Chemical tools for analysis and manipulation of ceramidases in human disease
Final Report Abstract
Sphingolipids are important components of eukaryotic plasma membranes, especially in the form of sphingomyelin or glycosphingolipids. Ceramide is a sphingolipid that occurs in cells as an intermediate in the biosynthesis or degradation of complex glycosphingolipids or of the abundant sphingomyelin. Ceramide also has a well-documented role as a signalling molecule that can induce programmed cell death or inflammatory responses or suppress multidrug resistance and other pathways that limit the efficacy of pharmacological cancer treatments. Interestingly, different ceramide species have also been correlated with coronary heart disease, diabetes and metabolic syndrome and there is published evidence that the concentration and composition of plasma ceramides are more predictive of coronary heart disease than cholesterol. The various ceramidases play a key role in the ceramide concentration in different cellular compartments and also outside the cell. In addition to the lysosomal acid ceramidase (ASAH1) and the neutral ceramidase (ASAH2), there are also three alkaline ceramidases (ACER 1-3). The individual roles of the individual ceramidases are only partially known and specific inhibitors are lacking. In the past research project, various fluorescent ceramides were synthesized with the aim of obtaining selective probes for the different ceramidases. It was hoped that these probes would make it easier to investigate the activities of the individual ceramidases and thus develop new and specific inhibitors for the individual ceramidases. In fact, we succeeded in synthesizing a series of double-labeled “FRET ceramides” in the course of the project. Some of these ceramides showed specific cleavage by individual ceramidases and could therefore be used as substrates in vitro as well as in cellulo experiments. For the first time, the probes allowed comparatively simple high-throughput screening to be realized. Indeed, we succeeded in identifying potent inhibitors of ACER3 and neutral ceramidase (nCDase). No inhibitors were previously known for ACER3 in particular. Based on the screening hits, we were able to develop improved derivatives with increased potency and biological half-life and demonstrate their specific inhibition in living cells. In addition to these published results, we synthesized well over a hundred derivatives of the various inhibitors with the aim of improving them. This process was supported by molecular docking. The evaluation of the different compounds is still ongoing.
Publications
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Discovery and Mechanism of Action of Small Molecule Inhibitors of Ceramidases. Angewandte Chemie International Edition, 61(2).
Healey, Robert D.; Saied, Essa M.; Cong, Xiaojing; Karsai, Gergely; Gabellier, Ludovic; Saint‐Paul, Julie; Del Nero, Elise; Jeannot, Sylvain; Drapeau, Marion; Fontanel, Simon; Maurel, Damien; Basu, Shibom; Leyrat, Cedric; Golebiowski, Jérôme; Bossis, Guillaume; Bechara, Cherine; Hornemann, Thorsten; Arenz, Christoph & Granier, Sebastien
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Neutral ceramidase-active site inhibitor chemotypes and binding modes. Bioorganic Chemistry, 139; 106747.
Coant, Nicolas; Bickel, John D.; Rahaim, Ronald; Otsuka, Yuka; Choi, Yong-Mi; Xu, Ruijuan; Simoes, Michael; Cariello, Chris; Mao, Cungui; Saied, Essa M.; Arenz, Christoph; Spicer, Timothy P.; Bannister, Thomas D.; Tonge, Peter J.; Airola, Michael V.; Scampavia, Louis; Hannun, Yusuf A.; Rizzo, Robert C. & Haley, John D.
