Discovery of a novel class of benzazepinone Nav1.7 blockers: Potential treatments for neuropathic pain

Article abstract of DOI:10.1016/j.bmcl.2007.05.076

A series of benzodiazepines and benzazepinones were synthesized and evaluated as potential sodium channel blockers in a functional, membrane potential-based assay. One member of the benzazepinone series, compound 47, displayed potent, state-dependent block of hNa_v1.7, and was orally efficacious in a rat model of neuropathic pain.

Full text of DOI:10.1016/j.bmcl.2007.05.076

Bioorganic & Medicinal Chemistry Letters 17 (2007) 4630–4634
Discovery of a novel class of benzazepinone Nav1.7
blockers: Potential treatments for neuropathic pain
Scott B. Hoyt,a,* Clare London,a David Gorin,a Matthew J. Wyvratt,a Michael H. Fisher,a
Catherine Abbadie,b John P. Felix,c Maria L. Garcia,c Xiaohua Li,a Kathryn A. Lyons,a
Erin McGowan,b D. Euan MacIntyre,b William J. Martin,b Birgit T. Priest,c Amy Ritter,b
McHardy M. Smith,c Vivien A. Warren,c Brande S. Williams,c
Gregory J. Kaczorowskic and William H. Parsonsa
aDepartment of Medicinal Chemistry, Merck Research Laboratories, Rahway, NJ 07065, USA
bDepartment of Pharmacology, Merck Research Laboratories, Rahway, NJ 07065, USA
cDepartment of Ion Channels, Merck Research Laboratories, Rahway, NJ 07065, USA
Received 18 April 2007; revised 22 May 2007; accepted 24 May 2007
Available online 27 May 2007
Abstract—A series of benzodiazepines and benzazepinones were synthesized and evaluated as potential sodium channel blockers in a
functional, membrane potential-based assay. One member of the benzazepinone series, compound 47, displayed potent, state-depen-
dent block of hNav1.7, and was orally efficacious in a rat model of neuropathic pain.
Ó 2007 Elsevier Ltd. All rights reserved.
Neuropathic pain is a chronic, debilitating pain state
that results from injury to the peripheral or central ner-
vous system. It affects more than 1.6 million people in
the United States, and can be triggered by a variety of
events or conditions, including diabetes, shingles, and
chemotherapy.1 Because few effective therapies exist, pa-
tients suffering neuropathic pain are often prescribed
anticonvulsants or topical anesthetics as treatment.
Optimized for other indications, these agents typically
offer only modest pain relief, and frequently elicit
dose-limiting CNS-based side effects such as sedation
and impairment.
the treatment of neuropathic pain, thereby providing
clinical validation for this approach (Fig. 1).5,6
Recent data from human genetic studies have implicated
hNav1.7, a subtype located primarily in the PNS, as a
key constituent in pain signaling. Individuals with gain
of function mutations in SCN9A, the gene that encodes
hNav1.7, experience bouts of intense pain that are either
evoked by mild stimuli or spontaneous in nature.7,8 As
such, their symptoms resemble those presented by neuro-
pathic pain patients. In contrast, individuals with loss of
function mutations in SCN9A have a complete inability
to sense pain, yet retain all other normal nerve func-
tions.9,10 Collectively, these studies provide compelling ge-
netic validation for hNav1.7 as an important pain target.
Neuropathic pain signaling begins with the aberrant fir-
ing of action potential bursts in damaged nerve tissue.
The initiation and propagation of these action potentials
typically require the opening of voltage-gated sodium
channels (Nav1.x). Because they can inhibit action po-
tential firing, Nav1 blockers have been investigated as
treatments for neuropathic pain.2–4 Weak blockers such
as carbamazepine and lidocaine have shown efficacy in
Channel blockers destined for the clinic must inhibit
aberrant neuronal signaling while leaving normal nerve
O
H2N
O
H
N
O
N
N
N
N
H
O
N
NH
Boc
Keywords: Sodium channel; Neuropathic pain; Nav1.7; Benzazepi-
none; Benzodiazepine.
Carbamazepine
Lidocaine
1
*
Corresponding author. Tel.: +1 732 594 3753; fax: +1 732 594
5350; e-mail: scott_hoyt@merck.com
Figure 1. Sodium channel blockers.
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.05.076

S. B. Hoyt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4630–4634
4631
functions intact. We believe that this can be achieved, in
large part, via state-dependent channel block. Voltage-
gated sodium channels exist in three main conforma-
tional states: resting, open, and inactivated. In healthy
nerve and cardiac tissue, these channels reside predomi-
nantly in the resting state. In damaged nerve tissue, on
the other hand, they accumulate in the inactivated state.
Compounds that selectively bind and stabilize that inac-
tivated state should block signaling in damaged tissue
preferentially, thereby minimizing the potential for
mechanism-based adverse effects.
pact cardiac function. Because block of hERG K+ chan-
nels has been associated with potentially lethal
ventricular arrhythmias, compounds were tested in a
binding assay that measures displacement of 35S-labeled
MK-0499, a known hERG blocker.13 Similarly, because
block of Cav1.2 Ca2+ channels can lead to unsafe de-
creases in mean arterial pressure, compounds were
screened in a binding assay that measures displacement
3
of H-diltiazem (DLZ), a known Cav1.2 blocker.14
Initial SAR efforts were aimed at paring 1 down to the
minimum structure required for potent hNav1.7 block.
We began by varying the sidechain R1 substituent. As
shown in Table 1, when the R1 phenyl present in 1
was replaced with a cyclohexyl group (compound 9) or
deleted entirely (compound 8), a dramatic loss in
hNav1.7 potency was observed. If the alkyl tether con-
necting R1 to the molecule was shortened or lengthened
(10 and 11), potency was also diminished. While fluori-
nation of the R1 phenyl was tolerated (12 and 13),
replacement of the phenyl with various aromatic hetero-
cycles led to weaker channel block (14–16). Taken to-
gether, these data indicate that a lipophilic aromatic
group is required at this site.
Our goal is to develop potent, state-dependent hNav1.7
blockers as treatments for neuropathic pain. Toward
that end, we initiated a screening of the Merck sample
collection and identified benzodiazepine 1 (Fig. 1) as a
starting point for optimization studies. The modification
of 1, described herein, has led to the discovery of a po-
tent, state-dependent hNav1.7 blocker that displays
good oral efficacy in a rat model of neuropathic pain.
Initial analogs of 1 were synthesized using the proce-
dures outlined in Scheme 1. The starting material for
this sequence, (R)-3-amino-1-methyl-5-phenyl-1,3-dihy-
dro-benzo[e][1,4]diazepin-2-one 2, has been described
previously.11 Coupling of 2 with N-Boc-D-phenylalanine
furnished protected amide 1. Acid-catalyzed amine
deprotection then gave 3, a central intermediate that
could be coupled with various sulfonyl chlorides, chlo-
roformates or carbonyl chlorides to yield products such
as 4, 5, and 6. Alternatively, 3 could participate in reduc-
tive amination to afford amines such as 7.
Having elucidated the requirements for R1, we turned
our attention toward the sidechain R2 site. In a bid to
reduce structural complexity, we prepared analogs
wherein the N-Boc group (Table 2, compound 17) or
the entire –NH(Boc) moiety (compound 18) had been
excised. Unfortunately, each of these afforded only weak
hNav1.7 functional block. An N-methylated derivative
(19), a series of sulfonamides (20, 4, and 21) and an
amine (7) that retained the terminal tert-butyl group of
1 were also prepared and found to be much less potent
than N-Boc derivative 1. These results suggested that
the sidechain N–H might play an important role in
channel binding, and might need to be maintained with-
in a specific pKa range. With that in mind, we narrowed
the scope of our efforts and prepared a series of carba-
mates and amides. In the carbamate series, methyl deriv-
ative 5 displayed only weak channel block, but benzyl
Once synthesized, compounds were then assayed for
their ability to block hNav1.7. The extent of channel
block was determined in a functional, membrane poten-
tial-based assay that measures the fluorescence reso-
nance energy transfer (FRET) between two
membrane-associated dyes. Specific details of the exper-
imental protocols employed have recently been de-
scribed.12 Selected compounds were also screened
against several other ion channels that are known to im-
Table 1. Effect of the R1 group on hNav1.7 potency
O
O
O
N
N
a
b
NH2
O
N
H
O
N
N
N
R1
NH
Boc
N
H
N
NH
Boc
2
1
O
O
O
O
N
N
R1
hNav1.7 (IC50, nM)
c
Compound
N
H
N
H
1
8
CH2Ph
Me
125
>1000
>1000
375
+
N
N
NH3
NHR
TFA-
9
CH2(c-Hex)
Ph
4 (R = SO2i-Pr)
5 (R = CO2Me)
6 (R = (CO)Ad)
3
10
11
12
13
14
15
16
CH2CH2Ph
CH2(2-F-Ph)
CH2(4-F-Ph)
CH2(4-thiazolyl)
CH2(3-pyridyl)
CH2(4-imidazolyl)
238
87
7 (R = CH2CH2t-Bu)
71
684
Scheme 1. Reagents and condition: (a) N-Boc-D-Phe, CDI, DMF,
60 °C (89%); (b) TFA, CH2Cl2; (c) i-PrSO2Cl, i-Pr2NEt, CH2Cl2; or
MeO2CCl, K2CO3, CH2Cl2; or 1-adamantyl carbonyl chloride,
K2CO3, CH2Cl2; or 3,3-dimethylbutyraldehyde, NaCNBH3, MeOH.
978
>1000

4632
S. B. Hoyt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4630–4634
Table 2. Effect of the R2 group on hNav1.7 potency
F3C
H
N
O
O
N
O
O
N
b
c
NHR
NHTr
N
H
R2
N
27 (R = H)
28 (R = Tr)
29
a
d or e
R2
hNav1.7 (IC50, nM)
F3C
F3C
Compound
X
Y
1
17
18
19
20
4
NH(Boc)
NH2
125
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
150
N
N
f
NHTr
N
H
H
NH
NMe(Boc)
NH(SO2Me)
NH(SO2i-Pr)
NH(SO2Ph)
Boc
31 (X = O; Y = O)
32 (X = O; Y = H,H)
33 (X = H,H; Y = O)
30
21
7
Scheme 2. Reagents and conditions: (a) TrCl, Et3N, DMF (60%); (b)
NaH, THF, CF3CH2OTf, 0 °C to RT (82%); (c) LAH, THF, 0–60 °C
(99%); (d) HCl, MeOH; N-Boc-D-Phe, CDI, DMF, 60 °C; (e) HCl,
MeOH; N-Boc-D-phenylalaninal, NaCNBH3, MeOH; (f) HCl, MeOH;
N-Boc-D-Phe, BOP, i-Pr2NEt, THF (64%—2 steps).
NH(CH2CH2t-Bu)
NH(CO2Me)
NH(CO2Bn)
NH(CO)c-Pr
NH(CO)t-Bu
NH(CO)Ad
5
22
23
24
6
>1000
380
420
tetrahydro-1H-[1]-benzazepin-2-one 27, was prepared
according to the procedure of Armstrong and cowork-
ers, then tritylated to yield compound 28.15 Sequential
treatment of 28 with sodium hydride and trifluoroethyl
triflate resulted in lactam alkylation to give 29. Acid-cat-
alyzed N-detritylation of 29 then furnished an amine
that could undergo amide coupling with N-Boc-D-phen-
ylalanine to yield 31 (step d), or reductive amination
with N-Boc-D-phenylalaninal to provide 32 (step e).
Alternatively, intermediate 29 could be treated with lith-
ium aluminum hydride to afford 30, the product of lac-
tam reduction. Acid-catalyzed amine deprotection
followed by BOP-mediated amide coupling then deliv-
ered 33. Compounds listed in Tables 3–5 were synthe-
sized according to these procedures using the
appropriate commercially available starting materials.
In several instances (46 and 47), compounds were pre-
pared using non-commercially available amino acids
that had been synthesized via the method of Schol-
lkopf.16 Analogs with (S) stereochemistry at the 3-amino
stereocenter (e.g., 48) were prepared from the enantio-
mer of 27, known compound (S)-3-amino-2,3,4,5-tetra-
hydro-1H-[1]-benzazepin-2-one.17
carbamate 22 proved to be as potent as 1. Likewise, in
the amide series, the smaller cyclopropyl analog 23
proved weak, while the larger tert-butyl and adamantyl
amides 24 and 6 were more potent. This initial survey
suggested that a range of amides and carbamates might
be tolerated at R2, so long as they terminated in a bulky
alkyl or aryl group.
We next probed the effect of alterations to the benzodi-
azepine core. With an eye toward simplification, we
truncated the tricyclic core of 1 and obtained 25, a com-
pound that maintained potent hNav1.7 block (Table 3).
Taking this approach a step further, we deleted the left-
hand phenyl ring from 25. The resulting derivative 26
proved less potent, thus underscoring the importance
of that ring in binding and/or conformational con-
straint. Because it seemed to offer numerous advantages,
the benzazepinone core embodied in 25 was employed in
all subsequent designs.
Analogs in this new benzazepinone series were synthe-
sized according to the procedures outlined in Scheme
2. The requisite starting material, (R)-3-amino-2,3,4,5-
Our final set of core modifications focused on the ben-
zazepinone lactam region. We began by replacing the
lactam N-methyl of 25 with a set of small alkyl groups
(Table 4, compounds 31, 34, and 35). Of these, the
N-isopropyl analog 35 displayed optimal hNav1.7 block-
ade. We also probed the importance of the lactam car-
bonyl in channel binding. When compared with
N-methyl lactam 25, the reduced derivative 36 was nota-
bly less potent. In contrast, the N-trifluoroethyl lactam
31 and its reduced counterpart 33 were essentially equi-
potent. Thus, it may be that while the lactam carbonyl is
not required per se, electron density on the lactam nitro-
gen must be attenuated in order to maintain potent
hNav1.7 block. Finally, we investigated the role of the
exocyclic amide in channel binding. When compared
with amide 31, the corresponding amine 32 showed both
reduced hNav1.7 potency and increased MK-0499 activ-
ity. From these studies, it appeared that an optimal
Table 3. Modifications to the benzodiazepine core
O
R3
=
NH
Boc
O
O
N
N
NHR3
NHR3
Compound
hNav1.7 (IC50, nM)
25
192
26
>1000

S. B. Hoyt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4630–4634
4633
Table 4. Modifications to the benzazepinone lactam region
design would incorporate an N-isopropyl benzazepinone
core coupled to an amide sidechain.
R4
X
Y
N
At this point, we had identified the essential structural
features required for potent block of hNav1.7. Because
substitution at the R1 position was known to affect bind-
ing, and because we had made significant changes to the
lead structure since our first survey of R1, we ran a fo-
cused re-optimization of that domain. Consistent with
our earlier results, fluorination of the R1 phenyl ring
was well tolerated (compounds 37–41), with the 3-fluor-
ophenyl and 2,6-difluorophenyl derivatives 38 and 41
proving particularly potent (Table 5). Incorporation of
the larger trifluoromethyl group at the 2-, 3- or 4-posi-
tions (42–44) led, in each case, to derivatives less potent
than their fluorinated counterparts. Potency of the 2-tri-
fluoromethyl analog could, however, be improved by
additional fluorination at the 3- or 6-position (45 and
46), or by conversion to a 2-trifluoromethoxy group
(47). In accordance with trends that have been observed
generally in this series, the S,R diastereomer of 47 (com-
pound 48) maintained good hNav1.7 block, while the
R,S diastereomer 49 proved to be less potent. In general,
compounds from this series were notably free of activity
in the MK-0499 and DLZ counterscreens.
N
H
NH
Boc
Compound R4
X
Y
hNav1.7 MK-0499
(IC50, nM) (% inhibition
at 10 lM)
25
31
34
35
36
33
32
CH3
0
0
192
150
16
16
15
6
CH2CF3
CH2c-Pr
i-Pr
0
0
0
0
78
45
0
H,H
0
CH3
0
>1000
145
885
28
40
79
CH2CF3 H,H
CH2CF3
0
H,H
0
Table 5. Optimization of the benzazepinone R1 substituent
O
O
N
*
N
H
R1
*
NH
Boc
Compound R1
/
* *
hNav1.7
(IC50, nM)
(% inhibition
at 10 lM)
Several members from this series were submitted to
pharmacokinetic (PK) assays. The first compound to
be profiled, 2-F-phenyl derivative 37, displayed rather
modest oral bioavailability and exposure (Table 6). For-
tunately, analogs of 37 that incorporated larger groups
at the phenylalanine 2-position fared better. The 2-
CF3-phenyl derivative 42, for instance, exhibited a two-
fold increase in oral exposure. Related compounds 46
and 47 maintained or improved upon that exposure level
and also featured reduced rates of clearance. The deriv-
ative with the best overall profile, 47, was submitted for
PK determination in dog and showed similar results, al-
beit with lower bioavailability, in that species.
MK-0499 DLZ
35
37
38
39
40
41
42
43
44
45
46
47
48
49
Ph
2-F-Ph
R,R
R,R
R,R
R,R
R,R
R,R
R,R
R,R
45
44
6
8
8
11
0
3-F-Ph
4-F-Ph
9
98
0
0
2,5-di-F-Ph
2,6-di-F-Ph
2-CF3-Ph
3-CF3-Ph
4-CF3-Ph
23
14
6
24
36
4
137
295
34
15
19
R,R >1000
2-CF3-3-F-Ph R,R
2-CF3-6-F-Ph R,R
65
32
39
43
18
20
0
87
81
3500a
1725a
2-OCF3-Ph
2-OCF3-Ph
2-OCF3-Ph
R,R
S,R
R,S
35
32
Because it was the most promising compound to emerge
from this series, 47 was selected for further study. For
reasons outlined above, it was important to determine
214
a IC50, nM.
Table 6. Pharmacokinetic data for selected compounds
O
O
N
N
H
R1
NH
Boc
R1
Species
Dosea (iv/po)
F%
AUCN
Cmax
Clp
T1/2
b
c
d
e
Target
37
42
46
47
48
47
2-F-Ph
2-CF3-Ph
Rat
Rat
Rat
Rat
Rat
Dog
1.0/3.0
1.0/3.0
1.0/3.0
1.0/3.0
1.0/3.0
0.38/1.5
14
35
28
24
15
9
0.12
0.22
0.24
0.31
0.09
0.27
0.10
0.30
0.33
0.30
0.14
0.07
41
50
32
24
55
15
1.5
2.1
2.5
2.3
2.1
4.2
2-CF3-6-F-Ph
2-OCF3-Ph
2-OCF3-Ph
2-OCF3-Ph
a mg/kg.
b lM h/mpk.
c lM.
d mL/min/kg.
e Hours.

4634
S. B. Hoyt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4630–4634
whether 47 blocked hNav1.7 in a state-dependent man-
ner. Compound 47 was therefore analyzed by whole cell
electrophysiology to determine its binding to hNav1.7
inactive and resting states (Ki and Kr, respectively).18
Analysis via this technique yielded a Ki = 40 nM and a
Kr of >3 lM, indicating that block of hNav1.7 by 47 is
highly state-dependent.
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nan, J.; Raashid, Y.; Al-Gazali, L.; Hamamy, H.; Valente,
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R. M. Tetrahedron Lett. 1987, 28, 521.
12. Felix, J. P.; Williams, B. S.; Priest, B. T.; Brochu, R. M.;
Dick, I. E.; Warren, V. A.; Yan, L.; Slaughter, R. S.;
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The ability of 47 to inhibit neuronal firing in vivo was
examined in the rat peripheral axotomy model.19 In this
assay, the sciatic nerve was sectioned at mid-thigh to
generate a neuroma. Three to 10 days later, the nerve
was exposed and electrodes were connected to neuronal
filaments that were firing spontaneously. After the base-
line firing frequency was established, test compound was
administered as an iv infusion. Firing frequency was
then measured for an additional 60 min post-dose.
When dosed iv at 2 mg/kg, 47 produced essentially com-
plete block of spontaneous neuronal firing at all time-
points studied (% inhibition at 10, 30, and
60 min = 100%, 100%, 93%, n = 6).
Finally, compound 47 was submitted for pharmacody-
namic profiling in a rat spinal nerve ligation model of
neuropathic pain.20 In that assay, rats underwent surgical
ligation and transection of the L5 spinal nerve as a means
of initiating neuropathy. Tactile allodynia was assessed
both before and 1 week after surgery using calibrated
Von Frey filaments. In rats that presented significant
allodynia, test compound was administered orally, and
reversal of allodynia was then determined. When dosed
orally at 10 mg/kg, compound 47 elicited significant
reversal of allodynia at both 2 and 4 h post-dose (% rever-
sal at 2, 4 h = 43%, 40%, n = 8), indicating that it is in-
deed efficacious in a rat model of neuropathic pain.
In summary, we have discovered a structurally novel
class of benzazepinone-based sodium channel blockers.
An exemplar of this class displayed potent, state-depen-
dent block of hNav1.7 in vitro, blocked spontaneous neu-
ronal firing in vivo, and was orally efficacious in a rat
model of neuropathic pain. Future work in this series will
focus on further improvements to pharmacokinetics and
pharmacodynamics, and will be reported in due course.
16. Schollkopf, U. Tetrahedron 1983, 39, 2085.
17. Chang, C.-Y.; Yang, T.-K. Tetrahedron: Asymmetry 2003,
14, 2081.
Acknowledgments
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333.
20. Chaplan, S. R.; Bach, F. W.; Pogrel, J. W.; Chung, J. M.;
Yaksh, T. L. J. Neurosci. Methods 1994, 53, 55.
We thank Ramona Gray and Joe Leone for their out-
standing technical contributions to this work. We also
thank Joseph L. Duffy, Deborah E. Pan, and Christian
Stevenson for their valuable assistance in proofreading
this document.