Efficacy and safety of 4 weeks' treatment with combined fluticasone furoate/vilanterol in a single inhaler given once daily in COPD: a placebo-controlled randomised trial

Background: Fluticasone furoate/vilanterol (FF/VI) is a novel once-daily (OD) inhaled corticosteroid/long-acting beta(2) agonist combination in development for chronic obstructive pulmonary disease (COPD) and asthma. Trial design: A multicentre, randomised, double-blind, parallel-group, placebo-controlled study. Methods: Participants were patients with moderate-to-severe COPD treated with placebo or FF/VI 400/25 mu g OD for 4 weeks. Study objectives were to assess the safety and efficacy of FF/VI 400/25 mg OD administered for 4 weeks via a novel dry powder inhaler. Co-primary end points were change from baseline in weighted mean (wm) heart rate 0-4 h postdose at day 28 and the incidence of adverse events (AEs). Secondary end points included change from baseline in trough forced expiratory volume in one second (FEV1) (23-24 h postdose; day 29) and wm FEV1 (0-4 h postdose; day 28). Patients were randomised to receive FF/VI 400/25 mg or placebo in a 2: 1 ratio; all patients and investigators were blinded to active or placebo treatment. Results: 60 patients (mean age 64 years) were randomised (FF/VI: n=40; placebo: n=20), and all contributed data to the analysis. Mean screening post-bronchodilator FEV1 per cent predicted was comparable between groups (FF/VI: 58.5%; placebo: 60.1%). The wm heart rate 0-4 h postdose was similar between groups (difference: 0.6 beats per minute; 95% CI -3.9 to 5.1). More on-treatment AEs were reported in the FF/VI group (68%) compared with the placebo group (50%). The most common drug-related AEs in the FF/VI group were oral candidiasis (8%) and dysphonia (5%). There were no clinically relevant effects on laboratory values, including glucose and potassium, or on vital signs or ECGs/Holters. The FF/VI group had statistically greater improvements compared with placebo in trough FEV1 (mean difference 183 ml) and 0-4 h postdose wm FEV1 (mean difference 236 ml). Conclusion: FF/VI has a good safety and tolerability profile and improves lung function compared with placebo in patients with COPD. Trial registration number: clinical trials. gov-NCT00731822

Efficacy and safety of 4 weeks' treatment with combined fluticasone furoate/vilanterol in a single inhaler given once daily in COPD: a placebo-controlled randomised trial

Background. Fluticasone furoate/vilanterol (FF/VI) is a novel once-daily (OD) inhaled corticosteroid/long-acting β2 agonist combination in development for chronic obstructive pulmonary disease (COPD) and asthma. Trial design. A multicentre, randomised, double-blind, parallel-group, placebo-controlled study. Methods. Participants were patients with moderate-to-severe COPD treated with placebo or FF/VI 400/25 μg OD for 4 weeks. Study objectives were to assess the safety and efficacy of FF/VI 400/25 μg OD administered for 4 weeks via a novel dry powder inhaler. Co-primary end points were change from baseline in weighted mean (wm) heart rate 0–4 h postdose at day 28 and the incidence of adverse events (AEs). Secondary end points included change from baseline in trough forced expiratory volume in one second (FEV1) (23–24 h postdose; day 29) and wm FEV1 (0–4 h postdose; day 28). Patients were randomised to receive FF/VI 400/25 μg or placebo in a 2:1 ratio; all patients and investigators were blinded to active or placebo treatment. Results. 60 patients (mean age 64 years) were randomised (FF/VI: n=40; placebo: n=20), and all contributed data to the analysis. Mean screening post-bronchodilator FEV1 per cent predicted was comparable between groups (FF/VI: 58.5%; placebo: 60.1%). The wm heart rate 0–4 h postdose was similar between groups (difference: 0.6 beats per minute; 95% CI −3.9 to 5.1). More on-treatment AEs were reported in the FF/VI group (68%) compared with the placebo group (50%). The most common drug-related AEs in the FF/VI group were oral candidiasis (8%) and dysphonia (5%). There were no clinically relevant effects on laboratory values, including glucose and potassium, or on vital signs or ECGs/Holters. The FF/VI group had statistically greater improvements compared with placebo in trough FEV1 (mean difference 183 ml) and 0–4 h postdose wm FEV1 (mean difference 236 ml). Conclusion. FF/VI has a good safety and tolerability profile and improves lung function compared with placebo in patients with COPD.publishedVersio

Exosomes in the nose induce immune cell trafficking and harbour an altered protein cargo in chronic airway inflammation

Background: Exosomes are nano-sized extracellular vesicles participating in cell-to-cell communication both in health and disease. However, the knowledge about the functions and molecular composition of exosomes in the upper airways is limited. The aim of the current study was therefore to determine whether nasal exosomes can influence inflammatory cells and to establish the proteome of nasal lavage fluid-derived exosomes in healthy subjects, as well as its alterations in individuals with chronic airway inflammatory diseases [asthma and chronic rhinosinusitis (CRS)]. Methods: Nasal lavage fluid was collected from 14 healthy subjects, 15 subjects with asthma and 13 subjects with asthma/CRS. Exosomes were isolated with differential centrifugation and the proteome was analysed by LC-MS/MS with the application of two exclusion lists as well as using quantitative proteomics. Ingenuity Pathways Analysis and GO Term finder was used to predict the functions associated with the exosomal proteome and a migration assay was used to analyse the effect on immune cells by nasal exosomes. Results: Firstly, we demonstrate that nasal exosomes can induce migration of several immune cells, such as monocytes, neutrophils and NK cells in vitro. Secondly, a mass spectrometry approach, with the application of exclusion lists, was utilised to generate a comprehensive protein inventory of the exosomes from healthy subjects. The use of exclusion lists resulted in the identification of similar to 15 % additional proteins, and increased the confidence in similar to 20 % of identified proteins. In total, 604 proteins were identified in nasal exosomes and the nasal exosomal proteome showed strong associations with immune-related functions, such as immune cell trafficking. Thirdly, a quantitative proteomics approach was used to determine alterations in the exosome proteome as a result of airway inflammatory disease. Serum-associated proteins and mucins were more abundant in the exosomes from subjects with respiratory diseases compared to healthy controls while proteins with antimicrobial functions and barrier-related proteins had decreased expression. Conclusions: Nasal exosomes were shown to induce the migration of innate immune cells, which may be important as the airway epithelium is the first line of defence against pathogens and allergens. The decreased expression in barrier and antimicrobial exosomal proteins in subjects with airway diseases, could possibly contribute to an increased susceptibility to infections, which have important clinical implications in disease progression.11138Ysciescopu

Two distinct extracellular RNA signatures released by a single cell type identified by microarray and next-generation sequencing.

Cells secrete extracellular RNA (exRNA) to their surrounding environment and exRNA has been found in many body fluids such as blood, breast milk and cerebrospinal fluid. However, there are conflicting results regarding the nature of exRNA. Here, we have separated 2 distinct exRNA profiles released by mast cells, here termed high-density (HD) and low-density (LD) exRNA. The exRNA in both fractions was characterized by microarray and next-generation sequencing. Both exRNA fractions contained mRNA and miRNA, and the mRNAs in the LD exRNA correlated closely with the cellular mRNA, whereas the HD mRNA did not. Furthermore, the HD exRNA was enriched in lincRNA, antisense RNA, vault RNA, snoRNA, and snRNA with little or no evidence of full-length 18S and 28S rRNA. The LD exRNA was enriched in mitochondrial rRNA, mitochondrial tRNA, tRNA, piRNA, Y RNA, and full-length 18S and 28S rRNA. The proteomes of the HD and LD exRNA-containing fractions were determined with LC-MS/MS and analyzed with Gene Ontology term finder, which showed that both proteomes were associated with the term extracellular vesicles and electron microscopy suggests that at least a part of the exRNA is associated with exosome-like extracellular vesicles. Additionally, the proteins in the HD fractions tended to be associated with the nucleus and ribosomes, whereas the LD fraction proteome tended to be associated with the mitochondrion.We show that the 2 exRNA signatures released by a single cell type can be separated by floatation on a density gradient. These results show that cells can release multiple types of exRNA with substantial differences in RNA species content. This is important for any future studies determining the nature and function of exRNA released from different cells under different conditions.112212Ysciescopu

Acidic microenvironment plays a key role in human melanoma progression through a sustained exosome mediated transfer of clinically relevant metastatic molecules

Background: Microenvironment cues involved in melanoma progression are largely unknown. Melanoma is highly influenced in its aggressive phenotype by the changes it determinates in its microenvironment, such as pH decrease, in turn influencing cancer cell invasiveness, progression and tissue remodelling through an abundant secretion of exosomes, dictating cancer strategy to the whole host. A role of exosomes in driving melanoma progression under microenvironmental acidity was never described. Methods: We studied four differently staged human melanoma lines, reflecting melanoma progression, under microenvironmental acidic pHs pressure ranging between pH 6.0-6.7. To estimate exosome secretion as a function of tumor stage and environmental pH, we applied a technique to generate native fluorescent exosomes characterized by vesicles integrity, size, density, markers expression, and quantifiable by direct FACS analysis. Functional roles of exosomes were tested in migration and invasion tests. Then we performed a comparative proteomic analysis of acid versus control exosomes to elucidate a specific signature involved in melanoma progression. Results: We found that metastatic melanoma secretes a higher exosome amount than primary melanoma, and that acidic pH increases exosome secretion when melanoma is in an intermediate stage, i.e. metastatic non-invasive. We were thus able to show that acidic pH influences the intercellular cross-talk mediated by exosomes. In fact when exposed to exosomes produced in an acidic medium, pH naïve melanoma cells acquire migratory and invasive capacities likely due to transfer of metastatic exosomal proteins, favoring cell motility and angiogenesis. A Prognoscan-based meta-analysis study of proteins enriched in acidic exosomes, identified 11 genes (HRAS, GANAB, CFL2, HSP90B1, HSP90AB1, GSN, HSPA1L, NRAS, HSPA5, TIMP3, HYOU1), significantly correlating with poor prognosis, whose high expression was in part confirmed in bioptic samples of lymph node metastases. Conclusions: A crucial step of melanoma progression does occur at melanoma intermediate -stage, when extracellular acidic pH induces an abundant release and intra-tumoral uptake of exosomes. Such exosomes are endowed with pro-invasive molecules of clinical relevance, which may provide a signature of melanoma advancement

Control of Allergic Rhinitis and Asthma Test (CARAT) can be used to assess individual patients over time

BACKGROUND: The Control of Allergic Rhinitis and Asthma Test (CARAT10) has been proposed as the first tool to implement the Allergic Rhinitis and its Impact on Asthma initiative guidelines in clinical practice. To serve this purpose, it must have adequate properties to assess the control of an individual over time. This study aimed to prospectively assess the test-retest reliability, responsiveness and longitudinal validity of CARAT10. METHODS: Adults with asthma and allergic rhinitis were enrolled at 4 outpatient clinics of Portuguese central hospitals. At each of the two visits, 4 to 6 weeks apart, patients filled out CARAT10 and additional questionnaires, followed by a medical evaluation blinded to the questionnaires' answers. RESULTS: From the 62 patients included, 51 patients completely filled out CARAT10 at both visits. The test-retest reliability, computed as an intra-class correlation coefficient, was 0.82. Regarding responsiveness, a significant change (p = 0.002) of CARAT10 score in clinically unstable patients was observed (95%CI -5.08; -1.31) and the Guyatt's responsiveness index was 1.54. As for the longitudinal validity assessment, the correlation coefficients of the changes of CARAT10 scores with those of ACQ5 and symptoms VAS ranged from 0.49 to 0.65, while with the physician assessment of control they ranged from 0.31 to 0.41. CONCLUSION: CARAT10 has good test-retest reliability, responsiveness and longitudinal validity. It can be used to assess control of allergic rhinitis and asthma, both to compare groups in clinical studies and to evaluate individual patients in clinical practice

Microguards and micromessengers of the genome

The regulation of gene expression is of fundamental importance to maintain organismal function and integrity and requires a multifaceted and highly ordered sequence of events. The cyclic nature of gene expression is known as ‘transcription dynamics’. Disruption or perturbation of these dynamics can result in significant fitness costs arising from genome instability, accelerated ageing and disease. We review recent research that supports the idea that an important new role for small RNAs, particularly microRNAs (miRNAs), is in protecting the genome against short-term transcriptional fluctuations, in a process we term ‘microguarding’. An additional emerging role for miRNAs is as ‘micromessengers’—through alteration of gene expression in target cells to which they are trafficked within microvesicles. We describe the scant but emerging evidence that miRNAs can be moved between different cells, individuals and even species, to exert biologically significant responses. With these two new roles, miRNAs have the potential to protect against deleterious gene expression variation from perturbation and to themselves perturb the expression of genes in target cells. These interactions between cells will frequently be subject to conflicts of interest when they occur between unrelated cells that lack a coincidence of fitness interests. Hence, there is the potential for miRNAs to represent both a means to resolve conflicts of interest, as well as instigate them. We conclude by exploring this conflict hypothesis, by describing some of the initial evidence consistent with it and proposing new ideas for future research into this exciting topic

Distinguishing Asthma Phenotypes Using Machine Learning Approaches.

Asthma is not a single disease, but an umbrella term for a number of distinct diseases, each of which are caused by a distinct underlying pathophysiological mechanism. These discrete disease entities are often labelled as asthma endotypes. The discovery of different asthma subtypes has moved from subjective approaches in which putative phenotypes are assigned by experts to data-driven ones which incorporate machine learning. This review focuses on the methodological developments of one such machine learning technique-latent class analysis-and how it has contributed to distinguishing asthma and wheezing subtypes in childhood. It also gives a clinical perspective, presenting the findings of studies from the past 5 years that used this approach. The identification of true asthma endotypes may be a crucial step towards understanding their distinct pathophysiological mechanisms, which could ultimately lead to more precise prevention strategies, identification of novel therapeutic targets and the development of effective personalized therapies

Comparing the effects of two inhaled glucocorticoids on allergen-induced bronchoconstriction and markers of systemic effects, a randomised cross-over double-blind study

<p>Abstract</p> <p>Background</p> <p>Inhaled glucocorticoids are efficient in protecting against asthma exacerbations, but methods to compare their efficacy vs systemic effects have only been attempted in larger multi-centre studies. The aim of the current study was therefore to directly compare the effects of two separate inhaled glucocorticoids, mometasone and budesonide, to compare the effects on the early and late asthmatic responses to inhaled allergen in patients with mild allergic asthma, and sputum eosinophils, and to relate the clinical positive effects to any systemic effects observed.</p> <p>Methods</p> <p>Twelve patients with documented early and late asthmatic responses (EAR and LAR) to inhaled allergen at a screening visit were randomized in a double-blind fashion to treatment with mometasone (200 μg × 2 or 400 μg × 2), budesonide (400 μg × 2) or placebo in a double-blind crossover fashion for a period of seven days. Challenge with the total allergen dose causing both an EAR and LAR was given on the last day of treatment taken in the morning. Lung function was assessed using FEV1, and systemic glucocorticoid activity was quantified using 24 h urinary cortisol.</p> <p>Results</p> <p>Mometasone and budesonide attenuate both EAR and LAR to allergen to a similar degree. No significant dose-related effects on the lung function parameters were observed. Both treatments reduced the relative amount of sputum eosinophils (%) after allergen. At the dose of 800 μg daily, mometasone reduced 24 h urinary cortisol by approximately 35%. Both drugs were well tolerated.</p> <p>Conclusions</p> <p>Mometasone and budesonide are equieffective in reducing early and late asthmatic responses induced by inhaled allergen challenge. Mometasone 800 μg given for seven days partially affects the HPA axis.</p