Caspase 3 (Ki = 0.23 nM); Caspase-8 (Ki = 0.92 nM); Caspase-7 (Ki = 1.6 nM); Caspase-10 (Ki = 12 nM); Caspase-1 (Ki = 18 nM); Caspase-6 (Ki = 31 nM); Caspase-9 (Ki = 60 nM); Caspase-4 (Ki = 132 nM); Caspase-5 (Ki = 205 nM); Caspase-2 (Ki = 1710 nM)

Ac-DEVD-CHO is a potent inhibitor of caspase-3 (Ki = 230 pM). In contrast, this aldehyde only slightly inhibits caspase-2 (Ki = 1.7 μM) and exhibits poor cleavage of the tetrapeptide substrate. With Ki values ranging from 1 to 300 nM, Ac-DEVD-CHO significantly inhibits Group III caspases[1]. Even when administered after the onset of ischemia, Ac-DEVD-CHO'scaspase-3inhibition significantly improves post-ischemic contractile recovery of stunned myocardium in the isolated working-heart rat model. Ac-DEVD-CHO'sprotectivemechanism(s) seem to operate independently of apoptosis. Ac-DEVD-CHO[2] did not inhibit troponin I cleavage.

Receiving Ac-DEVD-CHO at the time of MI causes a 61% decrease in the expression of activated caspase-3 in cardiomyocytes (p<0.05) and an 84% decrease in cardiomyocyte apoptosis in young animals. Caspase inhibition, however, had no impact on cardiomyocyte apoptosis or activated caspase-3 expression in the aging mice[4]. Ac-DEVD-CHO inhibited and/or postponed the development of photoreceptor cell damage in rats, as well as slows the disease's progression in rd gene-carrying mice, which typically experience retinal degeneration in their early years of life[2].

Studies with peptide-based and macromolecular inhibitors of the caspase family of cysteine proteases have helped to define a central role for these enzymes in inflammation and mammalian apoptosis. A clear interpretation of these studies has been compromised by an incomplete understanding of the selectivity of these molecules. Here we describe the selectivity of several peptide-based inhibitors and the coxpox serpin CrmA against 10 human caspases. The peptide aldehydes that were examined (Ac-WEHD-CHO, Ac-DEVD-CHO, Ac-YVAD-CHO, t-butoxycarbonyl-IETD-CHO, and t-butoxycarbonyl-AEVD-CHO) included several that contain the optimal tetrapeptide recognition motif for various caspases. These aldehydes display a wide range of selectivities and potencies against these enzymes, with dissociation constants ranging from 75 pM to >10 microM. The halomethyl ketone benzyloxycarbonyl-VAD fluoromethyl ketone is a broad specificity irreversible caspase inhibitor, with second-order inactivation rates that range from 2.9 x 10(2) M-1 s-1 for caspase-2 to 2.8 x 10(5) M-1 s-1 for caspase-1. The results obtained with peptide-based inhibitors are in accord with those predicted from the substrate specificity studies described earlier. The cowpox serpin CrmA is a potent (Ki < 20 nM) and selective inhibitor of Group I caspases (caspase-1, -4, and -5) and most Group III caspases (caspase-8, -9, and -10), suggesting that this virus facilitates infection through inhibition of both apoptosis and the host inflammatory response[1].

OCLs are incubated with RANKL and treated with 0.5 mM SIN for 24 hours, either with or without the particular caspase-3 inhibitor Ac-DEVD-CHO (10 μM). After the treatment, the cells are rinsed with PBS and stained for 15 min with 10 μM Hoechst 33258 dye. A fluorescent microscope is used to take pictures of the staining cells. By counting the number of cells with apoptotic nuclear condensation in each well, the differences are measured[4].

One hundred and two male mice are subjected to cecal ligation and puncture or sham operation. The animals are assigned into three equal groups (n=34) according to random number table: sham group, model group, and caspase-3 inhibitor (CI) group. Thirty minutes before CLP, Ac-DEVD-CHO (4 μg/g) is injected subcutaneously in CI group. The levels of blood urea nitrogen (BUN) and creatinine (Cr) are determined, and the concentrations of tumor necrosis factor-α (TNF-α), interleukins (IL-6 and IL-10) are measured by enzyme linked immunosorbent assay (ELISA), the renal cell apoptosis rate is determined by flow cytometry. The 4-day and 7-day survival rates of three groups of mice are observed[5].

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3 mg/kg; i.p.

C57Bl6 mice.

[1]. J Biol Chem . 1998 Dec 4;273(49):32608-13.

[2]. J Am Coll Cardiol . 2001 Dec;38(7):2063-70.

[3]. Mol Cell Biol . 2006 Nov;26(21):7880-91.

[4]. Cardiovasc Ther . 2013 Dec;31(6):e102-10.

[5] Acta Histochem. 2003, 36(4):263-270.

Ac-Asp-Glu-Val-Asp-H is a tetrapeptide consisting of two L-aspartic acid residues, an L-glutamyl residue and an L-valine residue with an acetyl group at the N-terminal and with the C-terminal carboxy group reduced to an aldehyde. It is an inhibitor of caspase-3/7. It has a role as a protease inhibitor.

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Studies with peptide-based and macromolecular inhibitors of the caspase family of cysteine proteases have helped to define a central role for these enzymes in inflammation and mammalian apoptosis. A clear interpretation of these studies has been compromised by an incomplete understanding of the selectivity of these molecules. Here we describe the selectivity of several peptide-based inhibitors and the coxpox serpin CrmA against 10 human caspases. The peptide aldehydes that were examined (Ac-WEHD-CHO, Ac-DEVD-CHO, Ac-YVAD-CHO, t-butoxycarbonyl-IETD-CHO, and t-butoxycarbonyl-AEVD-CHO) included several that contain the optimal tetrapeptide recognition motif for various caspases. These aldehydes display a wide range of selectivities and potencies against these enzymes, with dissociation constants ranging from 75 pM to >10 microM. The halomethyl ketone benzyloxycarbonyl-VAD fluoromethyl ketone is a broad specificity irreversible caspase inhibitor, with second-order inactivation rates that range from 2.9 x 10(2) M-1 s-1 for caspase-2 to 2.8 x 10(5) M-1 s-1 for caspase-1. The results obtained with peptide-based inhibitors are in accord with those predicted from the substrate specificity studies described earlier. The cowpox serpin CrmA is a potent (Ki < 20 nM) and selective inhibitor of Group I caspases (caspase-1, -4, and -5) and most Group III caspases (caspase-8, -9, and -10), suggesting that this virus facilitates infection through inhibition of both apoptosis and the host inflammatory response. [1]

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Objectives: The aim of this study was to investigate whether the caspase-3 inhibitor Ac-DEVD-CHO functionally improves stunned myocardium. Background: Degradation of troponin I contributes to the pathogenesis of myocardial stunning, whereas the role of apoptosis is unknown. Caspase-3 is an essential apoptotic protease that is specifically inhibited by Ac-DEVD-CHO. Methods: Isolated working hearts of rats were exposed to 30 min of low-flow ischemia, followed by 30 min of reperfusion. Ac-DEVD-CHO (0.1 to 1 micromol/l) was added 15 min before ischemia/reperfusion or 5 min before reperfusion. Cardiac output, external heart power, left ventricular (LV) developing pressure and contractility (dp/dt(max)) were measured. Apoptosis was assessed by TUNEL staining and internucleosomal deoxyribonucleic acid fragmentation. Caspase-3 processing and troponin I cleavage were determined by immunoblotting. Caspase-3 activity was measured using a fluorogenic substrate. Results: The addition of Ac-DEVD-CHO before ischemia/reperfusion or before reperfusion dose-dependently and significantly (p < 0.05) improved post-ischemic recovery of cardiac output, external heart power, LV developing pressure and dp/dt(max), compared with the vehicle (0.01% dimethyl sulfoxide). Ac-DEVD-CHO was similarly effective when given before reperfusion. Ac-DEVD-CHO blocked ischemia/reperfusion-induced caspase-3 activation, but cardiomyocyte apoptosis was unaffected. Troponin I cleavage was not inhibited by Ac-DEVD-CHO. Conclusions: Caspase-3 is activated in stunned myocardium. Inhibition of caspase-3 by Ac-DEVD-CHO significantly improves post-ischemic contractile recovery of stunned myocardium, even when given after the onset of ischemia. The mechanism(s) of protection by Ac-DEVD-CHO appear to be independent of apoptosis. Inhibition of caspase-3 is a novel therapeutic strategy to improve functional recovery of stunned myocardium.[2]

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The apoptosome, a heptameric complex of Apaf-1, cytochrome c, and caspase-9, has been considered indispensable for the activation of caspase-9 during apoptosis. By using a large panel of genetically modified murine embryonic fibroblasts, we show here that, in response to tumor necrosis factor (TNF), caspase-8 cleaves and activates caspase-9 in an apoptosome-independent manner. Interestingly, caspase-8-cleaved caspase-9 induced lysosomal membrane permeabilization but failed to activate the effector caspases whereas apoptosome-dependent activation of caspase-9 could trigger both events. Consistent with the ability of TNF to activate the intrinsic apoptosis pathway and the caspase-9-dependent lysosomal cell death pathway in parallel, their individual inhibition conferred only a modest delay in TNF-induced cell death whereas simultaneous inhibition of both pathways was required to achieve protection comparable to that observed in caspase-9-deficient cells. Taken together, the findings indicate that caspase-9 plays a dual role in cell death signaling, as an activator of effector caspases and lysosomal membrane permeabilization.[3]

C20H30N4O11

502.47

502.191

C, 47.81; H, 6.02; N, 11.15; O, 35.02

169332-60-9

644345

N-Acetyl-Asp-Glu-Val-Asp-al; Ac-Asp-Glu-Val-Asp-al; N-acetyl-L-alpha-aspartyl-L-alpha-glutamyl-L-valyl-L-aspart-1-al

Ac-DEVD-al

White to off-white solid powder

1.374g/cm3

1021.1ºC at 760mmHg

571.3ºC

0mmHg at 25°C

1.535

-2.6

7

11

16

35

843

4

O=C([C@]([H])(C([H])([H])C([H])([H])C(=O)O[H])N([H])C([C@]([H])(C([H])([H])C(=O)O[H])N([H])C(C([H])([H])[H])=O)=O)N([H])[C@]([H])(C(N([H])[C@]([H])(C([H])=O)C([H])([H])C(=O)O[H])=O)C([H])(C([H])([H])[H])C([H])([H])[H]

UMBVAPCONCILTL-MRHIQRDNSA-N

InChI=1S/C20H30N4O11/c1-9(2)17(20(35)22-11(8-25)6-15(29)30)24-18(33)12(4-5-14(27)28)23-19(34)13(7-16(31)32)21-10(3)26/h8-9,11-13,17H,4-7H2,1-3H3,(H,21,26)(H,22,35)(H,23,34)(H,24,33)(H,27,28)(H,29,30)(H,31,32)/t11-,12-,13-,17-/m0/s1

(4S)-4-[[(2S)-2-acetamido-3-carboxypropanoyl]amino]-5-[[(2S)-1-[[(2S)-1-carboxy-3-oxopropan-2-yl]amino]-3-methyl-1-oxobutan-2-yl]amino]-5-oxopentanoic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: 100 mg/mL (199.02 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.

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 (Please use freshly prepared in vivo formulations for optimal results.)