PEBP1 Wardens Ferroptosis by Enabling Lipoxygenase Generation of Lipid Death Signals.

Ferroptosis is a form of programmed cell death that is pathogenic to several acute and chronic diseases and executed via oxygenation of polyunsaturated phosphatidylethanolamines (PE) by 15-lipoxygenases (15-LO) that normally use free polyunsaturated fatty acids as substrates. Mechanisms of the altered 15-LO substrate specificity are enigmatic. We sought a common ferroptosis regulator for 15LO. We discovered that PEBP1, a scaffold protein inhibitor of protein kinase cascades, complexes with two 15LO isoforms, 15LO1 and 15LO2, and changes their substrate competence to generate hydroperoxy-PE. Inadequate reduction of hydroperoxy-PE due to insufficiency or dysfunction of a selenoperoxidase, GPX4, leads to ferroptosis. We demonstrated the importance of PEBP1-dependent regulatory mechanisms of ferroptotic death in airway epithelial cells in asthma, kidney epithelial cells in renal failure, and cortical and hippocampal neurons in brain trauma. As master regulators of ferroptotic cell death with profound implications for human disease, PEBP1/15LO complexes represent a new target for drug discovery.

Climate change impacts on respiratory health: exposure, vulnerability, and risk.

Anthropogenic climate change adversely impacts human health. In this perspective, we examine the impact of climate change on respiratory health risk. We describe five respiratory health threats-heat, wildfires, pollen, extreme weather events, and viruses-and discuss their impact on health outcomes in a warming climate. The risk of experiencing an adverse health outcome occurs at the intersection of exposure and vulnerability, consisting of sensitivity and adaptive capacity. Exposed individuals and communities most at risk are those with high sensitivity and low adaptive capacity, as influenced by the social determinants of health. We call for the implementation of a transdisciplinary strategy for accelerating respiratory health research, practice, and policy in the context of climate change.

Are We Meeting the Promise of Endotypes and Precision Medicine in Asthma?

While the term asthma has long been known to describe heterogeneous groupings of patients, only recently have data evolved which enable a molecular understanding of the clinical differences. The evolution of transcriptomics (and other 'omics platforms) and improved statistical analyses in combination with large clinical cohorts opened the door for molecular characterization of pathobiologic processes associated with a range of asthma patients. When linked with data from animal models and clinical trials of targeted biologic therapies, emerging distinctions arose between patients with and without elevations in type 2 immune and inflammatory pathways, leading to the confirmation of a broad categorization of type 2-Hi asthma. Differences in the ratios, sources, and location of type 2 cytokines and their relation to additional immune pathway activation appear to distinguish several different (sub)molecular phenotypes, and perhaps endotypes of type 2-Hi asthma, which respond differently to broad and targeted anti-inflammatory therapies. Asthma in the absence of type 2 inflammation is much less well defined, without clear biomarkers, but is generally linked with poor responses to corticosteroids. Integration of "big data" from large cohorts, over time, using machine learning approaches, combined with validation and iterative learning in animal (and human) model systems is needed to identify the biomarkers and tightly defined molecular phenotypes/endotypes required to fulfill the promise of precision medicine.

Discovering selective antiferroptotic inhibitors of the 15LOX/PEBP1 complex noninterfering with biosynthesis of lipid mediators.

Programmed ferroptotic death eliminates cells in all major organs and tissues with imbalanced redox metabolism due to overwhelming iron-catalyzed lipid peroxidation under insufficient control by thiols (Glutathione (GSH)). Ferroptosis has been associated with the pathogenesis of major chronic degenerative diseases and acute injuries of the brain, cardiovascular system, liver, kidneys, and other organs, and its manipulation offers a promising new strategy for anticancer therapy. This explains the high interest in designing new small-molecule-specific inhibitors against ferroptosis. Given the role of 15-lipoxygenase (15LOX) association with phosphatidylethanolamine (PE)-binding protein 1 (PEBP1) in initiating ferroptosis-specific peroxidation of polyunsaturated PE, we propose a strategy of discovering antiferroptotic agents as inhibitors of the 15LOX/PEBP1 catalytic complex rather than 15LOX alone. Here we designed, synthesized, and tested a customized library of 26 compounds using biochemical, molecular, and cell biology models along with redox lipidomic and computational analyses. We selected two lead compounds, FerroLOXIN-1 and 2, which effectively suppressed ferroptosis in vitro and in vivo without affecting the biosynthesis of pro-/anti-inflammatory lipid mediators in vivo. The effectiveness of these lead compounds is not due to radical scavenging or iron-chelation but results from their specific mechanisms of interaction with the 15LOX-2/PEBP1 complex, which either alters the binding pose of the substrate [eicosatetraenoyl-PE (ETE-PE)] in a nonproductive way or blocks the predominant oxygen channel thus preventing the catalysis of ETE-PE peroxidation. Our successful strategy may be adapted to the design of additional chemical libraries to reveal new ferroptosis-targeting therapeutic modalities.

Increased Muc5AC and Decreased Ciliated Cells in Severe Asthma Partially Restored by Inhibition of IL-4Rα Receptor.

BACKGROUND: The role of IL-13 on the airway epithelium in severe asthma leading to airway remodeling remains poorly understood. OBJECTIVE: To study IL-13 induced airway remodeling on goblet cells and cilia in the airway epithelium in severe asthma and the impact of an anti-IL4Rα antibody, dupilumab, in vitro. METHODS: Quantitative CT (qCT) lungs and endobronchial biopsies and brushings were obtained in 51 participants (22 severe, 11 non-severe asthma and 18 healthy participants) in the Severe Asthma Research Program (SARPIII) and measured for mucin and cilia related proteins. Epithelial cells were differentiated in air-liquid interphase (ALI) with IL-13 +/-dupilumab and assessed for mucin, cilia, cilia beat frequency (CBF) and epithelial integrity (transepithelial electrical resistance, TEER). RESULTS: Increased Muc5AC (Δ+263.2±92.7 lums/EpiArea) and decreased ciliated cells (Δ-0.07±0.03 Foxj1+cells/EpiArea) were observed in biopsies from severe asthma when compared to healthy (p<0.01 and p=0.047 respectively). RNAseq of epithelial cell brushes confirmed a Muc5AC increase with a decrease in a 5-gene cilia-related mean in severe asthma compared to healthy (all p<0.05). IL-13 (5 ng/mL) differentiated ALI cultures of healthy and asthmatic (severe and non-severe participants) increased Muc5AC, decreased cilia (α-acytl-tubulin) in healthy (Δ+6.5±1.5%, Δ-14.1±2.7%; all p<0.001 respectively) and asthma (Δ+4.4±2.5%, Δ-13.1±2.7%; p=0.084, p<0.001 respectively); decreased epithelial integrity (TEER) in healthy (-140.9±21.3 [ohms], p<0.001) while decreasing CBF in asthma (Δ-4.4±1.7 [Hz], p<0.01). When dupilumab was added to ALI with IL-13, there was no significant decrease in Mu5AC but there was restoration of cilia in healthy and asthma participants (absolute increase of 67.5% and 32.5% cilia, all p<0.05 respectively) while CBF increased (Δ+3.6±1.1 [Hz], p<0.001) and TEER decreased (only in asthma Δ-37.8±16.2 [ohms] p<0.05). CONCLUSIONS: IL-13 drives features of airway remodeling in severe asthma which are partially reversed by inhibiting IL-4Rα receptor in vitro.

A Novel Air Trapping Segment Score Identifies Opposing Effects of Obesity and Eosinophilia on Air Trapping in Asthma.

Rationale: Density thresholds in computed tomography (CT) lung scans quantify air trapping (AT) at the whole-lung level but are not informative for AT in specific bronchopulmonary segments. Objectives: To apply a segment-based measure of AT in asthma to investigate the clinical determinants of AT in asthma. Methods: In each of 19 bronchopulmonary segments in CT lung scans from 199 patients with asthma, AT was categorized as present if lung attenuation was less than -856 Hounsfield units at expiration in ⩾15% of the lung area. The resulting AT segment score (0-19) was related to patient outcomes. Measurements and Main Results: AT varied at the lung segment level and tended to persist at the patient and lung segment levels over 3 years. Patients with widespread AT (⩾10 segments) had more severe asthma (P < 0.05). The mean (±SD) AT segment score in patients with a body mass index ⩾30 kg/m2 was lower than in patients with a body mass index <30 kg/m2 (3.5 ± 4.6 vs. 5.5 ± 6.3; P = 0.008), and the frequency of AT in lower lobe segments in obese patients was less than in upper and middle lobe segments (35% vs. 46%; P = 0.001). The AT segment score in patients with sputum eosinophils ⩾2% was higher than in patients without sputum eosinophilia (7.0 ± 6.1 vs. 3.3 ± 4.9; P < 0.0001). Lung segments with AT more frequently had airway mucus plugging than lung segments without AT (48% vs. 18%; P ⩽ 0.0001). Conclusions: In patients with asthma, air trapping is more severe in those with airway eosinophilia and mucus plugging, whereas those who are obese have less severe trapping because their lower lobe segments are spared.

Mitochondrial DNA copy number variation in asthma risk, severity, and exacerbations.

BACKGROUND: Asthma pathophysiology is associated with mitochondrial dysfunction. Mitochondrial DNA copy number (mtDNA-CN) has been used as a proxy of mitochondrial function, with lower levels indicating mitochondrial dysfunction in population studies of cardiovascular diseases and cancers. OBJECTIVES: We investigated whether lower levels of mtDNA-CN are associated with asthma diagnosis, severity, and exacerbations. METHODS: mtDNA-CN is evaluated in blood from 2 cohorts: UK Biobank (UKB) (asthma, n = 39,147; no asthma, n = 302,302) and Severe Asthma Research Program (SARP) (asthma, n = 1283; nonsevere asthma, n = 703). RESULTS: Individuals with asthma have lower mtDNA-CN compared to individuals without asthma in UKB (beta, -0.006 [95% confidence interval, -0.008 to -0.003], P = 6.23 × 10-6). Lower mtDNA-CN is associated with asthma prevalence, but not severity in UKB or SARP. mtDNA-CN declines with age but is lower in individuals with asthma than in individuals without asthma at all ages. In a 1-year longitudinal study in SARP, mtDNA-CN was associated with risk of exacerbation; those with highest mtDNA-CN had the lowest risk of exacerbation (odds ratio 0.333 [95% confidence interval, 0.173 to 0.542], P = .001). Biomarkers of inflammation and oxidative stress are higher in individuals with asthma than without asthma, but the lower mtDNA-CN in asthma is independent of general inflammation or oxidative stress. Mendelian randomization studies suggest a potential causal relationship between asthma-associated genetic variants and mtDNA-CN. CONCLUSION: mtDNA-CN is lower in asthma than in no asthma and is associated with exacerbations. Low mtDNA-CN in asthma is not mediated through inflammation but is associated with a genetic predisposition to asthma.

Utility of eosinophil peroxidase as a biomarker of eosinophilic inflammation in asthma.

BACKGROUND: The relative utility of eosinophil peroxidase (EPX) and blood and sputum eosinophil counts as disease biomarkers in asthma is uncertain. OBJECTIVE: We sought to determine the utility of EPX as a biomarker of systemic and airway eosinophilic inflammation in asthma. METHODS: EPX protein was measured by immunoassay in serum and sputum in 110 healthy controls to establish a normal reference range and in repeated samples of serum and sputum collected during 3 years of observation in 480 participants in the Severe Asthma Research Program 3. RESULTS: Over 3 years, EPX levels in patients with asthma were higher than normal in 27% to 31% of serum samples and 36% to 53% of sputum samples. Eosinophils and EPX correlated better in blood than in sputum (rs values of 0.74 and 0.43, respectively), and high sputum EPX levels occurred in 27% of participants with blood eosinophil counts less than 150 cells/µL and 42% of participants with blood eosinophil counts between 150 and 299 cells/µL. Patients with persistently high sputum EPX values for 3 years were characterized by severe airflow obstruction, frequent exacerbations, and high mucus plug scores. In 59 patients with asthma who started mepolizumab during observation, serum EPX levels normalized in 96% but sputum EPX normalized in only 49%. Lung function remained abnormal even when sputum EPX normalized. CONCLUSIONS: Serum EPX is a valid protein biomarker of systemic eosinophilic inflammation in asthma, and sputum EPX levels are a more sensitive biomarker of airway eosinophilic inflammation than sputum eosinophil counts. Eosinophil measures in blood frequently miss airway eosinophilic inflammation, and mepolizumab frequently fails to normalize airway eosinophilic inflammation even though it invariably normalizes systemic eosinophilic inflammation.

Asthma Pathogenesis: Phenotypes, Therapies, and Gaps: Summary of the Aspen Lung Conference 2023.

Although substantial progress has been made in our understanding of asthma pathogenesis and phenotypes over the nearly 60-year history of the Aspen Lung Conferences on asthma, many ongoing challenges exist in our understanding of the clinical and molecular heterogeneity of the disease and an individual patient's response to therapy. This report summarizes the proceedings of the 2023 Aspen Lung Conference, which was organized to review the clinical and molecular heterogeneity of asthma and to better understand the impact of genetic, environmental, cellular, and molecular influences on disease susceptibility, heterogeneity, and severity. The goals of the conference were to review new information about asthma phenotypes, cellular processes, and cellular signatures underlying disease heterogeneity and treatment response. The report concludes with ongoing gaps in our understanding of asthma pathobiology and provides some recommendations for future research to better understand the clinical and basic mechanisms underlying disease heterogeneity in asthma and to advance the development of new treatments for this growing public health problem.

A novel DNase assay reveals low DNase activity in severe asthma.

Secreted deoxyribonucleases (DNases), such as DNase-I and DNase-IL3, degrade extracellular DNA, and endogenous DNases have roles in resolving airway inflammation and guarding against autoimmune responses to nucleotides. Subsets of patients with asthma have high airway DNA levels, but information about DNase activity in health and in asthma is lacking. To characterize DNase activity in health and in asthma, we developed a novel kinetic assay using a Taqman probe sequence that is quickly cleaved by DNase-I to produce a large product signal. We used this kinetic assay to measure DNase activity in sputum from participants in the Severe Asthma Research Program (SARP)-3 (n = 439) and from healthy controls (n = 89). We found that DNase activity was lower than normal in asthma [78.7 relative fluorescence units (RFU)/min vs. 120.4 RFU/min, P < 0.0001]. Compared to patients with asthma with sputum DNase activity in the upper tertile activity levels, those in the lower tertile of sputum DNase activity were characterized clinically by more severe disease and pathologically by airway eosinophilia and airway mucus plugging. Carbamylation of DNase-I, a post-translational modification that can be mediated by eosinophil peroxidase, inactivated DNase-I. In summary, a Taqman probe-based DNase activity assay uncovers low DNase activity in the asthma airway that is associated with more severe disease and airway mucus plugging and may be caused, at least in part, by eosinophil-mediated carbamylation.NEW & NOTEWORTHY We developed a new DNase assay and used it to show that DNase activity is impaired in asthma airways.

Protein-Protein interactive networks identified in bronchoalveolar lavage of severe compared to nonsevere asthma.

INTRODUCTION: Previous bronchoalveolar lavage fluid (BALF) proteomic analysis has evaluated limited numbers of subjects for only a few proteins of interest, which may differ between asthma and normal controls. Our objective was to examine a more comprehensive inflammatory biomarker panel in quantitative proteomic analysis for a large asthma cohort to identify molecular phenotypes distinguishing severe from nonsevere asthma. METHODS: Bronchoalveolar lavage fluid from 48 severe and 77 nonsevere adult asthma subjects were assessed for 75 inflammatory proteins, normalized to BALF total protein concentration. Validation of BALF differences was sought through equivalent protein analysis of autologous sputum. Subjects' data, stratified by asthma severity, were analysed by standard statistical tests, principal component analysis and 5 machine learning algorithms. RESULTS: The severe group had lower lung function and greater health care utilization. Significantly increased BALF proteins for severe asthma compared to nonsevere asthma were fibroblast growth factor 2 (FGF2), TGFα, IL1Ra, IL2, IL4, CCL8, CCL13 and CXCL7 and significantly decreased were platelet-derived growth factor a-a dimer (PDGFaa), vascular endothelial growth factor (VEGF), interleukin 5 (IL5), CCL17, CCL22, CXCL9 and CXCL10. Four protein differences were replicated in sputum. FGF2, PDGFaa and CXCL7 were independently identified by 5 machine learning algorithms as the most important variables for discriminating severe and nonsevere asthma. Increased and decreased proteins identified for the severe cluster showed significant protein-protein interactions for chemokine and cytokine signalling, growth factor activity, and eosinophil and neutrophil chemotaxis differing between subjects with severe and nonsevere asthma. CONCLUSION: These inflammatory protein results confirm altered airway remodelling and cytokine/chemokine activity recruiting leukocytes into the airways of severe compared to nonsevere asthma as important processes even in stable status.

Low CC16 mRNA Expression Levels in Bronchial Epithelial Cells Are Associated with Asthma Severity.

Rationale: CC16 is a protein mainly produced by nonciliated bronchial epithelial cells (BECs) that participates in host defense. Reduced CC16 protein concentrations in BAL and serum are associated with asthma susceptibility. Objectives: Few studies have investigated the relationship between CC16 and asthma progression, and none has focused on BECs. In this study, we sought to determine if CC16 mRNA expression levels in BECs are associated with asthma severity. Methods: Association analyses between CC16 mRNA expression levels in BECs (242 asthmatics and 69 control subjects) and asthma-related phenotypes in Severe Asthma Research Program were performed using a generalized linear model. Measurements and Main Results: Low CC16 mRNA expression levels in BECs were significantly associated with asthma susceptibility and asthma severity, high systemic corticosteroids use, high retrospective and prospective asthma exacerbations, and low pulmonary function. Low CC16 mRNA expression levels were significantly associated with high T2 inflammation biomarkers (fractional exhaled nitric oxide and sputum eosinophils). CC16 mRNA expression levels were negatively correlated with expression levels of Th2 genes (IL1RL1, POSTN, SERPINB2, CLCA1, NOS2, and MUC5AC) and positively correlated with expression levels of Th1 and inflammation genes (IL12A and MUC5B). A combination of two nontraditional T2 biomarkers (CC16 and IL-6) revealed four asthma endotypes with different characteristics of T2 inflammation, obesity, and asthma severity. Conclusions: Our findings indicate that low CC16 mRNA expression levels in BECs are associated with asthma susceptibility, severity, and exacerbations, partially through immunomodulation of T2 inflammation. CC16 is a potential nontraditional T2 biomarker for asthma development and progression.

Living in environmental justice areas worsens asthma severity and control: Differential interactions with disease duration, age at onset, and pollution.

BACKGROUND: Impoverished and historically marginalized communities often reside in areas with increased air pollution. OBJECTIVE: We evaluated the association between environmental justice (EJ) track and asthma severity and control as modified by traffic-related air pollution (TRAP). METHODS: We performed a retrospective study of 1526 adult asthma patients in Allegheny County, Pa, enrolled in an asthma registry during 2007-20. Asthma severity and control were determined using global guidelines. EJ tract designation was based on residency in census tracts with ≥30% non-White and/or ≥20% impoverished populations. TRAP exposures (NO2 and black carbon) for each census tract were normalized into pollution quartiles. Generalized linear model analyses determined the effect of EJ tract and TRAP on asthma. RESULTS: TRAP exposure in the highest quartile range was more frequent among patients living in an EJ tract (66.4% vs 20.8%, P < .05). Living in an EJ tract increased the odds of severe asthma in later onset asthma. The odds of uncontrolled asthma increased with disease duration in all patients living in EJ tracts (P < .05). Living in the highest quartile of NO2 also increased the odds of uncontrolled asthma in patients with severe disease (P < .05), while there was no effect of TRAP on uncontrolled asthma in patients with less severe disease (P > .05). CONCLUSIONS: Living in an EJ tract increased the odds of severe and uncontrolled asthma and was influenced by age at onset, disease duration, and potentially by TRAP exposure. This study underscores the need to better understand the complex environmental interactions that affect lung health in groups that have been economically and/or socially marginalized.

Bronchial epithelial cell transcriptional responses to inhaled corticosteroids dictate severe asthmatic outcomes.

BACKGROUND: Inhaled corticosteroids (CSs) are the backbone of asthma treatment, improving quality of life, exacerbation rates, and mortality. Although effective for most, a subset of patients with asthma experience CS-resistant disease despite receiving high-dose medication. OBJECTIVE: We sought to investigate the transcriptomic response of bronchial epithelial cells (BECs) to inhaled CSs. METHODS: Independent component analysis was performed on datasets, detailing the transcriptional response of BECs to CS treatment. The expression of these CS-response components was examined in 2 patient cohorts and investigated in relation to clinical parameters. Supervised learning was used to predict BEC CS responses using peripheral blood gene expression. RESULTS: We identified a signature of CS response that was closely correlated with CS use in patients with asthma. Participants could be separated on the basis of CS-response genes into groups with high and low signature expression. Patients with low expression of CS-response genes, particularly those with a severe asthma diagnosis, showed worse lung function and quality of life. These individuals demonstrated enrichment for T-lymphocyte infiltration in endobronchial brushings. Supervised machine learning identified a 7-gene signature from peripheral blood that reliably identified patients with poor CS-response expression in BECs. CONCLUSIONS: Loss of CS transcriptional responses within bronchial epithelium was related to impaired lung function and poor quality of life, particularly in patients with severe asthma. These individuals were identified using minimally invasive blood sampling, suggesting these findings may enable earlier triage to alternative treatments.

Determinants of lung function across childhood in the Severe Asthma Research Program (SARP) 3.

BACKGROUND: Children with asthma are at risk for low lung function extending into adulthood, but understanding of clinical predictors is incomplete. OBJECTIVE: We sought to determine phenotypic factors associated with FEV1 throughout childhood in the Severe Asthma Research Program 3 pediatric cohort. METHODS: Lung function was measured at baseline and annually. Multivariate linear mixed-effects models were constructed to assess the effect of baseline and time-varying predictors of prebronchodilator FEV1 at each assessment for up to 6 years. All models were adjusted for age, predicted FEV1 by Global Lung Function Initiative reference equations, race, sex, and height. Secondary outcomes included postbronchodilator FEV1 and prebronchodilator FEV1/forced vital capacity. RESULTS: A total of 862 spirometry assessments were performed for 188 participants. Factors associated with FEV1 include baseline Feno (B, -49 mL/log2 PPB; 95% CI, -92 to -6), response to a characterizing dose of triamcinolone acetonide (B, -8.4 mL/1% change FEV1 posttriamcinolone; 95% CI, -12.3 to -4.5), and maximal bronchodilator reversibility (B, -27 mL/1% change postbronchodilator FEV1; 95% CI, -37 to -16). Annually assessed time-varying factors of age, obesity, and exacerbation frequency predicted FEV1 over time. Notably, there was a significant age and sex interaction. Among girls, there was no exacerbation effect. For boys, however, moderate (1-2) exacerbation frequency in the previous 12 months was associated with -20 mL (95% CI, -39 to -2) FEV1 at each successive year. High exacerbation frequency (≥3) 12 to 24 months before assessment was associated with -34 mL (95% CI, -61 to -7) FEV1 at each successive year. CONCLUSIONS: In children with severe and nonsevere asthma, several clinically relevant factors predict FEV1 over time. Boys with recurrent exacerbations are at high risk of lower FEV1 through childhood.

Safety and Efficacy of Dexpramipexole in Eosinophilic Asthma (EXHALE): A randomized controlled trial.

BACKGROUND: There is a need for new and effective oral asthma therapies. Dexpramipexole, an oral eosinophil-lowering drug, has not previously been studied in asthma. OBJECTIVE: We sought to evaluate the safety and efficacy of dexpramipexole in lowering blood and airway eosinophilia in subjects with eosinophilic asthma. METHODS: We performed a randomized, double-blind, placebo-controlled proof-of-concept trial in adults with inadequately controlled moderate to severe asthma and blood absolute eosinophil count (AEC) greater than or equal to 300/µL. Subjects were randomly assigned (1:1:1:1) to dexpramipexole 37.5, 75, or 150 mg BID (twice-daily) or placebo. The primary end point was the relative change in AEC from baseline to week 12. Prebronchodilator FEV1 week-12 change from baseline was a key secondary end point. Nasal eosinophil peroxidase was an exploratory end point. RESULTS: A total of 103 subjects were randomly assigned to dexpramipexole 37.5 mg BID (N = 22), 75 mg BID (N = 26), 150 mg BID (N = 28), or placebo (N = 27). Dexpramipexole significantly reduced placebo-corrected AEC week-12 ratio to baseline, in both the 150-mg BID (ratio, 0.23; 95% CI, 0.12-0.43; P < .0001) and the 75-mg BID (ratio, 0.34; 95% CI, 0.18-0.65; P = .0014) dose groups, corresponding to 77% and 66% reductions, respectively. Dexpramipexole reduced the exploratory end point of nasal eosinophil peroxidase week-12 ratio to baseline in the 150-mg BID (median, 0.11; P = .020) and the 75-mg BID (median, 0.17; P = .021) groups. Placebo-corrected FEV1 increases were observed starting at week 4 (nonsignificant). Dexpramipexole displayed a favorable safety profile. CONCLUSIONS: Dexpramipexole demonstrated effective eosinophil lowering and was well tolerated. Additional larger clinical trials are needed to understand the clinical efficacy of dexpramipexole in asthma.

CCL5 is a potential bridge between type 1 and type 2 inflammation in asthma.

BACKGROUND: Type 1 (T1) inflammation (marked by IFN-γ expression) is now consistently identified in subsets of asthma cohorts, but how it contributes to disease remains unclear. OBJECTIVE: We sought to understand the role of CCL5 in asthmatic T1 inflammation and how it interacts with both T1 and type 2 (T2) inflammation. METHODS: CCL5, CXCL9, and CXCL10 messenger RNA expression from sputum bulk RNA sequencing, as well as clinical and inflammatory data were obtained from the Severe Asthma Research Program III (SARP III). CCL5 and IFNG expression from bronchoalveolar lavage cell bulk RNA sequencing was obtained from the Immune Mechanisms in Severe Asthma (IMSA) cohort and expression related to previously identified immune cell profiles. The role of CCL5 in tissue-resident memory T-cell (TRM) reactivation was evaluated in a T1high murine severe asthma model. RESULTS: Sputum CCL5 expression strongly correlated with T1 chemokines (P < .001 for CXCL9 and CXCL10), consistent with a role in T1 inflammation. CCL5high participants had greater fractional exhaled nitric oxide (P = .009), blood eosinophils (P < .001), and sputum eosinophils (P = .001) in addition to sputum neutrophils (P = .001). Increased CCL5 bronchoalveolar lavage expression was unique to a previously described T1high/T2variable/lymphocytic patient group in the IMSA cohort, with IFNG trending with worsening lung obstruction only in this group (P = .083). In a murine model, high expression of the CCL5 receptor CCR5 was observed in TRMs and was consistent with a T1 signature. A role for CCL5 in TRM activation was supported by the ability of the CCR5 inhibitor maraviroc to blunt reactivation. CONCLUSION: CCL5 appears to contribute to TRM-related T1 neutrophilic inflammation in asthma while paradoxically also correlating with T2 inflammation and with sputum eosinophilia.

Strikingly High Activity of 15-Lipoxygenase Towards Di-Polyunsaturated Arachidonoyl/Adrenoyl-Phosphatidylethanolamines Generates Peroxidation Signals of Ferroptotic Cell Death.

The vast majority of membrane phospholipids (PLs) include two asymmetrically positioned fatty acyls: oxidizable polyunsaturated fatty acids (PUFA) attached predominantly at the sn2 position, and non-oxidizable saturated/monounsaturated acids (SFA/MUFA) localized at the sn1 position. The peroxidation of PUFA-PLs, particularly sn2-arachidonoyl(AA)- and sn2-adrenoyl(AdA)-containing phosphatidylethanolamines (PE), has been associated with the execution of ferroptosis, a program of regulated cell death. There is a minor subpopulation (≈1-2 mol %) of doubly PUFA-acylated phospholipids (di-PUFA-PLs) whose role in ferroptosis remains enigmatic. Here we report that 15-lipoxygenase (15LOX) exhibits unexpectedly high pro-ferroptotic peroxidation activity towards di-PUFA-PEs. We revealed that peroxidation of several molecular species of di-PUFA-PEs occurred early in ferroptosis. Ferrostatin-1, a typical ferroptosis inhibitor, effectively prevented peroxidation of di-PUFA-PEs. Furthermore, co-incubation of cells with di-AA-PE and 15LOX produced PUFA-PE peroxidation and induced ferroptotic death. The decreased contents of di-PUFA-PEs in ACSL4 KO A375 cells was associated with lower levels of di-PUFA-PE peroxidation and enhanced resistance to ferroptosis. Thus, di-PUFA-PE species are newly identified phospholipid peroxidation substrates and regulators of ferroptosis, representing a promising therapeutic target for many diseases related to ferroptotic death.

Investigations of a combination of atopic status and age of asthma onset identify asthma subphenotypes.

OBJECTIVE: Subphenotypes of asthma may be determined by age onset and atopic status. We sought to characterize early or late onset atopic asthma with fungal or non-fungal sensitization (AAFS or AANFS) and non-atopic asthma (NAA) in children and adults in the Severe Asthma Research Program (SARP). SARP is an ongoing project involving well-phenotyped patients with mild to severe asthma. METHODS: Phenotypic comparisons were performed using Kruskal-Wallis or chi-square test. Genetic association analyses were performed using logistic or linear regression. RESULTS: Airway hyper-responsiveness, total serum IgE levels, and T2 biomarkers showed an increasing trend from NAA to AANFS and then to AAFS. Children and adults with early onset asthma had greater % of AAFS than adults with late onset asthma (46% and 40% vs. 32%; P < 0.00001). In children, AAFS and AANFS had lower % predicted FEV1 (86% and 91% vs. 97%) and greater % of patients with severe asthma than NAA (61% and 59% vs. 43%). In adults with early or late onset asthma, NAA had greater % of patients with severe asthma than AANFS and AAFS (61% vs. 40% and 37% or 56% vs. 44% and 49%). The G allele of rs2872507 in GSDMB had higher frequency in AAFS than AANFS and NAA (0.63 vs. 0.55 and 0.55), and associated with earlier age onset and asthma severity. CONCLUSIONS: Early or late onset AAFS, AANFS, and NAA have shared and distinct phenotypic characteristics in children and adults. AAFS is a complex disorder involving genetic susceptibility and environmental factors.

Genetic analyses of chr11p15.5 region identify MUC5AC-MUC5B associated with asthma-related phenotypes.

OBJECTIVE: Genome-wide association studies (GWASs) have identified single nucleotide polymorphisms (SNPs) in chr11p15.5 region associated with asthma and idiopathic interstitial pneumonias (IIPs). We sought to identify functional genes for asthma by combining SNPs and mRNA expression in bronchial epithelial cells (BEC) in the Severe Asthma Research Program (SARP). METHODS: Correlation analyses of mRNA expression of six candidate genes (AP2A2, MUC6, MUC2, MUC5AC, MUC5B, and TOLLIP) and asthma phenotypes were performed in the longitudinal cohort (n = 156) with RNAseq in BEC, and replicated in the cross-sectional cohort (n = 155). eQTL (n = 114) and genetic association analysis of asthma severity (426 severe vs. 531 non-severe asthma) were performed, and compared with previously published GWASs of IIPs and asthma. RESULTS: Higher expression of AP2A2 and MUC5AC and lower expression of MUC5B in BEC were correlated with asthma, asthma exacerbations, and T2 biomarkers (P < 0.01). SNPs associated with asthma and IIPs in previous GWASs were eQTL SNPs for MUC5AC, MUC5B, or TOLLIP, however, they were not in strong linkage disequilibrium. The risk alleles for asthma or protective alleles for IIPs were associated with higher expression of MUC5AC and lower expression of MUC5B. rs11603634, rs12788104, and rs28415845 associated with moderate-to-severe asthma or adult onset asthma in previous GWASs were not associated with asthma severity (P > 0.8). CONCLUSIONS: SNPs associated with asthma in chr11p15.5 region are not associated with asthma severity neither with IIPs. Higher expression of MUC5AC and lower expression of MUC5B are risk for asthma but protective for IIPs.