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R. A. Smith et al. / Bioorg. Med. Chem. Lett. 11 (2001) 2951–2954
8. Marquis, R. W.; Yamashita, D. S.; Ru, Y.; LoCastro,
Acknowledgements
S. M.; Oh, H.-J.; Erhard, K. F.; DesJarlais, R. L.; Head, M. S.;
Smith, W. W.; Zhao, B.; Janson, C. A.; Abdel-Meguid, S. S.;
Tomaszek, T. A.; Levy, M. A.; Veber, D. F. J. Med. Chem.
1998, 41, 3563.
9. Marquis, R. W.; Ru, Y.; Zeng, J.; Trout, R. E. L.; LoCas-
tro, S. M.; Gribble, A. D.; Witherington, J.; Fenwick, A. E.;
Garnier, B.; Tomaszek, T.; Tew, D.; Hemling, M. E.; Quinn,
C. J.; Smith, W. W.; Zhao, B.; McQueney, M. S.; Janson,
C. A.; D’Alessio, K.; Veber, D. F. J. Med. Chem. 2001, 44,
725.
We would like to thank Ulrike Moehler (student intern)
for assistance with the parallel syntheses, Anthony
Paiva and Lee Huang for LC–MS analyses, and Sook-
hee Ha for computationalstudies. We are aslo grateful
to Suresh Katti, Brian Dixon, David Campbell, and
Robert Schoenleber for helpful discussions.
10. Falgueyret, J.-P.; Oballa, R. M.; Okamoto, O.; Weso-
lowski, G.; Aubin, Y.; Rydzewski, R. M.; Prasit, P.; Riendeau,
D.; Rodan, S. B.; Percival, M. D. J. Med. Chem. 2001, 44, 94.
11. Katunuma, N.; Matsui, A.; Inubushi, T.; Murata, E.;
Kakegawa, H.; Ohba, Y.; Turk, D.; Turk, V.; Tada, Y.; Asao,
T. Biochem. Biophys. Res. Commun. 2000, 267, 850.
References and Notes
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15. All data reported herein reflect samples having high purity
(>95%) and confirmed identity (LC/UV/MS); IC50 values
reflect samples with further characterization (1H NMR, MS).
The cathepsin K assay conditions (pH 5.5, 100 mM sodium
acetate, 5 mM EDTA, 20 mM cysteine) were the same as
described elsewhere.2 Test compounds in 50% DMSO were
added in a dosed out, duplicate manner spanning a 2 log range
(2.5% finalDMSO concentration). Celavage of an added
substrate, benzyloxycarbonyl-Leu-Arg-7-amido-4-methyl-cou-
marin HCl(3 mM, 1/2 Km), was initiated by adding recombi-
nantly expressed cathepsin K at a concentration of 31 ng/mL.
This 100 mL reaction was incubated with shaking for 45 min at
room temperature before fluorescence (excitation/emission
360/460 nm) was measured with a Perkin–Elmer CytoFluor II
plate reader. As a control measure, several a-bromomethyl
ketones, as used in the synthesis of analogues of 2, were tested
in the cathepsin K assay; no inhibition was observed in these
experiments at up to 5 mM concentration.
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16. The backbone chiralcarbons of 2 and other analogues in
this article have (S,S) chirality. Other stereoisomers investi-
gated were found to be significantly less effective inhibitors.