Discovery of Potent, Selective, and State-Dependent NaV1.7 Inhibitors with Robust Oral Efficacy in Pain Models: Structure-Activity Relationship and Optimization of Chroman and Indane Aryl Sulfonamides

Article abstract of DOI:10.1021/acs.jmedchem.0c00361

Voltage-gated sodium channel NaV1.7 is a genetically validated target for pain. Identification of NaV1.7 inhibitors with all of the desired properties to develop as an oral therapeutic for pain has been a major challenge. Herein, we report systematic structure-activity relationship (SAR) studies carried out to identify novel sulfonamide derivatives as potent, selective, and state-dependent NaV1.7 inhibitors for pain. Scaffold hopping from benzoxazine to chroman and indane bicyclic system followed by thiazole replacement on sulfonamide led to identification of lead molecules with significant improvement in solubility, selectivity over NaV1.5, and CYP2C9 inhibition. The lead molecules 13, 29, 32, 43, and 51 showed a favorable pharmacokinetics (PK) profile across different species and robust efficacy in veratridine and formalin-induced inflammatory pain models in mice. Compound 51 also showed significant effects on the CCI-induced neuropathic pain model. The profile of 51 indicated that it has the potential for further evaluation as a therapeutic for pain.

Full text of DOI:10.1021/acs.jmedchem.0c00361

Subscriber access provided by BIU Pharmacie | Faculté de Pharmacie, Université Paris V
Article
Discovery of Potent, Selective and State-Dependent NaV1.7 Inhibitors
with Robust Oral Efficacy in Pain Models: Structure-Activity Relationship
and Optimization of Chroman and Indane Aryl Sulfonamides
Vidya Ramdas, Rashmi Talwar, Vijay Kanoje, Rajesh Loriya, Moloy Banerjee, pradeep patil, Advait
Arun Joshi, laxmikant datrange, Amit Kumar Das, Deepak Sahebrao Walke, Vaibhav Kalhapure, Talha
Khan, Ganesh Gote, Usha Dhayagude, Shreyas Deshpande, Javed Shaikh, Ganesh Chaure, Ravindra
Pal, Santosh Parkale, Sachin Suravase, Smita Bhoskar, rajesh gupta, anil kalia, rajesh yeshodharan,
mahammad azhar, jagadeesh daler, vinod mali, geetika sharma, amitesh kishore, rupali vyawahare,
Gautam Agarwal, himani pareek, sagar budhe, arun nayak, dnyaneshwar warude, praveen gupta, parag
joshi, sneha joshi, sagar darekar, dilip pandey, akshaya wagh, Prashant Nigade, Maneesh Mehta, Vinod
Patil, Dipak Modi, shashikant pawar, mahip verma, Minakshi Singh, sudipto das, Jayasagar Gundu, kumar
nemmani, mark bock, Sharad Sharma, Dhananjay Bakhle, Rajender Kumar Kamboj, and Venkata P. Palle
J. Med. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jmedchem.0c00361 • Publication Date (Web): 05 May 2020
Downloaded from pubs.acs.org on May 5, 2020
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a service to the research community to expedite the dissemination
of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in
full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully
peer reviewed, but should not be considered the official version of record. They are citable by the
Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore,
the “Just Accepted” Web site may not include all articles that will be published in the journal. After
a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web
site and published as an ASAP article. Note that technical editing may introduce minor changes
to the manuscript text and/or graphics which could affect content, and all legal disclaimers and
ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or
consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W.,
Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the course
of their duties.

Page 1 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
pandey, dilip; Lupin Ltd, Novel Drug Discovery & Development
wagh, akshaya; Lupin Ltd, Novel Drug Discovery & Development
Nigade, Prashant; Lupin Ltd, Novel Drug Discovery & Development
Mehta, Maneesh; Lupin Ltd, Novel Drug Discovery & Development
Patil, Vinod; Lupin Ltd, Novel Drug Discovery & Development
Modi, Dipak; Lupin Ltd, Novel Drug Discovery & Development
pawar, shashikant; Lupin Ltd, Novel Drug Discovery & Development
verma, mahip; Lupin Ltd, Novel Drug Discovery & Development
Singh, Minakshi; Lupin Ltd, Novel Drug Discovery & Development
das, sudipto; Lupin Ltd, Novel Drug Discovery & Development
Gundu, Jayasagar; Lupin Ltd, Novel Drug Discovery & Development
nemmani, kumar; Lupin Ltd, Novel Drug Discovery & Development
bock, mark; Lupin Ltd, Novel Drug Discovery & Development
Sharma, Sharad; Lupin Ltd, Novel Drug Discovery & Development
Bakhle, Dhananjay; Lupin Ltd, Novel Drug Discovery & Development
Kamboj, Rajender; Lupin Ltd, Novel Drug Discovery & Development
Palle, Venkata P.; Lupin Ltd, Novel Drug Discovery & Development
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 2 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
42x48mm (300 x 300 DPI)
ACS Paragon Plus Environment

Page 3 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
Discovery of Potent, Selective and State-Dependent
Na 1.7 Inhibitors with Robust Oral Efficacy in Pain
V
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Models: Structure-Activity Relationship and
Optimization of Chroman and Indane Aryl Sulfonamides
Vidya Ramdas,* Rashmi Talwar,Vijay Kanoje,Rajesh M. Loriya, Moloy Banerjee,Pradeep Patil, Advait Arun
Joshi, Laxmikant Datrange, Amit Kumar Das, Deepak Sahebrao Walke, Vaibhav Kalhapure, Talha Khan,
Ganesh Gote, Usha Dhayagude, Shreyas Deshpande, Javed Shaikh, Ganesh Chaure, Ravindra R. Pal,
Santosh Parkale, Sachin Suravase, Smita Bhoskar, Rajesh V. Gupta, Anil Kalia, Rajesh Yeshodharan,
Mahammad Azhar, Jagadeesh Daler, Vinod Mali, Geetika Sharma, Amitesh Kishore, Rupali Vyawahare,
Gautam Agarwal, Himani Pareek, Sagar Budhe, Arun Nayak,ꝉ Dnyaneshwar Warude, Praveen Kumar
Gupta, Parag Joshi, Sneha Joshi, Sagar Darekar, Dilip Pandey, Akshaya Wagh, Prashant B. Nigade, Maneesh
Mehta, Vinod Patil, Dipak Modi, Shashikant Pawar, Mahip Verma, Minakshi Singh, Sudipto Das, Jayasagar
Gundu, Kumar Nemmani, Mark Bock, Sharad Sharma, Dhananjay Bakhle, Rajender Kumar Kamboj,
Venkata P. Palle
Novel Drug Discovery & Development, Lupin Ltd., Lupin Research Park, Survey No. 46 A /47 A, Village Nande, Taluka
Mulshi, Pune 412115, India
ꝉDeceased, 11th March 2019
ABSTRACT
Voltage gated sodium channel Na
V
1.7 is a genetically validated target for pain. Identification of Na 1.7
V
inhibitors with all the desired properties to develop as an oral therapeutic for pain has been a major
challenge. Herein, we report systematic SAR studies carried out to identify novel sulfonamide derivatives
as potent, selective and state-dependent Na
to chroman and indane bicyclic system followed by thiazole replacement on sulfonamide led to
identification of lead molecules with significant improvement in solubility, selectivity over Na 1.5 and
V
1.7 inhibitors for pain. Scaffold hopping from benzoxazine
V
CYP2C9 inhibition. The lead molecules 13, 29, 32, 43 and 51 have shown favorable PK profile across
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 4 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
different species and robust efficacy in veratridine and formalin induced inflammatory pain models in
mice. Compound 51 has also shown significant effect in CCI induced neuropathic pain model. Profile of 51
has indicated that it has the potential for further evaluation as a therapeutic for pain.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
INTRODUCTION
Pain is a major health problem, greatly reduces quality of life and imparts huge health costs and economic
loss to society. One fourth of the world’s population suffers from chronic pain. Pain sensations originate
in peripheral neurons such as nociceptors. Increased pain sensations can result from altered electrical
excitability of neurons in both peripheral and central neurons. As a therapeutic class, sodium channel
blockers such as lidocaine, amitriptyline, and lamotrigine have been widely used to treat disorders where
the chosen therapeutic approach is designed to decrease neuronal excitability. Such disorders involving
hyper-excitability or increased neuronal sensitivity to stimulation include pain as well.1
Voltage‐gated sodium channels (VGSCs) are key elements of excitable cells responsible for the generation
2
and transmission of electrical activity in central and peripheral nervous system. VGSCs consist of a pore-
forming alpha subunit and a stabilizing beta subunit. Nine isoforms of the alpha subunit have been
3
identified till date (Na
V
1.1 to Na
V
1.9) and are encoded by SCN genes. All these nine members of the VGSC
family have high homology of amino acid sequence in the extracellular and transmembrane domain. The
sodium channels have also been further classified based on their sensitivity to the puffer fish toxin
(
tetrodotoxin, TTX). Channels Na
V
1.8, Na
V
1.9 and Na 1.5 are TTX resistant (TTX-R) whereas the remaining
V
channels are sensitive to TTX and classified as TTX sensitive (TTX-S). VGSCs mediate the influx of sodium
ions into the cell in response to membrane depolarization leading to generation of the action potential to
relay information of stimuli from periphery to the central nervous system. Thus, voltage-gated ion
channels play a major role in determining excitability of neurons.2,3
ACS Paragon Plus Environment

Page 5 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
Na
V
1.7 is one of the highly expressed TTX-sensitive sodium channels in dorsal root ganglion cells and is
believed to be an important contributor to signaling in both nociceptive and non-nociceptive neurons.
There is a strong genetic evidence linking Na 1.7 and the pain processing pathway and the evidence comes
mostly in the form of mutations in human SCN9A, the gene encoding Na 1.7. The gain-of-function
V
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
V
1.7 causes spontaneous pain syndromes such as inherited erythromelalgia,4
mutations in human Na
V
5
6
paroxysmal extreme pain disorder, and small fiber neuropathy. In contrast, the loss-of-function
mutations in human Na 1.7 lead to a rare condition known as congenital insensitivity to pain (CIP), which
manifests in the form of inability of individuals to sense pain while not significantly impacting motor or
V
7
8
cognitive function. Patients with CIP experience no pain following a wide variety of stimuli.
Similarly, deletion of Na
V
1.7 in sensory and sympathetic neurons of mice leads to a pain-free congenital
1.7-null
insensitivity to pain (CIP) phenotype similar to that described in humans. Human and mouse Na
V
mutants are apparently normal, suggesting that this channel is an excellent analgesic drug target for acute,
9
inflammatory and neuropathic pain. Non-selective Na
V
blockers such as mexiletine and carbamazepine
have long been used clinically for the treatment of neuropathic pain. Limitations with these therapies are
marginal response and adverse effects seen in patients and these are due to marginal potency and non-
selectivity against other sodium channels or resting state blockade of channels.10 The ion channel moves
from closed state to open through concerted movements and then to an inactivated state where the
channel stops to conduct ions. Disease states are characterized by activated state of high frequency firing
channels, however the normal somatosensory system is characterized by low frequency firing channels.
Hence identification of Na
increased affinity for a slow-inactivated channel state vs. fast-inactivated may lead to functional selectivity
and exhibit a better safety window. Thus, a potent, selective and state-dependent Na 1.7 blocker
V
1.7 inhibitors that exhibit a different affinity for each of these states and show
V
rationalize as an effective and safe therapeutic option for treatment of chronic pain in humans.11 One of
the primary challenges associated with these efforts has been the identification of isoform-selective
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 6 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
inhibitors, in particular those that demonstrate suitable levels of selectivity over Na
expressed in cardiac myocytes and plays a key role in cardiovascular function.12
V
1.5, which is
Many aryl and acyl sulfonamide derivatives have been reported as potent and selective Na
inhibitors.13 They have been shown to bind to voltage sensing domain4 of Na
by locking it into the inactivated state.14 However, identification of a selective Na
V
1.7
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
V
1.7 and block the channel
V
1.7 inhibitor which
shows robust efficacy in rodent pain models without any metabolic liability to develop as an oral
therapeutic for pain has been a major challenge for long time since the SAR on sulfonamide derivatives
were found to be non-additive for multi parameter optimization.15 Most of the initial lead molecules
reported by Pfizer,16 Amgen,17 Merck,18 Xenon19 and BMS20 were containing either 1,2,4-thiadiazole or
thiazole substituted arylsulfonamide analogs, A-F, Fig. 1. Most of these compounds suffer from high
CYP3A4 and/or CYP2C9 inhibition.16-21 In addition, many of these sulfonamide derivatives lack good
pharmacokinetic profile across rodent and non-rodent species suitable for development as an oral
therapeutic for pain.13,15 From our own internal SAR studies and also from the literature reports which
appeared while we were working on the project, it was clear that the CYP3A4 inhibition could overcome
by replacement of 1,2,4-thiadiazole with other heteroaryl substitution on sulfonamide, compounds A, F,
&
G-I, Fig. 1.16b,17d,19a,b,20b However, CYP2C9 inhibition was still a challenge and in addition heteroaryl
modification on sulfonamide led to new challenges such as decrease in potency and selectivity although
the extent of compromise on potency and selectivity was dependent on the overall scaffold.17,18b,19a,b
Herein, we report systematic SAR studies carried out to identify potent, selective and orally bioavailable
chroman and indane sulfonamide derivatives as state-dependent Na
CYP liabilities, Fig. 2. In addition, we report complete characterization of lead molecules 13, 29, 32, 43 and
1 including detailed pharmacokinetic properties across the species and in-vivo efficacy of selected lead
V
1.7 blockers without any significant
5
molecules in mouse models of veratridine induced pain, formalin induced inflammatory pain and CCI
induced neuropathic pain.
ACS Paragon Plus Environment

Page 7 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
V
Figure 1. Structures of Selected Aryl Sulfonamide Na 1.7 Inhibitors
RESULTS AND DISCUSSION
The objective was to identify a potent, selective and state dependent Na
safe oral therapeutic for chronic pain. To meet this objective we have set out a target product profile with
the criteria as Na 1.7 potency: IC50 ≤ 10 nM; Na 1.5 selectivity: IC50 >10 μM; No significant CYP inhibition
<50% at 10M); No significant hERG inhibition; State dependent block of target; Acceptable oral
V
1.7 blocker which will serve as a
V
V
(
bioavailability; Significant efficacy in veratridine, formalin induced nociception and/or CCI induced pain
model; and ED50 that allows a good therapeutic window. Initially SAR study was carried out on benzoxazine
core substituted with thiazole sulfonamide. Various analogs were prepared with different aryl, heteroaryl
and heterocyclyl groups as pendant substitution on terminal phenyl ring, Fig. 2. Most of these compounds
have shown very good potency (1-10 nM) and moderate selectivity over Na 1.5. However, these
V
compounds suffer from high CYP2C9 inhibition, poor solubility and poor pharmacokinetic profile in spite
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 8 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
of their good microsomal stability. Replacement of the thiazole group with different 5-6 membered
heteroaryl substitution on sulfonamide was not tolerated on this core.17e
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0

Very good potency
Moderate selectivity
High CYP2C9 inhibition
Poor solubility


Very good potency
Moderate selectivity
High CYP2C9 inhibition
Improved solubility

Good potency
Desired selectivity


Low CYP2C9 inhibition
Very good solubility

Poor Pharmacokinetics

Desired Pharmacokinetics

Desired Pharmacokinetics
Figure 2. Central Bicyclic Core Switch from Benzoxazine to Chroman for Multi-parameter Optimization
Hence, it was decided to alter the central bicyclic core. By considering multi-parameter optimization such
as physchem and ADME properties, it was decided to explore other bicyclic systems such as chroman as
central bicyclic core, since substituted chroman analogs are expected to possess an altered conformation
compared to the corresponding substituted benzoxazine analogs. We were interested to explore the role
3
of additional sp carbon on solubility and other ADME properties, the effect of conformational change on
potency and selectivity and CYP inhibition profile. Herein, we report the outcome of a systematic and
elaborate SAR study on multi parameter optimization of this series.
First to check the translation of potency and selectivity, a series of chroman compounds were prepared
by keeping the thiazole substitution constant on sulfonamide. A range of heteroaryl, heterocyclyl, aryl and
substituted alkyl groups were tried as pendant R group substitution on the terminal phenyl ring. Absence
of pendant R group or substitution such as –Cl, alkyl, or branched alkyl as pendant group led to very less
potent compounds. Alkoxy alkyl and fluoro alkyl of different chain length were found to be very potent
for Na
V
1.7, however less selective over Na 1.5 (data not shown). Various compounds (1-12) having
V
heteroaryl, heterocyclyl or substituted aryl as pendant R group were prepared and tested for their activity
ACS Paragon Plus Environment

Page 9 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
against Na
V
1.7 and Na
V
1.5. Since most of these compounds were prepared as racemic and then the
isomers were separated by chiral column chromatography at the final stage, the potency of both the
enantiomers were tested for many compounds with different R groups, and it was consistently found to
be one of the isomers was significantly more potent than the other isomer. As a representation, for some
of the R groups, activities of both the isomers were shown in Table 1. The absolute stereochemistry of the
compounds 1a and 1b were determined to be ‘S’ and ‘R’ configuration, respectively, by using vibrational
circular dichroism (VCD). Determination of absolute configuration of few additional pairs of isomers using
VCD has shown that the more potent isomer consistently possess alpha orientation (below plane in drawn
structures) of terminal phenyl group with respect to the bicyclic central core (chroman). This finding was
further confirmed without any ambiguity through a single crystal X-ray of sulfonamide intermediate and
its conversion to the lead compounds (refer supporting information for the structure and scheme).
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Most of the compounds have shown very good potency (IC50 <10nM) for Na
over Na 1.5 was not in the expected lines (IC50 >10M), Table 1. Only a few compounds with R groups
such as triazole (4b), morpholinone 8 and oxazolinone 9 have shown very good selectivity over Na 1.5.
Also, for the less active isomer, similar right ward shift of activity for Na 1.5 subtype was observed.
V
1.7, however, the selectivity
V
V
V
Table 1. Profile of Chroman Compounds Substituted with Thiazole Sulfonamide
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 10 of 66
CYP3A4
(% Inhibition)
CYP2C9
(% Inhibition)
Na
IC50
nM
V
1.7
Na 1.5
V
Compd
isomer)
Solubility
(M)
R
IC50
LogD
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
(
a
nMa,#,*
@1M @10M @1M @10M
1
a (S)
1b (R)
a (S)
2b (R)
a (S)
3b (R)
a (S)
4b (R)
a (S)
67.8
0.4
Inactive
812
2.2
2.2
3.3
3.3
2.9
2.9
2.3
2.3
NT
65
83
11
2
26
28
NT
64
42
38
NT
19
37
19
51
46
NT
88
54
45
NT
42
70
48
92
93
NT
99
91
88
NT
87
97
90
2
60.5
1.6
61%
7
NT
6
67%
15
3
850
2.3
79%
130
61
0
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
629
7
4
139
0.5
Inactive
Inactive
63%
154
171
NT
21
NT
5
5
15.4
0.4
10
6
5b (R)
154
3.2
6 (R)
4.5
362
3.5
7
15
33
38
86
7a (S)
222.6
0.5
69%
124
4.5
4.5
147
39
-3
5
17
35
68
62
94
94
7b (R)
8
(R)
(R)
7.8
Inactive
Inactive
1.8
2.1
695
295
0
6
0
1
32
60
9
35.5
12
19
10 (R)
11 (R)
12 (R)
6.2
0.8
2.9
50%
4089
983
2.5
1.4
2.1
99
-17
1
1
2
4
68
83
52
288
176
33
10
7
33
a
#
FLIPR membrane potential assay; Inactive refers to <10% inhibition at 10 M; * % Inhibition at 10 M; NT – Not Tested
Very good potency and the preference of one stereoisomer over the other for activity encouraged us to
explore this scaffold in depth for optimization of other properties. When we tested the solubility of these
3
compounds, as expected these compounds with chiral sp carbon have shown improved solubility over
the corresponding benzoxazine analogs. Then, we tested these compounds for inhibition profile of two
important CYP enzymes (3A4 and 2C9). Again, most of these thiazole compounds in spite of central core
modification with a wide range of R group variants have shown high CYP2C9 inhibition while CYP3A4
inhibition was not significant at 10M (except for compound 2b), Table 1. To our disappointment both
the isomers have shown similar CYP inhibition profile. The conformational difference between the
isomers did not lead to any difference in CYP inhibition profile. These trends were consistent throughout
ACS Paragon Plus Environment

Page 11 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
this series of compounds, where one of the isomers was found to be several fold more potent than the
other isomer, but both the isomers had similar CYP inhibition profile. Next we were interested in looking
at the pharmacokinetic profile of chroman based analogs as it was one of the challenges observed in
earlier benzoxazine series. A few best compounds were tested for their pharmacokinetic profile and to
our delight remarkable improvement in overall PK profile was observed for chroman analogs, Table 2.
Compound 4b has shown low t1/2 due to high clearance, however all the three compounds 1b, 4b and 5b
have shown very good exposures and oral bioavailability. Modification of the central bicyclic core from
benzoxazine to chroman helped in improving the solubility and pharmacokinetic profile. The SAR outcome
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
has clearly indicated that still we have two major challenges in hand, which are selectivity over Na 1.5 and
V
CYP2C9 inhibition. For optimizing these two liabilities, we have decided to modify the substitution on
sulfonamide. First, we have kept the pendant R group, N-methyl pyrazole as constant and tried various
substituted or unsubstituted five and six membered heteroaryls as we have attempted in the case of
benzoxazine core. The data for six membered heteroaryl sulfonamide derivatives, 13-20 is shown in Table
3
. To our fortune, compounds 13-15 & 18 having 2-aminopyrimidine, 4-F-2-aminopyrimidine, 4-
aminopyrimidine and 6-F-2-aminopyridine, respectively have shown good potency and very good
selectivity over Na 1.5. In addition, compounds 13 and 14 showed low levels of CYP2C9 inhibition at 10M
<40%), Table 3. The potency for Na 1.7 was on the lower side for pyridazine (16) and pyrazine (17)
V
(
V
compounds. Compounds 18-20 having F-pyridines as substitution on sulfonamide have shown better
selectivity over compound 1b, however, they
Table 2. Metabolic Rate and Pharmacokinetic Profile of Chroman Compounds in Male BALB/c Mice
Metabolic Rate
PK IV (1 mpk)
PK PO (10 mpk)
(
nmol/min/mg)
Compd
CL
Cmax
μM)
AUClast
T1/2
Vss
Cmax
(μM)
AUClast
MLM RLM HLM
(ml/min/
kg)
%F
(
(h*μM)
(h)
(l/kg)
(h*μM)
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 12 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
1
4
5
b
b
b
0.1
0.04 0.05
1.3
2.2
0.8
2.2
0.9
2.0
2.0
0.4
2.8
14
37
14
2.2
1.0
2.9
6
15
7.1
19
68
79
95
0.02 0.02 0.02
3.0
4.2
0.04 0.05 0.03
Vehicle: IV: DMA (10%): PEG 300 (20%): 20% Captisol in NS (QS); PO: Tween 80 (1%): 0.5% CMC solution (QS)
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
did not show improvement in CYP2C9 inhibition. Overall, this SAR outcome delineated a way forward to
overcome both selectivity and CYP2C9 inhibition challenges while retaining the potency for Na 1.7
V
through modification of substitution on aryl sulfonamide of chroman scaffold. Hence, it was decided to
prepare a focused library of compounds with the best pendant R groups and four different heteroaryl
substitution on sulfonamide. The substitutions were chosen based on Na 1.7 potency, selectivity and low
V
CYP inhibition profile. The resultant matrix of compounds 21-50 have shown that
Table 3. Profile of Chroman Compounds Substituted with Different Heteroaryl Sulfonamide
CYP3A4
% Inhibition)
CYP2C9
(% Inhibition)
Na 1.5
V
Na
IC50 nM
V
1.7
Solubility
(M)
(
Compd
R
IC50
LogD
a
nMa,#,*
@
1M
@10M
@1M @10M
1b
13
14
0.4
812
2.2
3.7
2.5
83
2
28
46
7
93
36
28
17.8
24.4
Inactive
Inactive
532
463
6
3
15
0
13
1
5
6
4.3
Inactive
34%
1.8
2.2
1.8
1180
915
0
3
37
8
10
19
32
69
56
74
1
40.2
86.3
17
Inactive
1174
19
68
18
19
20
0.6
20
Inactive
46%
3.6
3.8
3.2
27
29
46
5
13
4
21
54
23
33
31
33
86
87
81
6.5
60%
ACS Paragon Plus Environment

Page 13 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
a
#
FLIPR membrane potential assay; Inactive refers to <10% inhibition at 10 M; *34% inhibition at 10M
the SAR is not additive. The tolerability of sulfonamide substitution varies with respect to the pendant R
group. In the entire matrix, most of the compounds having heteroaryl group as pendant group were found
to be potent and selective except for compounds 21 (2-methyl oxazole is pendant R group) and 25-27
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(triazole is pendant R group) with a strikingly low potency for Na
V
1.7, Table 4. Compounds with p-F phenyl
3
9-42 have shown significant decrease in potency. Except for morpholine, compounds 43-46, many
saturated heterocyclic groups such as morpholinone (data not shown), oxazolinone (data not shown) and
thiamorpholine (47-50) were not tolerated as pendant R groups for modification of thiazole on
sulfonamide, significant decrease in potency was observed like triazole compounds. Selected potent
compounds were tested for their solubility and CYP inhibition profile. All the compounds except for 6-
fluoropyridyl sulfonamide derivatives have shown very good solubility, Table 5. Most of the compounds
with 6-fluoropyridyl and 4-aminopyrimidinyl substitution on the sulfonamide have shown high CYP2C9
inhibition at least at 10M, Table 5. Poor solubility and high CYP inhibition of most of the 6-fluoropyridyl
sulfonamide compounds indicates that 6-fluorpyridyl behaves the same way as thiazole on sulfonamide.
Compounds with 4-F-2-aminopyrimidine substitution on the sulfonamide have shown moderate to high
CYP2C9 inhibition at 10M depending on the pendant R group. However, 2-aminopyrimidinyl sulfonamide
compounds have not shown significant CYP2C9 inhibition even at 10M, except for compound 43, where
it was found to be moderate, 54% inhibition at 10M, Table 5.
Table 4. Potency & Selectivity of Focused Library of Chroman Sulfonamide Compounds
R’
Parameter
R
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 14 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Compd
1.7 IC50 nM
13
14
15
18
Na
V
17.8
24.4
4.3
0.6
#
,
Na
V
1.5 IC nM *
Inactive
21
47.3
Inactive
22
25.9
Inactive
23
1.8
Inactive
24
1.6
50
Compd
1.7 IC50 nM
Na
V
#
,
Na
V
1.5 IC nM *
Inactive
25
698
Inactive
26
601
1168
27
54
44%
28
1.1
50
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Compd
1.7 IC50 nM
#,
Na
V
Na
V
1.5 IC nM *
Inactive
29
Inactive
30
2.8
Inactive
31
0.11
Inactive
50
Compd
1.7 IC50 nM
Na
V
21
#
,
Na
V
1.5 IC nM *
58%
32
1.4
Inactive
33
8.3
Inactive
34
8.7
50
Compd
1.7 IC50 nM
35
6.6
Na
V
#
,
Na
V
1.5 IC nM *
Inactive
Inactive
37
2.1
1103
38
12
1312
50
Compd
1.7 IC50 nM
#,
36
Na
V
2
.7
Na
V
1.5 IC nM *
56%
Inactive
40
162
2516
41
808
50
Compd
1.7 IC50 nM
39
30.4
42
77
Na
V
#
,
Na
V
1.5 IC nM *
1069
43
7.2
Inactive
44
11.5
50%
45
3
64%
46
2.3
50
Compd
1.7 IC50 nM
Na
V
#
,
Na
V
1.5 IC nM *
34%
47
Inactive
48
248
49
14.5
188
50
173
50
Compd
1.7 IC50 nM
#,
Na
V
164
83
Na 1.5 IC nM *
V
Inactive
Inactive
Inactive
Inactive
50
#
Inactive refers to <10% inhibition at 10 M; *% Inhibition at 10 M
Table 5. Solubility & CYP Profile of Focused Library of Chroman Sulfonamide Compounds
R’
Parameter
R
Compd
LogD
Solubility (M)
13
3.7
532
14
2.5
463
15
1.8
1180
18
3.6
27
ACS Paragon Plus Environment

Page 15 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
%
%
CYP3A4 inhbn
1/ 10 µM
CYP2C9 inhbn
6
/15
/36
3/-1
-2/37
10/69
5/21
@
7
13/28
33/86
@
1/ 10 µM
Compd
LogD
21
2.8
245
22
2.8
281
23
NT
NT
24
4
13
Solubility (M)
CYP3A4 inhbn
1/ 10 µM
CYP2C9 inhbn
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
%
-3/12
-5/3
43/86
86/98
1/44
@
%
2/24
4/41
27/88
@
1/ 10 µM
Compd
LogD
29
2.8
408
30
2.9
198
31
2.3
742
Solubility (M)
% CYP3A4 inhbn
@ 1/ 10 µM
% CYP2C9 inhbn
@ 1/ 10 µM
Compd
11/14
3/48
8/38
24/67
90/99
0/18
32
2.3
280
33
2.5
225
34
3
181
35
3.7
7
LogD
Solubility (M)
%
CYP3A4 inhbn
1/ 10 µM
CYP2C9 inhbn
0
/8
-3/7
47/87
90/99
10/13
22/85
@
%
-1/28
2/68
@
1/ 10 µM
Compd
LogD
36
2.9
225
37
3
233
38
3
181
Solubility (M)
CYP3A4 inhbn
1/ 10 µM
%
-2/29
-1/28
6/49
-4/11
0/44
@
%
CYP2C9 inhbn
@ 1/ 10 µM
Compd
4/33
43
2.7
91
44
2.8
184
45
2
1174
46
4.1
31
LogD
Solubility (M)
CYP3A4 inhbn
1/ 10 µM
CYP2C9 inhbn
1/ 10 µM
%
1
1/19
3/54
3/32
13/64
74/97
3/27
@
%
1
83/98
58/95
@
While gaining the knowledge from SAR studies on chroman series, the best combination of pendant
groups and heteroaryl sulfonamide were also tried simultaneously on indane core to understand the
effect of terminal phenyl ring trajectory on Na 1.7 inhibition. Out of which, selected examples are shown
V
in Table 6. We have observed good translation of SAR from chroman series to indane, except for
compound 54 wherein morpholine was a pendant R group.
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 16 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Table 6. Profile of Indane Compounds Substituted with Pyrimidine Sulfonamide
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
CYP3A4
% Inhibition)
CYP2C9
(% Inhibition)
Na
IC50
nM
V
1.7
Na 1.5
V
Solubility
M)
(
Compd
R
IC50
LogD
(
a
nMa,#,*
@1M @10M @1M @10M
51
52
53
54
10.3
6.9
60%
75%
2.4
3.2
2.8
2.9
316
40
5
6
1
1
11
11
11
3
3
5
8
7
-8
41
47
43
35.6
105
Inactive
2792
47
12
a
#
FLIPR membrane potential assay; Inactive refers to <10% inhibition at 10 M; * % Inhibition at 10 M
The best compounds with respect to potency for Na
significant CYP inhibition at 10M were chosen for assessing the state dependent blockage of Na
Na 1.5 and detailed in-vitro ADMET profile. Data shown for selected compounds in Table 7 & 8. In manual
patch-clamp electrophysiology assay, all the tested compounds have shown state-dependent inhibition
of Na 1.7, however they did not show any use-dependent effect, which is true with known sulfonamide
V V
1.7, selectivity over Na 1.5, good solubility and no
V
1.7,
V
V
derivatives.17a Compound 29 has shown resting state block with IC50 of 2.3 M and compound 43 has
shown moderate resting state block of 57% inhibition at 10 M. The other three lead molecules 13, 32 &
5
1 did not show significant resting state inhibition, Table 7. These lead molecules exhibited low potency
for fast inactivated state of hNa 1.5 with an inhibition of <50% for 1st pulse and these values were shifted
V
up to 80% for 10th pulse at 10µM, Table 7. All the five lead molecules have shown very good metabolic
stability in liver microsomes across the species except for compound 43 in mice liver microsomes, Table
8
. The chroman and indane compounds have shown good permeability in Caco-2 assay and moderate to
ACS Paragon Plus Environment

Page 17 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
high plasma protein binding across the species, Table 8. All these compounds did not show any cytotoxicity
or hERG inhibition (no to very low (<10%) inhibition) at 10 M (data not shown).
Table 7. State Dependent Properties of Lead Compounds
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Na
V
1.5
1.7 IC50 (nM)a
1.7a
Na
V
Na
V
a
(
% Inhibition@10M)
Use
Compd
Fast
Resting
IC50 (M) or %
Inhibition@10M
Fast Fast
Slow
Inactivation
Dependent
Inhibition
(25p)
Inactivation
1p)
Inactivation
(1p)
Inactivation
(10p)
(
13
29
32
43
51
15.9±0.9
15.9 ± 1.2
39.6 ± 3.9
35.1±1.3
33±2.4
45.3±1.3
44.4 ± 2.2
134 ± 8.4
67.4±3.2
99.7±5.8
29.8±1.23
34.3 ± 1.5
108 ± 4.6
52.1±2.8
71±4.9
9.5±0.43µM
2.28±0.28M
38±2% @10M
57±2%@10µM
~8.1±0.58µM
34±2%
46±2%
29±2%
19±1%
40±1%
64±2%
80±1%
57±2%
71±2%
65±1%
a
Manual patch clamp EP assay; IC50 values are at V1/2 of slow inactivation and fast inactivation.
Table 8. Metabolic Stability, Permeability and Plasma Protein Binding of Lead Molecules
Permeability
Caco-2 Papp (nm/sec)
Metabolic Rate (nmol/min/mg)
% PPB
Rat Human
Compd
MLM
0
RLM
0.01
0
DLM
0.01
0.03
0.02
0.02
0.02
MoLM
0.02
0.01
0.02
0.03
0.03
HLM
0
A-B
280
418
395
376
315
B-A
268
334
368
254
367
ER
1
Mice
88.5
1
3
9
91
95.4
97.1
98.3
97.6
99.7
2
0
0
0.9
0.9
0.7
1.2
95.4 93.4
98 99.2
32
43
51
0.04
0.16
0.03
0.02
0.02
0.04
0
0.02
0.03
94.1 92.8
92.9 99.3
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 18 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
These five lead compounds were also subjected to detailed in-vivo pharmacokinetic profiling in rodent
and non-rodent species. All the compounds have shown very good oral bioavailability with low clearance
and good t1/2 in mice, rats and dogs, Table 9. These compounds have shown emesis in dogs post iv and/or
po dosing. This could be a species specific phenomenon. To ensure, pharmacokinetic profile of compound
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
5
1 was determined following oral gavage to cynomolgus monkeys at a higher dose of 150 mg/kg to a
group of 6 animals (3 male and 3 female). No emesis was observed. The mean Tmax was found to be 2.0h
and 1.5h in male and female, respectively. The mean Cmax and AUClast were found to be similar in male
(66±8µM and 684±122h*µM) and female (69±5µM and 601±12h*µM) suggesting no gender difference.
Table 9. Pharmacokinetic Profile of Lead Molecules
IV (1 mpk)
PO (10 mpk)
Compd Species
AUClast
T1/2
Vss
AUClast
Cmax
CL
Cmax
(μM)
%
F
(μM)
(ml/min/kg)
(h*μM)
(h)
4.2
(l/kg)
(h*μM)
Mice
2
12
11±3
59±20
12
2.7
0.9
15
13±6
10±1.7
17
107
114±45
117±41
171
89
100
40±4
>100
74
13
29
32
Rat
Dog*
Mice
Rat
2.2±0.2
6.9±1
2
4.2±1
7.8±3.8
3.2
3±0.8
0.5±0.3
2.6
0.9±0.1
0.3±0
0.8
1.7±0.5
5.7±0.7
3
15±2
70±13
7.6
9.8±5
24±4
1.5
2±0
1.2±0.4
0.5±0
0.6
6.6±1.8
14±2
18
111±29
218±87
63
Dog*
Mice
Rat
0.3±0.1
3.9
78±22
83
2.6±0.2 5.5±0.8 1.9±0.3
5.5±0.9
2±1
0.8±0.1
0.4±0.1
0.81
19±6
8.5±3
9.6
112±11
50±30
62
>100
51±16
50
Dog*
Mice
Rat
6.5±1
2
19±10
12
2.5±0.6
4.3
2.5
43
2.1±0.2 5.9±0.6 2.8±0.2
5.1±0.4
0.5 ± 0
3.3
1.2±0.1
0.5 ± 0
0.8
2.1±0.5
nd
19±3
nd
31
Dog
4.9 ± 1
2
48 ± 1
8.5
11 ± 2
2.8
nd
Mice
Rat
18
84
99
51
2.5±0.5
7.9±5
2.7±0.3
4.9±2.6
1.0±0.3
10.9±0.7
46±7
58
Vehicle (Mouse, Rat & Dog): IV: DMA (10%): PEG300 (20%): 20%Captisol in NS (q.s.); PO: Tween80 (1%): 0.5%CMC solution (QS)
*PO at 5 mpk & emesis observed in all cases; nd - not done
Taken together, systematic SAR studies on chroman series for parallel optimization of several parameters
(as summarized in Fig. 3.) led to identification of many novel, potent and selective Na 1.7 inhibitors with
V
good solubility and good pharmacokinetic profile in rodent and non-rodent species devoid of any
ACS Paragon Plus Environment

Page 19 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
significant CYP inhibition, which is a major liability seen with many known Na 1.7 inhibitors. Based on the
V
overall profile of lead molecules, all the lead molecules 13, 29, 32, 43 and 51 were tested for efficacy at
0 mpk p.o. dose to understand the translation of in-vitro potency to in-vivo efficacy in different pain
models of mice. We have used in-house developed veratridine induced pain model in mice as a primary
model to assess the target engagement of Na 1.7. Veratridine acts as a neurotoxin and binds to voltage-
3
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
V
gated sodium channels leading to persistent activation and inhibits transitioning into the inactivated state,
thus stimulating a pain response.9a Compounds were administered orally 30 min prior to sub-planter
injection of veratridine. All of these compounds demonstrated statistically significant reduction in time
spent in nociceptive behavior (time spent in licking and/or flinching) compared to vehicle post veratridine
injection at 30 mpk po dose. Secondly, formalin induced inflammatory pain model was used as an efficacy
model. In this model, formalin generates a biphasic pain response, an acute phase (or Phase I) reflective
of direct activation of nociceptors and a tonic phase (or Phase II) related to inflammatory responses.22
Lead compounds were administered orally at 30 mpk dose 30 minutes prior to formalin challenge, the
total time spent in nociceptive behavior at 0 – 5 min (phase I) and 10 - 40 min interval (phase II) for each
mouse was measured and it was found that all the tested compounds have shown profound effect on
decrease in nociceptive behavior in phase II. The brain exposures of the compounds 13, 29, 32 and 43
were measured from PK-PD study at 30mpk dose (at 75 min post dose of compounds) and found to be
minimal (<1M) with 0.5%, 1.8%, 0.7%, and 4.5%, respectively, of the plasma exposure, whereas for
compound 51, the brain exposure was estimated to be 2.1M, which is 7% of plasma exposure. When
these lead molecules were assessed at 30 mpk oral dose in CCI induced neuropathic pain model in mice,23
to our puzzle, only compounds 29 and 51 have shown effect on paw-withdrawal threshold (mechanical
allodynia) using von Frey filaments (data not shown). However, compounds 29, 32 and 43 were not chosen
for further profiling due to resting state block of compound 29 and 43 and relatively inferior PK (low t1/2
in both rodent and non-rodent) profile of compound 32. The best two compounds 13 and 51 based on
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 20 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
overall profile were chosen for dose response studies. Compound 13 has shown flat dose response in
veratridine induced pain model at 3, 10 and 30 mpk p.o. dose. However, in formalin induced inflammatory
pain model, compound 13 demonstrated a very good dose-dependent reversal of nociceptive effects in
the acute phase of the experiment (phase I) and the tonic phase of the experiment (phase II) at 3, 10 and
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
3
0 mpk p.o. dose, compared to vehicle treated animals, with statistically significant reversal of nociceptive
pain at all the doses in the tonic phase, Fig. 4. Plasma concentration were measured at the end of the
study and the unbound drug concentration was determined to be 0.728M at 3 mpk dose which is 22 fold
higher than the mNa 1.7 IC50 (32.6 ± 1.6 nM). Compound 13 did not show any effect even at 100 mpk dose
V
in CCI induced neuropathic pain model. However, compound 51 demonstrated a dose dependent
reduction in nociceptive effects in both veratridine induced pain model and in formalin induced
inflammatory pain model at 3, 10 and 30 mpk, p.o. dose, compared to vehicle treated animals, Fig. 5.
Plasma concentration were measured at the end of formalin study and the unbound drug concentration
was determined to be 0.343M at 3 mpk dose which is 10 fold above the mNa 1.7 IC50 (33 ± 1.8 nM).
V
Compound 51 has also shown significant effect in CCI induced neuropathic pain model at 100 mpk p.o.
dose, Fig. 5.
ACS Paragon Plus Environment

Page 21 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
•
•
•
Enantiomers have shown significant difference in potency
NCEs with a-orientation of Phenyl is active isomer
3
Introduction of chiral sp carbon improved solubility & ADME properties
Thiazoles
Thiadiazoles
Substituted pyridines
Pyrimidines
Pyridazine
Pyrazine
Oxazole
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
CF replacement with
3
-Cl, bulky alkyl, alkoxy
led to decrease in potency
Isoxazole
Hetero aryl modification
Outcome
•
•
•
•
•
Significant role in selectivity over Na 1.5
Dialed out CYP3A4 inhibition
Significantly improved CYP2C9 inhibition
Improved PK profile
Overall best profile with 2-amino pyrimidine
v
R group modification
Outcome
Various substitutions tolerated –aryl, heteroaryl,
heterocyclyl, halide, substituted alkyl, etc. on 2-amino
thiazole sulfonamide
•
•
Heteroaryls are tolerated on 2-amino pyrimidine sulfonamide
Figure 3. Overall SAR Summary of Chroman Series
Figure 4. Effect of Compound 13 in Different Pain Models in Mice; a) Dose response effect at 3, 10, 30 mpk, p.o. on veratridine
induced nociceptive behaviors; b) Dose response effect at 3, 10, 30 mpk, p.o. on phase I and II of formalin induced nociceptive
behaviors; Results are presented as the mean ± SEM response (n = 8–10/group) and were analyzed by One way ANOVA followed
by Dunnett’s post-hoc tests; **P < 0.01, ***P < 0.001, compared to respective vehicle group.
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 22 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Figure 5. Effect of Compound 51 in Different Pain Models in Mice; a) Dose response effect at 3, 10, 30 mpk, p.o. on veratridine
induced nociceptive behaviors; b) Dose response effect at 3, 10, 30 mpk, p.o. on phase I and II of formalin induced nociceptive
behaviors; c) Response of compound 51 at 100 mpk, p.o. in CCI induced allodynia. Gabapentine (60 mpk, ip) was used as the
positive control. Results are presented as the mean ± SEM response (n = 10–13/group) and were analyzed by One way ANOVA
followed by Dunnett’s post-hoc tests; **P < 0.01, ***P < 0.001, compared to respective vehicle group.
Table 10. State-dependent block of Compound 51 across species for Na 1.7
V
IC50 (nM)/% Inhibition*
State
Human
Monkey
Mouse
33 ± 1.8
Slow Inactivation
Fast Inactivation (1P)
Use Dependence (25P)
Resting
33 ± 2.4
60 ± 1.3
99.7 ± 5.8
71 ± 4.9
195 ± 11
118 ± 12
145 ± 13
89 ± 4.5
~8.1 ± 0.58 µM
31 ± 3% @10 µM
36 ± 1 % @10 µM
*
Manual patch clamp EP assay; IC50 values are at V1/2 of slow inactivation and fast inactivation.
Table 11. Selectivity of Compound 51 over Other Na Isoforms
V
%
Inhibition at 10 M*
Parameter
Resting
State (1
Pulse)
hNa
V
1.1
hNa
V
1.2
hNa
V
1.3
hNa 1.4 hNa 1.5
V
V
hNa
V
1.6
hNa
V
1.8
hNa 1.9
V
st
8 ± 2
72
13 ± 2
6 ± 1
7.7 ± 1
10 ± 1
40 ± 1
32 ± 1
59 ± 2
3 ± 1
6 ± 1
Inactivated
7
.1 ± 1.1
nd
State (2
51 ± 2
8.1 ± 0.6
27 ± 2
Not done
µM (IC50
)
Pulse)
*
Manual patch clamp EP assay; 2nd pulse at V1/2 of inactivation
ACS Paragon Plus Environment

Page 23 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
Having identified compound 51 as a highly desirable lead compound over compound 13 due to its efficacy
in both inflammatory and neuropathic pain models, compound 51 was chosen for testing potency across
rodent and non-rodent species and for extensive selectivity profiling across human Na
In manual patch clamp electro physiology assay, compound 51 exhibited state-dependent block of human,
monkey and mouse Na 1.7 with a similar potency range, Table 10. Pleasingly, it has also shown good
selectivity against other Na isoforms and minimal inhibition at 10 M for resting state across different
Na isoforms, Table 11. Further, no significant inhibition observed at 10 M in Eurofins CEREP ExpresS
V
channel subtypes.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
V
V
V
panel of 55 targets including GPCRs, other ion channels and transporters.
CHEMISTRY
Synthesis of chroman sulfonamide derivatives started with 7-bromochroman-4-one 55. A palladium
catalyzed thiolation24 converted 55 to 7-(benzylthio)chroman-4-one 56. The palladium catalyzed
Barluenga reaction25 was used for the elaboration of chroman ring at the 4-position with various
substituted phenyl groups, Scheme 1. To convert the 7-(benzylthio)chroman-4-one 56 to a suitable
25
substrate for the Barluenga reaction, tosylhydrazide 57 was prepared from 56 and then coupled with 2-
substituted 1-bromo-4-(trifluoromethyl)benzene 58a, b to get 7-(benzylthio)-4-substituted-2H-chromene
5
9a, b. The thiobenzyl group was then oxidized in situ to the sulfonyl chloride using sulfuryl chloride and
then converted to the stable 2H-chromene-7-sulfonates 60a, b.26
Scheme 1: Synthesis of Compounds 1, 4, 13-20 & 25-28
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 24 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
i
o
o
Reagents and conditions: (a) BnSH, Pd
2
(dba)
3
, ( Pr)
2
NEt, 80 C, 1 h, 60%; (b) Tosylhydrazide, MeOH, 80 C,
o
16 h, 94%; (c) Na
2
CO
3
, PdCl
2
(dppf).DCM, Dioxane-H
2
O (85:15), 100 C, 5-8 h 47-76%; (d) (i) SO
2
Cl , AcOH,
2
o
o
cat. H
2
O, DCM, -5 C , 1 h; (ii) Pentafluorophenol, Et
3
N, DCM, 0 C, 1 h, 60-71%; (e) H
2
(3 atm), 10% Pd-C,
o
EtOAc, 4-6 h, rt, 83-94%; (f) (i) HetArNH
2
, LiHMDS (1M in THF), 0 C to rt, 1-2 h; (ii) Chiral separation, 9-
44%.
The double bond was then reduced to generate the chiral center of the 4-substituted chroman ring 61a,
b by using hydrogen in presence of 10% Pd-C. The racemic compounds 61a, b was then reacted with
various heteroaryl amines in the presence of LiHMDS to give the final sulfonamide derivatives. At this
stage, the enantiomers of the individual compounds were separated by preparative chiral HPLC to obtain
chirally pure active isomers of chroman compounds 1, 4, 13-20, & 25-28.
Scheme 2: Synthesis of Compounds 2-3, 5-12, 21-24, 29-31, 36-50
ACS Paragon Plus Environment

Page 25 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
o
Reagents and conditions: (a) (i) NCS, AcOH-H
2
O (4:1), 0 C to rt, 45 min; (ii) Pentafluorophenol, Et
3
N, DCM,
0 oC, 45 min, 65%; (b) Tosylhydrazide, MeOH, 80 C, 16 h, 94%; (c) Na
o
, PdCl
(dppf).DCM, Dioxane-H
, LiHMDS
, 16 h, 79%.
O
2
CO
3
2
2
o
(
85:15), 100 C, 1-2 h, 38-61%; (d) H
2
(3 atm), 10% Pd-C, EtOAc, 4-6 h, rt, 50-90%; (e) (i) HetArNH
2
o
(
1M in THF), 0 C to rt, 1-2 h, 9-69%; (ii) chiral separation; (f) HCHO, AcOH cat., NaBH(OAc)
3
Alternatively, intermediate 56 was first oxidized in situ to the corresponding sulfonyl chloride with N-
chlorosuccinimide and then converted to the stable pentafluoroester 62, Scheme 2. This ester was
converted to the tosylhydrazide 63 and then coupled with various 2-substituted 1-bromo-4-
(
trifluoromethyl)benzene 58c-m to give the 2H-chromene-7-sulfonates 60c-m. The double bond was
reduced to get the racemic 4-substituted chroman compounds 61c-m by using hydrogen in presence of
0% Pd-C, Scheme 2. The enantiomers were either separated at this step and the purified enantiomers
1
were separately reacted with various heteroaryl amines in the presence of LiHMDS to give the final
sulfonamide compounds or the amine addition reactions were done on the racemic 61c-m and then the
enantiomers of the final compounds were separated by preparative chiral HPLC.
Scheme 3: Preparation of Compounds 32-35
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 26 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Reagents and conditions: (a) [(t-Bu)
3
PH]BF
4
, PdCl
2
(dppf).DCM, 2-methyl-5-(4,4,5,5-tetramethyl-1,3,2-
o
o
dioxaborolan-2-yl)pyridine, Dioxane, 100 C, 2 h, 91%; (b) HetArNH
2
, LiHMDS (1M in THF), 0 C to rt, 1-2
h, 9-78%.
Compounds 32-35 were synthesized from (S)-enantiomer of 61l (obtained by separation of the
enantiomers of racemic 61l using chiral column) by following the reaction sequence as shown in Scheme
3
. Suzuki coupling of 61l(S) with 2-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine gave
the pentafluoro intermediate 61n, which was then converted to the final sulfonamide compounds 32-35
using different heteroaryl amines as described in earlier scheme.
Scheme 4: Preparation of Compounds 51-54
ACS Paragon Plus Environment

Page 27 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
o
Reagents and conditions: (a) Tosylhydrazide, MeOH, 80 C, 16 h, 90%; (b) Na
2
CO
3
, PdCl (dppf).DCM,
2
o
o
Dioxane-H
2
O (85:15), 100 C, 4-6 h, 51-58%; (c) (i) NCS, AcOH-H
2
O (4:1), 0 C to rt, 45 min; (ii)
o
Pentafluorophenol, Et
3
N, DCM, 0 C, 45 min, 53-55%; (d) H
2
(1 atm), 10% Pd-C, EtOAc, rt, 16 h, 87-91%;
(
e) N-(2,4-dimethoxybenzyl)pyrimidin-2-amine (for 68a) or pyrimidin-2-amine (for 68b) LiHMDS (1M in
o
THF), 0 C to rt, 1-2 h, 35-55%; Chiral Separation for 54; (f) (i) [(t-Bu)
3
PH]BF
4
, PdCl (dppf).DCM,
2
o
heteroarylboronic acid, Dioxane, 100 C, 1-2 h; (ii) TFA, DCM, rt; (iii) Chiral Separation, 23-39%.
Scheme 4 depicts the reaction sequence for the synthesis of the indane sulfonamide derivatives and the
transformations were carried out by following the same chemistry already described for the chroman
sulfonamide compounds in Scheme 1. The starting material in this case was 5-(benzylthio)-2,3-dihydro-
1
H-inden-1-one 64. The keto compound was first converted to the tosyl hydrazide 65 followed by coupling
with either 1-bromo-2-chloro-4-(trifluoromethyl)benzene 58l or 4-(2-bromo-5-(trifluoromethyl)phenyl)
morpholine 58m to give the substituted 1H-indene compounds 66a and 66b, respectively. The thiobenzyl
group of 66a,b was then oxidized in situ to the sulfonyl chloride with N-chlorosuccinimide and then
ACS Paragon Plus Environment

Journal of Medicinal Chemistry
Page 28 of 66
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
converted to the stable 1H-indene pentafluoroester 67a,b which on reduction provided the racemic
indane derivatives 68a,b. The pentafluoroester derivative 68a was then reacted with N-(2,4-
dimethoxybenzyl)pyrimidin-2-amine in the presence of LiHMDS to give the sulfonamide derivative 69. The
racemic compound 69 was converted to the final compounds 51-53 by Suzuki coupling with the respective
boronic acids or esters, followed by de-protection of the dimethoxybenzyl group on the sulfonamide and
separation of the racemic compounds into required enantiomers by preparative chiral HPLC. Compound
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
5
4 with the morpholine side chain was obtained from 68b by base mediated reaction with pyrimidin-2-
amine and subsequent separation of the (S)-enantiomer by preparative chiral HPLC.
CONCLUSION
Chroman based sulfonamide derivatives have been identified as potent, selective and state-dependent
Na 1.7 inhibitors with very good PK and low CYP inhibition profile through systematic SAR studies and
V
parallel optimization approach. Switching from benzoxazine scaffold to chroman bicyclic system with a
3
sp chiral carbon led to significant improvement in solubility and overall pharmacokinetic profile.
Replacement of thiazole with other heteroaryl substitution on sulfonamide was tolerated on chroman
core with many pendant groups and also resulted in significant improvement in selectivity over Na 1.5
V
and lowered CYP2C9 inhibition. It is important to note that the modification of heteroaryl substitution on
sulfonamide led to steep decrease in activity on many other scaffolds.17b,e,18b,19a,b Two major challenges
were addressed with this one modification on sulfonamide, however extensive SAR studies with synthesis
of a large number of NCEs in a library fashion were conducted to identify lead molecules with defined
target product profile since the SAR was not additive. Also, good translation of SAR was observed on
similar bicyclic system such as indane core. More importantly, the lead molecules 13, 29, 32, 43 and 51
have shown favorable PK profile in both rodent and non-rodent species and robust efficacy translation in
veratridine induced pain and formalin induced inflammatory pain models in mice. However, to our puzzle,
ACS Paragon Plus Environment

Page 29 of 66
Journal of Medicinal Chemistry
1
2
3
4
5
6
7
8
9
only selected leads have shown efficacy in CCI induced neuropathic pain model in mice. Finally, the
detailed characterization of compound 51 has indicated that it has the desired properties for further
evaluation as a therapeutic agent for pain.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
EXPERIMENTAL SECTION
Chemistry. All reagents were purchased from commercial suppliers and used without purification unless
otherwise noted. All anhydrous reactions were performed under a nitrogen atmosphere using dry
solvents. Silica gel column chromatography was carried out on ISCO CombiFlash Companion with
prepacked columns. Nuclear magnetic resonance ( H & 13C NMR) spectra were recorded on a Bruker
AvanceV-III 400 MHz spectrometer. All spectra were determined in the solvents indicated, and chemical
shifts are reported in δ units downfield from the internal standard tetramethylsilane (TMS) with inter
proton coupling constants reported in hertz (Hz). All NMR spectra were analyzed using the MestReNova
1
1
0.0 program software. Liquid chromatography-mass spectrometry (LC-MS) analyses were performed
either on an Agilent 1200 Series LC instrument coupled to an Agilent Iontrap MS instrument or a Thermo
Scientific Accela LC instrument coupled to a Thermo Scientific MSQ Plus instrument. Preparative HPLC
were performed on a Gilson GX 281 series using a YMC ODS-A column or YMC Triat column (50 mm X 250
mm, 10 µm) using either acid (0.1 % formic acid) or base (0.1% NH
4
OH) modified MeCN-H O gradients.
2
Most of the final compounds were prepared as racemic mixture and separated using preparative chiral
HPLC at the final or pre-final stage. Preparative chiral separations and enatiomeric purity determination
were carried out using Knauer Azura 2.1L purification system with different columns and mobile phase.
Methods and conditions for chiral separation of all the relevant compounds are provided in the supporting
information. Compound purity was determined by reverse phase HPLC performed on an Agilent 1290
Infinity equipped with either a Zorbax Eclipse Plus C18, 50 X 2.1 mm, 1.8 µm or an Acquity Beh C18, 50 X
2
.1 mm, 1.7 µm column using either acid (0.1 % formic acid) or base (0.1% NH
4
OH) modified MeCN-H O
2
gradients. Purity of the final compounds were determined by HPLC and were greater than 95% for all the
compounds except for compounds 23 (94%), 42 (94%) and 45 (93%). The synthetic procedures and the
analytical data for the intermediates 58a-m are provided in the supporting information.
(
S
&
R)-4-(2-(1-methyl-1H-pyrazol-5-yl)-4-(trifluoromethyl)phenyl)-N-(thiazol-2-yl)chromane-7-
sulfonamide (1a & 1b). Thiazol-2-amine (25.5 g, 255 mmol) was dissolved in 2L THF and the solution was
cooled to 0 °C and a 1M solution of LiHMDS (242 mL, 242 mmol) was added to it slowly. After the addition,
ACS Paragon Plus Environment