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Objective: Bronchial thermoplasty is an effective intervention to improve respiratory symptoms and to reduce the rate of exacerbations in uncontrolled severe asthma. A reduction in airway smooth muscle is arguably the most widely discussed mechanisms accounting for these clinical benefits. Yet, this smooth muscle reduction should also translate into an impaired response to bronchodilator drugs. A pilot study was designed to address this question. Hypothesis: Thermoplasty decreases the response to a bronchodilator. Methods: Lung functions measured by spirometry and respiratory mechanics measured by oscillometry (tremoFlo, Thorasys) were examined pre- and post-bronchodilator (salbutamol, 400 μg) before and after at least 1 year of thermoplasty in 8 patients (7 women) with uncontrolled severe asthma. For each readout, a two-way ANOVA was used to assess the effect of thermoplasty, salbutamol, and their interaction. All procedures were approved by the IUCPQ Ethics Committee. Data: Consistent with previous studies, thermoplasty yielded no benefits in terms of baseline lung function and respiratory mechanics, despite improving symptoms based on two questionnaires assessing the level of asthma control (Asthma Control Questionnaire – ACQ-5, and Asthma Control Test – ACT-5). The response to salbutamol was not affected by thermoplasty based on spirometric readouts, including forced expiratory volume in 1 s (FEV1), forced vital capacity (FVC) and FEV1/FVC ratio. However, a significant interaction was observed between thermoplasty and salbutamol for two oscillometric readouts. The reactance of the respiratory system at 5 Hz (Xrs5) increased from -3.2 ±2.6 to -1.9 ± 1.9 cmH2O ·s/L in response to salbutamol pre-thermoplasty, versus from -2.8 ± 2.3 to -2.2 ± 1.9 cmH2O·s/L post-thermoplasty (interaction, p=0.04). Similarly, the reactance area (Ax) decreased from 32.6 ± 29.0 to 12.8 ±18.2 cmH2O/L in response to salbutamol pre-thermoplasty, versus 27.2 ± 29.4 to 15.7 ± 20.1 cmH2O/L post-thermoplasty (interaction, p=0.02). The salbutamol-induced changes in resistance at 5 Hz (Rrs5), resistance at 19 Hz (Rrs19), Rrs5 minus Rrs19, and resonant frequency were not significantly affected by thermoplasty. Summary of results: While spirometry was unable to detect changes in the response to salbutamol caused by thermoplasty, two oscillometric readouts, namely Xrs5 and Ax, confirmed that the bronchodilator response is attenuated post-thermoplasty. Conclusion: Despite its small size, this study demonstrates that thermoplasty attenuates the response to a bronchodilator. We argue that this result is a physiological proof of therapeutic efficacy, consistent with the well-described effect of thermoplasty in reducing the amount of airway smooth muscle. FQSR (Fondation Quebecoise en Sante Respiratoire); Canadian Institutes of Health Research (PJT-387910); FRQS (Fonds de recherche du Quebec - Sante); and Boston Scientific Inc This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Background Bronchial thermoplasty is a device-based treatment for subjects ≥18 years with severe asthma poorly controlled with inhaled corticosteroids and long-acting beta-agonists. The Post-FDA Approval Clinical Trial Evaluating Bronchial Thermoplasty in Severe Persistent Asthma (PAS2) study collected data on severe asthmatics undergoing this procedure. Research Question What are the 5-year efficacy and safety results in severe asthmatics who have undergone bronchial thermoplasty? Study Design and Methods This was a prospective, open-label, observational, multi-center study conducted in the United States and Canada. Subjects aged 18-65, taking inhaled corticosteroids ≥1000μg/day (beclomethasone or equivalent) and long-acting β-agonists ≥80μg/day (salmeterol or equivalent) were included. Severe exacerbations, hospitalization, emergency department visits, and medication usage were evaluated for the 12 months prior to and at years 1-5 post-treatment. Spirometry was evaluated at baseline and at years 1-5 post-treatment. Results 284 subjects were enrolled at 27 centers; 227 subjects (80%) completed 5 years of follow-up. By year 5 post-treatment, the proportion of subjects with severe exacerbations, emergency department visits, and hospitalizations was 42.7%, 7.9%, and 4.8%, respectively, compared to 77.8%, 29.4%, and 16.1% in the 12 months prior to treatment. The proportion of subjects on maintenance oral corticosteroids decreased from 19.4% at baseline to 9.7% at 5 years. Analyses of subgroups based on baseline clinical and biomarker characteristics revealed a statistically significant clinical improvement among all subgroups. Interpretation Five years after treatment, subjects experienced decreases in severe exacerbations, hospitalizations, emergency department visits and corticosteroid exposure. All subgroups demonstrated clinically significant improvement, suggesting that bronchial thermoplasty improves asthma control in different asthma phenotypes.

PDF file - 589K, Probe sets significantly associated with smoking in the discovery set

PDF file - 242K, Table S2. Clinical characteristics of patients in the first replication set (UBC). Table S3. Clinical characteristics of patients in the second replication set (Groningen). Figure S1. Volcano plots showing the impact of smoking, COPD, and lung cancer on gene expression in the lung. Figure S2. Comparison of gene expression recovery following smoking cessation between the discovery set (Laval) and UBC. Figure S3. Comparison of gene expression recovery following smoking cessation between the discovery set (Laval) and Groningen. Figure S4. Expression of SERPIND1 in lung parenchyma of smoker and never-smoker by immunochemistry. Figure S5. Enrichment plot for slowly reversible gene in UBC. Figure S6. Enrichment plot for slowly reversible gene in Groningen

The endocannabinoids 2-arachidonoyl-glycerol and N-arachidonoyl-ethanolamine are lipids regulating many physiological processes, notably inflammation. Endocannabinoid hydrolysis inhibitors are now being investigated as potential anti-inflammatory agents. In addition to 2-arachidonoyl-glycerol and N-arachidonoyl-ethanolamine, the endocannabinoidome also includes other monoacylglycerols and N-acyl-ethanolamines such as 1-linoleoyl-glycerol (1-LG) and N-linoleoyl-ethanolamine (LEA). By increasing monoacylglycerols and/or N-acyl-ethanolamine levels, endocannabinoid hydrolysis inhibitors will likely increase the levels of their metabolites. Herein, we investigated whether 1-LG and LEA were substrates for the 15-lipoxygenase pathway, given that both possess a 1Z,4Z-pentadiene motif, near their omega end. We thus assessed how human eosinophils and neutrophils biosynthesized the 15-lipoxygenase metabolites of 1-LG and LEA. Linoleic acid (LA), a well-documented substrate of 15-lipoxygenases, was used as positive control. N-13-hydroxy-octodecadienoyl-ethanolamine (13-HODE-EA) and 13-hydroxy-octodecadienoyl-glycerol (13-HODE-G), the 15-lipoxygenase metabolites of LEA and 1-LG, were synthesized using Novozym 435 and soybean lipoxygenase. Eosinophils, which express the 15-lipoxygenase-1, metabolized LA, 1-LG, and LEA into their 13-hydroxy derivatives. This was almost complete after five minutes. Substrate preference of eosinophils was LA > LEA > 1-LG in presence of 13-HODE-G hydrolysis inhibition with methyl-arachidonoyl-fluorophosphonate. Human neutrophils also metabolized LA, 1-LG, and LEA into their 13-hydroxy derivatives. This was maximal after 15–30 s. Substrate preference was LA ≫ 1-LG > LEA. Importantly, 13-HODE-G was found in humans and mouse tissue samples. In conclusion, our data show that human eosinophils and neutrophils metabolize 1-LG and LEA into the novel endogenous 15-lipoxygenase metabolites 13-HODE-G and 13-HODE-EA. The full biological importance of 13-HODE-G and 13-HODE-EA remains to be explored.

2-Arachidonoyl-glycerol (2-AG) is an endocannabinoid with anti-inflammatory properties. Blocking 2-AG hydrolysis to enhance CB2 signaling has proven effective in mouse models of inflammation. However, the expression of 2-AG lipases has never been thoroughly investigated in human leukocytes. Herein, we investigated the expression of seven 2-AG hydrolases by human blood leukocytes and alveolar macrophages (AMs) and found the following protein expression pattern: monoacylglycerol (MAG lipase; eosinophils, AMs, monocytes), carboxylesterase (CES1; monocytes, AMs), palmitoyl-protein thioesterase (PPT1; AMs), α/β-hydrolase domain (ABHD6; mainly AMs), ABHD12 (all), ABHD16A (all), and LYPLA2 (lysophospholipase 2; monocytes, lymphocytes, AMs). We next found that all leukocytes could hydrolyze 2-AG and its metabolites derived from cyclooxygenase-2 (prostaglandin E2-glycerol [PGE2-G]) and the 15-lipoxygenase (15-hydroxy-eicosatetraenoyl-glycerol [15-HETE-G]). Neutrophils and eosinophils were consistently better at hydrolyzing 2-AG and its metabolites than monocytes and lymphocytes. Moreover, the efficacy of leukocytes to hydrolyze 2-AG and its metabolites was 2-AG ≥ 15-HETE-G >> PGE2-G for each leukocyte. Using the inhibitors methylarachidonoyl-fluorophosphonate (MAFP), 4-nitrophenyl-4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate (JZL184), Palmostatin B, 4′-carbamoylbiphenyl-4-yl methyl(3-(pyridin-4-yl)benzyl)carbamate, N-methyl-N-[[3-(4-pyridinyl)phenyl]methyl]-4′-(aminocarbonyl)[1,1′-biphenyl]-4-yl ester carbamic acid (WWL70), 4′-[[[methyl[[3-(4-pyridinyl)phenyl]methyl]amino]carbonyl]oxy]-[1,1′-biphenyl]-4-carboxylic acid, ethyl ester (WWL113), tetrahydrolipstatin, and ML349, we could not pinpoint a specific hydrolase responsible for the hydrolysis of 2-AG, PGE2-G, and 15-HETE-G by these leukocytes. Furthermore, JZL184, a selective MAG lipase inhibitor, blocked the hydrolysis of 2-AG, PGE2-G, and 15-HETE-G by neutrophils and the hydrolysis of PGE2-G and 15-HETE-G by lymphocytes, two cell types with limited/no MAG lipase. Using an activity-based protein profiling (ABPP) probe to label hydrolases in leukocytes, we found that they express many MAFP-sensitive hydrolases and an unknown JZL184-sensitive hydrolase of ∼52 kDa. Altogether, our results indicate that human leukocytes are experts at hydrolyzing 2-AG and its metabolites via multiple lipases and probably via a yet-to-be characterized 52 kDa hydrolase. Blocking 2-AG hydrolysis in humans will likely abrogate the ability of human leukocytes to degrade 2-AG and its metabolites and increase their anti-inflammatory effects in vivo.

Background Bronchial thermoplasty (BT) is a novel technique used in the treatment of subjects with severe refractory asthma. Radiofrequency is provided to airway walls during bronchoscopy in order to reduce airway remodeling. Several clinical studies have reported an improvement in subjects’ symptoms following BT. However, how BT affects the airway architectures and inflammatory mediators in the airways has not been yet fully elucidated. Methods Fourteen subjects with severe asthma were recruited in this study according to the criteria of ATS severe asthma definition. The study subjects undertook bronchial biopsy during the bronchoscopy procedure at baseline and 6 weeks after the initial BT treatment. The obtained samples were stained with antibodies for α-smooth muscle actin (α-SMA); protein gene product (PGP) 9.5, a specific nerve marker; von Willebrand factor (vWF), a marker for blood vessels; interleukin-17A (IL-17A) and transforming growth factor-β1 (TGF-β1). Results The expression of α-SMA and PGP9.5 were significantly reduced post-BT. There was no significant difference in the number of blood vessels between baseline and post-BT. In addition, BT did not affect the production of IL-17A and TGF-β1 in the airways. The changes in the expression of α-SMA and PGP9.5 had no significant correlation with the improvement of pulmonary function. Conclusion and Clinical Relevance: This study suggests that BT reduces airway smooth muscle mass and the airway innervation without affecting vasculature and the production of inflammatory mediators in the airways of subjects with severe asthma.

Leukotriene B4 (LTB4) plays a prominent role in innate immunity as it induces phagocyte recruitment, the release of antimicrobial effectors, and as it potentiates the ingestion and killing of pathogens. In humans, LTB4 has a short half-life and is rapidly metabolized by leukocytes, notably into 20-OH- and 20-COOH-LTB4 by neutrophils. Although these LTB4 metabolites bind to the BLT1 receptor with high affinity, they activate neutrophils to a much lower extent than LTB4. We thus postulated that LTB4 metabolites could dampen BLT1-mediated responses, therefore limiting the impact of LTB4 on human neutrophil functions. We found that 20-OH-LTB4 and 20-COOH-LTB4 inhibited all of the LTB4-mediated neutrophil responses we tested (migration, degranulation, leukotriene biosynthesis). The potencies of the different compounds at inhibiting LTB4-mediated responses were 20-OH-LTB4 = CP 105,696 (BLT1 antagonist) > > 20-COOH-LTB4 ≥ resolvin E1 (RVE1). In contrast, the fMLP- and IL-8-mediated responses we tested were not affected by the LTB4 metabolites or RVE1. 20-OH-LTB4 and 20-COOH-LTB4 also inhibited the LTB4-mediated migration of human eosinophils but not that induced by 5-KETE. Moreover, using 20-COOH-LTB4, LTB4, and LTB4-alkyne, we show that LTB4 is a chemotactic, rather than a chemokinetic factor for both human neutrophils and eosinophils. In conclusion, our data indicate that LTB4 metabolites and RVE1 act as natural inhibitors of LTB4-mediated responses. Thus, preventing LTB4 ω-oxidation might result in increased innate immunity and granulocyte functions.

Bronchial thermoplasty is a device-based treatment for subjects ≥ 18 years of age with severe asthma poorly controlled with inhaled corticosteroids and long-acting beta-agonists. The Post-FDA Approval Clinical Trial Evaluating Bronchial Thermoplasty in Severe Persistent Asthma (PAS2) study collected data on patients with severe asthma undergoing this procedure.What are the 5-year efficacy and safety results in patients with severe asthma who have undergone bronchial thermoplasty?This was a prospective, open-label, observational, multicenter study conducted in the United States and Canada. Subjects 18 to 65 years of age who were taking inhaled corticosteroids ≥ 1,000 μg/d (beclomethasone or equivalent) and long-acting beta-agonists ≥ 80 μg/d (salmeterol or equivalent) were included. Severe exacerbations, hospitalization, ED visits, and medication usage were evaluated for the 12 months prior to and at years 1 through 5 posttreatment. Spirometry was evaluated at baseline and at years 1 through 5 posttreatment.A total of 284 subjects were enrolled at 27 centers; 227 subjects (80%) completed 5 years of follow-up. By year 5 posttreatment, the proportion of subjects with severe exacerbations, ED visits, and hospitalizations was 42.7%, 7.9%, and 4.8%, respectively, compared with 77.8%, 29.4%, and 16.1% in the 12 months prior to treatment. The proportion of subjects on maintenance oral corticosteroids decreased from 19.4% at baseline to 9.7% at 5 years. Analyses of subgroups based on baseline clinical and biomarker characteristics revealed a statistically significant clinical improvement among all subgroups.Five years after treatment, subjects experienced decreases in severe exacerbations, hospitalizations, ED visits, and corticosteroid exposure. All subgroups demonstrated clinically significant improvement, suggesting that bronchial thermoplasty improves asthma control in different asthma phenotypes.ClinicalTrials.gov; No.: NCT01350336; URL: www.gov.

Severe acute respiratory syndrome coronavirus 2 is responsible for coronavirus disease 2019 (COVID‐19). While COVID‐19 is often benign, a subset of patients develops severe multilobar pneumonia that can progress to an acute respiratory distress syndrome. There is no cure for severe COVID‐19 and few treatments significantly improved clinical outcome. Dexamethasone and possibly aspirin, which directly/indirectly target the biosynthesis/effects of numerous lipid mediators are among those options. Our objective was to define if severe COVID‐19 patients were characterized by increased bioactive lipids modulating lung inflammation. A targeted lipidomic analysis of bronchoalveolar lavages (BALs) by tandem mass spectrometry was done on 25 healthy controls and 33 COVID‐19 patients requiring mechanical ventilation. BALs from severe COVID‐19 patients were characterized by increased fatty acids and inflammatory lipid mediators. There was a predominance of thromboxane and prostaglandins. Leukotrienes were also increased, notably LTB4, LTE4, and eoxin E4. Monohydroxylated 15‐lipoxygenase metabolites derived from linoleate, arachidonate, eicosapentaenoate, and docosahexaenoate were also increased. Finally yet importantly, specialized pro‐resolving mediators, notably lipoxin A4 and the D‐series resolvins, were also increased, underscoring that the lipid mediator storm occurring in severe COVID‐19 involves pro‐ and anti‐inflammatory lipids. Our data unmask the lipid mediator storm occurring in the lungs of patients afflicted with severe COVID‐19. We discuss which clinically available drugs could be helpful at modulating the lipidome we observed in the hope of minimizing the deleterious effects of pro‐inflammatory lipids and enhancing the effects of anti‐inflammatory and/or pro‐resolving lipid mediators.

BACKGROUNDSevere Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) is the infectious agent responsible for Coronavirus disease 2019 (COVID-19). While SARS-CoV-2 infections are often benign, there are also severe COVID-19 cases, characterized by severe bilobar pneumonia that can decompensate to an acute respiratory distress syndrome, notably characterized by increased inflammation and a cytokine storm. While there is no cure against severe COVID-19 cases, some treatments significantly decrease the severity of the disease, notably aspirin and dexamethasone, which both directly or indirectly target the biosynthesis (and effects) of numerous bioactive lipids.OBJECTIVEOur working hypothesis was that severe COVID-19 cases necessitating mechanical ventilation were characterized by increased bioactive lipid levels modulating lung inflammation. We thus quantitated several lung bioactive lipids using liquid chromatography combined to tandem mass spectrometry.RESULTSWe performed an exhaustive assessment of the lipid content of bronchoalveolar lavages from 25 healthy controls and 33 COVID-19 patients necessitating mechanical ventilation. Severe COVID-19 patients were characterized by increased fatty acid levels as well as an accompanying inflammatory lipid storm. As such, most quantified bioactive lipids were heavily increased. There was a predominance of cyclooxygenase metabolites, notably TXB2 >> PGE2 ∼ 12-HHTrE > PGD2. Leukotrienes were also increased, notably LTB4, 20-COOH-LTB4, LTE4, and eoxin E4. 15-lipoxygenase metabolites derived from linoleic, arachidonic, eicosapentaenoic and docosahexaenoic acids were also increased. Finally, yet importantly, specialized pro-resolving mediators, notably lipoxin A4 and the D-series resolvins, were also found at important levels, underscoring that the lipid storm occurring in severe SARS-CoV-2 infections involves pro- and anti-inflammatory lipids.CONCLUSIONSOur data unmask the important lipid storm occurring in the lungs of patients afflicted with severe COVID-19. We discuss which clinically available drugs could be helpful at modulating the lipidome we observed in the hope of minimizing the deleterious effects of pro-inflammatory lipids and enhancing the effects of anti-inflammatory and/or pro-resolving lipids.

To identify susceptibility loci and candidate causal genes of asthma, we performed a genome-wide association study (GWAS) in UK Biobank on a broad asthma definition (n = 56,167 asthma cases and 352,255 controls). We then carried out functional mapping through transcriptome-wide association studies (TWAS) and Mendelian randomization in lung (n = 1,038) and blood (n = 31,684) tissues. The GWAS revealed 72 asthma-associated loci from 116 independent significant variants (PGWAS 20) including a stop-gain mutation in the filaggrin (FLG) gene. The top lung TWAS gene on 17q12-q21 was GSDMB (PTWAS=1.42E-54). Other TWAS genes of interest include TSLP on 5q22, RERE on 1p36, CLEC16A on 16p13, and IL4R on 16p12, which all replicated in GTEx lung (n = 515). A novel risk locus was also revealed by the lung asthma TWAS on 1q23.3 with the putative gene encoding the gamma chain of the high-affinity IgE receptor (FCER1G, PTWAS=2.13E-6), which was also replicated in GTEx lung (PTWAS=3.71E-7). By testing a comprehensive set of cells and tissues, we then demonstrated that the largest fold enrichment of regulatory and functional annotations among asthma-associated variants was in the blood. We mapped 485 eQTL-regulated genes associated with asthma in the blood and 50 of them were shown to be causally associated with asthma by Mendelian randomization. Prioritization of druggable genes revealed known (IL4R, TSLP, IL6, TNFSF4) and potentially new therapeutic targets for asthma.

To identify candidate causal genes of asthma, we performed a genome-wide association study (GWAS) in UK Biobank on a broad asthma definition (n = 56,167 asthma cases and 352,255 controls). We then carried out functional mapping through transcriptome-wide association studies (TWAS) and Mendelian randomization in lung (n = 1,038) and blood (n = 31,684) tissues. The GWAS reveals 72 asthma-associated loci from 116 independent significant variants (PGWAS, Kim Valette et al. perform a genomic study on asthma integrating genome-wide association study, functional mapping using lung and blood transcriptome-wide profiles, as well as Mendelian randomization. They show candidate causal genes expressed in lung and blood tissues that are putative therapeutic targets for asthma.

Aim: This study aimed to characterize DNA methylation (DNA-me) in promoter region of IL33, IL1RL1 and CCL26 in asthma and their impacts on transcriptional activity in bronchial epithelial cells (BECs). Patients & methods: We performed bis-pyrosequencing, quantitative real-time PCR and sequencing in BECs from ten asthmatic and ten control individuals. Results: We detected lower DNA-me levels of IL33 and CCL26 in asthmatic than control BECs. No correlation was found between methylation and expression levels. Interestingly, carriers of a mutative allele in a haplotype within the promoter of IL33 had a lower IL33 DNA-me level and CCL26 gene expression correlated with eosinophil count. Conclusion: These findings highlight the importance of investigating both epigenetic and genetic mechanisms in understanding the epithelial immune response in asthma.

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Background Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection can lead to a variety of clinical outcomes ranging from the absence of symptoms to severe acute respiratory disease and ultimately death. A feature of patients with severe Coronavirus disease 19 (COVID-19) is the abundance of inflammatory cytokines in the blood. Elevated levels of cytokine are predictive of infection severity and clinical outcome. In contrast, studies aimed at defining the driving forces behind the inflammation in lungs of severed COVID-19 subjects remain scarce. Objective To analyze and compare the plasma and bronchoalveolar lavages (BALs) of patients with severe COVID-19 (n=45) for the presence of cytokines and lipid mediators of inflammation (LMI). Methods Cytokines were measured using Luminex multiplex assay and LMI measured using LC-MS/MS. Results We reveal high concentrations of numerous cytokines/chemokines/LMI in BALs of patients with severe COVID-19. Among the 13 most abundant mediators in BALs, 11 were chemokines with CXCL1 and CXCL8 being 200 times more abundant than IL-6 and TNF-α. Eicosanoids were also elevated in lungs of severe COVID-19 subjects. Consistent with the presence chemotactic molecules, BALs were enriched for neutrophils, lymphocytes and eosinophils. Inflammatory cytokines/LMI in plasma showed limited correlations with those present in BALs, arguing that circulating inflammatory molecules may not be a reliable proxy of the inflammation occurring in the lungs of severe COVID-19 patients. Conclusions Our findings indicate that lungs hyperinflammation of severe COVID-19 patients is fueled by excessive production of chemokines and eicosanoids. Therapeutic strategies to dampen inflammation in COVID-19 patients should be tailored accordingly., Graphical abstract

Background The Airways Disease Endotyping for Personalized Therapeutics (ADEPT) study profiled patients with mild, moderate, and severe asthma and nonatopic healthy control subjects. Objective We explored this data set to define type 2 inflammation based on airway mucosal IL-13–driven gene expression and how this related to clinically accessible biomarkers. Methods IL-13–driven gene expression was evaluated in several human cell lines. We then defined type 2 status in 25 healthy subjects, 28 patients with mild asthma, 29 patients with moderate asthma, and 26 patients with severe asthma based on airway mucosal expression of (1) CCL26 (the most differentially expressed gene), (2) periostin, or (3) a multigene IL-13 in vitro signature (IVS). Clinically accessible biomarkers included fraction of exhaled nitric oxide (Feno) values, blood eosinophil (bEOS) counts, serum CCL26 expression, and serum CCL17 expression. Results Expression of airway mucosal CCL26, periostin, and IL-13–IVS all facilitated segregation of subjects into type 2–high and type 2–low asthmatic groups, but in the ADEPT study population CCL26 expression was optimal. All subjects with high airway mucosal CCL26 expression and moderate-to-severe asthma had Feno values (≥35 ppb) and/or high bEOS counts (≥300 cells/mm 3 ) compared with a minority (36%) of subjects with low airway mucosal CCL26 expression. A combination of Feno values, bEOS counts, and serum CCL17 and CCL26 expression had 100% positive predictive value and 87% negative predictive value for airway mucosal CCL26–high status. Clinical variables did not differ between subjects with type 2–high and type 2–low status. Eosinophilic inflammation was associated with but not limited to airway mucosal type 2 gene expression. Conclusion A panel of clinical biomarkers accurately classified type 2 status based on airway mucosal CCL26, periostin, or IL-13–IVS gene expression. Use of Feno values, bEOS counts, and serum marker levels (eg, CCL26 and CCL17) in combination might allow patient selection for novel type 2 therapeutics.

The endocannabinoids 2-arachidonoyl-glycerol and N-arachidonoyl-ethanolamine mediate an array of pro- and anti-inflammatory effects. These effects are related, in part, to their metabolism by eicosanoid biosynthetic enzymes. For example, N-arachidonoyl-ethanolamine and 2-arachidonoyl-glycerol can be metabolized by cyclooxygenase-2 into PG-ethanolamide (PG-EA) and PG-glycerol (PG-G), respectively. Although PGE2 is a recognized suppressor of neutrophil functions, the impact of cyclooxygenase-derived endocannabinoids such as PGE2-EA or PGE2-G on neutrophils is unknown. This study’s aim was to define the effects of these mediators on neutrophil functions and the underlying cellular mechanisms involved. We show that PGE2-G, but not PGE2-EA, inhibits leukotriene B4 biosynthesis, superoxide production, migration, and antimicrobial peptide release. The effects of PGE2-G were prevented by EP1/EP2 receptor antagonist AH-6809 but not the EP4 antagonist ONO-AE2-227. The effects of PGE2-G required its hydrolysis into PGE2, were not observed with the non-hydrolyzable PGE2-serinol amide, and were completely prevented by methyl-arachidonoyl-fluorophosphate and palmostatin B, and partially prevented by JZL184 and WWL113. Although we could detect six of the documented PG-G hydrolases in neutrophils by quantitative PCR, only ABHD12 and ABHD16A were detected by immunoblot. Our pharmacological data, combined with our protein expression data, did not allow us to pinpoint one PGE2-G lipase, and rather support the involvement of an uncharacterized lipase and/or of multiple hydrolases. In conclusion, we show that PGE2-G inhibits human neutrophil functions through its hydrolysis into PGE2, and by activating the EP2 receptor. This also indicates that neutrophils could regulate inflammation by altering the balance between PG-G and PG levels in vivo.

The endocannabinoid (eCB) 2-arachidonoyl-gycerol (2-AG) modulates immune responses by activating cannabinoid receptors or through its multiple metabolites, notably eicosanoids. Thus, 2-AG hydrolysis inhibition might represent an interesting anti-inflammatory strategy that would simultaneously increase the levels of 2-AG and decrease those of eicosanoids. Accordingly, 2-AG hydrolysis inhibition increased 2-AG half-life in neutrophils. Under such setting, neutrophils, eosinophils, and monocytes synthesized large amounts of 2-AG and other monoacylglycerols (MAGs) in response to arachidonic acid (AA) and other unsaturated fatty acids (UFAs). Arachidonic acid and UFAs were ~1000-fold more potent than G protein-coupled receptor (GPCR) agonists. Triascin C and thimerosal, which, respectively, inhibit fatty acyl-CoA synthases and acyl-CoA transferases, prevented the UFA-induced MAG biosynthesis, implying glycerolipid remodeling. 2-AG and other MAG biosynthesis was preceded by that of the corresponding lysophosphatidic acid (LPA). However, we could not directly implicate LPA dephosphorylation in MAG biosynthesis. While GPCR agonists poorly induced 2-AG biosynthesis, they inhibited that induced by AA by 25%-50%, suggesting that 2-AG biosynthesis is decreased when leukocytes are surrounded by a pro-inflammatory entourage. Our data strongly indicate that human leukocytes use AA and UFAs to biosynthesize biologically significant concentrations of 2-AG and other MAGs and that hijacking the immune system with 2-AG hydrolysis inhibitors might diminish inflammation in humans.

Background: Bronchial Thermoplasty (BT) reduces exacerbations and improves asthma quality of life questionnaire (AQLQ) scores in asthma [1]. Studies have shown reductions in airway smooth muscle (ASM) mass and reticular basement membrane (RBM) thickness after BT, but are limited by small sample size. Aims: To investigate airway remodeling responses to BT, and compare these to clinical outcomes. Methods: Pre- and post-BT data was pooled for 99 patients at 7 centres. ASM mass and RBM thickness were measured on bronchial biopsies, and compared to clinical outcomes. Results: Mean (SD) age was 48.4 years (11.7) and BMI was 29.0 kg/m2 (6.4). Baseline FEV1 was 68.8% predicted (23.1) and blood eosinophils were 0.25x10-9/L (0.26). 59 of 99 patients were taking maintenance oral Prednisolone (mean dose 25.7mg). In response to BT, mean (SD) annual exacerbation rate decreased from 6.4 (5.5) to 1.1 (1.7) (p= Conclusion: ASM mass and RBM thickness reduce rapidly following BT, and effects persist beyond 1 year. The observed improvements in airway remodeling parameters are not related to improvements in clinical outcomes. Other bronchial mucosal changes or mechanisms may play a role in the clinical response to BT. Reference: 1) Castro et al, AJRCCM. 2010;181(2):116-24

Background: BT is an endoscopic treatment for uncontrolled asthma. The long-term efficacy and safety of BT beyond 5 years is not known. Objective: To evaluate the efficacy and safety of BT at 10+ years follow-up. Methods: BT10+ is an international, multi-center, ≥10yrs follow-up study on subjects who were previously enrolled in the AIR, RISA and AIR2 trials. Demographics, quality of life, lung function, severe exacerbations (SE, defined as exacerbations requiring systemic corticosteroids) and healthcare utilisation for the previous year were collected. Results: Of the 429 subjects enrolled in the previous studies, 192 subjects had a follow-up at 10.6-15.8 years (12.1 median) post-treatment at 16 centers: 137 treated with BT during a previous study, 18 control/sham treated with BT post-study, and 37 control/sham only. Rates for hospitalisations, emergency visits and SE for BT subjects from the year prior to BT to the year prior to the BT 10+ visit are shown in Figure 1. No increases in hospitalisations or ER visits were observed compared to baseline and SE were stable compared to Year 1. No deterioration in rates were shown for control/sham only and post-study BT patients. Quality of life and spirometry results were comparable between Years 1, 5, and 10+ for all groups. Conclusion: The BT10+ study suggests that efficacy of bronchial thermoplasty is sustained over 10 years. Funding: Boston Scientific

Background Omalizumab is an established anti-IgE therapy for the treatment of allergic diseases that prevents IgE from binding to its receptor. QGE031 is an investigational anti-IgE antibody that binds IgE with higher affinity than omalizumab. Objective This study compared the effects of QGE031 with those of omalizumab on clinical efficacy, IgE levels, and FceRI expression in a clinical model of allergic asthma. Methods Thirty-seven patients with mild allergic asthma were randomized to subcutaneous omalizumab, placebo, or QGE031 at 24, 72, or 240 mg every 2 weeks for 10 weeks in a double-blind, parallel-group multicenter study. Inhaled allergen challenges and skin tests were conducted before dosing and at weeks 6, 12, and 18, and blood was collected until 24 weeks after the first dose. Results QGE031 elicited a concentration- and time-dependent change in the provocative concentration of allergen causing a 15% decrease in FEV 1 (allergen PC 15 ) that was maximal and approximately 3-fold greater than that of omalizumab ( P = .10) and 16-fold greater than that of placebo ( P = .0001) at week 12 in the 240-mg cohort. Skin responses reached 85% suppression at week 12 in the 240-mg cohort and were maximal at week 18. The top doses of QGE031 consistently suppressed skin test responses among subjects but had a variable effect on allergen PC 15 (2-fold to 500-fold change). QGE031 was well tolerated. Conclusion QGE031 has greater efficacy than omalizumab on inhaled and skin allergen responses in patients with mild allergic asthma. These data support the clinical development of QGE031 as a treatment of asthma.

CONTEXT High eosinophil (EOS) counts is a key feature of asthma. EOS notably affect the asthmatic response by the mean of their lipidome. Mouse models have been developed in the hope of defining new pharmacological targets to treat asthma. However, many pinpointed targets could not be translated into clinics, underscoring that key differences exist between mice and humans. Herein, we compared the ability of human (h) and mouse (m) EOS to synthesize key bioactive lipids from arachidonic acid (AA) and docosahexaenoic acid (DHA). RESULTS In response to AA, hEOS synthesized both LTC4 and LTB4 while mEOS only synthesized LTB4. The synthesis of the 15-lipoxygenase products 15-HETE and 12-HETE also differed, hEOS producing them at a 5:1 ratio while mEOS producing them at a 1:2 ratio. mEOS consistently synthetized more PGE2 than hEOS. As for DHA, mEOS produced ten times the levels of specialized pro-resolving mediators (resolvins, maresins and protectins) than hEOS. In contrast, hEOS produced important amounts of the endocannabinoid 2-AG and its congener 2-DHG, a biosynthetic pathway that was 10 – 50 times less important in mEOS. CONCLUSIONS Our data show that the lipidomes of hEOS and mEOS differ, notably by their production of leukotrienes, their opposite 15-HETE/12-HETE ratio, their ability to synthesize specialized pro-resolving mediators (mEOS > hEOS), and the inability of mEOS to synthesize anti-inflammatory endocannabinoids. We thus propose that in mouse models of asthma, the impact of LTB4 is likely overrated, the role of specialized pro-resolving mediators amplified, and the role of endocannabinoids underrated, compared to asthmatics.

BACKGROUND The endocannabinoids 2-AG and AEA are lipids regulating many physiological processes, notably inflammation. The endocannabinoidome includes other monoacylglycerols (MAG) and N-acyl-ethanolamines (NAE) such as 1-linoleoyl-glycerol (1-LG) and N-linoleoyl-ethanolamine (LEA). Endocannabinoid hydrolysis inhibitors are now being tested as potential anti-inflammatory agents. By increasing MAG and/or NAE levels, these inhibitors will likely increase the levels of their metabolites. Herein we investigated whether 1-LG and LEA were substrates for the 15-lipoxygenase pathway, which is strongly involved in asthma and its severity. We thus assessed how human eosinophils and neutrophils biosynthesized the 15-lipoxygenase metabolites of 1-LG and LEA. Linoleic acid (LA), a well-documented substrate of 15-lipoxygenases, was used as positive control. RESULTS We synthesized the putative 15-lipoxygenase metabolites of 1-LG and LEA using Novozym435 and soybean lipoxygenase. Eosinophils, which express the 15-lipoxygenase-1, metabolized LA, 1-LG, and LEA into their 13-hydroxy derivatives. This was almost complete after 5 minutes. Substrate preference of eosinophils was LA>LEA>1-LG. Human neutrophils, which express the 15-lipoxygenase-2, also metabolized LA, 1-LG, and LEA into their 13-hydroxy derivatives. This was maximal after 30 seconds. Substrate preference was LA≫1-LG>LEA. Importantly, the new 15-lipoxygenase metabolites we disclose were found in tissues from humans and mice. CONCLUSIONS We successfully showed that human eosinophils and neutrophils transforms 1-LG and LEA into novel 15-lipoxygenase metabolites. How these new metabolites modulate the inflammatory cascade is now being explored.

The endocannabinoid 2-arachidonoyl-gycerol (2-AG) modulates immune responses by activating cannabinoid receptors or through its multiple metabolites, notably eicosanoids. Thus, 2-AG hydrolysis inhibition might represent an interesting anti-inflammatory strategy that would simultaneously increase the levels of 2-AG and decrease those of eicosanoids. Accordingly, 2-AG hydrolysis inhibition increased 2-AG half-life in neutrophils. Under such setting, neutrophils, eosinophils and monocytes synthesized large amounts of 2-AG and other monoacylglycerols (MAGs) in response to arachidonic acid and other unsaturated fatty acids (UFAs). Arachidonic acid and UFAs were ~1000-fold more potent than GPCR agonists at stimulating MAG biosynthesis. Triascin C and thimerosal, which respectively inhibit fatty acyl-CoA synthases and acyl-CoA transferases, prevented the UFA-induced MAG synthesis, implying glycerolipid remodeling is essential in this process. 2-AG and other MAG biosynthesis was preceded by that of the corresponding lysophosphatidic acid (LPA). However, we could not directly implicate LPA dephosphorylation in MAG biosynthesis. While the GPCR agonists PAF, fMLP and LTB4 poorly or did not induced 2-AG biosynthesis, they inhibited that induced by AA by 25–50%, suggesting that 2-AG biosynthesis is decreased when leukocytes are surrounded by a pro-inflammatory entourage. Our data conclusively indicate that human leukocytes use AA and UFAs to biosynthesize biologically significant concentrations of 2-AG and other MAGs and that hijacking the immune system with 2-AG hydrolysis inhibitors might diminish inflammation in humans.

Leukotriene B

Summary Background The role of interleukin 13 in airway inflammation and remodelling in asthma is unclear. Tralokinumab is a human monoclonal antibody that neutralises interleukin 13. We aimed to evaluate whether tralokinumab would have an effect on airway eosinophilic infiltration, blood and sputum eosinophil concentrations, eosinophil activation, and airway remodelling. Methods We did a multicentre, double-blind, randomised, placebo-controlled phase 2 trial at 15 centres across the UK, Denmark, and Canada. We enrolled participants of either sex aged 18–75 years with inadequately controlled moderate-to-severe asthma for 12 months or more, requiring treatment with inhaled corticosteroids at a stable dose. We randomly assigned participants (1:1) to receive tralokinumab (300 mg) or placebo by an interactive web-based system or voice response system. Participants and study personnel were masked to treatment allocation. Both tralokinumab and placebo were administered subcutaneously every 2 weeks. The primary outcome measure was change from baseline to week 12 in bronchial biopsy eosinophil count. Secondary outcome measures included change in blood and sputum eosinophil counts. Exploratory outcomes included fractional exhaled nitric oxide (FENO) and blood IgE concentrations. Safety analyses were carried out in all participants who received study drug. This trial is registered with ClinicalTrials.gov, number NCT02449473, and with the European Clinical Trials Database, EudraCT 2015-000857-19. Findings Between Sept 25, 2015, and June 21, 2017, 224 participants were enrolled and screened. Of these participants, 79 were randomly assigned to receive tralokinumab (n=39) or placebo (n=40). Tralokinumab did not significantly affect bronchial eosinophil count compared with placebo at week 12 (treatment effect ratio 1·43, 95% CI 0·63–3·27; p=0·39). Compared with placebo, tralokinumab did not significantly affect blood eosinophil count (treatment effect ratio 1·21, 95% CI 1·00–1·48; p=0·055) or sputum eosinophil count (0·57, 0·06–6·00; p=0·63), but FENO concentration (0·78, 0·63–0·96; p=0·023) and total blood IgE concentration (0·86, 0·77–0·97; p=0·014) were significantly reduced. 33 (85%) of 39 patients receiving tralokinumab and 32 (80%) of 40 receiving placebo reported at least one adverse event during the treatment period. No deaths in either treatment group were observed. Treatment-related adverse events occurred more frequently in the tralokinumab group than in the placebo group (11 [28%] of 39 vs seven [18%] of 40). Interpretation Tralokinumab did not significantly affect eosinophilic inflammation in bronchial submucosa, blood, or sputum compared with placebo, but did reduce FENO and IgE concentrations. These results suggest interleukin 13 is not crucial for eosinophilic airway inflammation control in moderate-to-severe asthma. Funding AstraZeneca.

Introduction Indeterminate pulmonary nodules (IPNs) lack clinical or radiographic features of benign etiologies and often undergo invasive procedures unnecessarily, suggesting potential roles for diagnostic adjuncts using molecular biomarkers. The primary objective was to validate a multivariate classifier that identifies likely benign lung nodules by assaying plasma protein expression levels, yielding a range of probability estimates based on high negative predictive values (NPVs) for patients with 8 to 30 mm IPNs. Methods A retrospective, multicenter, case-control study was performed using multiple reaction monitoring mass spectrometry, a classifier comprising five diagnostic and six normalization proteins, and blinded analysis of an independent validation set of plasma samples. Results The classifier achieved validation on 141 lung nodule-associated plasma samples based on predefined statistical goals to optimize sensitivity. Using a population based nonsmall-cell lung cancer prevalence estimate of 23% for 8 to 30 mm IPNs, the classifier identified likely benign lung nodules with 90% negative predictive value and 26% positive predictive value, as shown in our prior work, at 92% sensitivity and 20% specificity, with the lower bound of the classifier's performance at 70% sensitivity and 48% specificity. Classifier scores for the overall cohort were statistically independent of patient age, tobacco use, nodule size, and chronic obstructive pulmonary disease diagnosis. The classifier also demonstrated incremental diagnostic performance in combination with a four-parameter clinical model. Conclusions This proteomic classifier provides a range of probability estimates for the likelihood of a benign etiology that may serve as a noninvasive, diagnostic adjunct for clinical assessments of patients with IPNs.

Bronchial thermoplasty is a recent endoscopic technique for the treatment of severe asthma. It is an innovative treatment whose clinical efficacy and safety are beginning to be better understood. Since this is a device-based treatment, the evaluation procedure of risks and benefits is different that for pharmaceutical products; safety aspects, regulatory requirements, study design and the assessment of the magnitude of effects may all be different. The mechanism of action and optimal patient selection need to be assessed further in rigorous clinical and scientific studies. This technique is in harmony with the development of personalised medicine in the 21st century. It should be developed further in response to the numerous challenges and needs not yet met in the management of severe asthma.

Bronchial thermoplasty is an endoscopic therapy for severe asthma. The previously reported, randomised sham-controlled AIR2 (Asthma Intervention Research 2) trial showed a significant reduction in severe asthma exacerbations, emergency department visits and hospitalisations after bronchial thermoplasty. More “real-world” clinical outcome data is needed. This article compares outcomes in bronchial thermoplasty subjects with 3 years of follow-up from the ongoing, post-market PAS2 (Post-FDA Approval Clinical Trial Evaluating Bronchial Thermoplasty in Severe Persistent Asthma) study with those from the AIR2 trial. 279 subjects were treated with bronchial thermoplasty in the PAS2 study. We compared the first 190 PAS2 subjects with the 190 bronchial thermoplasty-treated subjects in the AIR2 trial at 3 years of follow-up. The PAS2 subjects were older (mean age 45.9 versus 40.7 years) and more obese (mean body mass index 32.5 versus 29.3 kg·m−2) and took higher doses of inhaled corticosteroids (mean dose 2301 versus 1961 μg·day−1). More PAS2 subjects had experienced severe exacerbations (74% versus 52%) and hospitalisations (15.3% versus 4.2%) in the 12 months prior to bronchial thermoplasty. At year 3 after bronchial thermoplasty, the percentage of PAS2 subjects with severe exacerbations, emergency department visits and hospitalisations significantly decreased by 45%, 55% and 40%, respectively, echoing the AIR2 results. The PAS2 study demonstrates similar improvements in asthma control after bronchial thermoplasty compared with the AIR2 trial despite enrolling subjects who may have had poorer asthma control., A comparison of the results obtained thus far in PAS2 with AIR2 evaluating bronchial thermoplasty in severe asthma http://ow.ly/sB0X30csDuE

We collated 129 non-overlapping risk loci for chronic obstructive pulmonary disease (COPD) from the GWAS literature. Using recent and complementary integrative genomics approaches, combining GWAS and lung eQTL results, we identified 12 novel COPD loci and corresponding causal genes. In addition, we mapped candidate causal genes for 60 out of the 129 GWAS-nominated loci as well as for four sub-genome-wide significant COPD risk loci derived from the largest GWAS on COPD. Mapping causal genes in lung tissue represents an important contribution on the genetics of COPD, enriches our biological interpretation of GWAS findings, and brings us closer to clinical translation of genetic associations.

Background: Bronchial thermoplasty (BT) is an endoscopic treatment for patients with asthma that is not well controlled with optimal doses of inhaled corticosteroids and long-acting beta-agonists. The BT procedure delivers radiofrequency energy to the airways leading notably to a persistent reduction in airway smooth muscle and sub-epithelial collagen deposition. In 3 randomized controlled trials (AIR, RISA, and AIR2 trials) of patients with moderate to severe asthma, those who underwent BT were followed for 5 years post-BT and experienced fewer severe asthma exacerbations (SE), ER visits and hospitalizations and had improved quality of life. The BT10+ study will assess the safety and efficacy of BT in patients who previously participated in these 3 trials and who have reached ≥10 year follow-up. Methods: BT10+ is an international, multi-center, follow-up for subjects from AIR, RISA and AIR2. Subjects who participated in any of these 3 trials will be eligible to enroll in BT10+ when they reach 10 years from their index 6-week follow-up visit. Control and sham subjects will also be invited to participate. Efficacy and safety data of BT will be collected during a single study visit. The primary effectiveness endpoint is durability of BT treatment effect determined by comparing SEs during years 1 and 5 to the year prior to the BT10+ study visit. The primary safety endpoint is absence of abnormal CT scan findings attributable to BT. Conclusion: The BT10+ study is designed to assess the safety and efficacy of BT over the long term (10+ years) in patients who participated in three landmark clinical trials. Funder: Boston Scientific Corporation.

Summary A fungal protease (Alp1/ Asp f13 ) from Aspergillus fumigatus was detected in the airways of subjects with asthma but not controls, which correlated strongly with disease severity, respiratory dysfunction, and steroid use.

Allergic rhinitis is a common chronic disorder characterized by immunoglobulin E-mediated inflammation. To identify new genes associated with this trait, we performed genome- and epigenome-wide association studies and linked marginally significant CpGs located in genes or its promoter and SNPs located 1 Mb from the CpGs, by identifying cis methylation quantitative trait loci (mQTL). This approach relies on functional cellular aspects rather than stringent statistical correction. We were able to identify one gene with significant cis-mQTL for allergic rhinitis, caudal-type homeobox 1 (CDX1). We also identified 11 genes with marginally significant cis-mQTLs (p

Acute effects of bronchial thermoplasty on small airways and lung parenchyma: good or bad? http://ow.ly/T6Uu309keuf

Bronchial thermoplasty is an endoscopic therapy for severe asthma. The previously reported, randomised sham-controlled AIR2 (Asthma Intervention Research 2) trial showed a significant reduction in severe asthma exacerbations, emergency department visits and hospitalisations after bronchial thermoplasty. More “real-world” clinical outcome data is needed. This article compares outcomes in bronchial thermoplasty subjects with 3 years of follow-up from the ongoing, post-market PAS2 (Post-FDA Approval Clinical Trial Evaluating Bronchial Thermoplasty in Severe Persistent Asthma) study with those from the AIR2 trial. 279 subjects were treated with bronchial thermoplasty in the PAS2 study. We compared the first 190 PAS2 subjects with the 190 bronchial thermoplasty-treated subjects in the AIR2 trial at 3 years of follow-up. The PAS2 subjects were older (mean age 45.9 versus 40.7 years) and more obese (mean body mass index 32.5 versus 29.3 kg·m −2 ) and took higher doses of inhaled corticosteroids (mean dose 2301 versus 1961 μg·day −1 ). More PAS2 subjects had experienced severe exacerbations (74% versus 52%) and hospitalisations (15.3% versus 4.2%) in the 12 months prior to bronchial thermoplasty. At year 3 after bronchial thermoplasty, the percentage of PAS2 subjects with severe exacerbations, emergency department visits and hospitalisations significantly decreased by 45%, 55% and 40%, respectively, echoing the AIR2 results. The PAS2 study demonstrates similar improvements in asthma control after bronchial thermoplasty compared with the AIR2 trial despite enrolling subjects who may have had poorer asthma control.

Recent studies identified three genetic loci reproducibly associated with lung cancer in populations of European ancestry, namely 15q25, 5p15 and 6p21. The goals of this study are first to confirm whether these loci are associated with lung cancer in a French Canadian population and second to identify disease-associated single nucleotide polymorphisms (SNPs) influencing messenger RNA (mRNA) expression levels of genes in the lung, that is expression quantitative trait loci (eQTLs). SNPs were genotyped in 420 patients undergoing lung cancer surgery and compared with 3151 controls of European ancestry. Genome-wide gene expression levels in non-tumor lung tissues of the same 420 patients were also measured to identify eQTLs. Significant eQTLs were then followed-up in two replication sets (n = 339 and 363). SNPs found in the three susceptibility loci were associated with lung cancer in the French Canadian population. Strong eQTLs were found on chromosome 15q25 with the expression levels of CHRNA5 (P = 2.23 × 10(-) (22) with rs12907966). The CHRNA5-rs12907966 eQTL was convincingly validated in the two replication sets (P = 3.46 × 10(-) (16) and 2.01 × 10(-) (15)). On 6p21, a trend was observed for rs3131379 to be associated with the expression of APOM (P = 3.58 × 10(-) (4)) and validated in the replication sets (P = 1.11 × 10(-) (8) and 6.84 × 10(-) (4)). On 5p15, no significant eQTLs were found. This study confirmed that chromosomes 15q25, 5p15 and 6p21 harbored susceptibility loci for lung cancer in French Canadians. Most importantly, this study suggests that the risk alleles at 15q25 and 6p21 may mediate their effect by regulating the mRNA expression levels of CHRNA5 and APOM in the lung.

Background: Patients with severe refractory asthma treated with bronchial thermoplasty (BT), a bronchoscopic procedure that improves asthma control by reducing excess airway smooth muscle, were followed up for 5 years to evaluate long-term safety of this procedure. Objectives: To assess long-term safety of BT for 5 years. Methods: Patients with asthma aged 18 to 65 years requiring high-dose inhaled corticosteroids (ICSs) (>750 μg/d of fluticasone propionate or equivalent) and long-acting β2-agonists (LABAs) (at least 100 μg/d of salmeterol or equivalent), with or without oral prednisone (≤30 mg/d), leukotriene modifiers, theophylline, or other asthma controller medications were enrolled in the Research in Severe Asthma (RISA) Trial. Patients had a prebronchodilator forced expiratory volume in 1 second of 50% or more of predicted, demonstrated methacholine airway hyperresponsiveness, had uncontrolled symptoms despite taking maintenance medication, abstained from smoking for 1 year or greater, and had a smoking history of less than 10 pack-years. Results: Fourteen patients (of the 15 who received active treatment in the RISA Trial) participated in the long-term follow-up study for 5 years. The rate of respiratory adverse events (AEs per patient per year) was 1.4, 2.4, 1.7, and 2.4, respectively, in years 2 to 5 after BT. There was a decrease in hospitalizations and emergency department visits for respiratory symptoms in each of years 1, 2, 3, 4, and 5 compared with the year before BT treatment. Measures of lung function showed no deterioration for 5 years. Conclusion: Our findings suggest that BT is safe for 5 years after BT in patients with severe refractory asthma. Trial Registration: clinicaltrials.gov Identifier: NCT00401986. © 2013 American College of Allergy, Asthma and Immunology. Published by Elsevier Inc. All rights reserved.

To the Editor: A number of models have been proposed to predict spirometric lung function using age, sex, height and self-reported ethnicity [1, 2]. These models are particularly important to derive per cent predicted lung function and establish the severity of lung diseases such as chronic obstructive pulmonary disease (COPD) [3]. Compared to self-reported race and/or ethnicity, genetic data can provide more accurate and objective measurements of ancestry and has the potential to alleviate some of the problems related to the lack of consensus on the definition of race and ethnicity worldwide [4]. A recent report suggested that genetically determined ancestry improves predicted lung-function measurements in African Americans [5]. Here, we test whether genetic ancestry derived from genome-wide genotyping data is useful to predict lung function in a diverse European population. The population used in this study is part of a lung expression quantitative trait loci (eQTL) mapping study [6, 7]. Briefly, research participants were recruited from three academic sites: Laval University (Quebec, QC, Canada), University of British Columbia (Vancouver, BC, Canada), and Groningen University (Groningen, the Netherlands), henceforth referred to as Laval, UBC, and Groningen, respectively. All subjects were genotyped for ∼1.2 million single nucleotide polymorphisms (SNPs) using the Illumina Human1M-Duo BeadChip (Illumina, Inc., San Diego, CA, USA). Subjects from Laval and UBC provided written informed consent and the study was approved by the ethics committees of the respective study sites. At Groningen, the study protocol was consistent with the research code of the University Medical Center Groningen and Dutch national ethical and professional guidelines (“Code of conduct; Dutch federation of biomedical scientific societies”; http://www.federa.org). Prior to the analysis, standard genotyping quality controls and exclusion of patients with disorders that affected lung function (other than COPD) were …

Background and Objectives: Asthma is a heterogeneous disease where patient severity can be classified according to various models based on numerous variables. Large collections of well-phenotyped subjects are needed to find distinct clusters of patients for personalized medicine and future genetic studies. The objective of this study is to describe the collection of the Quebec City Case-Control Asthma Cohort and to identify homogeneous subgroups of asthma patients based on clinical characteristics. Methods: This cohort is part of an ongoing project initiated in 2007 to elucidate the genetic basis of asthma. All subjects are randomly recruited at the same site following advertisements. Subjects are unrelated French Canadian white adults 18 years of age or older. Each participant underwent a spirometry, methacholine challenge, and allergy skin-prick tests. Blood was collected for DNA, cell counts and total serum IgE measurements. So far, 982 subjects have been recruited and classified as cases (n = 566) or controls (n = 416). We performed factor and cluster analyses on collected phenotypes from this set to identify subgroups of phenotypically similar asthmatic patients. Results: Factor analysis with 13 variables led to the selection of five factors: lung function, numbers of allergens, blood eosinophil percentage, smoking status and age. K-means cluster analysis on the reduced dataset produced four significantly different groups with the following characteristics: smoking history, low atopy and low lung function, high atopy, and young non-smoking with average atopy. Conclusions: The Quebec City Case-Control Asthma Cohort is a new resource for local and collaborative clinical and genetic research on asthma. This new collection reveals distinct multivariate phenotypes of adult asthma that are likely to be important for future genetic studies and targeted therapies.

Summary Background Benralizumab is a humanised, anti-interleukin 5 receptor α monoclonal antibody that directly and rapidly depletes eosinophils, reduces asthma exacerbations, and improves lung function for patients with severe eosinophilic asthma. The objective of this trial was to assess the safety and efficacy of benralizumab for patients with mild to moderate, persistent asthma. Methods In this randomised, double-blind, placebo-controlled, phase 3 trial, we recruited patients aged 18–75 years, weighing at least 40 kg, and with a postbronchodilator reversibility in forced expiratory volume in 1 s (FEV 1 ) of at least 12% at screening, from 52 clinical research centres in six countries. Patients must have been receiving either low- to medium-dosage inhaled corticosteroids (ICS) or low-dosage ICS plus long-acting β 2 agonist fixed-combination therapy at screening, had a morning prebronchodilator FEV 1 of more than 50% to 90% predicted at screening, and had one or more of the following symptoms within the 7 days before randomisation: a daytime or night-time asthma symptom score of at least 1 for at least 2 days, rescue short-acting β 2 agonist use for at least 2 days, or night-time awakenings due to asthma for at least one night. We converted patients' ICS treatments to 180 μg or 200 μg budesonide dry powder inhaler twice daily for the entire duration of the study using the approved dosages in the patients' respective countries and randomly allocated them (1:1; stratified by blood eosinophil count [ vs ≥300 cells per μL] and region [USA vs the rest of the world]) with an interactive web-based voice response system to receive subcutaneous placebo or benralizumab 30 mg injections every 4 weeks for 12 weeks. All patients and investigators involved in patient treatment or clinical assessment and those assessing outcomes were masked to treatment allocation. The primary endpoint was change from baseline prebronchodilator FEV 1 at week 12. Efficacy analyses used an intention to treat approach. This trial is registered with ClinicalTrials.gov, number NCT02322775. Findings Between Feb 2, 2015, and April 24, 2015, we enrolled 351 patients, with 211 (60%) randomly assigned (105 [50%] to placebo and 106 [50%] to benralizumab). Benralizumab resulted in an 80 mL (95% CI 0–150; p=0·04) greater improvement (least-squares mean difference) in prebronchodilator FEV 1 after 12 weeks than did placebo (placebo group: 2246 mL [SD 768] at baseline vs 2261 mL [796] at week 12, change from baseline of 0 mL; benralizumab group: 2248 mL [606] vs 2310 mL [670], 70 mL). 44 (42%) patients in the benralizumab group had adverse events compared with 49 (47%) in the placebo group. The most common adverse events for both groups were nasopharyngitis (eight [8%] patients in each group) and upper respiratory tract infections (five [5%] patients in each group). Serious adverse events occurred in two (2%) patients each in the benralizumab (pancytopenia and a suicide attempt, both considered unrelated to treatment) and placebo (cervix carcinoma and colon adenoma) groups. Interpretation This study suggests that active and modifiable disease processes might be ongoing in patients with mild to moderate, persistent asthma receiving ICS. Although the lung function improvement observed does not warrant use of benralizumab in this population because it did not reach the minimum clinically important difference of 10%, further studies to assess this finding should be considered. Funding AstraZeneca.

The endocannabinoids 2-arachidonoyl-glycerol and

LTB4 is an inflammatory lipid mediator mainly biosynthesized by leukocytes. Since its implication in inflammatory diseases is well recognized, many tools to regulate its biosynthesis have been developed and showed promising results in vitro and in vivo, but mixed results in clinical trials. Recently, the mTOR pathway component p70S6 kinase 1 (p70S6K1) has been linked to LTC4 synthase and the biosynthesis of cysteinyl-leukotrienes. In this respect, we investigated if p70S6K1 could also play a role in LTB4 biosynthesis. We thus evaluated the impact of the p70S6K1 inhibitors PF-4708671 and LY2584702 on LTB4 biosynthesis in human neutrophils. At a concentration of 10 μM, both compounds inhibited S6 phosphorylation, although neither one inhibited the thapsigargin-induced LTB4 biosynthesis, as assessed by the sum of LTB4, 20-OH-LTB4, and 20-COOH-LTB4. However, PF-4708671, but not LY2584702, inhibited the ω-oxidation of LTB4 into 20-OH-LTB4 by intact neutrophils and by recombinant CYP4F3A, leading to increased LTB4 levels. This was true for both endogenously biosynthesized and exogenously added LTB4. In contrast to that of 17-octadecynoic acid, the inhibitory effect of PF-4708671 was easily removed by washing the neutrophils, indicating that PF-4708671 was a reversible CYP4F3A inhibitor. At optimal concentration, PF-4708671 increased the half-life of LTB4 in our neutrophil suspensions by 7.5 fold, compared to 5 fold for 17-octadecynoic acid. Finally, Michaelis-Menten and Lineweaver-Burk plots indicate that PF-4708671 is a mixed inhibitor of CYP4F3A. In conclusion, we show that PF-4708671 inhibits CYP4F3A and prevents the ω-oxidation of LTB4 in cellulo, which might result in increased LTB4 levels in vivo.

Background: Bronchial thermoplasty (BT) improves asthma symptoms by reducing the airway smooth muscle (ASM) mass. A durable efficacy and safety of BT on asthma control has been documented out to 5 years in patients with severe persistent asthma. The ASM reduction has been demonstrated in bronchial biopsies taken 3 weeks or 3 months post-BT. BT also decreased type 1 collagen deposition and sub-epithelial basement membrane thickening. The persistence of these structural changes over time with durable clinical benefits observed in clinical trials remains uncertain. Objectives: To assess long-term efficacy of BT on ASM mass and sub-epithelial collagen deposition. Methods: Nine subjects (age: 30-70 years; FEV1: 60-105% of predicted; ICS: 500-1500µg fluticasone equivalent) with persistent asthma underwent BT procedures. Bronchial biopsies were taken before (baseline), and at 3-14 weeks and 27-48 months after BT. Histology and immunohistochemistry studies were performed. Clinical evaluation was performed at 12 and ≥27 months post-BT. Results: ASM area decreased from 11.8% ±1.24% at baseline to 4.7± 0.95% at week 3 post BT and this decrease persisted at ≥27 months post-BT: 4.6±1.05%, P=0.0002 compared to baseline. The type I collagen deposition decreased from 6.7±0.4 µm at baseline to 4.5±0.5 µm 3 weeks post-BT and remained unchanged at long-term followup: 4.6±0.5 µm (P=0.003). At 12 and ≥27 months, BT improved significantly the Asthma Control Scoring Score compared to baseline and decreased the number of severe exacerbations. Conclusion: These data showed long-term (≥27 months) benefits from BT in terms of ASM mass reduction, sub-epithelial collagen deposition and asthma control.

Background: Elite swimmers9 airways show remodeling and inflammation similar to mild asthma. Aim: We hypothesized that after swimming retirement, airway inflammation and remodeling would decrease. We aimed to compare swimmers9 airways during their training career and two years after retirement. Methods: Six swimmers had bronchial biopsy samplings, methacholine challenge (MIT), and an eucapnic voluntary hyperpnea test (EVH) during and two years after the end of their swimming career (V1 and V2). Airway hyperresponsiveness (AHR) was defined as a PC 20 1 post-EVH. Biomarkers were analyzed by immunohistochemistry: Tenascin was expressed as % staining of the sub-epithelial basement membrane area, Collagen III as % staining in the whole tissue biopsy, and CD3 and Tryptase as number of cells/mm 2 . Results: Characteristics of swimmers were: Age (years): 20 ± 1 at V1 vs 25 ± 1 at V2 (p 1 (%): 119 ± 12 at V1 vs 116 ± 16 at V2 (p=0.58); Forced Vital Capacity (L): 127 ± 10 at V1 vs 126 ± 11 at V2 (p=0.70). Two swimmers had AHR on both visits, two had no AHR, one had no more AHR and one developed AHR at V2. Collagen type III was 3.1 ± 2.7% at V1 vs 9.2 ± 4.2% at V2 (p=0.04)) and CD3 (768 ± 323 cells/mm 2 at V1 vs 368 ± 170 cells/mm 2 at V2 (p=0.02).Tenascin and Tryptase did not change significantly. Conclusion: A decrease in lymphocyte count was observed after retirement, associated to an increase in the proportion of collagen III, which may reflect the persistence of remodeling in a less inflamed airway.

Introduction: Benralizumab is a humanized monoclonal antibody against the interleukin-5 receptor alpha subunit that results in direct, rapid, and nearly complete depletion of eosinophils via antibody-dependent cell-mediated cytotoxicity. It is being evaluated for treatment of severe, uncontrolled asthma with eosinophilic inflammation. Aims and Objectives: The purpose of the BISE study was to investigate the efficacy and safety of benralizumab in patients (pts) with mild to moderate, persistent asthma (NCT02322775). Methods: In this double-blind, placebo (PBO)-controlled, multicenter study, 211 pts aged 18-75 years with pre-bronchodilator (pre-BD) FEV1 >50%–≤90%, post-BD FEV1 reversibility ≥12%, and evidence of asthma symptoms within 7 days before randomization received benralizumab 30 mg or PBO subcutaneously every 4 weeks for 8 weeks. The primary endpoint was change from baseline in pre-BD FEV1 at Week 12. Results: At Week 12, pts receiving benralizumab demonstrated greater improvements in pre-BD FEV1 vs. pts receiving PBO (table). Fewer benralizumab-treated pts reported adverse events on treatment vs. PBO-treated pts (37/106 vs. 47/105). Conclusions: Benralizumab treatment significantly improved pre-BD FEV1 in pts with mild to moderate, persistent asthma, with tolerability comparable to PBO during the 12-week study.