Journal of Medicinal Chemistry
Article
chloroform-d) δ 9.31 (s, 1H), 9.19 (d, J = 7.4 Hz, 1H), 6.55 (d, J =
1.7 Hz, 1H), 6.51 (d, J = 1.9 Hz, 1H), 6.22 (s, 1H), 4.49 (s, 2H),
3.79−3.64 (m, 1H), 3.40 (d, J = 16.0 Hz, 1H), 2.83 (s, 2H), 2.55 (s,
3H), 2.40 (s, 2H), 2.19 (d, J = 11.2 Hz, 2H), 2.09−2.00 (m, 2H),
1.44 (dd, J = 18.9, 9.5 Hz, 4H), 1.10 (s, 6H). 13C NMR (101 MHz,
CDCl3) δ 193.6, 172.2, 151.4, 150.1, 149.3, 143.1, 141.5, 139.9, 117.2,
108.2, 105.4, 103.9, 100.8, 69.5, 60.7, 52.3, 50.2, 37.7, 35.8, 33.4 (2),
29.9 (2), 28.3 (2), 13.3. HRMS (ESI) m/z [M + H] calcd for
C26H33N4O4, 465.2502, found 465.2501.
8-Morpholino-3-propyl-6-(3,6,6-trimethyl-4-oxo-4,5,6,7-tetrahy-
dro-1H-indazol-1-yl)isoquinolin-1(2H)-one (5a). Yield 65%, 20 mg;
1H NMR (400 MHz, chloroform-d) δ 9.53 (s, 1H), 7.10 (d, J = 2.0
Hz, 1H), 7.05 (d, J = 2.1 Hz, 1H), 6.25 (s, 1H), 4.00 (t, J = 4.5 Hz,
4H), 3.22 (s, 4H), 2.86 (s, 2H), 2.57 (s, 5H), 2.43 (s, 2H), 1.76 (h, J
= 7.4 Hz, 2H), 1.12 (s, 6H), 1.03 (t, J = 7.3 Hz, 3H). 13C NMR (126
MHz, CDCl3) δ 193.4, 162.1, 155.4, 150.4, 149.1, 143.1, 142.8, 141.9,
117.5, 115.4, 112.9, 109.7, 104.3, 67.2 (2), 53.4 (2), 52.4, 37.6, 35.9,
34.8, 28.4 (2), 21.2, 13.5, 13.5. HRMS (ESI) m/z [M + H] calcd for
C26H33N4O3, 449.2553, found 449.2557.
1.01 (t, J = 7.4 Hz, 3H). 13C NMR (101 MHz, CDCl3) δ 193.5, 165.8,
151.3, 149.9, 149.1, 143.1, 142.1, 141.6, 140.8, 107.6, 105.3, 105.0,
100.2, 52.4, 46.3, 37.7, 35.8, 34.9 (2), 32.1, 31.6, 31.4, 28.4 (2), 21.2
(2), 13.5, 13.4. HRMS (ESI) m/z [M + Na] calcd for C28H37N5O2Na,
498.2845, found 498.2823.
8-((2-(Diethylamino)ethyl)(methyl)amino)-3-propyl-6-(3,6,6-tri-
methyl-4-oxo-4,5,6,7-tetrahydro-1H-indazol-1-yl)isoquinolin-
1(2H)-one (5g). Yield 45%, 14 mg; 1H NMR (500 MHz, chloroform-
d) δ 9.22 (s, 1H), 7.03 (d, J = 2.0 Hz, 1H), 7.00 (d, J = 2.0 Hz, 1H),
6.21 (s, 1H), 3.44−3.37 (m, 2H), 3.02 (s, 3H), 2.86 (s, 2H), 2.77 (t, J
= 7.4 Hz, 2H), 2.57 (s, 3H), 2.56−2.48 (m, 6H), 2.42 (s, 2H), 1.81−
1.71 (m, 2H), 1.12 (s, 6H), 1.03 (t, J = 7.3 Hz, 3H), 0.97 (t, J = 7.1
Hz, 6H). 13C NMR (126 MHz, CDCl3) δ 193.5, 162.0, 155.2, 150.1,
149.1, 142.9, 141.5, 117.3, 114.5, 111.4, 110.0, 104.3, 55.5, 52.4, 50.4,
47.3 (2), 41.7, 37.5, 35.9, 35.0, 29.9, 28.4 (2), 21.2, 13.6, 13.5 (2),
11.7. HRMS (ESI) m/z [M + H] calcd for C29H42N5O2, 492.3338,
found 492.3352.
General Procedure for the Syntheses of 6a and 6b. To a
solution of 4d/5b (0.1 mmol, 1 equiv) in 10 mL of ethanol was
introduced palladium (10% on activated carbon) (10 mol %) in a
pressure reactor vessel. After multiple cycles of degassing under
vacuum, hydrogen gas was introduced at 200 psi and the reactor
vessel was heated to 90 °C for 24 h. Upon completion, the reaction
mixture was passed through a plug of Celite and solvent was removed
under vacuum. Subsequently, preparative TLC was performed (SiO2,
5% MeOH in dichloromethane) to obtain the desired compounds.
8-(((1r,4r)-4-Hydroxycyclohexyl)amino)-3-propyl-6-(3,6,6-tri-
methyl-4-oxo-4,5,6,7-tetrahydro-1H-indazol-1-yl)-3,4-dihydroiso-
quinolin-1(2H)-one (6a). White amorphous solid, 20 mg, yield 42%.
1H NMR (400 MHz, chloroform-d) δ 8.78 (d, J = 7.5 Hz, 1H), 6.56
8-(4-Hydroxypiperidin-1-yl)-3-propyl-6-(3,6,6-trimethyl-4-oxo-
4,5,6,7-tetrahydro-1H-indazol-1-yl)isoquinolin-1(2H)-one (5b).
1
Yield 55%, 18 mg; H NMR (500 MHz, chloroform-d) δ 9.03 (s,
1H), 6.98 (d, J = 3.3 Hz, 2H), 6.15 (s, 1H), 3.86 (s, 1H), 3.41 (s,
2H), 2.87 (s, 2H), 2.78 (s, 2H), 2.52−2.44 (m, 5H), 2.35 (s, 2H),
2.12−2.03 (m, 2H), 1.92−1.81 (m, 2H), 1.67 (h, J = 7.4 Hz, 2H),
1.05 (s, 6H), 0.95 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3)
δ 193.5, 162.0, 155.8, 150.3, 149.1, 142.9, 142.4, 141.8, 117.4, 115.5,
112.4, 110.1, 104.2, 53.4, 52.4 (2), 37.6, 35.9, 34.9 (2), 30.9, 29.7,
28.4 (2), 21.1, 13.5, 13.5. HRMS (ESI) m/z [M + H] calcd for
C27H35N4O3, 463.2709, found 463.2710.
(d, J = 2.0 Hz, 1H), 6.49 (d, J = 1.9 Hz, 1H), 5.72 (s, 1H), 3.75 (dt, J
= 9.6, 4.9 Hz, 1H), 3.60 (m, J = 8.4, 6.8, 5.0, 2.7 Hz, 1H), 3.40−3.27
(m, 1H), 2.91 (dd, J = 15.4, 4.2 Hz, 1H), 2.83 (s, 2H), 2.74 (dd, J =
15.3, 10.5 Hz, 1H), 2.55 (s, 3H), 2.41 (s, 2H), 2.23−1.98 (m, 4H),
1.64−1.52 (m, 2H), 1.50−1.36 (m, 6H), 1.12 (s, 6H), 0.97 (t, J = 7.2
Hz, 3H). 13C NMR (101 MHz, CDCl3) δ 193.4, 168.7, 150.7, 150.0,
148.9, 142.3, 142.0, 117.2, 108.5, 107.9, 104.1, 69.6, 53.4, 52.3, 50.5,
50.2, 37.7, 37.2, 35.8, 35.6, 33.6, 30.2, 30.1 (2), 28.4, 18.6, 13.9, 13.4.
HRMS (ESI) m/z [M + H] calcd for C28H39N4O3, 479.3017, found
479.3041.
8-(4-Methylpiperazin-1-yl)-3-propyl-6-(3,6,6-trimethyl-4-oxo-
4,5,6,7-tetrahydro-1H-indazol-1-yl)isoquinolin-1(2H)-one (5c).
1
Yield 55%, 18 mg; H NMR (400 MHz, chloroform-d) δ 9.56 (s,
1H), 9.35 (d, J = 7.3 Hz, 1H), 7.17 (d, J = 2.0 Hz, 1H), 7.02 (d, J =
2.1 Hz, 1H), 6.19 (d, J = 1.9 Hz, 1H), 2.87 (s, 2H), 2.80 (s, 1H), 2.57
(s, 3H), 2.52 (t, J = 7.4 Hz, 2H), 2.43 (s, 2H), 2.33 (s, 3H), 2.26 (t, J
= 10.5 Hz, 2H), 2.16−2.08 (m, 2H), 1.79−1.58 (m, 7H), 1.13 (s,
6H), 1.01 (t, J = 7.4 Hz, 3H). 13C NMR (101 MHz, CDCl3) δ 193.5,
165.8, 151.2, 149.9, 149.0, 143.1, 142.1, 141.6, 140.8, 107.7, 105.3,
105.0, 100.2, 52.4, 46.3, 37.7, 35.8, 34.9 (2), 32.1, 31.6, 31.4, 28.4 (2),
21.2 (2), 13.5, 13.4. HRMS (ESI) m/z [M + Na] calcd for
C28H37N5O2Na, 498.2845, found 498.2823.
8-(4-Hydroxypiperidin-1-yl)-3-propyl-6-(3,6,6-trimethyl-4-oxo-
4,5,6,7-tetrahydro-1H-indazol-1-yl)-3,4-dihydroisoquinolin-1(2H)-
1
one (6b). White amorphous solid, 24 mg, yield 47%; H NMR (400
8-(Piperazin-1-yl)-3-propyl-6-(3,6,6-trimethyl-4-oxo-4,5,6,7-tet-
MHz, chloroform-d) δ 6.94 (d, J = 2.1 Hz, 1H), 6.85 (d, J = 2.1 Hz,
1H), 6.01−5.90 (m, 1H), 3.88 (dd, J = 8.5, 4.6 Hz, 1H), 3.66−3.54
(m, J = 10.5, 6.6, 3.2 Hz, 1H), 3.50−3.32 (m, 3H), 3.05−2.88 (m,
3H), 2.82−2.71 (m, 3H), 2.54 (s, 3H), 2.42−2.36 (m, 2H), 2.16−
1.98 (m, 1H), 1.99−1.74 (m, 2H), 1.65−1.52 (m, 2H), 1.51−1.37
(m, J = 14.3, 7.2, 5.1 Hz, 2H), 1.10 (d, J = 6.6 Hz, 6H), 0.97 (t, J = 7.2
Hz, 3H). 13C NMR (101 MHz, CDCl3) δ 193.4, 165.2, 153.8, 150.1,
149.0, 143.0, 141.1, 118.1, 117.2, 114.0, 111.8, 67.4, 52.3, 50.2, 50.1,
49.5, 37.4, 36.9, 36.8, 35.9, 34.3, 34.2, 28.5, 28.3, 18.8, 13.9, 13.4.
HRMS (ESI) m/z [M + H] calcd for C27H37N4O3, 465.2860, found
465.2864.
Fluorescence Polarization Assay. The assay was performed in a
96-well format in black, flat-bottom plates (Santa Cruz Biotechnol-
ogy) with a final volume of 100 μL. Twenty-five microliters of assay
buffer (20 mM HEPES, pH 7.3, 50 mM KCl, 5 mM MgCl2, 20 mM
Na2MoO4, 2 mM DTT, 0.1 mg/mL BGG, and 0.01% NP-40), 25 μL
of assay buffer containing 6 nM FITC-GDA (fluorescent tracer, stock
in DMSO and diluted in assay buffer), and 50 μL of assay buffer
containing 10 nM of Hsp90β(Enzo, ADI-SPP-777), Hsp90α(Enzo,
ADI-SPP-776), Grp94(Enzo, ADI-SPP-766), or 50 nM Trap1 (Enzo,
ADI-SPP-848) was added to each well. Compounds were tested in
triplicate wells (1% DMSO final concentration). For each plate, wells
containing buffer only (background), tracer in buffer only (low
polarization control), and protein and tracer in buffer with 1% DMSO
(high polarization control) were included. Plates were incubated at
4 °C with rocking for 2 h. Polarization values (in mP units) were
measured at 37 °C with an excitation filter at 485 nm and an emission
rahydro-1H-indazol-1-yl)isoquinolin-1(2H)-one (5d). Yield 45%, 14
1
mg; H NMR (400 MHz, chloroform-d) δ 9.59 (s, 1H), 7.09 (d, J =
2.0 Hz, 1H), 7.04 (d, J = 2.0 Hz, 1H), 6.17 (s, 1H), 3.13 (s, 6H), 2.79
(s, 2H), 2.50 (s, 5H), 2.35 (s, 2H), 2.27−2.01 (m, 2H), 1.68 (h, J =
7.4 Hz, 2H), 1.05 (s, 6H), 0.95 (t, J = 7.3 Hz, 3H). 13C NMR (126
MHz, CDCl3) δ 193.4, 161.7, 155.8, 150.3, 149.1, 143.0, 142.2, 141.9,
117.4, 115.5, 112.7, 109.8, 104.3, 54.3, 52.4 (2), 46.1 (2), 37.6, 35.9,
34.9, 28.5 (2), 21.2, 13.5, 13.5. HRMS (ESI) m/z [M + H] calcd for
C26H34N5O2, 448.2713, found 448.2706.
tert-Butyl-4-(1-oxo-3-propyl-6-(3,6,6-trimethyl-4-oxo-4,5,6,7-tet-
rahydro-1H-indazol-1-yl)-1,2-dihydroisoquinolin-8-yl)piperazine-
1-carboxylate (5e). Yield 53%, 20 mg; 1H NMR (500 MHz,
chloroform-d) δ 8.93 (s, 1H), 7.01 (d, J = 2.0 Hz, 1H), 6.96 (d, J =
2.1 Hz, 1H), 6.16 (d, J = 2.0 Hz, 1H), 3.75 (s, 4H), 3.08 (s, 4H), 2.79
(s, 2H), 2.51−2.43 (m, 5H), 2.35 (s, 2H), 1.66 (h, J = 7.4 Hz, 2H),
1.42 (s, 9H), 1.05 (s, 6H), 0.94 (t, J = 7.3 Hz, 3H). 13C NMR (126
MHz, CDCl3) δ 193.4, 161.8, 155.4, 154.9, 150.4, 149.1, 143.0, 142.5,
141.9, 117.5, 115.6, 112.9, 110.0, 104.2, 79.8, 53.0 (2), 52.4 (2), 37.6,
35.9, 34.9, 31.6, 28.5 (4), 21.1, 13.5, 13.5. HRMS (ESI) m/z [M + H]
calcd for C31H42N5O4, 548.3237, found 548.3226.
8-((1-Methylpiperidin-4-yl)amino)-3-propyl-6-(3,6,6-trimethyl-4-
oxo-4,5,6,7-tetrahydro-1H-indazol-1-yl)isoquinolin-1(2H)-one (5f).
Yield 40%, 13 mg; 1H NMR (400 MHz, chloroform-d) δ 9.56 (s, 1H),
9.35 (d, J = 7.3 Hz, 1H), 6.60 (d, J = 2.0 Hz, 1H), 6.53 (d, J = 2.1 Hz,
1H), 6.19 (d, J = 1.9 Hz, 1H), 2.87 (s, 2H), 2.80 (s, 1H), 2.57 (s,
3H), 2.52 (t, J = 7.4 Hz, 2H), 2.43 (s, 2H), 2.33 (s, 3H), 2.26 (t, J =
10.5 Hz, 2H), 2.16−2.08 (m, 2H), 1.79−1.58 (m, 7H), 1.13 (s, 6H),
1554
J. Med. Chem. 2021, 64, 1545−1557