Evidence for the Involvement of Lipid Rafts and Plasma Membrane Sphingolipid Hydrolases in Pseudomonas aeruginosa

Cystic fibrosis (CF) is the most common autosomal genetic recessive disease caused by mutations of gene encoding for the cystic fibrosis transmembrane conductance regulator. Patients with CF display a wide spectrum of symptoms, the most severe being chronic lung infection and inflammation, which lead to onset of cystic fibrosis lung disease. Several studies indicate that sphingolipids play a regulatory role in airway inflammation. The inhibition and downregulation of GBA2, the enzyme catabolizing glucosylceramide to ceramide, are associated with a significant reduction of IL-8 production in CF bronchial epithelial cells. Herein, we demonstrate that GBA2 plays a role in the proinflammatory state characterizing CF cells. We also report for the first time that Pseudomonas aeruginosa infection causes a recruitment of plasma membrane-associated glycosphingolipid hydrolases into lipid rafts of CuFi-1-infected cells. This reorganization of cell membrane may be responsible for activation of a signaling cascade, culminating in aberrant inflammatory response in CF bronchial epithelial cells upon bacterial infection. Taken together, the presented data further support the role of sphingolipids and their metabolic enzymes in controlling the inflammatory response in CF

Evidence for the Involvement of Lipid Rafts and Plasma Membrane Sphingolipid Hydrolases in Pseudomonas aeruginosa Infection of Cystic Fibrosis Bronchial Epithelial Cells

Cystic fibrosis (CF) is the most common autosomal genetic recessive disease caused by mutations of gene encoding for the cystic fibrosis transmembrane conductance regulator. Patients with CF display a wide spectrum of symptoms, the most severe being chronic lung infection and inflammation, which lead to onset of cystic fibrosis lung disease. Several studies indicate that sphingolipids play a regulatory role in airway inflammation. The inhibition and downregulation of GBA2, the enzyme catabolizing glucosylceramide to ceramide, are associated with a significant reduction of IL-8 production in CF bronchial epithelial cells. Herein, we demonstrate that GBA2 plays a role in the proinflammatory state characterizing CF cells. We also report for the first time that Pseudomonas aeruginosa infection causes a recruitment of plasma membrane-associated glycosphingolipid hydrolases into lipid rafts of CuFi-1-infected cells. This reorganization of cell membrane may be responsible for activation of a signaling cascade, culminating in aberrant inflammatory response in CF bronchial epithelial cells upon bacterial infection. Taken together, the presented data further support the role of sphingolipids and their metabolic enzymes in controlling the inflammatory response in CF

GM1 as adjuvant of innovative therapies for cystic fibrosis disease

Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein is expressed at the apical plasma membrane (PM) of different epithelial cells. The most common mutation responsible for the onset of cystic fibrosis (CF), F508del, inhibits the biosynthesis and transport of the protein at PM, and also presents gating and stability defects of the membrane anion channel upon its rescue by the use of correctors and potentiators. This prompted a multiple drug strategy for F508delCFTR aimed simultaneously at its rescue, functional potentiation and PM stabilization. Since ganglioside GM1 is involved in the functional stabilization of transmembrane proteins, we investigated its role as an adjuvant to increase the effectiveness of CFTR modulators. According to our results, we found that GM1 resides in the same PM microenvironment as CFTR. In CF cells, the expression of the mutated channel is accompanied by a decrease in the PM GM1 content. Interestingly, by the exogenous administration of GM1, it becomes a component of the PM, reducing the destabilizing effect of the potentiator VX-770 on rescued CFTR protein expression/function and improving its stabilization. This evidence could represent a starting point for developing innovative therapeutic strategies based on the co-administration of GM1, correctors and potentiators, with the aim of improving F508del CFTR function

Drug Survival of Interleukin (IL)‑17 and IL‑23 Inhibitors for the Treatment of Psoriasis: A Retrospective Multi‑country, Multicentric Cohort Study

Background: Drug survival, defined as the length of time from initiation to discontinuation of a given therapy, allows comparisons between drugs, helps to predict patient's likelihood of remaining on a specific treatment, and achieving the best decision for each patient in daily clinical practice. Objective: The aim of this study was to provide data on drug survival of secukinumab, ixekizumab, brodalumab, guselkumab, tildrakizumab, and risankizumab in a large international cohort, and to identify clinical predictors that might have an impact on the drug survival of these drugs. Methods: This was a retrospective, multicentric, multi-country study that provides data of adult patients with moderate to severe psoriasis who started treatment with an interleukin (IL)-17 or IL-23 inhibitor between 1 February 2015 and 31 October 2021. Data were collected from 19 distinct hospital and non-hospital-based dermatology centers from Canada, Czech Republic, Italy, Greece, Portugal, Spain, and Switzerland. Kaplan-Meier estimator and proportional hazard Cox regression models were used for drug survival analysis. Results: A total of 4866 treatment courses (4178 patients)-overall time of exposure of 9500 patient-years-were included in this study, with 3164 corresponding to an IL-17 inhibitor (secukinumab, ixekizumab, brodalumab) and 1702 corresponding to an IL-23 inhibitor (guselkumab, risankizumab, tildrakizumab). IL-23 inhibitors had the highest drug survival rates during the entire study period. After 24 months of treatment, the cumulative probabilities of drug survival were 0.92 (95% confidence interval [CI] 0.89-0.95) for risankizumab, 0.90 (95% CI 0.88-0.92) for guselkumab, 0.80 (95% CI 0.76-0.84) for brodalumab, 0.79 (95% CI 0.76-0.82) for ixekizumab, and 0.75 (95% CI 0.73-0.77) for secukinumab. At 36 months, only guselkumab [0.88 (95% CI 0.85-0.91)], ixekizumab [0.73 (95% CI 0.70-0.76)], and secukinumab [0.67 (95% CI 0.65-0.70)] had more than 40 patients at risk of drug discontinuation. Only two drugs had more than 40 patients at risk of drug discontinuation at 48 months, with ixekizumab demonstrating to have a higher cumulative probability of drug survival [0.71 (95% CI 0.68-0.75)] when compared with secukinumab [0.63 (95% CI 0.60-0.66)]. Secondary failure was the main cause for drug discontinuation. According to the final multivariable model, patients receiving risankizumab, guselkumab, and ixekizumab were significantly less likely to discontinue treatment than those receiving secukinumab. Previous exposure to biologic agents, absent family history of psoriasis, higher baseline body mass index (BMI), and higher baseline Psoriasis Area and Severity Index (PASI) were identified as predictors of drug discontinuation. Conclusion: The cumulative probability of drug survival of both IL-17 and IL-23 inhibitors was higher than 75% at 24 months, with risankizumab and guselkumab demonstrating to have overall cumulative probabilities ≥ 90%. Biological agent chosen, prior exposure to biologic agents, higher baseline BMI and PASI values, and absence of family history of psoriasis were identified as predictors for drug discontinuation. Risankizumab, guselkumab, and ixekizumab were less likely to be discontinued than secukinumab

Effects of the scaffold proteins liprin-α1, β1 and β2 on invasion by breast cancer cells

Background InformationThe expression of the scaffold protein liprin-1 is upregulated in human breast cancer. This protein is part of a molecular network that is important for tumour cell invasion in vitro. Liprin-1 promotes invasion by supporting the protrusive activity at the leading edge of the migrating tumour cell and the degradation of the extracellular matrix by invadopodia. In this study, we have addressed the role of liprin-1 in the invasive process in vivo and of liprin-proteins in tumor cell motility.ResultsThe human tumour cell line MDA-MB-231 expresses liprin-1 and is able to promote the formation of metastasis in mice. Liprin- proteins may hetero-oligomerize with the members of the subfamily of the liprin- adaptor proteins. Analysis of the role of liprin-1 and liprin-2 has shown that while liprin-1 contributes positively to tumour cell motility in vitro; liprin-2 has a negative effect on both cell motility and invasion. Interestingly, we also observed differential effects on the ability of tumour cells to degrade the extracellular matrix, which is required for efficient invasion by tumour cells. In addition, analysis of the formation of lung metastases in vivo revealed that while the overexpression of liprin-1 in MDA-MB-231 cells did not evidently affect the metastatic process, silencing of the endogenous protein strongly impaired the formation of metastases by two independent invasion assays, without inhibiting the growth of primary tumours.ConclusionsOur data support an important role of distinct liprin family members in the regulation of tumour cell invasion, highlighting pro-invasive and anti-invasive effects by liprin-1 and liprin-2, respectively.SignificanceOur results indicate the importance of liprins in breast cancer cell invasion, and are expected to lead to future investigations on the mechanisms underlying the effects of distinct liprin proteins in different processes linked to tumor cell migration and invasion

Gangliosides and the Treatment of Neurodegenerative Diseases: A Long Italian Tradition

Gangliosides are glycosphingolipids which are particularly abundant in the plasma membrane of mammalian neurons. The knowledge of their presence in the human brain dates back to the end of 19th century, but their structure was determined much later, in the middle of the 1950s. From this time, neurochemical studies suggested that gangliosides, and particularly GM1 ganglioside, display neurotrophic and neuroprotective properties. The involvement of GM1 in modulating neuronal processes has been studied in detail by in vitro experiments, and the results indicated its direct role in modulating the activity of neurotrophin-dependent receptor signaling, the flux of calcium through the plasma membrane, and stabilizing the correct conformation of proteins, such as α-synuclein. Following, in vivo experiments supported the use of ganglioside drugs for the therapy of peripheral neuropathies, obtaining very positive results. However, the clinical use of gangliosides for the treatment of central neurodegeneration has not been followed due to the poor penetrability of these lipids at the central level. This, together with an ambiguous association (later denied) between ganglioside administration and Guillain-Barrè syndrome, led to the suspension of ganglioside drugs. In this critical review, we report on the evolution of research on gangliosides, on the current knowledge on the role played by gangliosides in regulating the biology of neurons, on the past and present use of ganglioside-based drugs used for therapy of peripheral neuropathies or used in human trials for central neurodegenerations, and on the therapeutic potential represented by the oligosaccharide chain of GM1 ganglioside for the treatment of neurodegenerative diseases

Treatment of Psoriasis Patients with Latent Tuberculosis Using IL-17 and IL-23 Inhibitors : A Retrospective, Multinational, Multicentre Study

Tuberculosis has a major global impact. Immunocompetent hosts usually control this disease, resulting in an asymptomatic latent tuberculosis infection (LTBI). Because TNF inhibitors increase the risk of tuberculosis reactivation, current guidelines recommend tuberculosis screening before starting any biologic drug, and chemoprophylaxis if LTBI is diagnosed. Available evidence from clinical trials and real-world studies suggests that IL-17 and IL-23 inhibitors do not increase the risk of tuberculosis reactivation. To evaluate psoriasis patients with treated or untreated newly diagnosed LTBI who received IL-17 and IL-23 inhibitors and the tolerability/safety of tuberculosis chemoprophylaxis. This is a retrospective, observational, multinational study from a series of 14 dermatology centres based in Portugal, Spain, Italy, Greece and Brazil, which included adult patients with moderate-to-severe chronic plaque psoriasis and newly diagnosed LTBI who were treated with IL-23 or IL-17 inhibitors between January 2015 and March 2022. LTBI was diagnosed in the case of tuberculin skin test and/or interferon gamma release assay positivity, according to local guideline, prior to initiating IL-23 or IL-17 inhibitor. Patients with prior diagnosis of LTBI (treated or untreated) or treated active infection were excluded. A total of 405 patients were included; complete/incomplete/no chemoprophylaxis was administered in 62.2, 10.1 and 27.7% of patients, respectively. The main reason for not receiving or interrupting chemoprophylaxis was perceived heightened risk of liver toxicity and hepatotoxicity, respectively. The mean duration of biological treatment was 32.87 ± 20.95 months, and only one case of active tuberculosis infection (ATBI) was observed, after 14 months of treatment with ixekizumab. The proportion of ATBI associated with ixekizumab was 1.64% [95% confidence interval (CI): 0-5.43%] and 0% for all other agents and 0.46% (95% CI 0-1.06%) and 0% for IL-17 and IL-23 inhibitors, respectively (not statistically significant). The risk of tuberculosis reactivation in patients with psoriasis and LTBI does not seem to increase with IL-17 or IL-23 inhibitors. IL-17 or IL-23 inhibitors should be preferred over TNF antagonists when concerns regarding tuberculosis reactivation exists. In patients with LTBI considered at high risk for developing complications related to chemoprophylaxis, this preventive strategy may be waived before initiating treatment with IL-17 inhibitors and especially IL-23 inhibitors

GM1 Oligosaccharide Crosses the Human Blood–Brain Barrier In Vitro by a Paracellular Route

International audienceGanglioside GM1 (GM1) has been reported to functionally recover degenerated nervous system in vitro and in vivo, but the possibility to translate GM1's potential in clinical settings is counteracted by its low ability to overcome the blood-brain barrier (BBB) due to its amphiphilic nature. Interestingly, the soluble and hydrophilic GM1-oligosaccharide (OligoGM1) is able to punctually replace GM1 neurotrophic functions alone, both in vitro and in vivo. In order to take advantage of OligoGM1 properties, which overcome GM1's pharmacological limitations, here we characterize the OligoGM1 brain transport by using a human in vitro BBB model. OligoGM1 showed a 20-fold higher crossing rate than GM1 and time-concentration-dependent transport. Additionally, OligoGM1 crossed the barrier at 4 °C and in inverse transport experiments, allowing consideration of the passive paracellular route. This was confirmed by the exclusion of a direct interaction with the active ATP-binding cassette (ABC) transporters using the "pump out" system. Finally, after barrier crossing, OligoGM1 remained intact and able to induce Neuro2a cell neuritogenesis by activating the TrkA pathway. Importantly, these in vitro data demonstrated that OligoGM1, lacking the hydrophobic ceramide, can advantageously cross the BBB in comparison with GM1, while maintaining its neuroproperties. This study has improved the knowledge about OligoGM1's pharmacological potential, offering a tangible therapeutic strategy

Biomolecular index of therapeutic efficacy in psoriasis treated by anti-TNF alpha agents.

BACKGROUND: Clinical or quality of life assessments are currently available for psoriasis severity evaluation and therapeutic response. Laboratory scores focused to measure and follow treatment efficacy are lacking at present. METHODS: Design a microscopic and biomolecular score to monitor skin disease severity and clinical response to anti-psoriatic treatments. A susceptibility gene analysis on cellular retinoic acid binding protein-II (CRABP-II), acting on keratinocyte differentiation, was also performed. A Molecular Index of Therapeutic Efficacy (MITE) was defined by assembling morphometric/semiquantitative measurement of epidermal thickness, immunohistochemical Ki-67, keratin17 and CRABP-II expression of lesional and non-lesional psoriatic skin biopsies before and after anti-tumor necrosis factor (TNF)α therapies. A 0-12 MITE score was correlated with Psoriasis Area and Severity Index (PASI)/Psoriasis Disability Index (PDI) scores and inflammation. Three CRABP- II SNPs were analyzed by TaqMan assay. RESULTS: All parameters were highly expressed in psoriatic lesions and reduced after 12 weeks of anti-TNFα treatments. MITE score strongly correlated with PASI and PDI values either before or after therapies (p<<0.001 and p<0.001, respectively). Conversely, MITE values did not change after treatments of non-responder patients. CRABP-II did not resulted a psoriatic susceptibility gene for the SNPs probes analyzed. CONCLUSIONS: MITE score variations corresponded to the patients' clinical improvement following anti-TNFα treatments, with significant statistical correlation among MITE, PASI and PDI scores. If confirmed in a larger series and/or in different hyperproliferative and inflammatory dermatoses, MITE score could be proposed as additional monitoring system to evaluate treatment protocols in skin disorders and targeted biomolecular pathways supporting clinical efficacy