The discovery of tetrahydropyridine analogs as hNav1.7 selective inhibitors for analgesia

Article abstract of DOI:10.1016/j.bmcl.2017.03.043

hNav1.7 small molecular inhibitors have attracted lots of attention by its unique analgesic effect. Herein, we report the design and synthesis of a novel series of tetrahydropyridine analogs as hNav1.7 inhibitors for analgesia. Detail structural–activity

Full text of DOI:10.1016/j.bmcl.2017.03.043

Accepted Manuscript
The Discovery of Tetrahydropyridine analogs as hNav1.7 selective inhibitors
for analgesia
Wentao Wu, Zhixiang Li, Guangwen Yang, Mingxing Teng, Jian Qin, Zhijing
Hu, Lijuan Hou, Liang Shen, Haiheng Dong, Yang Zhang, Jian Li, Shuhui Chen,
Jingwei Tian, Jianzhao Zhang, Liang Ye
PII:
S0960-894X(17)30274-3
DOI:
http://dx.doi.org/10.1016/j.bmcl.2017.03.043
BMCL 24795
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
5 January 2017
6 March 2017
16 March 2017
Please cite this article as: Wu, W., Li, Z., Yang, G., Teng, M., Qin, J., Hu, Z., Hou, L., Shen, L., Dong, H., Zhang,
Y., Li, J., Chen, S., Tian, J., Zhang, J., Ye, L., The Discovery of Tetrahydropyridine analogs as hNav1.7 selective
inhibitors for analgesia, Bioorganic & Medicinal Chemistry Letters (2017), doi: http://dx.doi.org/10.1016/j.bmcl.
2017.03.043
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Bioorganic & Medicinal Chemistry Letters
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The Discovery of Tetrahydropyridine analogs as hNav1.7 selective inhibitors for
analgesia
Wentao Wua, Zhixiang Lia, Guangwen Yanga, Mingxing Tenga, Jian Qina, Zhijing Hua, Lijuan Houa, Liang
Shena, Haiheng Donga, Yang Zhanga,∗ Jian Lia, Shuhui Chena, Jingwei Tianb,∗ Jianzhao Zhangb, Liang Yeb
a WuXi AppTec (Shanghai) Co., Ltd., 288 FuTe Zhong Road, Shanghai 200131, People’s Republic of China
bKey Laboratory of Molecular Pharmacology and Drug Evaluation, Ministry of Education, Collaborative Innovation Center of Advanced Drug Delivery System
and Biotech Drugs in Universities of Shandong, Yantai University, Yantai 264005, People's Republic of China
ARTICLE INFO
ABSTRACT
Article history:
Received
hNav1.7 small molecular inhibitors have attracted lots of attention by its unique analgesic effect.
Herein, we report the design and synthesis of a novel series of tetrahydropyridine analogs as
hNav1.7 inhibitors for analgesia. Detail structural–activity relationship (SAR) studies were
undertaken towards improving hNav1.7 activity, in vitro ADME, and in vivo PK profiles. These
efforts resulted in the identification of compound (-)-15h, a highly potent and selective hNav1.7
inhibitor with good ADME and PK profiles.
Revised
Accepted
Available online
Keywords:
2009 Elsevier Ltd. All rights reserved.
Voltage-gated sodium channal
hNav1.7
Tetrohydropyridine
Pain
Analgesia
Pain is the most common symptom for which patients seek
medical attention. Although current analgesic drugs help many,
only an estimated one in four of those with pain achieves
adequate relief. Many marketed pain-relief drugs are also often
associated with poor tolerability, long-term safety, as well as
concerns over abuse potential1. This unmet medicine needs
required sustained development of drugs with novel mechanism
of actions2. Voltage-gated sodium channel (Navs) are
transmembrane ion channel proteins. The importance of sodium
channel inhibitors as a class of therapeutics is highlighted by
their successful exploitations in clinic as anticonvulsants,
antiarrhythmics, local anesthetics, and analgesics3. However,
non-selective modulation of sodium channels can be associated
with undesirable side effects such as cardiac toxicity due to
inhibition of hNav1.5 channel4. Among the nine Nav1 channels
(Nav1.1 to Nav1.9) identified up to date, Nav1.7 is
predominantly expressed in dorsal root ganglion and trigeminal
ganglia neurons5. Inherited loss of function mutations of hNav1.7
in humans were reported to cause congenital inability to
experience pain. For this reason, hNav1.7 inhibitors have
attracted lots of attention for new analgesic drug developments,
although the similar amino acid sequence homology between the
nine Nav subtypes made locating such a selective hNav1.7
companies reported their achievements in identifying and
developing selective hNav1.7 Of all,
inhibitors6.
arylsulfonamide-based hNav1.7 inhibitors are most promising by
their high potency and good Nav subtype selectivity. A
representative of this chemotype is PF-05089771, which was
once in phase 2 clinical trial developed by Pfizer Inc. for
erythromelalgia and diabetic peripheral neuropathic pains. Part of
our pain research programs were interested in identifying a
hNav1.7 selective inhibitor. Herein, we reported the discovery of
a new series of tetrahydropyridine analogs as potent and subtype
selective hNav1.7 inhibitors.
Figure 1. Structures of PF-05089771, hit compound (-)-1 and its
tetrahydropyridine analog 2a.
In our previous works, a hit compound (-)-1 was identified
(Figure 1), which showed good potency (IC50: 0.024 µM) against
i—nh—ibit—ors very challenging. Recently, several pharmaceutical
hNav1.7, along with >100 fold selectivity over hNav1.5 (IC50
>
∗ Corresponding author. Tel.: +86-021-50172812; fax: +86-21-50461000; e-mail: zhang_yang@wuxiapptec.com
∗ Corresponding author. Tel.: +86-535-3808268; fax: +86-535-3808268; e-mail: Tianjingwei@luye.com

10 µM) and the hERG (human ether a-go-go-related gene)
potassium channel (IC50 > 10 µM) (Table 1). However, further
profiling of compound (-)-1 in rat indicated poor
pharmacokinetic properties such as high clearance, relatively low
bioavailability (18.4%) and high volume of distribution
(Vdss=8.9 L/kg). The observed low bioavailability might be due
to poor permeability of compound (-)-1, with Papp (A to B) only
at 0.06 x 10-6 cm/s in Caco-2 assay. The primary amino group
anchored on cyclohexene ring was speculated for this liability.
The high volume of distribution of compound (-)-1 observed in
rat PK also brought up a concern of brain penetration, which was
confirmed in the rat PK studies upon analysis of brain and
plasma samples at 2 hours postdose. The brain to plasma (B/P)
ratio of compound (-)-1 was determined to be 0.26, which was
not desirable considering those common CNS side effects
observed in pain therapeutic areas. To address these undesirable
properties, the primary amine was repositioned on the six
member ring with the rationale to reduce the basicity of
compound (-)-1. It is well documented in literature that basicity
plays key roles in determining compounds’ permeability and
volume of distribution7. A unique tetrahydropyridine analog 2a
was prepared and evaluated in in vitro and in vivo. Although the
in vitro potency of 2a (hNav1.7 IC50: 0.22 µM) showed a 10-fold
decrease as comparison to (-)-1, it showed significantly improved
Caco-2 cell line permeability with Papp (A to B) at 3.2 x 10-6
cm/s and excellent systemic exposure was observed in rat PK
(AUC0-last: 3.56 µM·hr) via intravenous bolus injection (IV)
administration at 1 mg/kg. More importantly, the Vdss was
reduced from 8.9L/kg of (-)-1 to 0.6L/kg, and oral bioavailability
was also improved to 64.8% when oral gavage dosing at 10
mg/Kg. The B/P ratio of compound 2a was determined to be
0.017 with no significant CNS distribution, which might reduce
the potential CNS side effects. Further in vivo pharmacodynamic
studies indicated (-)-2a, one of the enantiomers of 2a, displayed
significant analgesic effect in rat formalin model by oral gavage
administration (PO) at a dose of 100 mg/kg. Analysis of the
associated PK/PD studies disclosed that when the unbound
plasma concentrations at terminal point covered above 4x
hNav1.7 IC50, analgesic effect could be observed (see
supplementary information, SI page 72). Encouraged by the
overall profiles of this newly identified chemotype compound,
optimization of the structure–activity relationship around 2a was
carried out.
converted to enol trifilate 8 by treatment with NaHMDS and
PhNTf2. Suzuki coupling of 8 with corresponding boric acid
result in substituted tetrahydropyridine
9 smoothly. After
removal of TBS protection with TBAF, alcohol 10 was obtained,
which was further coupled with the known sulfonyl amine 119,
followed by deprotection under acidic condition to yield final
product 2.
To obtain enantiopure products of compound 2a, a lipase
catalyzed kinetic resolution of racemic intermediate 10a10 was
employed to prepare chiral alcohol (+)-10a and acetate (-)-13a
(Scheme 2). Alcoholysis of (-)-13a under basic condition
afforded the other enantiomer (-)-10a at high ee value in good
yield. Both (-)-2a and (+)-2a were obtained following the
conditions described in Scheme 1. The hNav1.7 inhibition
potency of compound (-)-2a showed about 2-fold increase over
racemic 2 with IC50 at 0.090 µM, and its enantiomer (+)-2a lost
of potency completely with IC50 >10 µM (Table 2).
Scheme 1. General route to prepare compound 2.
Table1. Profiles of compound (-)-1 and 2a
Compound
(-)-1
2a
hNav1.7 IC50 (µM):
hNav1.5 IC50 (µM):
hERG IC50 (µM):
Dose (mg/kg)
0.024
>10
10.3
0.5
0.22
>10
-
1.0
2.7
Scheme 2. Lipase catalytic kinetic resolution of alcohol 10.
t1/2 (hr)
3.6
Rat PK
(i.v.)
Our first region in SAR exploration around compound 2a is
the top left phenyl ring and the structure modification’s impact of
this region on hNav1.7 inhibition was evaluated (Table 2). Non
F-substituted phenyl compound 2b showed about 2 fold hNav1.7
potency decrease as comparison to 4-F analog 2a. Relocation of
the fluorine to 3-position afforded compound 2f, which had equal
potency with 2a. Further modification of the mono-substituent
functional groups, either electron-withdrawing or electron-
donating group (such as Cl, Me, OMe, etc), were not tolerated
with 2 to10 fold potency decrease regardless of their substitution
position (2c-2e, 2g and 2h). Di-substituted compounds were also
prepared (2i-2k), and bioassay results indicated 2i (3,4-diF) and
2k (2-Cl-4-F) had comparable hNav1.7 potency with 2a, whereas
2,4-diF substituted compound 2i lost most of hNav1.7 potency.
Vdss (L/Kg)
Cl (mL/min/kg)
AUC0-last (µM)
Dose (mg/kg)
tmax (hr)
8.9
39.2
0.34
2.0
1.7
0.25
18.4%
0.6
4.5
3.56
10
3.8
41.78
64.8%
Rat PK
(p.o.)
AUC0-last (µM)
F (%)
An efficient synthetic route was designed for compound 2a as
depicted in Scheme 1. Protected the benzylic alcohol of
commercial available starting material 3 with TBS, followed by
hydrogenation to afford Piperidin-3-ol 58. The amino group of 5
was protected with Boc to generate alcohol 6. Ketone 7 was
obtained via IBX oxidation of alcohol 6, which was further

Heterocycles were further examined as replacement of phenyl of
2a (2l-2p). Comparison to 2a, chloro-substituted thiophene 2l
had 2 fold potency decrease. The basic pyridine derivative 2m
resulted in over 10 fold potency decrease. Furan analog 2n,3,6-
dihydro-2H-pyran 2p and pyrazole analog 2o were also not
tolerated with significant potency drops.
Cl
Cl
F
0.42
0.70
0.25
0.64
4.80
14a
14b
14c
14d
14e
Table 2. SAR study for phenyl substituent
Cl
Cl
Compound ID
R
hNav1.7 IC50 (µM)
4-F-Ph
4-F-Ph
0.22
0.090
>10
2a
(-)-2a
(+)-2a
2b
Next, we tried to replace the middle O-linker of 2a with N-
Linker (Table 4). Compound (-)-15a showed comparable in vitro
potency with (-)-2a, but surprisingly switching the right 5-amino-
thiazole moiety to 2-amino-thioazole derivative (-)-15b showed 3
fold potency increase with hNav1.7 IC50 at 0.031µM. Encouraged
by this intriguing result, we further examined the SAR with
variations on the two phenyl rings in the left and middle portion
of (-)-15b. Unfortunately, none of these modifications could offer
potency raise, but more or less compound (-)-15c to (-)-15f
suffered from 2 to 10 folds of potency drops.
4-F-Ph
Ph
0.44
0.9
4-Cl-Ph
4-Me-Ph
4-OMe-Ph
3-F-Ph
2c
1.67
3.1
2d
2e
0.25
1.03
0.58
0.34
1.45
0.17
2f
3-MeO-Ph
3-Cl-Ph
3,4-diF-Ph
2,4-diF-Ph
2-Cl-4-F-Ph
2g
2h
Table 4. Profiles of N-Linker analogs
2i
2j
2k
0.59
3.92
1.54
>10
2l
2m
2n
2o
hNav1.7
IC50
(µM)
Compound
ID
Linker
R1
R2
Het
O
N
O
N
N
N
N
N
4-F
4-F
4-F
4-F
4-F
4-F
5-Cl-2-F
5-Cl-2-F
5-Cl-2-F
5-Cl-2-F
2-F-5-F
5-Cl
0.09
0.13
(-)-2a
(-)-15a
14a
2p
3.92
0.42
Then we turned our effort to optimize the right heteroaryl
group, which was adjacent to sulfonamide pharmacophore and
had been demonstrated to have significant effect on hNav1.7
potency in previous literature report11. A few representative
compounds on SAR evaluation of this region were summarized
in Table 3. The result indicated five member heteroaryls, such as
thiazole 14a, 1,3,4-thiadiazole 14b and 1,2,4-thiadiazole 14c,
potencies were tolerated with hNav1.7 IC50 <1 µM, but no
potency enhancement obtained over compound 2a. Pyridine 14d
is also tolerated with IC50 value at 0.64 µM, but pyrimidine 14e
resulted in potency decrease dramatically with hNav1.7 IC50 at
4.80 µM.
(-)-15b
(-)-15c
(-)-15d
15e
0.031
0.070
0.090
0.58
2-Cl-
4-F
5-Cl-2-F
Table 3. SAR of right hetero aryl modification
4-Cl 5-Cl-2-F
0.039
(-)-15f
Compound (-)-15b was then advanced to rat PK studies and it
demonstrated acceptable in vivo profiles, with good exposure
(AUC0-last: 3025 nM.hr), low clearance (5.5 mL/min/kg) at 0.5
mpk IV dosing and good oral bioavailability (49.8%) at 2 mg/kg
PO dosing. However, further examinations disclosed that (-)-15b
had severe CYP450 enzyme inhibition on 2C9 and 2C19 subtype,
with IC50 at 0.033 µM and 0.386 µM respectively. The potential
hNav1.7 IC50
(µM)
Compound ID
X
Het

drug-drug interaction restricted its further development. As this
CYP450 enzyme inhibition was not observed on any other of the
5-amino-thioazole analogs, we hypothesized 2-amino-thioazole
had a positive impact on CYP450 enzyme inhibition. Docking
models of (-)-15b with CYP2C9 and CYP2C19 enzymes were
performed to explain this phenomenon. Docking results (Figure
2) clearly showed that two hydrogen bonds were formed through
the N atom of 2-amino-thioazole of (-)-15b with Asn204 and
Arg108 of CYP2C9 respectively. For CYP2C19 model, one
Hydrogen bond was observed between the O atom of sulfonyl
group of (-)-15b and Asn107 of the enzyme.
Table 5. Further optimization of N-Linker analog (-)-15b
Compound
Het
(-)-15b
(-)-15g
(-)-15h
a)
hNav1.7 IC50 (µM):
hNav1.5 IC50 (µM):
hERG IC50 (µM):
1A2
0.031
-
-
>50
0.033
0.39
17.2
>50
0.013
-
-
>50
0.71
7.0
4.15
37.5
0.011
>10
4.1
>50
0.60
16.9
32.1
40.1
CYP
IC50
(µM):
2C9
2C19
2D6
3A4
Dose
0.5
3.0
3.0
(mg/kg)
t1/2 (hr)
Vdss (L/Kg)
Cl
1.6
0.27
1.59
0.22
6.9
0.16
Rat PK
(i.v.)
5.5
7.4
0.4
(mL/min/kg)
AUC0-last
(µM.hr)
Dose
b).
3.03
13.01
203.19
2.0
0.5
10
3.0
10
2.0
(mg/kg)
tmax (hr)
AUC0-last
(µM.hr)
F (%)
Rat PK
(p.o.)
6.0
9.69
22.6%
99.1
14.6%
49.8%
Figure 2. a). Docking of (-)-15b with CYP2C9. b). Docking of (-)-15b
with CYP2C19.
With these docking information in mind, we envisioned an
additional electron-withdrawing group on the thiazole ring could
reduce this CYP450 inhibition through electronic and steric
effect. Compound (-)-15g and (-)-15h were synthesized
following the route shown in the Scheme 3. To our delight, both
compounds showed 3 fold boost on hNav1.7 potency with greater
than a thousand-fold selectivity over hNav1.5 (Table 5). More
importantly, significantly improved CYP450 inhibition profiles
were observed as we expected. CYP2C9 IC50 of (-)-15g and (-)-
15h had over 20 folds decrease as comparison to the non-
substituted analog (-)-15b. CYP2C19 IC50 of (-)-15g was 7.0 µM
and (-)-15h was 16.9 µM, also demonstrated 20~40 fold
decrease. Further rat pharmacokinetic studies indicated (-)-15h
had good PK properties with IV dosing at 3 mg/kg: long t1/2 (6.9
hr), low clearance (0.4 mL/min/kg) and high AUC0-last (203.19
µM.hr). Although the oral bioavailability was moderate (14.6%)
at 10 mg/kg PO dose, its plasma exposure is still very satisfactory
(99.1µM·hr).
Scheme 3. Synthesis of compound (-)-15g and (-)-15h.
In conclusion, a new series of highly selective hNav1.7
inhibitors specified by unique tetrahydropyridine architecture
was discovered. Through fully structure-active relationship study,
we successfully identified a promising compound (-)-15h, which
demonstrated excellent hNav1.7 potency, good subtype
selectivity over hNav1.5 and good ADME/PK profiles. Further
research of compound (-)-15h including in vivo toxicity and
pharmacology studies are in progress and will be reported in the
future.

Acknowledgments
We appreciate Zhiqiang Liao, Ying Chen from Biology
Department, WuXi AppTec for their bioassay support, and
Guoping Hu from DDSU Department, WuXi AppTec for CADD
support. This work was supported by National Natural Science
Foundation of China (NO.81473188).
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The Discovery of Tetrahydropyridine
analogs as hNav1.7 selective inhibitors for
analgesia
Wentao Wu, Zhixiang Li, Guangwen Yang, Mingxing Teng, Jian Qin, Zhijing Hu, Lijuan Hou, Liang Shen,
Haiheng Dong, Yang Zhang, Jian Li, Shuhui Chen, Jingwei Tian, Jianzhao Zhang, Liang Ye
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