Journal of Medicinal Chemistry
Article
H, q), 6.65 (1 H, d), 7.49−7.75 (5 H, m). HRMS (TOF) for
C13H15O2Br (M+): calcd, 282.0255; found, 282.0272.
dichloride DCM complex (45.7 mg, 0.06 mmol) were suspended in
degassed 1,4-dioxane (3 mL). The resulting suspension was evacuated
and purged with nitrogen 5 times and then heated to 90 °C for 5 h. The
reaction mixture was filtered through Celite, washing through with
DCM, then THF. The filtrate was concentrated to provide the title
compound (625 mg, 100%), which was used without further
trans-(1S,2S)/(1R,2R)-Ethyl 2-(4-Bromophenyl)cyclopropane-
carboxylate (38, R1 = H, Et Ester). Trimethylsulfoxonium iodide
(56.1 g, 255 mmol) was added in one portion to potassium tert-butoxide
(27.8 g, 235 mmol) in DMSO (405 mL) at rt under nitrogen. The
resulting suspension was heated to 50 °C and stirred for 1 h. The pale
yellow solution was cooled to rt, and a solution of 37 (R1 = H, Et ester)
(50 g, 196 mmol) in DMSO (135 mL) was added dropwise over 1 h.
The reaction was stirred at rt overnight. The reaction mixture was
diluted with diethyl ether and washed with saturated aqueous NH4Cl.
The organic layer was dried over MgSO4, filtered, and evaporated to
afford the title compound (36.8 g, 70%) as a yellow oil which was used
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purification. H NMR (400 MHz, DMSO-d6) δ 1.15 (6H, s), 1.22−
1.31 (6H, m), 1.61−1.71 (1H, m), 1.72−1.78 (1H, m), 2.11−2.22 (1H,
m), 3.59 (1H, t), 7.06 (2H, d), 7.52 (1H, d); m/z (M − H)− = 287.
6-Bromo-3-methyl-1-p-tolyl-1H-indazole (48, R2 = H, R4 =
Me). Copper(II) acetate (2.58 g, 14.2 mmol) was added to a stirred
mixture of 6-bromo-3-methyl-1H-indazole (2.00 g, 9.48 mmol), p-
tolylboronic acid (1.93 g, 14.2 mmol), pyridine (2.3 mL, 28.4 mmol),
and DCM (40 mL). The mixture was stirred at rt for 4 h under air. 35%
ammonia solution (25 mL) was added, and the mixture was stirred for
15 min. The organic layer was separated and concentrated in vacuo. The
crude product was purified by flash silica chromatography, with elution
gradient 0−5% EtOAc in heptane. Pure fractions were evaporated to
dryness and solid was washed with a small amount of diethyl ether to
afford the title compound (1.32 g, 46%) as a solid. 1H NMR (400 MHz,
DMSO-d6) δ 2.38 (3H, s), 2.57 (3H, s), 7.33−7.4 (3H, m), 7.57−7.62
(2H, m), 7.80 (1H, d), 7.89 (1H, d); m/z (M + H)+ = 301.
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without further purification. H NMR (400 MHz, DMSO-d6) δ 1.19
(3H, t), 1.36 (1H, ddd), 1.41−1.5 (1H, m), 1.93 (1H, ddd), 2.42 (1H,
ddd), 4.05−4.13 (2H, m), 7.11−7.17 (2H, m), 7.41−7.47 (2H, m).
trans-(1S,2S)/(1R,2R)-2-(4-Bromophenyl)cyclopropane-
carboxylic Acid (39, R1 = H). Lithium hydroxide hydrate (9.99 g, 238
mmol) was added to 38 (R1 = H, Et ester) (21.4 g, 79.4 mmol) in THF
(225 mL) and water (75 mL). The resulting mixture was stirred at rt
overnight. The reaction mixture was diluted with water (125 mL) and
washed with diethyl ether (125 mL). The organic phase was extracted
with water (60 mL). The combined aqueous phases were acidified with 2
M HCl (aq). The resulting suspension was extracted with DCM (600
mL). The organic phase was filtered through a phase separation
cartridge and evaporated to afford the title compound (17.8 g, 93%) as a
(1S,2S)-2-(4-(3-Methyl-1-p-tolyl-1H-indazol-6-yl)phenyl)-
cyclopropanecarboxylic Acid (34). 1,1′-Bis(di-tert-butyl-
phosphino)ferrocenepalladium(II) dichloride (19 mg, 0.03 mmol)
and tripotassium phosphate (251 mg, 1.2 mmol) were added to 42 (R1
= H) (138 mg, 0.48 mmol) and 48 (R2 = H, R4 = Me) (120 mg, 0.40
mmol) in degassed MeCN (1 mL) and water (1 mL) at 20 °C under
nitrogen. The resulting mixture was further degassed and stirred within a
microwave reactor at 100 °C for 1 h. The reaction mixture was acidified
with acetic acid (0.25 mL) and extracted into DCM/MeOH (9:1
mixture). The organic phase was passed through a PL-thiol MP SPE 500
mg/6 mL cartridge, and the solid was washed with a 9:1 mixture of
DCM/MeOH. The filtrate was concentrated. The crude product was
purified by preparative HPLC (Waters XBridge Prep C18 OBD column,
5 μm silica, 50 mm diameter, 150 mm length), using decreasingly polar
mixtures of water (containing 0.5% ammonia) and MeCN as eluents.
Fractions containing the desired compound were evaporated to dryness
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cream solid. H NMR (400 MHz, DMSO-d6) δ 1.32 (1H, ddd), 1.42
(1H, ddd), 1.77−1.84 (1H, m), 2.38 (1H, ddd), 7.13 (2H, d), 7.44 (2H,
d), 12.30 (1H, s); m/z (M − H)− = 241.
(S)-3-((1S,2S)-2-(4-Bromophenyl)cyclopropanecarbonyl)-4-
isopropyloxazolidin-2-one (40, R1 = H). Oxalyl dichloride (6.67
mL, 77.7 mmol) was added dropwise to 39 (R1 = H) (17.8 g, 74.0
mmol) and DMF (0.03 mL, 0.37 mmol) in DCM (150 mL) under
nitrogen. The resulting mixture was stirred at rt for 1.5 h. The reaction
mixture was evaporated to dryness and azeotroped with toluene. The
resulting crude acid chloride was dissolved in DCM (150 mL).
Triethylamine (20.6 mL, 148 mmol) was added followed by portionwise
addition of (S)-4-isopropyloxazolidin-2-one (9.56 g, 74.0 mmol). The
mixture was stirred at rt under nitrogen for 64 h. The reaction mixture
was diluted with DCM and washed sequentially with water and saturated
brine. The organic layer was filtered through a phase separation cartridge
and evaporated. The crude product was purified by flash silica
chromatography,with elution gradient 20% heptane in DCM. Pure
fractions were evaporated to dryness to afford the title compound as a
white solid after trituration with ether/heptane (7.13 g, 27%). 1H NMR
(400 MHz, DMSO-d6) δ 0.79 (3H, d), 0.84 (3H, d), 1.49 (1H, ddd),
1.59 (1H, ddd), 2.16 (1H, dp), 3.38 (1H, ddd), 4.25−4.35 (2H, m), 4.39
(1H, dt), 7.12−7.18 (2H, m), 7.43−7.48 (2H, m).
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to afford the title compound (106 mg, 70%) as solid. H NMR (400
MHz, DMSO-d6) δ 1.38 (1H, ddd), 1.46 (1H, dt), 1.82−1.88 (1H, m),
2.38 (3H, s), 2.41−2.46 (1H, m), 2.59 (3H, s), 7.27 (2H, d), 7.37 (2H,
d), 7.49 (1H, dd), 7.66 (4H, dd), 7.83 (1H, s), 7.88 (1H, d), 12.28 (1H,
s); 13C NMR (126 MHz, DMSO-d6) 12.1, 17.2, 21.0, 24.8, 25.5, 108.1,
120.9, 121.6, 122.3, 124.0, 127.0, 127.8, 130.5, 135.8, 137.8, 138.8, 139.8,
139.9, 140.3, 143.6, 174.3; m/z (M + H)+ = 383. HRMS (ES+) for
C25H23N2O2 (MH+): calcd 383.1760; found, 383.1742. αD24.5 +204.8 (c
0.62, EtOH).
Biological Protocols. Human and Mouse GPR120 Dynamic
Mass Redistribution (DMR) Assays. CHO FlpIn cryopreserved cells
stably expressing human or mouse GPR120 were seeded in 30 μL of
growth medium (DMEM/F-12 glutaMAX, 10% FCS, and 300 μg/mL
hygromycin) into 384-well Corning EPIC fibronectin coated plates 48 h
prior to assay and incubated at 37 °C in a CO2 incubator. The growth
medium was changed, 24 h prior to assay, to 30 μL of medium
containing no serum, and the plates were returned to the CO2 incubator.
The medium was again changed, 2 h prior to assay, to 30 μL of assay
buffer containing 0.5% DMSO (HBSS, 20 mM HEPES, 0.01% BSA at
pH 7.4), and the plates were returned to the CO2 incubator. Plates were
equilibrated in the EPIC instrument and multiple baseline measure-
ments were taken over a period of 10 min prior to adding 15 μL of test or
control substance to the wells. Following addition, further measure-
ments were taken over a period of 15 min to establish the agonist effect
of test substance on the DMR of the cells.
(1S,2S)-2-(4-Bromophenyl)cyclopropanecarboxylic Acid (41,
R1 = H). Hydrogen peroxide (50 wt %, 2.03 mL, 35.2 mmol) was added
dropwise to a solution of lithium hydroxide hydrate (0.591 g, 14.1
mmol) in water (8 mL) at rt. The resulting solution was stirred for 30
min and then cooled to 0 °C. This cooled solution was added
portionwise to a cooled solution of 40 (R1 = H) (3.1 g, 8.80 mmol) in
THF (40 mL) and water (10 mL). The internal temperature remained
below 10 °C throughout the addition. The resulting solution was stirred
for 3.5 h. Sodium sulfite (4.44 g, 35.2 mmol) in water (20 mL) was
added. The mixture was extracted with EtOAc (2 × 50 mL). The
aqueous phase was then acidified with 2 M HCl. The aqueous phase was
extracted with EtOAc (3 × 60 mL) and the combined organic layers
were dried over MgSO4, filtered, and evaporated to afford crude product.
This was recrystallized from diethyl ether/heptane to give the title
compound (1.39 g, 66%). 1H NMR (400 MHz, DMSO-d6) δ 1.31 (1H,
ddd), 1.42 (1H, ddd), 1.80 (1H, ddd), 2.38 (1H, ddd), 7.1−7.15 (2H,
m), 7.41−7.46 (2H, m), 12.29 (1H, s); m/z (M − H)− = 241.
Test compounds were administered to the plates in duplicate in a 10-
point concentration−response format with final concentrations from 16
μM to 0.8 nM. Two control agonist compounds were assessed in each
assay run to determine interassay variability. Each plate contained 32
maximum and minimum controls. A maximal concentration of a control
agonist compound was used to define the assay maximum. The
minimum control wells received 15 μL of assay buffer and DMSO
(1S,2S)-2-(4-(4,4,5,5-Tetramethyl-1,3,2-dioxaborolan-2-yl)-
phenyl)cyclopropanecarboxylic Acid (42, R1 = H). Potassium
acetate (330 mg, 3.36 mmol), 41, R1 = H (270 mg, 1.12 mmol),
4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bi(1,3,2-dioxaborolane) (313 mg,
1.23 mmol), and 1,1′-bis(diphenylphosphino)ferrocenepalladium(II)
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J. Med. Chem. XXXX, XXX, XXX−XXX