Studies on Resorcylic Acid Lactone LL-Z1640-2
by column chromatography (gradient elution with EtOAc/hexane)
Triazole Analogue 3: To a solution of triazole macrolactone 15
(72 mg, 0.17 mmol) in MeOH/THF (1:1 v/v, 10 mL) was added 1 n
HCl (5 mL) and the mixture was stirred for 48 h. The mixture was
to give azide 14 (93 mg, 77%) as a clear oil. R
f
= 0.8 (EtOAc/
hexane, 50%); [α]2
3
= –29.8 (c = 1.26, CHCl
=
D
3
). IR (film): ν˜ max
–
1 1
2
(
987, 2940, 1723, 1611, 1284, 1205, 1161, 1052 cm . H NMR extracted with EtOAc (3ϫ 5 mL) and the organic layers were com-
400 MHz, CDCl ): δ = 7.51 (d, J = 15.8 Hz, 1 H), 6.74 (d, J = bined, washed with saturated NaHCO and brine, and dried with
.4 Hz, 1 H), 6.35 (d, J = 2.5 Hz, 1 H), 6.17 (ddd, J = 15.8, 7.9, MgSO . The solvent was evaporated under reduced pressure and
3
3
2
5
4
1
4
.8 Hz, 1 H), 4.36–4.26 (m, 2 H), 3.85 (s, 3 H), 3.44 (dd, J = 16.0, the crude product was purified by column chromatography (EtOAc
.0 Hz, 1 H), 3.32 (dd, J = 16.0, 4.0 Hz, 1 H), 2.61–2.45 (m, 2 H), then MeOH/EtOAc, 10%) to afford triazole 3 (50 mg, 76%) as a
.70 (d, J = 0.8 Hz, 6 H), 1.52 (s, 3 H), 1.38 (s, 3 H) ppm.
): δ = 164.8, 160.2, 158.7, 143.3,
31.0, 129.4, 108.8, 108.5, 105.1, 103.7, 100.4, 76.5, 76.2, 55.6, 51.1,
3.0, 28.1, 25.7, 25.5, 25.4 ppm. HRMS (TOF-ESI): m/z calcd. for
1
3
C
pale-yellow amorphous solid. R
f
= 0.28 (EtOAc, 100%); m.p.
2
1
NMR (100 MHz, DEPT, CDCl
1
3
3
162 °C (dec.); [α]
D
= –44.1 (c = 1.08, DMSO). IR (KBr): ν˜ max
=
–
1
1
3409, 2928, 1645, 1609, 1255, 1210, 1159, 1041 cm . H NMR
(400 MHz, DMSO): δ = 10.16 (s, 1 H), 7.82 (s, 1 H), 6.47 (d, J =
2.3 Hz, 1 H), 6.30 (d, J = 2.3 Hz, 1 H), 6.19–6.07 (m, 1 H), 5.74
+
C
20
H
25
N
3
O
6
[M ] 403.1743; found 403.1733.
(
d, J = 15.9 Hz, 1 H), 5.47–5.36 (m, 1 H), 5.22 (d, J = 4.9 Hz, 1
H), 4.94 (d, J = 4.7 Hz, 1 H), 4.59 (br. s, 1 H), 4.30 (br. d, J =
3.3 Hz, 1 H), 3.83 (d, J = 3.1 Hz, 1 H), 3.72 (s, 3 H), 3.02–2.88
m, 2 H), 2.07 (dt, J = 15.5, 4.2 Hz, 1 H), 1.90 (br. s, 1 H), 1.40 (d,
5
-(E)-3-{(4S,5R)-5-{{4-[(S)-2-Hydroxypropyl-1H-1,2,3-triazol-1-
yl]methyl}-2,2-dimethyl-1,3-dioxolan-4-yl}prop-1-en-1-yl}-7-meth-
oxy-2,2-dimethyl-4H-benzo[d][1,3]dioxin-4-one (16): To a stirred
solution of azide 14 (180 mg, 0.45 mmol) in tert-butyl alcohol
1
(
1
3
J = 6.3 Hz, 3 H) ppm. C NMR (100 MHz, CDCl
3
): δ = 167.9,
61.1, 157.5, 143.4, 137.6, 130.8, 127.6, 123.9, 112.0, 101.6, 100.1,
1.4, 70.8, 70.7, 55.2, 50.7, 35.7, 31.4, 20.3 ppm. HRMS (TOF-
(
1 mL) and H
butyl alcohol (1 mL) was added sodium ascorbate (270 mg,
.4 mmol) and CuSO ·5H O (110 mg, 0.46 mmol). The reaction
was stirred under an argon atmosphere for 18 h and then diluted
with EtOAc (4 mL) and H O (4 mL). The organic phase was sepa-
rated and the aqueous phase was extracted with EtOAc (2 ϫ
0 mL). The combined organic layers were dried with MgSO and
2
O (2 mL), was added 7 (58 mg, 0.69 mmol) in tert-
1
7
1
4
2
+
23 3 6
ESI): m/z calcd. for C19H N O [M ] 389.1587; found 389.1597.
2
Kinase Profiling: The kinase profiling was carried out with Dis-
coveRX using KINOMEscan technology (see the Supporting In-
formation).
1
4
concentrated under reduced pressure. The crude product was puri-
fied by column chromatography (EtOAc/hexane, 50 to 100%) to
Determination of IC50 of Triazole Analogue 3: Determination of the
IC50 values of MNK1/2 was carried out according to the pro-
cedures reported previously.[ MNK1 and MNK2 genes (pur-
chased from GenScript USA Inc) were induced with 1.0 mm IPTG
and grown at 25 °C for 6 h. The proteins were purified by using
affinity chromatography on a Bio-Scale Mini Profinity GST car-
tridge. The GST tag was cleaved by using PreScission protease. The
proteins were concentrated to 10–15 mg/mL (equivalent to 285–
afford triazole 16 (170 mg, 78%) as a clear oil. R
f
= 0.36 (EtOAc,
00%); [α]2
D 3
= –38.5 (c = 1.06, CHCl ). IR (film): ν˜ max = 3384,
3c]
3
1
2
–
1 1
987, 2938, 1722, 1605, 1574, 1277, 1162, 1068 cm . H NMR
): δ = 7.56 (s, 1 H), 7.51 (d, J = 15.9 Hz, 1 H),
(400 MHz, CDCl
3
6
1
.73 (d, J = 2.4 Hz, 1 H), 6.36 (d, J = 2.4 Hz, 1 H), 6.15 (ddd, J =
5.8, 7.5, 5.9 Hz, 1 H), 4.65 (dd, J = 13.7, 3.0 Hz, 1 H), 4.63–4.50
(
9
m, 1 H), 4.46 (dd, J = 13.4, 6.2 Hz, 1 H), 4.33 (dd, J = 13.7,
.6 Hz, 1 H), 4.14 (m, 1 H), 3.85 (s, 3 H), 3.13 (s, 1 H), 2.87 (dd,
J = 15.0, 3.4 Hz, 1 H), 2.74 (dd, J = 15.0, 8.5 Hz, 1 H), 2.71–2.62
m, 1 H), 2.61–2.52 (m, 1 H), 1.69 (s, 6 H), 1.55 (s, 3 H), 1.35 (s, 3
4
–
28 μm) and flash-frozen in liquid nitrogen and stored in buffer at
80 °C. The IMAP 10000-tp FP assay kit was purchased
from Molecular Devices, and JH3 peptide (5-FAM-
TATKSGSTTKNRFVV-NH ) was synthesized by GenScript USA
(
13
H), 1.26 (d, J = 6.2 Hz, 3 H) ppm. C NMR (100 MHz, DEPT,
CDCl ): δ = 164.9, 160.3, 158.7, 145.2, 143.2, 131.3, 128.6, 109.1,
08.6, 105.1, 103.7, 100.4, 76.1, 76.1, 67.1, 55.6, 50.8, 34.8, 33.0,
8.2, 25.6, 25.5, 25.5, 22.8 ppm. HRMS (TOF-ESI): m/z calcd. for
2
Inc. Buffer B contains: 20 mm HEPES/KOH pH 7.4, 10 mm MgCl
2
,
3
0.5 mm DTT. IMAP progressive binding solution was prepared
1
2
freshly on the assay day according to the description in the assay
kit. 2.0 μL of 5.0 μm activated MNK2 (or 4.0 μL of 5.0 μm acti-
vated MNK1), 2.0 μL of compound at 10-fold concentrations, and
buffer B to a total volume of 20 μL were added to a 384-well plate
and incubated at 30 °C for 1 h. 2.0 μL of 1.0 μm JH3 and 2.0 μL
of 200 μm ATP were then added to each well and the plate was
incubated at 30 °C for another 2 h for MNK2 (or 6 h for MNK1).
The reaction was terminated with 60 μL of 1-fold IMAP binding
solution. After 30 min incubation at room temperature, fluores-
cence polarization was measured by the Safire microplate reader.
The raw data were analyzed using GraphPad Prism 5.0 to obtain
the IC50 values.
+
C
25
H
33
N
3
O
7
[M ] 487.2319; found 487.2325.
Triazole Macrolactone 15: A solution of triazole 16 (150 mg,
.30 mmol) in anhydrous THF (10 mL) was added dropwise to a
stirred suspension of NaH (120 mg, 3 mmol) in anhydrous THF
21 mL) and the resulting mixture was stirred for 16 h. The reaction
was quenched carefully by dropwise addition of H O until efferves-
cence ceased. The mixture was extracted with EtOAc (3ϫ 5 mL)
and the organic layers were combined and dried with MgSO . The
0
(
2
4
solvent was evaporated under reduced pressure and the crude prod-
uct was purified by column chromatography (EtOAc/hexane, 20–
5
1
2
1
4
0%) to afford 15 (82 mg, 63%) as a white solid. R
f
= 0.71 (EtOAc,
). IR (film): ν˜ max = 3075,
987, 1643, 1605, 1255 cm . H NMR (400 MHz, CDCl ): δ =
00%); [α]2
3
= –46.2 (c = 0.73, CHCl
Supporting Information (see footnote on the first page of this arti-
D
3
–1
1
1
13
3
cle): Copies of H and C NMR spectra of all new compounds,
1.60 (s, 1 H), 7.46 (s, 1 H), 6.43–6.37 (m, 3 H), 5.72–5.63 (m, 2 H),
.84 (ddd, J = 11.0, 4.0, 2.8 Hz, 1 H), 4.61 (dd, J = 13.8, 11.1 Hz, 1
kinase profiling data (% control) for triazole analogue 3.
H), 4.39–4.31 (m, 1 H), 4.26 (dd, J = 13.8, 2.5 Hz, 1 H), 3.79 (s, 3
H), 3.42 (dd, J = 15.1, 5.2 Hz, 1 H), 2.92 (dd, J = 15.2, 3.6 Hz, 1 Acknowledgments
H), 2.63 (dd, J = 16.2, 3.5 Hz, 1 H), 2.29–2.22 (m, 1 H), 1.51 (t, J
=
3.2 Hz, 6 H), 1.41 (s, 3 H) ppm. 13C NMR (100 MHz, CDCl
3
):
This study was supported by the Institute of Chemical and Engi-
neering Sciences (ICES), Agency for Science, Technology and Re-
search (A*STAR), Singapore. Ms Jin Xu, Institute of Chemical
and Engineering Sciences is thanked for carrying out the IC50 as-
says on MNK kinases.
δ = 170.4, 164.9, 163.7, 142.4, 141.7, 131.6, 127.6, 122.4, 108.0,
1
2
07.8, 104.1, 100.2, 78.0, 74.0, 69.8, 55.4, 49.7, 30.6, 30.3, 28.4,
5.6, 19.5 ppm. HRMS (TOF-ESI): m/z calcd. for C22
27 3 6
H N O
+
[M] 429.1900; found 429.1901.
Eur. J. Org. Chem. 2014, 7239–7244
© 2014 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
7243