Discovery of non-zwitterionic aryl sulfonamides as Nav1.7 inhibitors with efficacy in preclinical behavioral models and translational measures of nociceptive neuron activation

Article abstract of DOI:10.1016/j.bmc.2017.08.012

Since zwitterionic benzenesulfonamide Na_v1.7 inhibitors suffer from poor membrane permeability, we sought to eliminate this characteristic by replacing the basic moiety with non-basic bicyclic acetals and monocyclic ethers. These efforts led to

Full text of DOI:10.1016/j.bmc.2017.08.012

Accepted Manuscript
Discovery of non-zwitterionic aryl sulfonamides as Nav1.7 inhibitors with effi-
cacy in preclinical behavioral models and translational measures of nociceptive
neuron activation
Yong-Jin Wu, Jason Guernon, Andrea McClure, Guanglin Luo, Ramkumar
Rajamani, Alicia Ng, Amy Easton, Amy Newton, Clotilde Bourin, Dawn Parker,
Kathleen Mosure, Omar Barnaby, Matthew G. Soars, Ronald J. Knox, Michele
Matchett, Rick Pieschl, James Herrington, Ping Chen, D.V. Sivarao, Linda J.
Bristow, Nicholas A. Meanwell, Joanne Bronson, Richard Olson, Lorin A.
Thompson, Carolyn Dzierba
PII:
S0968-0896(17)31327-5
DOI:
http://dx.doi.org/10.1016/j.bmc.2017.08.012
BMC 13915
Reference:
Bioorganic & Medicinal Chemistry
To appear in:
Received Date:
Revised Date:
Accepted Date:
28 June 2017
4 August 2017
8 August 2017
Please cite this article as: Wu, Y-J., Guernon, J., McClure, A., Luo, G., Rajamani, R., Ng, A., Easton, A., Newton,
A., Bourin, C., Parker, D., Mosure, K., Barnaby, O., Soars, M.G., Knox, R.J., Matchett, M., Pieschl, R., Herrington,
J., Chen, P., Sivarao, D.V., Bristow, L.J., Meanwell, N.A., Bronson, J., Olson, R., Thompson, L.A., Dzierba, C.,
Discovery of non-zwitterionic aryl sulfonamides as Nav1.7 inhibitors with efficacy in preclinical behavioral models
and translational measures of nociceptive neuron activation, Bioorganic & Medicinal Chemistry (2017), doi: http://
dx.doi.org/10.1016/j.bmc.2017.08.012
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Graphical Abstract
Discovery of non-zwitterionic aryl sulfonamides as
Nav1.7 inhibitors with efficacy in preclinical behavioral
models and translational measures of nociceptive neuron
activation
Leave this area blank for abstract info.
Yong-Jin Wu,*,a Jason Guernon,a Andrea McClure,a Guanglin Luo,a Ramkumar Rajamani,a Alicia Ng,b Amy
Easton,a Amy Newton,a Clotilde Bourin,a Dawn Parker,a Kathleen Mosure,a Omar Barnaby,a Matthew G. Soars,a
Ronald J. Knox,a Michele Matchett,a Rick Pieschl,a James Herrington,a Ping Chen,a D.V. Sivarao,a Linda J.
Bristow,a Nicholas A. Meanwell,a Joanne Bronson,a Richard Olson,a Lorin A. Thompson,a and Carolyn Dzierbaa
aBristol-Myers Squibb Research and Development, 5 Research Parkway, Wallingford, CT 06492-7660, USA; bOne Squibb
Drive, New Brunswick, NJ 08903, USA

Bioorganic & Medicinal Chemistry
journal homepage: www.els evier.com
Discovery of non-zwitterionic aryl sulfonamides as Nav1.7 inhibitors with efficacy in
preclinical behavioral models and translational measures of nociceptive neuron
activation
Yong-Jin Wu,*,a Jason Guernon,a Andrea McClure,a Guanglin Luo,a Ramkumar Rajamani,a Alicia Ng,b
Amy Easton,a Amy Newton,a Clotilde Bourin,a Dawn Parker,a Kathleen Mosure,a Omar Barnaby,a Matthew
G. Soars,a Ronald J. Knox,a Michele Matchett,a Rick Pieschl,a James Herrington,a Ping Chen,a D.V.
Sivarao,a Linda J. Bristow,a Nicholas A. Meanwell,a Joanne Bronson,a Richard Olson,a Lorin A.
Thompson,a and Carolyn Dzierbaa
aBristol-Myers Squibb Research and Development, 5 Research Parkway, Wallingford, CT 06492-7660, USA
bOne Squibb Drive, New Brunswick, NJ 08903, USA
ARTICLE INFO
ABSTRACT
Article history:
Received
Since zwitterionic benzenesulfonamide Nav1.7 inhibitors suffer from poor membrane
permeability, we sought to eliminate this characteristic by replacing the basic moiety with non-
basic bicyclic acetals and monocyclic ethers. These efforts led to the discovery of the non-
zwitterionic aryl sulfonamide 49 as a selective Nav1.7 inhibitor with improved membrane
permeability. Despite its moderate cellular activity, 49 exhibited robust efficacy in mouse
models of neuropathic and inflammatory pain and modulated translational electromyogram
measures associated with activation of nociceptive neurons.
Received in revised form
Accepted
Available online
Keywords:
benzenesulfonamide
2009 Elsevier Ltd. All rights reserved.
Nav1.7 inhibitor
membrane permeability
neuropathic and inflammatory pain
translational electromyogram
nociceptive flexion reflex (NFR)
1. Introduction
(1)14 and benzenesulfonamides 215 and 316 as potent, isoform-
selective Nav1.7 inhibitors with efficacy in various rodent pain
There is substantial genetic evidence linking the voltage-gated
sodium channel Nav1.7 and its encoding SCN9A gene to painful
disorders in humans. For example, gain-of-function mutations in
the SCN9A gene lead to painful conditions such as inherited
erythromelalgia and paroxysmal extreme pain disorder1,2 whereas
loss-of-function mutations in the SCN9A gene result in
congenital insensitivity to pain, a rare disorder characterized by a
complete inability to sense painful stimuli.3-5 Apart from
anosmia, humans and mice deficient in Nav1.7 are apparently
normal,6,7 suggesting that Nav1.7 inhibitors would not incur
safety-related, on-target toxicity. The Nav1.7 isoform is
expressed predominantly in the peripheral nervous system (PNS)
and, therefore, a non-brain penetrant inhibitor would be expected
to have reduced potential for eliciting central nervous system
(CNS)-related adverse effects.8 Taken together, Nav1.7 is viewed
as a promising analgesic drug target for acute, inflammatory and
neuropathic pain.9-12 As block of the cardiac sodium channel
Nav1.5 may lead to arrhythmia,13 recent efforts have focused on
the development of isoform-selective inhibitors. These efforts
have culminated in the discovery of the sulfonamide GNE-131
models. These compounds bind to a unique site in the voltage
sensor domain 4 (VSD4) of Nav1.7 that is distinct from the pore
binding sites of tetrodotoxin or local anesthetics.17 The crystal
structure of a chimera consisting of human Nav1.7 VSD4 and a
bacterial pore domain in complex with an aryl sulfonamide has
been recently disclosed.18 However, 1 and 2 are highly protein
bound (>99%) while benzenesulfonamides 2 and 3 suffer from
poor membrane permeability. The impact of membrane
permeability on the oral bioavailability of these compounds is not
clear, but we have recently shown that poor membrane
permeability can contribute to poor exposure in dorsal root
ganglion (DRG) where the Nav1.7 channel is highly expressed.19
Inhibition of channel function in this region may be a primary
site of action for the expression of an analgesic effect. The poor
membrane permeability associated with 2 and 3 may be the result
of their zwitterionic character. In order to improve the
permeability properties of 3, we sought to eliminate the
zwitterionic character by removing either the acidic or the basic
moiety. However, the sulfonamide functionality and the thiazole
ring that form the acidic element are essential for binding to the

protein, via a salt bridge to a positively charged arginine in the
domain 4 voltage sensor.17,18 Consequently, we undertook to
replace the basic amine with non-basic heteroatoms, an effort that
led to the discovery of the bicyclic analogs 4-7 as well as the
monocyclic compounds 8. This report describes the synthesis and
biological activities of these analogs.
a Swern oxidation. Debenzylation was followed by spontaneous
intramolecular ketalization to furnish acetal 18. Removal of the
Boc group gave amine 19 as the TFA salt, which was used
directly in the coupling reaction with 11 to give analog 4 after
deprotection with TFA.
Scheme 2. Reagents and conditions: (a) NaH, Br(CH2)2OBn,
THF/DMF (1:1), rt, 65%; (b) LiAlH4, THF, 0 °C, 93%; (c) Boc2O,
DCM, rt, 66%; (d) (COCl)2, DMSO, DCM, -78 °C; Et3N, -78 °C to
rt, 82%; (e) 10% Pd(OH)2, H2 balloon, MeOH, rt, 100%; (f) TFA,
DCM, rt, 100%; (g) 11, K2CO3, DMF, rt; (h) TFA, DCM, rt, 21%
over 2 steps.
Figure 1. Structures of Nav1.7 inhibitors
Scheme 3 describes the preparation of ketal analog 5 from
aldehyde 16. This aldehyde was converted to the TFA salt of
amine 21 in four steps: addition of methylmagnesium bromide;
Swern oxidation; catalytic hydrogenation; and Boc deprotection.
Coupling of amine 21 with 11 gave analog 5 after deprotection
with TFA. Both ketal amines 19 (Scheme 2) and 21 showed no
appreciable decomposition when exposed to a range of acidic
conditions at 60 oC for 12 h.
2. Chemistry
Scheme
1
describes
a
general
synthesis
of
benzenesulfonamides 32-71 (Table 1) from compound 10, which
was readily available through reductive amination of 2,4-
dimethoxybenzaldehyde with 2-aminothiazole. Treatment of 10
with benzenesulfonyl chloride 9 under basic conditions afforded
benzenesulfonamide 11. The nucleophilic aromatic substitution
reaction of an amine with 11 using K2CO3 as the base gave amine
12, which underwent deprotection with TFA to afford
compounds 32-71. Enantiomers 49 and 50 were obtained from
their racemate by chiral HPLC separation and the absolute
configuration of 50 was confirmed by single crystal X-ray
crystallography.20
Boc
O
HN
NH2
O
c,d
a,b
e,f
5
16
O
BnO
OBn
Me
21
20
Scheme 3. Reagents and conditions: (a) MeMgBr, THF, rt,
93%; (b) (COCl)2, DMSO, DCM, -78 °C; Et3N, -78 °C to rt, 60%;
(c) 20% Pd(OH)2, H2 balloon, MeOH, rt, 93%; (d) TFA, DCM, rt,
100%; (e) 11, K2CO3, DMF, rt; (f) TFA, DCM, rt, 41% over 2
steps.
Scheme 4 describes the preparation of ketal analog 6
from amine 27. This amine was prepared in a similar
manner to amine 19 with the exception that the double
alkylation of ethyl 2-cyanoacetate was carried out in a
stepwise fashion to give the bis-alkylated product 22.
The cis configuration of 26 was confirmed by the
observation of a nOe effect between the acetal proton
and the two protons adjacent to the amino group.
Interestingly, there was no trans isomer of 26 formed in
the intramolecular ketalization step. Coupling of amine
27 with 11 gave analog 6 after deprotection with TFA.
Scheme 1. Reagents and conditions: (a) LHMDS, THF, -78
°C, 55%; (b) RNH2, K2CO3, DMF, rt, 20-75%; (c) TFA, DCM, rt,
50-95%.
Scheme 2 describes the preparation of acetal analog 4. Double
alkylation of ethyl 2-cyanoacetate (13) with 1-bromo-2-
benzyloxyethane gave 14. The ester and cyano groups were both
reduced with LiAlH4 and the amine was protected with a Boc
group to give alcohol 15 which was converted to aldehyde 16 via

since heteroaryl sulfonamides are known to exhibit potent CYP
inhibition, especially toward CYP3A4.15,21
The acetal 4 inhibited hNav1.7 channel activity with an IC50
value of 108 nM, with 190-fold selectivity over the hNav1.5
isoform. This compares with 2.2 nM and >13,000-fold,
respectively, for reference compound 3. The PAMPA
permeability of 4 was dependent on pH with higher permeability
observed at pH 5.5 than at 7.4 while the profile for 3 was the
reverse. Surprisingly, the homologous ketal analog 5 was
significantly less active than prototype 4, reflecting a tight
hydrophobic pocket in the acetal-binding region. However, ring
expansion of 4 to the 5,6-fused bicyclic acetal 6 was better
tolerated, reducing activity by only 6-fold. The bicyclic tertiary
alcohol 7 maintained Nav1.7 activity but selectivity over the
Nav1.5 isoform was reduced by ~4-fold. Since the reference
compound 1 is based on a lipophilic adamantanyl moiety, we
examined the effect of incorporation of
a functionalized
Scheme 4. Reagents and conditions: (a) NaH, Br(CH2)2OBn,
THF/DMF (1:1), rt, 74%; (b) NaH, Br(CH2)3OBn, THF/DMF
(1:1), rt, 70%; (c) LiAlH4, THF, 0 ºC, 59%; (d) Boc2O, DCM, rt,
59%; (e) Dess-Martin reagent, DCM, rt, 83%; (f) 20% Pd(OH)2,
H2 balloon, MeOH, rt, 75%; (g) TFA, DCM, rt, 100%; (h) 11,
K2CO3, DMF, rt; (i) TFA, DCM, rt, 40% over 2 steps.
adamantanyl group into our aryl sulfonamide inhibitors.
Moderate inhibitory activity was obtained with the adamantanol
analog 33, while the other adamantyl regioisomer 32 showed
much reduced inhibitory potency. Overall, within the bicyclic
series, acetal 4 showed the best activity, good selectivity over the
Nav1.5 isoform, and improved permeability compared to 3.
Scheme 5 describes the preparation of analog 7. Ethyl 2-
1,3-
We next investigated the monocyclic ether analogs. With
respect to the 3-substituted tetrahydrofuran analogs 34–38, a one
to three methylene spacer between the heterocycle and the amino
group resulted in comparable Nav1.7 inhibitory activity while
direct linkage, as exemplified by 34 and 35, resulted in a
significant reduction in potency. The position of the ring oxygen
(36 vs. 39) exerted little effect on activity and the introduction of
an additional oxygen to form an acetal, as in 40, maintained
activity.
oxocyclopentanecarboxylate
was
alkylated
with
diiodopropane, and the resulting iodide 28 was subjected to
SmI2-induced ring closure to give hydroxyl ester 29. Treatment
of 29 with LiNH2 gave amide 30, which was reduced to the
primary amine 31 with LiAlH4. Coupling of amine 31 with 11
gave analog 7 after deprotection with TFA.
Both the 3- and 4-substituted tetrahydropyran series of
analogs also showed a preference for a 1-2 methylene spacer over
a direct linkage as found in 41 and 48. The introduction of
methyl groups adjacent to the 3-substituted tetrahydropyranyl
oxygen atom, as in compound 43, was detrimental to activity
presumably due to steric interference with the channel protein.
Enantioselective recognition was observed with the 3-substituted
pyran analogs 49 and 50 where the R-enantiomer 49 (IC50 = 76
nM) was 5-fold more potent than the S-enantiomer 50 (IC50 = 328
nM) and also more selective over the hNav1.5 isoform. Both tert-
butyl and phenyl group at the α-position, as in 51 and 53,
respectively, were well tolerated, with the diastereoselectivity
dependent on the nature of the alkyl substituents: 3-fold for tert-
butyl (51 vs. 52) and 17-fold for phenyl (53 vs. 54). With respect
to the position of the ring oxygen, the 3-substituted
tetrahydropyran analogs 49 and 55 showed the best Nav1.7
inhibitory activity. The two oxetane analogs 56 and 57 that were
examined showed poor inhibitory activity. Among the
cyclohexanone derivatives 58-68, the 3-cyclohexanone analog 58
was optimal, followed by the 4-cyclohexanone methyloxime 63.
Finally, we explored several cyclic sulfone analogs represented
by 69-71 of which the most potent was 70, which exhibited an
IC50 value of 37 nM; however, poor PAMPA permeability
precluded further evaluation of this compound.
Scheme 5. Reagents and conditions: (a) I(CH2)3I (6 equiv.),
K2CO3, acetone, reflux,3 days, 62%; (b) SmI2, HMPA, THF, 0
°C, 2 h, 99%; (c) LiNH2, THF, reflux, 1 h, 45%; (d) LiAlH4, THF,
reflux, 1 h, 99%; (e) 11, K2CO3, DMF, rt, 45%; (f) TFA, DCM, rt,
36%.
3. Results and Discussion
The inhibitory activity of these compounds towards hNav1.7
and hNav1.5 channels was assessed using an automated 384-well
IonWorks Barracuda® patch-clamp assay designed to measure
potency on the inactivated channel. The Barracuda data generally
correlated well with manual patch clamp electrophysiology
results (data not shown). The PAMPA permeability of these
compounds was determined at both pH 5.5 and 7.4 in order to
cover the pH ranges in the intestinal lumen. Differences in
permeability at the two pH levels may indicate a potential for pH-
limited absorption. The inhibitory activity of these compounds
toward cytochrome P450 isoforms was also monitored closely
Table 1. hNav1.7 and hNav1.5 inhibition IC50, PAMPA permeability, and CYP3A4 inhibition IC50 dataa

hNav1.7
hNav1.5
Papp
CYP3A4
IC50 (µM)e
>20
Compd.
3
R
IC50 (nM)b
IC50 (nM)c
pH 5.5/7.4d
2.2
>30,000
18,300
NT
8.0/142
267/71
NT
>20
NT
NT
4
5
108
3400
6
7
614
169
12,800
750
NT
6.3
684/584
NT
6.3
32
33
916
260
19,400
NT
NT
>30,000
NT
NT
34
1400
931
NT
NT
NT
35f
>30,000
>20
NT
36
37
188
221
3,200
401/268
NT
10,000
>20
38
39
40
133
189
242
800
325/600
NT
NT
>20
NT
4,500
NT
272/225
41
42
336
127
>30,000
4,000
NT
>20
NT
422/270
43g
706
1,900
NT
>20
44
45
46
47
48
120
237
74
2,400
3,900
3,200
3,700
7,700
1120/736
610/610
211/520
NT
>20
>20
>20
NT
874
796
NT
>20
NT
49
50
76
6,400
5,000
477/377
NT
328

4.9
5.5
51h
52i
156
415
>30,000
>30,000
NA/677
NT/439
3.6
NT
>20
53h
54h
77
NT
NT
428/735
NT
1300
55
56
50
2,000
NT/385
NT
NT
NT
6500
14,800
57
1500
>30,000
NT
>20
>20
>20
16.5
58
52
4,700
3,500
1,800
285/233
216/162
216/162
59h
60h
281
151
61
62
162
150
6,100
2,700
451/419
538/207
>20
13.7
63
64
94
3,600
6,700
608/392
NT
NT
444
>20
>20
NT
65h
66h
156
380
1,300
730
546/207
464/101
67
68
407
348
3,200
NT
NT
NT
NT
NT
69
6400
NT
NT
>20
>20
70
71
37
6,000
NT
2.0/1.0
0/0
266
aAll values are the mean of at least two separate assay determinations. bhNav1.7 data were collected using an Ion Works Barracuda®
automated electrophysiology platform using a protocol where cells were held at at -60 mV. chNav1.5 data were collected on the same
platform using a protocol where cells were held at -50 mV. dApparent permeability at both pH 5.5 and 7.4 (nm/s). eFor assay conditions,
f
g
h
see Ref. 22. 3:2 mixture of endo/exo isomers. single isomer with relative stereochemistry undetermined. Relative stereochemistry
arbitrarily drawn and undetermined. NT: not tested.

N-(1,2,4-Thiadiazol-5-yl)benzenesulfonamide
Nav1.7
cellular IC50 value determined by electrophysiology (mNav1.7
IC50 = 54 nM).
inhibitors generally show significant inhibition of the CYP
isoforms 2C9 and 3A4.15,21 For example, reference compound 2
inhibited CYP2C9 and CYP3A4 with IC50 values of 0.17 and
0.077 µM, respectively.21 In contrast, the N-(thiazol-2-
yl)benzenesulfonamides shown in Table 1 showed low levels of
inhibition across the major human P450 isozymes (IC50 values
>20 µM for most of the analogs), with the highest level of
inhibition observed toward CYP2C9 (IC50 ~1 µM). We believe
that for these N-thiazolyl-benzenesulfonamides, the thiazole
heterocycle together with the amine linkage contributed to a
favorable CYP inhibition profile, an observation consistent with
that in other series of benzenesulfonamides which we have
recently described.19
Figure 2. Effect of 49 (100 mpk, PO) in the mouse CCI-
induced cold hyperalgesia model. Left: Latency to paw
withdrawal/licking response. Right: Percent reversal of cold
hyperalgesia. Results are presented as the mean SEM and
were analyzed by one way ANOVA followed by Dunnett’s post-
hoc tests; ***P<0.001 versus CCI/vehicle (N = 10-12/group).
As a result of its relatively potent cellular activity and
selectivity over the Nav1.5 isoform in the Barracuda assay and
good permeability in the PAMPA assay as well as a moderate
efflux ratio of 3 in the Caco-2 assay,23 compound 49 was tested
in a manual patch clamp electrophysiology (EP) assay that
measured affinities at resting and partially inactivated states of
the channel (holding potentials of −100 mV and −60 mV,
respectively) to determine the state-dependence of Nav1.7
channel block. The manual EP IC50 values were 78 and 54 nM
for the human and mouse Nav1.7 isoforms, respectively, at −60
mV holding potential, which is thought to be representative of
pain states.12 Virtually no activity was observed at the resting
state (−100 mV) of the channel and, as such, 49 was determined
to be a state-dependent inhibitor of Nav1.7. Compound 49 was
also tested for inhibition of sodium channel currents recorded
from cell bodies of individual sensory neurons dissociated from
Compound 49 was also tested for efficacy in the
complete Freund’s
adjuvant (CFA)
model
of
inflammatory pain in mice.25 This model has been used
to test Nav1.7 knockout mice, which do not develop CFA-
induced thermal hyperalgesia.6 Mice received an
intraplantar injection of either saline (sham group) or
CFA (20 µL) into the right hindpaw and thermal
hyperalgesia was tested
3 days later using the
Hargreaves test. Subjects receiving CFA showed a
marked reduction in latency to paw withdrawal
following application of a noxious thermal stimulus
compared to sham controls. CFA mice treated with 49
showed a dose-dependent increase in paw withdrawal
latency which was significant at 30 and 60 mpk at both
30 and 90 minutes post-dose (Figure 3). These results
demonstrated that 49 reversed established thermal
hyperalgesia in CFA mice; the reversal ranged from 39-
52% at the 30 and 60 mpk doses. In a similar
experiment, ibuprofen (positive control) at 300 mpk (PO)
reversed thermal hyperalgesia by 90%.
rat DRG.
In this assay, 49 inhibited fast-inactivating,
tetrodotoxin-sensitive (TTX-S) sodium currents measured in
adult mouse DRG sensory neurons with an IC50 of 34 nM.
Compound 49 was administered to mice as either an IV bolus
(2 mpk) or PO (30 mpk) to generate exposure data that would
support further testing in mouse efficacy models. Following an
IV dose, compound 49 exhibited moderate clearance (13
mL/min/kg) and a volume of distribution at steady state of 0.6
L/kg, which was consistent with total body water. After a 30 mpk
PO dose, compound 49 exhibited a Cmax of 37 18 µM, a half-
life of 3 hours and an oral bioavailability of 47%.
Compound 49 was evaluated in the mouse chronic
constriction injury (CCI) model in which three ligatures are
loosely tied around the left sciatic nerve to simulate conditions of
nerve compression injury.24 In contrast to mice receiving sham
surgery (i.e. the sciatic nerve is exposed but not ligated) CCI
mice show increased sensitivity to cold and a reduced latency to
exhibit nociceptive-like behaviors when placed on a cold plate
maintained at 5 ºC. Mice receiving CCI or sham surgery were
tested 5 weeks later and were treated orally with either vehicle or
49 30 minutes prior to testing. Figure 2 (left) shows that an oral
dose of 100 mpk 49 significantly increased the latency to elicit a
paw withdrawal/licking response in CCI mice. Figure 2 (right)
shows that the cold hyperalgesia response in CCI mice was
reversed by 86% following treatment with 100 mpk 49. In a
similar experiment, gabapentin (positive control) at 30 mpk (PO)
showed 80% reversal of CCI induced cold hyperalgesia.
Figure 3. Effect of PO dosing of 49 on latency to paw
withdrawal in the mouse CFA thermal hyperalgesia model at
30 minutes and 90 minutes post-dose. Results are presented as
the mean SEM and were analyzed at each time point by one
way ANOVA followed by Dunnett’s post-hoc tests; **P<0.01,
***P<0.001 versus CFA/vehicle (N = 11-15/group).
The total plasma concentration of 49, measured 30 minutes
after treatment with 100 mpk PO in mice, was 32 µM,
corresponding to a free plasma concentration of 0.74 µM (mouse
plasma protein binding = 2.3% free). The free plasma
concentration of 49 corresponding to an 86% reversal of cold
hyperalgesia in CCI mice was ~14-fold higher than the mouse
On completion of testing in the CFA model a subset of
animals (N = 3/dose) were euthanized and plasma and brain
collected for measurement of drug concentrations. At ~ 90

minutes post-dose, the mean ( SD) total plasma concentrations
of 49 were 0.75 0.09 µM, 1.13 0.54 µM and 2.56 0.13 µM
at the 10 mpk, 30 mpk and 60 mpk doses, respectively. The
corresponding mean plasma free concentrations of 49 were 17
nM, 26 nM and 59 nM, respectively. Interestingly, the free
plasma concentration of 49 (26 nM) corresponding to the partial
reversal of thermal hyperalgesia in CFA was only half that of the
mouse cellular IC50 value (EP mNav1.7 IC50 = 54 nM). At all
doses, the brain concentration of 49 was below the assay limit of
quantification (<25 nM) suggesting that 49 shows low
penetration into the CNS. Measureable brain concentrations of 49
were achieved in mice dosed orally at a dose of 100 mpk and
confirmed that 49 is a peripherally-restricted molecule with a
brain/plasma ratio of <0.02 in mice (Table 2).
phase II nociceptive behaviors (Figure 4). We also collected
plasma, DRG and trigeminal ganglion (TGG) from experimental
animals at 60 minutes post-dose and on satellite animals at 30
minutes post-dose. The total plasma concentrations of 49 were 13
and 7.3 µM at 30 and 60 minutes post-dose, respectively, and the
free plasma concentrations were 0.30 and 0.17 µM (mouse
plasma protein binding = 2.3% free). The free plasma
concentration of 49 corresponding to 36% reversal of phase II
nociceptive behaviors was 3- to 6-fold higher than the EP mouse
cellular IC50 value.
This compound achieved DRG
concentrations of 1.1 and 0.7 µM, respectively, at 30 and 60
minutes post-dose, and TGG concentrations of approximately 6
µM at both time points (Table 3). These results support our
hypothesis that improving membrane permeability can enhance
DRG and TGG exposure. The free DRG concentration of 49
(mouse DRG protein binding = 10% free) was 1- to 2-fold higher
than the EP mouse cellular IC50 value. However, protein binding
data specifically in mouse TGG tissues are needed to estimate the
free TGG concentration/mouse Nav1.7 IC50 ratios corresponding
to the reversal of phase II nociceptive behaviors in the formalin
test.
Table 2. Brain and plasma concentrations after oral dosing of
49 (100 mpk)a
Time point
Post-dose
Plasma Conc.
(µM)
Brain Concc
(µM)
0.12 (0.06)
Brain/Plasma
Ratio
0.017 (0.006)
30 minutes
90 minutes
7.34 (1.11)
3.67 (0.96)
0.042 (0.012) 0.012 (0.003)
aCompound dosed PO to male mice in 0.5% Methocel/0.1%
Tween/99.4 water. Results are presented as the mean (SD), N =
3/group.
Compound 49 was then examined in the formalin test model
measures
which
spontaneous,
nociceptive-like
paw
flinching/licking responses after peripheral nociceptor activation.
This model was chosen for in vivo efficacy screening since
formalin-induced nociceptive behaviors are abolished in global
Nav1.7 knockout mice.6
Male CD1 mice were injected
subcutaneously in the dorsal surface of the right hind paw with
20 µL of a 5% formalin solution and nociceptive behavior was
recorded for 45 minutes using an automated system (Automated
Nociception Analyzer, UCSD, La Jolla, CA, USA). Formalin
injection elicits a biphasic behavioral response: phase I occurs
during the first 5 min and results from direct nociceptor
activation whereas phase II occurs 15-45 minutes after injection
and is thought to involve spinal sensitization. Treatment with
compound 49 (30 mpk, PO) 15 minutes prior to formalin
injection had no effect on phase I responses but significantly
reversed phase II nociceptive behaviors by 36% (Figure 4). In
this assay, mexiletine (30 mpk, PO), a non-selective sodium
channel blocker which was used as the positive control, abolished
Figure 4. Effect oral administration of 49 (30 mpk) on phase
I and II formalin-induced nociceptive behaviors. Mexiletine (30
mpk, PO) was used as the positive control. Results are
presented as the mean
SEM response (n=12-14/group) and
were analyzed by ANOVA followed by Dunnett’s post-hoc tests;
*P<0.05, ***P<0.001 compared to vehicle/formalin group. Mex:
Mexiletine.
Table 3. Plasma, DRG and TGG concentrations after oral dosing of 49 (30 mpk)a
Time point
Post-dose
30 min
Plasma
Free Plasma
Conc. (µM)b
0.30 (0.05)
0.17 (0.09)
DRG
Free DRG
Conc. (µM)c
0.11 (0.02)
0.07 (0.01)
TGG
Conc. (µM)
13.0 (2.1)
7.3 (4.0)
Conc. (µM)
1.10 (0.21)
0.66 (0.19)
Conc. (µM)
5.8 (0.6)
5.0 (3.3)
60 min
aCompound dosed PO to male mice in 0.5% Methocel/0.1% Tween/99.4% water (10 mL/kg). Results are presented as the mean
(SD), N = 3–9/group. Calculated using unbound fraction in mouse plasma (2.3%). Calculated using unbound fraction in mouse
b
c
DRG
(10%).
Encouraged by the efficacy observed in the mouse CCI
measured in the ipsilateral biceps femoris using
electromyography (EMG). The NFR is a translational
measure of nociceptive neuron activation that can be
used as a pharmacodynamic marker across multiple
species. In humans, the stimulus intensity at which the
NFR is evoked is highly correlated with the subjective
and CFA and the formalin test models, we further
investigated the effect of compound 49 on the electrical
threshold for the nociceptive flexion reflex (NFR) in
mice. The NFR is a muscle withdrawal reflex, primarily
in response to activation of A-delta nociceptive neurons,
that is elicited by electrical stimulation to the foot and is
experience of pain.26 As
a
spinal reflex, NFR

encompasses activity through afferent and efferent
nerve fibers as well as neurotransmission across
multiple synapses within the spinal cord and offers
multiple sites for pharmacological modulation.
Interestingly, clinically-effective pain medications of
diverse mechanisms of action elevate the NFR in
humans,27 presumably by engaging one or more of these
sites. Pertinent to the role of sodium channels in
mediating NFR, in human subjects local application of
the non-selective sodium channel blocker lidocaine in
the area of electrical stimulation26 or following
intravenous administration28 robustly elevated the NFR
threshold. We first validated the NFR assay in mice
using systemic administration of mexiletine. An IP dose
of mexiletine (10 mpk) elevated the A-delta current
threshold to greater than 15 mA (a ceiling response in
this assay) in comparison to 0.2 mA obtained after
vehicle injection (N = 5). This effect was of rapid onset,
manifest after 15 minutes post-dose and lasted for at
least 60 minutes post-dose.29 Clonidine (0.1 mpk, SC), an
(8.94 2.5 mA) compared to a vehicle injection (0.71
0.33 mA). For the evaluation of 49, cohorts of mice were
first dosed with vehicle (IP),31 and the NFR threshold
was determined at 15 minute intervals up to an hour.
This was performed in order to both establish and
confirm the stability of the measurement. In the control
cohort, this was followed by a second vehicle injection
and further testing for a second hour. In the treatment
cohort, the second injection was compound 49 (10 or 30
mpk, IP). As shown in Figure 5 (left), compound 49
elicited a significant and dose-dependent elevation of the
NFR threshold when compared to vehicle. The free
plasma concentration of compound 49 obtained 60
minutes after the second injection from individual mice
correlated strongly with the corresponding NFR
threshold (Figure 5, right). The total plasma levels at 60
minutes post-dose were 29 7.2 µM and 82 42.7 µM,
and the free drug concentrations were calculated to be
0.66 0.17 µM and 1.9 0.98 µM for the 10 and 30 mpk
doses, respectively. More studies are needed to further
understand the PK/PD relationship.
adrenergic agonist that has been previously shown to
2
be effective in humans30 also elevated the NFR threshold
Figure 5. Left: Effect of 49 on the NFR threshold following IP administration. Right: Correlation between the NFR threshold and
free plasma 49 concentration in individual mice ~60 minutes after IP dosing. NFR threshold results are presented as the mean
SEM and were analyzed by repeated measures 2 way ANOVA followed by Dunnett’s post-hoc tests; **P<0.01, ****P=0.0001 versus
vehicle treatment (N = 8-9/group).
microelectrodes filled with internal solution containing (in mM)
50 CsCl, 90 CsF, 10 NaF, 2 MgCl2, 10 EGTA, 10 HEPES, pH
4. Conclusion
Benzenesulfonamide-based Nav1.7 inhibitors showed poor
membrane permeability that we attributed to their zwitterionic
character, stimulating an approach to optimization that relied
upon removing the basic amine moiety. This design concept
7.2 with CsOH had resistances of 1-4 MOhms. The external
solution was (in mM): 150 NaCl, 4 KCl, 1 MgCl2, 1.8 CaCl2, 10
HEPES, 10 glucose, pH 7.4 with NaOH. Series resistance was
kept below 10 MOhms and electronically compensated at least
50%. Currents were acquired at 50 kHz via an EPC-10 amplifier
with Patchmaster software (HEKA Instruments Inc., Bellmore,
NY). Leakage subtraction was performed with the P/N method.
Solution exchange was performed with a 16-well Dynaflow
Resolve glass chip (Cellectricon, Sweden). The holding potential
was -70 mV unless otherwise indicated. Following a 100 msec
prepulse to -130 mV, Nav1.7 current was elicited by a 10 msec
step to +10 mV. Peak inward current was measured with
PatchMaster software and percent inhibition calculated. IC50
values were obtained by fits of the Hill equation to the average
percent inhibition data. Data are presented as mean S.E.M.
resulted in
a series of non-zwitterionic aryl sulfonamide
derivatives as selective Nav1.7 inhibitors that exhibited improved
membrane permeability at pH 5.5. This work led to the discovery
of 49 as a selective Nav1.7 channel inhibitor that demonstrated
efficacy following oral administration in three mouse models of
pain. Moreover, in the NFR assay, compound 49 showed a robust
elevation of the threshold to elicit an EMG response to
nociceptor neuron activation that was highly correlated with
plasma exposure paving a path for clinical translation. These
results indicate that non-zwitterionic aryl sulfonamide Nav1.7
inhibitors have the potential to be developed for the treatment of
pain.
Automated Electrophysiology Methods: Ion Works
Barracuda population patch clamp (PPC). Population patch
clamp (PPC) measurements from HEK 293 cells expressing
Nav1.7 and Nav1.5 channels were performed using an IonWorks
Barracuda instrument (Molecular Devices Corporation, Union
City, CA) using a proprietary PPC PatchPlate™ substrate
(Molecular Devices Corporation) with 64 patch clamp apertures
per well. The ability to average Nav1.x currents from 64
recordings from each well greatly improves data consistency and
5. Experimental Section
Patch clamp electrophysiology. Nav1.7 currents were
recorded using whole-cell patch clamp of HEK293 cells stably
expressing recombinant human or mouse Nav1.7. Glass

recording success rates in the measurement of Nav1.x mediated
ionic currents. Calculated leak current was digitally subtracted
from the total cell Nav1.x current for each sample data point
acquired.
recirculating blanket to maintain body temperature at
approximately 37°C. Anesthesia was maintained (1.5-2.5%
isoflurane) throughout surgery. An approximately 1.5 cm
incision was made down the center of the thigh and the common
sciatic nerve was exposed through blunt dissection. Three
ligatures of 5-0 chromic gut were loosely tied proximal to the
trifurcation approximately 1mm apart so that the epineural
circulation was preserved. The muscle was closed with 4-0
sutures, and the skin was closed with wound clips. All mice
Nav1.7 currents were elicited by a voltage clamp protocol
designed to strongly bias the Nav1.7 channels toward their
inactivated state as follows. From a holding potential of –60 mV
cells were briefly hyperpolarized to -100 mV for 1.25 sec, then
stepped to -20 mV for 20 sec to inactivate the channels. This was
followed by a relatively brief hyperpolarization to -100 mV for
300 ms, then a 20 msec test pulse to -20 mV to elicit the Nav1.7
current used to measure the pharmacology of all test compounds.
Compounds were incubated for 600 sec between the pre- and
post-compound reads. The external recording solution used was
(in mM) 137 NaCl, 4 KCl, 1 MgCl2, 1.8 CaCl2, 10 Hepes, 10
glucose, pH to 7.4 with NaOH, and the internal solution used was
(in mM) 100 K-gluconate, 40KCl, 3.2 MgCl2, 5 EGTA, 10
HEPES pH to 7.2 with KOH. The same solutions were used to
record Nav1.5 currents, with the following voltage clamp
protocol. Nav1.5 currents were elicited by a voltage clamp
protocol designed to bias the Nav1.5 channels to their inactivated
state as follows. From holding potential of –40 mV cells were
briefly hyperpolarized to -100 mV for 300 ms, then stepped to -
10 mV for 20 sec to inactivate the channels. This was followed
by a relatively brief hyperpolarization to -100 mV for 30 ms, then
a 20 msec test pulse to -10 mV to elicit the Nav1.5 current used to
measure the pharmacology of all test compounds.
received
1 mL of a warmed lactated Ringer's solution,
subcutaneously for fluid replacement at the end of surgery. 5
weeks following surgery mice were dosed (10 mpk) orally with
the compound or vehicle and tested 30 min post-treatment on a
5°C cold plate (IITC model # 400, Series 8). Upon lifting of the
left hind paw, the timer was stopped and the mouse removed
from the plate.
Complete
Freund’s
adjuvant
(CFA)-induced
inflammatory pain model and thermal test. Efficacy in the
complete Freund’s adjuvant (CFA)-induced inflammatory pain
model in mice was also tested. 7-8 weeks mice were acclimated
to the Hargreave's equipment (IITC Plantar, set to 30 °C, 60%
intensity, max temperature of 28.5°C) for at least 60 min each
day for up to 3 days prior to testing. On the last day of
acclimation, the baseline of thermal escape latency was taken by
applying a radiant heat stimulus to the plantar surface of each
hind paw. Upon lifting of the paw, the light beam was shut off
and the timer stopped. The average of 3 latencies was recorded.
Each mouse was then injected with 20 µL CFA (Sigma F5881)
subcutaneously into the intraplantar region of the right paw. 72 h
later, mice were dosed (10 mpk) orally with the compound or
vehicle and, at both 30 and 90 min post-treatment, thermal escape
latency was taken by applying a radiant heat stimulus to the
plantar surface of each hind paw. Upon lifting of the paw, the
light beam is shut off and the timer stopped. The average of 3
latencies was recorded.
HEK 293 cells expressing Nav1.7 and Nav1.5 channels, were
used (Essen Biosciences, Ann Arbor, Michigan). Cells were
cultured in T-175 flasks and passaged every 2 to 3 days at 1:3 to
1:6 seeding density dilutions. Cells were grown to 70% to 90%
confluence in a flask and removed from the incubator (37° C, 5%
CO2) 1 to 3 days after plating. Growth medium was aspirated
from the culture flasks. Cells were gently rinsed with 10 mL of
Phosphate buffered saline (PBS) to remove residual media. Next
a total of 2 mL TrypLE (Gibco) solution was added, and the
flasks containing cells were sat for 3 min at RT, after which, the
cells became visibly rounded and were easily dislodged from the
bottom of the flask with a few brief taps on a solid surface. A
total of 8 mL of media was added to the flask to inactivate the
TrypLE, and the mixture was centrifuged at 910 rpm for 4 min.
The cell supernatant was decanted, and the cell pellets were
resuspended in 5-6 mL of external solution followed by gentle
trituration using a 10 mL pipette, and transferred to a 15 mL
conical tube and immediately brought to the IW Barracuda
instrument. The cell suspension had a final concentration of ∼2
to 3 million cells per mL; this corresponds to 10,000 cells added
per well.
Formalin test. Persistent pain was induced with an injection
of formalin in the dorsum of one hind paw. Formalin-induced
flinching was quantified with an automated system (Automated
Nociception Analyzer, UCSD, La Jolla, CA, USA). A metal band
was glued on the right hind paw, the mouse was then acclimated
for 60 min in an individual Plexiglas test chamber. Each mouse
was dosed orally with the compound (10 mL/kg) or vehicle, 15
min before formalin injection. The right hind paw was injected
with 20 µL of 5% formalin (SC of formalin w/v in saline,
Sigma), and the animal was immediately placed back into the
chamber. Each time the band breaks the electromagnetic field
located under the mouse a signal was generated and nociceptive
behavior, paw flinches and licking, were recorded with the
automated system for 45 minutes.
Peak membrane currents were analyzed with IW Barracuda
software and exported to Excel for further analysis.
Concentration response curve fitting was performed with BMS
in-house software. IC50 values were obtained by fits of the Hill
equation to the average percent inhibition data plotted versus
compound concentration. Concentration-response curves for all
test compounds were fitted to a 4-parameter equation: % of
control = 100 (1 + ([drug]/IC50)p)–1, where IC50 is the
concentration of drug required to inhibit current by 50% and p is
the Hill slope.
Electrical threshold for the nociceptive flexion reflex
(NFR). Urethane anesthetized male mice were equipped with
wire electrodes in the sural field for transcutaneous electrical
stimulations (ES) and the biceps femoris on the same side was
used for recording extracellular muscle action potentials (APs)
using fine wire or needle electrodes. Brief ES (e.g. 0.1-0.4 ms
pulses; 0.1-15 mA; 200 Hz; 25 ms) were delivered at 30 s
intervals till a reproducible response was obtained across three
consecutive trials. Lowest current required to produce clear A-
delta related APs was noted as NFR threshold. After vehicle (IP)
injection, the threshold was determined at 15 min intervals up to
an hour. In control mice, this was followed by a second vehicle
injection and testing for another hour. In treatment mice, the
second injection was compound 49 (10 or 30 mpk). Threshold
responses were compared between the second vehicle and the
Chronic constriction injury (CCI) and cold plate test. CCI
of the sciatic nerve was induced by ligation of the left sciatic
nerve. 8-9 weeks old mice were initially anesthetized by
inhalation of 1.5-4% isoflurane in an induction chamber. The left
hind limb was shaved and prepared with betadine prior to
surgery. The animals were then placed on a warm water

drug using a 2-way ANOVA followed by a Dunnett’s test. Blood
was withdrawn from the mouse immediately after the 60 min
threshold evaluation and the plasma was isolated for exposure
analysis.
4.8 Hz, 1H), 6.86 - 6.82 (m, 2H), 6.87 (s, 1H), 6.25 (s, 1H), 5.40
(s, 1H), 3.82 (t, J = 6.8 Hz, 4H), 2.55 (s, 2H), 1.94 - 1.84 (m,
4H). LC/MS m/z: (M + H)+ calcd for C16H18ClFN3O4S2, 434.04;
found 434.0.
Chemistry. Proton magnetic resonance (1H NMR) spectra
were recorded on either a Bruker Avance 300 or 400 or a JEOL
Eclipse 500 spectrometer and are reported in ppm relative to the
reference solvent of the sample in which they were run. HPLC
and LC/MS analyses were conducted using a Shimadzu SCL-
5-Chloro-2-fluoro-4-(((6a-methylhexahydrofuro[2,3-
b]furan-3a-yl)methyl)amino)-N-(thiazol-2-
yl)benzenesulfonamide (5). Prepared from 21 using the
1
procedures described for 4. H NMR (500 MHz, DMSO-d6) δ
7.62 (d, J = 7.3 Hz, 1H), 7.26 (d, J = 4.8 Hz, 1H), 6.93 (d, J =
12.8 Hz, 1H), 6.82 (d, J = 4.4 Hz, 1H), 5.60 (br s, 1H), 3.84 -
3.71 (m, 4H), 2.55 (s, 2H), 2.11 - 1.88 (m, 4H), 1.33 (s, 3H).
LC/MS m/z: (M + H)+ calcd for C17H20ClFN3O4S2, 448.06; found
448.0.
−vis detector at 220 nm
10A liquid chromatograph and a SPD UV
with the MS detection performed with either a Micromass
Platform LC spectrometer or Waters Micromass ZQ
spectrometer. All flash column chromatography was performed
−60 µm). All
a
on EM Science silica gel 60 (particle size of 40
5-Chloro-2-fluoro-4-(((hexahydro-2H-furo[2,3-b]pyran-3a-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide
reagents were purchased from commercial sources and used
without further purification unless otherwise noted. All reactions
were performed under an inert atmosphere. HPLC analyses were
performed using the following conditions. All final compounds
had an HPLC purity of ≥ 95% unless specifically mentioned.
(6).
Step A: A mixture of 27 TFA salt (58 mg, 0.37 mmol), 11 (187
mg, 0.41 mmol) and Cs2CO3 (264 mg, 0.81 mmol) in DMF (3.7
mL) was stirred at 65 °C for 12 h. The reaction mixture was
filtered through a pad of Celite and the filtrate was concentrated
to give crude 5-chloro-N-(2,4-dimethoxybenzyl)-2-fluoro-4-
(((hexahydro-2H-furo[2,3-b]pyran-3a-yl)methyl)amino)-N-
(thiazol-2-yl)benzenesulfonamide, which was used directly for
the next step. MS: 596.0 (M - H)+. Step B: A mixture of the crude
5-chloro-N-(2,4-dimethoxybenzyl)-2-fluoro-4-(((hexahydro-2H-
furo[2,3-b]pyran-3a-yl)methyl)amino)-N-(thiazol-2-
HPLC Methods. Analytical HPLC analyses were carried out
following methods A and B, and preparatory reverse-phase scale
purifications were performed using method C.
Method A. A linear gradient using 5% acetonitrile, 95%
water, and 0.05% TFA (solvent A) and 95% acetonitrile, 5%
water, and 0.05% TFA (solvent B) (t = 0 min, 10% B; t = 15 min,
100% B) was employed on a SunFire C18 3.5 µm 3.5 mm × 150
mm column. Flow rate was 0.5 mL/min, and UV detection was
set to 220 nm. The LC column was maintained at ambient
temperature.
yl)benzenesulfonamide obtained from Step A and TFA (129 µL,
1.7 mmol) in DCM (0.84 mL) was stirred at rt for 30 min. DCM
was removed, and crude product was purified by preparative
HPLC (Method C) to give 6 (35 mg, 21% over 2 steps). 1H NMR
(500 MHz, DMSO-d6) δ 7.58 (d, J = 7.3 Hz, 1H), 7.26 (d, J =
4.8 Hz, 1H), 6.87 - 6.77 (m, 2H), 6.24 (br t, J = 6.1 Hz, 1H), 4.70
(s, 1H), 3.97 - 3.87 (m, 2H), 3.77 - 3.65 (m, 1H), 3.35 - 3.10 (m,
3H), 1.99 - 1.86 (m, 1H), 1.76 - 1.66 (m, 3H), 1.61 - 1.41 (m,
Method B. A linear gradient using 5% acetonitrile, 95%
water, and 0.05% TFA (solvent A) and 95% acetonitrile, 5%
water, and 0.05% TFA (solvent B) (t = 0 min, 10% B; t = 15 min,
100% B (20 min)) was employed on a XBridge Ph 3.5 µm 3.0
mm × 150 mm column. Flow rate was 0.5 mL/min, and UV
detection was set to 220 nm. The LC column was maintained at
ambient temperature.
2H), 1.23 (s, 3H). LC/MS m/z: (M
C17H20ClFN3O4S2, 448.06; found 448.0.
+
H)+ calcd for
5-Chloro-2-fluoro-4-(((6a-hydroxyoctahydropentalen-3a-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide
Prepared from 31 using the procedures described for 4. 1H NMR
(500 MHz, DMSO-d6) δ 7.58 (d, J = 7.3 Hz, 1H), 7.24 (d, J =
4.4 Hz, 1H), 6.82 (d, J = 4.6 Hz, 1H), 6.80 - 6.74 (m, 1H), 6.19
(br s, 1H), 3.24 (d, J = 5.9 Hz, 2H), 1.72 - 1.63 (m, 5H), 1.62 -
1.52 (m, 4H), 1.48 - 1.31 (m, 5H). LC/MS m/z: (M + H)+ calcd
for C18H22ClFN3O3S2, 446.08; found 446.0.
(7).
Method C. Column: Waters XBridge C18, 19 x 200 mm, 5
µm particles; Guard Column: Waters XBridge C18, 19 x 10 mm,
5 µm particles; Mobile Phase A: water with 20 mM ammonium
acetate; Mobile Phase B: 95:5 methanol:water with 20 mM
ammonium acetate; Gradient: 25-65% B over 40 minutes, then a
5-minute hold at 100% B; Flow: 20 mL/min.
Methyl
4-(benzyloxy)-2-(2-(benzyloxy)ethyl)-2-
5-Chloro-2-fluoro-4-(((hexahydrofuro[2,3-b]furan-3a-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide
cyanobutanoate (14). To a solution of methyl 2-cyanoacetate
(0.89 mL, 10.1 mmol) in DMF (20 mL) at rt was added sodium
hydride (0.97 g, 24.2 mmol), and the reaction mixture was stirred
at rt for 15 min. Tetrabutylammonium iodide (0.37 g, 1.0 mmol)
and ((2-bromoethoxy)methyl)benzene (3.35 ml, 21.19 mmol)
were added, and the reaction mixture was stirred at 90 °C for 3 h.
Water was added and the aqueous layer was extracted with ether
(x3). The combined organic layers were washed with brine, dried
over anhydrous sodium sulfate, and filtered, and the filtrate was
concentrated to give the crude product. The crude product was
purified by silica gel chromatography eluting with 0-30%
EtOAc/hexanes to give 14 (2.4 g, 65 % yield) as a colorless oil.
1H NMR (500 MHz, CDCl3) δ 7.41 - 7.27 (m, 10H), 4.49 (s, 4H),
3.78 - 3.69 (m, 5H), 3.45 (s, 3H), 2.49 - 2.40 (m, 2H), 2.06 (dt, J
= 14.2, 4.7 Hz, 2H). LC/MS m/z: (M + H)+ calcd for C22H26NO4,
368.19; found 368.3.
(4).
Step A: A mixture of 19 TFA salt (9 mg, 0.06 mmol), 11 (32 mg,
0.07 mmol) and Cs2CO3 (45 mg, 0.14 mmol) in DMF (0.63 mL)
was stirred at 65 °C for 12 h. The reaction mixture was filtered,
and the crude product was purified by preparative TLC on silica
gel eluting with 80% EtOAc/hexanes to give 5-chloro-N-(2,4-
dimethoxybenzyl)-2-fluoro-4-(((hexahydrofuro[2,3-b]furan-3a-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (20 mg,
54%) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.77 (d, J =
6.9 Hz, 1H), 7.40 (d, J = 3.5 Hz, 1H), 7.23 (d, J = 8.2 Hz, 1H),
6.98 (d, J = 3.5 Hz, 1H), 6.43 - 6.30 (m, 3H), 5.47 (s, 1H), 5.21
(s, 2H), 5.05 - 4.95 (m, 1H), 4.09 (dd, J = 8.8, 5.0 Hz, 4H), 3.82 -
3.70 (m, 7H), 3.35 (d, J = 5.2 Hz, 2H), 2.14 - 1.98 (m, 4H). 584.2
(M
+
H)+. Step B:
A
solution of 5-chloro-N-(2,4-
dimethoxybenzyl)-2-fluoro-4-(((hexahydrofuro[2,3-b]furan-3a-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (20 mg,
0.034 mmol) and TFA (0.05 mL) in DCM (0.34 mL) was stirred
at rt for 1 h. The solvents were removed, and residue was purified
tert-Butyl
(4-(benzyloxy)-2-(2-(benzyloxy)ethyl)-2-
(hydroxymethyl)butyl)carbamate (15). Step A: To a solution of
14 (400 mg, 1.1 mmol) THF (5 mL) and ether (5 mL) at rt was
1
via preparative HPLC (Method C) to give 4 (6 mg). H NMR
(500 MHz, DMSO-d6) δ 7.59 (d, J = 7.3 Hz, 1H), 7.25 (d, J =

added LiAlH4 (83 mg, 2.2 mmol), and the reaction mixture was
heated under reflux for 4 h. Sodium sulfate decahydrate was
added, and the reaction mixture was stirred at rt for 1 h. The
mixture was filtered, and the filtrate was evaporated to give crude
2-(aminomethyl)-4-(benzyloxy)-2-(2-(benzyloxy)ethyl)butan-1-
ol (349 mg, 93%). This material was used directly for the next
reaction. Step B: To a solution of crude 2-(aminomethyl)-4-
(benzyloxy)-2-(2-(benzyloxy)ethyl)butan-1-ol (830 mg, 0.41
mmol) DCM (5 mL) at rt was added Boc2O (0.62 mL, 2.7 mmol),
and the reaction mixture was stirred at rt for 5 h. DCM was
removed, and the residue was purified by silica gel
chromatography eluting with 0-30% EtOAc/hexanes to give 15
hydroxyethyl)butyl)carbamate (109 mg, 0.24 mmol) in DCM (1.6
mL) was added dropwise. The mixture was stirred for 15 min and
then triethylamine (232 µL, 1.67 mmol) was added. The mixture
was stirred for 15 min and allowed to warm to room temperature.
The crude product was purified by prep TLC eluting with 40%
EtOAc/hexanes to give 20 (65 mg, 60%) as a colorless oil. 1H
NMR (500 MHz, CDCl3) δ 7.43 - 7.20 (m, 11H), 4.43 (s, 4H),
3.57 - 3.35 (m, 6H), 2.15 (s, 3H), 2.05 - 1.95 (m, 2H), 1.93 - 1.85
(m, 2H), 1.43 (s, 9H). LC/MS m/z: (M + H)+ calcd for
C27H38NO5, 456.27; found 456.3.
(6a-Methylhexahydrofuro[2,3-b]furan-3a-yl)methanamine
TFA salt (21). Step A: A suspension of 20 (65 mg, 0.14 mmol)
and 20% palladium hydroxide on carbon (40 mg) in methanol
(1.8 mL) was hydrogenated with a hydrogen balloon for 1 h. The
reaction mixture was filtered through a pad of Celite and the
1
(0.71 g, 66%). H NMR (400 MHz, CDCl3) δ 7.43 - 7.20 (m,
10H), 4.48 (s, 4H), 3.66 - 3.52 (m, 4H), 3.31 (d, J = 7.3 Hz, 2H),
3.09 (d, J = 6.6 Hz, 2H), 1.73 - 1.52 (m, 4H), 1.49 - 1.38 (m,
9H). LC/MS m/z: (M + H)+ calcd for C26H38NO5, 444.27; found
444.3.
filtrate was evaporated to give
methylhexahydrofuro[2,3-b]furan-3a-yl)methyl)carbamate (34
tert-butyl ((6a-
1
mg, 93%) as a colorless oil. H NMR (500 MHz, CDCl3) δ 4.63
tert-Butyl
(4-(benzyloxy)-2-(2-(benzyloxy)ethyl)-2-
formylbutyl)carbamate (16). A solution of (COCl)2 (0.21 mL,
2.4 mmol) in DCM (5.4 mL) was cooled to -78°C and a solution
of DMSO (0.32 mL, 4.5 mmol) in DCM (5.4 mL) was added
dropwise. The mixture was stirred for 10 min, and a solution of
15 (710 mg, 1.6 mmol) in DCM (5.4 mL) was added dropwise.
The mixture was stirred for 15 min and then triethylamine (1.6
mL, 11.2 mmol) was added. The mixture was stirred for 15 min
and allowed to warm to room temperature. The crude product
was purified by silica gel chromatography eluting with 0-10%
(br. s., 1H), 4.02 - 3.87 (m, 4H), 3.28 (d, J = 6.0 Hz, 2H), 2.10 -
1.88 (m, 4H), 1.51 - 1.45 (m, 9H), 1.41 (s, 3H). Step B: A
solution of crude tert-butyl ((6a-methylhexahydrofuro[2,3-
b]furan-3a-yl)methyl)carbamate (18 mg, 0.07 mmol) and TFA
(0.05 mL) in DCM (0.7 mL) was stirred at rt for 2 h. DCM was
removed to give 21 TFA salt (13 mg, 100%). This material was
used directly for the next reaction. H NMR (500 MHz, CD3OD)
δ 4.02 - 3.91 (m, 4H), 3.07 (s, 2H), 2.14 - 2.03 (m, 4H), 1.36 (s,
3H).
1
1
EtOAc/hexanes to give 16 (0.58 g, 82%) as a colorless oil. H
Methyl
5-(benzyloxy)-2-(2-(benzyloxy)ethyl)-2-
NMR (500 MHz, CDCl3) δ 9.54 (s, 1H), 7.41 - 7.25 (m, 12H),
4.45 (s, 4H), 3.60 - 3.46 (m, 4H), 1.99 - 1.80 (m, 4H), 1.49 - 1.39
(m, 9H). LC/MS m/z: (M + H)+ calcd for C26H36NO5, 442.26;
found 442.3.
cyanopentanoate (22). Step A: methyl 2-cyanoacetate (23 g,
232 mmol) was added dropwise to a suspension of sodium
hydride (60%, 2.79 g, 69.7 mmol) in DMF (100 mL) and THF
(100 mL) at 0 ºC, and the reaction mixture was warmed to rt and
stirred at rt for 30 min. ((2-Bromoethoxy)methyl)benzene (10 g,
46.5 mmol) was added, and the reaction mixture was stirred at rt
for 12 h. Water was added and the aqueous layer was extracted
with ether (x3). The combined organic layers were washed with
brine, dried over anhydrous sodium sulfate, and filtered, and the
filtrate was evaporated to give the crude product. Excessive
methyl 2-cyanoacetate was removed by distillation under high
vacuum at 120-130 ºC, and remaining residue was purified by
silica gel chromatography eluting with 0-50% EtOAc/hexanes to
give methyl 4-(benzyloxy)-2-cyanobutanoate (8 g, 74%) as a
colorless liquid. 1H NMR (300 MHz, CDCl3) δ 7.3 (m, 5H), 4.51
(s, 2H), 3.70 (s, 3H), 3.6-3.9 (m, 3H), and 2.3 (m, 2H). LC/MS
m/z: (M + H)+ calcd for C13H16NO3, 234.11; found 234.4. Step B:
To a suspension of sodium hydride (60%, 9 g, 225 mmol) in THF
(170 mL) and DMF (170 mL) was added methyl 4-(benzyloxy)-
2-cyanobutanoate (35 g, 150 mmol) at 0 ºC, and the reaction
tert-Butyl
((hexahydrofuro[2,3-b]furan-3a-
yl)methyl)carbamate (18). A suspension of 16 (27 mg, 0.06
mmol) and 20% palladium hydroxide on carbon (17 mg) in
methanol (3 mL) was hydrogenated with a hydrogen balloon for
1 h. The reaction mixture was filtered through a pad of Celite and
the filtrate was concentrated to give 18 (15 mg, 100%) as a
1
colorless oil. H NMR (500 MHz, CDCl3) δ 5.34 (s, 1H), 4.06 -
3.94 (m, 1H), 3.41 - 3.29 (m, 1H), 2.01 (dt, J = 12.6, 8.6 Hz,
1H), 1.88 (ddd, J = 12.7, 6.0, 4.0 Hz, 1H), 1.49 (s, 9H).
(Hexahydrofuro[2,3-b]furan-3a-yl)methanamine TFA salt
(19). To a solution of 18 (15 mg, 0.06 mmol) in DCM (1.2 mL)
at rt was added TFA (95 µL, 1.23 mmol), and the reaction
mixture was stirred at rt for 3 h. The solvents were removed to
give 19 TFA salt (10 mg, 100%) as yellowish oil. 1H NMR (500
MHz, CD3OD) δ 5.40 (s, 1H), 4.08 - 4.00 (m, 4H), 3.20 (s, 2H),
2.13 - 1.99 (m, 4H).
mixture
was
stirred at
rt
for
30
min.
((3-
Bromopropoxy)methyl)benzene (37.8 g, 165 mmol) was added,
and the reaction mixture was stirred at rt for 12 h. Water was
added and the aqueous layer was extracted with ether (x3). The
combined organic layers were washed with brine, dried over
anhydrous sodium sulfate, and filtered, and the filtrate was
evaporated to give the crude product. The crude product was
purified by silica gel chromatography eluting with 0-40%
EtOAc/hexanes to give 22 (40 g, 70%) as a colorless oil. 1H
NMR (400 MHz, CDCl3) δ 7.3 (m, 10H), 4.48 (2H, s), 4.47 (2H,
s), 3.7 (m, 2H), 3.57 (3H, s), 3.48 (m, 1H), 2.42 (m, 1H), 1.8-2.1
(m, 4H), and 1.6 (m, 1H). LC/MS m/z: (M + H)+ calcd for
C23H28NO4, 382.20; found 382.4.
tert-Butyl
(2-acetyl-4-(benzyloxy)-2-(2-
(benzyloxy)ethyl)butyl)carbamate (20). Step A: To a solution
of 16 (113 mg, 0.26 mmol) in THF (1.3 mL) at rt was added
MeMgBr in THF (0.77 mL, 0.77 mmol), and the reaction mixture
was stirred at rt for 2 h. Saturated ammonium chloride was added
and the aqueous layer was extracted with EtOAc (x3). The
combined organic layers were washed with brine, dried over
anhydrous sodium sulfate, and filtered, and the filtrate was
concentrated to give
tert-butyl (4-(benzyloxy)-2-(2-
(benzyloxy)ethyl)-2-(1-hydroxyethyl)butyl)carbamate (109 mg,
93%) as a colorless oil. This material was used directly for the
next reaction. Step B: A solution of (COCl)2 (42 µL, 0.48 mmol)
in DCM (1.6 mL) was cooled to -78°C and a solution of DMSO
(68 µL, 0.95 mmol) in DCM (1.6 mL) was added dropwise. The
mixture was stirred for 10 min, and a solution of tert-butyl (4-
(benzyloxy)-2-(2-(benzyloxy)ethyl)-2-(1-
tert-Butyl
(5-(benzyloxy)-2-(2-(benzyloxy)ethyl)-2-
(hydroxymethyl)pentyl)carbamate (23). Step A: To a solution
of 22 (15 g, 39.3 mmol) in THF (150 mL) was added LiAlH4

(1M in THF, 74.7 mL, 74.7 mmol) at 0 ºC, and the reaction
mixture was stirred at 0 ºC for 1 h. The reaction mixture was
poured to ice slowly, and EtOAc was added. The resulting
suspension was filtered, and the organic layer was dried over
Na2SO4, filtered and concentrated to give crude 2-(aminomethyl)-
5-(benzyloxy)-2-(2-(benzyloxy)ethyl)pentan-1-ol (8.3 g, 59%).
MS: 358.2 (M + H)+. This crude material was used directly for
Step B. Step B: To a solution of the 2-(aminomethyl)-5-
(benzyloxy)-2-(2-(benzyloxy)ethyl)pentan-1-ol (3.3 g, 9.23
mmol) in DCM (10 mL) was added Boc2O (3.2 mL, 13.9 mmol),
and the reaction mixture was stirred at rt for 12 h. Water was
added, the aqueous layer was extracted with DCM (x3), and the
combined organic layers were dried over Na2SO4, filtered and
= 7.1 Hz, 3H). LC/MS m/z: (M + H)+ calcd for C11H18IO3,
325.03; found 325.0.
Ethyl
(3aSR,6aSR)-6a-hydroxyhexahydropentalene-
3a(1H)-carboxylate (29). To a solution of SmI2 (0.1 M in THF)
(154 mL, 15.4 mmol) was added HMPA (12.9 mL, 74.0 mmol),
and the reaction mixture was stirred at rt for 15 min during which
the mixture went from dark blue to deep purple. The mixture was
cooled to 0 °C and a solution of 28 (1g, 3.1 mmol) in THF (103
mL) was added dropwise over 2 h. When the addition was
complete, the mixture was quenched by the addition of saturated
aqueous sodium bicarbonate and extracted with EtOAc (x3). The
combined organic extracts were washed with brine, dried over
MgSO4, filtered and concentrated. The crude product was
purified by silica gel chromatography eluting with 0-50%
concentrated.
chromatography eluting with 30-80% EtOAc/hexanes to give 23
The residue was purified by silica gel
1
EtOAc/hexanes to give 29 (496 mg, 77%). H NMR (500 MHz,
1
(2.5 g, 59%). H NMR (300 MHz, CDCl3) δ 7.3 (m, 10H), 5.17
CDCl3) δ 4.19 (q, J = 7.2 Hz, 2H), 2.86 (s, 1H), 2.41 - 2.31 (m,
2H), 1.90 - 1.74 (m, 6H), 1.65 (s, 1H), 1.63 - 1.52 (m, 4H), 1.30
(t, J = 7.2 Hz, 3H). LC/MS m/z: (M + H)+ calcd for C11H19O3,
199.13; found 199.0.
(apparent br t, 1H), 4.48 (4H, s), 3.86 (t, J = 7.5 Hz, 1H), 2.9-3.7
(m, 8H), 1.36 (s, 9H), 1.1-1.6 (m, 4H). LC/MS m/z: (M + H)+
calcd for C27H40NO5, 458.29; found 458.3.
tert-Butyl
(5-(benzyloxy)-2-(2-(benzyloxy)ethyl)-2-
(3aSR,6aSR)-6a-Hydroxyhexahydropentalene-3a(1H)-
carboxamide (30). To a mixture of 29 (50 mg, 0.25 mmol) in
THF (0.84 mL) was added LiNH2 (41 mg, 1.77 mmol), and the
reaction mixture was heated under reflux for 1 h. The mixture
was diluted with EtOAc and washed with water and then brine.
The organic layer was dried over MgSO4, filtered and
formylpentyl)carbamate (24). A solution of 23 (9 g, 19.67
mmol) in DCM (130 mL) was added Dess-Martin periodinane
(10.9 g, 25.6 mmol) and the reaction mixture was stirred at rt at
for 2 h. The reaction mixture was filtered and washed with
EtOAc. The filtrate was washed with water, dried over Na2SO4,
and concentrated. The crude product was purified by silica gel
chromatography eluting with 10-30 EtOAc/hexanes to give 24
(7.5 g, 83%) as a white solid. 1H NMR (300 MHz, CDCl3) δ 9.50
(s, 1H), 7.3 (m, 10H), 5.02 (apparent br t, 1H), 4.48 (s, 2H), 4.44
(s, 2H), 3.3-3.6 (m, 6H), 1.5-1.9 (m, 6H), and 1.36 (s, 9H).
LC/MS m/z: (M + H)+ calcd for C27H38NO5, 456.27; found 456.5.
1
concentrated to give 30 (20 mg, 45%). H NMR (500 MHz,
CDCl3) δ 5.81 (br s, 1H), 5.48 (br s, 1H), 4.00 (s, 1H), 2.18 (dt, J
= 12.4, 7.7 Hz, 2H), 1.99 - 1.78 (m, 6H), 1.73 - 1.57 (m, 6H).
LC/MS m/z: (M + H)+ calcd for C9H16NO2, 170.12; found 170.0.
(3aSR,6aRS)-6a-(Aminomethyl)hexahydropentalen-
3a(1H)-ol (31). To a mixture of 6a-hydroxyoctahydropentalene-
3a-carboxamide (151 mg, 0.89 mmol) in THF (9 mL) was added
LiAlH4 (102 mg, 2.7 mmol) slowly (gas evolution). When the
addition was complete and gas evolution ceased, the mixture was
heated under reflux for 1 h. The mixture was cooled down to rt,
and sodium sulfate decahydrate (200 mg) was added. The
resulting mixture was stirred at rt for 12 h and then filtered
through a pad of Celite. The filtrate was concentrated to give 31
tert-Butyl (((3aSR,7aRS)-hexahydro-2H-furo[2,3-b]pyran-
3a-yl)methyl)carbamate (26). A suspension of 24 (5 g, 11
mmol) and 20% palladium hydroxide on carbon (0.5 g) in
methanol (50 mL) was hydrogenated with a hydrogen baloon for
3 h. The reaction mixture was filtered through a pad of Celite and
the filtrate was concentrated. The crude product was purified by
silica gel chromatography eluting with 10-40% EtOAC/hexanes
1
to give 26 (2.1 g, 75%) as a white solid. H NMR (400 MHz,
1
(145 mg, 99%). H NMR (500 MHz, CDCl3) δ 2.86 (s, 2H), 1.75
CDCl3) δ 4.75 (br s, 1H), 4.1 (m,1H), 4.0 (m, 1H), 3.9 (m, 1H),
3.4 (m, 1H), 3.1 (m, 2H), 2.0 (m, 1H), 1.5-1.9 (m, 5H), 1.47 (s,
9H). 99315-471. 13H NMR (100 MHz, CDCl3) δ 156.4, 103.8,
79.3, 67.0, 63.7, 46.9, 45.4, 29.4, 28.3, 27.2, and 20.9.
(td, J = 6.7, 2.3 Hz, 4H), 1.70 - 1.59 (m, 4H), 1.55 - 1.46 (m,
2H), 1.43 - 1.35 (m, 2H). 13C NMR (126 MHz, CDCl3) δ 90.6,
52.3, 47.9, 43.0, 38.1, 23.0.
Compounds 32-71 were prepared using the procedures
described for 4.
((3aSR,7aRS)-Tetrahydro-4H-furo[2,3-b]pyran-3a(7aH)-
yl)methanamine (27). To a solution of 26 (200 mg, 0.78 mmol)
in DCM (7.8 mL) at rt was added TFA (0.60 mL, 7.77 mmol),
and the reaction mixture was stirred at rt for 2 h. DCM was
removed to give 27 TFA salt (150 mg, 100 % yield) as a yellow
oil. This materil was used directly for the next reaction. 1H NMR
(400 MHz, CD3OD) δ 4.80 (s, 1H), 4.18 - 4.09 (m, 1H), 4.03 (td,
J = 8.5, 6.8 Hz, 1H), 3.94 - 3.86 (m, 1H), 3.54 - 3.44 (m, 1H),
3.03 - 2.89 (m, 2H), 2.22 - 2.08 (m, 1H), 1.94 - 1.66 (m, 4H),
1.63 - 1.52 (m, 1H).
5-Chloro-2-fluoro-4-((((1S,3R,4s,5S,7S)-4-
hydroxyadamantan-1-yl)methyl)amino)-N-(thiazol-2-
1
yl)benzenesulfonamide (32). H NMR (500 MHz, DMSO-d6) δ
7.56 (d, J = 7.0 Hz, 1H), 7.21 (d, J = 4.8 Hz, 1H), 6.77 (d, J =
4.4 Hz, 1H), 6.72 (d, J = 13.2 Hz, 1H), 5.87 (br t, J = 6.2 Hz,
1H), 3.58 (br s, 1H), 2.90 (d, J = 6.6 Hz, 2H), 1.89 - 1.77 (m,
6H), 1.66 - 1.54 (m, 4H), 1.42 (br s, 2H), 1.16 (br d, J = 11.4 Hz,
2H). LC/MS m/z: (M + H)+ calcd for C20H24ClFN3O3S2, 472.09;
found 472.0.
Ethyl 1-(3-iodopropyl)-2-oxocyclopentanecarboxylate (28).
To a mixture of 1,3-diiodopropane (4.4 mL, 38.4 mmol) and
ethyl 2-oxocyclopentanecarboxylate (0.95 ml, 6.4 mmol) in
acetone (16 mL) was added K2CO3 (2.3 g, 16.7 mmol), and the
reaction mixture was heated under reflux for 3 days. The mixture
was filtered, and the filtrate was then concentrated. The residue
was purified by silica gel chromatography eluting with 0-50%
5-Chloro-2-fluoro-4-(((1R,3S,5R,7)-3-hydroxyadamantan-
1-yl)amino)-N-(thiazol-2-yl)benzenesulfonamide (33). 1H
NMR (500 MHz, DMSO-d6) δ 7.62 (d, J = 7.3 Hz, 1H), 7.26 (d,
J = 4.8 Hz, 1H), 6.88 (d, J = 13.2 Hz, 1H), 6.83 (d, J = 4.8 Hz,
1H), 3.38 (br s, 1H), 3.36 (br s, 1H), 2.21 (br s, 2H), 1.93 - 1.78
(m, 6H), 1.70 - 1.42 (m, 6H). LC/MS m/z: (M + H)+ calcd for
C19H24ClFN3O3S2, 458.08; found 458.0.
1
EtOAc/hexanes to give 28 (1.44 g, 62%). H NMR (500 MHz,
CDCl3) δ 4.25 - 4.10 (m, 2H), 3.16 (qt, J = 9.6, 6.8 Hz, 2H), 2.56
- 2.40 (m, 2H), 2.34 - 2.24 (m, 1H), 2.08 - 1.86 (m, 5H), 1.82 -
1.72 (m, 1H), 1.71 - 1.64 (m, 1H), 1.71 - 1.64 (m, 1H), 1.26 (t, J
( )-5-Chloro-2-fluoro-4-((tetrahydrofuran-3-yl)amino)-N-
(thiazol-2-yl)benzenesulfonamide (34). 1H NMR (500 MHz,

DMSO-d6) δ 7.65 - 7.55 (m, 1H), 7.27 (d, J = 4.8 Hz, 1H), 6.83
(d, J = 4.4 Hz, 1H), 6.72 (d, J = 12.8 Hz, 1H), 6.07 (br d, J = 6.2
Hz, 1H), 4.19 - 4.11 (m, 1H), 3.92 - 3.82 (m, 2H), 3.71 (td, J =
8.3, 5.9 Hz, 1H), 3.64 (dd, J = 9.0, 4.2 Hz, 1H), 2.23 (dd, J =
12.7, 6.8 Hz, 1H), 1.91 (br d, J = 5.9 Hz, 1H). LC/MS m/z: (M +
H)+ calcd for C13H14ClFN3O3S2, 378.01; found 377.9.
5-Chloro-2-fluoro-4-((tetrahydro-2H-pyran-4-yl)amino)-
1
N-(thiazol-2-yl)benzenesulfonamide (41). H NMR (400 MHz,
CDCl3) δ 13.0 (br s, 1H), 7.88 (d, J = 7.3 Hz, 1H), 7.19 (d, J =
4.5 Hz, 1H), 6.52 (d, J = 4.5 Hz, 1H), 6.35 (d, J = 12.3 Hz, 1H),
4.81 - 4.69 (m, 1H), 4.02 (dt, J = 11.9, 3.6 Hz, 2H), 3.59 - 3.44
(m, 3H), 2.02 (br d, J=11.0 Hz, 2H), 1.67 - 1.50 (m, 2H). LC/MS
m/z: (M + H)+ calcd for C14H16ClFN3O3S2, 392.03; found 391.9.
( )-4-(7-Oxabicyclo[2.2.1]heptan-2-ylamino)-5-chloro-2-
fluoro-N-(thiazol-2-yl)benzenesulfonamide (35). It is a 3:2
5-Chloro-2-fluoro-4-(((tetrahydro-2H-pyran-4-
1
mixture of endo/exo isomers. Endo isomer: H NMR (400 MHz,
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (42). 1H
NMR (500 MHz, DMSO-d6) δ 7.57 (d, J = 7.3 Hz, 1H), 7.26 (d,
J = 4.8 Hz, 1H), 6.82 (d, J = 4.8 Hz, 1H), 6.68 (d, J = 12.8 Hz,
1H), 6.42 (br s, 1H), 3.83 (br dd, J = 11.2, 3.1 Hz, 3H), 3.36 -
3.21 (m, 1H), 3.09 (t, J = 6.6 Hz, 2H), 1.87 - 1.79 (m, 1H), 1.58
(br d, J = 11.4 Hz, 2H), 1.23 - 1.14 (m, 2H). LC/MS m/z: (M +
H)+ calcd for C15H18ClFN3O3S2, 406.05; found 406.0.
CDCl3) δ 7.87 (d, J = 8.0 Hz, 1H), 7.19 (d, J = 4.0 Hz, 1H), 6.53
(d, J = 4.5 Hz, 1H), 6.26 (d, J = 12.3 Hz, 1H), 4.91 (br d, J = 6.5
Hz, 1H), 4.74 - 4.67 (m, 1H), 4.44 (d, J = 5.3 Hz, 1H), 3.57 -
3.45 (m, 1H), 2.10 (dd, J=12.8, 7.5 Hz, 1H), 1.94 - 1.51 (m, 3H),
1.50 - 1.46 (m, 1H), 1.16 (dd, J = 12.5, 4.3 Hz, 1H). Exo isomer:
1H NMR (400 MHz, CDCl3) δ 7.89 (d, J = 8.0 Hz, 1H), 7.20 (d, J
= 4.0 Hz, 1H), 6.53 (d, J = 4.5 Hz, 1H), 6.35 (d, J = 12.0 Hz,
1H), 4.96 (br d, J = 4.8 Hz, 1H), 4.74 - 4.68 (m, 1H), 3.81 (br dd,
J = 10.5, 5.0 Hz, 1H), 2.52 - 2.37 (m, 1H), 1.93 - 1.41 (m, 5H).
LC/MS m/z: (M + H)+ calcd for C15H16ClFN3O3S2, 404.03; found
404.0.
5-Chloro-4-(((2,6-dimethyltetrahydro-2H-pyran-4-
yl)methyl)amino)-2-fluoro-N-(thiazol-2-
1
yl)benzenesulfonamide (43). H NMR (500 MHz, DMSO-d6) δ
7.57 (dd, J = 7.3, 1.8 Hz, 1H), 7.24 (d, J = 4.6 Hz, 1H), 6.80 (d, J
= 4.5 Hz, 1H), 6.65 (dd, J = 13.1, 2.5 Hz, 1H), 3.72 – 3.61 (m,
1H), 3.06 (t, J = 6.6 Hz, 1H), 2.13 (s, 1H), 1.63 (d, J = 12.8 Hz,
1H), 1.51 (d, J = 13.7 Hz, 1H), 1.24 (td, J = 13.1, 5.3 Hz, 1H),
1.08 (d, J = 6.2 Hz, 3H), 1.03 (d, J = 6.1 Hz, 3H), 0.75 (q, J =
11.9 Hz, 1H). LC/MS m/z: (M + H)+ calcd for C17H22ClFN3O3S2,
434.08; found 434.08; 433.9 (M + H)+.
( )-5-Chloro-2-fluoro-4-(((tetrahydrofuran-3-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (36).
1H NMR (500 MHz, DMSO-d6) δ 7.58 (d, J = 7.3 Hz, 1H), 7.26
(d, J = 4.4 Hz, 1H), 6.83 (d, J = 4.8 Hz, 1H), 6.71 (d, J = 13.2
Hz, 1H), 6.50 (br s, 1H), 3.76 (td, J = 8.0, 5.7 Hz, 1H), 3.70 -
3.64 (m, 1H), 3.64 - 3.57 (m, 1H), 3.46 (dd, J = 8.6, 5.0 Hz, 1H),
3.17 (br t, J = 7.2 Hz, 2H), 2.58 - 2.53 (m, 1H), 2.58 - 2.53 (m,
1H), 2.58 - 2.53 (m, 1H), 1.94 (br dd, J = 12.3, 5.7 Hz, 1H), 1.59
(dd, J = 12.7, 5.3 Hz, 1H). LC/MS m/z: (M + H)+ calcd for
C14H16ClFN3O3S2, 392.03; found 392.0.
5-Chloro-2-fluoro-4-((2-(tetrahydro-2H-pyran-4-
yl)ethyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (44). 1H
NMR (500 MHz, DMSO-d6) δ 7.57 (d, J = 7.3 Hz, 1H), 7.26 (d,
J = 4.8 Hz, 1H), 6.83 (d, J = 4.8 Hz, 1H), 6.61 (d, J = 12.8 Hz,
1H), 6.31 (br. s., 1H), 3.82 (dd, J = 11.4, 3.3 Hz, 2H), 3.31 - 3.13
(m, 2H), 2.50 (2H, m), 1.66 - 1.45 (m, 5H), 1.17 (dd, J = 1 1.6,
3.9 Hz, 2H). LC/MS m/z: (M + H)+ calcd for C16H20ClFN3O3S2,
434.08; found 420.06; 420.0 (M + H)+.
( )-5-Chloro-2-fluoro-4-((2-(tetrahydrofuran-2-
yl)ethyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (37). 1H
NMR (500 MHz, DMSO-d6) δ 7.58 (d, J = 7.3 Hz, 1H), 7.26 (d,
J = 4.8 Hz, 1H), 6.82 (d, J = 4.8 Hz, 1H), 6.61 (d, J = 13.2 Hz,
1H), 6.41 (br t, J = 4.8 Hz, 1H), 3.86 - 3.74 (m, 2H), 3.67 - 3.55
(m, 1H), 3.31 - 3.14 (m, 1H), 2.05 - 1.93 (m, 1H), 1.89 - 1.64 (m,
5H), 1.52 - 1.38 (m, 1H). LC/MS m/z: (M + H)+ calcd for
C15H18ClFN3O3S2, 406.05; found 405.9.
( )-5-Chloro-2-fluoro-4-(((tetrahydro-2H-pyran-2-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (45).
1H NMR (500 MHz, DMSO-d6) δ 7.57 (d, J = 7.3 Hz, 1H), 7.24
(d, J = 4.4 Hz, 1H), 6.82 (d, J = 4.4 Hz, 1H), 6.69 (d, J = 12.8
Hz, 1H), 6.14 (t, J = 6.1 Hz, 1H), 3.87 (br d, J = 11.4 Hz, 1H),
3.49 - 3.30 (m, 1H), 3.27 - 3.14 (m, 2H), 1.76 (br s, 1H), 1.59 (br
d, J = 11.7 Hz, 1H), 1.50 - 1.39 (m, 3H), 1.26 - 1.16 (m, 1H).
LC/MS m/z: (M + H)+ calcd for C15H18ClFN3O3S2, 434.08; found
406.05; 406.0 (M + H)+.
( )-5-Chloro-2-fluoro-4-((3-(tetrahydrofuran-3-
yl)propyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (38). 1H
NMR (500 MHz, DMSO-d6) δ 7.57 (d, J = 7.3 Hz, 1H), 7.25 (d,
J = 4.8 Hz, 1H), 6.82 (d, J = 4.4 Hz, 1H), 6.63 (d, J = 13.2 Hz,
1H), 6.37 - 6.32 (m, 1H), 3.77 (t, J = 7.7 Hz, 1H), 3.70 (td, J =
8.3, 4.8 Hz, 1H), 3.60 (q, J = 7.7 Hz, 1H), 3.22 - 3.15 (m, 2H),
2.13 (dt, J = 14.8, 7.5 Hz, 1H), 2.00 - 1.93 (m, 1H), 1.60 - 1.48
(m, 2H), 1.45 - 1.32 (m, 3H). LC/MS m/z: (M + H)+ calcd for
C16H20ClFN3O3S2, 420.06; found 420.0.
( )-4-(((1,4-Dioxan-2-yl)methyl)amino)-5-chloro-2-fluoro-
1
N-(thiazol-2-yl)benzenesulfonamide (46). H NMR (500 MHz,
DMSO-d6) δ 7.60 (d, J = 7.0 Hz, 1H), 7.22 (d, J = 4.3 Hz, 1H),
6.81 (d, J = 4.6 Hz, 1H), 6.71 (d, J = 13.1 Hz, 1H), 6.08 (br. s.,
1H), 3.79 - 3.41 (m, 6H), 3.34 - 3.22 (m, 4H). LC/MS m/z: (M +
H)+ calcd for C14H16ClFN3O4S2, 434.08; found 408.03; 407.9 (M
+ H)+.
( )-5-Chloro-2-fluoro-4-(((tetrahydrofuran-2-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (39).
1H NMR (500 MHz, DMSO-d6) δ 7.58 (d, J = 7.3 Hz, 1H), 7.26
(d, J = 4.8 Hz, 1H), 6.83 (d, J = 4.8 Hz, 1H), 6.74 (d, J = 13.2
Hz, 1H), 6.19 (t, J = 5.5 Hz, 1H), 4.08 - 3.98 (m, 1H), 3.81 - 3.72
(m, 1H), 3.67 - 3.56 (m, 1H), 3.29 - 3.13 (m, 1H), 2.5 (1H, m),
2.01 - 1.74 (m, 3H), 1.67 - 1.53 (m, 1H). LC/MS m/z: (M + H)+
calcd for C14H16ClFN3O3S2, 392.03; found 391.9.
( )-5-Chloro-2-fluoro-4-((2-(tetrahydro-2H-pyran-2-
yl)ethyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (47). 1H
NMR (500 MHz, CDCl3) δ 13.10 (br s, 1H), 7.86 (d, J = 7.2 Hz,
1H), 7.19 (d, J = 4.4 Hz, 1H), 6.51 (d, J = 4.6 Hz, 1H), 6.31 (d, J
= 12.5 Hz, 1H), 5.82 (br s, 1H), 4.02 (dt, J = 11.3, 2.1 Hz, 1H),
3.48 - 3.38 (m, 2H), 3.31 (dq, J=12.3, 6.1 Hz, 1H), 3.25 - 3.16
(m, 1H), 1.90 - 1.76 (m, 3H), 1.65 - 1.47 (m, 4H), 1.45 - 1.34 (m,
1H). LC/MS m/z: (M + H)+ calcd for C16H20ClFN3O3S2, 420.06;
found 420.0 (M + H)+.
4-(((1,3-Dioxolan-2-yl)methyl)amino)-5-chloro-2-fluoro-N-
(thiazol-2-yl)benzenesulfonamide (40). 1H NMR (500 MHz,
DMSO-d6) δ 7.58 (d, J = 7.3 Hz, 1H), 7.26 (d, J = 4.8 Hz, 1H),
6.83 (d, J = 4.8 Hz, 1H), 6.78 (d, J = 12.8 Hz, 1H), 6.26 (br t, J
= 5.9 Hz, 1H), 4.97 (t, J = 3.9 Hz, 1H), 3.97 - 3.87 (m, 2H), 3.86
- 3.77 (m, 2H), 3.41 (dd, J = 6.1, 4.2 Hz, 1H). LC/MS m/z: (M +
H)+ calcd for C13H14ClFN3O4S2, 394.01; found 393.9.
( )-5-Chloro-2-fluoro-4-((tetrahydro-2H-pyran-3-
yl)amino)-N-(thiazol-2-yl)benzenesulfonamide (48). 1H NMR
(500 MHz, DMSO-d6) δ 7.60 (d, J = 7.0 Hz, 1H), 7.24 (d, J =
4.4 Hz, 1H), 6.86 - 6.70 (m, 2H), 5.69 (br d, J = 8.1 Hz, 1H),

3.77 (dd, J = 11.0, 2.6 Hz, 1H), 3.73 - 3.67 (m, 1H), 3.64 - 3.52
(m, 1H), 1.99 - 1.85 (m, 1H), 1.71 - 1.46 (m, 3H). LC/MS m/z:
(M + H)+ calcd for C14H16ClFN3O3S2, 392.03; found 391.9
NMR (500 MHz, DMSO-d6) δ 7.60 (d, J = 7.3 Hz, 1H), 7.26 (d,
J = 4.4 Hz, 1H), 6.89 - 6.75 (m, 2H), 6.40 (br t, J = 5.5 Hz, 1H),
4.44 (d, J = 5.9 Hz, 2H), 4.21 (d, J = 5.9 Hz, 2H), 2.51 (s, 2H),
1.27 (s, 3H). LC/MS m/z: (M + H)+ calcd for C14H16ClFN3O3S2,
392.03; found 392.0.
(R)-5-Chloro-2-fluoro-4-(((tetrahydro-2H-pyran-3-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (49). 1H
NMR (500 MHz, DMSO-d6) δ 7.57 (d, J = 7.0 Hz, 1H), 7.25 (d,
J = 4.8 Hz, 1H), 6.82 (d, J = 4.8 Hz, 1H), 6.66 (d, J = 12.8 Hz,
1H), 6.40 (br s, 1H), 3.77 - 3.67 (m, 2H), 3.18 - 3.02 (m, 3H),
1.84 (br s, 1H), 1.82 - 1.75 (m, 1H), 1.62 - 1.51 (m, 1H), 1.48 -
1.35 (m, 1H), 1.23 (br d, J = 9.9 Hz, 1H). LC/MS m/z: (M + H)+
calcd for C15H18ClFN3O3S2, 406.05; found 406.0.
4-(2-Oxaspiro[3.3]heptan-6-ylamino)-5-chloro-2-fluoro-N-
(thiazol-2-yl)benzenesulfonamide (57). 1H NMR (500 MHz,
CDCl3) δ 7.91 (d, J=7.3 Hz, 1H), 7.18 (d, J = 4.6 Hz, 1H), 6.59 -
6.49 (m, 2H), 4.14 (s, 1H), 3.85 (s, 2H), 3.45 (s, 2H), 1.94 - 1.86
(m, 2H), 1.58 (dd, J = 4.6, 1.8 Hz, 2H). LC/MS m/z: (M + H)+
calcd for C15H16ClFN3O3S2, 404.03; found 404.0.
(S)-5-Chloro-2-fluoro-4-(((tetrahydro-2H-pyran-3-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (50).
1H NMR and MS data are identical to those of 49.
5-Chloro-2-fluoro-4-(((3-oxocyclohexyl)methyl)amino)-N-
(thiazol-2-yl)benzenesulfonamide (58). 1H NMR (500 MHz,
DMSO-d6) δ 7.62 - 7.53 (m, 1H), 7.26 (d, J = 4.8 Hz, 1H), 6.82
(d, J = 4.8 Hz, 1H), 6.70 (d, J = 13.2 Hz, 1H), 6.47 (br t, J = 5.9
Hz, 1H), 3.28 - 3.03 (m, 2H), 2.34 - 2.25 (m, 2H), 2.21 - 2.03 (m,
3H), 2.00 - 1.95 (m, 1H), 1.88 - 1.78 (m, 1H), 1.54 (dt, J = 12.6,
3.8 Hz, 1H), 1.41 - 1.28 (m, 1H). LC/MS m/z: (M + H)+ calcd for
C16H18ClFN3O3S2, 418.05; found 418.0.
4-((((2SR,3RS)-2-(tert-Butyl)tetrahydro-2H-pyran-3-
yl)methyl)amino)-5-chloro-2-fluoro-N-(thiazol-2-
1
yl)benzenesulfonamide (51). H NMR (500 MHz, DMSO-d6) δ
7.58 (d, J = 7.3 Hz, 1H), 7.21 (d, J = 4.4 Hz, 1H), 6.76 (d, J =
4.4 Hz, 1H), 6.57 (d, J = 12.8 Hz, 1H), 6.18 (br d, J = 4.0 Hz,
1H), 3.85 (td, J = 7.1, 3.1 Hz, 1H), 3.18 - 3.06 (m, 1H), 2.78 (d, J
= 6.6 Hz, 1H), 2.00 - 1.91 (m, 1H), 1.73 - 1.62 (m, 1H), 1.58 -
1.44 (m, 2H), 1.42 - 1.30 (m, 1H), 0.91 (s, 9H). LC/MS m/z: (M
+ H)+ calcd for C19H26ClFN3O3S2, 462.11; found 462.0.
5-Chloro-2-fluoro-4-((((1SR,3SR)-3-
hydroxycyclohexyl)methyl)amino)-N-(thiazol-2-
1
yl)benzenesulfonamide (59). H NMR (500 MHz, DMSO-d6) δ
7.56 (d, J = 7.3 Hz, 1H), 7.17 (d, J = 4.4 Hz, 1H), 6.71 (d, J =
4.4 Hz, 1H), 6.54 (d, J = 13.2 Hz, 1H), 6.27 - 6.04 (m, 1H), 3.86
(br s, 1H), 3.14 - 2.93 (m, 2H), 2.07 - 1.96 (m, 1H), 1.66 - 1.48
(m, 4H), 1.45 - 1.30 (m, 2H), 1.22 - 1.11 (m, 1H), 1.01 - 0.89 (m,
1H). LC/MS m/z: (M + H)+ calcd for C16H20ClFN3O3S2, 420.06;
found 420.0.
4-((((2SR,3SR)-2-(tert-Butyl)tetrahydro-2H-pyran-3-
yl)methyl)amino)-5-chloro-2-fluoro-N-(thiazol-2-
yl)benzenesulfonamide (52). 1H NMR (500 MHz, DMSO-d6) δ
7.59 (d, J = 7.3 Hz, 1H), 7.26 (d, J = 4.4 Hz, 1H), 6.82 (d, J =
4.4 Hz, 1H), 6.48 (d, J = 12.8 Hz, 1H), 6.40 - 6.30 (m, 1H), 4.03
(br dd, J = 11.0, 4.4 Hz, 1H), 3.48 - 3.33 (m, 1H), 3.11 (d, J =
1.8 Hz, 1H), 2.25 - 2.13 (m, 1H), 1.88 - 1.69 (m, 2H), 1.59 - 1.48
(m, 1H), 1.33 - 1.19 (m, 1H), 0.94 (s, 9H). LC/MS m/z: (M + H)+
calcd for C19H26ClFN3O3S2, 462.11; found 462.0.
5-Chloro-2-fluoro-4-((((1RS,3SR)-3-
hydroxycyclohexyl)methyl)amino)-N-(thiazol-2-
yl)benzenesulfonamide (60). 1H NMR (400 MHz, CDCl3) δ 7.86
(d, J = 7.3 Hz, 1H), 7.17 (d, J = 4.5 Hz, 1H), 6.52 (d, J = 4.5 Hz,
1H), 6.30 (d, J = 12.3 Hz, 1H), 4.92 (br t, J = 5.0 Hz, 2H), 3.69 -
3.57 (m, 1H), 3.13 - 3.03 (m, 2H), 2.06 - 1.97 (m, 2H), 1.90 -
1.81 (m, 1H), 1.79 - 1.65 (m, 2H), 1.39 - 1.14 (m, 2H), 1.05 -
0.85 (m, 2H). LC/MS m/z: (M + H)+ calcd for C16H20ClFN3O3S2,
420.06; found 420.0.
5-Chloro-2-fluoro-4-((((2SR,3RS)-2-phenyltetrahydro-2H-
pyran-3-yl)methyl)amino)-N-(thiazol-2-
1
yl)benzenesulfonamide (53). H NMR (500 MHz, DMSO-d6) δ
7.49 (d, J = 7.3 Hz, 1H), 7.41 - 7.30 (m, 5H), 7.25 (d, J = 4.8 Hz,
1H), 6.81 (d, J = 4.8 Hz, 1H), 6.03 (br t, J = 5.3 Hz, 1H), 5.96 (d,
J = 13.2 Hz, 1H), 4.04 (d, J = 9.9 Hz, 1H), 3.98 - 3.89 (m, 1H),
3.53 - 3.41 (m, 1H), 3.02 - 2.92 (m, 1H), 2.73 - 2.64 (m, 1H),
1.98 (br d, J = 12.8 Hz, 1H), 1.94 - 1.83 (m, 1H), 1.63 (br d, J =
3.3 Hz, 2H), 1.42 - 1.26 (m, 1H). LC/MS m/z: (M + H)+ calcd for
C21H22ClFN3O3S2, 482.08; found 482.0.
4-((1,4-Dioxaspiro[4.5]decan-7-ylmethyl)amino)-5-chloro-
2-fluoro-N-(thiazol-2-yl)benzenesulfonamide (61). 1H NMR
(500 MHz, DMSO-d6) δ 7.61 - 7.53 (m, 1H), 7.25 (d, J = 4.8 Hz,
1H), 6.82 (d, J = 4.4 Hz, 1H), 6.64 (d, J = 13.2 Hz, 1H), 6.44 -
6.37 (m, 1H), 3.88 - 3.77 (m, 4H), 3.08 (dt, J = 12.6, 6.4 Hz,
2H), 1.90 - 1.77 (m, 1H), 1.74 - 1.58 (m, 5H), 1.43 - 1.29 (m,
2H), 1.17 (t, J = 12.5 Hz, 1H), 0.96 - 0.81 (m, 1H). LC/MS m/z:
(M + H)+ calcd for C18H22ClFN3O3S2, 462.07; found 462.0.
5-Chloro-2-fluoro-4-((((2RS,3RS)-2-phenyltetrahydro-2H-
pyran-3-yl)methyl)amino)-N-(thiazol-2-
1
yl)benzenesulfonamide (54). H NMR (500 MHz, DMSO-d6) δ
7.50 (d, J = 7.3 Hz, 1H), 7.36 (d, J = 4.4 Hz, 4H), 7.30 - 7.20 (m,
2H), 6.81 (d, J = 4.4 Hz, 1H), 6.16 (br t, J = 5.3 Hz, 1H), 5.81 (d,
J = 12.8 Hz, 1H), 4.68 (d, J = 2.2 Hz, 1H), 4.13 (br dd, J = 11.0,
3.7 Hz, 1H), 3.59 (s, 1H), 2.79 - 2.64 (m, 1H), 2.74 (s, 1H), 2.23
- 2.13 (m, 1H), 1.99 - 1.86 (m, 2H), 1.83 - 1.74 (m, 1H), 1.44 -
1.32 (m, 1H). LC/MS m/z: (M + H)+ calcd for C21H22ClFN3O3S2,
482.08; found 482.0.
5-Chloro-2-fluoro-4-(((4-oxocyclohexyl)methyl)amino)-N-
(thiazol-2-yl)benzenesulfonamide (62). 1H NMR (500 MHz,
DMSO-d6) δ 7.61 - 7.54 (m, 1H), 7.23 (d, J = 4.8 Hz, 1H), 6.79
(d, J = 4.4 Hz, 1H), 6.70 (d, J = 13.2 Hz, 1H), 6.47 (br t, J = 5.1
Hz, 1H), 3.16 (t, J = 6.6 Hz, 2H), 2.36 (td, J = 13.8, 6.2 Hz, 2H),
2.20 (br d, J = 14.3 Hz, 2H), 2.13 - 1.95 (m, 3H), 1.44 - 1.30 (m,
2H). LC/MS m/z: (M + H)+ calcd for C16H18ClFN3O3S2, 418.05;
found 418.0.
( )-5-Chloro-2-fluoro-4-((2-(tetrahydro-2H-pyran-3-
yl)ethyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (55). 1H
NMR (500 MHz, DMSO-d6) δ 7.57 (d, J = 7.3 Hz, 1H), 7.24 (d,
J = 4.8 Hz, 1H), 6.80 (d, J = 4.8 Hz, 1H), 6.60 (d, J = 12.8 Hz,
1H), 6.31 (br. s., 1H), 3.85 - 3.69 (m, 2H), 3.19 (d, J = 6.2 Hz,
1H), 3.00 (t, J = 10.3 Hz, 1H), 2.50 (2H, m), 1.84 (d, J = 12.5
Hz, 1H), 1.62 - 1.11 (m, 6H). LC/MS m/z: (M + H)+ calcd for
C16H20ClFN3O3S2, 420.06; found 420.0.
5-Chloro-2-fluoro-4-(((4-
(methoxyimino)cyclohexyl)methyl)amino)-N-(thiazol-2-
yl)benzenesulfonamide (63). H NMR (500 MHz, DMSO-d6) δ
7.65 - 7.52 (m, 1H), 7.25 (d, J = 4.4 Hz, 1H), 6.81 (d, J = 4.4 Hz,
1H), 6.67 (d, J = 13.2 Hz, 1H), 6.44 (br t, J = 5.5 Hz, 1H), 3.70
(s, 3H), 3.09 (br t, J = 6.2 Hz, 2H), 3.03 - 2.96 (m, 1H), 2.25 (br
d, J = 14.7 Hz, 1H), 2.05 (td, J = 13.4, 4.4 Hz, 1H), 1.91 - 1.72
(m, 5H), 1.17 - 0.98 (m, 2H). LC/MS m/z: (M + H)+ calcd for
C17H21ClFN4O3S2, 447.07; found 447.0.
1
5-Chloro-2-fluoro-4-(((3-methyloxetan-3-
yl)methyl)amino)-N-(thiazol-2-yl)benzenesulfonamide (56). 1H

4-(((4-((Benzyloxy)imino)cyclohexyl)methyl)amino)-5-
chloro-2-fluoro-N-(thiazol-2-yl)benzenesulfonamide (64). 1H
NMR (500 MHz, DMSO-d6) δ 7.61 - 7.54 (m, 1H), 7.39 - 7.28
(m, 5H), 7.25 (d, J = 4.4 Hz, 1H), 6.81 (d, J = 4.4 Hz, 1H), 6.67
(d, J = 13.2 Hz, 1H), 6.44 (br s, 1H), 4.98 (s, 2H), 3.09 (br t, J =
6.4 Hz, 3H), 2.26 (br d, J = 14.3 Hz, 1H), 2.06 (td, J = 13.4, 4.4
Hz, 1H), 1.95 - 1.70 (m, 5H), 1.17 - 0.93 (m, 2H). LC/MS m/z:
(M + H)+ calcd for C23H25ClFN4O3S2, 523.10; found 523.0.
1H), 2.90 - 2.81 (m, 2H), 2.78 - 2.70 (m, 1H), 2.30 - 2.21 (m,
1H), 1.90 - 1.79 (m, 2H). LC/MS m/z: (M + H)+ calcd for
C14H16ClFN3O4S2, 440.00; found 439.9.
6. Acknowledgments
We thank Mr. Rick Rampulla and members of the Department
of Discovery Synthesis of Biocon Bristol-Myers Squibb
Research Center (BBRC) for scaling up intermediates 11 and 26.
5-Chloro-2-fluoro-4-((((1RS,4RS)-4-
hydroxycyclohexyl)methyl)amino)-N-(thiazol-2-
1
yl)benzenesulfonamide (65). H NMR (500 MHz, DMSO-d6) δ
7.56 (d, J = 7.3 Hz, 1H), 7.23 (d, J = 4.8 Hz, 1H), 6.79 (d, J =
4.4 Hz, 1H), 6.60 (d, J = 12.8 Hz, 1H), 6.31 (br s, 1H), 3.74 (br s,
1H), 3.06 (t, J = 6.4 Hz, 2H), 1.65 - 1.53 (m, 3H), 1.43 - 1.34 (m,
7H). LC/MS m/z: (M + H)+ calcd for C16H20ClFN3O3S2, 420.06;
found 420.0.
7. References
1. Dib-Hajj, S. D.; Yang, Y.; Black, J. A.; Waxman, S. G. The
Nav1.7 sodium channel: From molecule to man. Nat. Rev.
Neurosci. 2013, 14, 49–62.
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Mutations in SCN9A, encoding a sodium channel alpha subunit,
in patients with primary erythermalgia. J. Med. Genet. 2004, 41,
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3. Cox, J. J.; Reimann, F.; Nicholas, A. K.; Thornton, G.; Roberts,
E.; Springell, K.; Karbani , G.; Jafri, H.; Mannan, J.; Raashid, Y.;
Al-Gazali, L.; Hamamy, H.; Valente, E.; Gorman, S.; Williams,
R.; McHale, D. P.; Wood, J. N.; Gribble, F. M.; Woods, C. G. A
SCN9A channelopathy causes congenital inability to experience
pain. Nature 2006, 444, 894–898.
5-Chloro-2-fluoro-4-((((1SR,4SR)-4-
hydroxycyclohexyl)methyl)amino)-N-(thiazol-2-
yl)benzenesulfonamide (66). 1H NMR (500 MHz, CDCl3) δ 7.87
(d, J = 7.2 Hz, 1H), 7.17 (d, J = 4.6 Hz, 1H), 6.52 (d, J = 4.6 Hz,
1H), 6.31 (d, J = 12.4 Hz, 1H), 4.94 - 4.84 (m, 1H), 3.66 - 3.55
(m, 1H), 3.03 (t, J = 6.3 Hz, 2H), 2.09 - 2.01 (m, 2H), 1.87 (br d,
J = 12.8 Hz, 2H), 1.65 - 1.50 (m, 1H), 1.35 - 1.24 (m, 2H), 1.16 -
0.97 (m, 2H). LC/MS m/z: (M + H)+ calcd for C16H20ClFN3O3S2,
420.06; found 420.0.
4. Ahmad, S.; Dahllund, L.; Eriksson, A. B.; Hellgren, D.; Karlsson,
U.; Lund, P.; Meijer, I. A.; Meury, L.; Mills, T.; Moody, A.;
Morinville, A.; Morten, J.; O’Donnell1, D.; Raynoschek, C.;
Salter, H.; Rouleau, G. A.; Krupp, J. J. A stop codon mutation in
SCN9A causes lack of pain sensation. Hum. Mol. Genet. 2007, 16,
2114–2121.
4-((1,4-Dioxaspiro[4.5]decan-8-ylmethyl)amino)-5-chloro-
2-fluoro-N-(thiazol-2-yl)benzenesulfonamide (67). 1H NMR
(500 MHz, DMSO-d6) δ 7.57 (d, J = 7.3 Hz, 1H), 7.25 (d, J =
4.8 Hz, 1H), 6.82 (d, J = 4.4 Hz, 1H), 6.64 (d, J = 12.8 Hz, 1H),
6.43 - 6.32 (m, 1H), 3.84 (s, 4H), 3.07 (br t, J = 6.2 Hz, 2H), 1.79
- 1.54 (m, 5H), 1.49 - 1.32 (m, 2H), 1.29 - 1.05 (m, 2H). LC/MS
m/z: (M + H)+ calcd for C18H22ClFN3O4S2, 462.07; found 462.0.
5. Goldberg Y. P.; MacFarlane J.; MacDonald M. L.; Thompson J.;
Dube M.; Mattice M.; Fraser R.; Young C.; Hossain S.; Pape T.;
Payne B.; Radomski C.; Donaldson G.; Ives E.; Cox J.;
Younghusband H. B.; Green R.; Duff A.; Boltshauser E.; Grinspan
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NMR (500 MHz, DMSO-d6) δ 7.57 (d, J = 7.3 Hz, 1H), 7.26 (d,
J = 4.4 Hz, 1H), 6.82 (d, J = 4.8 Hz, 1H), 6.59 (d, J = 12.8 Hz,
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1.59 (m, 4H), 1.52 - 1.33 (m, 5H), 1.27 - 1.07 (m, 2H). LC/MS
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(70). H NMR (500 MHz, DMSO-d6) δ 7.58 (d, J = 7.0 Hz, 1H),
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