Indol-3-yl-tetramethylcyclopropyl ketones: Effects of indole ring substitution on CB2 cannabinoid receptor activity

Article abstract of DOI:10.1021/jm7011613

A series of potent indol-3-yl-tetramethylcyclopropyl ketones have been prepared as CB₂ cannabinoid receptor ligands. Two unsubstituted indoles (5, 32) were the starting points for an investigation of the effect of indole ring substitutions on CB₂ and CB₁ binding affinities and activity in a CB₂ in vitro functional assay. Indole ring substitutions had varying effects on CB₂ and CB₁ binding, but were generally detrimental to agonist activity. Substitution on the indole ring did lead to improved CB₂/CB₁ binding selectivity in some cases (i.e., 7-9,15-20). All indoles with the morpholino-ethyl side chain (32-43) exhibited weaker binding affinity and less agonist activity relative to that of their tetrahydropyranyl-methyl analogs (5-31). Several agonists were active in the complete Freund's adjuvant model of chronic inflammatory thermal hyperalgesia (32, 15).

Full text of DOI:10.1021/jm7011613

1904
J. Med. Chem. 2008, 51, 1904–1912
Indol-3-yl-tetramethylcyclopropyl Ketones: Effects of Indole Ring Substitution on CB2
Cannabinoid Receptor Activity
Jennifer M. Frost,* Michael J. Dart, Karin R. Tietje, Tiffany R. Garrison, George K. Grayson, Anthony V. Daza,
Odile F. El-Kouhen, Loan N. Miller, Lanlan Li, Betty B. Yao, Gin C. Hsieh, Madhavi Pai, Chang Z. Zhu,
Prasant Chandran, and Michael D. Meyer
Neurological Diseases Research, Global Pharmaceutical Research & DeVelopment, Abbott Laboratories, R472, AP9A, 100 Abbott Park Rd.,
Abbott Park, Illinois 60064
ReceiVed September 17, 2007
A series of potent indol-3-yl-tetramethylcyclopropyl ketones have been prepared as CB2 cannabinoid receptor
ligands. Two unsubstituted indoles (5, 32) were the starting points for an investigation of the effect of
indole ring substitutions on CB2 and CB1 binding affinities and activity in a CB2 in vitro functional assay.
Indole ring substitutions had varying effects on CB2 and CB1 binding, but were generally detrimental to
agonist activity. Substitution on the indole ring did lead to improved CB2/CB1 binding selectivity in some
cases (i.e., 7-9, 15-20). All indoles with the morpholino-ethyl side chain (32-43) exhibited weaker binding
affinity and less agonist activity relative to that of their tetrahydropyranyl-methyl analogs (5-31). Several
agonists were active in the complete Freund’s adjuvant model of chronic inflammatory thermal hyperalgesia
(32, 15).
Introduction
of the most well-known examples from the Huffman laboratories
is 2, in which the amino alkyl group is replaced by a propyl
side chain.17 Makriyannis and co-workers have also disclosed
many aminoalkylindoles, including the CB2 inverse agonist
AM630, as well as the CB2-selective ligand 3.18 Compound 3,
although demonstrating efficacy in a range of preclinical pain
models,18,19 has also exhibited some unique characteristics
including an opioid receptor dependency20 not shared by other
CB2-selective ligands21 and varied activities in in vitro functional
assays.22,23
Merck has disclosed a selective CB2 ligand, 4, which has
the acyl group attached to the indole nitrogen and a morpholino-
ethyl side chain in the 3-indole position.24 Researchers at Bristol-
Myers Squibb have reported numerous 3-amide indoles25 and
related indolopyridones.26 There are several other reported
indole-related cannabinoid ligands, including those described
by patent applications from Organon,27 Hoffman-La
Roche,28GlaxoSmithKline,29 Sanofi-Synthelabo,30 and Schering-
Plough.31
The cannabinoid receptors are members of the G-protein
coupled receptor (GPCR)a family of receptors. The cannabinoid
1 receptor (CB1) is found in the central nervous system as well
as the periphery,1 whereas the cannabinoid 2 receptor (CB2) is
found mainly in the periphery, particularly in the immune
system.2 Evidence of CB2 receptor expression in the CNS has
recently emerged.3 Activation of the CB1 receptor is thought to
mediate the psychotropic effects associated with nonselective
agonists such as ∆9-tetrahydrocannabinol (∆9-THC), the prin-
ciple active component of marijuana. In animal studies, activa-
tion of either the CB1 or CB2 receptor will result in analgesic
activity,4 and several studies have established that CB2-selective
agonists (relative to CB1) exhibit efficacy in many rodent pain
models and lack the CB1-mediated CNS effects at analgesic
doses.5 Specifically, CB2-selective agonists have shown efficacy
in preclinical models of neuropathic and inflammatory pain.5,6
Cannabinoid receptor agonists are also being investigated for
use in cancer,7 multiple sclerosis,8 osteoporosis,9 Alzheimer’s
disease,10 liver disease,11 and diabetes.12
Aminoalkylindoles (AAIs) and indoles lacking the amine
functionality are well-established structural motifs in cannab-
inoid research.13 Initial work by Sterling Winthrop led to
pravadoline14 and later to the potent but nonselective cannab-
inoid ligand 1 (Figure 1).15 Huffman and co-workers have
subsequently published on numerous indole cannabinoid
ligands13 with their work focusing on varying the indole nitrogen
substitution and the 3-aryl acyl substituents. Importantly, the
work of Huffman and co-workers led to the realization that the
amino alkyl side chain, which was previously thought to be
necessary for interaction with the cannabinoid receptors,16 could
be replaced with groups lacking the amino functionality. One
Several structure–activity studies have explored indole nucleus
16
substitutions, both with respect to activity at the CB1 and
CB225a receptors. In a study of the CB1 receptor structure-activity
relationship (SAR), Eissenstat and co-workers16 reported that
only small groups (hydrogen, methyl) were tolerated in the
2-indole position. This conclusion is supported by the work of
Huffman and co-workers.13 The Eissenstat group also reported
that substitution in the 5-indole position (methyl, methoxy,
fluoro, bromo) was detrimental to activity in both their binding
and functional assays. Substitution at the 6-indole position
(methyl, methoxy, bromo) resulted in ligands with binding
affinity but no functional activity. Substitution at the 7-indole
position (methyl, methoxy, fluoro) gave compounds with modest
improvements in binding and functional activities relative to
the unsubstituted analogs.
* To whom correspondence should be addressed. Phone: 847-937-0721.
Fax: 847-937-9195. E-mail: jennifer.frost@abbott.com.
Hynes and co-workers have reported on the effects of indole
ring substitution on CB2 activity in their 3-amide indole series.25a
A C-7 methoxy group increased CB2 binding affinity, and a
C-2 substituent other than hydrogen resulted in decreased CB2
binding affinity. The 7-methoxy derivatives were used to
a Abbreviations: FLIPR, fluorescence imaging plate reader; CFA,
complete Freund’s adjuvant; PEG, poly(ethylene glycol); GPCR, G-protein
coupled receptor; CB1, cannabinoid 1 receptor; CB2, cannabinoid 2 receptor;
HEPES, 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid; BSA, bovine
serum albumin; PWL, paw withdrawal latency.
10.1021/jm7011613 CCC: $40.75
2008 American Chemical Society
Published on Web 02/27/2008

Indol-3-yl-tetramethylcyclopropyl Ketones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1905
Figure 1. Literature compounds.
was assessed using the CFA model of chronic inflammatory
thermal hyperalgesia. A solution of CFA in phosphate-buffered
saline was injected into the plantar surface of the right hind
paw in rats. Thermal hyperalgesia was assessed 48 h post CFA
injection.
Results and Discussion
As discussed above, the starting points of our investigation
of the substitution of the indole ring were parent compounds 5
and 32. Indole 5 proved to be one of the most potent compounds
in these series with subnanomolar human CB2 binding affinity
(Ki ) 0.21 nM) and potent (EC50 ) 9 nM), full agonist efficacy
in the FLIPR assay. Indole 5 also had relatively high affi-
nity for the human CB1 receptor (Ki ) 12 nM) and exhibited
only moderate selectivity (57-fold) for the CB2 receptor. The
potency and efficacy of 5 at the CB2 receptor made it an
appealing lead, despite its affinity for the CB1 receptor.
Therefore, the SAR of modifications at the 4–7 positions of the
indole ring was investigated (Tables 1 and 3).
Indole 32, with the morpholinoethyl side chain, was more
selective than the tetrahydropyranylmethyl derivative 5 (CB1/
CB2 ) 192), and 32 was also a potent (EC50 ) 17 nM) agonist
in the FLIPR calcium flux assay. The improved selectivity of
32 as well as its efficacy in in vivo models (vide infra) made it
another interesting target for SAR studies with modifications
at the 4–7 positions of the indole ring (Table 2).
Looking first to derivatives of 5 (Table 1), analog 6 with
electron-withdrawing fluorine substitution in the 4–7 positions
exhibited lower CB2 and CB1 binding affinities, less potency
in the FLIPR functional assay and lower selectivity for CB2 vs
CB1 (13-fold) than 5. The 5-fluoro derivative 7 displayed low
nanomolar binding affinity for CB2 and dramatically improved
binding selectivity relative to 5 or 6. Also, there was a marked
decrease in functional potency and efficacy in the FLIPR assay
as the 5-indole substitution changed from fluoro (7) to chloro
(8) to bromo (9) suggesting that increased size of the 5-sub-
stituent decreases CB2 in vitro functional activity. This is
supported by a comparison of 16, 20, and 24, which all exhibit
potent binding affinity, but where 16 (5-hydroxy) is a potent
agonist, 20 (5-methoxy) is a weak agonist, and 24 (5-benzyloxy)
lacks agonist efficacy in the FLIPR assay. All 5-halogen
substituted derivatives exhibited high levels of CB2 binding
selectivity.
Interrogation of monosubstitution at the 6-indole position also
revealed a pronounced effect of substituent size on functional
activity. The 6-chloro and 6-hydroxy analogs, 10 and 17,
respectively, were moderately potent and efficacious agonists,
whereas the 6-bromo (11), 6-methyl (12), 6-trifluoromethyl (13),
6-methylsulfonyl (14), and 6-methoxy (21) analogs all lacked
agonist activity in the FLIPR functional assay. Interestingly,
the 6-benzyloxy analog (25) exhibited weak partial agonist
Figure 2. Indol-3-yl-tetramethylcyclopropyl ketones.
investigate other indole substituents. Substitutions at the C-4
and C-6 positions were reportedly not tolerated. Only chloro
and fluoro substitutions were described for C-5, with the
5-chloro substitution leading to significantly decreased CB2
binding affinity relative to that of the 5-hydrogen analog.
The published literature has been highly biased toward
3-indolyl-acyl aromatic substitutions; however, work in our
laboratories led us to re-evaluate the use of nonaromatic rings
in this position.32 Herein we describe the SAR of indole ring
substitution on human CB2 and human CB1 binding affinity and
in vitro CB2 functional activity for a series of 3-tetramethyl-
cyclopropyl ketone substituted indoles (Figure 2). For the
purpose of this discussion, N-1 substitution was limited to the
morpholino-ethyl and the methyl-tetrahydropyran side chains
(5 and 32). The morpholino-ethyl moiety is a well-known
structural motif in cannabinoid literature,13,15,24 and the tet-
rahydropyranyl methyl group was envisioned to be a truncated
version of the morpholino-ethyl moiety. The in vivo activity of
selected compounds in the complete Freund’s adjuvant (CFA)
model of chronic inflammatory thermal hyperalgesia will also
be discussed.
Chemistry. All of the indoles discussed in this paper were
synthesized by coupling of the appropriately substituted indole
with 2,2,3,3-tetramethylcyclopropanecarbonyl chloride (Scheme
1) using EtMgBr and ZnCl2.33 This resulted in a separable
mixture of the C-3-acylated and the N-acylated products.
The C-3-acylated product then underwent N-alkylation with
either the mesylate of (tetrahydro-2H-pyran-4-yl)methanol or
2-morpholinoethanol.
Biology. The binding affinity of this series of indole ligands
was evaluated at recombinant human CB1 and human CB2
receptors through competition binding against [3H]-56 (CP
55,940).34 In vitro functional activity was assessed in an
HEK293 cell line coexpressing the human CB2 receptor and a
chimeric GRq/o5 protein to facilitate redirection of the GRi/o
signaling to intracellular calcium release responses and enable
measurement of calcium mobilization using a fluorescence
imaging plate reader (FLIPR) as previously described.22,35
Maximal efficacy (% max) in the FLIPR assay was determined
relative to the response elicited by 10 µM 56. In vivo activity

1906 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Frost et al.
Scheme 1. General Synthesis of Indol-3-yl-tetramethylcyclopropyl Ketones
Table 1. In Vitro Biological Activity of Tetrahydropyranyl-methyl Series
Human CB2 Binding
Human CB1 Binding
CB2 FLIPR
EC50 (nM)
Compd
R4
R5
R6
R7
pKi ( SEM
Ki (nM)
pKi ( SEM
Ki (nM)
CB1/CB2
(SEM range)
% max
1 (WIN 55,212-2)
2 (JWH-015)
8.89 ( 0.06
7.45 ( 0.06
7.94 ( 0.10
9.67 ( 0.12
8.31 ( 0.07
8.80 ( 0.17
8.52 ( 0.08
8.18 ( 0.27
8.60 ( 0.19
9.19 ( 0.08
9.83 ( 0.12
8.46 ( 0.08
8.53 ( 0.10
8.41 ( 0.11
8.72 ( 0.11
8.07 ( 0.20
8.50 ( 0.05
8.48 ( 0.09
8.34 ( 0.05
9.29 ( 0.27
9.90 ( 0.10
8.03 ( 0.11
8.90 ( 0.22
9.05 ( 0.06
8.52 ( 0.07
8.75 ( 0.09
9.16 ( 0.12
7.67 ( 0.07
8.99 ( 0.05
1.3
35
7.88 ( 0.12
5.92 ( 0.10
5.90 ( 0.25
7.91 ( 0.18
7.18 ( 0.25
5.92 ( 0.08
6.30 ( 0.12
5.72 ( 0.10
7.86 ( 0.30
7.88 ( 0.06
7.56 ( 0.28
7.16 ( 0.18
5.90 ( .05
6.41 ( 0.10
6.69 ( 0.15
>5.52
6.61 ( 0.09
5.96 ( 0.20
6.55 ( 0.10
7.40 ( 0.20
7.35 ( 0.11
>5
6.62 ( 0.19
7.48 ( 0.02
5.96 ( 0.06
5.55 ( 0.11
6.11 ( 0.15
6.40 ( 0.22
7.55 ( 0.19
13.3
1204
1269
12
10
34
110
57
13
756
171
292
5.6
20
187
20
423
99
86–163
634–961
>10,000
7–12
37–43
22–51
92–172
280–501
10–32
>10,000
>10,000
>10,000
>10,000
64–82
74 ( 4
75 ( 5
3 (AM1241)
11.5
0.21
5.0
1.6
3.0
6.6
2.5
0.65
0.15
3.5
3.0
3.9
1.9
8.5
3.2
3.3
4.6
0.51
0.12
9.3
1.3
0.88
3.1
1.8
0.7
21
-
5
6
7
8
H
F
H
F
H
F
H
F
133 ( 9
66
88 ( 3
H
H
H
H
H
H
H
H
F
H
H
H
Cl
Br
H
H
H
H
H
H
H
H
H
H
H
OH
H
H
H
OCH3
H
1210
512
1930
14
13
28
88 ( 4
Cl
Br
H
69 ( 7
9
54 ( 10
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
60 ( 6
H
-
-
-
-
H
H
H
H
CH2
CF3
SO2CH3
H
70
1270
388
204
>3,000
246
1090
282
40
OH
H
H
94 ( 4
OH
H
H
H
OH
H
107
>353
77
330
61
7–15
15–22
78 ( 5
107 ( 4
H
>10,000
81–101
61–141
>10,000
>10,000
383–620
>10,000
135–199
>10,000
>10,000
>10,000
21–35
-
OCH3
H
H
OCH3
H
H
H
OCH3
H
37 ( 7
35 ( 3
-
-
45 ( 6
-
45 ( 4
-
-
-
H
H
OBn
H
H
H
H
H
H
78
H
45
375
>1075
183
38
353
1558
1119
19
H
H
>10000
238
33
1095
2804
783
395
28
OBn
H
H
H
OBn
H
H
H
OBn
H
H
OBn
OH
OH
OCH3
OCH3
OH
H
H
106 ( 4
62 ( 1
H
1.0
28
18–41
31
H
NH2
H
H
8.03 ( 0.17
9.3
>6
>1000
108
10–21
80 ( 8
activity. All of the 6-substituted analogs exhibited high affinity
for the CB2 receptor. The 6-methylsulfonyl derivative (14) and
5-hydroxy (17) derivative exhibited low affinity for the CB1
receptor, and all other 6-substituted analogues investigated had
higher binding affinity (<100 nM) for the CB1 receptor.
Substitutions at the 4 and 7 positions of the indole ring were
also investigated. The 4-hydroxy analog (15) was a potent, fully
efficacious agonist in the FLIPR assay. In contrast, although
the 7-hydroxy analog (18) demonstrated high affinity for the
CB2 binding site and moderate affinity for the CB1 receptor, it
failed to exhibit agonist activity. Similar trends were obtained
with the 4- (19) and 7-methoxy (22) and the 4- (23) and
high affinity for the CB2 receptor, weaker or no affinity for the
CB1 receptor, and weak agonist activity in the FLIPR assay.
The 7-substituted analogs (22, 26) exhibited high CB2 and CB1
receptor affinity but lacked agonist activity in the FLIPR assay.
Several 5,6-bis-substituted indole analogues were also inves-
tigated (27-30). Analogues 27 and 28 had good affinity for
CB2 but did not demonstrate agonist activity in the FLIPR assay,
which is consistent with the corresponding monosubstituted
analogues (21, 24). The diol 29 had lower affinity and was less
selective than its monosubstituted analogues (16, 17). Perhaps
the most interesting bis-substituted analogue was 30, which
maintained high potency and near full agonist efficacy in the
FLIPR assay (18–41 nM, 62%), whereas the similar 5-hydroxy,
7-benzyloxy (26) analogs. Both 4-substituted analogs exhibited

Indol-3-yl-tetramethylcyclopropyl Ketones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1907
Table 2. In Vitro Biological Activity of Morpholino-ethyl Series
Human CB2 Binding
Human CB1 Binding
CB2 FLIPR
EC50 (nM)
Compd
R4
R5
R6
R7
R2
pKi ( SEM
Ki (nM)
pKi ( SEM
Ki (nM)
CB1/CB2
(SEM range)
max
32
33
34
35
36
37
38
39
40
41
42
43
H
H
H
H
H
H
H
H
H
OH
H
OCH3
H
OBn
H
H
H
H
H
H
H
OH
H
OCH3
H
OBn
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
CH3
H
8.36 ( 0.17
7.04 ( 0.11
6.96 ( 0.09
6.63 ( 0.09
8.80 ( 0.04
<6
4.4
92
109
237
1590
>1000
2.9
27
297
6.07 ( 0.14
<5
845
192
14–21
71 ( 4
>10,000
>10,000
>3,500
1285
>10000
986
>10,000
>10,000
>3,500
>2,200
4077
>109
>92
15
>10,000
186–280
>10,000
>10,000
>10,000
>10,000
227–497
>10,000
50–380
-
<5
62 ( 1
-
-
-
>5.46
5.89 ( 0.11
<5
6.00 ( 0.04
<5
<5
>5.46
>5.66
5.39 ( 0.01
0.81
8.53 ( 0.07
7.57 ( 0.09
6.53 ( 0.11
7.49 ( 0.11
<6
340
>370
>34
>109
-
-
60 ( 6
-
41 ( 5
-
-
NO2
NH2
NHC(O)CH3
H
H
H
32
>1000
>1000
>10,000
>10,000
H
<6
-
Table 3. In Vitro Biological Activity of Additional Tetrahydropyranyl-methyl Analogues
human CB2 binding
human CB1 Binding
CB2 FLIPR
compd
R5
R6
pK i ( SEM
Ki (nM)
pK i ( SEM
Ki (nM)
CB1/CB2
EC50 (nM)(SEM range)
max
44
45
46
47
48
49
50
51
52
53
54
55
O(CH2)4OH
O(CH2)4Br
CN
CH2NH2
CH2OH
CH2OCH3
C(O)OCH3
Ph
H
H
H
H
H
H
H
H
H
H
H
H
6.58 ( 0.11
7.14 ( 0.11
7.52 ( 0.26
<6
6.95 ( 0.09
7.59 ( 0.012
6.62 ( 0.07
7.23 ( 0.09
8.30 ( 0.012
7.21 ( 0.26
8.86 ( 0.40
9.39 ( 0.13
260
72
30
>1000
113
26
238
58
5.0
62
1.4
0.41
<5
>10,000
>10,000
3326
>10,000
>10,000
1702
>10,000
1875
31
>38
>139
111
>10,000
>10,000
226–299
>10,000
73–122
<5
5.48 ( 0.05
<5
<5
5.77 ( 0.08
<5
5.73 ( 0.27
7.50 ( 0.27
<4
65 ( 3
>88
65
>42
32
6.2
>16
18
53 ( 6
50 ( 5
41–108
>10,000
>10,000
45–61
>10,000
>10,000
>10,000
CN
67 ( 7
CH2NH2
C(O)OCH3
Ph
>1,000
25
228
7.60 ( 0.17
6.64 ( 0.16
556
6-methoxy analogue, 28, the 5-methoxy analogue, 20, and the
6-methoxy analogue, 21, exhibited little or no agonist activity
in FLIPR.
derivative 38 being the one example with binding affinity
similar to that of 32. Finally, as is consistent with the
previously reported work,13,16,25a substitution at the C-2
indole position resulted in decreased binding affinity at both
the CB2 and CB1 receptors and decreased functional activity
at the CB2 receptor (39 vs 32).
The effects of substitutions on the 5 or 6 positions of the
indole ring were further investigated in the tetrahydropyranyl-
methyl series as shown in Table 3. As demonstrated in Tables
1 and 2, indole substituents larger than a methyl group generally
resulted in loss of agonist activity in the FLIPR assay. Analogues
with smaller substituents such as the cyano group (46, 52) did
retain agonist activity, as did the 5-hydroxymethyl and 5-meth-
oxymethyl analogues (48, 49); however, these compounds were
only partial agonists in the FLIPR assay. With regard to binding,
larger groups were tolerated and less polar substitutions
displayed better CB2 activity than more polar substitutions (i.e.,
As shown in Table 2, the morpholino-ethyl derivatives
were also examined. Analogue 32 is a CB2-selective com-
pound with high affinity for the CB2 receptor and potent
activity in the FLIPR assay, and it proved to be the most
interesting analogue in this group of morpholino-ethyl
derivatives. In most cases where direct comparisons can be
made between the morpholino-ethyl series and the tetrahy-
dropyranyl-methyl series, the analogues in the latter group
demonstrate higher affinity for CB2, as well as greater potency
and efficacy in the FLIPR assay (33 vs 16, 34 vs 17, 35 vs
20). When comparing the substituted indoles in the mor-
pholino-ethyl series with the parent 32, it is clear that all
indole substitution is detrimental to agonist activity and, in
most cases, to binding affinity as well, with the 6-benzyloxy

1908 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Frost et al.
Figure 3. Dose response of 32 in CFA model of chronic inflammatory
pain. Thermal hyperalgesia was assessed 48 h post CFA injection; n
) 6 for each dose; vehicle is 5% DMSO/95% poly(ethylene glycol)
(PEG); *p < 0.05 vs vehicle control; **p < 0.01 vs vehicle control.
Figure 4. Activity of 32 in CFA model with pretreatment with 57
and pretreatment with 58. Thermal hyperalgesia was assessed 48 h post
CFA injection; n ) 6 for each dose; vehicle is 5% DMSO/95% PEG;
**p < 0.01 vs vehicle control.
45 vs 44 and 48 vs 47). With respect to binding affinity, the 6
position appears to be more tolerant of larger substitutions than
the 5 position (i.e., 54 vs 50 and 55 vs 51). Polar substitutions
also appear to be better tolerated in the 6 position (i.e., 53 vs
47). However, agonist efficacy was generally lost when large
substituents were introduced into any position on the indole
nucleus.
In Vitro SAR Summary. Clearly, the effect of indole ring
substitution on CB2 functional activity is sensitive to sub-
stituent size, with the effect varying by location of the indole
substituent. For example, in the 4- and 5-indole positions
the methoxy group is tolerated for partial agonist activity
(19, 20), but this substituent is not tolerated in the 6 or 7
positions (21, 22). Binding at both the CB2 and CB1 receptors
is less sensitive to substituent size with aryl groups (51, 55,
23-26) and relatively large alkyl groups being tolerated (44,
45). The morpholino-ethyl analogues (Table 2) were generally
weaker than the tetrahydropyranyl analogues (Tables 1 and
3). All substitution in both series led to decreased CB2
functional activity in the FLIPR assay but had less of an
effect on binding affinities.
Figure 5. Dose response of 15 in CFA model of chronic inflammatory
pain. Thermal hyperalgesia was assessed 48 h post CFA injection; n
) 6 for each dose; vehicle is 5% DMSO/95% PEG, **p < 0.01 vs
vehicle control.
CB2 antagonist 58 completely blocked activity of 32 in this
model, providing strong evidence that the effect of 32 is
mediated through activation of the CB2 receptor and not CB1.
A second analogue (15), exhibiting a selectivity and efficacy
profile comparable to those of 32, was selected for evaluation
in the CFA model of chronic inflammatory thermal hyperalgesia.
Compound 15, an agonist in the FLIPR assay (EC50 ) 73 nM,
94% response), exhibited high affinity for the CB2 receptor (Ki
) 3.9 nM) and 100-fold selectivity vs CB1, and as shown in
Figure 5, it also demonstrated robust dose-dependent efficacy
in the CFA model. By contrast, analogue 55, which demon-
strated a comparable radioligand binding affinity and selectivity
profile but failed to exhibit agonist efficacy in the FLIPR
functional assay, was inactive (data not shown) in the CFA
inflammatory pain model.
In summary, two series of indole derivatives were prepared
and evaluated for human CB2 affinity, selectivity against CB1
affinity, and agonist activity at the human CB2 receptor. The
two lead compounds, 5 and 32, both demonstrated high affinity
for the CB2 receptor, and both were agonists in the FLIPR
functional assay. The tetrahydropyranyl-methyl analogue 5 was
more potent than the corresponding morpholino-ethyl derivative
32, but 5 also exhibited high affinity for the CB1 receptor (Ki
) 12 nM). To further investigate the SAR of these compounds,
numerous indole-substituted analogues were prepared. In the
tetrahydropyranyl-methyl series, several analogues exhibited
binding selectivity for CB2 better than that of 5 (i.e., 7, 15–17,
29–31, 48, 52) while still maintaining high affinity for the CB2
binding site and good potency and efficacy in the FLIPR assay.
In the morpholino-ethyl series, all indole-substituted analogues
Comparing our series with the previously reported work of
Eissenstat16 and Hynes,25a several similarities in SAR trends
exist. For example, Eissenstat and co-workers demonstrated that
C-5 substituents gave weaker CB1 binding affinity, and this was
also true with our compounds (i.e., 7–9, 16, 24 vs 5). A C-7
16
methoxy substituent was reported to enhance both CB1 and
25a
CB2 binding affinity, whereas in our series, C-7 methoxy
derivative 22 exhibited weaker CB1 binding affinity and
improved CB2 binding affinity relative to that of the parent
analogue 5.
In Vivo Characterization. An accumulating body of
evidence suggests the potential utility of CB2-selective
agonists in the treatment of pain. To test this hypothesis,
several CB2-selective analogues exhibiting in vitro agonist
efficacy were selected for evaluation in the CFA model of
chronic inflammatory thermal hyperalgesia. Compound 32,
which exhibited high radioligand binding affinity for the CB2
receptor (Ki ) 4.4 nM), very good selectivity (CB1/CB2 )
192), and high potency and near full efficacy in the FLIPR
assay (EC50 ) 17 nM, 71% maximal response), was also
efficacious in the CFA model of chronic inflammatory
thermal hyperalgesia (Figure 3). The dependence on CB2
receptor activation was demonstrated by coadministration of
32 with the selective CB1 and CB2 receptor antagonists 57
(SR141716A)36 and 58 (SR144528),37 respectively (Figure
4). Pretreatment with the CB1 antagonist 57 resulted in no
reduction in efficacy for 32, whereas pretreatment with the

Indol-3-yl-tetramethylcyclopropyl Ketones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1909
and the maximum time of exposure was set at 20.48 s to limit
possible tissue damage. The elapsed time until a brisk withdrawal
of the hind paw from the thermal stimulus was recorded automati-
cally using photodiode motion sensors. The right and left hind paw
of each rat was tested in 3 sequential trials at approximately 5 min
intervals. Paw withdrawal latency (PWL) was calculated as the
mean of the two shortest latencies. PWL was measured 30 min
post administration in both the CFA-treated (ipsilateral) and
uninjected paw (contralateral).
Chemistry. Proton NMR spectra were obtained on a General
Electric QE 300 or QZ 300 MHz instrument with chemical shifts
(δ) reported relative to tetramethylsilane as an internal standard.
Elemental analyses were performed by Robertson Microlit Labo-
ratories or Quantitative Technologies, Inc. Column chromatography
was carried out on silica gel 60 (230–400 mesh). Thin-layer
chromatography was performed using 250 mm silica gel 60 glass-
backed plates with F254 as indicator. All materials were com-
mercially available and were obtained from Aldrich unless otherwise
specified.
2,2,3,3-Tetramethylcyclopropanecarbonyl Chloride. To a flask
containing 2,2,3,3-tetramethylcyclopropane carboxylic acid (13.5
g, 95 mmol) was added 30 mL of thionyl chloride (410 mmol,
excess). This solution was warmed to reflux and was stirred for
2 h. The mixture was then cooled to ambient temperature and
concentrated under reduced pressure. The residue was diluted three
times with 10 mL of benzene and concentrated to remove any
remaining thionyl chloride. This was repeated two additional times,
and the material was used without further purification or charac-
terization.
Tetrahydro-2H-pyran-4-ylmethyl Methanesulfonate. To tet-
rahydropyran-4-methanol (Combi-Blocks, Inc., 0.15 g, 1.2 mmol)
in 10 mL of tetrahydrofuran (THF) at 0 °C was added triethylamine
(0.56 mL, 4.1 mmol) followed by methanesulfonyl chloride (0.15
mL, 1.9 mmol). The mixture was stirred at 0 °C for 10 min, the
ice-bath was removed, and the reaction mixture was stirred at 23
°C for an additional 1.5 h. The reaction mixture was filtered though
Celite with THF and concentrated under reduced pressure. The
crude tetrahydro-2H-pyran-4-ylmethyl methanesulfonate was used
without further purification or characterization.
exhibited lower affinity and lower potency compared to those
of 32, with only three analogues (34, 39, and 41) showing any
agonist activity in the FLIPR assay. Overall, agonist activity
was highly dependent on indole substituent size, particularly in
the 6- and 7-indole positions. CB2 binding affinity proved much
less sensitive to substituent size, especially in the tetrahydro-
pyranyl-methyl series where even aryl-substituted indoles
retained high binding affinity (i.e., 23-26, 51, 55).
Compound 32 is a novel, high affinity ligand for the CB2
receptor, exhibiting selectivity versus the CB1 binding site. It
displays full agonist efficacy in an in vitro functional assay and
is active in a model of chronic inflammatory pain, an effect
that is selectively blocked by pretreatment with a CB2 antagonist
and not by a CB1 antagonist. A more detailed characterization
of the in vitro and in vivo properties of this ligand has been
published,38 and a description of the effects of further variations
of the indole nitrogen side chain in the 3-cycloalkyl acyl indole
series will be forthcoming.
Experimental Section
Radioligand Binding Assays. Membrane samples prepared from
HEK cells stably expressing the human CB2 receptor and the CHO
cells stably expressing the human CB1 receptor were used to
perform radioligand binding assays using [3H]-56 as previously
described.35 Briefly, competition experiments were conducted using
0.5 nM [3H]-56 in the presence of variable concentrations of test
compounds in an assay buffer containing 50 mM Tris-HCl, pH 7.4,
2.5 mM EDTA, 5 mM MgCl2, and 0.05% fatty acid free BSA.
After 90 min of incubation at 30 °C, the reactions were terminated
by rapid vacuum filtration through UniFilter-96 GF/C filter plates
(Perkin-Elmer, Boston, MA) and six washes with cold assay buffer,
and the filter plates were air-dried. The bound activity was counted
in a TopCount using Microscint-20 (Perkin-Elmer). Nonspecific
binding was defined by 10 µM unlabeled 56. Ki values from
competition binding assays were determined with one site binding
or one site competition curve fitting using the MDL Assay Explorer
software(SanRamon,CA).Dataarepresentedasmeanvalues ( standard
error of the mean (SEM) of at least three independent experiments,
each of which was performed in duplicate.
Fluorescence Imaging Plate Reader Functional Assays. FLIPR
assays were performed using HEK cells stably coexpressing the
chimeric GRq/o5 protein with the human CB2 receptor.35 Briefly,
cells were seeded at 75,000 cells per well 1 day prior to the assay,
and assays were performed with no-wash dye (FLIPR Calcium
Assay Kit, Molecular Device, Sunnyvale, CA) following vendor’s
instruction. Variable concentrations of test compounds (0.3 nM to
10 µM) and positive control 56 (at 10 µM final concentration) or
vehicle negative control were added to cells in the presence of assay
buffer (10 mM HEPES, pH 7.4, 130 mM NaCl, 5 mM KCl, 0.05%
BSA), and fluorescence responses were measured immediately with
a FLIPR machine. Net peak responses were compared with that of
10 µM 56 and expressed as percentages of the 56-evoked response.
EC50 values were analyzed with sigmoidal dose response curve
fitting using MDL Assay Explorer software. Data are presented as
mean values ( standard error of the mean (SEM) of at least three
independent experiments, each of which was performed in duplicate.
Chronic Inflammatory Pain Model. Chronic inflammatory
thermal hyperalgesia was induced by injection of 150 µL of a 50%
solution of CFA in phosphate-buffered saline into the plantar surface
of the right hind paw in rats. Thermal hyperalgesia was assessed
48 h post CFA injection.
2-Morpholin-4-ylethyl Methanesulfonate. A solution of 4-(2-
hydroxylethyl)-morpholine (5.1 mL, 42 mmol), triethylamine (17
mL, 124 mmol), and methanesulfonyl chloride (4.8 mL, 62 mmol)
in 100 mL of THF was processed as described in the procedure
for the tetrahydro-2H-pyran-4-ylmethyl methanesulfonate to give
the crude 2-morpholin-4-ylethyl methanesulfonate, which was used
without further purification or characterization.
1H-Indol-3-yl(2,2,3,3-tetramethylcyclopropyl)methanone. To
a solution of indole (11 g, 95 mmol) in 30 mL of dichloromethane
at ambient temperature was added 105 mL of a 1 M solution of
ethyl magnesium bromide in THF (105 mmol) dropwise via syringe
pump. After the addition was complete, the solution was stirred
for 15 min at which time 105 mL of a 1 M solution of ZnCl2 in
Et2O (105 mmol) was added. The mixture was stirred for an
additional 30 min, and then 2,2,3,3-tetramethylcyclopropanecar-
bonyl chloride (95 mmol) in 50 mL of dichloromethane was added
via cannula. The mixture was stirred for 6 h at ambient tempera-
ture and then was quenched with 50 mL of saturated, aqueous
NH4Cl and diluted with 50 mL dichloromethane. The layers were
separated, and the aqueous layer was extracted with 3 × 30 mL
dichloromethane. The combined organics were washed with 20 mL
of H2O, dried over anhydrous Na2SO4, filtered, and concentrated
under reduced pressure. The crude material was purified via column
chromatography (SiO2, 50% ethyl acetate/hexanes) to give 9.7 g
of the major regioisomer 1H-indol-3-yl(2,2,3,3-tetramethylcyclo-
propyl)methanone (40 mmol, 42% yield) and 6.1 g of the minor
regioisomer of 1-[(2,2,3,3-tetramethylcyclopropyl)carbonyl]-1H-
indole (25 mmol, 27% yield). 1H NMR (major product) (300 MHz,
CD3OD) δ ppm 1.32 (s, 6 H), 1.33 (s, 6 H), 2.14 (s, 1 H), 7.12–7.24
(m, 2 H), 7.38–7.46 (m, 1 H), 8.02 (s, 1 H), 8.19–8.25 (m, 1 H);
1H NMR (minor product) (300 MHz, CD3OD) δ ppm 1.29 (s, 6
H), 1.34 (s, 6 H), 1.94 (s, 1 H), 6.66 (dd, J ) 3.7, 0.7 Hz, 1 H),
Thermal hyperalgesia was determined using a commercially
available thermal paw stimulator (UARDG, University of California,
San Diego, CA) as described by Hargreaves et al.39 Rats were
placed into individual plastic cubicles mounted on a glass surface
maintained at 30 °C and allowed a 20 min habituation period. A
thermal stimulus, in the form of radiant heat, emitted from a focused
projection bulb, was then applied to the plantar surface of each
hind paw. The stimulus current was maintained at 4.50 ( 0.05 A,

1910 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Frost et al.
7.16–7.32 (m, 2 H), 7.51–7.58 (m, 1 H), 7.67 (d, J ) 3.7 Hz, 1 H),
8.32–8.39 (m, 1 H); MS (major and minor regioisomers) (DCI/
NH3) m/z 242 (M + H)+.
(m, 3 H), 7.44–7.50 (m, 2 H), 8.06 (d, J ) 8.8 Hz, 1 H), 8.09 (d,
J ) 3.1 Hz, 1 H); MS (DCI/NH3) m/z 348 (M + H)+.
The (6-benzyloxy-1H-indol-3-yl)-(2,2,3,3-tetramethylcyclopro-
pyl)methanone (0.90 g, 2.6 mmol), tetrahydro-2H-pyran-4-ylmethyl
methanesulfonate (4.4 mmol), and NaH (60% dispersion in mineral
oil, 0.31 g, 7.8 mmol) in 15 mL of DMF were processed as
described in the procedure for 5 to provide 25 (0.87 g, 2.0 mmol,
75% yield). 1H NMR (300 MHz, CDCl3) δ ppm 1.29 (s, 6 H),
1.34 (s, 6 H), 1.34–1.51 (m, 4 H), 1.90 (s, 1 H), 1.98–2.12 (m, 1
H), 3.30 (dt, J ) 11.7, 2.4 Hz, 2 H), 3.91–4.00 (m, 2 H), 3.93 (d,
J ) 7.1 Hz, 2 H), 5.15 (s, 2 H), 6.81 (d, J ) 2.4 Hz, 1 H), 7.01
(dd, J ) 8.8, 2.0 Hz, 1 H), 7.29–7.43 (m, 3 H), 7.43–7.49 (m, 2
H), 7.50 (s, 1 H), 8.28 (d, J ) 8.8 Hz, 1 H); MS (DCI/NH3) m/z
446 (M + H)+; Anal. (C29H35NO3) C, H, N.
[1-(2-Morpholin-4-ylethyl)-1H-indol-3-yl](2,2,3,3-tetrameth-
ylcyclopropyl)methanone p-Toluenesulfonic Acid (32). 1H-Indol-
3-yl(2,2,3,3-tetramethylcyclopropyl)methanone (5.0 g, 21 mmol),
2-morpholin-4-ylethyl methanesulfonate (42 mmol), and NaH (60%
dispersal in mineral oil, 4.2 g, 104 mmol) in 40 mL of dimethyl-
formamide were processed as described in the procedure for 5 to
provide 6.6 g of [1-(2-morpholin-4-ylethyl)-1H-indol-3-yl](2,2,3,3-
tetramethylcyclopropyl)methanone 18.6 mmol, 90% yield). 1H
NMR (300 MHz, CD3OD) δ ppm 1.33 (s, 12 H), 2.13 (s, 1 H),
2.46–2.54 (m, 4 H), 2.79 (t, J ) 6.4 Hz, 2 H), 3.61–3.71 (m, 4 H),
4.37 (t, J ) 6.4 Hz, 2 H), 7.16–7.30 (m, 2 H), 7.45–7.53 (m, 1 H),
8.11 (s, 1 H), 8.20–8.30 (m, 1 H), MS (DCI/NH3) m/z 355 (M +
H)+.
To [1-(2-morpholin-4-ylethyl)-1H-indol-3-yl](2,2,3,3-tetrameth-
ylcyclopropyl)methanone (6.6 g, 19 mmol) in 25 mL of EtOAc
and 5 mL of EtOH was added p-toluenesulfonic acid monohydrate
(3.5 g, 19 mmol). No precipitate formed after 10 min of stirring,
so the crude material was concentrated under reduced pressure and
dried under reduced pressure to give 9.4 g of the title compound
(18 mmol, 96% yield). 1H NMR (MeOH-d4, 300 MHz) δ 1.33 (s,
6H), 1.34 (s, 6H), 2.15 (s, 1H), 2.36 (s, 3H), 3.40 (m, 4H), 3.68
(dd, J ) 7.1, 7.1 Hz, 2H), 3.90 (m, 4H), 4.73 (dd, J ) 7.1, 7.1 Hz,
2H), 7.23 (br d, J ) 7.8 Hz, 2H), 7.26 (ddd, J ) 8.1, 8.1, 1.4 Hz,
1H), 7.33 (ddd, J ) 7.1, 7.1, 1.0 Hz, 1H), 7.56 (br d, J ) 8.1 Hz,
1H), 7.72 (br d, J ) 8.5 Hz, 2H), 8.15 (s, 1H), 8.29 (dt, J ) 7.8,
1.0 Hz, 1H); MS (DCI/NH3) m/z 355 (M + H)+; Anal.
(C22H30N2O2 ·C7H8O3S) C, H, N.
[5-Hydroxy-1-(2-morpholin-4-ylethyl)-1H-indol-3-yl](2,2,3,3-
tetramethylcyclopropyl)methanone (33). A mixture of 37 (1.2
g, 2.5 mmol) and Pd/C (10 wt % palladium on activated carbon,
120 mg) in 50 mL of EtOH was processed as described in the
procedure for 17 to provide 33 (0.85 g, 2.3 mmol, 92% yield). 1H
NMR (CDCl3, 300 MHz) δ ppm 1.30 (s, 6 H), 1.34 (s, 6 H), 1.87
(s, 1 H), 2.41–2.58 (m, 4 H), 2.70–2.84 (m, 2 H), 3.66–3.81 (m, 4
H), 4.16–4.28 (m, 2 H), 4.84–4.98 (m, 1 H), 6.87 (dd, J ) 8.8, 2.4
Hz, 1 H), 7.21 (d, J ) 8.8 Hz, 1 H), 7.73 (s, 1 H), 7.88 (d, J ) 2.7
Hz, 1 H); MS (DCI/NH3) m/z 371 (M + H)+; Anal. (C22H30N2O3)
C, H, N.
[1-(Tetrahydro-2H-pyran-4-ylmethyl)-1H-indol-3-yl](2,2,3,3-
tetramethylcyclopropyl)methanone (5). To a solution of 1H-indol-
3-yl(2,2,3,3-tetramethylcyclopropyl)methanone (0.15 g, 0.62 mmol)
in 8 mL of DMF at 0 °C was added NaH (60% dispersion in mineral
oil, 0.12 g, 3.1 mmol). This mixture was stirred at 0 °C for 10
min, warmed to ambient temperature, and allowed to stir for 30
min. The solution was again cooled to 0 °C and tetrahydro-2H-
pyran-4-ylmethyl methanesulfonate (2.1 mmol) in 5 mL of DMF
was added via cannula. The ice-bath was removed after the addition
was complete, and the reaction mixture was warmed to 45 °C and
stirred for 2 h. The mixture was cooled to ambient temperature,
diluted with 10 mL of ethyl acetate and quenched with 10 mL of
saturated, aqueous NH4Cl and 5 mL of H2O. The layers were
separated, the aqueous layer was extracted with 3 × 5 mL ethyl
acetate, and the combined organics were dried over anhydrous
Na2SO4, filtered, concentrated under reduced pressure, and purified
via column chromatography (SiO2, 50% hexanes in EtOAc) to give
0.19 g of 5 (0.56 mmol, 90% yield). 1H NMR (300 MHz, CDCl3)
δ ppm 1.31 (s, 6H), 1.35 (s, 6H), 1.46 (m, 4H), 1.94 (s, 1H), 2.16
(m, 1H), 3.33 (dt, J ) 11.5, 2.4 Hz, 2H), 3.98 (dd, J ) 10.5, 3.1
Hz, 2H), 4.04 (d, J ) 7.5 Hz, 2H), 7.27 (m, 2H), 7.33 (m, 1H),
7.61 (s, 1H), 8.40 (m, 1H); MS (DCI/NH3) m/z 340 (M + H)+;
Anal. (C22H29NO2) C, H, N.
[6-Hydroxy-1-(tetrahydro-2H-pyran-4-ylmethyl)-1H-indol-3-
yl](2,2,3,3-tetramethylcyclopropyl)methanone (17). A mixture of
25 (0.64 g, 1.4 mmol) and Pd/C (10 wt % palladium on activated
carbon, 100 mg) in 20 mL of EtOH and 10 mL of EtOAc was
stirred under 1 atm of H2 (balloon) for 16 h. The system was purged
with an inert nitrogen atmosphere. The mixture was filtered,
concentrated under reduced pressure, and purified via column
chromatography (SiO2, 50% hexanes in EtOAc) to provide 17 (0.48
1
g, 1.35 mmol, 94% yield). H NMR (300 MHz, CDCl3) δ ppm
1.29 (s, 6 H), 1.34 (s, 6 H), 1.38–1.58 (m, 4 H), 1.89 (s, 1 H),
2.06–2.21 (m, 1 H), 3.33 (dt, J ) 11.8, 2.2 Hz, 2 H), 3.95 (d, J )
7.1 Hz, 2 H), 3.97–4.04 (m, 2 H), 4.67 (s, 1 H), 6.76–6.81 (m, 2
H), 7.50 (s, 1 H), 8.25 (d, J ) 9.2 Hz, 1 H); MS (DCI/NH3) m/z
356 (M + H)+; Anal. (C22H29NO3) C, H, N.
[6-Methoxy-1-(tetrahydro-2H-pyran-4-ylmethyl)-1H-indol-3-
yl](2,2,3,3-tetramethylcyclopropyl)methanone (21). To a solution
of 17 (0.15 g, 0.42 mmol) in 10 mL of THF was added NaH (60%
dispersion in mineral oil, 51 mg, 1.3 mmol) followed by CH3I (39
µL, 0.63 mmol). The mixture was stirred at ambient temperature
for 18 h and then was quenched with 3 mL of saturated aqueous
NH4Cl. The mixture was diluted with 10 mL of EtOAc, the layers
were separated, and the aqueous layer was extracted with 3 × 3
mL of EtOAc. The combined organic extracts were dried over
anhydrous Na2SO4, filtered, concentrated under reduced pressure,
and purified via column chromatography (SiO2, 30% hexanes in
1
EtOAc) to provide 21 (86 mg, 0.23 mmol, 55% yield). H NMR
(300 MHz, CDCl3) δ ppm 1.30 (s, 6 H), 1.34 (s, 6 H), 1.34–1.63
(m, 4 H), 1.90 (s, 1 H), 2.05–2.24 (m, 1 H), 3.34 (dt, J ) 11.7, 2.4
Hz, 2 H), 3.88 (s, 3 H), 3.94–4.02 (m, 2 H), 3.97 (d, J ) 7.5 Hz,
2 H), 6.77 (d, J ) 2.4 Hz, 1 H), 6.92 (dd, J ) 8.8, 2.0 Hz, 1 H),
7.51 (s, 1 H), 8.28 (d, J ) 8.8 Hz, 1 H); MS (DCI/NH3) m/z 370
(M + H)+; Anal. (C23H31NO3) C, H, N.
[6-(Benzyloxy)-1-(tetrahydro-2H-pyran-4-ylmethyl)-1H-indol-
3-yl](2,2,3,3 tetramethylcyclopropyl)methanone (25). A mixture
of 6-benzyloxyindole (Lancaster, 2.0 g, 9.0 mmol), ethylmagnesium
bromide (1.0 M solution in THF, 11 mL, 11 mmol), zinc chloride
(1.0 M solution in Et2O, 11 mL, 11 mmol), and 2,2,3,3-tetrameth-
ylcyclopropanecarbonyl chloride (13.4 mmol) in 30 mL of dichlo-
romethane was processed as described in the procedure for 1H-
indol-3-yl(2,2,3,3-tetramethylcyclopropyl)methanone to provide (6-
benzyloxy-1H-indol-3-yl)-(2,2,3,3-tetramethyl-cyclopropyl)-
methanone (2.0 g, 5.8 mmol, 64% yield). 1H NMR (300 MHz,
DMSO-d6) δ ppm 1.25 (s, 12 H), 2.19 (s, 1 H), 5.13 (s, 2 H), 6.86
(dd, J ) 8.6, 2.2 Hz, 1 H), 6.99 (d, J ) 2.0 Hz, 1 H), 7.29–7.43
(5-Hydroxy-1-(2-morpholinoethyl)-1H-indol-3-yl)(2,2,3,3-tet-
ramethylcyclopropyl)methanone (35). Compound 33 (0.15 g, 0.41
mmol), Cs2CO3 (0.4 g, 1.2 mmol), and CH3I (51 µL, 0.61 mmol)
in 5 mL of DMF were combined and stirred at ambient temperature
for 72 h. The mixture was quenched with 3 mL of saturated,
aqueous NH4Cl and diluted with 5 mL of EtOAc. The layers were
separated, and the aqueous layer was extracted with 3 × 3 mL of
EtOAc. The combined organic extracts were washed with 1 × 5
mL of saturated aqueous NaCl, dried over anhydrous Na2SO4,
filtered, concentrated under reduced pressure, and recrystallized with
4:1 hexanes/EtOAc to provide 35 (75 mg, 0.20 mmol, 48% yield).
1H NMR (MeOH-d4, 300 MHz) δ ppm 1.33 (s, 12 H), 2.10 (s, 1
H), 2.47–2.53 (m, 4 H), 2.77 (t, J ) 6.4 Hz, 2 H), 3.63–3.69 (m,
4 H), 3.84 (s, 3 H), 4.33 (t, J ) 6.4 Hz, 2 H), 6.89 (dd, J ) 8.8,
2.7 Hz, 1 H), 7.38 (d, J ) 8.8 Hz, 1 H), 7.81 (d, J ) 2.4 Hz, 1 H),
8.06 (s, 1 H); MS (DCI/NH3) m/z 385 (M + H)+; Anal.
(C23H32N2O3) C, H, N.

Indol-3-yl-tetramethylcyclopropyl Ketones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1911
Glia in Alzheimer’s Disease Brains. J. Neurosci. 2003, 23, 11136–
11141.
(5-(Benzyloxy)-1-(2-morpholinoethyl)-1H-indol-3-yl)(2,2,3,3-
tetramethylcyclopropyl)methanone (37). The (5-benzyloxy-1H-
indol-3-yl)-(2,2,3,3tetramethyl-cyclopropyl)methanone (from the
procedure for 24) (1.1 g, 3.0 mmol), 2-morpholin-4-ylethyl meth-
anesulfonate (5.1 mmol), and NaH (60% dispersion in mineral oil,
0.36 g, 9.1 mmol) in 25 mL of DMF were processed as described
in the procedure for 5 to provide 37 (1.2 g, 2.6 mmol, 86% yield).
1H NMR (300 MHz, CDCl3) δ ppm 1.31 (s, 6 H), 1.36 (s, 6 H),
1.90 (s, 1 H), 2.39–2.59 (m, 4 H), 2.72–2.87 (m, 2 H), 3.63–3.81
(m, 4 H), 4.13–4.31 (m, 2 H), 5.14 (s, 2 H), 7.01 (dd, J ) 9.0, 2.5
Hz, 1 H), 7.22–7.28 (m, 1 H), 7.29–7.44 (m, 3 H), 7.45–7.52 (m,
2 H), 7.75 (s, 1 H), 8.07 (d, J ) 2.4 Hz, 1 H); MS (DCI/NH3) m/z
461 (M + H)+; Anal. (C23H31NO2) C, H, N.
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