Aza-Peptide Michael Acceptors:  A New Class of Inhibitors Specific for Caspases and Other Clan CD Cysteine Proteases

Ekici, Özlem Doğan; Götz, Marion G.; James, Karen E.; Li, Zhao Zhao; Rukamp, Brian J; Asgian, Juliana L.; Caffrey, Conor R.; Hansell, Elizabeth; Dvořák, Jan; McKerrow, James H.; Potempa, Jan; Travis, James C.; Mikolajczyk, Jowita; Salvesen, Guy S.; Powers, James C.

doi:10.1021/jm049938j

Cited by 79 publications

(82 citation statements)

References 25 publications

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“…The legumain substrate was purchased from AnaSpec Biological Inc (Fremont, CA, USA). Synthetic legumain inhibitor RR-11a 37,38 was synthesized by Wuxi Apptec Inc. (Beijing, China). The legumain-specific substrate Ala-Ala-Asn (AAN), TAT, the AAN-TAT conjugate at the fourth lysine of the TAT sequence and the FITC-labelled AAN-TAT were custom synthesized by Ziyu peptide Inc (Shanghai, China).…”

Section: Methodsmentioning

confidence: 99%

Legumain protease-activated TAT-liposome cargo for targeting tumours and their microenvironment

et al. 2014

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Specific targeting and cellular internalization are key properties for carriers of antitumor therapeutic agents. Here, we develop a drug carrier through the attachment of substrate of endoprotease legumain, alanine-alanine-asparagine (AAN), to cell-penetrating peptides (TAT, trans-activating factor). The addition of the AAN moiety to the fourth lysine in the TAT creates a branched peptide moiety, which leads to a decrease in the transmembrane transport capacity of TAT by 72.65%. Legumain efficiently catalyses the release of TAT-liposome from the AAN-TAT-liposome and thereby recovers the penetrating capacity of TAT. Doxorubicin carried by the AAN-TAT-liposome led to an increase in the tumoricidal effect of doxorubicin and a reduction in its systemic adverse effects in comparison with doxorubicin carried by a control delivery system. Thus, the specific targeting and high efficiency of this delivery platform offers a novel approach to limit the toxicity of anticancer agents as well as increasing their efficacy in cancer therapy.

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Section: Methodsmentioning

confidence: 99%

Legumain protease-activated TAT-liposome cargo for targeting tumours and their microenvironment

et al. 2014

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“…Using our solid phase method, we synthesized a series of ABPs containing the previously reported epoxide and Michael acceptor reactive groups. 9,11 We also reasoned that since the peptide acyloxymethyl ketones (AOMK) were effective probes of caspases that an AOMK warhead could be used to make a potentially interesting new class of aza-peptide electrophiles. 5 We therefore synthesized aza peptide ABPs containing either dimethyl or dichloro benzoic acid leaving groups to examine the effects of reactivity of the electrophile on target labeling.…”

Section: Resultsmentioning

confidence: 99%

“…Recent work by Powers and co-workers has demonstrated the utility of aza-peptide electrophiles as potent and selective inhibitors of clan CD cysteine proteases. [9][10][11][12] In particular, compounds containing an aza-asparate (aza-Asp) in the P1 position and either epoxide or Michael acceptor warheads were shown to be potent caspase inhibitors with very low crossreactivity towards other cysteine proteases. 10,11 We recently developed a solid-phase synthesis methodology that permits facile introduction of multiple electrophilic warheads onto the aza-Asp peptide backbone.…”

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confidence: 99%

“…However, when these aza-Asp ABPs were incubated with proteomes containing high lysosomal protease activity, both probes efficiently labeled legumain and cathepsin B. Reactivity with the clan CA protease cathepsin B was particularly unexpected as previous in vitro kinetic studies using purified enzymes identified these scaffolds as selective for inhibition of clan CD proteases. 10,11 In addition, the novel class of aza-Asp AOMKs, while significantly less potent as ABPs, could be engineered to label multiple caspases by introduction of an optimized peptide binding sequence. These ABPs were more selective caspase labels, presumably as a result of their slow labeling kinetics.…”

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confidence: 99%

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Specificity of aza-peptide electrophile activity-based probes of caspases

et al. 2006

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Activity-Based Probes (ABPs) are small molecules that form stable covalent bonds with active enzymes thereby allowing detection and quantification of their activities in complex proteomes. A number of ABPs that target proteolytic enzymes have been designed based on well-characterized mechanism-based inhibitors. We describe here the evaluation of a novel series of ABPs based on the aza-aspartate inhibitory scaffold. Previous in vitro kinetic studies showed that this scaffold has a high degree of selectivity for the caspases, clan CD cysteine proteases activated during apoptotic cell death. Aza-aspartate ABPs containing either an epoxide or Michael acceptor reactive group were potent labels of executioner caspases in apoptotic cell extracts. However they were also effective labels of the clan CD protease legumain and showed unexpected crossreactivity with the clan CA protease cathepsin B. Interestingly, related aza peptides containing an acyloxymethyl ketone reactive group were relatively weak but highly selective labels of caspases. Thus azapeptide electrophiles are valuable new ABPs for both detection of a broad range of cysteine protease activities and for selective targeting of caspases. This study also highlights the importance of confirming the specificity of covalent protease inhibitors in crude proteomes using reagents such as the ABPs described here.

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“…Hence these are recognized as novel therapeutic targets for CNS diseases in which cell death occurs mainly by an apoptosis mechanism [55]. The presence of near absolute specificity for residue at P1 is a distinctive feature of this family of enzymes and so in an effort to obtain greater potency and selectivity a variety of azapeptide Michael acceptors with P1 Asp residue were synthesized as potential inhibitors for caspases [56,57].…”

Section: Various Azapeptide Scaffolds As Peptidomimeticsmentioning

confidence: 99%

A Review on Azapeptides: The Promising Peptidomimetics

Begum¹,

Sujatha²,

Prasad³

et al. 2017

Asian J. Chem.

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Peptidomimetics, the mimics of natural peptides are considered as promising therapeutics with potential applications in modern medicine. Among the various structural variants of peptidomimetics that have been designed and synthesized, the azapeptides serve as the interesting peptide backbone modifications owing to their diversified biological and pharmacokinetic properties. The aim of this review is to highlight the significance of azapeptide in enhancement of stability and bioavailability, represent the various scaffolds of azapeptides as peptidomimetics, notify their significance as cysteine and serine protease inhibitors, provide an insight on the various biologically significant azapeptides and emphasize the main features on the conventional to modified synthetic strategies of azapeptides.

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Aza-Peptide Michael Acceptors: A New Class of Inhibitors Specific for Caspases and Other Clan CD Cysteine Proteases

Cited by 79 publications

References 25 publications

Legumain protease-activated TAT-liposome cargo for targeting tumours and their microenvironment

Legumain protease-activated TAT-liposome cargo for targeting tumours and their microenvironment

Specificity of aza-peptide electrophile activity-based probes of caspases

A Review on Azapeptides: The Promising Peptidomimetics

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