Monocyclic thiophenes as protein tyrosine phosphatase 1B inhibitors: Capturing interactions with Asp48

Article abstract of DOI:10.1016/j.bmcl.2006.06.051

A series of monocyclic thiophenes was designed and synthesized as PTP1B inhibitors. Guided by X-ray co-crystal structural information and computational modeling, rational design led to key interactions with Asp48 and improved inhibitory potency against PT

Full text of DOI:10.1016/j.bmcl.2006.06.051

Bioorganic & Medicinal Chemistry Letters 16 (2006) 4941–4945
Monocyclic thiophenes as protein tyrosine phosphatase
1B inhibitors: Capturing interactions with Asp48
Zhao-Kui Wan,a,* Jinbo Lee,a Weixin Xu,a David V. Erbe,b Diane Joseph-McCarthy,a
Bruce C. Followsa and Yan-Ling Zhangb
aChemical and Screening Sciences, Wyeth Research, 200 Cambridge Park Drive, Cambridge, MA 02140, USA
bCardiovascular and Metabolic Diseases, Wyeth Research, 200 Cambridge Park Drive, Cambridge, MA 02140, USA
Received 25 April 2006; revised 13 June 2006; accepted 14 June 2006
Available online 27 June 2006
Abstract—A series of monocyclic thiophenes was designed and synthesized as PTP1B inhibitors. Guided by X-ray co-crystal struc-
tural information and computational modeling, rational design led to key interactions with Asp48 and improved inhibitory potency
against PTP1B.
Ó 2006 Elsevier Ltd. All rights reserved.
Protein tyrosine phosphatase (PTP1B) is an intracellular
protein tyrosine phosphatase expressed in insulin
responsive tissues1 and shown to be a negative regulator
of insulin signaling as evidenced by studies with PTP1B
knockout mice2 as well as studies using anti-sense oligo-
nucleotides.3 It is thus considered as one of the most
promising therapeutic targets2,3 among the large PTPase
family for potential treatment of type 2 diabetes and
obesity. Vast efforts toward the development of PTP1B
inhibitors have been disclosed.4,5 We have recently
reported ring-fused thiophenes as PTP1B inhibitors
(Fig. 1).6 A more potent and efficient monocyclic thio-
phene series is reported herein.
The X-ray co-crystal structure of PTP1B and 2 is
shown in Figure 3. In this structure, the WPD-loop
is in the catalytically competent closed conformation
and the aromatic thiophene ring is p-stacking between
Phe182 and Tyr46, while the C2 and C3 carboxylates
provide opportunities for a salt bridge formation with
Lys120 and Arg221, respectively. It was also observed
that the ether oxygen at the C3 position forms a net-
work of water-mediated hydrogen bonds with Ala217,
Arg221, and other residues in the active site. More
interestingly, compared to the fused thiophene sys-
tems, the C4 bromo group more tightly packs against
Ile219, likely contributing to the better inhibition
against PTP1B. Replacement of this group with small-
er or larger substituents led to diminished inhibitory
activity against PTP1B (Scheme 1 and Table 1) as pre-
dicted by modeling. Smaller groups such as H or Me
(Table 1) have reduced van der Waals and hydropho-
bic interaction with Ile219, while the larger groups are
not sterically accommodated as revealed by the X-ray
co-crystal structure and subsequent molecular
modeling.
While the fused thiophenes were shown to be reversible
and competitive PTP1B inhibitors, close examination of
X-ray co-crystal structures suggested that the second
and the third fused ring might not be necessary for bind-
ing to the enzyme active site. Indeed, removal of the
fused rings from the parent compound 1 (R = H,
Ki = 230 lM) afforded the monocyclic thiophene 2 with
comparable activity (Ki = 160 lM, Fig. 2). Compound 2
was also shown to be a reversible and competitive
PTP1B inhibitor.
In contrast, introduction of the C5-substitution turned
out to be much more productive. For instance, incorpo-
ration of an additional bromo group on the C5 position
gave compound 10 (Ki = 18 lM, Table 2) synthesized
according to the procedures in Scheme 1, displaying a
9-fold increase in inhibition against PTP1B in compari-
son with compound 2 (Ki = 160 lM, Table 1). Replacing
the bromo group with a phenyl ring in compound 11
Keywords: Protein tyrosine phosphatase 1B (PTP1B).
*
Corresponding author. Tel.: +1 617 665 5635; fax: +1 617 665
5682; e-mail: zwan@wyeth.com
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.06.051

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Z.-K. Wan et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4941–4945
Table 1. The effect of C4-substitution on the inhibition against PTP1B.
O
S
N
HO
O
R
S
O
HO
O
O
R
1
OH
O
OH
Figure 1. Bicyclic thiophenes as PTP1B inhibitors.
Compound
R
Kia (lM)
O
2
S
HO
2
6
7
8
Br
H
160
>1000
>1000
>1000
4
O 3
Br
Me
Ph
O
2
OH
a Enzymatic assay6 was carried out at room temperature in 96-well
plates. To an assay buffer, containing 50 mM of 3,4-dimethyl gluta-
rate, 1 mM EDTA, 1 mM TCEP, and 0.01% Triton (pH 7.0 with an
ionic strength of 0.15 M adjusted by sodium chloride), were added
pNPP then compounds (at eight different concentrations). The
reaction was initiated by addition of PTP1B (1–299) at a final con-
centration of 10 or 100 nM. The initial rate of PTPase-catalyzed
hydrolysis of pNPP was measured by following the absorbance
change at 405 nm. IC50 value was determined under fixed pNPP
concentration of 1 mM. All the assays were carried out in duplicate
or triplicate and the average results are presented.
Figure 2. Reversible and competitive monocyclic thiophene 2 as
PTP1B inhibitor.
C4,5-Di-aryl thiophene, that is, 16 (Ar = Ph), was some-
times the major product when aqueous THF was used.
16 did not show any inhibition against PTP1B.
The X-ray co-crystal structure of 11 and PTP1B was
successfully obtained (Fig. 4). It was found that the
binding mode of 11 was similar to that of compound 2
(Fig. 3). The structural information has provided guid-
ance for further optimization of this series of com-
pounds. It was envisioned that substitution off the
para position on the phenyl ring 7 would provide oppor-
tunities to capture interactions with Arg47 and Asp48
and thus enhance inhibition against PTP1B. In addition,
the possibility of achieving modest selectivity against
TCPTP by interacting with the residues in the YRD
motif (residues 46–48) has been reported before.
Figure 3. X-ray co-crystal structure of 2 and PTP1B (PBD code:
2HB1).
O
O
O
S
S
S
O
HO
i
iii
Arg47 and Asp48 are close to the catalytic site, interac-
tion of which has proven to be important in achieving
inhibitor selectivity over most of the PTPs as reported
by Zhang and co-workers.8 The para-position on the
O
O
O
R
R
R1
HO
O
O
O
OH
3a: R = H
3b: R = Me
3c: R = Br
˚
C5-phenyl ring is 4.7 A away from Asp48 (measured
4a: R = Br
5: R = Ph
2, 6 - 8
from the para-carbon of the phenyl to the acid moiety
of Asp48) as revealed by X-ray co-crystal structure of
11 and PTP1B (Fig. 4). It was predicted by molecular
modeling that a hydrogen bond would exist between
the Asp48 and a donor at the para-position of the phen-
yl ring. In addition, it is also possible to introduce inter-
actions with Arg47 that has been shown to significantly
improve potency against PTP1B.9
ii
Scheme 1. Reagents: (i) BrCH2CO2Me, K2CO3, DMF (99%); (ii)
Pd(PPh3)4, PhB(OH)2, KF, THF/H2O (78%); (iii) LiOH, THF/H2O
(90–99%).
(Ki = 3.2 lM, Table 2) led to another 5- to 6-fold
increase in potency.
To confirm this hypothesis, compound 17 (Table 3) was
synthesized according to Scheme 2. Indeed, a 10-fold in-
crease in potency was quickly realized (Table 3). The
existence of a hydrogen bond between the hydroxyl
group of 17 and Asp48 was confirmed by X-ray crystal-
lography (Fig. 5).
A panel of aryl motifs was then examined. It was shown
that a variety of aromatic groups could be accommodat-
ed at the C5-position (Scheme 2 and Table 2). It was
also noticed that selective Suzuki coupling could be
accomplished with non-aqueous solvent such as THF.

Z.-K. Wan et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4941–4945
4943
Table 2. The effect of C5-substitution on the inhibition against PTP1B
Table 3. The importance of a H-bond with Asp48
O
O
S
S
R
Ar
HO
HO
O
Br
O
Br
O
O
OH
Ar
OH
Kia (lM)
Compound
X
Kia (lM)
Compound
X
R
OH
10
Br
Br
18
17
Br
0.3
3.0
1.0
0.57
OMe
11
12
13
14
15
Br
Br
Br
Br
Br
3.2
1.4
4.0
3.3
5.0
18
19
20
Br
Br
Br
S
OH
N
N
H
a See Table 1 for enzymatic assay conditions.
O
a See Table 1 for enzymatic assay conditions.
O
O
S
O
Br
S
S
R
Ar
O
HO
HO
i, iii
O
Br
O
Br
O
Ar
or ii, iii
O
O
O
O
OH
OH
9
R = Br, Ar
10 - 15
16
Scheme 2. Reagents and condition: (i) Pd(PPh3)4, PhB(OH)2, KF,
THF; (ii) Pd(PPh3)4, PhB(OH)2, KF, THF/H2O or dioxane/H2O; (iii)
LiOH, THF/H2O, rt.
Figure 5. X-ray co-crystal structure of 17 with PTP1B (PBD code:
2H4G).
O
O
NH2
S
S
Br
O
O
i
O
O
Br
Br
O
O
O
O
9
21
ii
O
O
R
R
S
S
iii
HO
O
O
Br
O
Br
O
O
OH
O
22
23 - 36
Scheme 3. Reagents: (i) Pd(PPh3)4, ArB(OH)2, KF, THF/H2O
(70–78%); (ii) R0COCl, or R0SO2Cl or R0–N@C@O or R0OCOCl,
TEA, DCM (80–95%); (iii) LiOH, THF/H2O (80–99%).
Figure 4. A molecular surface on X-ray co-crystal structure of
compound 11 (PBD code: 2H4K). The surface is colored by lipophil-
icity with the more lipophilic regions shown in brown.

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Z.-K. Wan et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4941–4945
Table 4. The effect of the representative 5-para-substituted-phenyl
thiophenes on the inhibition against PTP1B
Other H-bond donors have been subsequently adopt-
ed to further optimize the hydrogen bond interactions
with Asp48 of PTP1B as well as balancing the overall
physical–chemical properties. An array of amines,
amides, sulfonamides, ureas, and carbamates were
thus synthesized (Scheme 3 and Table 4). The impor-
tance of a hydrogen-bond interaction between the
inhibitors and Asp48 was once again demonstrated.
For instance, elimination of hydrogen-bond donor
from 23 led to a more than 5-fold loss in potency
as observed in compound 24. Similarly, a 6-fold loss
was observed in compound 32 compared to parent
compound 31. In contrast, increasing the acidity of
the hydrogen-bond donor (NH group) strengthens
its H-bond interaction with Asp48 and thus improves
potency as in the case of amides (e.g., 25–27), sulfon-
amides (e.g., 30–31), carbamates (e.g., 33), and ureas
(e.g., 34). Consistently, the reversed amide 29 is 4-
fold less active than 28, presumably due to the loss
of the hydrogen bond between the amide NH and
Asp48.
O
R
S
HO
O
Br
O
OH
Compound
R
Kia (lM)
23
NH2
1.6
24
25
26
27
28
29
30
31
9.0
N
H
N
0.50
0.62
0.82
0.60
2.5
O
O
H
N
O
H
N
N
Incorporation of a tetramethylcyclohexyl group in 35
led to 6- to 7-fold increase in inhibition, likely resulting
from additional van der Waals interactions with the
relatively lipophilic region of the protein surface formed
by the hydrocarbon part of the side chain of Asp48, Val
49, and Met258 as suggested by molecular modeling. A
carboxylic group in 36 was designed to target electro-
static interaction with Arg47. Indeed, the presence of
this carboxylic group resulted in significant improve-
ment in inhibitory activity (compound 36, Ki = 0.14 lM,
Table 4).
O
O
H
N
O
N
H
H
N
0.74
0.20
S
O
O
CF3
CF3
H
N
These PTP1B inhibitors have shown good selectivity
against other PTPases except for the highly homolo-
gous T-cell PTPase (TCPTP). Representative examples
are shown in Table 5. In general, greater than 1000-
fold of selectivity was observed against LAR (com-
pounds 11, 24, 27, and 36, Table 5), and the selectiv-
ity against CD45 increases with the potency against
PTP1B. For instance, more than 236-fold selectivity
against CD45 was achieved with compound 36
(Ki(PTP1B) = 0.14 lM, Table 5), while compound 24
(Ki(PTP1B) = 0.3 lM, Table 5) only has a 40-fold selec-
S
O
O
Me
N
32
1.2
S
O
O
H
N
O
33
34
1.0
O
O
H
H
N
N
0.36
H
N
Table 5. Inhibitory activity of select compounds against various
protein tyrosine phosphatases*
35
0.25
0.14
Compound
PTP1ba
TCPTPb
CD45c
Kid (lM)
LARe
H
N
Ki (lM)
Ki (lM)
Ki (lM)
OH
36
24
27
11
36
1.6
1.0
64 (40)
55 (67)
26 (87)
33 (236)
>1000
320
O
O
0.82
0.30
0.14
0.68
0.32
0.18
a See Table 1 for enzymatic assay conditions.
>1000
>1000
a PTP1B (residues 1–299) concentration: 20 nM.
b TCPTP (residues 1–299) concentration: 20 nM.
Methylation of 17 in compound 18 reverted the potency
back to that of 11 (Table 3), which reassured the impor-
tance of the hydrogen bond. Consistent with modeling
prediction, a hydrogen-bond donor at the meta-position
in compounds 19 and 20 was less effective likely due to
less optimal directionality of hydrogen bond formation
(Table 3).
c CD45 (residues 484–1281, purchased from Biomol) concentration:
20 nM.
d Selectivity ratio is in parentheses.
e LAR (residues 1275–1613, purchased from Biomol) concentration:
20 nM.
* Enzymatic assay was carried out according to description on Table 1.

Z.-K. Wan et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4941–4945
4945
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tivity against CD45. Though it has been known that
interaction with the YRD motif could achieve modest
selectivity against TCPTP,8 there is no differentiation
in inhibitory potency against PTP1B and TCPTP with
this series of inhibitors.
In summary, a series of monocyclic thiophenes was dis-
covered to be competitive and reversible PTP1B inhibi-
tors with good potency and selectivity against other
PTPases with the exception of TCPTP. A key hydrogen
bond with Asp48 was proven to be pivotal in achieving
better inhibition against PTP1B. This interaction was
confirmed by X-ray co-crystal structures. Additional ef-
fort in further optimizing this monocyclic thiophene ser-
ies will be reported in due course.
Acknowledgments
The authors thank Drs. Steve Tam, James Tobin, and
Tarek Mansour for helpful discussions, Dr. Eddine
Saiah for reviewing this manuscript. We also thank
Drs. Nelson Huang, Walt Massefski, and Ying Ge,
Ms. Ning Pan, Ms. May Tam, and Ms. Eva Binnun
for technical support.
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