PLASMA LIPIDS AND AMINO ACIDS LINKED TO LOW SARS-CoV-2 SUSCEPTIBILITY

Background: The mechanisms driving SARS-CoV-2 susceptibility remain poorly understood, especially the factors determining why a subset of unvaccinated individuals remain uninfected despite high-risk exposures. Method(s): We studied an exceptional group of unvaccinated healthcare workers heavily exposed to SARS-CoV-2 ('nonsusceptible') from April to June 2020, who were compared against 'susceptible' individuals to SARS-CoV-2, including uninfected subjects who became infected during the follow-up, and hospitalized patients with different disease severity providing samples at early disease stages. We analyzed plasma samples using different mass spectrometry technique and obtained metabolites and lipids profiles. Result(s): We found that the metabolite profiles were predictive of the selected study groups and identified lipids profiles and metabolites linked to SARS-CoV-2 susceptibility and COVID-19 severity. More importantly, we showed that non-susceptible individuals exhibited unique metabolomics and lipidomic patterns characterized by upregulation of most lipids -especially ceramides and sphingomyelin-and amino acids related to tricarboxylic acid cycle and mitochondrial metabolism, which could be interpreted as markers of low susceptibility to SARS-CoV-2 infection. Lipids and metabolites pathways analysis revealed that metabolites related to energy production, mitochondrial and tissue dysfunction, and lipids involved in membrane structure and virus infectivity were key markers of SARS-CoV-2 susceptibility. Conclusion(s): Lipid and metabolic profiles differ in 'nonsusceptible' compared to individuals susceptible to SARS-CoV-2. Our study suggests that lipid profiles are relevant actors during SARS-CoV-2 pathogenesis and highlight certain lipids relevant to understand SARS-CoV-2 pathogenesis. (Figure Presented).

Sphingolipid metabolism and its relationship with cardiovascular, renal and metabolic diseases.

Sphingolipids (sphingomyelin, glycolipids, gangliosides) are located in cell membranes, plasma, and lipoproteins. In patients with cardiovascular, renal, and metabolic diseases, the profile of sphingolipids and their metabolites (ceramide, sphingosine, and sphingosine-1-phosphate) is modified, and these changes may explain the alterations in some cellular responses such as apoptosis. Furthermore, sphingosine and sphingosine-1-phosphate have been suggested to prevent COVID-19. This review also briefly mentions the techniques that allow us to study sphingolipids and their metabolites.

Los esfingolípidos (esfingomielina, glucolípidos y gangliósidos) se localizan en las membranas celulares, el plasma y las lipoproteínas. En pacientes con enfermedades cardiovasculares, renales y metabólicas, el perfil de los esfingolípidos y sus metabolitos (ceramida, esfingosina y esfingosina-1-fosfato) se modifica, y estos cambios pueden explicar las alteraciones en algunas respuestas celulares, como la apoptosis. Además, se ha sugerido que la esfingosina y la esfingosina-1-fosfato previenen la COVID-19. En esta revisión también se mencionan brevemente las técnicas que permiten el estudio de los esfingolípidos y sus metabolitos.

Pulmonary Phospholipid Components as Promising Natural Inhibitors against COVID-19 Mpro;Molecular Docking Analysis Based Study

The ongoing pandemic of COVID-19 caused by the severe acute respiratory syndrome SARS-CoV-2 has become a global crisis. Phospholipids are structural components of mammalian cell membranes that suppress viral attachment to the plasma membrane and subsequent replication in lung cells. Using the molecular docking approach, the inhibitory activity of phosphatidylcholine, dipalmitoylphosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidyl-inositol, lysobisphosphatidic acid and sphingomyelin against SARS CoV-2 by targeting main protease (Mpro, PDB code: 6LU7) has been investigated. All phospholipids established excellent binding to Mpro active bocket by forming several H-bonds with the catalytic amino acids Cys145 and His4, as well as various amino acids involved in the bocket. Furthermore, a potent binding affinity is increased from -7.01 to -9.16 kcal/mol compared to compound N3 (N-[(5methylisoxazol-3-yl)carbonyl]alanyl-L (where L = valyl-N-1-(1R,2Z)-4-(benzyloxy)-4-oxo-1-{[(3R)-2-oxopyrrolidin-3-yl]methyl}but-2-enyl)-L-leucinamide), a peptide linker, inhibitor for Covid-19 main protease. Co-crystalline ligand of enzyme 6LU7 of -9.99 kcal/mol. The sphingomyelin has the same binding affinity to main protease when compared to compound N3. These findings implied that the selected compounds have the potential to be developed as novel SARS-CoV-2 inhibitors. Therefore, improved, well-designed, potent and structurally and pharmacokinetically effective drugs are urgently needed. Further investigations should focus on validating and finalizing effective drugs for COVID-19 beyond preliminary in silico and in vivo screening. © 2022 Chemical Publishing Co.. All rights reserved.

Effect of Subacute Angiotensin II Elevations on Sphingolipid Levels and Lung Injury in Mice

Background: There is a paucity of therapies for acute lung injury (ALI) induced by respiratory viruses. A previously demonstrated key mechanism of ALI, particularly in the setting of severe acute respiratory syndrome coronavirus infections, has been ascribed to decreased cell surface angiotensin converting enzyme 2 (ACE2) leading to increased circulating levels of angiotensin II (Ang2). In turn, supraphysiological Ang2 levels trigger a cascade of events that culminates with endothelial injury in the systemic circulation via acid sphingomyelinase (ASMase) activation. ASMase has been implicated in several models of ALI, but its specific involvement in Ang2-induced ALI is unknown. ASMase hydrolyzes sphingomyelin to pro-apoptotic, edemagenic ceramide, which can be metabolized to endothelial-protective sphingosine-1-phosphate (S1P). Therefore, the ratio of ceramide/S1P can determine endothelial cell fate and lung vascular permeability. We hypothesized that ceramide levels are increased relative to S1P in mice with Ang2-induced ALI. Methods: Following a published protocol of Ang2-induced ALI (Wu et al, 2017), we delivered Ang2 via osmotic pumps (1 ug/kg/min, 7 days;Ang2-mice), using saline (sham) or untreated C57BL/6 mice as controls. We evaluated pulmonary function (FlexiVent);albumin, IgM (ELISA), and inflammatory cell abundance in bronchoalveolar lavage fluid (BALF);and lung parenchyma inflammation and fibrosis (Ashcroft score) on H/E-stained lungs. Sphingolipid levels in lungs and plasma were measured by tandem liquid chromatography/mass spectrometry. Results: Inspiratory capacity, lung compliance, and body weight all decreased in Ang2-mice (by 13-14%, p<0.05 each) compared to sham. Lung pressure-volume loops exhibited a right-shift in Ang2- vs. sham or untreated mice. There was no significant change in BALF albumin, IgM, or inflammatory cells, or in lung histology inflammation or fibrosis scores in Ang2-mice. Compared to sham, S1P levels were significantly increased in plasma and unlavaged lung in Ang2-mice, decreasing ceramide/S1P ratios (from 3.1 to 2.0, and 26 to 20, respectively, p<0.05 each). Conclusions: Sustained subacute systemic elevations of Ang2 increased lung stiffness, but did not cause severe ALI in mice. Lung and circulatory elevations of S1P but not ceramide may have protected against lung edema and inflammatory injury. Although the cause of increased lung stiffness in this model remains to be elucidated, it is notable that chronic (months) supraphysiological elevations of either Ang2 or S1P have been associated with lung fibrosis. In conclusion, a second-hit injury may be necessary to augment the susceptibility of murine lung to Ang2-induced endothelial damage and inflammation relevant to coronavirus.

eP280: Continued improvement in adults with acid sphingomyelinase deficiency after 2 years of olipudase alfa in the ASCEND placebo-controlled trial

Introduction: Acid sphingomyelinase deficiency (ASMD), also historically known as Niemann-Pick disease A (OMIM #257200) and B (OMIM#607616), is a rare and debilitating lysosomal storage disease caused by pathogenic variants in SMPD1 gene. Deficient activity of the lysosomal enzyme acid sphingomyelinase (ASM) leads to sphingomyelin accumulation in various organs. Visceral manifestations of ASMD include interstitial lung disease and pulmonary dysfunction, splenomegaly, hepatomegaly, dyslipidemia, thrombocytopenia, and anemia and are present across ASMD phenotypes (ASMD type A, B and A/B). In more severe cases of ASMD (ASMD type A), there are also central nervous system manifestations. No disease-specific treatment is currently approved for patients with ASMD. Olipudase alfa, an intravenous-recombinant-human ASM, is in late-stage development (Sanofi Genzyme) for the non-central-nervous-system manifestations of ASMD in children and adults. Two open-label trials, a phase 1b trial in 5 adults (NCT01722526) and a phase 1/2 trial in 20 children with chronic ASMD (ASCEND-Peds, NCT02292654) demonstrated improvement of pulmonary function, reduction of liver and spleen volume, reversal of dyslipidemia, decreased disease biomarkers, and in children, improved growth. A phase 2/3 placebo-controlled trial, the ASCEND study (NCT02004691) in 36 adults with ASMD who had splenomegaly and pulmonary dysfunction, has completed its primary analysis. Olipudase-alfa-treated patients compared to placebo-treated patients (1:1 randomization) had statistically significant increases in percent-predicted diffusing capacity of carbon monoxide (DLCO) and statistically significant decreases in spleen and liver volume after 1 year of placebo or olipudase alfa. Thirty-five of 36 patients continued in an open-label trial extension including 17 of the 18 patients who initially received placebo in the first year and all 18 patients who received olipudase alfa. Here we report Year 2 results of the ASCEND trial for the former placebo group after 1 year of olipudase alfa treatment and for the initial olipudase alfa group after 2 years of olipudase alfa treatment. Methods: All patients underwent gradual dose-escalation to 3.0 mg/kg every 2 weeks for approximately 14 weeks when starting olipudase alfa. Efficacy outcomes include percent-predicted DLCO, spleen volume, liver volume, lung high-resolution computerized tomography (HRCT) scores for ground glass appearance, histopathologic clearance of sphingomyelin in the liver, platelet count, plasma lyso-sphingomyelin, liver function, and lipid profile. Change from baseline results are presented as least-square mean (analysis of covariance [ANCOVA]) percent change ± standard error of the mean (SEM), except for ground glass appearance, which is the least-square mean ANCOVA absolute change from baseline, and percent liver tissue area occupied by sphingomyelin and plasma lyso-sphingomyelin, which are presented as mean changes ± standard deviation (SD). Absolute values at Baseline, Year 1, and Year 2 are presented as mean ± SD (Table). Results: Overall, 33 of 35 patients completed Year 2 of ASCEND;one former placebo patient withdrew due to COVID-19 travel restrictions, and one continuing olipudase alfa patient withdrew consent. COVID-19 travel restrictions also resulted in at least one missed assessment in six patients. In Year 2, improvements for patients in the former placebo group paralleled the olipudase alfa group in the primary analysis while clinical improvement continued for patients who received 2 years of olipudase alfa (Table). For patients in the former placebo group, percent-predicted DLCO increased by 28.0 ±6.2% (n=10);spleen volume decreased by 36.0 ±3.0% (n=11);liver volume decreased by 30.7 ±2.5% (n=11), and platelet count increased by 21.7 ±6.4% (n=15). In patients with 2 years of olipudase alfa treatment, percent-predicted DLCO increased by 22.2 ±3.4% (n=17) at Year 1 and 28.5±6.2% at Year 2 (n=10);spleen volume decreased by 39.5 ±2.4% (n=17) at Year 1 and 47.0 ±2.7% (n=14) at Year 2 liver volume decreased by 27.8 ±2.5% (n=17) at Year 1 and 33.4 ±2.2% (n=14) at Year 2, and platelet count increased by 16.6 ±4.0% at Year 1 (n=18) and 24.9 ±6.9% (n=13) at Year 2. HRCT ground glass appearance score decreased 0.30 ±0.5 (n=14) at Year 2 for patients in the former placebo group and decreased by 0.45 ±0.13 (n=18) at Year 1 and 0.48 ±0.07 (n=16) at Year 2 for patients continuing to receive olipudase alfa. Liver sphingomyelin clearance at Year 2 was 93.3 ±5.0% (n=10) for patients in the former placebo group and 92.7 ±5.8% at Year 1 (n=13) and 98.4 ±2.0% at Year 2 (n=10) for patients continuing to receive olipudase alfa. Plasma lyso-sphingomyelin decreased by 79.4 ±11.3% (n=14) for patients in the former placebo group and by 78.0 ±11.1% (n=18) at Year 1 and 64.4 ±28.5% (n=15) at Year 2 for patients continuing to receive olipudase alfa;several patients had transient increases due to missed infusions. Alanine aminotransferase decreased by 45.2 ±34.4% (n=15) for patients in the former placebo group, and by 36.5 ±8.4% (n=18) in Year 1 and 32.0 ±10.2% (n=12) in Year 2 for patients continuing to receive olipudase alfa. For patients in the former placebo group, high-density lipoprotein cholesterol (HDL-C) increased by 59.7 ±9.7% (n=14) and low-density lipoprotein cholesterol (LDL-C) decreased by 27.5 ±6.8% (n=13) in Year 2. For patients continuing to receive olipudase alfa, HDL-C increased by 40.0 ±6.8% (n=18) in Year 1 and 64.4 ±10.5% (n=12) in Year 2 and LDL-C decreased by 25.8 ±4.8% (n=18) in Year 1 and 23.0 ±7.1% (n=12) in Year 2. Overall, 99% of treatment-emergent adverse events were mild or moderate, with one treatment-related serious adverse event (extrasystoles in patient with previously documented cardiomyopathy). No patient discontinued due to an adverse event. Conclusion: During Year 2 of ASCEND, patients crossing over from placebo to olipudase alfa had the same magnitude and time course of clinical improvement seen in patients receiving olipudase alfa for 1 year, while continuing olipudase-alfa patients had sustained or further improvements. Olipudase alfa reduced sphingomyelin storage in the liver and lyso-sphingomyelin in plasma. Clinically, olipudase alfa improved pulmonary function, reduced splenomegaly and hepatomegaly, and improved liver function and dyslipidemia for up to 2 years. These results are consistent with the published 30- and 42-month data for adults reported in the long-term extension of the open-label Phase 1b study. Treatment with olipudase alfa reduces manifestations of chronic ASMD in adults and has sustained efficacy. [Formula presented]

Two-year results of the ASCEND trial of olipudase alfa adults with chronic acid sphingomyelinase deficiency show parallel improvements in former placebo patients and further improvement in continuing olipudase alfa patients

Acid sphingomyelinase deficiency (ASMD) is a rare debilitating lysosomal disease characterized by pulmonary dysfunction, hepatosplenomegaly, and dyslipidemia. Olipudase alfa, intravenous-recombinant-human ASM, is in late-stage development (Sanofi Genzyme) for non-central-nervous-system manifestations of ASMD. We report 2-year outcomes for 33/36 ASMD adults with splenomegaly (mean baseline spleen volume: 11.3 multiples of normal [MN]) and respiratory impairment (mean baseline percent-predicted-diffusing capacity for carbon monoxide [DLCO]: 49.3%) who participated in the 1-year double-blind placebo-controlled primary-analysis period [PAP] of the ASCEND trial of olipudase alfa (NCT02004691) and completed a second year in the open-label extension. Patients underwent gradual dose-escalation to 3.0 mg/kg/2-weeks. During the PAP, olipudase-alfa-treated compared to placebo-treated patients (1:1 randomization) had statistically significant increases in DLCO and decreases in spleen and liver volume. One placebo patient withdrew during year-1. In year-2, improvements in former placebo patients paralleled the olipudase-alfa group in the PAP (all values: ANCOVA LS-mean percent change from trial baseline ± SEM): DLCO increased 28.0 ± 6.2% (n = 10);spleen volume decreased 36.0 ± 3.0% (n = 11);liver volume decreased 30.7 ± 2.5% (n = 11). Olipudase-alfa patients who received 2 years of treatment continued improving: DLCO increased 28.5 ± 6.2%, n = 10 (year-1 increase: 22.2 ± 3.4%, n = 17);spleen volume decreased 47.0 ± 2.7%, n = 14 (year-1 decrease: 39.5 ± 2.4%, n = 17), liver volume decreased 33.4 ± 2.2%, n = 14 (year-1 decrease: 27.8 ± 2.5, n = 17). Improvements in dyslipidemia, liver function, liver sphingomyelin clearance, and plasma lyso-sphingomyelin in former placebo patients paralleled those seen in olipudase-alfa patients in the PAP;continuing olipudase-alfa patients maintained these benefits in year-2. Overall, 99% of treatment-emergent adverse events were mild/moderate, with one treatment-related serious adverse event. During year-2, six patients missed ≥1 assessments and one patient discontinued due to COVID-19 travel restrictions;one additional patient discontinued (withdrawal of consent). In summary, during year-2 of ASCEND, crossover-placebo patients improved to a similar extent as olipudase-alfa patients in year-1 and patients continuing on olipudase alfa showed sustained or further improvements.

P.0404 Rapid antidepressant response to first-line selective serotonin reuptake inhibitors in post-COVID-19 depression

Introduction: Depression was reported in 30–40% of patients at one, three, and six months following COVID-19 [1]. The host immune response to SARS-CoV-2 infection and related severe systemic inflammation seems to be the main mechanism contributing to the development of post-COVID depression. Emerging literature suggests anti-inflammatory and antiviral properties of antidepressants in the treatment of SARS-CoV-2 infection [2]. We hypothesized that post-COVID depression, triggered by infection and sustained by systemic inflammation, could particularly benefit from antidepressants. Thus, the present study aims to investigate the efficacy of SSRI in treating post-COVID depression. Methods: We included 58 adults patients who showed depressive episodes in the six months following COVID-19. We excluded patients if they showed: other psychiatric comorbidities, ongoing treatment with antidepressants or neuroleptics, somatic disease and medications known to affect mood. The severity of depression was rated at baseline and after for four weeks from the start of the treatment on the Hamilton Depression Rating Scale (HDRS) and response was considered when the patients achieved a 50% HDRS reduction after treatment. Statistical analyses to compare group means and frequencies (Student's t-test, Pearson χ2 test) were performed. To investigate changes in HDRS scores over time, repeated measures ANOVAs (according to sex, mood disorder history, and antidepressant molecule) were performed. Results: We found that 53 (91%) patients showed a clinical response to antidepressant treatment. Age, sex, mood disorder history, and hospitalization for COVID did not affect the response rate. Patients were treated with sertraline (n=26), citalopram (n=18), paroxetine (n=8), fluvoxamine (n=4), and fluoxetine (n=2). From baseline to follow-up, patients showed a significant decrease over time of HDRS score (F=618.90, p<0.001), irrespectively of sex (0.28, p=0.599), mood disorder history (F=0.04, p=0.834), and drug used (F=1.47, p=0.239). Discussion: Common knowledge highlights that among antidepressant-treated patients, response rates are moderate (40–60%). On the contrary, we observed a rapid response to the first-line antidepressants in more than 90% of patients irrespectively of clinical variables, thus suggesting a higher antidepressant response rate in post-COVID depression. The pathophysiology of post-COVID neuropsychiatric sequelae mainly entails severe systemic inflammation and subsequent neuroinflammation. In this context, we have previously found that one and three months after COVID-19, the severity of depression was predicted by the baseline systemic immune-inflammation index (SII) [3,4]. Furthermore, we found a protective effect of the IL-1β and IL-6 receptor antagonist against post-COVID depression possibly associated with their effect in dampening SII [5]. Mounting evidence suggests that antidepressants may a) decrease markers of inflammation;b) may inhibit acid sphingomyelinase preventing the infection of epithelial cells with SARS-CoV-2;c) may prevent the COVID-19 related cytokine storm by stimulating the σ-1 receptor;d) may exert antiviral effects via lysosomotropic properties;e) may inhibit platelets activation [2]. In conclusion, we hypothesized that post-COVID depression could particularly benefit from antidepressants since this molecules have anti-inflammatory and antiviral properties, pass the BBB and accumulate in the CNS, thus preventing the neuro-inflammation triggered by SARS-CoV-2 and associated with post-COVID depression. No conflict of interest

Sphingolipidomics in Translational Sepsis Research-Biomedical Considerations and Perspectives.

Scientific Background: Sphingolipids are a highly diverse group of lipids with respect to physicochemical properties controlling either structure, distribution, or function, all of them regulating cellular response in health and disease. Mass spectrometry, on the other hand, is an analytical technique characterizing ionized molecules or fragments thereof by mass-to-charge ratios, which has been prosperingly developed for rapid and reliable qualitative and quantitative identification of lipid species. Parallel to best performance of in-depth chromatographical separation of lipid classes, preconditions of precise quantitation of unique molecular species by preprocessing of biological samples have to be fulfilled. As a consequence, "lipid profiles" across model systems and human individuals, esp. complex (clinical) samples, have become eminent over the last couple of years due to sensitivity, specificity, and discriminatory capability. Therefore, it is significance to consider the entire experimental strategy from sample collection and preparation, data acquisition, analysis, and interpretation. Areas Covered: In this review, we outline considerations with clinical (i.e., human) samples with special emphasis on sample handling, specific physicochemical properties, target measurements, and resulting profiling of sphingolipids in biomedicine and translational research to maximize sensitivity and specificity as well as to provide robust and reproducible results. A brief commentary is also provided regarding new insights of "clinical sphingolipidomics" in translational sepsis research. Expert Opinion: The role of mass spectrometry of sphingolipids and related species ("sphingolipidomics") to investigate cellular and compartment-specific response to stress, e.g., in generalized infection and sepsis, is on the rise and the ability to integrate multiple datasets from diverse classes of biomolecules by mass spectrometry measurements and metabolomics will be crucial to fostering our understanding of human health as well as response to disease and treatment.

Differential role of sphingomyelin in influenza virus, rhinovirus and SARS-CoV-2 infection of Calu-3 cells.

Host cell lipids play a pivotal role in the pathogenesis of respiratory virus infection. However, a direct comparison of the lipidomic profile of influenza virus and rhinovirus infections is lacking. In this study, we first compared the lipid profile of influenza virus and rhinovirus infection in a bronchial epithelial cell line. Most lipid features were downregulated for both influenza virus and rhinovirus, especially for the sphingomyelin features. Pathway analysis showed that sphingolipid metabolism was the most perturbed pathway. Functional study showed that bacterial sphingomyelinase suppressed influenza virus and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication, but promoted rhinovirus replication. These findings suggest that sphingomyelin pathway can be a potential target for antiviral therapy, but should be carefully evaluated as it has opposite effects on different respiratory viruses. Furthermore, the differential effect of sphingomyelinase on rhinovirus and influenza virus may explain the interference between rhinovirus and influenza virus infection.