Aziridine analogs of [[trans-(epoxysuccinyl)-L-leucyl]amino]-4-guanidinobutane (E-64) as inhibitors of cysteine proteases.
Aziridine derivatives of E-64 have been synthesized, and their characterization against the cysteine proteases cathepsin B, cathepsin L, and papain is reported. The inhibition was found to be strongly pH-dependent, with maximum activity observed at pH 4, indicating that the protonated aziridinium ion form of the inhibitor is the more reactive form. At low pH, the peptide aziridine HO-(L)Az-Leu-NH-iAm inactivated papain with a second-order rate constant, kinac/Ki, of 7.0 x 10(4) M-1 s-1, a value very close to that observed with E-64 or with the corresponding epoxysuccinyl analog HO-(L)Eps-Leu-NH-iAm. This demonstrates that with the correct peptide sequence, aziridine analogs of E-64 can be good irreversible inhibitors of cysteine proteases. Substitution of the epoxysuccinyl moiety by an aziridine does not affect the specificity of inhibition against the three proteases used in this study. The D-diastereomer is the preferred (by 10-fold) diastereomer for the inhibition of cysteine proteases. The reactivity of both diastereomers of iBuNH-Az-LeuPro-OH against cathepsin B was also found to be much lower than that of iBuNH-(L)Eps-LeuPro-OH, which is a potent selective inhibitor of cathepsin B. These differences are attributed mainly to the presence of the protonated aziridine ring, which can modify the binding mode of aziridine analogs at the active site of cysteine proteases.
Martichonok
V
V
Department of Chemistry, University of Toronto, Lash Miller Laboratories, Canada.
Plouffe
C
C
Storer
A C
AC
Ménard
R
R
Jones
J B
JB
D000818:Animals; D001388:Aziridines / Q000737:chemistry / Q000494:pharmacology*; D002401:Cathepsin B / Q000037:antagonists & inhibitors; D056668:Cathepsin L; D002403:Cathepsins / Q000037:antagonists & inhibitors; D003546:Cysteine Endopeptidases; D015853:Cysteine Proteinase Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D010450:Endopeptidases; D006863:Hydrogen-Ion Concentration; D007930:Leucine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology; D010206:Papain / Q000037:antagonists & inhibitors; D051381:Rats; D013237:Stereoisomerism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D001388:Aziridines; D015853:Cysteine Proteinase Inhibitors; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D002401:Cathepsin B; D056668:Cathepsin L; C534280:Ctsl protein, rat; D010206:Papain; D007930:Leucine; C024974:E 64