Th2 Biomarkers Research Articles

Signature Proteins in Small Extracellular Vesicles of Granulocytes and CD4+ T-Cell Subpopulations Identified by Comparative Proteomic Analysis.

Several studies have described the proteomic profile of different immune cell types, but only a few have also analysed the content of their delivered small extracellular vesicles (sEVs). The aim of the present study was to compare the protein signature of sEVs delivered from granulocytes (i.e., neutrophils and eosinophils) and CD4+ T cells (i.e., TH1, TH2, and TH17) to identify potential biomarkers of the inflammatory profile in chronic inflammatory diseases. Qualitative (DDA) and quantitative (DIA-SWATH) analyses of in vitro-produced sEVs revealed proteome variations depending on the cell source. The main differences were found between granulocyte- and TH cell-derived sEVs, with a higher abundance of antimicrobial proteins (e.g., LCN2, LTF, MPO) in granulocyte-derived sEVs and an enrichment of ribosomal proteins (RPL and RPS proteins) in TH-derived sEVs. Additionally, we found differentially abundant proteins between neutrophil and eosinophil sEVs (e.g., ILF2, LTF, LCN2) and between sEVs from different TH subsets (e.g., ISG15, ITGA4, ITGB2, or NAMPT). A "proof-of-concept" assay was also performed, with TH2 biomarkers ITGA4 and ITGB2 displaying a differential abundance in sEVs from T2high and T2low asthma patients. Thus, our findings highlight the potential use of these sEVs as a source of biomarkers for diseases where the different immune cell subsets studied participate, particularly chronic inflammatory pathologies such as asthma or chronic obstructive pulmonary disease (COPD).

Serum inflammatory biomarkers associated with disease severity and response to dupilumab treatment in bullous pemphigoid: A cluster analysis

BackgroundDupilumab, a novel therapy targeting the T helper (Th) 2-mediated inflammation, is showing clinical benefits in treating bullous pemphigoid (BP). However, limited research investigated the serum biomarkers that reflect the inflammation alterations throughout the disease course. ObjectivesTo explore the changes of the serum inflammatory biomarkers under dupilumab therapy in BP and establish their correlations with disease severity and clinical outcomes. MethodsThis exploratory study evaluated serum samples from 40 patients with BP at baseline, 30 of these patients following 16-week dupilumab therapy, and 20 senior healthy controls. Serum levels of 29 cytokines and chemokines were quantified using the Magnetic Luminex Assay. ResultsTwo distinct clusters based on serum inflammatory profiles were identified. The first cluster, characterized by elevated levels of inflammatory activation, exhibited worse disease severity and poorer remission outcomes. Following the 16-week dupilumab therapy regimen, a significant suppression of Th2-mediated inflammation in the serum was observed, alongside a relative upregulation of Th1 responses. Patients treated with adjuvant systemic steroids exhibited an enhanced suppression of B cell activating factor compared to those receiving dupilumab alone. Significant correlations were unveiled between Th2 biomarkers and clinical scores, eosinophil counts, and anti-BP180 immunoglobulin G levels. Baseline levels of CCL18, Periostin, interleukin (IL)-6, and IL-16 constitute an optimal combination to distinguish between inflammatory clusters. ConclusionsCluster analysis of serum inflammatory biomarkers provided novel insights into the heterogeneity of the inflammation profiles in BP. Baseline levels of CCL18, Periostin, IL-6, IL-16 emerged as effective predictors for disease severity and therapy response to dupilumab.

Diagnostic Accuracy of Specific IgE Against Wheat and Rye in Flour-Induced Occupational Asthma

Assessment of IgE-mediated sensitization to flour allergens is widely used to investigate flour-induced occupational asthma. The diagnostic efficiency of detecting specific IgE antibodies (sIgEs) against wheat and rye flour, however, has not been thoroughly compared with other diagnostic procedures. We sought to evaluate the diagnostic accuracy of sIgE against wheat and rye compared with specific inhalation challenge (SIC) with flour as the reference standard. This retrospective multicenter study included 264 subjects who completed an SIC with flour in eight tertiary centers, of whom 205 subjects showed a positive SIC result. Compared with SIC, sIgE levels of 0.35 kUA/L or greater against wheat and rye provided similar sensitivities (84% to 85%, respectively), specificities (71% to 78%), positive predictive values (91% to 93%), and negative predictive values (56% to 61%). Increasing the threshold sIgE value to 5.10 kUA/L for wheat and to 6.20 kUA/L for rye provided a specificity of 95% or greater and further enhanced the positive predictive value to 98%. Among subjects with a positive SIC, those who failed to demonstrate sIgE against wheat and rye (n= 26) had significantly lower total serum IgE level and blood and sputum eosinophil counts and a lesser increase in postchallenge FeNO compared with subjects with a detectable sIgE. High levels of sIgE against wheat and/or rye flour strongly support a diagnosis of flour-induced occupational asthma without the need to perform an SIC. The absence of detectable sIgE against wheat and rye in subjects with a positive SIC seems to be associated with lower levels of TH2 biomarkers.

Identification of biologic-responsive phenotypes in elderly people with eosinophilic asthma

Asthma in the elderly is not as well studied as in younger age groups. Age-related immunosenescence may result in diminished TH2 inflammation, which raises a question about whether asthma in elderly patients responds well to anti-TH2 asthma biologics. We sought to determine whether asthma in elderly people has different TH2 biomarkers and clinical features compared to nonelderly people, and if disease in the 2 age groups responds differently to anti-TH2 biologics. We also aimed to identify treatment-responsive phenotypes with clinical and biomarker features that could be used to predict best response to biologics. A retrospective chart review was conducted for 56 patients (30 elderly [age ≥62 years] and 26 nonelderly [ages 18-59 years] subjects) with severe asthma treated with dupilumab or benralizumab. Differences in baseline characteristics and response to treatment were analyzed. Ahierarchical cluster analysis was also performed to identify treatment-responsive phenotypes. Significance threshold was P= .05 for all analyses. Baseline characteristics and TH2 biomarkers (blood eosinophil level, total IgE, aeroallergen sensitivity) were similar between elderly and nonelderly subjects. The disease in both groups responded well to biologics (improvement in ACT scores, decreased exacerbations, decreased need for prednisone), but no significant response difference was found based on age groups. Cluster analysis identified 3 phenotypes, as follows: cluster 1, youngest age, moderate eosinophil levels, lowest total IgE, few environmental allergies, and least response to biologics; cluster 2, intermediate age, lowest eosinophil level, highest IgE level, many environmental allergies, and an intermediate response to biologics; and cluster 3, oldest ages, highest eosinophil levels, high total IgE, few environmental allergies, and best response to biologics. These results confirm trends seen in another study utilizing cluster analyses showing that subjects with highest levels of IgE and eosinophils responded better to biologic treatment for asthma. Elderly people with asthma should be considered for biologic therapy no differently than younger people. There may be subgroups of patients with different biologic responses based on age, allergenicity, IgE, and eosinophil levels that could be used to predict treatment response.

Potential determinants of T helper 2 markers and their distribution in school-aged children

BackgroundThere is growing data on T helper 2 (Th2) biomarker determinants in adult populations. However, the determinants and typical range of these biomarkers have not been well studied in general populations of children. Therefore, we assessed the determinants and typical range of three Th2 biomarkers, including blood eosinophils, FeNO, and serum total IgE in 9-11-year-old children in a prospective birth cohort. MethodsWe examined the pre- and postnatal factors associated with Th2 biomarkers using multivariable logistic regression analysis (n = 428) and extended the results to the original cohort (n = 17,009) using inverse probability weighting. We also measured typical Th2 biomarker distribution in all examined children and healthy participants without allergic diseases (n = 180). ResultsAt age 9–11, wheeze (odds ratio (OR) 7.63), rhinitis (OR 3.14), and eczema (OR 2.46) were significantly associated with increased blood eosinophils. All three allergic conditions were associated with FeNO and total serum IgE, but the ORs were smaller than those for blood eosinophils. Secondhand smoking was inversely associated with the blood eosinophils (OR, 0.38). Similar results were found in the original cohort. Male sex and prenatal factors (maternal smoking and parental history of allergies) were not independent predictors of high Th2 levels. ConclusionsIn addition to wheezing and rhinitis, eczema and secondhand smoke exposure are independent factors for Th2 biomarker interpretation in children. Furthermore, the typical values and cutoff values of blood eosinophils in adults may not be applicable to children.

Smoking Status Modifies the Relationship between Th2 Biomarkers and Small Airway Obstruction in Asthma.

Background Cigarette smoking and Th2-inflammation are both crucial in the pathogenesis of asthma. However, it is unknown whether smoking can affect the association between Th2-inflammation and small airway obstruction in adults with asthma. Methods Adults diagnosed with asthma by a pulmonologist according to Global Initiative for Asthma guidelines were recruited from September 2016 to April 2018 to participate in this study. Participants were divided into two groups, the small airway obstruction group (those with FEF25–75% predicted value ≤ 65%) and the normal small airway function group (those with FEF25–75% predicted value > 65%). Final data analysis included 385 and 93 people in the Obstructive Group and the Normal Group, respectively. Total serum IgE level and blood eosinophil count were used as biomarkers of the Th2 phenotype. Results The Obstructive Group had a larger fraction of smokers, higher blood eosinophil count, and lower lung function than the Normal Group. Current-smoking status was associated with an increased risk of small airway obstruction (adjusted odds ratio = 4.677, 95% confidence interval [1.593–13.730]); and log-IgE level was associated with a decreased risk of small airway obstruction (0.403 [0.216–0.754]). Smoking status stratified analysis showed an association between log-IgE level and a decreased risk of small airway obstruction only in never-smoker asthmatics (0.487 [0.249–0.954]). Conclusions Current-smoking status and total serum IgE are, respectively, associated with small airway obstruction. Smoking status modifies the relationship between Th2 biomarkers and small airway function. These findings contribute to the understanding of risk factors associated with asthma endotyping.

Abstract 2793: The role of IL-10 in obesity-induced T-cell polarization in the prostate tumor microenvironment

Abstract Background: Prostate cancer remains the second leading cause of cancer related deaths in men and obesity has been shown to severely impact its prognosis. Previous studies in our laboratory have shown that CD4+ T cells polarized under obese conditions are prostate cancer promoting and that this is due in part to suppression of IFNγ and induction of IL-6 in the T cells. Objective: The goal of this study is to determine the role of IL-10 in mediating the impact of obese conditions on T cell phenotype. Methods: A series of in vitro studies were conducted to evaluate the effects of obesity on T cell function and polarization. Splenic CD4+ T-cells from 10-week-old, male, C57BI/6 mice were isolated and stimulated with anti-CD3+/CD28+ antibodies under obese conditions. Gene expression was measured using qPCR and AKT phosphorylation was measured using Western blot analysis. In vivo T-cell function was compared between obese and non-obese male mice. Deletion assays were used to determine the role of IL-10 in mediating the effects observed on T cell phenotype in response to obese conditions. Results: For both the in vitro and in vivo studies, obese conditions suppressed the expression of Th1 biomarkers (IFNγ, IL-2, and T-bet) and upregulated the expression of Th2 biomarkers (IL-6, IL-10, and GATA-3) in CD4+ T cells. Likewise, obese conditions in vitro and in vivo were associated with decreased AKT phosphorylation. Studies are on-going to determine whether inhibiting the IL-10 pathway restores proper T cell activation under obese conditions. Conclusion: Obesity induces T cell dysfunction characterized by a Th1 to Th2 phenotypic shift, resulting in a pro-tumorigenic phenotype, potentially mediated by paracrine IL-10 signaling. These results are important in understanding the mechanisms by which obesity promotes a more aggressive disease, thereby identifying potential targets for improving outcomes. The results from these studies will provide a better understanding of the role of IL-10 as a novel target for preventing prostate cancer in obese subjects and improving overall survival. Citation Format: Margaret Kappel, Michael Adkison, Alejandra De Angulo, Peyton Travis, Brittany Harlow, Christopher Jolly, Linda deGraffenried. The role of IL-10 in obesity-induced T-cell polarization in the prostate tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2793.

Facial erythema after the treatment of dupilumab in SLE patient

BackgroundDupilumab is a receptor antagonist binding to the alpha subunit of the interleukin-4 receptor. Through binding to it, dupilumab inhibits signaling of both IL-4 and IL-13, the representative Th2 biomarkers. Recently, in addition to the treatment effects for atopic dermatitis (AD), there is an emerging adverse event as facial erythema.Case presentationA twenty-seven-year-old female patient developed erythema and desquamation on the face and neck after dupilumab administration. She had AD on her arms, legs, and trunk before the treatment but there was no atopic clinical feature in her face and neck. With the treatment of dupilumab, her skin lesions of the body have improved from the beginning of the treatment. In the patch test, including dupilumab, there was no specific finding other than the 1+ response to neomycin on day 2. In the intradermal test to dupilumab, a positive result was observed 15 min later, but negative both days 1 and 2. The blood examination showed an elevation of both ANA as 1:80 and anti-phospholipid antibodies (Anti-cardiolipin IgM, IgG, and Anti- beta 2 GPI IgG). She was diagnosed with Systemic lupus erythematosus (SLE) based on diagnostic criteria by a rheumatologist.ConclusionDupilumab is an emerging therapeutic agent for AD, and treatment cases are increasing in Korea. However, there are several adverse events during the treatment of dupilumab. Herein, we report the unexpected adverse event during the treatment of dupilumab in SLE patients.

Characterization of chronic obstructive pulmonary disease in dairy farmers

BackgroundAlthough farming is often considered a risk factor for COPD, data regarding the burden and characteristics of COPD in dairy farmers are sparse and conflicting. ObjectivesTo characterize COPD in dairy farmers. Methods4788 subjects entered two parallel COPD screening programs, one in agricultural workers and one in general practice from 2011 to 2015. Subjects with COPD were invited to participate in the characterization phase of the study. Those who accepted were included in two subgroups: dairy farmers with COPD (DF-COPD) (n = 101) and non-farmers with COPD (NF-COPD) (n = 85). Patients with COPD were frequency-matched with subjects with normal spirometry for age, sex and tobacco smoking (pack-years and status) (DF-controls n = 98, NF-controls n = 89). All subjects from these four groups underwent lung function and exercise testing, questionnaires and blood analysis. ResultsThe frequency of COPD in dairy farmers was 8.0% using the GOLD criterion and 6.2% using the lower limit of normal criterion and was similar in non-farming subjects (7.3% and 5.2%, respectively) although dairy farmers had lower tobacco consumption (screening phase). DF-COPD had better pulmonary function, exercise capacity and quality of life, fewer symptoms and comorbidities than NF-COPD, and higher levels of some Th2 biomarkers (MCP-2, periostin) (characterization phase). In farmers, COPD was not related to occupational exposure factors, supporting the role of host factors. ConclusionCOPD secondary to organic dust exposure (dairy farming) appears less severe and associated with fewer comorbidities than COPD secondary to tobacco smoking.

Ruxolitinib Cream Suppresses Th2 Inflammation in Adult Patients With Atopic Dermatitis

Ruxolitinib cream, a selective inhibitor of Janus kinases (JAK)-1 and JAK2, potently inhibits proinflammatory cytokine signaling involved in atopic dermatitis (AD) pathogenesis. In a phase 2 trial, 1.5% ruxolitinib cream twice daily (BID) demonstrated efficacy at least comparable to triamcinolone. Rapid decreases in itching were observed. The effects of ruxolitinib cream on Th2 biomarkers and the relationship with clinical measures in this trial are reported below. Sera were collected from 111 patients treated with ruxolitinib cream 0.15% once daily (QD; n=17), 0.5% QD (n=20), 1.5% QD (n=15), 1.5% BID (n=18), triamcinolone BID (n=19), and vehicle BID (n=22). Serum immunoglobulin E (IgE) and thymus and activation-regulated chemokine (TARC/CCL17) were analyzed at baseline and Week 8. Differences between vehicle and each treatment arm were determined using mixed-model repeated measures; significance was conferred at P<0.05. Application of ruxolitinib cream 1.5% BID for 8 weeks significantly (P=0.0095) reduced TARC/CCL17 levels in circulation versus vehicle-treated patients. Additionally, total serum IgE levels were reduced in patients treated with 1.5% ruxolitinib cream (QD or BID). No material differences were observed with 0.15% QD or 0.5% QD ruxolitinib. Efficacy analyses by baseline TARC/CCL17 (≤522 vs >522 pg/mL) and total IgE subgroups (<200 vs ≥200 kU/L) were performed for percentage changes from baseline in Eczema Area and Severity Index (EASI). Levels of TARC/CCL17 and IgE at baseline did not predict ruxolitinib cream treatment response (percentage reductions in EASI). Treatment of patients with AD with ruxolitinib cream inhibited select Th2 biomarkers, suggesting its ability to modulate disease course.

Precision medicine in asthma: the role of bronchial thermoplasty.

The inflammatory makeup of severe asthma is heterogeneous. Identification of the predominant cellular endotype via biomarkers can aid in the selection of more advanced therapies. This review is clinically focused on how to use these biomarkers to help select between biologic agents and/or bronchial thermoplasty. Several Th2 biomarkers exist for the detection of eosinophilic disease; however, the best biomarker for clinical practice is debatable depending upon local resources. Currently, there are three federal drug agency-approved biologic agents (omalizumab, mepolizumab and reslizumab) to treat severe asthma with frequent exacerbations despite standard medical therapy. Several others are either in clinical trials or in the development phase for the treatment of eosinophilic asthma. To date, agents targeting neutrophilic inflammation have been largely unsuccessful. Bronchial thermoplasty has emerged as an option for the treatment of severe asthma. The appropriate selection of patients through the use of eosinophilic biomarkers has led to significant reductions in exacerbations with the use of mAb therapy. Bronchial thermoplasty has also shown reductions in asthma exacerbations and improved quality of life; however, it is unclear which patients may respond best to this intervention.

Biologics in practice: A unique opportunity for allergist/immunologist expertise

Biologics in practice: A unique opportunity for allergist/immunologist expertise

Biomarker Profiles in Asthma With High vs Low Airway Reversibility and Poor Disease Control

High bronchodilator reversibility in adult asthma is associated with distinct clinical characteristics. This analysis compares lung function, biomarker profiles, and disease control in patients with high reversibility (HR) and low reversibility (LR) asthma. A retrospective analysis was performed with data from two completed clinical trials of similar design. Patients were divided into HR and LR subgroups based on their response to bronchodilators (HR = ΔFEV1 postbronchodilator ≥ 20%). Blood eosinophil count, serum IgE level, and fraction of exhaled nitric oxide concentration, biomarkers commonly used to stratify patients into T-helper (Th)-2-high vs Th2-low phenotypes, were measured in patients with not well controlled (1.5 ≤ Asthma Control Questionnaire [ACQ] ≤ 2.143) and very poorly controlled (ACQ > 2.143) disease. The majority of patients in the HR and LR subgroups displayed Th2-low biomarker profiles and very poor disease control. HR was more frequently associated with Th2-high biomarker profiles (40.1% vs 29.4%, P = .006), lower lung function (FEV1, 63.5 ± 7.7% predicted vs 67.9 ± 8.4% predicted; P < .001), and atopy (93.7% vs 86.5%, P = .005). HR is a physiologic indicator of reduced lung function and is more often associated with elevations in Th2 biomarkers than LR in moderate to severe asthma. However, the majority of patients with HR and LR asthma in this analysis had a Th2-low biomarker profile. Moreover, a Th2-high biomarker profile was not associated with worse disease control.

Obesity shifts house dust mite-induced airway cellular infiltration from eosinophils to macrophages: effects of glucocorticoid treatment.

Although classically characterized by chronic airway inflammation with eosinophil infiltration, asthma is a complex and multifactorial condition with numerous clinical phenotypes. Epidemiological studies strongly support the link between obesity and asthma and suggest that obesity precedes and promotes asthma development, increases asthma severity, and reduces steroid responsivity. Using a house dust mite (HDM) model of airway hyperresponsiveness in C57BL/6 mice, we examined the effects of diet-induced obesity on allergic airway inflammation and its treatment with dexamethasone. When compared to lean mice treated with HDM, obese-HDM mice had reduced plasma adiponectin, an anti-inflammatory adipokine, lower eosinophil and higher macrophage infiltration into the lungs and bronchoalveolar lavage (BAL) fluid, increased expression of total, M1, and M2 macrophage markers in the lungs, and enhanced Th2 and non-Th2 cytokine expression in the lungs. While Th2-associated responses in obese-HDM mice were suppressed by systemic dexamethasone, several Th2-independent responses, including total and M1 macrophage markers in the lungs, and lung CXC-motif ligand 1 (CXCL1) levels, were not improved following dexamethasone treatment. Thus, HDM combined with obesity promotes mixed localized inflammatory responses (e.g., M1, M2, Th1, and Th2) and shifts the cellular infiltration from eosinophils to macrophages, which are less sensitive to dexamethasone regulation. Because obese asthmatics exhibit more severe symptoms, lack a predominance of Th2 biomarkers, and are predicted to experience more steroid resistance when compared to lean asthmatics, this model could be used to study blunted steroid responses in obese-HDM mice and to define the macrophages found in the lungs.

A qPCR‐based metric of Th2 airway inflammation in asthma

BackgroundUsing microarray profiling of airway epithelial cells, we previously identified a Th2-high molecular phenotype of asthma based on expression of periostin, CLCA1 and serpinB2 and characterized by specific inflammatory, remodeling, and treatment response features. The goal of the current study was to develop a qPCR-based assay of Th2 inflammation to overcome the limitations of microarray-based methods.MethodsAirway epithelial brushings were obtained by bronchoscopy from two clinical studies comprising 44 healthy controls and 62 subjects with asthma, 39 of whom were studied before and after a standardized 8 week course of inhaled corticosteroids (ICS). The qPCR-based expression of periostin, CLCA1 and serpinB2 were combined into a single metric.ResultsIn asthma, the three-gene-mean of periostin, CLCA1 and serpinB2 correlated with FeNO (r = 0.75, p = 0.0002), blood eosinophils (r = 0.58, p = 0.003) and PC20 methacholine (r = -0.65, p = 0.0006), but not total serum IgE (r = 0.33, p = 0.1). Higher baseline three-gene-mean correlated with greater improvement in FEV1 with ICS at 2, 4 and 8 weeks (all p < 0.05). By ROC analysis, the area under the curve (AUC) of the three-gene-mean for FEV1 improvement with ICS at 4 and 8 weeks was 0.94 and 0.87, respectively, which are higher than the AUCs of FeNO, blood eosinophils, IgE or PC20. Th2 airway inflammation as measured by this three-gene-mean also had predictive capacity for an improvement in symptoms.ConclusionsThe three-gene-mean of periostin, CLCA1 and serpinB2 in airway epithelial brushings identifies Th2-high and low populations, is correlated with other Th2 biomarkers, and performs well for prediction of FEV1 improvement with ICS. The three-gene-mean provides a measurement of Th2 airway inflammation that is clinically relevant and that can serve as a valuable tool to evaluate non-invasive biomarkers to predict treatment responses to existing and emerging asthma therapies.