strates.10,11 Recently reported inhibitors incorporate this type
of functionality at the P1′ position, and a structural examina-
tion of a series of these inhibitors defines a general trend
for caspase-3 of increasing inhibition with increasing surface
area.10 However, this trend was seen using inhibitors with
very similar P1′ groups and varied P4-P1 substituents. Thus,
to provide a clearer understanding of the P1′ position and
its importance to caspase inhibitor design, we report the
systematic substitution of the P1′ position and the effects of
the resulting peptides upon inhibition of caspase-3 and
caspase-7.
access to the desired compounds was envisioned through the
coupling of various â-amino-γ-keto acids to an appropriately
protected DEV tripeptide, followed by deprotection.
The crystal structures of caspase-312 and caspase-713 bound
to aldehyde 1 were used to guide selection of appropriate
P1′ substituents. The length of functionality in the P1′
position was limited to groups that would best interact with
the S1′ pocket, shown in Figure 2. There are slight differ-
To systematically probe the effect of P1′ substitution on
inhibition, we designed a series of inhibitors of the general
structure shown in Figure 1 that hold the optimal Group II
Figure 1. General ketone-containing Ac-DEVD peptide and its
interaction with the various substrate-binding sites (S4-S1′) of
caspase-3 and -7. By holding the P4-P1 amino acids constant, the
effect of substitutions in the P1′ position can explicitly be probed
and compared to aldehyde 1.
Figure 2. X-ray crystal structures of peptide 1 bound to caspase-3
and -7.12,13 The S1′ sites of these enzymes have differences in both
size and shape. Blue represents surface-exposed regions.
caspase recognition sequence (DEVD) constant while chang-
ing only the P1′ position. In this design, a ketone replaces
the amide carbonyl of the scissile bond; the active site
cysteine of caspases is known to attack such ketones to
reversibly form thio-hemiacetals.11 A range of R groups can
then project from the ketone into the S1′ pocket. Convenient
ences in the size and shape of the S1′ site between caspase-3
and caspase-7; caspase-3 appears to have a more shallow
yet broad S1′ pocket, while caspase-7 has a deeper and nar-
rower S1′ site. It was envisioned that these differences might
allow for some isozyme selectivity in the caspase inhibition.
(7) (a) Talanian, R. V.; Quinlan, C.; Trautz, S.; Hackett, M. C.;
Mankovich, J. A.; Banach, D.; Ghayur, T.; Brady, K. D.; Wong, W. W. J.
Biol. Chem. 1997, 272, 9677-9682. (b) Chereau, D.; Kodandapani, L.;
Tomaselli, K. J.; Spada, A. P.; Wu, J. C. Biochemistry 2003, 42, 4151-
4160.
To synthesize the designed inhibitors, a common protected
peptide trimer was coupled to various â-amino-γ-keto acids.
The protected peptide DEV trimer 5 could easily be obtained
in good yield and high purity using the FAAST solution-
phase peptide synthesis technique (Scheme 1).14 The requisite
â-amino-γ-keto acid modules 6a-o were synthesized either
through a method previously developed in our laboratory
based on Grignard addition to protected L-homoserine15 or
through a method based on the cuprate displacement of a
(8) (a) Han, Y.; Giroux, A.; Grimm, E. L.; Aspiotis, R.; Francoeur, S.;
Bayly, C. I.; Mckay, D. J.; Roy, S.; Xanthoudakis, S.; Vaillancourt, J. P.;
Rasper, D. M.; Tam, J.; Tawa, P.; Thornberry, N. A.; Paterson, E. P.; Garcia-
Calvo, M.; Becker, J. W.; Rotonda, J.; Nicholson, D. W.; Zamboni, R. J.
Bioorg. Med. Chem. Lett. 2004, 14, 805-808. (b) Han, Y.; Giroux, A.;
Colucci, J.; Bayly, C. I.; Mckay, D. J.; Roy, S.; Xanthoudakis, S.;
Vaillancourt, J.; Rasper, D. M.; Tam, J.; Tawa, P.; Nicholson, D. W.;
Zamboni, R. J. Bioorg. Med. Chem. Lett. 2005, 15, 1173-1180.
(9) For the effect of substitution in the P1′ position on the inhibition of
caspase-1, see: (a) Mjalli, A. M. M.; Chapman, K. T.; MacCoss, M.;
Thornberry, N. A. Bioorg. Med. Chem. Lett. 1993, 3, 2689-2692. (b) Mjalli,
A. M. M.; Chapman, K. T.; MacCoss, M. Bioorg. Med. Chem. Lett. 1993,
3, 2693-2698.
(10) Becker, J. W.; Rotonda, J.; Soisson, S. M.; Aspiotis, R.; Bayly, C.;
Francoeur, S.; Gallant, M.; Garcia-Calvo, M.; Giroux, A.; Grimm, E.; Han,
Y.; McKay, D.; Nicholson, D. W.; Peterson, E.; Renaud, J.; Roy, S.;
Thornberry, N.; Zamboni, R. J. Med. Chem. 2004, 47, 2466-2474.
(11) Brady, K. D.; Giegel, D. A.; Grinnell, C.; Lunney, E.; Talanian, R.
V.; Wong, W.; Walker, N. Bioorg. Med. Chem. 1999, 7, 621-631.
(12) Rotonda, J.; Nicholson, D. W.; Fazil, K. M.; Gallant, M.; Gareau,
Y.; Labelle, M.; Peterson, E. P.; Rasper, D. M.; Ruel, R.; Vaillancourt, J.
P.; Thornberry, N. A.; Becker, J. W. Nat. Struct. Biol. 1996, 3, 619-625.
(13) Wei, Y.; Fox, T.; Chambers, S. P.; Sintchak, J.; Coll, J. T.; Golec,
J. M.; Swenson, L.; Wilson, K. P. Charifson, P. S. Chem. Biol. 2000, 7,
423-432.
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Org. Lett., Vol. 7, No. 16, 2005