Activity-Based Imaging of Acid Ceramidase in Living Cells

Yadira F. Ordóñez, José Luís Abad, Mazen Aseeri, Josefina Casas, Virginie Garcia, Mireia Casasampere, Edward H. Schuchman, Thierry Levade, Antonio Delgado, Gemma Triola, Gemma Fabrias

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

Abstract

Acid ceramidase (AC) hydrolyzes ceramides into sphingoid bases and fatty acids. The enzyme is overexpressed in several types of cancer and Alzheimer's disease, and its genetic defect causes different incurable disorders. The availability of a method for the specific visualization of catalytically active AC in intracellular compartments is crucial for diagnosis and follow-up of therapeutic strategies in diseases linked to altered AC activity. This work was undertaken to develop activity-based probes for the detection of AC. Several analogues of the AC inhibitor SABRAC were synthesized and found to act as very potent (two-digit nM range) irreversible AC inhibitors by reaction with the active site Cys143. Detection of active AC in cell-free systems was achieved either by using fluorescent SABRAC analogues or by click chemistry with an azide-substituted analogue. The compound affording the best features allowed the unprecedented labeling of active AC in living cells.

Original languageEnglish
Pages (from-to)7736-7742
Number of pages7
JournalJournal of the American Chemical Society
Volume141
Issue number19
DOIs
StatePublished - 15 May 2019

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