Identification of potent, highly constrained CGRP receptor antagonists

Article abstract of DOI:10.1016/j.bmcl.2010.02.086

A novel series of potent CGRP receptor antagonists containing a central quinoline ring constraint was identified. The combination of the quinoline constraint with a tricyclic benzimidazolinone left hand fragment produced an analog with picomolar potency (

Full text of DOI:10.1016/j.bmcl.2010.02.086

Bioorganic & Medicinal Chemistry Letters 20 (2010) 2572–2576
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Identification of potent, highly constrained CGRP receptor antagonists
a
a
a
a
b
Craig A. Stump a, , Ian M. Bell , Rodney A. Bednar , John F. Fay , Steven N. Gallicchio , James C. Hershey ,
Richard Jelley a, Constantine Kreatsoulas c, Eric L. Moore d, Scott D. Mosser a, Amy G. Quigley a,
Shane A. Roller e, Christopher A. Salvatore d, Steven S. Sharik a, Cory R. Theberge a, C. Blair Zartman a,
Stefanie A. Kane d, Samuel L. Graham a, Harold G. Selnick a, Joseph P. Vacca a, Theresa M. Williams a
*
a Department of Medicinal Chemistry, Merck & Co., Inc., PO Box 4, West Point, PA 19486, USA
b Department of Central Pharmacology, Merck & Co., Inc., PO Box 4, West Point, PA 19486, USA
c Department of Chemistry Modeling and Informatics, Merck & Co., Inc., PO Box 4, West Point, PA 19486, USA
d Department of Pain Research, Merck & Co., Inc., PO Box 4, West Point, PA 19486, USA
e Department of Drug Metabolism, Merck & Co., Inc., PO Box 4, West Point, PA 19486, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A novel series of potent CGRP receptor antagonists containing a central quinoline ring constraint was
identified. The combination of the quinoline constraint with a tricyclic benzimidazolinone left hand frag-
ment produced an analog with picomolar potency (14, CGRP Ki = 23 pM). Further optimization of the tri-
cycle produced a CGRP receptor antagonist that exhibited subnanomolar potency (19, CGRP Ki = 0.52 nM)
and displayed a good pharmacokinetic profile in three preclinical species.
Received 17 January 2010
Revised 19 February 2010
Accepted 22 February 2010
Available online 25 February 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
CGRP
Migraine
Calcitonin gene-related peptide
Migraine is a disabling neurovascular disorder that affects
approximately 13% of the general population.1 A migraine attack
is characterized by unilateral head pain and is often accompanied
by phonophobia, photophobia, and nausea. Calcitonin gene-related
peptide (CGRP), a 37 amino acid neuropeptide, has been implicated
in the pathogenesis of migraine, and therefore, antagonism of the
CGRP receptor has been proposed as a novel approach for migraine
therapy.2 Clinical studies have demonstrated the effectiveness of
CGRP receptor antagonists for the acute treatment of migraine
with both the intravenously-administered olcegepant3 and the or-
ally-administered telcagepant.4 Furthermore, recently published
Phase III clinical results have shown telcagepant 300 mg to have
similar efficacy to a positive control (zolmitriptan 5 mg), with few-
er reported adverse events.5 Our research program continued to fo-
cus on developing non-peptidic, orally bioavailable CGRP receptor
antagonists as novel therapeutics.
spirohydantoin fragment with a spiroazaoxindole further im-
proved potency to provide 3.8 While compounds 2 and 3 are very
potent CGRP receptor antagonists, they exhibit poor overall phar-
macokinetic profiles. Calculated polar surface area (PSA) has been
shown to correlate with intestinal permeability.9 In a related series
of spirohydantoin compounds, an inverse relationship between
PSA and bioavailability has been observed.6 In an effort to reduce
the PSA of 2 (158 Å2) and 3 (149 Å2), replacement of the central
amide linker, one area of the molecules that had thus far eluded
modification, was investigated.
A torsion plot with respect to rotation around the central amide
of 1 indicated a low energy conformation that was planar with re-
spect to the amide carbonyl and aryl group of the spiroindane (i.e.,
U
= 0° or 180°). Substitution on the aryl group at the positions
ortho to the amide linker was predicted to bias one of two possible
planar conformations. To that end, the o-fluoro analogs 4 and 5
were synthesized, with each predicted to favor only one of the
low energy, planar conformations by approximately 3 kcal/mol
(Fig. 2). The similar potency of compounds 1 (Ki = 20 nM) and 4
A previous communication from these laboratories described
the discovery of novel CGRP receptor antagonist 1 (Fig. 1).6 Subse-
quent communications have described the evolution of both the
left and right hand fragments of 1 to produce more potent antago-
nists. Optimization of the benzimidazolinone fragment provided
the more potent tricyclic analog 2;7 whereas replacement of the
(Ki = 55 nM), and the loss of potency with compound
5
(Ki = 720 nM), suggested that the bioactive conformation with re-
spect to the central amide was the one depicted for structure 4
in Figure 2 (i.e.,
U = 0°).
In order to further elucidate the preferred binding mode of com-
pounds like 1, we sought to incorporate a constraint that would
* Corresponding author. Tel.: +1 215 652 1775; fax: +1 215 652 7310.
E-mail address: craig_stump@merck.com (C.A. Stump).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.02.086

C. A. Stump et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2572–2576
2573
mimic the proposed bioactive conformation of 1. Evaluation of the
two benzimidazole analogs 6 (Ki = 320 nM) and 7 (Ki = 6600 nM)
provided further evidence for the proposed bioactive conforma-
tion, albeit with the loss of activity observed with 6 suggesting
the presentation of the left hand fragment was suboptimal
(Fig. 3). The benzoxazole 8 suffered an additional loss of potency
(Ki = 6000 nM) compared to the corresponding benzimidazole 6
(Ki = 320 nM). It was anticipated that expansion of the constraint
to a six membered ring might better approximate the geometry
of the central amide of 1. Indeed, when the racemic quinoxaline
9 was evaluated, a modest increase in potency was observed
(Ki = 185 nM). The quinoline analog 10 exhibited an even greater
improvement in potency, with a Ki = 17 nM making it equipotent
to amide 1 and providing further support for the proposed bioac-
tive conformation.
The quinoline constraint was evaluated with a number of left
hand fragments, both with the spiroazaoxindole and spirohydan-
toin moieties (Table 1). In the related amide series of compounds,
the spiroazaoxindole imparted a significant improvement in po-
tency over the corresponding hydantoins.8 However, we antici-
pated that the more polar spirohydantoin moiety might help
improve the physical properties (e.g., solubility, protein binding)
of the lipophilic quinoline analogs, as their calculated log D7.4
(ACD/Labs 11.0) was typically at least one log unit lower than for
the corresponding spiroazaoxindoles. The simple 3,4-dihydro-2-
quinolinone 11 exhibited similar potency to the lead compound
10 (Ki = 8.9 nM). As anticipated, replacement of the spirohydantoin
with the spiroazaoxindole (12) provided an order of magnitude in-
crease in potency in both the binding and cell-based assays.
Combination of the quinoline with an optimized tricyclic benzim-
idazolinone7 produced subnanomolar CGRP receptor antagonist 13
(Ki = 0.30 nM). Once again, incorporation of the spiroazaoxindole
produced a significant increase in potency, with the corresponding
compound 14 exhibiting picomolar potency (Ki = 23 pM) in the
binding assay. Unfortunately, compound 14 showed no bioavail-
ability when dosed in rats (Table 2). In an effort to further reduce
the PSA of 14 (130 Å2), modification or deletion of one of the
remaining polar amides was investigated. Modification of either
the spirohydantoin or azaoxindole has been shown to drastically
reduce the potency of related compounds.8 Therefore, effort was
focused on modification of the amides of the tricyclic benzimidaz-
olinone rings. Simple methylation of the secondary amide in the
tricycle caused a marked loss of potency (15 and 16 vs 13 and
14), as well as a more pronounced serum shift in the functional
assay. Replacement of the benzimidazolinone with an indoline
served to reduce PSA and introduce a weakly basic amine into
the structure, which we hoped would improve solubility in acidic
environments.10 Indolines 17 (PSA = 116 Å2) and 18 (PSA =
108 Å2, data for most potent epimers shown) showed similar po-
tency in the binding assay to 15 and 16, however, the serum shift
observed in the functional assay was attenuated. Gratifyingly,
these indoline analogs demonstrated improved bioavailability
and reduced intrinsic clearance (Table 2) in the rat, especially in
the spirohydantoin series (rat F = 41%, Cl = 11 mL/min/kg). Addi-
tion of a 3-methyl substituent to the indoline produced analogs
19 and 20, which exhibited similar potency to the benzimidazo-
lone tricycles 13 and 14. Fortunately, the improved pharmacoki-
netic profile was maintained with addition of the methyl group.
Homologation of the 3-indoline substituent to an ethyl group pro-
vided no potency advantage (21 and 22), and in the case of spiro-
hydantoin 21 increased the serum shift in the functional assay
(IC50 = 44 nM).
The combination of the indoline tricycle with the spirohydan-
toin quinoline provided compounds with superior bioavailability
in rats (F = 34–65%) compared to the corresponding spiroazaoxin-
dole quinolines (F = 6–9%). Interestingly, it was also noted that
the difference in potency between the spirohydantoins 17 and 19
and spiroazaoxindoles 18 and 20 was somewhat less than the or-
der of magnitude we had typically observed. Based on the combi-
nation of potency and rat pharmacokinetics, compound 19 was
selected for more in-depth evaluation.
The sodium salt of compound 19 displayed a good overall phar-
macokinetic profile in rats, dogs, and rhesus monkeys (Table 3),
with low clearance in all three species, ranging from 1.6 mL/min/
Figure 1. Previously disclosed, potent CGRP receptor antagonists.
Figure 2. Plot of torsional energy strain for rotation about the central amide (C1–N–C2–C3) of 4 (solid line) and 5 (dashed line).

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C. A. Stump et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2572–2576
Figure 3. CGRP receptor antagonists with central constraints.
kg (dog) to 14 mL/min/kg (rat). After an intravenous dose, the plas-
ma half-life ranged from 1.6 h (rat) to 6.1 h (dog). The oral bioavail-
ability was good (34–78%) in all three species in a number of
different dosing vehicles. The good bioavailability observed when
19 was dosed as a suspension in methylcellulose (F = 38–59%) indi-
cates that poor aqueous solubility is not greatly limiting its absorp-
tion. In fact, only marginal effects were observed when 19 was
dosed in more solubilizing formulations, such as Imwitor:Tween
(rat F = 34%) or PEG400 (dog F = 78%, rhesus F = 44%).
Compound 19 was further tested in a primate model to evaluate
its ability to act in vivo as a CGRP receptor antagonist. When dosed in
rhesus monkeys in the capsaicin-induced dermal vasodilation phar-
Table 1
Comparison of CGRP receptor antagonist activity of spriroazaoxindole and spirohydantoin
O
O
N
N
A
NH
A
NH
N
N
O
Me
cAMP IC50 + HSa,d
(nM)
Compound CGRP Kia,b (nM)
cAMP IC50 (nM)
cAMP IC50 + HSa,d
(nM)
a,c
a,c
A
Compound CGRP Kia,b (nM) cAMP IC50
(nM)
O
N
11
13
8.9 (2)
50 (1)
850 (1)
12
14
1.4 0.33 (13)
4.7 1.6 (3)
81 61 (3)
O
N
N
0.30 (1)
1.7 0.55 (3)
14 11 (3)
0.023 0.0032 (4)
0.077 0.024 (3)
2.5 0.27 (3)
O
HN
O
N
N
15
3.0 0.25 (4)
1.5 0.21 (7)
33 20 (3)
7.3 4.6 (5)
1200 380 (3)
16
0.41 (2)
1.2 (1)
95 (1)
O
N
H3C
N
N
N
O
17
19
43 20 (5)
6.5 1.3 (4)
18
20
0.55 0.35 (12)
0.12 0.030 (4)
0.86 0.38 (9)
0.52 0.18 (3)
4.1 1.9 (9)
HN
H3C
0.52 0.068 (4) 2.2 0.42 (4)
1.3 0.27 (3)
O
HN
H3C
21
1.3 (1)
7.2 (2)
44 (2)
22
0.072 0.015 (4)
0.68 0.077 (3)
1.5 0.35 (3)
O
HN
a
b
c
Mean value standard deviation, where appropriate; number of replicates in parentheses.
Ki values for inhibition of 125I-hCGRP binding determined using membranes from HEK293 cells stably expressing human CLR/RAMP1.5
Inhibition of CGRP-induced cAMP production in HEK293 cells stably expressing human CLR/RAMP1.5
d
Inhibition of CGRP-induced cAMP production in HEK293 cells stably expressing human CLR/RAMP1in the presence of 50% human serum.5

C. A. Stump et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2572–2576
2575
The synthesis of compound 19 is shown in Scheme 1.11 Alkyl-
ation of dimethylmalonate with 2-chloro-1,3-dinitrobenzene, fol-
lowed by decarboxylation produced methyl (2,6-dinitrophenyl)
acetate. Further alkylation with ethyl bromoacetate provided the
2,6-dinitrophenyl substituted succinic diester, which underwent
reduction of the nitro groups and monocyclization upon treatment
with iron and ammonium chloride. The tricyclic core was formed
upon heating in xylenes and acetic acid. The 3-methyl substituent
was installed by alkylation with methyl iodide and sodium hy-
dride. Selective reduction of the oxindole carbonyl was achieved
in good yield upon treatment with DIBAH. Chiral resolution on a
ChiralPak AD column produced the tricyclic indoline intermediate
in eight total steps and overall 10% yield. The quinoline aldehyde
was produced in a two step sequence from the known spirohydan-
toin aniline.6 A Skraup-type reaction between the aniline and cro-
tonaldehyde, using p-chloranil as the oxidant, produced the
2-methylquinoline in good yield.12 Oxidation of the methyl group
with selenium dioxide produced the desired aldehyde. The final
reductive alkylation between the tricyclic indoline and quinoline
aldehyde was carried out using sodium cyanoborohydride in meth-
anol to produce 19.
Table 2
Pharmacokinetics of compounds in rats
Compound po dose
(mpk)
iv dosea
(mpk)
Fb
(%)
iv t1/2
(h)
Cl (mL/min/
kg)
11
12
14
17
18
19
20
21
22
10
10
5
10
5
10
10
10
10
2
2
1
2
1
2
2
2
2
10I
3I
0.70
0.35
0.45
1.1
0.32
1.6
0.62
1.1
0.61
38
56
40
11
19
14
28
44
11
0II
41III
8III
34III
6III
65I
9III
a
Compound dosed in DMSO.
Compound dosed orally in (I) 1% methylcellulose, or (II) 90% PEG 400:10%
b
water, or (III) 50% Imwitor:50% Tween.
Table 3
Pharmacokinetics of compound 19 (sodium salt)
Species po dose
(mpk)
iv dosea
(mpk)
Fb
(%)
iv t1/2
(h)
Cl (mL/min/
kg)
In conclusion, good evidence for the bioactive conformation of
CGRP receptor antagonists linked by a central amide was provided
by the use of conformational constraints. In particular, replace-
ment of the central amide with a quinoline ring produced a novel
series of potent CGRP receptor antagonists. Furthermore, the com-
bination of the quinoline constraint with a tricyclic benzimidazoli-
none left hand fragment produced analogs with picomolar potency
in both the radioligand binding assay and the cell-based functional
assay. Further optimization of the tricycle produced a subnanomo-
lar CGRP receptor antagonist (19) that also exhibited good in vivo
potency in a pharmacodynamic assay in rhesus monkeys. In addi-
tion, 19 displayed a good pharmacokinetic profile in three preclin-
ical species.
Rat
Dog
Rhesus
10
1
2
2
0.5
0.5
45
59
38
1.6
6.1
1.7
14
1.6
5.6
a
Compound dosed in DMSO.
Compound dosed orally in 1% methylcellulose.
b
macodynamic assay,4c plasma levels of 5.5
to inhibit ninety percent of the capsaicin-induced vasodilation re-
sponse (EC90 = 5.5 M). Thisvalue, although slightlyless potentthan
the number obtained for the clinical compound telcagepant, demon-
strated in vivo potency for this class of CGRP receptor antagonists.
lM of 19 were required
l
Scheme 1. Synthesis of compound 19. Reagents and conditions: (a) CH2(CO2Me)2, KOt-Bu, DMA, 90 °C, 90%; (b) LiCl, H2O, DMSO, 100 °C, 99%; (c) BrCH2CO2Et, Cs2CO3, DMF,
75%; (d) Fe, NH4Cl(aq), EtOH, 90 °C, 95%; (e) AcOH, xylenes, 140 °C, 75%; (f) NaH, CH3I, DMF, 0 °C, 65%; (g) DIBAH, THF, 0 °C, 70%; (h) crotonaldehyde, p-chloranil, 1-BuOH, HCl,
120 °C, 79%; (i) SeO2, dioxane, 100 °C, 87%; (j) NaBH3CN, AcOH, MeOH, 86%.

2576
C. A. Stump et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2572–2576
6. Bell, I. M.; Bednar, R. A.; Fay, J. F.; Gallicchio, S. N.; Hochman, J. H.; McMasters,
Acknowledgments
D. R.; Miller-Stein, C.; Moore, E. L.; Mosser, S. D.; Pudvah, N. T.; Quigley, A. G.;
Salvatore, C. A.; Stump, C. A.; Theberge, C. R.; Wong, B. K.; Zartman, C. B.; Zhang,
X.-F.; Kane, S. A.; Graham, S. L.; Vacca, J. P.; Williams, T. M. Bioorg. Med. Chem.
Lett. 2006, 16, 6165.
The authors thank Gary Sitko, Kimberly Michel, and Anne Taylor
for animal dosing; Ken Anderson and Cynthia Miller-Stein for PK
analytical data; the MRL West Point analytical chemistry, mass
spectrometry, and NMR spectroscopy groups for analytical and
spectroscopic data; and Donnette Staas for helpful discussions.
7. Theberge, C. R.; Bednar, R. A.; Bell, I. M.; Corcoran, H. A.; Fay, J. F.; Hershey, J. C.;
Johnston, V. K.; Kane, S. A.; Mosser, S.; Salvatore, C. A.; Williams, T. M.; Zartman,
C. B.; Zhang, X.-F.; Graham, S. L.; Vacca, J. P. Bioorg. Med. Chem. Lett. 2008, 18,
6122.
8. Stump, C. A.; Bell, I. M.; Bednar, R. A.; Bruno, J. G.; Fay, J. F.; Gallicchio, S. N.;
Johnston, V. K.; Moore, E. L.; Mosser, S.; Quigley, A. G.; Salvatore, C. A.;
Theberge, C. R.; Zartman, C. B.; Zhang, X.-F.; Kane, S. A.; Graham, S. L.; Vacca, J.
P.; Williams, T. M. Bioorg. Med. Chem. Lett. 2009, 19, 214.
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