N-[3-(2-dimethylaminoethyl).2-methyl-1H-indol-5-yl]-4-fluorobenzamide: A potent, selective, and orally active 5-HT1F receptor agonist potentially useful for migraine therapy

Article abstract of DOI:10.1021/jm0155190

Recent studies have demonstrated that selective 5-HT_1F receptor agonists inhibit neurogenic dural inflammation, a model of migraine headache, indicating that these compounds may be effective therapies for the treatment of migraine pain. This communication describes the synthesis and discovery of a novel compound, N-[3-(2-(dimethylamino)-ethyl)-2-methyl-1H-indol-5-yl]-4-fluorobenzamide (4), which possesses high binding affinity and selectivity at the 5-HT_1F receptor relative to more than 40 other serotonergic and nonserotonergic receptors examined.

Full text of DOI:10.1021/jm0155190

J . Med. Chem. 2001, 44, 4031-4034
4031
discovering effective migraine therapeutics. Sumatrip-
tan has moderate 5-HT1F receptor agonist activity (Ki
) 25.7 nM)5 and is active in the neurogenic dural
inflammation model (Figure 1). While the antimigraine
effect of sumatriptan may be occurring via 5-HT1F
receptor activation, the cardiovascular liabilities of this
agent most likely result from coronary arterial vaso-
constriction,6 mediated via a receptor other than the
5-HT1F receptor.4b This finding has stimulated consider-
able interest recently in the identification of potent
5-HT1F receptor agonists without cross-reactivity at the
5-HT1B receptor or 5-HT1D receptor, which could ulti-
mately lead to safer migraine therapy.7,8
Our effort in the search for more potent and selective
5-HT1F receptor agonists was directed toward the
tryptamine core structure. On the basis of the structure
features of compound 1 (LY334370),8 which has an
arylamido functional group at the C-5 position of the
indole, we introduced the same substituent at a similar
position of tryptamine, leading to compound 3 (Table
1). This analogue had a 5-HT1F affinity (Ki ) 3.8 nM)
higher than that of either serotonin (Ki ) 10 nM) or
sumatriptan (Ki ) 25.7 nM).5 Unfortunately, compound
3, like sumatriptan, lacked selectivity versus other
5-HT1 receptor subtypes.
A second modification was introduced by installation
of a methyl group at the C-2 position of the indole
nucleus, leading to a new analogue 4 (LY349950). A
reduction in binding affinity at the 5-HT1 receptor
subtypes would be expected on the basis of a report12
that 2-methyl-5-HT is more than 40-fold less potent in
binding than 5-HT itself. To our surprise, 4 retained
high affinity at the 5-HT1F receptor (Ki ) 8.2 nM). As
expected, the affinities for other 5-HT1 receptor subtypes
dropped significantly. The combination of these two
modifications on the aryl-substituted tryptamine core
led to the discovery of 4, a potent, selective, and orally
active 5-HT1F receptor agonist with a long duration of
action.
Ch em istr y. Compound 4 was prepared in a four-step
sequence shown in Scheme 1 starting from 4-nitroa-
niline 5. Coupling reaction of 5 with 4-fluorobenzoyl
chloride 6 gave an arylamide intermediate whose nitro
group was then reduced by hydrogenation to afford
compound 7. Diazotization of aniline 7 with sodium
nitrite in HCl solution followed by an in situ reduction
using SnCl2 yielded arylhydrazine 8 in good yield.9
When the Fischer indole synthesis is applied,10 the
reaction of hydrazine 8 with 1-N,N-dimethylaminopen-
tan-4-one 9 gave rise to 2,3,5-trisubstituted indole 4 in
good overall yield (44% from 5). It should be noted that
the Fischer indolization with an asymmetrically sub-
stituted ketone usually gives a mixture of two possible
indole products.11 The reaction of 8 with asymmetrically
substituted ketone 9, however, is chemoselective to give
only 4. No other possible indole product was isolated.
N-[3-(2-Dim eth yla m in oeth yl)-2-m eth yl-1H-
in d ol-5-yl]-4-flu or oben za m id e: A P oten t,
Selective, a n d Or a lly Active 5-HT1F
Recep tor Agon ist P oten tia lly Usefu l for
Migr a in e Th er a p y
Yao-Chang Xu,*,† Kirk W. J ohnson,† Lee A. Phebus,†
Marlene Cohen,† David L. Nelson,† Kathy Schenck,†
Clint D. Walker,† J ames E. Fritz,†
Stephen W. Kaldor,† Michael E. LeTourneau,†
Robert E. Murff,† J ohn M. Zgombick,‡
David O. Calligaro,† J ames E. Audia,† and
J ohn M. Schaus†
Lilly Research Laboratories, Eli Lilly and Company,
Indianapolis, Indiana 46285, and Synaptic Pharmaceutical
Corporation, 215 College Road, Paramus, New J ersey 07652
Received April 3, 2001
Abstr a ct: Recent studies have demonstrated that selective
5-HT1F receptor agonists inhibit neurogenic dural inflamma-
tion, a model of migraine headache, indicating that these
compounds may be effective therapies for the treatment of
migraine pain. This communication describes the synthesis
and discovery of a novel compound, N-[3-(2-(dimethylamino)-
ethyl)-2-methyl-1H-indol-5-yl]-4-fluorobenzamide (4), which
possesses high binding affinity and selectivity at the 5-HT1F
receptor relative to more than 40 other serotonergic and
nonserotonergic receptors examined.
Serotonin (5-hydroxytryptamine, 5-HT), a neurotrans-
mitter widely distributed in the brain and peripheral
tissues, is involved in the regulation of various physi-
ological functions such as mood, appetite, pain, sexual
behavior, blood pressure, and body temperature.1 Recent
advances in the molecular cloning of seven serotonin
receptor subfamilies (5-HT1, 5-HT2, 5-HT3, 5-HT4, 5-HT5,
5-HT6, and 5-HT7) have provided a plethora of poten-
tially important targets for drug discovery research.
Among these 5-HT receptors, the 5-HT1 receptor family
appears to be the most complex and has been further
subclassified into the 5-HT1A, 5-HT1B, 5-HT1D, 5-HT1E
,
and 5-HT1F subtypes.2
Sumatriptan (1) is the first 5-HT1 receptor agonist
approved for the clinical treatment of patients with
migraine headaches.3 Since the introduction of this and
other structurally related triptans, their exact mecha-
nism of action has been the subject of debate.2f,4 For
some time, 5-HT1B/1D receptor activation had been
believed to be involved in mediating the therapeutic
effects4c partly because triptans have high binding
affinity for these receptor subtypes (Table 1). Phebus
and colleagues have recently demonstrated that agonist
potency to inhibit trigeminal-stimulated dural plasma
extravasation in the guinea pig is highly correlated with
their affinities for the 5-HT1F receptor.5 This finding
indicates that activation of the 5-HT1F receptor instead
of the 5-HT1B/1D receptor could be the rational way for
In Vitr o a n d in Vivo P h a r m a cologica l Resu lts.
The affinity of 4 for cloned human serotonin receptors
in vitro (Tables 2 and 3) was determined by radioligand
binding studies using membranes from transfected
cells.12-14 Studies examining compound 4 binding to the
* To whom correspondence should be addressed. Phone: (317) 277-
2284. Fax: (317) 277-1125. E-mail: Xu_Yao-Chang@Lilly.com.
† Eli Lilly and Company.
‡ Synaptic Pharmaceutical Corporation.
10.1021/jm0155190 CCC: $20.00 © 2001 American Chemical Society
Published on Web 10/23/2001

4032 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 24
Letters
Ta ble 1. Binding Affinity and Selectivity of 5-HT1F Receptor Agonists at Human 5-HT1 Receptors
a
b
The Ki values are expressed as the mean values ( SEM of two to eight experiments. The Ki valus for 5-HT1A, 5-HT1B, and 5-HT1D
used for calculation of the selectivity are derived from at least two separate experiments.
Sch em e 1. Synthesis of N-[3-(2-Dimethylaminoethyl)-2-methyl-1H-indol-5-yl]-4-fluorobenzamide 4
native rat adrenergic, dopaminergic, benzodiazepine,
GABA, histamine H1, and muscarinic receptors were
conducted using methods referenced in Table 2. All
other binding assays (Table 3) were conducted by
Novascreen (Hanover, MD) using their protocols.
As shown in Tables 2 and 3, 4 possessed high affinity
and selectivity for the 5-HT1F receptor. With Ki being
8.2 nM at the 5-HT1F receptor, 4 displayed more than
32-fold selectivity versus the 44 binding sites examined.
Minimizing the affinity at the 5-HT1B and 5-HT1D
receptors was most important to potentially avoid the
cardiovascular side effects associated with current trip-
tan agents. 4 showed greater than 100-fold selectivity
for the 5-HT1F receptor relative to both 5-HT1B and
5-HT1D receptors.
Ta ble 2. Binding Profile of 4 to Neurotransmitter Receptor
Sites
a
binding site
5-HT1F
5-HT1A
5-HT1B
5-HT1D
5-HT1E
5-HT2A
5-HT2B
5-HT2C
5-HT4
5-HT6
5-HT7
R1-adrenergic
R2-adrenergic
â-adrenergic
dopamine D1
dopamine D2
Ki (nM)
species
radioligand
8.2 ( 1.2
265 ( 99
human
human
[3H] serotonin12
[3H] serotonin13
[3H] serotonin14
[3H] serotonin14
[3H] serotonin13
1060 ( 204 human
1620 ( 100 human
>4830
1000 ( 85
676 ( 118
human
human
human
[
125I] DOI15
[3H] serotonin15
2200 ( 390 human
[
125I] DOI15
380 ( 40
human
[3H] serotonin
[3H] LSD
1770 ( 130 human
1460 ( 102 human
[3H] serotonin16
[3H] prazosin17
[3H] rauwolscine18
[3H] DHA19
15600
6000
38000
>100000
38000
rat
rat
rat
rat
rat
rat
rat
rat
rat
[3H] SCH2339020
[3H] raclopride21
[3H] flunitrazepam22
[3H] muscimol23
[3H] pyrilamine24
[3H] QNB25
To determine the functional properties of 4, the
compound was evaluated for its ability to inhibit for-
skolin-stimulated adenylate cyclase in cell lines express-
ing the human 5-HT1F receptor. Responses were com-
pared to those elicited by 5-HT and sumatriptan.12 As
benzodiazepine >100000
GABA
histamine H1
muscarinic
>10000
12000
>100000
shown in Table 4, compound 4 was a potent (EC50
)
a
The Ki values are expressed as the mean ( SEM of two to
eight experiments. Single values indicate the average results of
two separate binding experiments.
6.0 nM) 5-HT1F receptor agonist (Emax ) 98%). The rank
order of agonist potency in this functional assay was 4
≈ 5-HT > sumatriptan.
Neurogenic dural extravasation and subsequent in-
flammation have been used as an animal model of
migraine. Inhibition of the inflammation by compounds
has been suggested to predict their clinical efficacy in

Letters
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 24 4033
Ta ble 3. Low Binding Activity (Ki > 1000 nM) of 4 at
Additional Sites
5-HT4
melatonin
kainate
opoid
angiotensin i
angiotensin ii
arg-vasopressin 1
arg-vasopressin 2
substance P
NK-3
somatostatin
neurotensin
VIP
galanin
calcium channels
(T and L)
5-HT uptake
NE uptake
DA uptake
adenosine
GABAA
GABAB
histamine
(H2 and H3)
nicotinic
bradykinin
chloride channels
CCK central
potassium channels
Ta ble 4. Functional Properties of 4, Serotonin, and
Sumatriptan at the Cloned Human 5-HT1F Receptor in Vitro
F igu r e 2. Duration of action of 4‚HCl and sumatriptan to
inhibit trigeminal stimulation induced dural extravasation in
the guinea pig.
a
a
compound
EC50 (nM)
Emax (% of 5-HT)
4
6.0 ( 0.7
7.9 ( 0.8
35 ( 5.0
98 ( 1.4
100
98 ( 2.0
serotonin12b
sumatriptan12b
a
EC50 and Emax values are expressed as mean values ( SEM
from at least two separate experiments performed in triplicate.
F igu r e 3. Effects of 4‚HCl and sumatriptan to contract the
rabbit saphenous vein.
contrast, 4 was fully effective at 1 h and remained
partially effective at 24 h after oral administration.
Sumatriptan is a known vasoconstrictor and was
developed on the basis of the vascular theory of mi-
graine.29 It is not known which serotonin receptor (5-
HT1B, 5-HT1D, or 5-HT1D-like) mediates the vasocon-
striction,4b which might be related to the cardiovascular
side effects of the drug. 4 was examined in vitro and
compared to sumatriptan using the isolated rabbit
saphenous vein to evaluate contractile activity (Figure
3). While sumatriptan produced concentration-depend-
ent contractile activity in this tissue, 4 did not constrict
this tissue (up to 10-4 M). These in vitro results suggest
that 4 does not possess the vasoconstrictive properties
in rabbit saphenous vein that might be potentially
linked to the cardiovascular side effects associated with
sumatriptan.6
F igu r e 1. Dose response of 4‚HCl and sumatriptan to inhibit
trigeminal stimulation induced dural extravasation in guinea
pig.
treating acute migraine.26 Several compounds such as
sumatriptan, naratriptan, and rizatriptan, being active
in the dural extravasation model,5 were eventually
found to be effective drugs for migraine with the
exception of CP-122288.27 4 was examined for its ability
to inhibit neurogenic dural inflammation following
either oral or intravenous administration in guinea
pigs.28 As shown in Figure 1, 4 potently inhibited
neurogenic dural inflammation by both routes with an
intravenous ID50 value of 1.4 ng/kg and an oral ID50
value of 4.3 ng/kg. Sumatriptan was less potent than 4
with ID50 values of 15 and 4600 ng/kg under the same
conditions following intravenous and oral administra-
tion, respectively.
In conclusion, 4 represents the discovery of a novel,
potent, and selective 5-HT1F receptor agonist. The
compound lacks the vasoconstriction properties in the
isolated rabbit saphenous vein and has good oral activity
and a long duration of action in the guinea pig neuro-
genic dural inflammation model of migraine.
Sumatriptan displayed a high oral-to-intravenous
ratio of ID50 values (4600/15) in the dural inflammation
model in guinea pig. In contrast 4 displayed a much
lower ratio (4.3
/1.4), indicative of the high oral bioavail-
Ack n ow led gm en t. We thank Mr. J ames D. Mar-
shall for running the large-scale hydrogenation reaction,
and Dr. Charles H. Mitch for proof-reading of the
manuscript.
ability of this compound. Clinically, two doses of
sumatriptan are often required to treat a migraine
attack. Figure 2 compares the duration of action for both
sumatriptan and 4 using the ID100 value following oral
administration from the dural inflammation model in
the guinea pig. Sumatriptan was fully effective at 1 h
but no longer active at 4 h after administration. In
Su p p or tin g In for m a tion Ava ila ble: NMR and MS data
for compounds 3 and 4. This material is available free of charge
via the Internet at http://pubs.acs.org.

4034 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 24
Letters
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