Development and Application of Reversible and Irreversible Covalent Probes for Human and Mouse Cathepsin-K Activity Detection, Revealing Nuclear Activity

Gourab Dey, Reut Sinai-Turyansky, Evalyn Yakobovich, Emmanuelle Merquiol, Jure Loboda, Nikhila Sridharan, Yael Houri-Haddad, David Polak, Simon Yona, Dusan Turk, Ori Wald, Galia Blum*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

Cathepsin-K (CTSK) is an osteoclast-secreted cysteine protease that efficiently cleaves extracellular matrices and promotes bone homeostasis and remodeling, making it an excellent therapeutic target. Detection of CTSK activity in complex biological samples using tailored tools such as activity-based probes (ABPs) will aid tremendously in drug development. Here, potent and selective CTSK probes are designed and created, comparing irreversible and reversible covalent ABPs with improved recognition components and electrophiles. The newly developed CTSK ABPs precisely detect active CTSK in mouse and human cells and tissues, from diseased and healthy states such as inflamed tooth implants, osteoclasts, and lung samples, indicating changes in CTSK's activity in the pathological samples. These probes are used to study how acidic pH stimulates mature CTSK activation, specifically, its transition from pro-form to mature form. Furthermore, this study reveals for the first time, why intact cells and cell lysate exhibit diverse CTSK activity while having equal levels of mature CTSK enzyme. Interestingly, these tools enabled the discovery of active CTSK in human osteoclast nuclei and in the nucleoli. Altogether, these novel probes are excellent research tools and can be applied in vivo to examine CTSK activity and inhibition in diverse diseases without immunogenicity hazards.

Original languageEnglish
Article number2401518
JournalAdvanced Science
Volume11
Issue number38
DOIs
StatePublished - 16 Oct 2024

Bibliographical note

Publisher Copyright:
© 2024 The Author(s). Advanced Science published by Wiley-VCH GmbH.

Keywords

  • activity-based probes
  • cathepsin K
  • covalent probes
  • imaging protease activity
  • proteases

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