K. A. Josef et al. / Bioorg. Med. Chem. Lett. 11 (2001) 2615–2617
2617
The tripeptides in the series, 3l and 3m, are also potent
inhibitors of the calpain I enzyme, but more impor-
tantly, are the most potent inhibitors identified in our
MOLT-4 cell assay to date. Therefore, these compounds
have greater cell permeability than the dipeptide a-keto-
hydroxamates in our assay.12 This result is consistent
with a-ketoamide and a-ketoester tripeptides described
by Li et al.2 in a platelet membrane permeability assay.
S. I.; Ator, M. A.; Bihovsky, R. Bioorg. Med. Chem. Lett.
1997, 7, 539. (f) Conroy, J. L.; Seto, C. T. J. Org. Chem. 1998,
63, 2367.
3. (a) Crawford, C.; Mason, R. W.; Wickstrom, P.; Shaw, E.
Biochem. J. 1988, 253, 751. (b) McGowan, E. B.; Becker, E.;
Detwiler, T. C. Biochem. Biophys. Res. Commun. 1989, 158,
432. (c) Huang, Z.; McGowan, E. B.; Detwiler, T. C. J. Med.
Chem. 1992, 35, 2048. (d) Harris, A. L.; Gregory, J. S.; May-
cock, A. L.; Graybill, T. L.; Osifo, I. K.; Schmidt, S. L.; Dolle,
R. E. Bioorg. Med. Chem. Lett. 1995, 5, 393. (e) Chatterjee, S.;
Josef, K.; Wells, G.; Iqbal, M.; Bihovsky, R.; Mallamo, J. P.;
Ator, M. A.; Bozyczko-Coyne, D.; Mallya, S.; Senadhi, S.;
Siman, R. Bioorg. Med. Chem. Lett. 1996, 6, 1237.
4. After this manuscript was accepted for publication, a paper
with similar subject matter was published. Donkor, I. O.;
Zheng, X.; Han, J.; Lacy, C.; Miller, D. D. Bioorg. Med.
Chem. Lett. 2001, 11, 1753.
5. Mallamo, J. P.; Bihovsky, R.; Josef, K. A. WO 00/16767,
2000. Chem. Abstr. 2001, 132, 237376.
6. As described by Harbeson et al. in ref 2b.
We have described a series of novel peptide a-keto-
hydroxamates that are potent inhibitors of recombinant
human calpain I. The activity of this series compares
favorably to that of previously described a-ketoamides.2
However, substituent changes on the P0 hydroxamates
do not affect the potency as significantly as changes to
the P0 amide substituents. These a-ketohydroxamates
are highly membrane permeable as demonstrated in a
whole-cell MOLT-4 assay.
7. Dess, D. B.; Martin, J. C. J. Org. Chem. 1983, 48, 4156.
8. For reference, 3a: 1H NMR (CDCl3) d 9.55 (br s, 1H), 7.20
(m, 10H), 6.82 (d, 1H), 5.40 (m, 1H), 5.03 (s, 2H), 4.95 (br s,
1H), 4.14 (m, 1H), 3.81 (s, 3H), 3.24 (dd, 1H), 2.96 (dd, 1H),
1.52 (m, 2H), 1.39 (m, 1H), 0. 83 (m, 6H). H NMR spectra
were recorded on a GE QE300 Plus spectrometer at 300 MHz
using tetramethylsilane as internal standard.
9. Meyer, S. L.; Bozyczko-Coyne, D.; Mallya, S. K.; Spais, C.
M.; Bihovsky, R.; Kawooya, J. K.; Lang, D. M.; Scott, R. W.;
Siman, R. Biochem. J. 1996, 314, 511. In vitro calpain I enzy-
mic activity was measured in a 96-well format using a con-
tinuous fluorimetric assay. In this assay, hydrolysis of
succinyl-Leu-Tyr-4-methoxy-2-naphthylamine (Suc-Leu-Tyr-
MNA; Enzyme Systems Products, Dublin, CA, USA;
Km=0.4 mM) was monitored by a Fluoroskan II fluorimeter
(Labsystems, Helsinki, Finland). Enzyme activity was deter-
mined by measuring the calcium-dependant increase in fluor-
escence at 430nm ( lexcitation=340nm) of 0.2 ꢁ1 mM substrate
in 0.2 mL total volume of 50 mM Tris/HCl, pH 7.5, containing
50mM NaCl, 1 mM EDTA, 1 mM EGTA, 5 mM b-mercap-
toethanol, 2.5% DMSO, and 5 mM CaCl2.
Acknowledgements
1
The authors would like to thank John Mallamo, Sankar
Chatterjee, and Derek Dunn for their support, and
helpful discussions. We would also like to thank Mark
Ator, Donna Bozyczko-Coyne, Satish Mallya, Beth
Ann McKenna, Teresa M. O’Kane, and Shobha E.
Senadhi for performing the biological assays. We would
also like to thank SmithKline Beecham for partial sup-
port in this project.
References and Notes
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1998, 8, 1707. (b) Chatterjee, S. Drugs Future 1998, 23, 1217.
(c) Donkor, I. O. Curr. Med. Chem. 2000, 7, 1171.
10. Chatterjee, S.; Ator, M. A.; Bozyczko-Coyne, D.; Josef,
K.; Wells, G.; Tripathy, R.; Iqbal, M.; Bihovsky, R.; Senadhi,
S. E.; Mallya, S.; O’Kane, T. M.; McKenna, B. A.; Siman, R.;
Mallamo, J. P. J. Med. Chem. 1997, 40, 3820.
2. (a) Medhi, S. Trends Biol. Sci. 1991, 16, 50. (b) Harbeson,
S. L.; Abelleira, S. M.; Akiyama, A.; Barrett, R., III; Carroll,
R. M.; Straub, J. A.; Tkacz, J. N.; Wu, C.; Musso, G. F. J.
Med. Chem. 1994, 37, 2918. (c) Li, Z.; Patil, G.; Golubski,
Z. E.; Hori, H.; Tehrani, K.; Foreman, J. E.; Eveleth, D. D.;
Bartus, R. T.; Powers, J. C. J. Med. Chem. 1993, 36, 3742. (d)
Peet, N. P.; Kim, H. O.; Marquart, A. L.; Angelastro, M. R.;
Nieduzak, T. R.; White, J. N.; Friedrich, D.; Flynn, G. A.;
Webster, M. E.; Vaz, R. J.; Linnik, M. D.; Koehl, J. R.;
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Chem. Lett. 1999, 9, 2365. (e) Iqbal, M.; Messina, P. A.;
Freed, B.; Das, M.; Chatterjee, S.; Tripathy, R.; Tao, M.;
Josef, K. A.; Dembofsky, B.; Dunn, D.; Griffith, E.; Siman,
R.; Senadhi, S. E.; Biazzo, W.; Bozyczko-Coyne, D.; Meyer,
11. Chatterjee, S.; Gu, Z. Q.; Dunn, D.; Tao, M.; Josef, K.;
Tripathy, R.; Bihovsky, R.; Senadhi, S. E.; O’Kane, T. M.;
McKenna, B. A.; Mallya, S.; Ator, M. A.; Bozyczko-Coyne,
D.; Siman, R.; Mallamo, J. P. J. Med. Chem. 1998, 41, 2663.
12. One reviewer suggested there could be a correlation of the
MOLT-4 cell assay IC50 values and some biophysical property
such as lipophilicity. Calculation of the log P values gave a
range of 4.5 to 7.3ꢂ1 for compounds with IC50<1 mM.
Although there appears to be no correlation with these data at
this time, we gratefully thank the reviewer for bringing this to
our attention.