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A. Kramer et al. / European Journal of Medicinal Chemistry 208 (2020) 112770
15
column chromatography with a mobile phase of n-hexane and
ethyl acetate (ratio gradually ranging from 5:1 to 1:1) led to the
desired compound as a bright yellow solid (51 mg, 59%). 1H NMR
mobile phase of ethyl acetate with 1% AcOH. The title compound
was obtained as a white solid (23 mg, 35%). 1H NMR (500 MHz,
DMSO‑d6):
d
12.35 (s, 1H, COOH), 9.01 (d, J ¼ 7.8 Hz, 1H, HetH), 8.75
(500 MHz, DMSO‑d6):
d
8.97 (d, J ¼ 7.8 Hz, 1H, HetH), 8.67e8.57 (m,
(d, J ¼ 1.5 Hz, 1H, HetH), 8.72 (s, 1H, PhH), 7.70 (d, J ¼ 8.1 Hz, 1H,
PhH), 7.55 (d, J ¼ 7.8 Hz, 1H, HetH), 7.42 (dd, J ¼ 8.1, 1.5 Hz, 1H, PhH),
4.40 (t, J ¼ 6.0 Hz, 2H, CH2), 4.10 (t, J ¼ 6.6 Hz, 2H, CH2), 3.93 (t,
J ¼ 6.6 Hz, 2H, CH2), 3.89 (t, J ¼ 6.0 Hz, 2H, CH2), 1.54 (s, 9H, (CH3)3)
2H, HetH, PhH), 7.51 (d, J ¼ 7.8 Hz, 1H, HetH), 7.33 (d, J ¼ 7.6 Hz, 1H
PhH), 7.25 (t, J ¼ 7.9 Hz, 1H, PhH), 6.69 (dd, J ¼ 8.3, 2.7 Hz, 1H, PhH),
4.31 (t, J ¼ 6.3 Hz, 2H, CH2), 4.08 (t, J ¼ 6.9 Hz, 2H, CH2), 3.92 (t,
J ¼ 6.9 Hz, 2H, CH2), 3.87 (t, J ¼ 6.3 Hz, 2H, CH2), 1.54 (s, 9H, (CH3)3)
ppm. 13C NMR (126 MHz, DMSO‑d6):
d 166.83, 158.82, 153.36,
ppm. 13C NMR (126 MHz, DMSO‑d6):
d
158.94, 152.70, 152.46,
152.45, 143.03, 142.93, 137.23, 136.50, 131.42, 117.64, 116.61, 112.61,
106.46, 104.15, 82.77, 67.51, 66.86, 66.58, 46.60, 27.67 ppm. MS
(ESIþ) m/z: 441.00 [M þ H]þ. HRMS m/z: [M þ H]þ calcd for
142.43, 136.29, 133.32, 129.38, 116.92, 114.41, 110.88, 107.29, 103.62,
82.64, 66.15, 65.84, 46.18, 27.66 ppm. MS (ESIþ) m/z: 397.20 [M þ
H]þ. HRMS m/z: [M þ H]þ calcd for C21H25N4O4, 397.18703; found
397.18689. HPLC: tR ¼ 18.047, purity ꢃ95%.
C
22H25N4O6, 441.17686; found 441.17510. HPLC: tR ¼ 15.137, purity
ꢃ95%.
Synthesis of Methyl 2-hydroxy-4-(5-{[2-(2-hydroxyethoxy)
ethyl]amino}pyrazolo[1,5-a]pyrimidin-3-yl)benzoate (30). Methyl
4-(5-{[(tert-butoxy)carbonyl][2-(2-hydroxyethoxy)ethyl]amino}
pyrazolo [1,5-a]pyrimidin-3-yl)-2-hydroxybenzoate (22) (75 mg,
0.16 mmol) was suspended in methylene chloride (2.5 mL) and
trifluoroacetic acid (0.75 mg, 6.50 mmol) was added slowly at 0 ꢀC.
After stirring for 16 h at room temperature the solvent was evap-
orated under vacuum. The crude product was purified by silica gel
column chromatography with a mobile phase of n-hexane and
ethyl acetate (ratio gradually ranging from 1:0 to 0:1). The white
solid obtained was the desired compound (50 mg, 85%). 1H NMR
Synthesis of Methyl 7,10-dioxa-13,17,18,21-tetraazatetracyclo
[12.5.2.12,⁶.01⁷,2⁰]docosa-1(20),2,4,6(22),14(21),15,18-heptaene-5-
carboxylate (33). To a 0 ꢀC cooled solution of 13-tert-butyl 5-methyl
7,10-dioxa-13,17,18,21-tetraazatetracyclo [12.5.2.12,⁶.01⁷,2⁰]docosa-1
(20),2,4,6(22),14(21),15,18-heptaene-5,13-dicarboxylate
(26)
(0.30 g, 0.66 mmol) in methylene chloride (12 mL) trifluoroacetic
acid (3.58 g, 31.36 mmol) was added dropwise and it was stirred for
16 h at room temperature. Afterwards the solvent was removed
under reduced pressure and the residue was taken up in methanol.
Potassium carbonate was added to the stirring solution. Subse-
quently, the solution was filtrated and the solvent was removed
again. The residue was taken up with ethyl acetate and it was
washed with water and brine. After drying over MgSO4 and
removal of the organic solvent under vacuum, the title compound
was yielded as a white solid (0.23 g, 99%). 1H NMR (500 MHz,
(500 MHz, DMSO‑d6):
d
10.62 (s, 1H, OH), 8.52 (d, J ¼ 7.6 Hz, 1H,
HetH), 8.44 (s, 1H, HetH), 7.88 (t, J ¼ 5.3 Hz, 1H, NH), 7.78e7.75 (m,
2H, PhH), 7.66 (dd, J ¼ 8.4, 1.6 Hz, 1H, PhH), 6.39 (d, J ¼ 7.6 Hz, 1H,
HetH), 3.89 (s, 3H, OCH3), 3.72e3.49 (m, 9H, (CH3)3) ppm. 13C NMR
(126 MHz, DMSO‑d6):
d
169.63, 160.93, 157.89, 156.33, 145.58,
DMSO‑d6):
d
8.83 (d, J ¼ 1.5 Hz, 1H, HetH), 8.57 (d, J ¼ 7.6 Hz, 1H,
142.43, 141.34, 135.58, 129.88, 115.81, 111.74, 108.03, 103.05, 100.52,
72.25, 68.41, 60.23, 52.25, 40.51 ppm. MS (ESIþ) m/z: 372.96 [M þ
H]þ. HRMS m/z: [M þ H]þ calcd for C18H21N4O5, 373.15065; found
373.15150. HPLC: tR ¼ 12.942, purity ꢃ95%.
HetH), 8.39 (s, 1H, PhH), 7.94 (t, J ¼ 5.4 Hz, 1H, NH), 7.70 (d,
J ¼ 8.1 Hz, 1H, PhH), 7.29 (dd, J ¼ 8.1, 1.4 Hz, 1H, PhH), 6.34 (d,
J ¼ 7.6 Hz, 1H, HetH), 4.37 (t, J ¼ 7.9 Hz, 2H, CH2), 4.00e3.97 (m, 2H,
CH2), 3.89e3.86 (m, 2H, CH2), 3.76 (s, 3H, OCH3), 3.56e3.50 (m, 2H,
Synthesis of 4-(5-{[(tert-Butoxy)carbonyl][2-(2-hydroxyethoxy)
ethyl]amino}pyrazolo[1,5-a]pyrimidin-3-yl)-2-hydroxybenzoic
acid (31). A solution of lithium hydroxide monohydrate (31 mg,
0.74 mmol) in water (1 mL) was added to a solution of methyl 4-(5-
{[(tert-butoxy)carbonyl][2-(2-hydroxyethoxy)ethyl]amino}pyr-
azolo [1,5-a]pyrimidin-3-yl)-2-hydroxybenzoate (22) (70 mg,
0.15 mmol) in tetrahydrofuran (5 mL) and it was stirred for 16 h at
50 ꢀC. The solvent was removed in vacuo and the residue was taken
up with water. Afterwards the solution was acidified with 1 N HCl
and it was extracted with ethyl acetate. The organic layers were
dried over MgSO4 and the solvent was evaporated. The crude
product was purified by silica gel column chromatography with a
mobile phase of petroleum ether and tetrahydrofuran with the
addition of 1% acetic acid (ratio of 1:2). The white solid (29 mg, 42%)
CH2) ppm. 13C NMR (126 MHz, DMSO‑d6):
d 165.66, 158.08, 156.18,
145.33, 141.81, 139.04, 135.82, 131.61, 115.64, 114.82, 110.45, 103.40,
100.21, 65.35, 65.27, 51.57 ppm. MS (ESIþ) m/z: 354.96 [M þ H]þ.
HRMS m/z: [M þ H]þ calcd for C18H19N4O4, 355.14008; found
355.14078. HPLC: tR ¼ 12.869, purity ꢃ95%.
Synthesis
of
7,10-Dioxa-13,17,18,21-tetraazatetracyclo
[12.5.2.12,⁶.01⁷,2⁰]docosa-1(20),2,4,6(22),14(21),15,18-heptaene-5-
carboxylic acid (34). In tetrahydrofuran (3 mL) methyl 7,10-dioxa-
13,17,18,21-tetraazatetracyclo
[12.5.2.12,⁶.01⁷,2⁰]docosa-1
(20),2,4,6(22),14(21),15,18-heptaene-5-carboxylate (33) (50 mg,
0.14 mmol) and lithium hydroxide monohydrate (30 mg,
0.71 mmol) were dissolved and water (0.6 mL) was added. The
reaction mix was stirred at 50 ꢀC for 16 h. After that, the solvent was
removed under reduced pressure and the residue was taken up in
water. By adding 10% hydrochloric acid pH 1 was set and the
precipitated solid was filtrated. Washing and drying of the filtrate
resulted in the white solid tile compound (47 mg, 99%). 1H NMR
was the title compound. 1H NMR (500 MHz, DMSO‑d6):
d 11.40 (s,
1H, OH), 9.00 (d, J ¼ 7.7 Hz, 1H, HetH), 8.76 (s, 1H, HetH), 7.80 (d,
J ¼ 8.2 Hz, 1H, PhH), 7.69e7.66 (m, 2H, PhH), 7.49 (d, J ¼ 7.7 Hz, 1H,
HetH), 4.54 (s, 1H, OH), 4.22 (t, J ¼ 6.4 Hz, 2H, CH2), 3.78 (t,
J ¼ 6.4 Hz, 2H, CH2), 3.49e3.43 (m, 4H, CH2), 1.53 (s, 9H, (CH3)3)
(500 MHz, DMSO‑d6):
d 12.24 (bs, 1H, COOH), 8.81 (s, 1H, HetH),
8.57 (d, J ¼ 7.6 Hz, 1H, HetH), 8.38 (s, 1H, PhH), 7.93 (t, J ¼ 5.4 Hz, 1H,
NH), 7.70 (d, J ¼ 8.1 Hz, PhH), 7.27 (dd, J ¼ 8.1, 1.4 Hz, 1H, PhH), 6.34
(d, J ¼ 7.6 Hz, 1H, HetH), 4.38 (t, J ¼ 7.1 Hz, 2H, CH2), 3.99 (t,
J ¼ 7.5 Hz, 2H, CH2), 3.89e3.86 (m, 2H, CH2), 3.56e3.50 (m, 2H, CH2)
ppm. 13C NMR (126 MHz, DMSO‑d6):
d 171.90, 161.71, 153.79, 152.83,
143.79, 143.04, 139.53, 136.58, 130.51, 116.07, 112.49, 106.40, 104.75,
82.54, 72.27, 67.98, 60.23, 45.83, 31.16, 29.83, 27.67 ppm. MS (ESIþ)
m/z: 458.98 [M þ H]þ. HRMS m/z: [M þ H]þ calcd for C22H27N4O7,
459.18743; found 459.18597. HPLC: tR ¼ 14.985, purity ꢃ95%.
Synthesis of 13-[(tert-Butoxy)carbonyl]-7,10-dioxa-13,17,18,21-
tetraazatetracyclo[12.5.2.12,⁶.01⁷,2⁰]docosa-
1(20),2,4,6(22),14(21),15,18-heptaene-5-carboxylic acid (32). The
desired compound was synthesized according to the procedure of
(31) using 13-tert-butyl 5-methyl 7,10-dioxa-13,17,18,21-
tetraazatetracyclo [12.5.2.12,⁶.01⁷,2⁰]docosa-1 (20),2,4,6(22),14(21),
15,18-heptaene-5,13-dicarboxylate (26) (68 mg, 0.15 mmol). Puri-
fication was done by silica gel column chromatography with a
ppm. 13C NMR (126 MHz, DMSO‑d6):
d 166.70,158.07, 156.14,145.27,
141.76, 138.69, 135.80, 131.82, 115.79, 115.61, 110.39, 103.50, 100.18,
65.31, 65.22, 63.89, 59.77 ppm. MS (ESIþ) m/z: 340.93 [M þ H]þ.
HRMS m/z: [M þ H]þ calcd for C17H17N4O4, 341.12443; found
341.12573. HPLC: tR ¼ 11.408, purity ꢃ95%.
Synthesis
of
Methyl
13-acetyl-7,10-dioxa-13,17,18,21-
tetraazatetracyclo[12.5.2.12,⁶.01⁷,2⁰]docosa-
1(20),2,4,6(22),14(21),15,18-heptaene-5-carboxylate (35). Acetyl
chloride (138 mg, 1.76 mmol) and N,N-diisopropylethylamine
(227 mg, 1.76 mmol) were added to a solution of methyl 7,10-dioxa-