Journal of Medicinal Chemistry
Article
temperature, the mixture was adjusted to a pH of 5 and extracted with
ethyl acetate, washed with brine, and the organic phase was dried over
Na2SO4. The solvent was removed and the residue was purified by
flash column chromatography on silica (80% PE in ethyl acetate
∼50% PE in ethyl acetate) to give a white solid 47d (0.35 g, 49%). 1H
NMR (500 MHz, DMSO-d6): δ 6.87−6.85 (m, 1H), 6.84 (s, 1H),
6.82 (d, J = 2.8 Hz, 1H), 3.68 (s, 3H), 3.45 (m, 2H), 3.44 (m, 4H),
2.94 (m, 4H). 13C NMR (126 MHz, DMSO): δ 173.04, 154.32,
152.01, 145.32, 124.49, 121.12, 116.45, 111.74, 79.40, 56.13, 50.34,
36.23, 28.53. MS (ESI): m/z 351.2 (M + H)+.
Step 2: tert-Butyl 4-(3-(2-amino-2-oxoethyl)-4-methoxyphenyl)-
piperazine-1-carboxylate (48d). CDI (0.29 g, 1.29 mmol) was added
to a solution of 47d (0.30 g, 0.86 mmol) in 2 mL of DMF. After
stirring for 0.5 h at room temperature, NH3 (0.6 mL 7 N in methanol
solution) was added and allowed to stir for another 1 h at rt. The
solvent was evaporated, and ethyl acetate (2 × 120 mL) was used to
extract the product. The combined organic layer was dried over
Na2SO4 and evaporated to give a white residue that was purified by
flash column chromatography on silica (80% PE in ethyl acetate
∼50% PE in ethyl acetate) to give a white solid 48d (0.22 g 70%). 1H
NMR (500 MHz, DMSO-d6): δ 7.17 (s, 1H), 6.92−6.69 (m, 4H),
3.69 (s, 3H), 3.69−3.43 (m, 4H), 3.31 (s, 2H), 2.95−2.93 (m, 4H),
1.42 (s, 9H). 13C NMR (126 MHz, DMSO): δ 172.60, 154.31,
151.93, 145.33, 125.50, 120.96, 116.16, 111.76, 79.39, 56.15, 50.40,
37.30, 28.53. MS (ESI): m/z 350.2 (M + H)+.
45.27, 41.73. HRMS (ESI): m/z calcd for C26H25N4O4 [M + H]+,
457.1876; found, 457.1794. Purity: 97.0%.
Compounds 25−30 were prepared following a similar synthetic
procedure of compound 28. Detailed spectral data are present in the
Scientific (Beijing, China).
Synthesis of 3-((1-Acryloylpiperidin-4-yl)amino)-4-(1H-
indol-3-yl)-1H-pyrrole-2,5-dione (34). Step 1: 3-Bromo-4-(1H-
indol-3-yl)-1H-pyrrole-2,5-dione (52). In a two-necked flask
equipped with a dropping funnel under argon, indole (0.30 g, 1.18
mmol) was dissolved in anhydrous THF (8 mL). A solution of 3 M
ethyl magnesium bromide in Et2O (1.57 mL, 4.71 mmol) was added
dropwise into the mixture, which was then heated to reflux for 2 h.
After cooling, a solution of 51 (0.55 g, 4.71 mmol) in anhydrous THF
(6 mL) was added dropwise over 1 h. Then, the reaction mixture was
stirred at rt for 1 h. The mixture was then hydrolyzed to pH = 9 with
aqueous HCl solution. After adding saturated aqueous NH4Cl
solution, the aqueous phase was extracted with ethyl acetate. The
combined organic layers were then washed with saturated aqueous
NaCl, dried over Na2SO4, and evaporated under vacuum. The residue
was purified by flash column chromatography on silica to yield 52
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(0.28 g, 82%), a yellow solid. H NMR (400 MHz, DMSO-d6): δ
12.10 (s, 1H), 11.35 (s, 1H), 8.03 (d, J = 2.9 Hz, 1H), 7.89 (dt, J =
8.1, 1.0 Hz, 1H), 7.51 (dt, J = 8.1, 1.0 Hz, 1H), 7.22 (ddd, J = 8.1, 7.0,
1.2 Hz, 1H), 7.14 (ddd, J = 8.1, 7.1, 1.2 Hz, 1H). 13C NMR (101
MHz, DMSO): δ 170.75, 167.99, 138.54, 137.01, 131.54, 125.05,
122.95, 122.77, 120.92, 115.13, 112.84, 104.25. MS (ESI): m/z 291.0
(M + H)+.
Step 3: tert-Butyl-4-(3-(4-(1H-indol-3-yl)-2,5-dioxo-2,5-dihydro-
1H-pyrrol-3-yl)-4-methoxyphenyl)piperazine-1-carboxylate (49d).
t
At 0 °C, BuOK (2.2 mL, 1 M in THF) was added to a solution of
48d (0.20 g, 0.57 mmol) and 39 (0.17 g, 0.85 mmol) in THF (4 mL).
The solution was stirred for 2 h at 10 °C, HCl (5 N) was added to
adjust pH to 5, the solvent was removed, the mixture was extracted
with ethyl acetate, and the organic phase was dried with Na2SO4. The
solvent was removed, and the residue was used in the next step
without further purification. MS (ESI): m/z 503.2 (M + H)+.
Step 4: 3-(1H-Indol-3-yl)-4-(2-methoxy-5-(piperazin-1-yl)-
phenyl)-1H-pyrrole-2,5-dion (50d). TFA (2 mL) was added to a
solution of 49d (0.12 g, 0.24 mmol) in DCM (2 mL) and the mixture
was stirred at rt for 15 min. TFA and the solvent were evaporated, the
mixture was extracted with ethyl acetate, washed by NaHCO3 and
brine, and the organic phase was dried with Na2SO4. The solvent was
removed, and the residue was purified by flash column chromatog-
raphy on silica (2% methanol in DCM ∼10% methanol in DCM) to
Step 2: tert-Butyl-4-((4-(1H-indol-3-yl)-2,5-dioxo-2,5-dihydro-1H-
pyrrol-3-yl)amino)piperidine-1-carboxylate (53). 52 (0.13 g, 0.45
mmol) and piperidin-4-amine (0.18 g, 0.89 mmol) were dissolved in
DMSO (1.5 mL), followed by the addition of DIEA (0.15 mL, 0.89
mmol). The mixture was heated at 126 °C overnight. After cooling,
the mixture was diluted with water and extracted with ethyl acetate.
The combined organic layers were then washed with brine, dried over
Na2SO4, and evaporated under vacuum. The residue was purified by
flash column chromatography on silica to yield 53 (0.11 g, 60%), a
yellow solid. 1H NMR (400 MHz, DMSO-d6): δ 11.21 (d, J = 2.5 Hz,
1H), 10.34 (s, 1H), 7.40 (d, J = 8.1 Hz, 1H), 7.37−7.27 (m, 2H),
7.14−7.06 (m, 1H), 7.04−6.95 (m, 1H), 6.86 (d, J = 9.0 Hz, 1H),
3.67 (m, 2H), 3.43 (m, 1H), 1.97 (m, 2H), 1.47 (m, 2H), 1.31 (s,
9H), 1.26 (m, 2H). 13C NMR (101 MHz, DMSO). 13C NMR (101
MHz, DMSO): δ 173.82, 169.57, 154.19, 143.23, 136.10, 128.71,
126.48, 121.70, 119.91, 119.37, 112.05, 104.59, 100.05, 93.48, 79.22,
50.31, 32.01, 28.52. MS (ESI): m/z 410.1 (M + H)+.
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give a yellow solid 50d (0.15 g, 52% for two steps). H NMR (400
MHz, DMSO-d6): δ 11.80 (s, 1H), 7.91 (s, 1H), 7.37 (d, J = 8.1 Hz,
1H), 7.02 (ddd, J = 8.2, 7.0, 1.1 Hz, 1H), 6.95 (dd, J = 9.0, 3.0 Hz,
1H), 6.88 (d, J = 9.1 Hz, 1H), 6.78 (d, J = 2.9 Hz, 1H), 6.65 (ddd, J =
8.2, 7.0, 1.1 Hz, 1H), 6.51−6.41 (m, 1H), 3.26 (s, 3H), 2.89−2.81
(m, 4H), 2.80−2.78 (m, 4H). 13C NMR (101 MHz, DMSO): δ
173.08, 172.67, 152.06, 145.93, 136.87, 134.69, 130.87, 128.29,
125.38, 122.39, 121.29, 121.12, 120.23, 119.96, 118.58, 113.08,
112.38, 106.29, 55.97, 50.91, 45.78. MS (ESI): m/z 403.2 (M + H)+.
Step 5: 3-(5-(4-Acryloylpiperazin-1-yl)-2-methoxyphenyl)-4-(1H-
indol-3-yl)-1H-pyrrole-2,5-dione (28). 50d (0.10 g, 0.24 mmol) was
dissolved in a mixture of THF (2 mL) and water (1 drop), followed
by adding DIEA (0.16 mL, 0.96 mmol) and acryloyl chloride (0.03
mL, 0.36 mmol). The resulting solution was stirred at rt for 10 min.
After removing the solvent, the mixture was extracted with ethyl
acetate, washed with brine, and the organic phase was dried over
Na2SO4. The solvent was removed, and the residue was purified by
flash column chromatography on silica (1% methanol in DCM ∼1.5%
methanol in DCM) to give a yellow solid 28 (0.09 g, 82% for two
Step 3: 3-((1-Acryloylpiperidin-4-yl)amino)-4-(1H-indol-3-yl)-1H-
pyrrole-2,5-dione (34). TFA (2 mL) was added to a solution of 53
(0.10 g, 0.24 mmol) in DCM (2 mL) and the mixture was stirred at rt
for 15 min. TFA and the solvent were evaporated, and the residue was
used in the next step without further purification. The residue (0.24
mmol) was dissolved in a mixture of THF (2 mL) and water (1 drop),
followed by the addition of DIEA (0.16 mL, 0.96 mmol) and acryloyl
chloride (0.03 mL, 0.36 mmol). The resulting solution was stirred at
rt for 10 min. After removing the solvent, the mixture was extracted
with ethyl acetate and washed with brine. The organic phase was
dried over Na2SO4. The solvent was removed, and the residue was
purified by flash column chromatography on silica (1% methanol in
DCM ∼1.5% methanol in DCM) to yield a yellow solid 34 (0.08 g,
1
90% for two steps). H NMR (400 MHz, DMSO-d6): δ 11.23 (s, 1
H), 10.43 (s, 1 H), 7.54−7.20 (m, 3 H), 7.09 (d, J = 7.1 Hz, 1 H),
7.01 (t, J = 7.0 Hz, 1 H), 6.64 (d, J = 7.5 Hz, 1 H), 5.75 (t, J = 14.6
Hz, 1 H), 5.27 (d, J = 7.8 Hz, 1 H), 3.87 (d, J = 12.2 Hz, 1 H), 2.93
(m, 1 H), 2.82 (m, 1 H), 2.56 (m, 1 H), 1.75 (m, 1 H), 1.57 (m, 2
H), 0.84 (m, 1 H). 13C NMR (101 MHz, DMSO): δ 173.79, 169.60,
164.78, 142.34, 136.13, 128.48, 128.46, 127.84, 127.35, 126.68,
121.84, 119.96, 119.59, 112.24, 104.14, 50.10, 49.39, 41.81, 30.50,
28.52. HRMS (ESI): m/z calcd for C20H21N4O3 [M + H]+, 365.1614;
found, 365.1591. Purity: 99.3%.
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steps). H NMR (400 MHz, DMSO-d6): δ 11.81 (s, 1H), 10.93 (s,
1H), 7.93 (d, J = 2.8 Hz, 1H), 7.38 (d, J = 8.1 Hz, 1H), 7.07−6.98
(m, 2H), 6.92 (d, J = 9.0 Hz, 1H), 6.84 (d, J = 2.9 Hz, 1H), 6.80 (dd,
J = 16.7, 10.5 Hz, 1H), 6.66 (t, J = 7.3 Hz, 1H), 6.45 (d, J = 8.1 Hz,
1H), 6.10 (dd, J = 16.7, 2.4 Hz, 1H), 5.68 (dd, J = 10.4, 2.4 Hz, 1H),
3.59 (m, 4H), 3.29 (s, 3H), 2.91 (m, 4H). 13C NMR (101 MHz,
DMSO-d6): δ 173.05, 172.62, 164.74, 152.47, 144.97, 136.88, 134.75,
130.96, 128.69, 128.10, 127.93, 125.33, 122.42, 121.25, 121.20,
120.60, 120.26, 119.13, 113.12, 112.42, 106.23, 55.98, 50.83, 50.22,
Enzymatic Assay. Kinases were purchased from Carna Bio-
sciences. The enzymatic activities of JAK3 were assessed using the
K
J. Med. Chem. XXXX, XXX, XXX−XXX