Brief Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 5 2327
tolerance of the N1-position to addition of the aminopropyl
linker, we consider this location the most appropriate to
attach large biophysical tags.
(neat) 3352, 3349, 2928, 1684, 1641, 1533, 1245, 1172, 1027,
748 cm-1; HRMS-ESI(þ) m/z calcd for C25H38N5O3 456.2975
[M þ H]þ, found 456.2990 [M þ H]þ. Anal. Calcd for C25H37-
N5O3: C 65.91%, H 8.19%, N 15.37%. Found C 65.52%,
H 8.27%, N 15.04%.
To a solution of carbamate (0.16 g, 0.35 mmol) in MeOH
(8 mL) was added dropwise a solution of 1.2 M HCl in MeOH
(10 mL) and stirred for 12 h at room temperature. The progress
of the reaction was monitored by TLC, and the solvents were
removed in vacuo and 3 times coevaporated with toluene to get
the crude product. The crude product was further purified
To confirm the validity of this hypothesis, we attached a
BODIPY FL fluorophore at this position (Scheme 3) to
create fluorescent analogue 16 (BFL-GR). Ki data show that
16 binds with high affinity to 5-HT3AR (Ki = 2.80 (
0.72 nM, n = 3) and can be visualized in HEK293 cells
expressing 5-HT3AR (Figure 3). Similar images have pre-
viously been obtained using fluorescein, rhodamine 6G, and
cyanine Cy5 dyes attached via a linker at a similar position of
ondansetron, another high-affinity 5-HT3AR antagonist.30
by crystallization (CH2Cl2/Et2O) to afford 7 2HCl (0.15 g,
3
1
0.34 mmol, 98%) as a white solid: mp 296-298 °C; H NMR
(CD3OD, 400 MHz) δ 1.58-1.71 (m, 3H), 1.90-2.10 (m, 3H),
2.21-2.39 (m, 4H), 2.55-2.71 (m, 2H), 2.96-3.01 (m, 5H), 3.74
(d, J = 10.2 Hz, 2H), 4.61-4.70 (m, 3H), 7.30 (t, J = 7.5 Hz,
1H), 7.49 (t, J = 7.7 Hz, 1H), 7.70 (d, J = 8.5 Hz, 1H), 8.22 (d,
J = 8.2 Hz, 1H); 13C NMR (CD3OD, 100 MHz) δ 15.2, 26.7,
31.1, 34.6, 40.9, 41.0, 41.9, 49.7, 57.8, 113.4, 125.6, 126.6, 127.1,
131.0, 137.5, 144.9, 166.8; IR (neat) 2884, 2749, 2676, 1630,
1553, 1208, 745 cm-1; HRMS-ESI(þ) m/z calcd for C20H30N5O
356.2450 [M - 2HCl þ H]þ, found 356.2445 [M - 2HCl þ H]þ.
Anal. Calcd for C20H31Cl2N5O: C 56.07%, H 7.29%, N
16.35%, Cl 16.55%. Found C 55.21%, H 7.35%, N 16.00%,
Cl 16.31%.
Conclusion
In summary, several novel derivatives of granisetron (1)
havebeen discovered. Somederivatives havehighaffinities for
the human 5-HT3AR, and most notably derivatives 2a, 5a, 5c,
and 7 equal or exceed the affinity measured for 1. The data of
our systematic structure-activity study show that substitu-
tion at three positions of granisetron are well tolerated and
that large functional biophysical tags can be attached at the
N1-position.
BFL-GR (16). To a solution of amine hydrochloride 7 2HCl
3
Experimental Section
(9 mg, 0.021 mmol) in anhydrous DMF (0.5 mL) was added
iPr2EtN (5.4 mg, 0.042 mmol) and stirred for 10 min. Then a
solution of BODIPY FL SE (5 mg, 0.012 mmol) in DMF (1 mL)
was added to the mixture and stirred at room temperature for
3.5 h. The progress of the reaction was monitored by TLC. The
DMF was removed in vacuo and the crude product was purified
by flash column chromatography (CH2Cl2 and then CH2Cl2/
MeOH/Et3N, 96:3:1) to afford 16 (7.9 mg, 0.012 mmol, 98%) as
an orange solid: mp 190-192 °C (dec); Rf = 0.42 (CH2Cl2/
General. Chemicals and solvents were either purchased from
commercial suppliers or purified by standard techniques. All
experiments involving air-sensitive reagents were performed
under an inert atmosphere in oven-dried glassware. Thin-layer
chromatography (TLC) was carried out on Merck silica gel
60 F254 plates, and compounds were visualized by irradiating
with UV light, by exposing to I2 vapors, and/or by staining with
cerium molybdate stain (Hanessian’s stain) followed by heating.
Flash chromatography was carried out using Matrex silica gel
60 unless otherwise stated. Infrared spectra were recorded neat
1
MeOH, 7:3); H NMR (CD3OD, 400 MHz) δ 1.07-1.18 (m,
3H), 1.40-1.59 (m, 4H), 1.88-2.05 (m, 6H), 2.15 (s, 3H), 2.32-
2.40 (m, 4H), 2.48-2.52 (m, 5H), 3.08-3.16 (m, 4H), 4.37 (t, J =
6.8 Hz, 2H), 4.47 (tt, J = 6.7 Hz, J = 11.8 Hz, 1H), 6.10 (s, 1H),
6.23 (d, J = 4.0 Hz, 1H), 6.86 (d, J = 4.0 Hz, 1H), 7.17 (t, J =
7.7 Hz, 1H), 7.26 (s, 1H), 7.33 (t, J = 7.4 Hz, 1H), 7.47 (d, J =
8.5 Hz, 1H), 8.10 (d, J = 8.2 Hz, 1H); 13C NMR (CD3OD, 100
MHz) δ 11.2, 14.6, 15.3, 25.6, 33.0, 36.2, 37.8, 40.4, 41.1, 47.6,
53.2, 110.9, 117.7, 123.0, 123.8, 125.8, 128.1, 129.6, 138.3, 139.6,
140.5, 145.2, 149.3, 164.5, 174.8; IR (neat) 2921, 1605, 1488,
1245, 1132, 1056, 974, 734 cm-1; HRMS-ESI(þ) m/z calcd for
C34H43BF2N7O2 630.3539 [M þ H]þ, found 630.3526 [M þ H]þ;
UV/vis/Fluo (MeOH) λmax abs = 506 nm, λmax emiss = 514 nm.
on a Nicolet AVATAR 320 FT-IR spectrometer. H and 13C
1
NMR spectra were recorded on a Bruker DPX-300, DPX-400,
and DRX-500. The chemical shifts are reported in δ (ppm),
and the residual signal of the solvent was used as the inter-
nal standard. High resolution mass spectra were obtained
using electrospray ionization mass (MS-ESI) technique on a
Bruker MicroTOF instrument. Purity was determined by ele-
mental analysis and/or HPLC; purity of key target compounds
was g95%.
1-(3-Aminopropyl)-N-[(3-endo)-9-methyl-9-azabicyclo-[3.3.1]-
non-3-yl]-1H-indazole-3-carboxamide Dihydrochloride (7 2HCl).
3
Compound 14 (0.3 g, 1.0 mmol) was dissolved in anhydrous
DMF/THF (1:5, 10 mL), cooled to 0 °C, and stirred for 5 min.
Then a solution of KOtBu (0.135 g, 1.1 mmol) in anhydrous
THF (2 mL) was added dropwise at 0 °C and stirred for 15 min,
followed by the addition of a solution of tert-butyl N-(3-bromo-
propyl)carbamate (0.27 g, 1.2 mmol) in anhydrous THF (2 mL).
The mixture was warmed to room temperature and stirred for
10 h. The progress of the reaction was monitored by TLC.
The solvents were removed under vacuo, and the residue was
extracted with EtOAc (3 ꢀ 20 mL). The combined organic
phases were dried over Na2SO4, filtered, and concentrated
to give the crude product. The crude product was further
purified by crystallization (CH2Cl2/Et2O) to afford the carba-
mate (0.36 g, 0.79 mmol, 78%) as a white solid: mp 156-158 °C;
Rf = 0.23 (CH2Cl2/MeOH, 7:3); 1H NMR (CD3OD, 400 MHz)
δ 1.14-1.17 (m, 3H), 1.45 (s, 9H), 1.54-1.65 (m, 3H), 2.02-2.21
(m, 4H), 2.44-2.52 (m, 2H), 2.55 (s, 3H), 3.11-3.14 (m, 4H),
4.54 (t, J = 7.7 Hz, 2H), 4.60 (tt, J=6.6 Hz, J = 11.5 Hz, 1H),
7.29 (t, J = 7.5 Hz, 1H), 7.47 (t, J=7.7 Hz, 1H), 7.64 (d, J=
8.5 Hz, 1H), 8.25 (d, J = 8.2 Hz, 1H); 13C NMR (CD3OD,
100 MHz) δ 15.1, 25.7, 28.8, 33.3, 34.8, 38.8, 40.7, 41.6, 47.7,
52.8, 110.9, 123.2, 123.6, 124.0, 128.0, 137.5, 142.5, 166.8; IR
Acknowledgment. The authors thank the EPSRC (EP/
E042139/1) and the Wellcome Trust (081925/Z/07/Z, A.J.T.
and S.C.R.L.) for financial support. S.C.R.L. is a Wellcome
Trust Senior Research Fellow in Basic Biomedical Science.
Supporting Information Available: Synthesis details and spec-
tral data for compounds other than 7 and 16, HPLC purity
assessment for all target compounds, CIF files of 4b and 5a,
experimental details for competition binding, and Ki determina-
tion. This material is available free of charge via the Internet at
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