Inhibitory Effect of di- and Tripeptidyl Aldehydes on Calpains and Cathepsins

Abstract

Eight different di- and tripeptidyl aldehyde derivatives, each having at its C-terminus an aldehyde analog of L-norleucine, L-methionine, or L-phenylalanine with a preceding L-leucine residue, were synthesized and tested for their inhibitory effects on several serine and cysteine endopeptidases. These compounds showed almost no inhibition of trypsin, and only weak inhibition of alpha-chymotrypsin and cathepsin H, while they exhibited marked inhibition of cathepsin B less than calpain II congruent to calpain I less than cathepsin L, being stronger in this order. The mode of inhibition of these cysteine proteinases was competitive for the peptide substrate used and inhibitor constants (Ki) were calculated from the Dixon plot. The best inhibitors found were: 4-phenyl-butyryl-Leu-Met-H for calpain I (Ki, 36 nM) and calpain II (Ki, 50 nM); acetyl-Leu-Leu-nLeu-H for cathepsin L (Ki, 0.5 nM); acetyl-Leu-Leu-Met-H for cathepsin B (Ki, 100 nM).

Cysteine protease inhibition prevents mitochondrial apoptosis-inducing factor (AIF) release

“…Indeed, all reagents precluding the AIF mitochondrial processing (including proteasome inhibitors) inhibit two major families of cystein proteases: calpains and cathepsins. [16][17][18] However, as calpains are inactive enzymes in the Atr calcium-depleted system (Figure 1c), our results point toward a cathepsin implication in Ca 2 þ -independent AIF processing. Thus, we looked for an individual cathepsin protease integrating the results presented above.…”

Inhibition of rat liver cathepsins B and L by the peptide aldehyde benzyloxycarbonyl-leucyl-leucyl-leucinal and its analogues

“…Furthermore, several di-and tripeptidyl aldehyde inhibitors were synthesized and their inhibitory effects investigated toward cysteine proteases including cathepsins B and L [1]. Among these, the most potent inhibitors were reported to be acetyl-Leu-Val-lysinal toward cathepsins B (IC 50 , 4 nM) [22] and acetyl-Leu-Leunorleucinal toward cathepsin L (K i , 0.5 nM) [23], respectively. Thus, several of the present peptide aldehydes appear to possess inhibitory potencies toward cathepsins B and L comparable with or somewhat weaker than those inhibitors.…”

In vivo degradation of N-myc in neuroblastoma cells is mediated by the 26S proteasome

“…Similar experiments were performed on three separate occasions and resulted in a mean increase in N-myc steady-state level of 4.5-, 4.6-and 4.7-fold. ALLnL not only inhibits the proteasome (Orlowski et al, 1993;Rock et al, 1994), but is also an inhibitor of calpain (Wang et al, 1994) and cathepsin (Sasaki et al, 1990). To con®rm that the results we obtained were due speci®cally to proteasome inhibition, we also examined the e ect of lactacystin (LC, 10 mM), a speci®c inhibitor of the proteasome (Fenteany et al, 1995), on the steady-state level of N-myc.…”

“…However, because ALLnL has been shown to inhibit neutral cysteine proteases such as calpain (Wang, 1990;Figuereido-Pereira et al, 1994), as well as acid cysteine proteases such as cathepsin (Sasaki et al, 1990;Wang et al, 1994), we examined additional membrane-permeant inhibitors. Cells were exposed for 4 h to either calpeptin (10 mg/ml), a slightly more potent calpain inhibitor than ALLnL, E64D (35 mM), a lipid-soluble cathepsin and calpain inhibitor or to CBZ (40 mM), a mixed cathepsin, calpain and proteasome inhibitor, similar in speci®city to ALLnL.…”