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Inhibition of acid sphingomyelinase by ambroxol prevents SARS-CoV-2 entry into epithelial cells

Carpinteiro et al., Journal of Biological Chemistry, doi:10.1016/j.jbc.2021.100701
Jan 2021  
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In Vitro study showing that ambroxol (a metabolite of bromhexine) inhibits SARS-CoV-2 infection.
Bromhexine efficacy may vary depending on the degree of TMPRSS-dependent fusion for different variants Peacock, Willett.
4 preclinical studies support the efficacy of bromhexine for COVID-19:
Carpinteiro et al., 31 Jan 2021, peer-reviewed, 10 authors.
In Vitro studies are an important part of preclinical research, however results may be very different in vivo.
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Abstract: RESEARCH ARTICLE Inhibition of acid sphingomyelinase by ambroxol prevents SARS-CoV-2 entry into epithelial cells Received for publication, January 10, 2021, and in revised form, April 15, 2021 Published, Papers in Press, April 23, 2021, Alexander Carpinteiro1,2, Barbara Gripp3, Markus Hoffmann4,5 , Stefan Pöhlmann4,5 , Nicolas Hoertel6, Michael J. Edwards7, Markus Kamler8 , Johannes Kornhuber9, Katrin Anne Becker1, and Erich Gulbins1,7, * From the 1Institute of Molecular Biology and 2Department of Hematology, University Hospital Essen, University of DuisburgEssen, Essen, Germany; 3Zentrum für Seelische Gesundheit des Kindes- und Jugendalters, Sana-Klinikum Remscheid GmbH, Remscheid, Germany; 4Infection Biology Unit, German Primate Center – Leibniz Institute for Primate Research, Göttingen, Germany; 5Faculty of Biology and Psychology, University of Göttingen, Göttingen, Germany; 6AP-HP.Centre-Université de Paris, Hôpital Corentin-Celton, Département de Psychiatrie, Issy-les-Moulineaux, and Université de Paris, INSERM, Institut de Psychiatrie et Neurosciences de Paris, UMR_S1266, and Faculté de Santé, UFR de Médecine, Paris, France; 7Department of Surgery, Medical School, University of Cincinnati, Cincinnati, Ohio, USA; 8Department of Thoracic and Cardiovascular Surgery, Division of Thoracic Transplantation, University Hospital Essen, Essen, Germany; and 9Department of Psychiatry and Psychotherapy, University Hospital of Erlangen, Friedrich-Alexander University Erlangen-Nürnberg (FAU), Erlangen, Germany Edited by Craig Cameron The acid sphingomyelinase/ceramide system has been shown to be important for cellular infection with at least some viruses, for instance, rhinovirus or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Functional inhibition of the acid sphingomyelinase using tricyclic antidepressants prevented infection of epithelial cells, for instance with SARS-CoV-2. The structure of ambroxol, that is, trans-4-[(2,4-dibromanilin-6-yl)methyamino]-cyclohexanol, a mucolytic drug applied by inhalation, suggests that the drug might inhibit the acid sphingomyelinase and thereby infection with SARS-CoV-2. To test this, we used vesicular stomatitis virus pseudoviral particles presenting SARS-CoV-2 spike protein on their surface (ppVSV-SARS-CoV-2 spike), a bona fide system for mimicking SARS-CoV-2 entry into cells. Viral uptake and formation of ceramide localization were determined by fluorescence microscopy, activity of the acid sphingomyelinase by consumption of [14C]sphingomyelin and ceramide was quantified by a kinase method. We found that entry of pp-VSV-SARS-CoV-2 spike required activation of acid sphingomyelinase and release of ceramide, events that were all prevented by pretreatment with ambroxol. We also obtained nasal epithelial cells from human volunteers prior to and after inhalation of ambroxol. Inhalation of ambroxol reduced acid sphingomyelinase activity in nasal epithelial cells and prevented pp-VSV-SARS-CoV-2 spikeinduced acid sphingomyelinase activation, ceramide release, and entry of pp-VSV-SARS-CoV-2 spike ex vivo. The addition of purified acid sphingomyelinase or C16 ceramide restored entry of pp-VSV-SARS-CoV-2 spike into ambroxol-treated epithelial cells. We propose that ambroxol might be suitable for clinical studies to prevent coronavirus disease 2019. Infections with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the..
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