Fluoxetine and Sertraline Potently Neutralize the Replication of Distinct SARS-CoV-2 Variants.

The pandemic caused by SARS-CoV-2 is still a major health problem. Newly emerging variants and long-COVID-19 represent a challenge for the global health system. In particular, individuals in developing countries with insufficient health care need easily accessible, affordable and effective treatments of COVID-19. Previous studies have demonstrated the efficacy of functional inhibitors of acid sphingomyelinase against infections with various viruses, including early variants of SARS-CoV-2. This work investigated whether the acid sphingomyelinase inhibitors fluoxetine and sertraline, usually used as antidepressant molecules in clinical practice, can inhibit the replication of the former and recently emerged SARS-CoV-2 variants in vitro. Fluoxetine and sertraline potently inhibited the infection with pseudotyped virus-like particles and SARS-CoV-2 variants D614G, alpha, delta, omicron BA.1 and omicron BA.5. These results highlight fluoxetine and sertraline as priority candidates for large-scale phase 3 clinical trials at different stages of SARS-CoV-2 infections, either alone or in combination with other medications.

Medications Modulating the Acid Sphingomyelinase/Ceramide System and 28-Day Mortality among Patients with SARS-CoV-2: An Observational Study.

Prior evidence indicates the potential central role of the acid sphingomyelinase (ASM)/ceramide system in the infection of cells with SARS-CoV-2. We conducted a multicenter retrospective observational study including 72,105 adult patients with laboratory-confirmed SARS-CoV-2 infection who were admitted to 36 AP-HP (Assistance Publique-Hôpitaux de Paris) hospitals from 2 May 2020 to 31 August 2022. We examined the association between the ongoing use of medications functionally inhibiting acid sphingomyelinase (FIASMA), which reduces the infection of cells with SARS-CoV-2 in vitro, upon hospital admission with 28-day all-cause mortality in a 1:1 ratio matched analytic sample based on clinical characteristics, disease severity and other medications (N = 9714). The univariate Cox regression model of the matched analytic sample showed that FIASMA medication use at admission was associated with significantly lower risks of 28-day mortality (HR = 0.80; 95% CI = 0.72-0.88; p < 0.001). In this multicenter observational study, the use of FIASMA medications was significantly and substantially associated with reduced 28-day mortality among adult patients hospitalized with COVID-19. These findings support the continuation of these medications during the treatment of SARS-CoV-2 infections. Randomized clinical trials (RCTs) are needed to confirm these results, starting with the molecules with the greatest effect size in the study, e.g., fluoxetine, escitalopram, and amlodipine.

Acid sphingomyelinase expression is associated with survival in resectable pancreatic ductal adenocarcinoma.

Pancreatic adenocarcinoma (PDAC) is one of the most common cancers worldwide. Unfortunately, the prognosis of PDAC is rather poor, and for instance, in the USA, over 47,000 people die because of pancreatic cancer annually. Here, we demonstrate that high expression of acid sphingomyelinase in PDAC strongly correlates with long-term survival of patients, as revealed by the analysis of two independent data sources. The positive effects of acid sphingomyelinase expression on long-term survival of PDAC patients were independent of patient demographics as well as tumor grade, lymph node involvement, perineural invasion, tumor stage, lymphovascular invasion, and adjuvant therapy. We also demonstrate that genetic deficiency or pharmacological inhibition of the acid sphingomyelinase promotes tumor growth in an orthotopic mouse model of PDAC. This is mirrored by a poorer pathologic response, as defined by the College of American Pathologists (CAP) score for pancreatic cancer, to neoadjuvant therapy of patients co-treated with functional inhibitors of the acid sphingomyelinase, in particular tricyclic antidepressants and selective serotonin reuptake inhibitors, in a retrospective analysis. Our data indicate expression of the acid sphingomyelinase in PDAC as a prognostic marker for tumor progression. They further suggest that the use of functional inhibitors of the acid sphingomyelinase, at least of tricyclic antidepressants and selective serotonin reuptake inhibitors in patients with PDAC, is contra-indicated. Finally, our data also suggest a potential novel treatment of PDAC patients with recombinant acid sphingomyelinase. KEY MESSAGES: Pancreatic ductal adenocarcinoma (PDAC) is a common tumor with poor prognosis. Expression of acid sphingomyelinase (ASM) determines outcome of PDAC. Genetic deficiency or pharmacologic inhibition of ASM promotes tumor growth in a mouse model. Inhibition of ASM during neoadjuvant treatment for PDAC correlates with worse pathology. ASM expression is a prognostic marker and potential target in PDAC.

Simultaneous targeting of mitochondrial Kv1.3 and lysosomal acid sphingomyelinase amplifies killing of pancreatic ductal adenocarcinoma cells in vitro and in vivo.

Pancreas ductal adenocarcinoma (PDAC) remains a malignant tumor with very poor prognosis and low 5-year overall survival. Here, we aimed to simultaneously target mitochondria and lysosomes as a new treatment paradigm of malignant pancreas cancer in vitro and in vivo. We demonstrate that the clinically used sphingosine analog FTY-720 together with PAPTP, an inhibitor of mitochondrial Kv1.3, induce death of pancreas cancer cells in vitro and in vivo. The combination of both drugs results in a marked inhibition of the acid sphingomyelinase and accumulation of cellular sphingomyelin in vitro and in vivo in orthotopic and flank pancreas cancers. Mechanistically, PAPTP and FTY-720 cause a disruption of both mitochondria and lysosomes, an alteration of mitochondrial bioenergetics and accumulation of cytoplasmic Ca2+, events that collectively mediate cell death. Our findings point to an unexpected cross-talk between lysosomes and mitochondria mediated by sphingolipid metabolism. We show that the combination of PAPTP and FTY-720 induces massive death of pancreas cancer cells, thereby leading to a substantially delayed and reduced PDAC growth in vivo. KEY MESSAGES: FTY-720 inhibits acid sphingomyelinase in pancreas cancer cells (PDAC). FTY-720 induces sphingomyelin accumulation and lysosomal dysfunction. The mitochondrial Kv1.3 inhibitor PAPTP disrupts mitochondrial functions. PAPTP and FTY-720 synergistically kill PDAC in vitro. The combination of FTY-720 and PAPTP greatly delays PDAC growth in vivo.

Risk of Death in Individuals Hospitalized for COVID-19 With and Without Psychiatric Disorders: An Observational Multicenter Study in France.

Background: Prior research suggests that psychiatric disorders could be linked to increased mortality among patients with COVID-19. However, whether all or specific psychiatric disorders are intrinsic risk factors of death in COVID-19 or whether these associations reflect the greater prevalence of medical risk factors in people with psychiatric disorders has yet to be evaluated. Methods: We performed an observational, multicenter, retrospective cohort study to examine the association between psychiatric disorders and mortality among patients hospitalized for laboratory-confirmed COVID-19 at 36 Greater Paris University hospitals. Results: Of 15,168 adult patients, 857 (5.7%) had an ICD-10 diagnosis of psychiatric disorder. Over a mean follow-up period of 14.6 days (SD = 17.9), 326 of 857 (38.0%) patients with a diagnosis of psychiatric disorder died compared with 1276 of 14,311 (8.9%) patients without such a diagnosis (odds ratio 6.27, 95% CI 5.40-7.28, p < .01). When adjusting for age, sex, hospital, current smoking status, and medications according to compassionate use or as part of a clinical trial, this association remained significant (adjusted odds ratio 3.27, 95% CI 2.78-3.85, p < .01). However, additional adjustments for obesity and number of medical conditions resulted in a nonsignificant association (adjusted odds ratio 1.02, 95% CI 0.84-1.23, p = .86). Exploratory analyses after the same adjustments suggested that a diagnosis of mood disorders was significantly associated with reduced mortality, which might be explained by the use of antidepressants. Conclusions: These findings suggest that the increased risk of COVID-19-related mortality in individuals with psychiatric disorders hospitalized for COVID-19 might be explained by the greater number of medical conditions and the higher prevalence of obesity in this population and not by the underlying psychiatric disease.

Cell-intrinsic ceramides determine T cell function during melanoma progression.

Acid sphingomyelinase (Asm) and acid ceramidase (Ac) are parts of the sphingolipid metabolism. Asm hydrolyzes sphingomyelin to ceramide, which is further metabolized to sphingosine by Ac. Ceramide generates ceramide-enriched platforms that are involved in receptor clustering within cellular membranes. However, the impact of cell-intrinsic ceramide on T cell function is not well characterized. By using T cell-specific Asm- or Ac-deficient mice, with reduced or elevated ceramide levels in T cells, we identified ceramide to play a crucial role in T cell function in vitro and in vivo. T cell-specific ablation of Asm in Smpd1fl/fl/Cd4cre/+ (Asm/CD4cre) mice resulted in enhanced tumor progression associated with impaired T cell responses, whereas Asah1fl/fl/Cd4cre/+ (Ac/CD4cre) mice showed reduced tumor growth rates and elevated T cell activation compared to the respective controls upon tumor transplantation. Further in vitro analysis revealed that decreased ceramide content supports CD4+ regulatory T cell differentiation and interferes with cytotoxic activity of CD8+ T cells. In contrast, elevated ceramide concentration in CD8+ T cells from Ac/CD4cre mice was associated with enhanced cytotoxic activity. Strikingly, ceramide co-localized with the T cell receptor (TCR) and CD3 in the membrane of stimulated T cells and phosphorylation of TCR signaling molecules was elevated in Ac-deficient T cells. Hence, our results indicate that modulation of ceramide levels, by interfering with the Asm or Ac activity has an effect on T cell differentiation and function and might therefore represent a novel therapeutic strategy for the treatment of T cell-dependent diseases such as tumorigenesis.

Antiviral and Anti-Inflammatory Activities of Fluoxetine in a SARS-CoV-2 Infection Mouse Model.

The coronavirus disease 2019 (COVID-19) pandemic continues to cause significant morbidity and mortality worldwide. Since a large portion of the world's population is currently unvaccinated or incompletely vaccinated and has limited access to approved treatments against COVID-19, there is an urgent need to continue research on treatment options, especially those at low cost and which are immediately available to patients, particularly in low- and middle-income countries. Prior in vitro and observational studies have shown that fluoxetine, possibly through its inhibitory effect on the acid sphingomyelinase/ceramide system, could be a promising antiviral and anti-inflammatory treatment against COVID-19. In this report, we evaluated the potential antiviral and anti-inflammatory activities of fluoxetine in a K18-hACE2 mouse model of SARS-CoV-2 infection, and against variants of concern in vitro, i.e., SARS-CoV-2 ancestral strain, Alpha B.1.1.7, Gamma P1, Delta B1.617 and Omicron BA.5. Fluoxetine, administrated after SARS-CoV-2 infection, significantly reduced lung tissue viral titres and expression of several inflammatory markers (i.e., IL-6, TNFα, CCL2 and CXCL10). It also inhibited the replication of all variants of concern in vitro. A modulation of the ceramide system in the lung tissues, as reflected by the increase in the ratio HexCer 16:0/Cer 16:0 in fluoxetine-treated mice, may contribute to explain these effects. Our findings demonstrate the antiviral and anti-inflammatory properties of fluoxetine in a K18-hACE2 mouse model of SARS-CoV-2 infection, and its in vitro antiviral activity against variants of concern, establishing fluoxetine as a very promising candidate for the prevention and treatment of SARS-CoV-2 infection and disease pathogenesis.

Antidepressant Use and Its Association with 28-Day Mortality in Inpatients with SARS-CoV-2: Support for the FIASMA Model against COVID-19.

To reduce Coronavirus Disease 2019 (COVID-19)-related mortality and morbidity, widely available oral COVID-19 treatments are urgently needed. Certain antidepressants, such as fluvoxamine or fluoxetine, may be beneficial against COVID-19. We included 388,945 adult inpatients who tested positive for SARS-CoV-2 at 36 AP−HP (Assistance Publique−Hôpitaux de Paris) hospitals from 2 May 2020 to 2 November 2021. We compared the prevalence of antidepressant use at admission in a 1:1 ratio matched analytic sample with and without COVID-19 (N = 82,586), and assessed its association with 28-day all-cause mortality in a 1:1 ratio matched analytic sample of COVID-19 inpatients with and without antidepressant use at admission (N = 1482). Antidepressant use was significantly less prevalent in inpatients with COVID-19 than in a matched control group of inpatients without COVID-19 (1.9% versus 4.8%; Odds Ratio (OR) = 0.38; 95%CI = 0.35−0.41, p < 0.001). Antidepressant use was significantly associated with reduced 28-day mortality among COVID-19 inpatients (12.8% versus 21.2%; OR = 0.55; 95%CI = 0.41−0.72, p < 0.001), particularly at daily doses of at least 40 mg fluoxetine equivalents. Antidepressants with high FIASMA (Functional Inhibitors of Acid Sphingomyelinase) activity seem to drive both associations. These treatments may reduce SARS-CoV-2 infections and COVID-19-related mortality in inpatients, and may be appropriate for prophylaxis and/or COVID-19 therapy for outpatients or inpatients.

Stress induces major depressive disorder by a neutral sphingomyelinase 2-mediated accumulation of ceramide-enriched exosomes in the blood plasma.

Major depressive disorder (MDD) is a very common, severe disease with a lifetime prevalence of ~ 10%. The pathogenesis of MDD is unknown and, unfortunately, therapy is often insufficient. We have previously reported that ceramide levels are increased in the blood plasma of patients with MDD and in mice with experimental MDD. Here, we demonstrate that ceramide-enriched exosomes in the blood plasma are increased in mice with stress-induced MDD. Genetic studies reveal that neutral sphingomyelinase 2 is required for the formation of ceramide-enriched exosomes in the blood plasma. Accordingly, induced deficiency of neutral sphingomyelinase 2 prevented mice from the development of stress-induced MDD. Intravenous injection of microparticles from mice with MDD or injection of ceramide-loaded exosomes induced MDD-like behavior in untreated mice, which was abrogated by ex vivo pre-incubation of purified exosomes with anti-ceramide antibodies or ceramidase. Mechanistically, injection of exosomes from mice with MDD or injection of ex vivo ceramide-loaded microparticles inhibited phospholipase D (PLD) in endothelial cells in vitro and in the hippocampus in vivo and thereby decreased phosphatidic acid in the hippocampus, which has been previously shown to mediate MDD by plasma ceramide. In summary, our data indicate that ceramide-enriched exosomes are released by neutral sphingomyelinase 2 into the blood plasma upon stress and mediate stress-induced MDD. KEY MESSAGES: Stress induces ceramide-enriched exosomes in the blood plasma. Ceramide-enriched exosomes mediate major depressive disorder (MDD). Deficiency of neutral sphingomyelinase 2 protects from stress-induced MDD. Neutralization or digestion of ceramide in exosomes prevents stress-induced MDD. Ceramide-enriched exosomes inhibit endothelial phospholipase D in the hippocampus.

Staphylococcus aureus α-Toxin Induces Acid Sphingomyelinase Release From a Human Endothelial Cell Line.

Staphylococcus aureus (S. aureus) is well known to express a plethora of toxins of which the pore-forming hemolysin A (α-toxin) is the best-studied cytolysin. Pore-forming toxins (PFT) permeabilize host membranes during infection thereby causing concentration-dependent effects in host cell membranes ranging from disordered ion fluxes to cytolysis. Host cells possess defense mechanisms against PFT attack, resulting in endocytosis of the breached membrane area and delivery of repair vesicles to the insulted plasma membrane as well as a concurrent release of membrane repair enzymes. Since PFTs from several pathogens have been shown to recruit membrane repair components, we here investigated whether staphylococcal α-toxin is able to induce these mechanisms in endothelial cells. We show that S. aureus α-toxin induced increase in cytosolic Ca2+ in endothelial cells, which was accompanied by p38 MAPK phosphorylation. Toxin challenge led to increased endocytosis of an extracellular fluid phase marker as well as increased externalization of LAMP1-positive membranes suggesting that peripheral lysosomes are recruited to the insulted plasma membrane. We further observed that thereby the lysosomal protein acid sphingomyelinase (ASM) was released into the cell culture medium. Thus, our results show that staphylococcal α-toxin triggers mechanisms in endothelial cells, which have been implicated in membrane repair after damage of other cell types by different toxins.

Ceramide and Related Molecules in Viral Infections.

Ceramide is a lipid messenger at the heart of sphingolipid metabolism. In concert with its metabolizing enzymes, particularly sphingomyelinases, it has key roles in regulating the physical properties of biological membranes, including the formation of membrane microdomains. Thus, ceramide and its related molecules have been attributed significant roles in nearly all steps of the viral life cycle: they may serve directly as receptors or co-receptors for viral entry, form microdomains that cluster entry receptors and/or enable them to adopt the required conformation or regulate their cell surface expression. Sphingolipids can regulate all forms of viral uptake, often through sphingomyelinase activation, and mediate endosomal escape and intracellular trafficking. Ceramide can be key for the formation of viral replication sites. Sphingomyelinases often mediate the release of new virions from infected cells. Moreover, sphingolipids can contribute to viral-induced apoptosis and morbidity in viral diseases, as well as virus immune evasion. Alpha-galactosylceramide, in particular, also plays a significant role in immune modulation in response to viral infections. This review will discuss the roles of ceramide and its related molecules in the different steps of the viral life cycle. We will also discuss how novel strategies could exploit these for therapeutic benefit.

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

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 (pp-VSV-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 spike-induced 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.

Interferon regulatory factor 8 regulates expression of acid ceramidase and infection susceptibility in cystic fibrosis.

Most patients with cystic fibrosis (CF) suffer from acute and chronic pulmonary infections with bacterial pathogens, which often determine their life quality and expectancy. Previous studies have demonstrated a downregulation of the acid ceramidase in CF epithelial cells resulting in an increase of ceramide and a decrease of sphingosine. Sphingosine kills many bacterial pathogens, and the downregulation of sphingosine seems to determine the infection susceptibility of cystic fibrosis mice and patients. It is presently unknown how deficiency of the cystic fibrosis transmembrane conductance regulator (CFTR) connects to a marked downregulation of the acid ceramidase in human and murine CF epithelial cells. Here, we employed quantitative PCR, western blot analysis, and enzyme activity measurements to study the role of IRF8 for acid ceramidase regulation. We report that genetic deficiency or functional inhibition of CFTR/Cftr results in an upregulation of interferon regulatory factor 8 (IRF8) and a concomitant downregulation of acid ceramidase expression with CF and an increase of ceramide and a reduction of sphingosine levels in tracheal and bronchial epithelial cells from both human individuals or mice. CRISPR/Cas9- or siRNA-mediated downregulation of IRF8 prevented changes of acid ceramidase, ceramide, and sphingosine in CF epithelial cells and restored resistance to Pseudomonas aeruginosa infections, which is one of the most important and common pathogens in lung infection of patients with CF. These studies indicate that CFTR deficiency causes a downregulation of acid ceramidase via upregulation of IRF8, which is a central pathway to control infection susceptibility of CF cells.

Integrative Genomic Analyses of Patient-Matched Intracranial and Extracranial Metastases Reveal a Novel Brain-Specific Landscape of Genetic Variants in Driver Genes of Malignant Melanoma.

BACKGROUND: Development of brain metastases in advanced melanoma patients is a frequent event that limits patients' quality of life and survival. Despite recent insights into melanoma genetics, systematic analyses of genetic alterations in melanoma brain metastasis formation are lacking. Moreover, whether brain metastases harbor distinct genetic alterations beyond those observed at different anatomic sites of the same patient remains unknown. EXPERIMENTAL DESIGN AND RESULTS: In our study, 54 intracranial and 18 corresponding extracranial melanoma metastases were analyzed for mutations using targeted next generation sequencing of 29 recurrently mutated driver genes in melanoma. In 11 of 16 paired samples, we detected nucleotide modifications in brain metastases that were absent in matched metastases at extracranial sites. Moreover, we identified novel genetic variants in ARID1A, ARID2, SMARCA4 and BAP1, genes that have not been linked to brain metastases before; albeit most frequent mutations were found in ARID1A, ARID2 and BRAF. Conclusion: Our data provide new insights into the genetic landscape of intracranial melanoma metastases supporting a branched evolution model of metastasis formation.

Ex vivo assay to evaluate the efficacy of drugs targeting sphingolipids in preventing SARS-CoV-2 infection of nasal epithelial cells.

This protocol enables the testing of drugs against infection of epithelial cells with SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2), using pseudo-typed replication deficient vesicular stomatitis virus particles (pp-VSV) presenting the SARS-CoV-2 spike protein. After treating human volunteers with amitriptyline, an approved antidepressant and inhibitor of the acid sphingomyelinase, freshly isolated nasal epithelial cells were infected ex vivo and infection levels were quantified. This protocol offers the possibility to rapidly test the efficacy of potential drugs in the fight against COVID-19. For complete details on the use and execution of this protocol, please refer to Carpinteiro et al. (2020).

Acid Ceramidase Rescues Cystic Fibrosis Mice from Pulmonary Infections.

Previous studies have shown that sphingosine kills a variety of pathogenic bacteria, including Pseudomonas aeruginosa and Staphylococcus aureus Sphingosine concentrations are decreased in airway epithelial cells of cystic fibrosis (CF) mice, and this defect has been linked to the infection susceptibility of these mice. Here, we tested whether the genetic overexpression of acid ceramidase rescues cystic fibrosis mice from pulmonary infections with P. aeruginosa We demonstrate that the transgenic overexpression of acid ceramidase in CF mice corresponds to the overexpression of acid ceramidase in bronchial and tracheal epithelial cells and normalizes ceramide and sphingosine levels in bronchial and tracheal epithelial cells. In addition, the expression of ß1-integrin, which is ectopically expressed on the luminal surface of airway epithelial cells in cystic fibrosis mice, an alteration that is very important for mediating pulmonary P. aeruginosa infections in cystic fibrosis, is normalized in cystic fibrosis airways upon the overexpression of acid ceramidase. Most importantly, the overexpression of acid ceramidase protects cystic fibrosis mice from pulmonary P. aeruginosa infections. Infection of CF mice or CF mice that inhaled sphingosine with P. aeruginosa or a P. aeruginosa mutant that is resistant to sphingosine indicates that sphingosine and not a metabolite kills P. aeruginosa upon pulmonary infection. These studies further support the use of acid ceramidase and its metabolite sphingosine as potential treatments of cystic fibrosis.

Pharmacological Inhibition of Acid Sphingomyelinase Prevents Uptake of SARS-CoV-2 by Epithelial Cells.

The acid sphingomyelinase/ceramide system plays an important role in bacterial and viral infections. Here, we report that either pharmacological inhibition of acid sphingomyelinase with amitriptyline, imipramine, fluoxetine, sertraline, escitalopram, or maprotiline or genetic downregulation of the enzyme prevents infection of cultured cells or freshy isolated human nasal epithelial cells with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or vesicular stomatitis virus (VSV) pseudoviral particles (pp-VSV) presenting SARS-CoV-2 spike protein (pp-VSV-SARS-CoV-2 spike), a bona fide system mimicking SARS-CoV-2 infection. Infection activates acid sphingomyelinase and triggers a release of ceramide on the cell surface. Neutralization or consumption of surface ceramide reduces infection with pp-VSV-SARS-CoV-2 spike. Treating volunteers with a low dose of amitriptyline prevents infection of freshly isolated nasal epithelial cells with pp-VSV-SARS-CoV-2 spike. The data justify clinical studies investigating whether amitriptyline, a safe drug used clinically for almost 60 years, or other antidepressants that functionally block acid sphingomyelinase prevent SARS-CoV-2 infection.

Sphingosine prevents binding of SARS-CoV-2 spike to its cellular receptor ACE2.

Sphingosine has been shown to prevent and eliminate bacterial infections of the respiratory tract, but it is unknown whether sphingosine can be also employed to prevent viral infections. To test this hypothesis, we analyzed whether sphingosine regulates the infection of cultured and freshly isolated ex vivo human epithelial cells with pseudoviral particles expressing SARS-CoV-2 spike (pp-VSV-SARS-CoV-2 spike) that served as a bona fide system mimicking SARS-CoV-2 infection. We demonstrate that exogenously applied sphingosine suspended in 0.9% NaCl prevents cellular infection with pp-SARS-CoV-2 spike. Pretreatment of cultured Vero epithelial cells or freshly isolated human nasal epithelial cells with low concentrations of sphingosine prevented adhesion of and infection with pp-VSV-SARS-CoV-2 spike. Mechanistically, we demonstrate that sphingosine binds to ACE2, the cellular receptor of SARS-CoV-2, and prevents the interaction of the receptor-binding domain of the viral spike protein with ACE2. These data indicate that sphingosine prevents at least some viral infections by interfering with the interaction of the virus with its receptor. Our data also suggest that further preclinical and finally clinical examination of sphingosine is warranted for potential use as a prophylactic or early treatment for coronavirus disease-19.