Identification of bidentate salicylic acid inhibitors of PTP1B

Article abstract of DOI:10.1021/acsmedchemlett.5b00171

PTP1B is a master regulator in the insulin and leptin metabolic pathways. Hyper-activated PTP1B results in insulin resistance and is viewed as a key factor in the onset of type II diabetes and obesity. Moreover, inhibition of PTP1B expression in cancer cells dramatically inhibits cell growth in vitro and in vivo. Herein, we report the computationally guided optimization of a salicylic acid-based PTP1B inhibitor 6, identifying new and more potent bidentate PTP1B inhibitors, such as 20h, which exhibited a > 4-fold improvement in activity. In CHO-IR cells, 20f, 20h, and 20j suppressed PTP1B activity and restored insulin receptor phosphorylation levels. Notably, 20f, which displayed a 5-fold selectivity for PTP1B over the closely related PTPσ protein, showed no inhibition of PTP-LAR, PRL2 A/S, MKPX, or papain. Finally, 20i and 20j displayed nanomolar inhibition of PTPσ, representing interesting lead compounds for further investigation.

Full text of DOI:10.1021/acsmedchemlett.5b00171

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Potent Tethered Bidentate Salicylic Acid Inhibitors of PTP1B
Sina Haftchenary,† ∥ [a] Andriana O. Jouk,† [a] Isabelle Aubry,† [b] Andrew M. Lewis,† [a] Melissa
Landry,[b] Daniel Ball,[a] Andrew E. Shouksmith,[a] Catherine V. Collins,[a] Michel L. Tremblay,* [b]
Patrick T. Gunning*[a]
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[a] Department of Chemical and Physical Sciences, University of Toronto Mississauga, Mississauga, ON L5L 1C6,
Canada [b] McGill Goodman Cancer Research Centre and Department of Biochemistry, McGill University, Montreal,
QC H3G 1Y6, Canada.
*Financial support was provided by NSERC (SH, DB), CFI (PTG), ORF (PTG), Canada Research Chair (PTG) and in part supported
by Canadian Institute of Health Research Operating Grant, MOP-62887 (to MLT).
†These authors contributed equally to this work
Keywords PTP1B, PTPσ, diabetes, insulin receptor, smallꢀmolecule inhibitor, salicylic acid, bidentate inhibitors
Supporting Information Placeholder
ABSTRACT: PTP1B is a master regulator in the insulin and leptin metabolic pathways. Hyper-activated PTP1B results in
insulin resistance and is viewed as a key factor in the onset of type II diabetes and obesity. Moreover, inhibition of PTP1B
expression in cancer cells dramatically inhibits cell growth in vitro and in vivo. Herein, we report the computationally
guided optimization of a salicylic acid-based PTP1B inhibitor 6, identifying new and more potent bidentate PTP1B inhibi-
tors, such as 20h, which exhibited a > 4-fold improvement in activity. In CHO-IR cells, 20f, 20h, and 20j suppressed
PTP1B activity and restored insulin receptor phosphorylation levels. Notably, 20f, which displayed a 5-fold selectivity for
PTP1B over the closely related PTPσ protein, showed no inhibition of PTP-LAR, PRL2 A/S, MKPX or papain. Finally, 20i
and 20j displayed nM inhibition of PTPσ, representing interesting lead compounds for further investigation.
Phosphorylation of regulatory and signaling proteins
within the cell is closely monitored by the coordinated
action of protein tyrosine kinases (PTKs) and the
superfamily of protein tyrosine phosphatases (PTPs). The
PTKs and PTPs are responsible for the addition and removal
of phosphate groups from specific tyrosine (Y) residues on
proteins, respectively. This partnership is responsible for the
regulation of cellular communication and homeostasis.1
Dysregulation of phosphorylation events are directly related
to malignancy states including, but not limited to; diabetes,2
cancer,3-5 neurological,6 metabolic7-9 and autoimmune
disorders.10 Historically, drug discovery programs have
predominately focused on identifying PTK inhibitors since
the prevailing dogma was that phosphatases were not as
attractive drug targets. PTPs were overlooked, and
considered housekeeping enzymes with broad specificity, as
compared to their PTK counterparts. However,
advancements in the past two decades in the field of
molecular biology uncovered a more profound role for PTPs
in cell regulation, growth, and the onset of human
disease.6,7,11 The importance of PTPs in signal transduction
and cellular homeostasis has led to a flurry of discoveries
which linked their aberrant activities to numerous
pathologies.12-15 These discoveries have validated PTPs as
interesting therapeutic targets.
adipose tissues, and implicated in the onset of Type II
Diabetes (T2D). PTP1B overexpression leads to persistent
dephosphorylation of the insulin receptor (IR), stimulating
the insulin-resistant phenotype in T2D and obesity. The
pivotal biological role of PTP1B has led to numerous
inhibitor programmes.8-9,16-19 Despite the identification of
many potent compounds, a PTP1B-inhibiting drug has yet to
reach the clinic. The most common strategy for inhibiting
PTP1B, or other PTPs, has been to target the active site.
However, deriving selectivity has been problematic, given
that PTP active sites have, in some cases, 100% sequence
homology.9,12,20 As a result, recent inhibitor designs have
focused on the simultaneous targeting of multiple
subpockets proximal to the active site.21-23 These domains
The most studied of the PTP enzymes is PTP1B (gene
code: PTPN1), which is overexpressed in muscle, liver, and
Figure 1. Chemical structures of previously described PTP1B
inhibitors.
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Optimization for Ligand Docking (GOLD) software (Figure 2).
Results indicated that the salicylic acid was binding at the
cyclohexylbenzyl group was binding within large
hydrophobic pocket in the active site (Ile219, Gly259, and
Val49, Figure 2), and the quinoline group was forming π-π
interactions with Phe182, as well as H-bonding between the
quinoline nitrogen and the peptide backbone. Since the N-
methyl group was not involved in binding and was oriented
towards the second pY binding site, it was selected for
attaching the second salicylic acid substituent.
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Figure 2. Docking image of compound 6 in the active site of
PTP1B (PDB: 1NWL). Protein: hydrophobic residues (blue) and
hydrophilic residues (red); Compounds: nitrogen (blue), oxygen
(red) and sulphur (gold).
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A
are less conserved amongst the PTP isoforms and are seen
as being a route to achieving enhanced target selectivity. In
the case of PTP1B, where a proximal phosphotyrosine (pY)-
binding site is located adjacent to the pY-binding active site,
inhibitors have been developed which possess two pY
groups or similar bivalent combinations of pY and more
drug-like pY bioisosteres (compounds 1-5, Figure 1).24
However, bivalent pY-containing inhibitors that exhibit high
PTP1B selectivity often exhibit poor pharmacological
profiles, likely due to the charged pY-mimicking
B
Figure 3. Comparative docking studies of 20f (cyan) and inhibitor
SP7343-SP7964 (green) bound to PTP1B (PDB: 1NWL). Protein:
hydrophobic residues (blue) and hydrophilic residues (red);
Compounds: nitrogen (blue), oxygen (red) and sulphur (gold).
Figure 4. (A) Structures of inhibitors 20a-20j; (B) preliminary
high-throughput kinetic enzyme activity screen for compounds
6 and 20a-j at 10 µM against phosphatases PTP1B, TC-PTP,
PTPσ (D1D2), PTP-LAR (D1D2), PRL2 A/S and MKPX, with the
protease papain present as a negative control. Experiments
were performed in duplicate. Graph bars represent percent
inhibition (100 = complete inhibition, 0 = no inhibition).
functionality and peptidic character.12
In previous efforts to target the pY binding site of
PTP1B, scaffolds have incorporated either a salicylic or
benzoic acid25 as a cell-permeable pY mimetic.26-28 Salicylic
acid-based inhibitors of PTP1B were reported, with IC50
values of 3-8 µM, and no inhibition of PTP-LAR, PRL2 A/S,
MKPX, or papain was observed (6, Figure 2).2 However, the
compounds displayed equipotent inhibition of another
clinically important phosphatase, PTPσ. PTPσ is a potential
target for the development of novel therapies for spinal
cord injury and multiple neurodegenerative diseases.29-31
Moreover, PTPσ inhibition has recently been identified as a
target for improving memory loss.32 In this study, it was
hypothesized that incorporation of a second carboxylate
group to compound 6 might provide a second pY mimetic
for more potent and selective targeting of PTP1B. It was
reasoned that such a scaffold might improve- cell
penetrating properties more than current pY mimetics
utilized in bidentate PTP1B inhibitors.22
Copper-catalyzed click chemistry methodology was
selected to couple the two components via a triazole linker
of varying alkyl chain length.33-34 As opposed to a flexible
and rather hydrophobic hydrocarbon linker, the flat triazole
provides additional polarity, rigidity and has been deemed
the ideal linker for development of bridged inhibitors in
recent literature.33-34 To study the binding mode of the
proposed di-salicylic acids, comparative in silico docking
analysis was conducted with PTP1B and a proposed
bidentate inhibitor 20f, superimposed with known
bidentate PTP1B inhibitor, SP7343-SP7964 (Ki= 4.1 mM,
Figure 3). Docking revealed that 20f bound analogously to
SP7343-SP7964, with both salicylic acids interacting with
the respective pY-binding sites. The interactions between
the cyclohexylbenzyl group of 6 and the hydrophobic pocket
of the active site were retained, despite the addition of the
second salicylate group. Next, the linker length between
To identify the site of carboxylate binding on 6, in silico
docking studies were performed with PTP1B using Genetic
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both salicylic acid groups was explored in silico with analysis
suggesting to incorporate linkers ranging from one to five
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carbons in length (Supporting information). The N-
sulfonamide aryl group, R, was either a quinoline, as
previously used in 6, or a tolyl group, as previous
investigations with this substituent had indicated activity
against PTP1B.2
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The focused library was screened for activity at 10 µM
using a previously described DiFMUP assay against the non-
receptor tyrosine phosphatases PTP1B and TC-PTP,
receptor-like tyrosine phosphatases PTPσ and PTP-LAR, the
dual specificity phosphatases PRL2 A/S (active mutant) and
MKPX, as well as a human cysteine-based protease papain
(used here as a control for oxidation). Compounds 20f-j
inhibited TC-PTP, PTP1B and PTPσ enzymatic activity by over
90% compared to the DMSO control. Compounds 20f-j
showed modest increases in activity as compared to 6
(Supporting information). Negligible effects were observed
against PRL2 A/S, PTP-LAR (D1D2), MKPX, or CD45D1D2.
MKPX and PRL2 A/S are dual specificity phosphatases that
recognize phosphorylated S and T residues as well as pY.
The selectivity may be the result of unfavourable binding
interactions with the catalytic pockets of PTP-LAR (D1D2),
MKPX, CD45D1D2, and PRL2 A/S. Tolyl inhibitors 20a-e were
significantly less active and less selective compared to the
quinoline analogues. This was likely due to poor water
solubility under the assay conditions and/or less favourable
binding interactions of the tolyl with the PTP1B active site
as compared to the quinoline derivatives (Comparative
docking studies provided in the supporting information).
Quinoline compounds 20f–j inhibited the receptor-like PTPs,
such as PTPσ, more potently than the non-receptor PTPs
like PTP1B. Thus, 20f-j were further evaluated to quantify
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Figure 5. Dose-dependent curves for inhibitors 20f-j. Inhibitors
were tested against an array of phosphatases. Individual plots
overlay inhibitor-specific data for the three phosphatases.
compared to PTP-LAR (D1D2), MKPX and CD45D1D2 (Figure
5). The limited discrimination of the compounds between
PTP1B and TC-PTP was anticipated, given a > 74% homology
in their catalytic domains and adjacent pockets. This
demonstrated for the first time a scaffold, which displayed
potency and selectivity for PTP1B over PTPσ. Moreover, this
study identified potent PTPσ inhibitors, 20i and 20j.
However, given there is no cell-based assay for assessing
the enzymatic activity of PTPσ, the inhibition profiles of 20i
and 20j against PTPσ could not be explored further at this
time.
The cytotoxicities of PTP1B inhibitors 20f, 20h, and 20j
were determined in CHO cells, which express the human
insulin receptor (CHO-IR). A Titer-Blue Cell viability assay
(Promega) showed no toxicity at 0.78 to 50 µM
concentrations
(Supporting
information).2
Having
established that the compounds were non-toxic, 20f, 20h,
and 20j were subjected to Western blot analysis against
PTP1B in CHO cells.2 Briefly, the insulin-signaling cascade
depends on the phosphorylation state of the insulin
receptor (IR). In this signal transduction pathway, insulin
binds to the IR on the extracellular plasma membrane and
promotes autophosphorylation of the IR (pIR). Since pIR is a
direct substrate for PTP1B, the IR phosphorylation state is
largely dependent on the activity levels of PTP1B within the
cytoplasm. Thus, monitoring pIR levels is a useful reporter
for PTP1B activity. CHO cells were incubated with increasing
concentrations of inhibitor in serum-free media, then
stimulated with insulin for 15 min. Western blot analysis
determined the relative levels of pIR (Figure 6). Compounds
20f, 20h, and 20j dose-dependently increased the pIR signal,
relative to the DMSO control (Figure 6A). Although
vanadate induced a strong response (50 μM), it suffers as a
Table 1. IC50 (µM) values for 6 and 20f-j against PTP1B and
PTPσ (Sigma) in dose-dependent enzymatic assays.
relative levels of PTP1B and PTPσ inhibition.
IC50 values were determined for each compound (Table
1, with representative curves shown in Figure 5). IC50 values
for 20f-j ranged from 0.6 - 6.1 μM. Two inhibitors, 20f and
20h, possessed > 2-fold improved potency for PTP1B over 6
(20f; IC50 = 3.6 μM, 20h; IC50 = 1.7 μM and 6; IC50 = 6.2 μM),
with 20h also displaying a 4-fold selectivity for PTP1B over
PTPσ. Notably, 20i was identified as the first nM PTPσ
inhibitor, also with approximately 4-fold selectivity over
PTP1B (PTP1B IC50 = 2.3 μM, PTPσ IC50 = 0.6 μM).
Compounds 20i and 20j were screened against a panel
of PTPs employing the DiFMUP assay. The dose-dependent
enzyme activity rate curves of 20i and 20j showed
significantly greater activity against PTP1B and PTPσ,
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therapeutic due to poor metabolic stability and low PTP1B inhibition and > 4-fold selectivity over PTPσ. Finally,
bioavailability.35 In addition, the compounds displayed
improved activity in cells relative to 6 (Supporting
information). The densitometry graph showed 20f, 20h, and
20j had a significant effect on the pIR/IR levels compared to
the DMSO control (Figure 6B).2
20i and 20j displayed nM inhibition for PTPσ and thus
represent interesting leads for further investigation against
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Figure 7. Michaelis–Menten rate data as a function of sub-
strate concentration for 20f and 6. Curves represent the hy-
drolysis of DiFMUP and subsequent fluorescent detection at
450 nm (excitation at 358 nm). The bottom curves show the
Lineweaver–Burke transformation detailing competitive inhi-
bition for 20f and 6.
this novel target.
Supporting Information Available
Figure 6. (A) The dose dependent response of compounds 20f,
20h, and 20j upon the phosphorylated insulin receptor was de-
termined by western blot. Cells were either treated with com-
pounds (5 – 50 μM), DMSO as a solvent control, 50 μM vanadate
(VN) or left unstimulated (NI); (B) Densitometry graph showing
the response of 20f, 20h, and 20j at 25 μM on the levels of pIR
and IR vs. a DMSO control, *P<0.05. Values correspond to mean
±standard error of the mean (n=4). Statistical analysis was per-
formed with two-tailed, unpaired, Student’s t test.
Information on compound synthesis, characterization, de-
tailed results, and experimental procedures can be found in
the supporting information. This material is available free
of charge via the Internet at http://pubs.acs.org
AUTHOR INFORMATION
Corresponding Author
*E-mail: patrick.gunning@utoronto.ca. Telephone: (905)
828-5354. Fax: (905) 569-5425.
*E-mail: michel.tremblay@mcgill.ca. Telephone: (514) 398-
7290, ext.
Finally, comparative Michaelis–Menten kinetic studies
were performed for 6 and 20f against PTP1B (Figure 7).
Compound 20f showed an approximate 3-fold greater
competitive inhibition (Ki = 1.83 ± 0.39 µM) compared to 6
(Ki = 5.66 ± 2.11 µM). In comparison to 6, the bidentate
approach yielded a more direct, competitive inhibitor of the
active site. In general, in the field of PTP inhibition, mixed
inhibitors tend to display a higher degree of toxicity,
attributed to their promiscuous mode of action.12,36-37 Thus,
we have optimized a mixed inhibitor, 6, to produce 20f a
competitive and presumed active site binder of PTP1B.
Notes
The authors declare no competing financial interest.
Present Addresses
∥ Center for the Science of Therapeutics, Broad Institute
75 Ames Street, Room 3022C, Cambridge, MA 02142, USA.
References
(1) Bauman, A. L.; Scott, J. D. Kinase- and phosphatase-
anchoring proteins: harnessing the dynamic duo. Nat. Cell
Biol. 2002, 8, 203-206.
(2) Haftchenary, S.; Ball, D. P.; Aubry, I.; Landry, M.;
Shahani, V. M.; Fletcher, S.; Page, B. D. G.; Jouk, A. O.;
Tremblay, M. L.; Gunning, P. T. Identification of a potent
salicylic acid-based inhibitor of tyrosine phosphatase
PTP1B. MedChemComm 2013, 4, 987-992.
(3) Haftchenary, S.; Avadisian, M.; Gunning, P. T.
Inhibiting aberrant Stat3 function with molecular
therapeutics: a progress report. Anticancer Drugs 2011, 22,
115-127.
In summary, identifying selective and potent small
molecule inhibitors of the PTP family of proteins (> 200
members) remains a daunting task due to the high degree
of sequence homology. Inspired by current approaches
utilizing bidentate scaffolds, a family of di-salicylic acid
derivatives, based on PTP1B inhibitor 6, were developed to
achieve more potent and selective inhibition of PTP1B. A
number of compounds were identified that displayed
improved potency (> 5-fold) and selectivity for PTP1B over
PTPσ (4-fold) relative to 6. Compounds 20f-j did not inhibit
other PTP family members such as PTP-LAR (D1D2). In
particular, 20h represents an interesting lead for further
investigation against PTP1B, as it demonstrated whole-cell
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(4) Östman, A.; Hellberg, C.; Böhmer, F. D. Protein-
tyrosine phosphatases and cancer. Nat. Rev. Cancer 2006,
6, 307-320.
(21) Guo, X.-L.; Shen, K.; Wang, F.; Lawrence, D. S.; Zhang,
Z.-Y. Probing the Molecular Basis for Potent and Selective
Protein-tyrosine Phosphatase 1B Inhibition. J. Biol. Chem.
2002, 277, 41014-41022.
(22) Low, J. L.; Chai, C. L. L.; Yao, S. Q. Bidentate
Inhibitors of Protein Tyrosine Phosphatases. Antioxid.
Redox Signaling 2014, 20, 2225-2250.
(23) Srinivasan, R.; Uttamchandani, M.; Yao, S. Q. Rapid
assembly and in situ screening of bidentate inhibitors of
protein tyrosine phosphatases. Org. Lett. 2006, 8, 713-716.
(24) Bialy, L.; Waldmann, H. Inhibitors of Protein
Tyrosine Phosphatases: Next-Generation Drugs? Angew.
Chem., Int. Ed. 2005, 44, 3814-3839.
(25) Haftchenary, S.; Luchman, H. A.; Jouk, A. O.; Veloso,
A. J.; Page, B. D. G.; Cheng, X. R.; Dawson, S. S.; Grinshtein,
N.; Shahani, V. M.; Kerman, K.; Kaplan, D. R.; Griffin, C.;
Aman, A. M.; Al-awar, R.; Weiss, S.; Gunning, P. T. Potent
Targeting of the STAT3 Protein in Brain Cancer Stem Cells:
A Promising Route for Treating Glioblastoma. ACS Med.
Chem. Lett. 2013, 4, 1102-1107.
(26) Fletcher, S.; Singh, J.; Zhang, X.; Yue, P. B.; Page, B. D.
G.; Sharmeen, S.; Shahani, V. M.; Zhao, W.; Schimmer, A.
D.; Turkson, J.; Gunning, P. T. Disruption of
Transcriptionally Active Stat3 Dimers with Non-
phosphorylated, Salicylic Acid-Based Small Molecules:
Potent in vitro and Tumor Cell Activities. ChemBioChem
2009, 10, 1959-1964.
(27) Page, B. D. G.; Fletcher, S.; Yue, P. B.; Li, Z. H.; Zhang,
X. L.; Sharmeen, S.; Datti, A.; Wrana, J. L.; Trudel, S.;
Schimmer, A. D.; Turkson, J.; Gunning, P. T. Identification
of a non-phosphorylated, cell permeable, small molecule
ligand for the Stat3 SH2 domain. Bioorg. Med. Chem. Lett.
2011, 21, 5605-5609.
(28) Zhang, X.; He, Y.; Liu, S.; Yu, Z.; Jiang, Z.-X.; Yang, Z.;
Dong, Y.; Nabinger, S. C.; Wu, L.; Gunawan, A. M.; Wang,
L.; Chan, R. J.; Zhang, Z.-Y. A salicylic acid-based small
molecule inhibitor for the oncogenic Src homology-2
domain containing protein tyrosine phosphatase-2 (SHP2).
J. Med. Chem. 2010, 53, 2482-2493.
1
2
3
4
5
6
7
8
(5) Hoekstra, E.; Peppelenbosch, M. P.; Fuhler, G. M. The
role of protein tyrosine phosphatases in colorectal cancer.
Biochim. Biophys. Acta 2012, 1826, 179-188.
(6) Muise, A. M.; Walters, T.; Wine, E.; Griffiths, A. M.;
Turner, D.; Duerr, R. H.; Regueiro, M. D.; Ngan, B. Y.; Xu,
W.; Sherman, P. M.; Silverberg, M. S.; Rotin, D. Protein-
tyrosine phosphatase sigma is associated with ulcerative
colitis. Curr. Biol. 2007, 17, 1212-1218.
(7) Kennedy, B. P.; Ramachandran, C. Protein tyrosine
phosphatase-1B in diabetes. Biochem. Pharmacol. 2000, 60,
877-883.
(8) Combs, A. P. Recent Advances in the Discovery of
Competitive Protein Tyrosine Phosphatase 1B Inhibitors for
the Treatment of Diabetes, Obesity, and Cancer. J. Med.
Chem. 2010, 53, 2333-2344.
(9) Johnson, T. O.; Ermolieff, J.; Jirousek, M. R. Protein
tyrosine phosphatase 1B inhibitors for diabetes. Nat. Rev.
Drug Discovery 2002, 1, 696-709.
(10) Yip, S. C.; Saha, S.; Chernoff, J. PTP1B: a double agent
in metabolism and oncogenesis. Trends Biochem. Sci. 2010,
35, 442-449.
(11) Freiss, G.; Vignon, F. Protein tyrosine phosphatases
and breast cancer. Crit. Rev. Oncol. Hemat. 2004, 52, 9-17.
(12) Zhang, S.; Zhang, Z. Y. PTP1B as a drug target: recent
developments in PTP1B inhibitor discovery. Drug Discovery
Today 2007, 12, 373-381.
(13) Elchebly, M.; Payette, P.; Michaliszyn, E.; Cromlish,
W.; Collins, S.; Loy, A. L.; Normandin, D.; Cheng, A.;
Himms-Hagen, J.; Chan, C. C.; Ramachandran, C.; Gresser,
M. J.; Tremblay, M. L.; Kennedy, B. P. Increased insulin
sensitivity and obesity resistance in mice lacking the
protein tyrosine phosphatase-1B gene. Science 1999, 283,
1544-1548.
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(14) Friedman, J. M. Obesity in the new millennium.
Nature 2000, 404, 632-634.
(15) Ahima, R. S.; Prabakaran, D.; Mantzoros, C.; Qu, D.;
Lowell, B. Role of leptin in the neuroendocrine response to
fasting. Nature 1996, 382, 250-252.
(29) Shen, Y.; Tenney, A. P.; Busch, S. A.; Horn, K. P.;
Cuascut, F. X.; Liu, K.; He, Z.; Silver, J.; Flanagan, J. G.
(16) Szczepankiewicz, B. G.; Liu, G.; Hajduk, P. J.
PTPsigma is
a
receptor for chondroitin sulfate
Discovery of
a
potent, selective protein tyrosine
proteoglycan, an inhibitor of neural regeneration. Science
2009, 326, 592-596.
phosphatase 1B inhibitor using a linked-fragment strategy.
J. Am. Chem. Soc. 2003, 125, 4087-4096.
(17) Stuible, M.; Zhao, L.; Aubry, I.; Schmidt-Arras, D.
Cellular inhibition of protein tyrosine phosphatase 1B by
uncharged thioxothiazolidinone derivatives. ChemBioChem
2007, 8, 179-186.
(18) Wilson, D. P.; Wan, Z. K.; Xu, W. X.; Kirincich, S. J.
Structure-based optimization of protein tyrosine
phosphatase 1B inhibitors: from the active site to the
second phosphotyrosine binding site. J. Med. Chem. 2007,
50, 4681-4698.
(19) Popov, D. Novel protein tyrosine phosphatase 1B
inhibitors: interaction requirements for improved
intracellular efficacy in type 2 diabetes mellitus and obesity
control. Biochem. Biophys. Res. Commun. 2011, 410, 377-381.
(20) Iversen, L. F.; Møller, K. B.; Pedersen, A. K.; Peters, G.
H. Structure determination of T cell protein-tyrosine
phosphatase. J. Biol. Chem. 2002, 277, 19982-19990.
(30) Coles, C. H.; Shen, Y. J.; Tenney, A. P.; Siebold, C.;
Sutton, G. C.; Lu, W. X.; Gallagher, J. T.; Jones, E. Y.;
Flanagan, J. G.; Aricescu, A. R. Proteoglycan-Specific
Molecular Switch for RPTP sigma Clustering and Neuronal
Extension. Science 2011, 332, 484-488.
(31) Thompson, K. M.; Uetani, N.; Manitt, C.; Elchebly, M.;
Tremblay, M. L.; Kennedy, T. E. Receptor protein tyrosine
phosphatase sigma inhibits axonal regeneration and the
rate of axon extension. Mol. Cell. Neurosci. 2003, 23, 681-
692.
(32) Horn, K. E.; Xu, B.; Gobert, D.; Hamam, B. N.;
Thompson, K. M.; Wu, C. L.; Bouchard, J. F.; Uetani, N.;
Racine, R. J.; Tremblay, M. L.; Ruthazer, E. S.; Chapman, C.
A.; Kennedy, T. E. Receptor protein tyrosine phosphatase
sigma regulates synapse structure, function and plasticity. J.
Neurochem. 2012, 122, 147-161.
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(33) Manetsch, R.; Krasiński, A.; Radić, Z.; Raushel, J.;
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Taylor, P.; Sharpless, K. B.; Kolb, H. C. In Situ Click
Chemistry: Enzyme Inhibitors Made to Their Own
Specifications. J. Am. Chem. Soc. 2004, 126, 12809-12818.
(34) Agalave, S. G.; Maujan, S. R.; Pore, V. S. Click
chemistry: 1, 2, 3-triazoles as pharmacophores. Chem. -
Asian J. 2011, 6, 2696-2718.
(35) Huyer, G.; Liu, S.; Kelly, J.; Moffat, J.; Payette, P.
Mechanism of inhibition of protein-tyrosine phosphatases
by vanadate and pervanadate. J. Biol. Chem. 1997, 272, 843-
851.
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(36) Cornish-Bowden, A. Why is uncompetitive inhibition
so rare?: A possible explanation, with implications for the
design of drugs and pesticides. FEBS lett. 1986, 203, 1-4.
(37) Shehzad, S. The potential effect of vanadium
compounds on glucose-6-phosphatase. Biosci. Horiz. 2013,
6, 1-11.
For Table of Contents Use Only.
Potent Tethered Bidentate Salicylic Acid Inhibitors of PTP1B
Sina Haftchenary,† ∥ [a] Andriana O. Jouk,† [a] Isabelle Aubry,† [b] Andrew M. Lewis,† [a] Melissa
Landry,[b] Daniel Ball,[a] Andrew E. Shouksmith,[a] Catherine V. Collins,[a] Michel L. Tremblay,* [b]
Patrick T. Gunning*[a]
Secondary pY
Binding Pocket
PTP1B
Primary pY
Binding Pocket
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Potent Tethered Bidentate Salicylic Acid Inhibitors of PTP1B
Sina Haftchenary,† ∥ [a] Andriana O. Jouk,† [a] Isabelle Aubry,† [b] Andrew M. Lewis,† [a] Melissa
Landry,[b] Daniel Ball,[a] Andrew E. Shouksmith,[a] Catherine V. Collins,[a] Michel L. Tremblay,* [b]
Patrick T. Gunning*[a]
The phosphatase family of proteins plays a fundamental role in cell biology. The dysregulation of their
activity has been highly implicated in many disorders including diabetes, cancer and neurodegenerative
disease. Protein tyrosine phosphatase 1B (PTP1B) has been identified as an important druggable target
for the treatment of cancer, with much work in the field highlighting a pressing need for potent and seꢀ
lective drugꢀlike inhibitors of PTP1B. To address this we designed and synthesized a small library of
molecules utilizing a clickꢀchemistry approach to selectively target this important diseaseꢀrelated proꢀ
tein.
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