Sulfonamides as Selective NaV1.7 Inhibitors: Optimizing Potency and Pharmacokinetics to Enable in Vivo Target Engagement

Article abstract of DOI:10.1021/acsmedchemlett.6b00243

Human genetic evidence has identified the voltage-gated sodium channel Na_V1.7 as an attractive target for the treatment of pain. We initially identified naphthalene sulfonamide 3 as a potent and selective inhibitor of Na_V1.7. Optimization to reduce biliary clearance by balancing hydrophilicity and hydrophobicity (Log D) while maintaining Na_V1.7 potency led to the identification of quinazoline 16 (AM-2099). Compound 16 demonstrated a favorable pharmacokinetic profile in rat and dog and demonstrated dose-dependent reduction of histamine-induced scratching bouts in a mouse behavioral model following oral dosing.

Full text of DOI:10.1021/acsmedchemlett.6b00243

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Letter
V
Sulfonamides as Selective Na1.7 Inhibitors: Optimizing Potency
and Pharmacokinetics to Enable in Vivo Target Engagement
Isaac E. Marx, Thomas A. Dineen, Jessica Able, Christiane Bode, Howard Bregman, Margaret Chu-
Moyer, Erin F. DiMauro, Bingfan Du, Robert S. Foti, Robert T. Fremeau, Jr., Hua Gao, Hakan Gunaydin,
Brian E. Hall, Liyue Huang, Thomas Kornecook, Charles R. Kreiman, Daniel S. La, Joseph Ligutti,
Min-Hwa Jasmine Lin, Dong Liu, Jeff S. McDermott, Bryan D. Moyer, Emily A Peterson, Jonathan
T. Roberts, Paul E. Rose, Jean Wang, Beth D. Youngblood, Violeta L. Yu, and Matthew M. Weiss
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.6b00243 • Publication Date (Web): 21 Sep 2016
Downloaded from http://pubs.acs.org on September 22, 2016
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Sulfonamides as Selective NaV1.7 Inhibitors: Optimizing Potency and
Pharmacokinetics to Enable in Vivo Target Engagement
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Isaac E. Marx*,†, Thomas A. Dineen†,#, Jessica Able‡, Christiane Bode†, Howard Bregman†, Margaret
ChuꢀMoyer†, Erin F. DiMauro†, Bingfan Du†,#, Robert S. Foti§, Robert T. Fremeau, Jr. ‡,#, Hua Gao±,
Hakan Gunaydin±,#, Brian E. Hallǁ, Liyue Huang§, Thomas Kornecook‡, Charles R. Kreiman†, Daniel S.
La†, Joseph Ligutti‡, MinꢀHwa Jasmine Lin§,#, Dong Liu‡, Jeff S. McDermott‡,#, Bryan D. Moyer‡,
Emily A. Peterson†,#, Jonathan T. Roberts§, Paul Roseǁ, Jean Wang‡, Beth D. Youngblood‡, Violeta Yu‡,
and Matthew M. Weiss†
†Department of Medicinal Chemistry, ±Department of Molecular Engineering, §Department of Pharmacokinetics and Drug
Metabolism, ‡Department of Neuroscience, and ǁDepartment of Biologics, Amgen, Inc., 360 Binney Street, Cambridge, Masꢀ
sachusetts 02142, and One Amgen Center Drive, Thousand Oaks, California 91320, United States
KEYWORDS: Sodium channel, NaV1.7, NaV1.5, pain, histamine scratching model
ABSTRACT: Human genetic evidence has identified the voltageꢀgated sodium channel NaV1.7 as an attractive target for the treatꢀ
ment of pain. We initially identified naphthalene sulfonamide 3 as a potent and selective inhibitor of NaV1.7. Optimization to reꢀ
duce biliary clearance by balancing hydrophilicity and hydrophobicity (LogD) while maintaining NaV1.7 potency led to the identiꢀ
fication of quinazoline 16 (AMꢀ2099). Compound 16 demonstrated a favorable pharmacokinetic profile in rat and dog, and demonꢀ
strated doseꢀdependent inhibition of histamineꢀinduced scratching bouts in a mouse pharmacodynamic model following oral dosing.
Human genetics has implicated the voltageꢀgated sodium
channel NaV1.7, which is expressed in nociceptive sensory
neurons in dorsal root ganglia (DRG),1 as a compelling target
for pain.2ꢀ4 The primary challenge associated with the develꢀ
opment of NaV1.7 inhibitors has historically been achieving
selectivity over the other eight NaV isoforms. These isoforms
are differentially expressed throughout the body but inhibition
of NaV1.5, which is expressed in cardiac tissue, is of particular
concern as it has been shown to prolong the cardiac QRS wave
in humans.5,6 Previous efforts, including our own, have met
with limited success.4 Here we report the characterization,
structureꢀactivity relationship (SAR) and optimization of a
series of sulfonamideꢀderived NaV1.7 inhibitors. These efforts
delivered an isoformꢀselective compound that was effective in
a histamineꢀinduced scratching model that is representative of
Nav1.7 target engagement.
compounds 1, 2, and 3 supported this hypothesis (Figure
1B),10 and we were pleased to find that 2 and 3 were potent
NaV1.7 inhibitors and showed greater than 200ꢀfold selectivity
over NaV1.5 (Figure 1C).11 In general, analogs in the naphthaꢀ
lene series demonstrated superior NaV1.7 inhibition compared
to the corresponding indole analogs, thus the naphthalene scafꢀ
fold was chosen for further optimization. Additional profiling
showed 3 also suffered from high clearance; however, we beꢀ
lieved that 3 represented a promising starting point for further
optimization that would ultimately deliver tool compounds to
evaluate the pharmacology of an orally bioavailable, isoform
selective, peripherally restricted NaV1.7 inhibitor.
The disparity observed between the low in vitro metabolic
turnover and high rat iv clearance of our initial set of comꢀ
pounds (see Supporting Info) led us to focus on better underꢀ
standing the clearance mechanism of this class of compounds.
Towards that end, a series of experiments were undertaken
that suggested hepatic transporters play a significant role in
the elimination of this class of compounds. First, compound 2
was rapidly removed from the medium when incubated with
attached rat hepatocytes and showed much higher apparent
uptake intrinsic clearance (CLint) compared to metabolic CLint
from suspended hepatocytes.12 A metabolite profiling study
with a compound closely related to compound 2, wherein the
paraꢀCF3 was replaced with a paraꢀCl, showed parent comꢀ
pound was the major observed peak in the bile following inꢀ
travenous administration to bile duct cannulated rats; no parent
or metabolites were detected in the urine (see Supporting Inꢀ
fo). Furthermore, CL and volume of distribution (Vdss) of a
structurally related heteroarylsulfonamide were reduced
Recently Pfizer and Icagen described a series of heteroarylꢀ
sulfonamide NaV1.7 inhibitors with high levels of selectivity
over NaV1.5.7ꢀ9 These results were reproduced by our group
and are exemplified by compound 1 (Figure 1A). The lack of
NaV1.5 activity was noteworthy and we envisioned this as a
good starting point for our own lead optimization efforts,
which would initially be aimed at addressing some of the liaꢀ
bilities and shortcomings associated with this class of comꢀ
pounds. Namely, this series suffered from low passive permeꢀ
ability and high clearance in rodents. We believed constrainꢀ
ing the linker within a bicyclic core such as indole 2 or naphꢀ
thalene 3 would afford a similar conformation and potentially
help address the pharmacokinetic liabilities of this class of
compounds. An overlay of global minima conformations of
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Figure 1: Design of Bicyclic Sulfonamide Scaffolds. (A) Representative example of a NaV1.7 selective heteroarylsulfonamide. (B) Overꢀ
lay of biaryl ether 1 (blue), indole 2 (green), and naphthalene 3 (orange) (pyrazole rings omitted for clarity). (C) Profile of initial indole 2
and naphthalene 3.
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approximately 80% in rats pretreated with a single dose of
rifampicin (50 mg/kg, iv), a known inhibitor of hepatic uptake
transporters OATP1B1 and OATP1B3. These results together
suggested that hepatic uptake and efflux transporters play a
significant role in the elimination of this class of heteroarylꢀ
sulfonamides. It is known that LogD and pKa both play a
significant role in determining the extent of biliary excretion
of compound into bile.13,14 With an eye towards occupying
drugꢀlike chemical space, a strategy was thus adopted that
focused on optimizing potency while targeting a cLogD range
of 2ꢀ3.15
As shown in Table 1, the heteroarylsulfonamide moiety had
a profound effect on potency and pharmacokinetics. Interestꢀ
ingly, upon intravenous dosing to rats compound 8 was found
to have high clearance. This result was attributed to the signifꢀ
icant drop in cLogD afforded by the orthoꢀmethoxy moiety
relative to the orthoꢀbromo substituent of 6 and we thus foꢀ
cused on modifying the polarity of the heteroarylsulfonamide
as a means of attaining compounds in the desired LogD range.
Given the acidity of the 5ꢀaminoꢀ1,2,4ꢀthiadiazole sulfonaꢀ
mide moiety (see Entry 8 above)18 it was reasoned that moduꢀ
lating the pKa of the sulfonamide could significantly impact
LogD and provide another handle to help modulate the rate of
biliary clearance. Indeed, compound 9, which exhibited a tenꢀ
fold loss of NaV1.7 potency, demonstrated a 100ꢀfold imꢀ
provement in total clearance, a manifestation that was attributꢀ
ed to the pKa differences between 8 and 9 and the correspondꢀ
ing increase in cLogD. Apart from 2ꢀaminothiazole (9) and 4ꢀ
aminopyrimidine (10), a dramatic loss in potency was obꢀ
served with aminoheterocycle modifications, representatives
of which are shown in Table 1 (entries 11-14).
Compounds were evaluated using a PatchXpress (PX) platꢀ
form and, in an effort to identify and evaluate compounds that
exhibited stateꢀdependent block, a voltage protocol that led to
inactivation of approximately 20% of the NaV1.7 channels was
employed.
As a first step towards exploring the SAR of the naphthaꢀ
lene series, we turned our attention to the substitution of the
southern ring (Table 1). Early SAR found this aryl ring was
required and that a paraꢀtrifluoromethyl group provided supeꢀ
rior potency. We were pleased to find that NaV1.5 selectivity
remained high and interestingly, despite the high homology
between human, rat, and mouse NaV1.7, compounds from this
series were generally equipotent on human and mouse NaV1.7
but much less potent against rat NaV1.7.16 Various lipophilic
groups were examined on the southern ring. Such groups
proved well tolerated in the para position, though impact on
clearance was minimal. Turning to the orthoꢀpyrazole moiety,
it was found that basic heterocycles afforded very potent
NaV1.7 inhibition (4); however, clearance for these highly
polar compounds, which have cLogD values in the 0ꢀ1 range,
remained high.17 Attempts to replace the pyrazole with a
smaller polar group such as phenol 5 were poorly tolerated,
but bromide 6 demonstrated only a modest loss of NaV1.7
potency while improving passive permeability and iv clearꢀ
ance relative to compounds 2-4. Methyl analog 7 was the first
compound with a truncated orthoꢀsubstituent to achieve subꢀ
100 nM potency, demonstrating that potent NaV1.7 inhibition
as well as high levels of NaV1.5 selectivity could be achieved
without the use of a large polar heterocycle. Furthermore,
methoxy analog 8 showed equivalent levels of NaV1.7 inhibiꢀ
tion and greatly improved passive permeability compared to
previous compounds. As the orthoꢀmethoxy moiety afforded
the best overall potency and selectivity profile of the substituꢀ
ents examined, this substituent was kept constant while optiꢀ
mizing the rest of the molecule to further improve the potency
and pharmacokinetic profile.
Despite the reduction in NaV1.7 potency afforded by the 2ꢀ
aminothiazole sulfonamide, our efforts to this point suggested
that consistent challenges were associated with designing a 5ꢀ
aminoꢀ1,2,4ꢀthiadiazole sulfonamide with an acceptable
pharmacokinetic profile; thus, we proceeded to further optiꢀ
mize NaV1.7 potency with the 2ꢀaminothiazole sulfonamide in
place by focusing on the central bicyclic core. In general, the
addition of polarity in the lower half of the core was wellꢀ
tolerated from a potency perspective, and permeability was
greatly improved relative to the parent naphthalene. Additionꢀ
ally, as expected compounds with lower cLogD values generꢀ
ally showed increased clearance, further strengthening the
association between cLogD and clearance. Interestingly, isoꢀ
quinoline 15 demonstrated improved NaV1.7 potency and unꢀ
bound clearance relative to 9 despite a decrease in cLogD.
Adding additional polarity in the form of quinazoline 16 led to
further improvements in NaV1.7 potency, though at the cost of
a slight increase in unbound clearance. Compounds 17 and
18, which demonstrated lower cLogDs, also demonstrated
higher unbound clearance, while shifting polarity to the upper
ring of the [6,6] system led to loss of NaV1.7 potency (e.g.,
naphthyridine 19). Ultimately, quinazoline 16 afforded the
best combination of NaV1.7 potency, permeability, and unꢀ
bound clearance. Overall, this series of bicyclic sulfonamides
demonstrated much improved pharmacokinetics in preclinical
species, enabling the identification of a tool compound to
evaluate the pharmacology of the sulfonamide series after oral
dosing.
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Table 1. Naphthalene Sulfonamide SAR
1
2
3
4
5
6
7
8
9
10
11
12
Rat
CL
iv
hNaV1.7
IC50
(µM)
hNaV1.5
IC50
(µM)
HLM/RLM
CLint
Papp
(10ꢀ6
Rat
PPB
(Fu)
Cpd R2
R1
pKa d
3.6
cLogD
1.6
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(CLu;
((µL/min)/mg)a cm/s)b
L/h/kg)c
5.4
0.037
0.81
>30
23
<14/<14
25/18
1.3
<2
0.10
4
5
0.001
ꢀ
3.6
1.7
0.16
21
<14/<14
15/<14
>5e
>5e
0.003
0.001
0.24 (80) 3.6
2.4
2.2
6
7
0.048
5.1
ꢀ
3.6
0.051
0.59
0.51
>30
12
15/18
17
12
27
9.5
0.001
0.001
0.003
ꢀ
5.20
3.7
6.7
-
1.7
3.4
2.2
2.8
8
0.049
(49)
20.5
>30
ꢀ
25/52
9
<14/<14
16/25
ꢀ
ꢀ
ꢀ
ꢀ
ꢀ
10
11
-
1.5
>30
1.4
>30
ꢀ
17/<14
46/37
36
28
27
ꢀ
ꢀ
ꢀ
2.2
3.4
4.1
12
13
14
-
-
21
<14/<14
7.2
aRat liver microsomal (RLM) and human microsomal (HLM) clearance. Compound concentration = 1 µM. Microsomal protein concenꢀ
tration = 250 µg/mL. Apparent permeability measured in parental MDCK cells. cCompounds were dosed intravenously at 0.5 mg/kg as a
b
solution in 100% DMSO to male rats. n = 2 animals per group. CLu = unbound clearance. dSee reference 15. eRecovery <50%.
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Table 2. Bicyclic Core SAR
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2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
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59
60
hNaV1.7
IC50 (µM)
hNaV1.5
IC50 (µM) cm/s)a
Papp (10ꢀ6 Rat PPB Rat iv CL
Cpd
X1
X2
X3
X4
cLogD
(Fu)
(CLu; L/h/kg)a
C(H)
C(H)
C(H)
C(H)
C(H)
N
C(H)
C(H)
0.59
0.30
20.5
>30
12
34
0.001
0.012
0.049 (49)
0.18 (15)
3.4
2.6
9
15
16b
N
C(H)
N
C(H)
0.16
>30
31
0.016
0.54 (34)
2.1
N
N
C(H)
N
C(H)
C(H)
N
0.32
0.15
2.5
>30
>30
>30
36
3.2
12
0.014
0.013
ꢀ
1.1 (81)
2.4 (160)
ꢀ
1.9
17
18
19
C(O)
C(H)
N(H)
C(H)
0.95
2.1
N
aSee notes in Table 1. pKa = 6.46.
b
Based on its overall balance of NaV1.7 potency, selectivity
and favorable rat pharmacokinetics, compound 16 (AMꢀ2099)
was further profiled. To understand the stateꢀdependence of
this series, 16 was evaluated using electrophysiology protocols
comparing channels in closed (fully nonꢀinactivated) and inacꢀ
tivated states. While an IC50 of 7.2 µM was determined when
inhibition was measured at a holding potential of ꢀ140 mV,
when cells were depolarized to a voltage yielding 20% chanꢀ
nel inactivation the NaV1.7 IC50 shifted to 0.14 µM and when
cells were depolarized to a voltage yielding ~90% channel
inactivation, the IC50 shifted to 0.009 µM, demonstrating the
steep stateꢀdependent block exhibited by 16. In heterologous
cells, comparable inhibition was observed across human,
mouse, dog, and cynomolgus monkey NaV1.7 with reduced
activity against rat NaV1.7 (Table 3); as such, it was decided to
evaluate the pharmacological activity of 16 in mice. The acꢀ
tivity of 16 on TTXꢀsensitive sodium channels recorded from
the cell bodies of sensory neurons dissociated from mouse
dorsal root ganglia was in good agreement with heterologously
expressed mouse NaV1.7 in stably transfected HEK293 cells.
Table 3. Cross Species NaV1.7 Activity of 16
Mouse
IC50
Monkey
IC50
Mouse
Rat IC50 Dog IC50
DRG IC50
(µM)a
(µM)a
(µM)a
(µM)a
(µM)b
0.18
0.14
±
0.18
0.14
±
0.16
0.091
±
0.13
0.049
±
3.5 ±1.3
aGenerated using automated PatchXpress at a holding potential
that gives 20% inactivation. bGenerated using manual electroꢀ
physiology at a holding potential that gives 20% inactivation.
The pharmacokinetic profile of compound 16 was explored
in more detail (Table 5). In rats 16 showed low total clearance
and moderate Vdss and halfꢀlife. In contrast, when dosed in
dogs 16 showed very low clearance, a low Vdss and long halfꢀ
life (18 h). Metabolic CLint in hepatocytes was comparable
between rats and dogs; further investigation is required to unꢀ
derstand mechanisms for the disparity in rates of iv clearance
across species. Bioavailability was high in rats and moderate
in dogs when dosed orally, and absorption was rapid in both
species. Compound 16 was administered intravenously to bile
duct catheterized rats (vide supra) to understand its clearance
mechanisms. Biliary excretion as parent accounted for 4.7% of
the dose,19 demonstrating that our strategy to reduce biliary
clearance was successful.
To evaluate the selectivity of 16 across the NaV isoforms,
whole cell manual patch clamp experiments on partially inacꢀ
tivated NaV channels (~20% inactivated) were conducted (Taꢀ
ble 4). Compound 16 was >100ꢀfold selective over NaV1.3,
NaV1.4, NaV1.5, and NaV1.8, while lower levels of selectivity
were observed against NaV1.1, NaV1.2, and NaV1.6. The diꢀ
minished levels of selectivity over NaV1.1 and NaV1.2 were
not of particular concern given these isoforms are predomiꢀ
nantly expressed in the CNS and this class of sulfonamides is
peripherally restricted (see below). Compound 16 demonꢀ
strated low affinity for hERG (>30 µM) and did not show
greater than 50% inhibition against a panel of 100 kinases (1
µM) and a broad CEREP panel (10 µM).
In light of the emergence of literature implicating NaV1.7 in
the itch pathway we opted to use a histamineꢀinduced scratchꢀ
ing model to measure NaV1.7 target engagement in vivo. It
has been shown that NaV1.7 knockout mice do not scratch
following a histamine challenge, demonstrating that histaꢀ
mineꢀinduced scratching is a NaV1.7ꢀdependent behavior,3 and
that gainꢀofꢀfunction NaV1.7 variants produce increased itch in
humans.20 The ease and simplicity with which this model can
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Table 4. Selectivity Profile of 16
1
2
3
4
hNaV1.1
(µM)a
hNaV1.2
(µM)a
hNaV1.3
(µM)a
hNaV1.4
(µM)a
hNaV1.5
(µM)a
hNaV1.6
(µM)a
hNaV1.7
(µM)a
hNaV1.8
(µM)a
hERG
(µM)
Ki
5
6
7
7.3 ± 0.91
2.1 ± 0.092 21 ± 0.21
17 ± 1.6
16 ± 2.5
3.7 ± 0.021
0.14 ± 0.021 >30
>30
aGenerated using manual electrophysiology at a holding potential that gives 20% inactivation.
8
9
10
11
12
13
14
15
16
17
18
19
Table 5. Pharmacokinetic Profiles and Plasma Protein Binding of 16
iva
pob
plasma protein
binding (Fu)
species
CL (L/h/kg)
Vdss (L)
t1/2 (h)
tmax (h)
%F
Cmax (µM)
rat
0.54 (34)
0.013 (1)
1.45
1.9
3.43
7.97
1.67
2.33
100
53
0.016
dog
0.313
17.9
0.014
a0.5 mg/kg as a solution in 100% DMSO (rat), 30% HPBCD/70% water/KOH at pH = 10 (dog). Clu = unbound clearance. 10 mg/kg
b
20
21
22
23
24
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60
(rat) and 1.7 mg/kg (dog) oral dose as a solution in 30% HPBCD/70% water/KOH at pH = 10.
[plasma]total
(µM)
[plasma]unb
(µM)
Dose (mg/kg)
5
3.1 ± 3.6
22 ± 13
0.11
0.76
5.1
20
60
150 ± 27
(A)
(B)
Figure 2: (A) Reduction of scratching behavior in a mouse histamineꢀinduced scratching model with vehicle, 16, and DPH (diphenhydraꢀ
mine, 30 mg/kg po dosing). **, p < 0.01, ***, p < 0.0001 versus vehicle group (oneꢀway ANOVA followed by Dunnet’s tests). (B) Total
and unbound plasma exposure levels of 16 30 minutes post histamine administration.
be translated in human studies makes it an appealing option
for measuring target engagement in vivo. 16 was dosed orally
to C57BL/6 male mice 120 minutes prior to intradermal adꢀ
ministration of histamine. Instances of scratching behavior
were then measured over a 30ꢀminute time period. 16 demonꢀ
strated a doseꢀdependent increase in plasma exposure with a
concomitant doseꢀdependent reduction in scratching bouts
compared to vehicleꢀtreated animals, with a statistically signifꢀ
icant reduction observed at the 60 mg/kg dose (Figure 2). It
should be noted that these effects occurred at doses and expoꢀ
sures that did not manifest in any reduction in locomotor activꢀ
ity in a mouse openꢀfield movement assay. Compound 16
demonstrated a brainꢀtoꢀplasma ratio of 0.008 in mice, indicatꢀ
ing that the compound was peripherally restricted and that the
pharmacodynamic effect was a result of NaV1.7 inhibition in
the PNS. The unbound concentration of 16 required to obꢀ
serve a significant reduction of itching behavior is noteworthy
and warrants further investigation.
high levels of NaV1.5 selectivity was designed. Based on
pharmacokinetic experiments that suggested hepatic transportꢀ
ers as the primary route of elimination, key strategies for reꢀ
ducing clearance were identified such as targeting an optimal
cLogD range by modulating overall hydrophobicity and the
pKa of the sulfonamide moiety. NaV1.7 potency was optiꢀ
mized through modification of the bicyclic core topology and
the polarity of the phenyl and sulfonamide substituents. Ultiꢀ
mately, optimally distributing polarity in the central core led to
the identification of 16, which demonstrated a favorable
pharmacokinetic profile, good selectivity over a range of NaV
isoforms, and doseꢀdependent reduction of scratching behavior
in a histamineꢀinduced scratching model. Further efforts to
improve the potency, pharmacokinetics, and efficacy of the
bicyclic sulfonamide series will be reported in due course.
In conclusion, utilizing compound 1 as a starting point, a
novel series of bicyclic sulfonamide NaV1.7 inhibitors with
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(2) DibꢀHajj, S. D.; Cummins, T. R.; Black, J. A.; Waxman, S. G.
From Genes to Pain: NaV1.7 and Human Pain Disorders. Trends in
Neurosci. 2007, 30, 555ꢀ563.
ASSOCIATED CONTENT
Supporting Information
1
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(3) Gingras, J.; Smith, S.; Matson, D. J.; Johnson, D.; Nye, K.; Couꢀ
ture, L.; Feric, E.; Yin, R.; Moyer, B. D.; Peterson, M. L.; Rottman, J.
B.; Beiler, R. J.; Malmberg, A. B.; McDonough, S. I. Global NaV1.7
Knockout Mice Recapitulate the Phenotype of Human Congenital
Indifference to Pain. PLoS ONE 2014, 9, e105895, 1ꢀ14.
(4) For recent review: Ruiz, M. L.; Kraus, R. L. VoltageꢀGated Sodiꢀ
um Channels: Structure, Function, Pharmacology, and Clinical Indiꢀ
cations. J. Med. Chem. 2015, 58, 7093ꢀ7118.
(5) Wood, J. N.; Boorman, J. P.; Okuse, K.; Baker, M. D. Voltageꢀ
Gated Sodium Channels and Pain Pathways. J. Neurobiol. 2004, 61,
55ꢀ71.
(6) Erdemli, G.; Kim, A. M.; Ju, H.; Springer, C.; Penland, R. C.;
Hoffmann, P. K. Cardiac Safety Implications of hNaV1.5 Blockade
and a Framework for PreꢀClinical Evaluation. Front. Pharmacol.
2012, 3, 6.
Supporting Information Available: Synthetic procedures of all
final compounds, description of in vitro and in vivo assays, and
additional experimental details for determining clearance mechaꢀ
nism. The Supporting Information is available free of charge on
the ACS Publications website at DOI: XXX.
AUTHOR INFORMATION
Corresponding Author
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*Phone: 617-914-6086. Fax: 617-914-4569. E-mail:
isaac.marx@biogen.com. Address: Biogen Inc., 225 Bin-
ney Street, Cambridge, MA 02142.
Present Addresses
(7) Beaudoin, S.; LauferꢀSweiler, M.; Markworth, C. J.; Marron, B.
E.; Millan, D. S.; Rawson, D. J.; Reister, S. M.; Sasaki, K.; Storer, R.
I.; Stupple, P. A.; Swain, N. A.; West, C. W.; Zhou, S. Sulfonamide
Derivatives. WO2010/079443A1, July 15, 2010.
# For T.A.D.: Novartis Institutes for Biomedical Research, 250
Massachusetts Avenue, Cambridge, MA 02139.
For B.D.: Mersana Therapeutics, 840 Memorial Drive, Camꢀ
bridge, MA 02139.
For R.T.F.: NeuroRx Consulting, 854 Massachusetts Avenue,
Unit 3, Cambridge, MA 02139.
For H.G.: Merck Research Laboratories, 33 Avenue Louis Pasꢀ
teur, Boston, MA, 02115.
For J.S.M.: Neuroscience Discovery, Lilly Research Laboratories,
307 East Merrill Street, Indianapolis, IN, 46285.
For E.A.P.: Biogen, 225 Binney Street, Cambridge, MA, 02142.
(8) For additional examples see: Focken, T.; Liu, S.; Chahal, N.;
Dauphinais, M.; Grimwood, M. E.; Chowdhury, S.; Hemeon, I.; Bichꢀ
ler, P.; Bogucki, D.; Waldbrook, M.; Bankar, G.; Sojo, L. E.; Young,
C.; Lin, S.; Shuart, N.; Kwan, R.; Pang, J.; Chang, J. H.; Safina, B. S.;
Sutherlin, D. P.; Johnson, J. P. Jr.; Dehnhardt, C. M.; Mansour, T. S.;
Oballa, R. M.; Cohen, C. J.; Robinette, C. L. Discovery of Aryl Sulꢀ
fonamides as IsoformꢀSelective Inhibitors of NaV1.7 with Efficacy in
Rodent Pain Models. ACS Med. Chem. Lett. 2016, 7, 277ꢀ282.
(9) Sun, S.; Jia, Q.; Zenova, A. Y.; Chafeev, M.; Zhang, Z.; Lin, S.;
Kwan, R.; Grimwood, M. E.; Chowdhury, S.; Young, C.; Cohen, C.
J.; Oballa, R. M. The Discovery of BenzenesulfonamideꢀBased Potent
and Selective Inhibitors of VoltageꢀGated Sodium Channel NaV1.7.
Bioorg. Med. Chem. Lett. 2014, 24, 4397ꢀ4401.
(10) Conformational analyses were performed using Chemical Comꢀ
puting Group’s Molecular Operating Environment (MOE) software.
(11) For synthetic routes and procedures for all compounds, see Supꢀ
porting Information.
(12) Huang, L.; Chen, A.; Roberts, J.; Janosky, B.; Be, X.; Berry, L.;
Lin, M.ꢀH. J. Use of Uptake Intrinsic Clearance from Attached Rat
Hepatocytes to Predict Hepatic Clearance for Poorly Permeable Comꢀ
pounds. Xenobiotica 2012, 42, 830ꢀ840.
(13) Chen, Y.; Cameron, K.; GuzmanꢀPerez, A.; Perry, D.; Li, D.;
Gao, H. Structure–Pharmacokinetic Relationship of In Vivo Rat Biliꢀ
ary Excretion. Biopharm. Drug Dispos. 2010, 31, 82ꢀ90.
(14) Tu, M.; Mathiowetz, A. M.; Pfefferkorn, J. A.; Cameron, K. O.;
Dow, R. L.; Litchfield, J.; Di, L.; Feng, B.; Liras, S. Medicinal
Chemistry Design Principles for Liver Targeting Through OATP
Transporters. Curr. Top. Med. Chem. 2013, 13, 857ꢀ866.
(15) Waring, M. J. Lipophilicity in Drug Discovery. Expert Opin.
Drug Discovery 2010, 5, 235ꢀ248.
(16) This disparity has previously been observed with related chemiꢀ
cal matter (see Ref. 8), suggesting a common binding site for this
class of structurally related sulfonamideꢀbased NaV1.7 inhibitors.
Studies to further understand this disparity will be reported elsewhere.
(17) cLogD(7.4) was calculated using daylight cLogP and an internal
pKa calculator.
Author Contributions
The manuscript was written through contributions of all authors.
All authors have given approval to the final version of the manuꢀ
script.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
The authors gratefully acknowledge Grace Bi and Larry Miller for
purification support; Paul Krolikowski and Steve Hollis for anaꢀ
lytical support; Yohannes Teffera for PK support; Christopher
Ilch and Kimberley Nye for in vivo support; Roman Shimanovich
and Melanie Cooke for formulations support; and Benjamin Milꢀ
gram for proofreading the manuscript.
ABBREVIATIONS
CLint, intrinsic clearance; CLu, unbound clearance; DRG, dorsal
root ganglion; hERG, human etherꢀaꢀgoꢀgoꢀrelated gene; HPBCD,
hydroxypropyl betaꢀcyclodextrin; Papp, apparent permeability;
PNS, peripheral nervous system; PPB, plasma protein binding;
SAR, structure activity relationship; TTX, tetrodotoxin; Vdss,
volume of distribution.
(18) pKa values were calculated from capillary electrophoresis data
obtained with pKa PRO Analyzer (AATI, Ames, IA) in the pH range
of 1.7 to 11.2.
(19) 0.02% and 0.60% of the dose was detected as parent in the urine
and feces, respectively.
(20) Devigili, G.; Eleopra, R.; Pierro, T.; Lombardi, R.; Rinaldo, S.;
Lettieri, C.; Faber, C. G.; Merkies, I. S. J.; Waxman, S. G.; Lauria, G.
Paroxysmal Itch Caused by GainꢀofꢀFunction NaV1.7 Mutation. Pain
2014, 155, 1702ꢀ1707.
REFERENCES
(1) ToledoꢀAral, J.J.; Moss, B. L.; He, Z.; Koszowski, A. G.;
Whisenand, T.; Levinson, S. R.; Wolf, J. J.; SilosꢀSantiago, I.;
Halegoua, S.; Mandel, G. Identification of PN1, a Predominant Voltꢀ
ageꢀDependent Sodium Channel Expressed Principally in Peripheral
Neurons. Proc. Natl. Acad. Sci. USA 1997, 94, 1527ꢀ1532.
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