A1.
[15] B. D. Hudson, I. G. Tikhonova, S. K. Pandey, T. Ulven, G. Milligan, J.
Biol. Chem. 2012, 287, 41195-41209.
[16] a) E. Sergeev, A. H. Hansen, D. Bolognini, K. Kawakami, T. Kishi, J.
Aoki, T. Ulven, A. Inoue, B. D. Hudson, G. Milligan, Sci. Rep. 2017, 7,
13741; b) E. Sergeev, A. H. Hansen, S. K. Pandey, A. E. MacKenzie,
B. D. Hudson, T. Ulven, G. Milligan, J. Biol. Chem. 2016, 291, 303-317.
[17] C. E. Augelli-Szafran, D. C. Horwell, C. Kneen, D. F. Ortwine, M. C.
Pritchard, T. S. Purchase, B. D. Roth, B. K. Trivedi, D. Hill, N. Suman-
Chauhan, L. Webdale, Bioorg. Med. Chem. 1996, 4, 1733-1745.
[18] M. R. Linder, S. Steurer, J. Podlech, Org. Synth. 2002, 79, 154.
[19] N. Ikemoto, D. M. Tellers, S. D. Dreher, J. Liu, A. Huang, N. R. Rivera,
E. Njolito, Y. Hsiao, J. C. McWilliams, J. M. Williams, J. D. Armstrong,
3rd, Y. Sun, D. J. Mathre, E. J. Grabowski, R. D. Tillyer, J. Am. Chem.
Soc. 2004, 126, 3048-3049.
Acknowledgements
This work was supported by the Danish Council for Strategic
Research (grant 0603-00452B) and the Lundbeck Foundation
(R181-2014-3247 and R307-2018-2950).
[20] K. B. Hansen, J. Balsells, S. Dreher, Y. Hsiao, M. Kubryk, M. Palucki,
N. Rivera, D. Steinhuebel, J. D. Armstrong, D. Askin, E. J. J.
Grabowski, Org. Process Res. Dev. 2005, 9, 634-639.
[21] M. T. Robak, M. A. Herbage, J. A. Ellman, Chem. Rev. 2010, 110,
3600-3740.
Keywords: FFA2 • GPR43 • GPCR • short-chain fatty acids •
[22] a) H. K. Dema, F. Foubelo, M. Yus, Helv. Chim. Acta 2012, 95, 1790-
1798; b) S. K. Kang, G. H. Cho, H. J. Leem, B. K. Soh, J. Sim, Y.-G.
Suh, Tetrahedron Asymmetry 2017, 28, 34-40.
inflammation
[23] C.-K. Lin, T.-J. Lu, Tetrahedron 2010, 66, 9688-9693.
[24] D. Hernández, K. B. Lindsay, L. Nielsen, T. Mittag, K. Bjerglund, S.
Friis, R. Mose, T. Skrydstrup, J Org Chem 2010, 75, 3283-3293.
[25] a) J. W. Choy, C. Bryant, C. M. Calvet, P. S. Doyle, S. S. Gunatilleke,
S. S. Leung, K. K. Ang, S. Chen, J. Gut, J. A. Oses-Prieto, J. B.
Johnston, M. R. Arkin, A. L. Burlingame, J. Taunton, M. P. Jacobson, J.
M. McKerrow, L. M. Podust, A. R. Renslo, Beilstein J. Org. Chem.
2013, 9, 15-25; b) M. Sinha, V. R. Dola, P. Agarwal, K. Srivastava, W.
Haq, S. K. Puri, S. B. Katti, Bioorg. Med. Chem. 2014, 22, 3573-3586.
[26] B. D. Hudson, M. E. Due-Hansen, E. Christiansen, A. M. Hansen, A. E.
Mackenzie, H. Murdoch, S. K. Pandey, R. J. Ward, R. Marquez, I. G.
Tikhonova, T. Ulven, G. Milligan, J. Biol. Chem. 2013, 288, 17296-
17312.
[1]
a) A. J. Brown, S. M. Goldsworthy, A. A. Barnes, M. M. Eilert, L.
Tcheang, D. Daniels, A. I. Muir, M. J. Wigglesworth, I. Kinghorn, N. J.
Fraser, N. B. Pike, J. C. Strum, K. M. Steplewski, P. R. Murdock, J. C.
Holder, F. H. Marshall, P. G. Szekeres, S. Wilson, D. M. Ignar, S. M.
Foord, A. Wise, S. J. Dowell, J. Biol. Chem. 2003, 278, 11312-11319;
b) E. Le Poul, C. Loison, S. Struyf, J. Y. Springael, V. Lannoy, M. E.
Decobecq, S. Brezillon, V. Dupriez, G. Vassart, J. Van Damme, M.
Parmentier, M. Detheux, J. Biol. Chem. 2003, 278, 25481-25489; c) N.
E. Nilsson, K. Kotarsky, C. Owman, B. Olde, Biochem. Biophys. Res.
Commun. 2003, 303, 1047-1052.
a) J. K. Tan, C. McKenzie, E. Marino, L. Macia, C. R. Mackay, Annu.
Rev. Immunol. 2017, 35, 371-402; b) T. Ulven, Front. Endocrinol. 2012,
3, 111; c) K. M. Maslowski, A. T. Vieira, A. Ng, J. Kranich, F. Sierro, D.
Yu, H. C. Schilter, M. S. Rolph, F. Mackay, D. Artis, R. J. Xavier, M. M.
Teixeira, C. R. Mackay, Nature 2009, 461, 1282-1287.
[2]
[27] M. E. Due-Hansen, S. K. Pandey, E. Christiansen, R. Andersen, S. V.
Hansen, T. Ulven, Org. Biomol. Chem. 2016, 14, 430-433.
15
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10.1002/cmdc.202100356
ChemMedChem
FULL PAPER
[28] A. H. Hansen, E. Sergeev, S. K. Pandey, B. D. Hudson, E.
Christiansen, G. Milligan, T. Ulven, J. Med. Chem. 2017, 60, 5638-
5645.
[29] P. D. Leeson, B. Springthorpe, Nat. Rev. Drug Discov. 2007, 6, 881-
890.
[30] E. Christiansen, M. E. Due-Hansen, C. Urban, M. Grundmann, J.
Schmidt, S. V. F. Hansen, B. D. Hudson, M. Zaibi, S. B. Markussen, E.
Hagesaether, G. Milligan, M. A. Cawthorne, E. Kostenis, M. U.
Kassack, T. Ulven, J. Med. Chem. 2013, 56, 982−992.
[31] M. Szostak, M. Spain, A. J. Eberhart, D. J. Procter, J. Am. Chem. Soc.
2014, 136, 2268-2271.
[34] J. Lu, N. Byrne, J. Wang, G. Bricogne, F. K. Brown, H. R. Chobanian,
S. L. Colletti, J. Di Salvo, B. Thomas-Fowlkes, Y. Guo, D. L. Hall, J.
Hadix, N. B. Hastings, J. D. Hermes, T. Ho, A. D. Howard, H. Josien,
M. Kornienko, K. J. Lumb, M. W. Miller, S. B. Patel, B. Pio, C. W.
Plummer, B. S. Sherborne, P. Sheth, S. Souza, S. Tummala, C.
Vonrhein, M. Webb, S. J. Allen, J. M. Johnston, A. B. Weinglass, S.
Sharma, S. M. Soisson, Nat. Struct. Mol. Biol. 2017, 24, 570-577.
[35] E. Christiansen, B. D. Hudson, A. H. Hansen, G. Milligan, T. Ulven, J.
Med. Chem. 2016, 59, 4849-4858.
[36] J. R. Hill, Curr. Protoc. Pharmacol. 2003, 23, 7.8.1-7.8.11.
[37] E. Rexen Ulven, T. Quon, E. Sergeev, N. Barki, M. Brvar, B. D.
Hudson, P. Dutta, A. H. Hansen, L. O. Bielefeldt, A. B. Tobin, C. J.
McKenzie, G. Milligan, T. Ulven, J. Med. Chem. 2020, 63, 3577-3595.
[32] Y. Combret, J. Duflos, G. Dupas, J. Bourguignon, G. Quéguiner,
Tetrahedron Asymmetry 1993, 4, 1635-1644.
[33] A. J. Ross, F. Dreiocker, M. Schäfer, J. Oomens, A. J. Meijer, B. T.
Pickup, R. F. Jackson, J. Org. Chem. 2011, 76, 1727-1734.
16
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Entry for the Table of Contents
The short-chain fatty acid receptor FFA2 is a promising drug target for metabolic and inflammatory diseases. We here present the
SAR exploration of an antagonist series, leading to the discovery of a more potent antagonist with favorable solubility and
pharmacokinetic properties.
17
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