Synthesis of fluorine substituted pyrazolopyrimidines as potential leads for the development of PET-imaging agents for the GABAA receptors

Article abstract of DOI:10.1016/j.bmc.2007.10.079

Neuroimaging of GABA_A receptors offers the potential for a better diagnosis of diseases related to a dysfunction of the GABAergic neurotransmission. A series of potent fluorinated analogues of the pyrazolopyrimidine Indiplon has been synthesized and evaluated in vitro as potential agents for imaging the GABA_A receptor by means of positron emission tomography (PET). The most promising compound N-(3-fluoropropyl)-N-[3-[3-(thiophene-2-carbonyl)-pyrazolo[1,5-a]pyrimidin-7-yl]-phenyl]-acetamide (5b) showed an IC₅₀ value of 2.78 ± 0.63 nM comparable to the lead compound Indiplon (IC₅₀ 3.29 ± 0.37 nM), thus making it an interesting candidate for further investigations. In addition to the fluorinated reference compounds, suitable precursors for ¹⁸F-radiolabelling studies have been synthesized.

Full text of DOI:10.1016/j.bmc.2007.10.079

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry 16 (2008) 1184–1194
Synthesis of fluorine substituted pyrazolopyrimidines as potential
leads for the development of PET-imaging agents
for the GABAA receptors
Alexander Hoepping,a,* Michael Diekers,a Winnie Deuther-Conrad,b
Matthias Scheunemann,b Steffen Fischer,b Achim Hiller,b
Florian Wegner,c Jo¨rg Steinbachb and Peter Brustb
aABX advanced biochemical compounds GmbH, H.-Gla¨ser-Str. 10-14, D-01454 Radeberg, Germany
bInstitute of Interdisciplinary Isotope Research, Permoserstr. 15, D-04318 Leipzig, Germany
cDepartment of Neurology, University of Leipzig, Liebigstr. 22a, D-04103 Leipzig, Germany
Received 19 June 2007; revised 15 October 2007; accepted 23 October 2007
Available online 26 November 2007
Abstract—Neuroimaging of GABAA receptors offers the potential for a better diagnosis of diseases related to a dysfunction of the
GABAergic neurotransmission. A series of potent fluorinated analogues of the pyrazolopyrimidine Indiplon has been synthesized
and evaluated in vitro as potential agents for imaging the GABAA receptor by means of positron emission tomography (PET). The
most promising compound N-(3-fluoropropyl)-N-[3-[3-(thiophene-2-carbonyl)-pyrazolo[1,5-a]pyrimidin-7-yl]-phenyl]-acetamide
(5b) showed an IC50 value of 2.78 0.63 nM comparable to the lead compound Indiplon (IC50 3.29 0.37 nM), thus making it
an interesting candidate for further investigations. In addition to the fluorinated reference compounds, suitable precursors for
18F-radiolabelling studies have been synthesized.
ꢀ 2007 Elsevier Ltd. All rights reserved.
1. Introduction
large number of structurally diverse compounds like
pyrazolopyrimidines, imidazopyridines9 and imidazol-
ones10 that act at these receptor sites as agonists,11 in-
verse agonists12 and antagonists13 were developed
(Fig. 1).
Impairment of GABAergic neurotransmission is as-
sumed to be crucial in the pathogenesis of a variety of
neurological disorders such as anxiety, sleep disorders,
or epilepsy. The inhibitory neurotransmitter c-aminobu-
tyric acid (GABA) binds to three classes of receptors:
GABAA and GABAC are ligand-gated ion channels
and GABAB are G-protein coupled receptors. Molecu-
lar imaging of neuroreceptors, in particular by positron
emission tomography (PET), is a powerful tool to study
the pathology of brain diseases in vivo. To date, the
majority of radiotracers suitable for GABAA receptor
imaging are structurally related to benzodiazepines, with
[11C]flumazenil,1,2 [18F]flumazenil3 and [18F]fluoroethyl-
flumazenil4–8 as the most widely used tracers. However,
benzodiazepine binding site ligands bind to a variety of
GABAA receptor subtypes, which are characterized by a
heterogeneity of expression and function. Therefore, a
NC
N
N
N
N
O
O
A
S
Zaleplon
N
B N
N
N C
N
D
O
O
Indiplon
N
N
Zolpidem
Keywords: GABAA; Indiplon; Fluorine-18; PET.
*
Figure 1. Selected structures of compounds with high affinity for the
GABAA receptor.
Corresponding author. Tel.: +49 3528 404164; fax: +49 3528
404165; e-mail: hoepping@abx.de
0968-0896/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2007.10.079

A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
1185
Novel ‘nonbenzodiazepines’ that exhibit high affinity to
the GABAA receptor display a high selectivity for recep-
tors containing the a1-subunit. Isoforms containing the
a1-subunit constitute about 60% of the whole GABAA
receptor population and represent an important phar-
maceutical target since sedative and hypnotic effects
are primarily mediated by these receptors.14 Compounds
that bind to the GABAA-a1 receptor and were devel-
oped for the treatment of insomnia are the imidazopyri-
dine Zolpidem, the cyclopyrrolone Zopiclone and the
pyrazolopyrimidines Indiplon and Zaleplon. The sub-
nanomolar binding affinity of Indiplon with Ki values
of 0.55 and 0.45 nM measured at rat cerebellar and cere-
bral cortex membranes is a prerequisite for the develop-
ment of imaging agents.15 Therefore, we chose Indiplon
as a lead structure for the development of PET-radiotra-
cers. Due to the superior imaging properties of 18F with
t1/2 = 109 min in contrast to t1/2 = 20 min of 11C we fo-
cused on the synthesis of fluorinated Indiplon deriva-
tives.16 Since Indiplon does not contain a fluorine
atom, it has to be incorporated without altering the bio-
logical properties. Hence, we developed several fluori-
nated pyrazolopyrimidines as reference compounds to
investigate their biological properties in terms of
in vitro affinity and specificity. Furthermore, corre-
sponding precursors for future 18F-labelling have been
synthesized.
ture methods.17 With Demethylindiplon in hand, we
envisioned a straightforward fluoroalkylation reaction
for the preparation of various fluoroalkyl derivatives
of Indiplon. However, yields of the corresponding fluo-
roalkyl derivatives were rather low due to the poor reac-
tivity of the demethyl derivative, caused by the
decreased nucleophilicity of the amide nitrogen. Thus,
the introduction of the fluoroalkyl group had to be per-
formed at an earlier stage of the reaction sequence
(Scheme 1). 3-Aminoacetophenone (1) was reacted with
various fluoroalkyl tosylates or halogenides to provide
the fluoroalkyl acetophenones 2a–c which were subse-
quently acylated with carboxylic acid anhydrides to
yield the amides 3a–e. Then the enaminones 4a–e as
the first key building block of the overall strategy were
synthesized by reacting 3a–e with N,N-dimethylformam-
ide dimethyl acetal (DMFDMA). In analogy to the syn-
thesis of Indiplon, the condensation of enaminones 4a–e
with the second key building block aminopyrazole 617
yielded the respective fluoroalkyl derivatives 5a–e under
acidic conditions.
The synthesis of potential precursors for 18F-labelling
was performed in a similar manner (Scheme 1). Again,
3-aminoacetophenone (1) was used as a starting mate-
rial. By reaction with various hydroxyalkyl halogenides,
neat or in aqueous solution, the x-hydroxyalkyl deriva-
tives 7b,c were obtained in good yields. Next, the x-
hydroxyalkyl derivatives were acylated with different
acetic acid anhydrides. Since a selective monoacylation
of the amino group was difficult to achieve, the N,O-
bis-acyl derivatives 8b,c were subjected to a mild O-
deacylation using potassium carbonate in methanol.
Compounds 9b,c were then converted to the enaminones
10b,c which upon reaction with the aminopyrazole 6
2. Results and discussion
2.1. Chemical synthesis
The synthesis of the lead compound Indiplon and its
demethyl derivative was performed according to litera-
R1
R1
R2
n
R1
R2
HN
n
NH2
N
n
b
a
1
2
3
1
2
3
2
2
2
3
2
3
2
3
2
3
F
F
F
F
F
F
F
F
OH
OH
OAc
OAc
OH
OH
-
-
-
2a
2b
2c
3a
3b
3c
3d
3e
7b
7c
8b
8c
9b
9c
O
O
O
- 5a
- 5b
- 5c
- 5d
- 5e
Acetyl
Acetyl
Acetyl
Propionyl
Isobutyroyl
-
2a-c
7b,c
3a-e
1
8b,c
9b,c
c
O
R1
-
R2
N
n
Acetyl
Acetyl
Acetyl
Acetyl
S
e
R2
N
N
d
N
N
NMe2
- 11b
- 11c
R1
n
O
12b
12c
OTos Acetyl
OTos Acetyl
5a-e
11b,c
12b,c
4a-e
10b,c
f
Scheme 1. Reagents and conditions: (a) for 2a: toluene-4-sulfonic acid 2-fluoroethyl ester, CaCO3, water, reflux 3.5 h (42%); for 2b: toluene-4-
sulfonic acid 3-fluoropropyl ester, CaCO3, water, reflux 3.5 h (51%); for 2c: 1-chloro-4-fluorobutane, CaCO3, water, reflux 3.5 h (13%); for 7b: 3-
bromo-1-propanol, CaCO3, water, reflux 2 h (46%); for 7c: 4-chloro-1-butanol, CaCO3, water, reflux 2 h (7%); (b) for 3a–c: acetic anhydride, DMAP,
pyridine, 0 ꢁC to rt 16 h (45%, 57% and 64%, resp.); for 3d: propionic anhydride, DMAP, pyridine, 0 ꢁC to rt 16 h (39%); for 3e: isobutyryl chloride,
DMAP, pyridine, 0 ꢁC to rt 16 h (13%); for 8b,c: acetic anhydride, DMAP, pyridine, rt 16 h (for 8b 100%, for 8c 97%); (c) K2CO3, methanol/water, rt
15 min (97%); (d) DMFDMA, reflux 16 h (for 4a 92%, for 4b 65%, for 4c 81%, for 4d 69%, for 4e 84%, for 10b 74% and for 10c 71%); (e) acetic acid,
6, 65 ꢁC 1.5 h (for 5a 44%, for 5b 46%, for 5c 47%, for 5d 55%, for 5e 41%, for 11b 58% and for 11c 56%); (f) toluene-4-sulfonic acid anhydride,
pyridine, dichloromethane, 0 ꢁC to rt 1.25 h (for 12b 65%, for 12c 55%).

1186
A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
yielded the x-hydroxyalkyl derivatives 11b,c of Indip-
lon. Potential labelling precursors 12b,c were obtained
by converting the x-hydroxy group of these compounds
into tosylate leaving groups.
DMFDMA to the enaminone 21 and further to the isox-
azole 22 proceeded quite smoothly. Ring opening of
isoxazole 22 yielded the cyano-enaminone 23 which
was converted to the benzoyl substituted aminopyrazole
24. Aminopyrazole 24 was then condensed with com-
pound N-[3-(3-dimethylamino-acryloyl)-phenyl]-N-methyl-
acetamide to furnish the target compound 25 in good
yields (Scheme 3).
In a second approach the methyl group was maintained
and the acetyl group was replaced by a 4-fluorobenzoyl
group. Here we started again from 3-aminoacetophe-
none (1) which was N-methylated according to literature
methods.17 The methylamino acetophenone 13 was then
acylated with the respective substituted benzoyl chlo-
rides to yield the fluoro-derivative 16a as reference and
the nitro-derivative 16b as the corresponding precursor
compound (Scheme 2).
2.2. In vitro binding assays
Different strategies for the synthesis of fluorinated pyr-
azolopyrimidines have been developed. First, we synthe-
sized a series of fluoroalkyl derivatives of Indiplon by
replacing the methyl group of the N-methylacetanilide
part with an x-fluorinated alkyl group. Three com-
pounds have been tested for their binding affinity at GA-
BAA receptor complexes (IC50 values are shown in
Table 1). Compound 5b has the highest affinity with
an IC50 of 2.78 0.63 nM on cerebellar GABAA recep-
tors of the adult rat. However, differences in binding
affinity between the fluoroethyl compound 5a (IC50
5.71 0.95 nM) as the compound with the shortest alkyl
chain and the fluorobutyl compound 5c (IC50
5.98 0.98 nM) with the longest alkyl chain are rather
small. The IC50 values for the fluoroalkyl compounds
Eventually, the acetyl group in Indiplon was substituted
by a fluoroethyl group. Thus, 13 was alkylated with
bromofluoroethane to afford fluoroethyl aminoaceto-
phenone 17. Enaminone 18 was obtained by refluxing
17 in DMFDMA. Condensation with aminopyrazole 6
furnished 19 in good yields (Scheme 2).
Another approach aimed at the replacement of the
thiophene ring A in Indiplon by a fluorophenyl ring.
Hence, the aminopyrazole building block had to be
modified. Conversion of 4-fluoroacetophenone (20) with
R3
HN
NH2
N
b
a
R3
O
O
O
- 16a
- 16b
- 19
4-Fluorobenzoyl
4-Nitrobenzoyl
2-Fluoroethyl
14a
14b
17
13
1
14a,b
17
O
R3
N
S
N
d
R3
N
N
c
N
NMe2
O
15a,b
18
16a,b
19
Scheme 2. Reagents and conditions: (a) MeI, K2CO3, DMF; (b) for 14a: 4-fluorobenzoyl chloride, DMF, triethylamine, rt 16 h (96%); for 14b: 4-
nitrobenzoyl chloride, DMF, triethylamine, rt 16 h (95%); for 17: toluene-4-sulfonic acid 2-fluoroethyl ester, CaCO3, water, reflux 3.5 h (41%); (c)
DMFDMA, reflux, 16 h (for 15a 80%, for 15b 89%, and for 18 86%); (d) acetic acid, 6, 65 ꢁC 1.5 h (for 16a 78%, for 16b 79% and for 19 35%).
O
O
N
O
F
b
c
a
NMe2
F
F
F
21
20
23
22
O
O
O
N
CN
e
N
N
F
d
N
N
H2N
N
NMe2
F
O
H
24
25
Scheme 3. Reagents and conditions: (a) DMFDMA, reflux 16 h (63%); (b) NH2OH Æ HCl, methanol, reflux 2.5 h (97%); (c) DMFDMA, reflux 16 h
(60%); (d) aminoguanidine nitrate, NaOH, ethanol, reflux 4 h (62%); (e) acetic acid, N-[3-(3-dimethylamino-acryloyl)-phenyl]-N-methyl-acetamide,
65 ꢁC 1.5 h (48%).

A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
1187
Table 1. Binding affinities of the newly synthesized Indiplon-derived
compounds at GABAA receptors isolated from adult rat brain tissue
(cerebellum) and labelled with [3H]flunitrazepam (ꢀ0.3 nM)
ring A in Indiplon with a 4-fluorophenyl ring in 25 (IC50
2.78 0.63 nM for 5b vs 53.3 18.3 nM for 25). The
introduction of an aromatic fluorine label is expected
to be rather stable against biodegradation. Moreover,
the carbonyl group in 4-position exerts an electron with-
drawing effect that facilitates a nucleophilic substitution
of a nitro or a trimethylammonium group by fluoride.
However, the low binding affinity of this compound pre-
cludes a further development of a tracer based on this
compound.
Compound
IC50 (nM)
Indiplon
ABX012
ABX016
ABX014
ABX017
ABX021
ABX006
ABX007
ABX008
ABX020
3.29 0.37
5.71 0.95
2.78 0.63
5.98 0.98
5.26 1.30
37.7 15.4
14.1 1.70
15.9 7.04
17.8 1.75
53.3 18.3
5a
5b
5c
5d
5e
16a
16b
19
25
2.3. Measurement of logD values
A high brain uptake is important for radiotracers de-
signed to monitor neuroreceptor distribution and den-
sity. The logP value gives a first estimate of the ability
of a compound to cross the blood–brain barrier.
Although not to be taken as an absolute criterion, logP
values within a range of 0.9–2.5 indicate a good brain
penetration of tracer compounds.18 Therefore, we deter-
mined the logD values of the Indiplon-derived com-
pounds by means of HPLC and compared these data
with calculated logP values (ChemOffice, Table 2).19
The logD values were found to be in the range between
2.2 and 2.7. Calculated logP values differ somewhat
from the measured values, the differences are probably
due to difficulties in fragmentation of these rather com-
plex compounds and protonation at physiological pH.
However, based on our experimental data we assume
that our newly synthesized compounds are able to cross
the blood–brain barrier in a sufficient amount.
The IC50 values were obtained from triplicates, estimated by nonlinear
regression analyses, and are given as means SD (n P 3).
are in the same order of magnitude as the IC50 value for
Indiplon (3.29 0.37 nM). These excellent binding affin-
ities make the fluoroalkyl compounds promising candi-
dates for the development of radiotracers. Next, we
investigated the influence of the alkanoic acid residue
of the N-methylanilide in the propanamide 5d and the
isobutyramide 5e. While the binding affinity of 5d is
comparable to that of the acetamides, it is nearly one or-
der of magnitude lower for 5e (IC50 5.26 1.3 nM and
37.7 15.4 nM, respectively). Apparently, the binding
affinity of the pyrazolopyrimidines is rather sensitive to-
wards changes of the acid residue at the amide group for
the fluoroalkyl compounds. On the other hand, the IC50
of 14.1 1.7 nM for the fluorobenzamide 16a does not
show such a drastic decrease in binding affinity. This re-
sult is complemented by a similar IC50 value of
15.9 7.0 nM obtained for the nitrobenzamide 16b.
Therefore, we assume that the overall steric bulk at
the amide nitrogen contributes to the receptor binding.
Another parameter affecting the receptor binding is
probably the amide carbonyl. In compound 19 that
lacks the carbonyl group we found a decreased binding
affinity. Though this effect is not very strong, the binding
affinity of 19 is decreased by a factor of about 6 in com-
parison to 5b (IC50 2.78 0.63 nM for 5b vs
17.8 1.75 nM for 19). We observed a very profound
change in binding affinity by substituting the thiophene
3. Conclusion
In conclusion, we present here the synthesis of a new ser-
ies of fluorinated pyrazolopyrimidines as potential PET
imaging agents for GABAA receptors. Some of these
compounds displayed excellent binding affinities
in vitro. In addition to these fluorinated reference
compounds we have synthesized suitable precursor com-
pounds. The latter provide a basis for a straightforward
18F-labelling currently under investigation in our
laboratories.
Table 2. Calculated logP and measured logD values (standard deviation given in parentheses)
Compound
logP
logD
Calculateda
Calculatedb
Measuredc
Measuredd
Indiplon
ABX012
ABX016
ABX014
ABX017
ABX021
ABX006
ABX007
ABX008
ABX020
2.57 (0.47)
2.76 (0.47)
2.87 (0.47)
3.32 (0.47)
3.52 (0.47)
4.09 (0.47)
4.46 (0.47)
—
2.68 (0.49)
2.84 (0.49)
2.90 (0.47)
3.35 (0.49)
3.52 (0.47)
4.09 (0.49)
4.46 (0.47)
—
2.26 (0.23)
2.62 (0.26)
2.35 (0.24)
2.48 (0.25)
—
2.23 (0.18)
—
5a
5b
5c
2.38 (0.18)
—
—
5d
5e
2.55 (0.25)
2.64 (0.25)
2.51 (0.25)
2.66 (0.26)
2.34 (0.24)
2.68 (0.41)
2.63 (0.24)
2.64 (0.23)
2.69 (0.25)
2.38 (0.24)
16a
16b
19
25
4.63 (0.47)
2.75 (0.47)
4.74 (0.49)
2.80 (0.49)
a Crippen’s fragmentation.20
b Viswanadhan’s fragmentation.21
c HPLC method 1 (C-18 reversed phase).
d HPLC method 2 (alkyl amide reversed phase).

1188
A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
4. Experimental
radioactivity). The incubation started with the applica-
tion of the membrane solution and was performed at
21 ꢁC for 60 min. The incubation was terminated by ra-
pid filtration through glass–fibre filters (GF/B, What-
man; presoaked in 0.3% polythyleneimine solution at
4 ꢁC for 90 min) on a cell harvester manifold (Brandel).
The filters were washed four times with ice-cold 50 mM
Tris–HCl, pH 7.4, at 4 ꢁC, and filter-bound radioactivity
was measured by liquid scintillation spectrometry.
4.1. Chemical synthesis
Chemicals were purchased from Sigma–Aldrich, Ger-
many, and used without further purification. Indiplon,
Demethylindiplon,
phen-2-yl-methanone (6), 3-N-methylaminoacetophenone
(13), and N-[3-(3-dimethylamino-acryloyl)-phenyl]-N-
methyl-acetamide were synthesized according to litera-
ture procedures.17 Reactions were routinely monitored
by TLC on precoated silica gel plates (ALUGRAM
(5-amino-1H-pyrazol-4-yl)-thio-
The concentration of the test compounds that inhibited
[3H]flunitrazepam binding by 50%, IC50, was determined
by nonlinear regression analyses with 10–12 concentra-
tions of the compound, each in triplicate. Each com-
pound was assayed in at least three independent
experiments and the respective IC50 values were given
as means standard deviation (SD).
sheets SIL G/UV254, Macherey-Nagel, Duren). Flash
¨
chromatography was conducted on silica gel (0.040–
0.063 mm, VWR International) columns. All melting
points were determined in open glass capillaries using
1
a Buchi apparatus and are uncorrected. H NMR spec-
¨
tra were recorded at a Bruker AV500 Ultra Shielded
spectrometer, using the hydrogenated residue of deuter-
ated solvents as internal standards. Chemical shifts are
reported in ppm, downfield from TMS (s, d, t, dd, td
and br for singlet, doublet, triplet, doublet of doublets,
triplet of doublets and broad, respectively). Elemental
analyses were performed on a EL analyser (Elementar
Analysensysteme, Germany) and were within 0.4% of
the theoretical values for C, H, and N. High-resolution
mass spectra were recorded with a Bruker Daltonics 7
Tesla APEX II spectrometer.
4.3. Measurement of logD values
logD values were determined with HPLC methods and
were performed according to the European guideline.22
HPLC system: Agilent Hewlett Packard 1100 with a bin-
ary pump, auto sampler, variable wavelength UV detec-
tor (228, 344; 220, 254 nm). Column for method 1:
Multisorb RP18-7 column (250 · 4 mm, 7 lm, CS Chro-
matography Service, Germany); for method 2: Supelco-
silTM ABZ+Plus (250 · 4 mm, 5 lm, Sigma–Aldrich,
Germany). Mobile phase: Solvent A: 5% acetoni-
trile + 20 mM ammonium acetate; solvent B: 80% aceto-
nitrile + 20 mM ammonium acetate. Gradient 1:
0–5 min (solvent A), 5–40 min (solvent B); Gradient 2:
0–5 min (solvent A), 5–50 min (solvent B), Gradient 3:
0–5 min (solvent A), 5–65 min (solvent B); flow rate
1 mL/min. The sample (2–20 lg/mL) was injected in a
4.2. In vitro binding assays
Adult female Sprague–Dawley rats (8–10 weeks old;
University of Leipzig) were used. Animal care and han-
dling through experimental procedures were in accor-
dance with the national regulations for animal
research. Rats were anaesthetised and sacrificed by
decapitation, their brains were rapidly dissected on ice,
and the cerebella were collected in 30 volumes of ice-
cold 50 mM Tris–HCl, pH 7.4, at 4 ꢁC, homogenised
with a glass–Teflon Potter (1000 rpm), and the crude
membrane fraction was collected by centrifugation at
20,000g at 4 ꢁC for 15 min. The resulting pellet was
washed two times by resuspending in the same volume
of fresh buffer and recentrifugation, and stored at 0.1 g
wet weight/mL fresh buffer at ꢁ25 ꢁC until the day of
the assay. For homogenate binding assays, the frozen
membranes were thawed, recentrifuged, washed another
two times according to the described protocol, and
resuspended at ꢀ0.05 g wet weight/mL 50 mM Tris–
HCl, pH 7.4, at 21 ꢁC (incubation buffer). [3H]Flunitraz-
epam binding was measured in a final volume of 1 mL,
consisting of 100 lL membrane suspension (ꢀ5 mg wet
weight = ꢀ100 lg protein), 100 ll N-methyl-[3H]fluni-
trazepam (specific activity 3600 GBq/mmol; Amersham,
GE Healthcare; final concentration in the assay
ꢀ0.3 nM), 100 lL of drug solution and incubation buf-
fer. Solutions of the test drugs were prepared by diluting
a 10 mM DMSO-stock with incubation buffer, and non-
specific binding of [3H]flunitrazepam was determined in
the presence of 10 lM Zolpidem (TOCRIS; 100 lM
stock in incubation buffer). Specific binding of the radio-
ligand (about 85%) was calculated by subtracting the
nonspecific from the total binding (about 5% of the total
volume of 1–5 lL of solvent
C (25% acetoni-
trile + 20 mM ammonium acetate). All runs were done
in triplicate. A good reproducibility was obtained
(DtR < 0.07 min). Before running test samples, the sys-
tem was validated by running standard samples. The fol-
lowing standard compounds were used for
referencing23,24: acetophenone (logP = 1.7–1.6), aniline
(0.9), anisole (2.11), benzene (2.13), benzonitrile (1.56),
benzyl alcohol (1.1), benzyl chloride (2.13), indole
(2.14), nitrobenzene (1.9), 2-nitrophenol (1.79) and tolu-
ene (2.73) either as single standard or mixture of three
(1: aniline, nitrobenzene, toluene; 2: anisole, benzyl chlo-
ride, toluene; 3: benzyl alcohol, indole, toluene). Analy-
sis was performed by determination of capacity factors
k0 and linear regression of logk0/logP. Linear regression
with n = 32 (number of runs) showed R < 0.94667 with
p < 0.0001 independent from the method and gradient.
4.4. General procedure for the synthesis of the fluoroalkyl
acetophenones (2a–c)
3-Aminoacetophenone (1) (3.95 g, 29.2 mmol) and cal-
cium carbonate (1.32 g, 13.2 mmol) were suspended in
water (100 mL) in an ultrasonic bath. The alkylating
agent (26.3 mmol, toluene-4-sulfonic acid 2-fluoroethyl
ester for 2a, toluene-4-sulfonic acid 3-fluoropropyl ester
for 2b and 1-chloro-4-fluorobutane for 2c) was added,
and the mixture was heated under reflux for 3.5 h. After

A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
1189
cooling to room temperature, aqueous (aq) 10% potas-
sium carbonate (50 mL) was added. The mixture was ex-
tracted with dichloromethane (3· 100 mL). The
combined organic phases were dried over sodium sulfate
and concentrated. The residue was purified by flash
chromatography.
J = 5.8 Hz, J2F = 47.1 Hz, CH2F), 3.87 (t, 2H,
J = 7.3 Hz, NCH2), 2.64 (s, 3H, CH3), 2.03–1.91 (m,
2H, CH2), 1.85 (br s, 3H, CH3).
4.5.3. N-(3-Acetyl-phenyl)-N-(4-fluorobutyl)-acetamide
(3c). Yield: 64%, yellow oil, Rf (dichloromethane/ethyl
acetate 5:1) = 0.24. 1H NMR (CDCl3, 500 MHz):
d = 7.94 (m, 1H, HAR), 7.77 (m, 1H, HAR), 7.56 (m,
1H, HAR), 7.39 (m, 1H, HAR), 4.45 (td, 2H,
J = 5.8 Hz, J2F = 47.0 Hz, CH2F), 3.78 (t, 2H,
J = 7.3 Hz, NCH2), 2.65 (s, 3H, CH3), 1.85 (br s, 3H,
CH3), 1.78–1.61 (m, 4H, CH2).
4.4.1. 1-[3-(2-Fluoroethylamino)-phenyl]-ethanone (2a).
Yield: 42%, yellow oil, Rf (ethyl acetate/dichlorometh-
ane 1:25) = 0.41. 1H NMR (CDCl3, 500 MHz):
d = 7.34–7.26 (m, 2H, HAR), 7.23 (m, 1H, HAR), 6.87–
6.83 (m, 1H, HAR), 4.65 (td, 2H, J = 4.7 Hz,
J2F = 47.2 Hz, CH2F), 3.51 (td, 2H, J = 4.9 Hz,
J2F = 26.7 Hz, NCH2), 2.58 (s, 3H, CH3).
4.5.4. N-(3-Acetyl-phenyl)-N-(3-fluoropropyl)-propiona-
mide (3d). Yield: 39%, yellow oil, Rf (dichlorometh-
ane/ethyl acetate 5:1) = 0.42. 1H NMR (CDCl3,
500 MHz): d = 7.94 (m, 1H, HAR), 7.78 (m, 1H,
HAR), 7.56 (m, 1H, HAR), 7.39 (m, 1H, HAR),
4.51 (td, 2H, J = 5.8 Hz, J2F = 47.0 Hz, CH2F), 3.87
(t, 2H, J = 7.3 Hz, NCH2), 2.65 (s, 3H, CH3),
2.10–1.92 (m, 4H, CH2), 1.06 (t, J = 7.3 Hz, 3H,
CH3).
4.4.2. 1-[3-(3-Fluoropropylamino)-phenyl]-ethanone (2b).
Yield: 51%, yellow oil, Rf (dichloromethane) = 0.36. H
1
NMR (CDCl3, 500 MHz): d = 7.30–7.24 (m, 2H, HAR),
7.20 (m, 1H, HAR), 6.83–6.80 (m, 1H, HAR), 4.60 (td,
2H, J = 5.6 Hz, J2F = 47.2 Hz, CH2F), 3.37 (t, 2H,
J = 6.7 Hz, NCH2), 2.58 (s, 3H, CH3), 2.09–1.97 (m,
2H, CH2).
4.4.3. 1-[3-(4-Fluorobutylamino)-phenyl]-ethanone (2c).
Yield: 13%, yellow oil, Rf (ethyl acetate/dichlorometh-
ane 1:25) = 0.33. 1H NMR (CDCl3, 500 MHz):
d = 7.32–7.22 (m, 2H, HAR), 7.29 (m, 1H, HAR), 6.82–
6.78 (m, 1H, HAR), 4.51 (td, 2H, J = 5.8 Hz,
J2F = 47.3 Hz, CH2F), 3.24 (t, 2H, J = 6.9 Hz, NCH2),
2.58 (s, 3H, CH3), 1.89–1.74 (m, 4H, CH2).
4.5.5. N-(3-Acetyl-phenyl)-N-(3-fluoropropyl)-isobutyra-
mide (3e). Yield: 94%, yellow oil, Rf (dichloromethane/
ethyl acetate 5:1) = 0.44. H NMR (CDCl3, 500 MHz):
d = 7.95 (m, 1H, HAR), 7.79 (m, 1H, HAR), 7.56 (m,
1H, HAR), 7.40 (m, 1H, HAR), 4.51 (td, 2H,
J = 5.9 Hz, J2F = 47.2 Hz, CH2F), 3.84 (t, 2H,
J = 7.3 Hz, NCH2), 2.64 (s, 3H, CH3), 2.44–2.34 (m,
1H, CH), 2.03–1.91 (m, 2H, CH2), 1.03 (d, 6H, CH3).
1
4.5. General procedure for the synthesis of the fluoroalkyl
acetophenones (3a–e)
4.6. General procedure for the synthesis of the enaminones
(4a–e)
A solution of 2a–c (14.8 mmol) and DMAP (183 mg,
1.5 mmol) in dry pyridine (75 mL) was cooled in an
ice bath and the acylating agent was added
(22.2 mmol, acetic anhydride for 3a–c, propionic anhy-
dride for 3d, isobutyryl chloride for 3e). After com-
plete addition, the ice bath was removed and the
mixture was stirred for 16 h at room temperature.
The mixture was poured onto ice/water and was ad-
justed to pH 1 with hydrochloric acid and extracted
with dichloromethane. The combined organic phases
were washed with aq sodium bicarbonate solution,
dried over sodium sulfate and concentrated. The resi-
due was purified by flash chromatography (dichloro-
methane/ethyl acetate 5:1).
A solution of 3a–e (8.5 mmol) in N,N-dimethylformam-
ide dimethyl acetal (DMFDMA, 30 mL) was heated at
120 ꢁC overnight. After evaporation of the solvent, the
residue was purified by flash chromatography (ethyl ace-
tate/methanol 9:1).
4.6.1. N-[3-(3-Dimethylamino-acryloyl)-phenyl]-N-(2-flu-
oroethyl)-acetamide (4a). Yield: 92%, yellow solid, mp
89.6–90.4 ꢁC, Rf (ethyl acetate/methanol 9:1) = 0.27.
1H NMR (CDCl3, 500 MHz): d = 7.90–7.82 (m, 2H,
HAR), 7.76 (br s, 1H, HAR), 7.47 (t, 1H, J = 7.8 Hz,
HAR), 7.33 (m, 1H, HAR), 5.67 (d, 1H, J = 12.3 Hz,
CH), 4.62 (td, 2H, J = 4.9 Hz, J2F = 47.4 Hz, CH2F),
4.01 (td, 2H, J = 5.0 Hz, J3F = 25.6 Hz, CH2F), 3.18
(br s, 3H, NCH3), 2.96 (br s, 3H, NCH3), 1.89 (s, 3H,
CH3).
4.5.1. N-(3-Acetyl-phenyl)-N-(2-fluoroethyl)-acetamide
(3a). Yield: 45%, yellow oil, Rf (dichloromethane/ethyl
acetate 10:1) = 0.21. 1H NMR (CDCl3, 500 MHz):
d = 7.95 (m, 1H, HAR), 7.84 (m, 1H, HAR), 7.55 (m,
1H, HAR), 7.47 (m, 1H, HAR), 4.64 (td, 2H,
J = 4.8 Hz, J2F = 47.4 Hz, CH2F), 4.00 (t, 2H,
J = 4.8 Hz, J3F = 26.3 Hz, NCH2), 2.63 (s, 3H, CH3),
1.89 (br s, 3H, CH3).
4.6.2. N-[3-(3-Dimethylamino-acryloyl)-phenyl]-N-(3-flu-
oropropyl)-acetamide (4b). Yield: 65%, yellow solid, mp
1
119–120 ꢁC, Rf (ethyl acetate/methanol 9:1) = 0.37. H
NMR (CDCl3, 500 MHz): d = 7.90–7.82 (m, 2H,
HAR), 7.72 (br s, 1H, HAR), 7.47 (t, 1H, J = 7.8 Hz,
HAR), 7.26 (m, 1H, HAR), 5.67 (d, 1H, J = 12.3 Hz,
CH), 4.50 (td, 2H, J = 5.9 Hz, J2F = 47.2 Hz, CH2F),
3.86 (m, 2H, NCH2), 3.19 (br s, 3H, NCH3), 2.96 (br
s, 3H, NCH3), 2.03–1.91 (m, 2H, CH2), 1.86 (s, 3H,
CH3). HRMS m/z calculated for C16H22FN2O2 (M+H)
293.1660, found 293.1661.
4.5.2. N-(3-Acetyl-phenyl)-N-(3-fluoropropyl)-acetamide
(3b). Yield: 57%, yellow oil, Rf (dichloromethane/ethyl
acetate 5:1) = 0.31. 1H NMR (CDCl3, 500 MHz):
d = 7.94 (m, 1H, HAR), 7.79 (m, 1H, HAR), 7.56 (m,
1H, HAR), 7.40 (m, 1H, HAR), 4.50 (td, 2H,

1190
A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
4.6.3. N-[3-(3-Dimethylamino-acryloyl)-phenyl]-N-(4-flu-
orobutyl)-acetamide (4c). Yield: 81%, yellow oil, Rf
(ethyl acetate/methanol 9:1) = 0.25. H NMR (CDCl3,
61.75, H: 4.20, N: 13.72, found C: 61.45, H: 4.04, N:
13.50.
1
500 MHz): d = 7.89–7.83 (m, 2H, HAR), 7.70 (br s, 1H,
HAR), 7.47 (t, 1H, J = 7.8 Hz, HAR), 7.24 (m, 1H,
HAR), 5.68 (d, 1H, J = 12.3 Hz, CH), 4.43 (td, 2H,
J = 5.8 Hz, J2F = 47.1 Hz, CH2F), 3.77 (m, 2H,
NCH2), 3.18 (br s, 3H, NCH3), 2.96 (br s, 3H, NCH3),
1.84 (s, 3H, CH3), 1.77–1.60 (m, 4H, CH2). HRMS m/z
calculated for C17H24FN2O2 (M+H) 307.1816, found
307.1818.
4.7.2. N-(3-Fluoropropyl)-N-[3-[3-(thiophene-2-carbonyl)-
pyrazolo[1,5-a]pyrimidin-7-yl]-phenyl]-acetamide
(5b).
Yield: 46%, yellow solid, mp 194–195 ꢁC, Rf (ethyl ace-
1
tate) = 0.30. H NMR (CDCl3, 500 MHz): d = 8.84 (d,
1H, J3H = 4.4 Hz, HAR), 8.72 (s, 1H, HAR), 8.1 (m, 1H,
HAR), 8.00 (m, 2H, HAR), 7.73–7.67 (m, 2H, HAR), 7.47
(m, 1H, HAR), 7.21 (m, 1H, HAR), 7.17 (d, 1H,
J3H = 4.4 Hz, HAR), 4.53 (dt, 2H, J2F = 47 Hz,
J3H = 5.8 Hz, CH2F), 3.92 (t, 2H, J3H = 7.3 Hz, N–
CH2), 2.09–1.95 (m, 5H, CH2 and CH3). 19F NMR
(CDCl3, 470 MHz): d = ꢁ219.99 ppm. Calculated C:
62.54, H: 4.53, N: 13.26, found C: 62.36, H: 4.52, N: 13.14.
4.6.4. N-[3-(3-Dimethylamino-acryloyl)-phenyl]-N-(3-flu-
oropropyl)-propionamide (4d). Yield: 69%, yellow solid,
mp
95.3–96.0 ꢁC,
Rf
(ethyl
acetate/methanol
9:1) = 0.46. 1H NMR (CDCl3, 500 MHz): d = 7.89–
7.82 (m, 2 H, HAR), 7.70 (br s, 1H, HAR), 7.47 (t, 1H,
J = 7.8 Hz, HAR), 7.25 (m, 1H, HAR), 5.67 (d, 1H,
J = 12.3 Hz, CH), 4.50 (td, 2H, J = 5.9 Hz,
J2F = 47.2 Hz, CH2F), 3.86 (m, 2H, NCH2), 3.19 (br s,
3H, NCH3), 2.96 (br s, 3H, NCH3), 2.07–1.92 (m, 4H,
CH2), 1.04 (t, 3H, J = 7.4 Hz, CH3). HRMS m/z calcu-
lated for C17H24FN2O2 (M+H) 307.1816, found
307.1818.
4.7.3. N-(4-Fluorobutyl)-N-[3-[3-(thiophene-2-carbonyl)-
pyrazolo[1,5-a]pyrimidin-7-yl]-phenyl]-acetamide
(5c).
Yield: 47%, yellow solid, mp 151–152 ꢁC, Rf (ethyl ace-
1
tate) = 0.26. H NMR (CDCl3, 500 MHz): d = 8.87 (d,
1H, J3H = 4.3 Hz, HAR), 8.73 (s, 1H, HAR), 8.11 (m, 1H,
HAR), 8.03–7.99 (m, 2H, HAR), 7.73–7.66 (m, 2H, HAR),
7.45 (m, 1H, HAR), 7.23 (m, 1H, HAR), 7.18 (d, 1H,
J3H = 4.3 Hz, HAR), 4.47 (dt, 2H, J2F = 41.5 Hz,
J3H = 5.7 Hz, CH2F), 3.84 (t, 2H, J3H = 7.1 Hz, N–CH2),
1.97 (s, 3H, CH3) 1.81–1.68 (m, 4H, CH2). 19F NMR
(CDCl3, 470 MHz): d = ꢁ218.57 ppm. Calculated C:
63.29, H: 4.85, N: 12.84, found C: 63.16, H: 4.91, N: 12.67.
4.6.5. N-[3-(3-Dimethylamino-acryloyl)-phenyl]-N-(3-flu-
oropropyl)-isobutyramide (4e). Yield: 84%, yellow solid,
mp 102.4–106.1 ꢁC, Rf (ethyl acetate/methanol
9:1) = 0.48. 1H NMR (CDCl3, 500 MHz): d = 7.89–
7.84 (m, 2H, HAR), 7.71 (br s, 1H, HAR), 7.47 (t, 1H,
J = 7.8 Hz, HAR), 7.26 (m, 1H, HAR), 5.68 (d, 1H,
J = 12.3 Hz, CH), 4.50 (td, 2H, J = 5.9 Hz,
J2F = 47.1 Hz, CH2F), 3.84 (m, 2H, NCH2), 3.19 (br s,
3H, NCH3), 2.97 (br s, 3H, NCH3), 2.46 (m, 1H, CH),
2.04-1.92 (m, 2H, CH2), 1.03(d, 6H, J = 6.7 Hz, CH3).
HRMS m/z calculated for C18H25FN2O2 (M+H)
321.1973, found 321.1972.
4.7.4. N-(3-Fluoropropyl)-N-[3-[3-(thiophene-2-carbonyl)-
pyrazolo[1,5-a]pyrimidin-7-yl]-phenyl]-propionamide (5d).
Yield: 55%, yellow solid, mp 171.0–171.8 ꢁC, Rf (ethyl
1
acetate/dichloromethane 1:1) = 0.50. H NMR (CDCl3,
500 MHz): d = 8.85 (d, 1H, J3H = 4.4 Hz, HAR), 8.73
(s, 1H, HAR), 8.11 (m, 1H, HAR), 8.04–7.97 (m, 2H,
HAR), 7.73–7.67 (m, 2H, HAR), 7.45 (m, 1H, HAR),
7.22 (m, 1H, HAR), 7.17 (d, 1H, J3H = 4.4 Hz, HAR),
4.54 (td, 2H, J2F = 47.1 Hz, J3H = 5.8 Hz, CH2F), 3.92
(t, 2H, J3H = 7.1Hz, N–CH2), 2.18 (m, 2H, CH2), 2.14
(m, 2H, CH2) 1.11 (m, 3H, CH3). 19F NMR (CDCl3,
470 MHz): d = ꢁ219.89 ppm. Calculated C: 63.29, H:
4.85, N: 12.84, found C: 63.02, H: 4.88, N: 12.66.
4.7. General procedure for the synthesis of the fluoroalkyl
pyrazolopyrimidines (5a–e)
A suspension of 4a–e (2.74 mmol) and (5-amino-1H-
pyrazol-4-yl)-thiophen-2-yl-methanone (6) (2.74 mmol)
in acetic acid (20 mL) was heated to 65 ꢁC for 1.5 h.
After cooling to room temperature the mixture was
poured onto ice/water and adjusted to pH 8 with sodium
bicarbonate before extraction with dichloromethane.
The combined organic phases were dried over sodium
sulfate and concentrated. The residue was purified by
flash chromatography.
4.7.5. N-(3-Fluoropropyl)-N-[3-[3-(thiophene-2-carbonyl)-
pyrazolo[1,5-a]pyrimidin-7-yl]-phenyl]-isobutyramide (5e).
Yield: 41%, yellow solid, mp 155.4–157.0 ꢁC, Rf (ethyl
1
acetate/dichloromethane 1:2) = 0.41. H NMR (CDCl3,
500 MHz): d = 8.86 (d, 1H, J3H = 4.4 Hz, HAR), 8.72
(s, 1H, HAR), 8.11 (m, 1H, HAR), 8.05–7.97 (m, 2H,
HAR), 7.74–7.68 (m, 2H, HAR), 7.46 (m, 1H, HAR),
7.23 (m, 1H, HAR), 7.17 (d, 1H, J3H = 4.4 Hz, HAR),
4.55 (td, 2H, J2F = 47.2 Hz, J3H = 5.8 Hz, CH2F), 3.90
(t, 2 H, J3H = 7.2 Hz, N–CH2), 2.59 (m, 1H, CH),
2.10–1.98 (m, 2H, CH2), 1.10 (d, 6H, J3H = 6.6 Hz,
CH3). 19F NMR (CDCl3, 470 MHz): d = ꢁ219.70 ppm.
Calculated C: 63.98, H: 5.15, N: 12.44, found C: 63.88,
H: 5.81, N: 12.31.
4.7.1. N-(2-Fluoroethyl)-N-[3-[3-(thiophene-2-carbonyl)-
pyrazolo[1,5-a]pyrimidin-7-yl]-phenyl]-acetamide (5a).
Yield: 44%, yellow solid, mp 179–180 ꢁC, Rf (ethyl ace-
tate/dichloromethane/methanol 10:10:1) = 0.39. 1H
NMR (CDCl3, 500 MHz): d = 8.84 (d, 1H,
J3H = 4.4 Hz, HAR), 8.72 (s, 1H, HAR), 8.1 (m, 1H,
HAR), 8.03 (m, 2H, HAR), 7.73–7.66 (m, 2H, HAR),
7.52 (m, 1H, HAR), 7.21 (m, 1H, HAR), 7.16 (d, 1H,
J3H = 4.4 Hz, HAR), 4.69 (td, 2H, J2F = 47 Hz,
J3H = 4.7 Hz, CH2F), 4.05 (td, 1H, J3F = 26 Hz,
J3H = 4.8 Hz, N–CH2), 2.02 (s, 3H, CH3). 19F NMR
(CDCl3, 470 MHz): d = ꢁ222.24 ppm. Calculated C:
4.8. General procedure for the synthesis of the alcohols
(7b,c)
3-Aminoacetophenone (1) (24.33 g, 180 mmol) and cal-
cium carbonate (8.1 g, 81 mmol) were suspended in

A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
1191
water (300 mL) in an ultrasonic bath under inert gas
atmosphere. The alkylating agent (162 mmol, 3-bro-
mo-1-propanol for 7b, 4-chloro-1-butanol for 7c) was
added and the mixture was heated under reflux for
2 h. After cooling to room temperature 10% aq
potassium carbonate (250 mL) was added before extrac-
tion with dichloromethane. The combined organic
phases were dried over sodium sulfate and concentrated.
The residue was purified by flash chromatography.
combined organic phases were dried over sodium sul-
fate, the solvent was evaporated and the product was
dried in vacuum.
4.10.1. N-(3-Acetyl-phenyl)-N-(3-hydroxy-propyl)-acet-
amide (9b). Yield: 97%, white solid, mp 51.4-53.2 ꢁC,
Rf (ethyl acetate) = 0.16. H NMR (CDCl3, 500 MHz):
1
d = 7.96 (m, 1H, HAR), 7.77 (m, 1H, HAR), 7.57 (m,
1H, HAR), 7.37 (m, 1H, HAR), 3.90 (m, 2 H, CH2),
3.69–3.64 (m, 3H, CH2,OH), 2.64 (s, 3H, CH3), 1.88
(s, 3H, CH3), 1.67 (m, 2H, CH2).
4.8.1.
1-[3-(3-Hydroxy-propylamino)-phenyl]-ethanone
(7b). Yield: 46%, yellow oil, Rf (ethyl acetate/dichloro-
methane/triethylamine 50:50:1) = 0.25. 1H NMR
(CDCl3, 500 MHz): d = 7.30–7.20 (m, 3H, HAR), 6.83
(m, 1H, HAR), 3.83 (t, J = 5.9 Hz, 2H, CH2), 3.33 (t,
J = 6.5 Hz, 2H, CH2), 2.57 (s, 3H, CH3), 1.91 (m, 2H,
CH2).
4.10.2. N-(3-Acetyl-phenyl)-N-(4-hydroxy-butyl)-acetam-
ide (9c). Yield: 97%, yellow oil, Rf (ethyl acetate) = 0.13.
1H NMR (CDCl3, 500 MHz): d = 7.94 (m, 2H, HAR),
7.77 (m,1H, HAR), 7.55 (m, 1H, HAR), 7.39 (m, 1H,
HAR), 3.77 (m, 2H, CH2), 3.66 (m, 2H, CH2), 2.64 (s,
3H, CH3), 1.84 (s, 3H, CH3), 1.65–1.55 (m, 4H, CH2).
4.8.2.
1-[3-(4-Hydroxy-butylamino)-phenyl]-ethanone
(7c). Yield: 7.4%, yellow oil, Rf (ethyl acetate) = 0.38.
1H NMR (CDCl3, 500 MHz): d = 7.29–7.18 (m, 3H,
HAR), 6.80 (m, 1H, HAR), 3.72 (m, 2H, CH2), 3.21 (m,
2H, CH2), 2.57 (s, 3H, CH3), 1.78–1.66 (m, 4H, CH2).
4.11. General procedure for the synthesis of the enami-
nones (10b,c)
A solution of 9b,c (41 mmol) in DMFDMA (70 mL)
was heated at 120 ꢁC overnight. After evaporation of
the solvent, the residue was purified by flash chromatog-
raphy (ethyl acetate/methanol 9:1–8:2).
4.9. General procedure for the synthesis of the acetamides
(8b,c)
To a solution of 7b,c (41.4 mmol) and DMAP (1.01 g,
8.3 mmol) in dry pyridine (40 mL), acetic anhydride
(12.8 g; 11.8 mL, 124 mmol) was added dropwise and
the mixture was stirred at room temperature for 16 h.
The mixture was added to ice/water and adjusted to
pH 1–2 with 3 N HCl before extraction with dichloro-
methane. The combined organic phases were washed
with 1 N HCl, saturated aq sodium bicarbonate and
brine. The solution was dried over sodium sulfate, the
solvent was evaporated and the product was dried in
vacuum.
4.11.1. N-[3-(3-Dimethylamino-acryloyl)-phenyl]-N-(3-
hydroxy-propyl)-acetamide (10b). Yield: 74%, yellow so-
lid, mp 143.4–144.7 ꢁC, Rf (ethyl acetate/methanol
9:1) = 0.16. 1H NMR (CDCl3, 500 MHz): d = 7.90–
7.83 (m, 2H, HAR), 7.70 (m, 1H, HAR), 7.48 (m, 1H,
HAR), 7.22 (m, 1H, HAR), 5.67 (d, 1H, J3H = 12.2 Hz,
CH), 3.92-3.73 (m, 3H, CH2, OH), 3.67 (m, 2H, CH2),
3.19 (br s, 3H, NCH3), 2.96 (br s, 3H, NCH3), 1.89 (s,
3H, CH3), 1.67 (m, 2H, CH2).
4.11.2. N-[3-(3-Dimethylamino-acryloyl)-phenyl]-N-(4-
hydroxy-butyl)-acetamide (10c). Yield: 71%, yellow oil,
Rf (ethyl acetate/methanol 9:1) = 0.12. 1H NMR
(CDCl3, 500 MHz): d = 7.86 (m, 2H, HAR), 7.71
(m,1H, HAR), 7.46 (m, 1H, HAR), 7.25 (m, 1H, HAR),
5.68 (d, 1H, J3H = 12.3 Hz, HAR), 3.77 (m, 2H, CH2),
3.65 (m, 2H, CH2), 3.19 (br s, 3H, NCH3), 2.96 (br s,
3H, NCH3), 1.96 (m, 3H, CH3), 1.73–1.58 (m, 4H, CH2).
4.9.1. Acetic acid 3-[acetyl-(3-acetyl-phenyl)-amino]-pro-
pyl ester (8b). Yield: 100%, yellow oil, Rf (ethyl acetate/
dichloromethane 1:1) = 0.33. 1H NMR (CDCl3,
500 MHz): d = 7.95 (m, 1H, HAR), 7.79 (m, 1H, HAR),
7.55 (m, 1H, HAR), 7.40 (m, 1H, HAR), 4.10 (t,
J = 6.5 Hz, 2H, CH2), 3.82 (m, 2H, CH2), 2.64 (s, 3H,
CH3), 2.01 (s, 3H, CH3), 1.90–1.83 (m, 5H, CH2/3).
4.12. General procedure for the synthesis of the x-
hydroxyalkyl derivatives of Indiplon (11b,c)
4.9.2. Acetic acid 3-[acetyl-(3-acetyl-phenyl)-amino]-butyl
ester (8c). Yield: 96%, yellow oil, Rf (ethyl ace-
tate) = 0.44. H NMR (CDCl3, 500 MHz): d = 7.94 (m,
2H, HAR), 7.77 (m,1H, HAR), 7.56 (m, 1H, HAR), 7.38
(m, 1H, HAR), 4.04 (m, 2H, CH2), 3.75 (m, 2H, CH2),
2.64 (s, 3H, CH3), 2.02 (s, 3H, CH3), 1.84 (br s, 3H,
CH3), 1.68–1.53 (m, 4H, CH2).
1
A suspension of 10b,c (8.5 mmol) and (5-amino-1H-pyr-
azol-4-yl)-thiophen-2-yl-methanone (6) 1.65 g (8.5
mmol) in acetic acid (50 mL) was heated to 65 ꢁC for
1.5 h. After cooling to room temperature the mixture
was poured onto ice/water and adjusted to pH 8 with so-
dium bicarbonate before extraction with dichlorometh-
ane. The combined organic phases were dried over
sodium sulfate and concentrated. The residue was puri-
fied by flash chromatography.
4.10. General procedure for the synthesis of the alcohols
(9b,c) (Deacetylation)
To a solution of 8b,c (42 mmol) in methanol (100 mL),
an aq 4 M potassium carbonate solution was added.
The mixture was stirred for 15 min at room temperature
and adjusted to pH 8 by adding saturated aq ammonium
chloride before extracting with dichloromethane. The
4.12.1. N-(3-Hydroxy-propyl)-N-[3-[3-(thiophene-2-car-
bonyl)-pyrazolo[1,5-a]-pyrimidin-7-yl]-phenyl]-acetam-
ide (11b). Yield: 58%, yellow solid, mp 210–211 ꢁC, Rf
1
(ethyl acetate/methanol 9:1) = 0.32. H NMR (CDCl3,

1192
A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
500 MHz): d = 8.85 (d, 1H, J3H = 4.4 Hz, HAR), 8.73 (s,
1H, HAR), 8.1 (m, 1H, HAR), 8.00 (m, 2H, HAR), 7.74–
7.68 (m, 2H, HAR), 7.44 (m, 1H, HAR), 7.22 (m, 1H,
HAR), 7.17 (d, 1H, J3H = 4.4 Hz, HAR), 3.96 (t, 2H,
J3H = 6.2 Hz, CH2), 3.73–3.59 (m, 3H, CH2, OH), 2.01
(s, 3H, CH3), 1.75 (m, 2H, CH2). HRMS m/z calculated
for C22H20N4O3S (M+H) 421.1329, found 421.1335.
CH3), 1.76–1.61 (m, 4H, CH2). Calculated C: 61.21,
H: 4.79, N: 9.52, found C: 60.52, H: 4.82, N: 9.29.
4.14. General procedure for the synthesis of the benzam-
ides (14a,b)
To a solution of 3-N-methylaminoacetophenone (13)
(13.4 mmol) in DMF (50 mL), the appropriate benzoyl
chloride (13.4 mmol) and triethylamine (14.7 mmol)
were added and the mixture was stirred at room temper-
ature overnight. The solution was poured onto ice/water
and extracted with ethyl acetate (3· 30 mL). The com-
bined organic phases were dried over sodium sulfate
and concentrated. The residue was purified by flash
chromatography (light petroleum/ethyl acetate: 4:1).
4.12.2. N-(4-Hydroxy-butyl)-N-[3-[3-(thiophene-2-car-
bonyl)-pyrazolo[1,5-a]-pyrimidin-7-yl]-phenyl]-acetamide
(11c). Yield: 56%, yellow solid, mp 178–180 ꢁC, Rf (ethyl
acetate/methanol 9:1) = 0.29. 1H NMR (CDCl3,
500 MHz): d = 8.85 (d, 1H, J3H = 4.3 Hz, HAR), 8.73
(s, 1H, HAR), 8.10 (m, 1H, HAR), 8.03–7.97 (m, 2 H,
HAR), 7.74–7.67 (m, 2H, HAR), 7.45 (m, 1H, HAR),
7.22 (m, 1H, HAR), 7.18 (d, 1H, J3H = 4.3 Hz, HAR),
3.84 (t, 2H, J3H = 7.3 Hz, CH2), 3.69 (m, 2H, CH2),
1.85 (m, 3H, CH3), 1.65–1.54 (m, 4H, CH2). HRMS
m/z calculated for C23H22N4O3S (M+H) 435.1485,
found 435.1486.
4.14.1. N-(3-Acetylphenyl)-N-methyl-4-fluorobenzamide
(14a). Yield: 96%, yellow solid, mp 64.8–67.1 ꢁC, Rf
(light petroleum/ethyl acetate 1:1) = 0.43. 1H NMR
(CDCl3, 500 MHz): d = 7.73 (dt, 1H, J = 7.7, 1.2 Hz),
7.67 (t, 1H, J = 1.9 Hz), 7.35–7.25 (m, 3H), 7.18 (m,
1H), 6.86(m, 2H), 3.51 (s, 3H), 2.51 (s, 3H).
4.13. General procedure for the synthesis of the x-tosyloxy
precursors (12b,c)
4.14.2. N-(3-Acetylphenyl)-N-methyl-4-nitrobenzamide
(14b). Yield: 95%, off-white solid, mp 117–118 ꢁC, Rf
(light petroleum/ethyl acetate 2:1) = 0.17. 1H NMR
(CDCl3, 500 MHz): d = 8.04 (d, 1H, J = 7.8 Hz), 7.76
(m, 1H), 7.70 (m, 1H), 7.45 (d, 1H, J = 8.6 Hz), 7.35 (t,
1H, J = 7.8 Hz), 7.19 (m, 1H), 3.54 (s, 1H), 2.53 (s, 1H).
A solution of 11b,c (2.4 mmol) and pyridine (0.57 mL,
7.1 mmol) in dry dichloromethane (100 mL) was cooled
in an ice bath under argon atmosphere. Toluene-4-sul-
fonic acid anhydride (1.0 g, 3.1 mmol) was added and
stirring was continued for 30 min. The cooling bath
was removed and stirring was continued for 45 min.
The mixture was successively washed with 1 N hydro-
chloric acid, aq sodium bicarbonate and brine. The or-
ganic phase was dried over sodium sulfate and
concentrated (bath temperature <30 ꢁC). The residue
was purified by flash chromatography (ethyl acetate).
After evaporation of the solvent the product was washed
with diethyl ether and pentane, and dried in vacuum.
Compounds 12b,c are sensitive to heat, handle below
25 ꢁC and store at ꢁ20 ꢁC.
HRMS m/z calculated for C16H14N2O4 (M+H)
299.1026, found 299.1029.
4.15. General procedure for the synthesis of the enami-
nones (15a,b)
A solution of 14a,b (4 mmol) in DMFDMA (5 mL) was
heated at 120 ꢁC overnight. After evaporation of the sol-
vent, the residue was purified by flash chromatography
(ethyl acetate/methanol 9:1).
4.13.1. Toluene-4-sulfonic acid 3-(acetyl-[3-[3-(thiophene-
2-carbonyl)-pyrazolo[1,5-a]-pyrimidin-7-yl]-phenyl]-amino)-
propyl ester (12b). Yield: 65%, yellow solid, mp 123–
127 ꢁC (dec.) Rf (ethyl acetate) = 0.26. 1H NMR
(CDCl3, 500 MHz): d = 8.86 (d, 1H, J3H = 4.4 Hz,
HAR), 8.73 (s, 1H, HAR), 8.12 (m, 1H, HAR), 8.06–7.96
(m, 2 H, HAR), 7.76–7.66 (m, 4H, HAR), 7.42 (m, 1H,
HAR), 7.32 (d, 1H, 1H, J3H = 8.1 Hz, HAR), 7.22 (m,
2H, HAR), 4.11 (t, 2H, J3H = 6.3 Hz, CH2), 3.83 (m,
2H, CH2), 2.43 (s, 3H, CH3), 2.05–1.92 (m, 5H, CH2,
CH3). Calculated C: 60.61, H: 4.56, N: 9.75, found C:
60.28, H: 4.56, N: 9.74.
4.15.1. (E)-N-(3-(3-(Dimethylamino)acryloyl)phenyl)-N-
methyl-4-fluoro-benzamide (15a). Yield: 80%, yellow so-
lid, mp 105–121 ꢁC, Rf (ethyl acetate/methanol
1
9:1) = 0.43. H NMR (CDCl3, 500 MHz) d = 7.78 (d,
1H, J = 12.3 Hz), 7.66 (m, 1H), 7.61 (t, 1H,
J = 1.8 Hz), 7.31 (m, 2H), 7.25 (t, 1H, J = 7.8 Hz),
7.05 (s, 1H), 6.84 (s, 2H), 5.48 (d, 1H, J = 12.5), 3.52
(s, 1H), 3.16 (s, 3H), 2.91 (s, 3H).
4.15.2. (E)-N-(3-(3-(Dimethylamino)acryloyl)phenyl)-N-
methyl-4-nitrobenzamide (15b). Yield: 89%, brownish
oil, Rf (ethyl acetate/methanol 9:1) = 0.44. 1H NMR
(CDCl3, 500 MHz) d = 8.02 (d, 2H, J = 8.6 Hz), 7.79 (d,
1H, J = 12.2 Hz), 7.68–7.65 (m, 2H), 7.46 (d, 1H,
J = 8.7 Hz), 7.26 (m, 1H), 7.05 (d, 1H, J = 7.2 Hz), 5.50
(d, 1H, J = 12.2 Hz), 3.55 (s, 3H), 3.17(s, 3H), 2.92(s, 3H).
4.13.2. Toluene-4-sulfonic acid 4-(acetyl-[3-[3-(thiophene-
2-carbonyl)-pyrazolo[1,5-a]-pyrimidin-7-yl]-phenyl]-amino)-
butyl ester (12c). Yield: 55%, yellow solid, mp 83–85 ꢁC
(dec.), Rf (ethyl acetate) = 0.28. 1H NMR (CDCl3,
500 MHz): d = 8.86 (d, 1H, J3H = 4.4 Hz, HAR), 8.74
(s, 1H, HAR), 8.12 (m, 1H, HAR), 8.04–7.95 (m, 2 H,
HAR), 7.76–7.67 (m, 4H, HAR), 7.42 (m, 1H, HAR),
7.31 (d, 1H, 1H, J3H = 8.1 Hz HAR), 7.22 (m, 2H,
HAR), 4.04 (t, 2H, J3H = 5.9 Hz, CH2), 3.78 (t, 2H,
J3H = 6.9 Hz, CH2), 2.43 (s, 3H, CH3), 1.95 (m, 3H,
4.16. General procedure for the synthesis of the pyrazol-
opyrimidines (16a,b)
A suspension of 15a,b (5 mmol) and (5-amino-1H-pyra-
zol-4-yl)-thiophen-2-yl-methanone (6) (5 mmol) in acetic

A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
1193
acid (50 mL) was heated at 65 ꢁC for 1.5 h. After cooling
to room temperature the mixture was poured onto ice/
water and adjusted to pH 8 with sodium bicarbonate be-
fore extraction with dichloromethane. The combined or-
ganic phases were dried over sodium sulfate and
concentrated. The residue was purified by flash chroma-
tography (ethyl acetate/methanol 9:1).
1H, HAR), 8.12 (m, 1H, HAR), 7.71 (m, 1H, HAR),
7.45 (m, 1H, HAR), 7.35 (m, 1H, HAR), 7.27 (m, 1H,
HAR), 7.22 (m, 1H, HAR), 7.13 (m, 1H, HAR), 6.96 (m,
1H, HAR), 4.67 (td, 2H, J2F = 47.1 Hz, J3H = 5.1 Hz,
CH2F), 3.74 (td, 1 H, J3F = 24.6 Hz, J3H = 5.1 Hz, N–
CH2), 3.11 (s, 3H, CH3). Calculated C: 63.14, H: 4.50,
N: 1214.73, found C: 62.90, H: 4.45, N: 14.34.
4.16.1. 4-Fluoro-N-methyl-N-(3-(3-(thiophene-2-carbonyl)
pyrazolo[1,5-a]pyrimidin-7-yl)phenyl)benzamide (16a).
Yield: 78%, yellow solid, mp 170.7–171.8 ꢁC, Rf (ethyl
4.16.6. 3-(Dimethylamino)-1-(4-fluorophenyl)prop-2-en-1-
one (21). A solution of 4-fluoroacetophenone (20) (1.5 g,
10.8 mmol) in DMFDMA (10 mL) was heated at 120 ꢁC
overnight. After evaporation of the solvent, the residue
was purified by flash chromatography (ethyl acetate).
Yield: 63%, yellow solid, mp 77.5–79.3 ꢁC, Rf (ethyl ace-
1
acetate) = 0.30. H NMR (CDCl3, 500 MHz): d = 8.77
(d, 1H, J = 4.4 Hz), 8.65 (s, 1H), 8.07 (dd, 1H, J = 3.7,
1.0 Hz), 7.80 (m, 2H), 7.71 (dd, 1H, J = 4.9, 1.1 Hz),
7.50 (t, 1H, J = 7.9 Hz), 7.39 (m, 2H), 7.30 (m, 1H),
7.20 (dd, 1H, J = 4.9, 3.8 Hz), 6.92 (m, 3H), 3.57 (s, 3H).
1
tate) = 0.28. H NMR (CDCl3, 500 MHz): d = 7.91 (m,
2H), 7.80 (d, 1H, J = 12.3 Hz), 7.07 (m, 2H), 5.67 (d,
1H, J = 12.3), 3.15 (s, 3H), 2.93 (s, 3H).
Calculated C: 65.78, H: 3.75, N: 12.27, found C: 65.51,
H: 3.70, N: 11.98.
4.16.7. 5-(4-Fluorophenyl)isoxazole (22). A solution of 21
(1.17 g, 6.04 mmol) and hydroxylamine hydrochloride
(420 mg, 6.04 mmol) in methanol (10 mL) was heated
at 65 ꢁC for 2.5 h. After cooling to room temperature
the mixture was diluted with water (10 mL) and ex-
tracted with dichloromethane (3· 10 mL). The com-
bined organic phases were dried over sodium sulfate
and concentrated. Yield: 97%, white solid, mp 53.4–
54.2 ꢁC, Rf (ethyl acetate) = 0.86. 1H NMR (CDCl3,
500 MHz): d = 8.28 (d, 1H, J = 1.8 Hz), 7.79 (m, 2H),
7.17 (m, 2H), 6.47 (d, 1H, J = 1.9 Hz).
4.16.2. N-Methyl-4-nitro-N-(3-(3-(thiophene-2-carbonyl)-
pyrazolo[1,5-a]pyrimidin-7-yl)phenyl)benzamide (16b).
Yield: 79%, yellow solid, mp 159–161 ꢁC, Rf (ethyl ace-
1
tate/light petroleum 4:1) = 0.28. H NMR (DMSO-d6,
500 MHz): d = 8.87 (d, 1H, J = 4.4 Hz), 8.74 (s, 1H),
8.19 (dd, 1H, J = 1.0, 3.8 Hz), 8.14 (d, 1H, J = 8.5 Hz),
8.05 (dd, 1H, J = 4.9 Hz, 1.0 Hz), 8.01 (s, 1H), 7.93
(m, 1H), 7.63 (d, 1H, J = 8.2 Hz), 7.54 (m, 1H), 7.42
(d, 1H, J = 4.4 Hz), 7.30 (dd, 1H, J = 4.9 Hz, 3.9 Hz),
3.48 (s, 1H).
4.16.8. 3-(Dimethylamino)-2-(4-fluorobenzoyl)acryloni-
trile (23). A solution of 22 (1.8 g, 11.0 mmol) in
DMFDMA (10 mL) was heated at 120 ꢁC overnight.
After cooling to room temperature the mixture was di-
luted with a mixture of dichloromethane/diethyl ether 1/
10 (40 mL). The product precipitates and was collected
by suction filtration. Yield: 60%, yellow solid, mp 150.8–
151.4 ꢁC. 1H NMR (CDCl3, 500 MHz): d = 7.96 (s,
1 H), 7.83 (m, 2H), 7.10 (s, 2H), 3.49 (s, 3H), 3.40 (s, 3H).
Calculated C: 62.10, H: 3.54, N:14.48, found C: 61.96,
H: 3.30, N: 14.35.
4.16.3. 1-[3-[(2-Fluoroethyl)methylamino]phenyl]ethanone
(17). It was prepared according to the general procedure
for acetophenones 6a–c. Yield: 41%, yellow oil, Rf
(dichloromethane) = 0.41.
1H
NMR
(CDCl3,
500 MHz): d = 7.35-7.28 (m, 3H, HAR), 7.23 (m, 1H,
HAR), 6.93 (m, 1H, HAR), 4.62 (td, 2H, J = 5.1 Hz,
J2F = 47.2 Hz, CH2F), 3.71 (td, 2H, J = 5.1 Hz,
J2F = 24.8 Hz, NCH2), 2.98 (s, 3H, CH3), 2.60 (s, 3H,
CH3). HRMS m/z calculated for C11H14FNO (M+Na)
218.0951, found 218.0950.
4.16.9. (5-Amino-1H-pyrazol-4-yl)(4-fluorophenyl)metha-
none (24). To a solution of 23 (1.96 g, 9 mmol) and
aminoguanidinium nitrate (1.54 g, 11.2 mmol) in etha-
nol (15 mL) was added sodium hydroxide (1.25 mL,
10 M). The mixture was refluxed for 4 h and the solvent
was removed. Then water (16 mL) was added and the
solution was stored at 2–8 ꢁC overnight. The precipitate
was collected by suction filtration and dissolved in ethyl
acetate (50 mL). The solution was dried over sodium
sulfate and filtered. The filtrate was concentrated and
dissolved in ethyl acetate (30 mL). Hexane was added
and the solution was allowed to crystallize. The precip-
itate was collected and dried in vacuum. Yield: 62%, yel-
low solid, mp 155.7–156.3 ꢁC. 1H NMR (DMSO-d6,
500 MHz): d = 12.09 (br s, 1H, NH), 7.80 (m, 2H,
HAR), 7.62 (br s, 1H), 7.32 (m, 2H, HAR), 6.71 (br s,
2H). HRMS m/z calculated for C10H8FN3O (M+H)
206.0724, found 206.0726.
4.16.4.
3-Dimethylamino-1-[3-[(2-fluoroethyl)methyl-
amino]phenyl]propenone 18. It was prepared according
to the general procedure for enaminones 4a–e. Yield:
86%, yellow oil, Rf (ethyl acetate/methanol 9:1) = 0.42.
1H NMR (CDCl3, 500 MHz): d = 7.79 (m, 1H, HAR),
7.30–7.18 (m, 3H, HAR), 6.83 (m, 1 H, HAR), 5.70 (d,
1H, J = 12.4 Hz, CH), 4.62 (td, 2H, J = 45.2 Hz,
J2F = 47.2 Hz, CH2F), 4.70 (td, 2H, J = 5.2 Hz,
J3F = 24.5 Hz, CH2F), 3.20–2.85 (m, 9 H, CH3). HRMS
m/z calculated for C14H19FN2O (M+H) 251.1554, found
251.1553.
4.16.5. (7-[3-[(2-Fluoroethyl)methylamino]phenyl]pyrazol-
o[1,5-a]pyrimidin-3-yl)-thiophen-2-yl-methanone (19). It
was prepared according to the general procedure for
pyrazolopyrimidines 5a–e. Yield: 35%, orange solid,
4.16.10. N-(3-(3-(4-Fluorobenzoyl)pyrazolo[1,5-a]pyrimi-
din-7-yl)phenyl)-N-methylacetamide (25). A suspension
of N-[3-(3-dimethylamino-acryloyl)-phenyl]-N-methyl-
acetamide (0.50 g, 2 mmol) and (5-amino-1H-pyrazol-
1
mp 126.7–127.5 ꢁC, Rf (ethyl acetate) = 0.75. H NMR
(CDCl3, 500 MHz): d = 8.80 (m, 1H, HAR), 8.72 (s,

1194
A. Hoepping et al. / Bioorg. Med. Chem. 16 (2008) 1184–1194
8. Lee, J. D.; Park, H. J.; Park, E. S.; Kim, D. G.; Rha, D.
W.; Kim, E. Y.; Kim, D. I.; Kim, J. J.; Yun, M.; Ryu, Y.
H.; Lee, J.; Jeong, J. M.; Lee, D. S.; Lee, M. C.; Park, C. I.
Neuroimage 2007, 34, 19.
9. Goodacre, S. C.; Street, L. J.; Hallett, D. J.; Crawforth, J.
M.; Kelly, S.; Owens, A. P.; Blackaby, W. P.; Lewis, R. T.;
Stanley, J.; Smith, A. J.; Ferris, P.; Sohal, B.; Cook, S. M.;
Pike, A.; Brown, N.; Wafford, K. A.; Marshall, G.;
Castro, J. L.; Atack, J. R. J. Med. Chem. 2006, 49, 35.
10. Grunwald, C.; Rundfeldt, C.; Lankau, H. J.; Arnold, T.;
Ho¨fgen, N.; Dost, R.; Egerland, U.; Hofmann, H. J.;
Unverferth, K. J. Med. Chem. 2006, 49, 1855.
4-yl)(4-fluorophenyl)methanone (24) (416 mg, 2 mmol)
in acetic acid (10 mL) was heated to 65 ꢁC for 1.5 h.
After cooling to room temperature the mixture was
poured onto ice/water and adjusted to pH 8 with sodium
bicarbonate before extraction with dichloromethane.
The combined organic phases were dried over sodium
sulfate and concentrated. The residue was purified by
flash chromatography. Yield: 48%, white solid, mp
121.5–123.2 ꢁC, Rf (ethyl acetate) = 0.30. 1H NMR
(CDCl3, 500 MHz): d = 8.81 (d, 1H, J = 4.3 Hz), 8.57
(s, 1H), 7.99 (m, 4H), 7.67 (t, 1H, J = 7.8 Hz), 7.47 (d,
1H, J = 7.7 Hz), 7.18 (m, 1H), 3.35 (s, 3H), 2.00 (s,
3H). Calculated C: 68.03, H: 4.41, N: 14.43, found C:
67.65, H: 4.36, N: 14.32.
11. Selleri, S.; Bruni, F.; Costagli, C.; Costanzo, A.; Guerrini,
G.; Ciciani, G.; Gratteri, P.; Besnard, F.; Costa, B.;
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P. M. J. Med. Chem. 2005, 48, 6756.
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K. L.; Carter, D. B.; Im, H. K.; Im, W. B.; Sethy, V. H.;
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W. Z.; Mickelson, J. W. J. Med. Chem. 1999, 42, 1123.
13. Frølund, B.; Jensen, L. S.; Guandalini, L.; Canillo, C.;
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14. McKernan, R. M.; Rosahl, T. W.; Reynolds, D. S.; Sur,
C.; Wafford, K. A.; Atack, J. R.; Farrar, S.; Myers, J.;
Cook, G.; Ferris, P.; Garrett, L.; Bristow, L.; Marshall,
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Acknowledgments
The authors thank Dr. C. Birkenmeyer (Institute of
Analytical Chemistry, University of Leipzig) for per-
forming the high-resolution mass spectra. We also thank
Dr. Gesine Kabuß (ABX GmbH) for proof reading the
manuscript.
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