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2. Elchebly, M. et al. Science 1999, 283, 1544.
3. Klaman, L. D. et al. Mol. Cell. Biol. 2000, 20, 5479.
4. Zinker, B. A. et al. Proc. Natl. Acad. Sci. U.S.A. 2002, 99,
11357.
(i.e., CD45, PTPc, SHP2, etc.). Unfortunately, selectivi-
ty over the closely homologous TC-PTP was poor (2-
fold).
5. Bialy, L.; Waldmann, H. Angew. Chem., Int. Ed. 2005, 44,
3814.
6. Liu, G. Curr. Med. Chem. 2003, 10, 1407.
7. Blaskovich, M. A.; Kim, H.-O. Expert Opin. Ther. Patents
2002, 12, 871.
8. During the course of this work a report appeared
describing sulfamic acids as phosphatase inhibitors. San-
kar et al.; Preparation of sodium sulfamic acid salts as
inhibitors of human cytoplasmic protein tyrosine phos-
phatases for the treatment of wounds and of damaged
tissues; US 2002-368901.
9. Tetrahydroisoquinolines have been reported to have
cdc25B inhibitory activity Fritzen, E. L. et al. Bioorg.
Med. Chem. Lett. 2000, 10, 649.
Complete removal of the 3-position substituent from 10i
provided 12 which showed a 5-fold loss in potency
(Fig. 2). Additional simplification of 10i by removal of
the constraining TIQ ring gave the benzyl amine deriva-
tive 11 which also suffered from a loss in potency (6-fold).
Interestingly, the X-ray structure of 11 showed a reversal
in the binding mode of the compound. In the case of 11,
the sulfamic acid residue of the propionic acid side chain
was shown to bind in the P0 pocket, while the benzyl
amine unit was extended into the P + 1 pocket.
High-throughput screening of the P&GP corporate
repository identified the sulfamic acid functionality as
a potential pTyr mimetic. Incorporation of a sulfamic
acid moiety at the 7-position of the 1,2,3,4-tetrahydro-
isoquinoline scaffold provided compounds which
showed promising initial PTP1B inhibitory properties.
X-ray crystallography provided a guide to inhibitor de-
sign by determining that side chains extending from the
2-position of the natural enantiomer pointed toward the
second aryl phosphate binding region (P + 1 pocket).
An initial survey of side chains extending from the 2-po-
sition of the TIQ scaffold indicated that a three-atom
linker between the TIQ ring system and the distal aryl
ring was preferred. A second investigation provided an
order of magnitude increase in potency by incorporating
an acidic functionality in the 3-position of the aryl ring.
10. Asante-Appiah, E. et al. J. Biol. Chem. 2001, 276, 26036.
11. Klopfenstein, S. R. et al. US 2004-0167183 A1.
12. Klopfenstein, S. R. et al. WO 2004-074256 A1.
13. Chaudhary, A.; Girgis, M.; Parshad, M.; Hu, B.; Har, D.;
ˇ
Repic, O.; Blacklock, T. J. Tetrahedron Lett. 2003, 44,
5543.
14. All compounds were fully characterized by 1H and 13C
NMR, HPLC, MS, and combustion analysis.
15. Phosphatase activity and kinetics: recombinant phospha-
tases (nM) were diluted in assay buffer (150 mM NaCl, 0–
0.1% BSA, 5 mM DTT, and 50 mM Tris–HCl) (pH 7) or
10 mM Na/Tris acetate (pH 6) and incubated with 10 lM
DiFMUP (Molecular Probes). After 15 min, the fluores-
cence increase was measured on a Victor V plate reader
(Wallac). Inhibitors were pre-incubated with enzyme for
10 min before substrate addition. Kinetic measurements
were measured using 3–5 concentrations of inhibitor
around the IC50 over a range of substrate concentrations
(3–500 lM). Data were used to determine Km/Ki/compet-
itive inhibition using GraphPad Prism software.
16. Crystal structures: preparation of recombinant PTP1B
catalytic domain (1–322) was essentially as described
earlier (see Ref. 21). Crystallization of apo-PTP1B for
soaking experiments, co-crystallization of PTP1B bound
to various inhibitors, X-ray data collection, and structure
solution are described in the Supplementary material.
17. All structures referred to in this article were deposited with
(3), 2F6Y (8g), 2F6V (8k), 2F6Z (10i), 2F71 (10j), 2F70
(11), and 2F6W (13-Supplementary material).
Simplification of the TIQ ring system by removal of the
3-carboxamide functionality reduced potency as did
removal of the constraining saturated 6-membered ring.
X-ray crystallography of these improved compounds re-
vealed the nature of the ligand–protein interactions and
should provide a stepping stone for further potency
enhancements in this series of compounds.
Supplementary material
Supplementary data associated with this article can be
18. Sarmiento, M.; Puius, Y. A.; Vetter, S. W.; Keng, Y.-F.;
Wu, L.; Zhao, Y.; Lawrence, D. S.; Almo, S. C.; Zhang,
Z.-Y. Biochemistry 2000, 39, 8171.
19. Shen, K.; Keng, Y.-F.; Wu, L.; Guo, X.-L.; Lawrence, D.
S.; Zhang, Z.-Y. J. Biol. Chem. 2001, 276, 47311.
20. Manuscript in preparation.
References and notes
21. Puius, Y. A.; Zhao, Y.; Sullivan, M.; Lawrence, D. S.;
Almo, S. C.; Zhang, Z.-Y. Proc. Natl. Acad. Sci. U.S.A.
1997, 94, 13420.
1. Cheng, A.; Dube, N.; Gu, F.; Tremblay, M. L. Eur. J.
Biochem. 2002, 269, 1050.