COVID-19 in a severe eosinophilic asthmatic receiving benralizumab - a case study.

INTRODUCTION: Only little is known about COVID-19 in patients with asthma. There is no data on COVID-19 in patients with severe asthma or patients with asthma who are treated with monoclonal antibodies. CASE STUDY: Here, we present the case of a severe eosinophilic asthmatic in whom benralizumab treatment, an anti-IL-5R monoclonal antibody, was initiated 2 years ago. Prior to benralizumab treatment, every viral infection had resulted in a prolonged course of oral corticosteroids (OCS). Since initiation of benralizumab, the patient has had good asthma control. Mid-March 2020, the patient developed high fever. RESULTS: A SARS-CoV-2-PCR (nasopharyngeal swab) was positive. The patient's symptoms subsided after few days. No OCS was needed. The asthma control questionnaire 6-item scale worsened moderately in the week of the infection and returned to normal levels thereafter. The asthma control test, measuring longer term asthma control, showed no decline. CONCLUSION: The course of COVID-19 was very mild in this particular patient with severe eosinophilic asthma. So far, there is no evidence that would suggest a more severe course of COVID-19 in patients with asthma. It is worth noting, that prior to the initiation of benralizumab this patient had multiple exacerbations per year triggered by viral infections (4/year), which all required OCS. Whilst only anecdotal, this case study provides the first evidence to support the current recommendation of continuing monoclonal antibodies in patients with severe asthma during the COVID-19 pandemic.

Effectiveness of mepolizumab therapy in patients with severe eosinophilic asthma: Austrian real-life data.

BACKGROUND: Mepolizumab was effective in several randomized clinical trials (RCTs) in patients with severe eosinophilic asthma, but evidence for symptom control in a real-world population is scarce. OBJECTIVE: To assess asthma symptom control, lung function, use of oral corticosteroids, and biomarkers after mepolizumab initiation in real-world clinical practice. METHODS: Thirty-five adult patients with severe eosinophilic asthma and inadequate asthma symptom control, including former smokers and patients with cardiac disease, were enrolled in a prospective single-arm real-world study. Asthma control tests (ACT), exacerbations, spirometry (pre-bronchodilator forced expiratory volume in 1 s [FEV1]), and oral corticosteroid doses were documented. Further assessments included peripheral blood eosinophil counts and adverse events. RESULTS: After mepolizumab initiation asthma symptom control was significantly improved with the median ACT score of 12.5 at baseline (interquartile range [IQR ]10.5-19.5) rising to 19 (15-22.5) after 4 weeks. The improvement was maintained throughout the observation period of 20 weeks. Likewise, exacerbations were reduced. After 8 weeks of mepolizumab daily OCS doses were reduced from 6.25 mg daily (0-20) at baseline to 2.5 mg daily (0-11.9) at week 8 (P < 0.001). FEV1 remained generally unchanged during the course of the study. Eosinophil counts rapidly declined and remained at a low level during the observation period. No new safety signals were observed in this study. CONCLUSION: Mepolizumab improved asthma symptom control and had a steroid-sparing effect. Efficacy in this real-world study was comparable to RCTs, despite a history of smoking and comorbidities in many of the patients included.

Safety, pharmacokinetics and pharmacodynamics of a novel anti-asthmatic drug, XC8, in healthy probands.

INTRODUCTION: XC8 (histamine glutarimide) is a novel agent which targets eosinophilic migration and mast cell degranulation and has shown anti-asthmatic effects in animal studies. OBJECTIVE: The objective of this placebo-controlled phase 1 study was to assess the safety of oral XC8 and to evaluate its pharmacokinetic and pharmacodynamic properties. METHODS: 32 healthy volunteers in three dose-escalation treatment groups (10 mg [n = 8], 50 mg [n = 8] and 200 mg [n = 16]) were randomized in a 3:1 ratio to XC8 or placebo respectively. The subjects received a single dose of the drug at Day 1 and then once-daily for 14 days (Days 8-21). RESULTS: No severe adverse events occurred. The number of adverse events was similar in the treatment arms compared to placebo and all subjects completed the study as planned. No clinically significant changes occurred in hematologic and biochemical blood tests in subjects receiving XC8. The pharmacokinetic data showed similar dose and time dependent mean plasma XC8 concentrations after single (Day 1) and multiple (Day 21) dosing. The mean maximum concentrations were 114-1993 ng/mL after single and 115-2089 ng/mL after multiple dosing. The mean times to maximum concentration were 0.68-1.01 and 0.67-0.98 h, respectively. There was no evidence for accumulation of XC8 after multiple dosing. CONCLUSION: XC8 was safe and well tolerated. A phase 2 study is being performed to further evaluate the potential role of XC8 in asthma treatment. TRIAL REGISTRATION: ClinicalTrials.gov, NCT02882217.

CD23 surface density on B cells is associated with IgE levels and determines IgE-facilitated allergen uptake, as well as activation of allergen-specific T cells.

BACKGROUND: Increasing evidence suggests that the low-affinity receptor for IgE, CD23, plays an important role in controlling the activity of allergen-specific T cells through IgE-facilitated allergen presentation. OBJECTIVE: We sought to determine the number of CD23 molecules on immune cells in allergic patients and to investigate whether the number of CD23 molecules on antigen-presenting cells is associated with IgE levels and influences allergen uptake and allergen-specific T-cell activation. METHODS: Numbers of CD23 molecules on immune cells of allergic patients were quantified by using flow cytometry with QuantiBRITE beads and compared with total and allergen-specific IgE levels, as well as with allergen-induced immediate skin reactivity. Allergen uptake and allergen-specific T-cell activation in relation to CD23 surface density were determined by using flow cytometry in combination with confocal microscopy and T cells transfected with the T-cell receptor specific for the birch pollen allergen Bet v 1, respectively. Defined IgE-allergen immune complexes were formed with human monoclonal allergen-specific IgE and Bet v 1. RESULTS: In allergic patients the vast majority of CD23 molecules were expressed on naive IgD+ B cells. The density of CD23 molecules on B cells but not the number of CD23+ cells correlated with total IgE levels (RS = 0.53, P = .03) and allergen-induced skin reactions (RS = 0.63, P = .008). Uptake of allergen-IgE complexes into B cells and activation of allergen-specific T cells depended on IgE binding to CD23 and were associated with CD23 surface density. Addition of monoclonal IgE to cultured PBMCs significantly (P = .04) increased CD23 expression on B cells. CONCLUSION: CD23 surface density on B cells of allergic patients is correlated with allergen-specific IgE levels and determines allergen uptake and subsequent activation of T cells.

Frequent occurrence of T cell-mediated late reactions revealed by atopy patch testing with hypoallergenic rBet v 1 fragments.

BACKGROUND: Late allergic reactions are common in the course of allergen-specific immunotherapy and even occur with allergy vaccines with reduced IgE reactivity. OBJECTIVE: We sought to study atopy patch test (APT) reactions and T-cell responses to the recombinant birch pollen allergen Bet v 1 and recombinant hypoallergenic T-cell epitope-containing Bet v 1 fragments in patients with birch pollen allergy with and without atopic dermatitis (AD). METHODS: A clinical study was conducted in 15 patients with birch pollen allergy with AD (group 1), 5 patients with birch pollen allergy without AD (group 2), 5 allergic patients without birch pollen allergy (group 3), and 5 nonallergic subjects (group 4) by performing skin prick tests and APTs with rBet v 1 and hypoallergenic rBet v 1 fragments. T-cell, cutaneous lymphocyte antigen (CLA)(+) and CCR4(+) T-cell and cytokine responses were studied by thymidine uptake, carboxyfluorescein diacetate succinimidyl ester staining, and Luminex technology, respectively. RESULTS: rBet v 1 and hypoallergenic rBet v 1 fragments induced APT reactions in not only most of the patients with birch pollen allergy with AD (11/15) but also in most of those without AD (4/5). Patients with birch pollen allergy with AD had higher Bet v 1-specific proliferation of CLA(+) and CCR4(+) T cells compared with patients with birch pollen allergy without AD. There were no differences in Bet v 1-specific CLA(+) and CCR4(+) proliferation and cytokine secretion in patients with and without APT reactions. CONCLUSION: Hypoallergenic rBet v 1 fragments induce T cell-dependent late reactions not only in patients with birch pollen allergy with AD but also in those without AD, which can be determined based on APT results but not based on in vitro parameters.

Development and characterization of a recombinant, hypoallergenic, peptide-based vaccine for grass pollen allergy.

BACKGROUND: Grass pollen is one of the most important sources of respiratory allergies worldwide. OBJECTIVE: This study describes the development of a grass pollen allergy vaccine based on recombinant hypoallergenic derivatives of the major timothy grass pollen allergens Phl p 1, Phl p 2, Phl p 5, and Phl p 6 by using a peptide-carrier approach. METHODS: Fusion proteins consisting of nonallergenic peptides from the 4 major timothy grass pollen allergens and the PreS protein from hepatitis B virus as a carrier were expressed in Escherichia coli and purified by means of chromatography. Recombinant PreS fusion proteins were tested for allergenic activity and T-cell activation by means of IgE serology, basophil activation testing, T-cell proliferation assays, and xMAP Luminex technology in patients with grass pollen allergy. Rabbits were immunized with PreS fusion proteins to characterize their immunogenicity. RESULTS: Ten hypoallergenic PreS fusion proteins were constructed, expressed, and purified. According to immunogenicity and induction of allergen-specific blocking IgG antibodies, 4 hypoallergenic fusion proteins (BM321, BM322, BM325, and BM326) representing Phl p 1, Phl p 2, Phl p 5, and Phl p 6 were included as components in the vaccine termed BM32. BM321, BM322, BM325, and BM326 showed almost completely abolished allergenic activity and induced significantly reduced T-cell proliferation and release of proinflammatory cytokines in patients' PBMCs compared with grass pollen allergens. On immunization, they induced allergen-specific IgG antibodies, which inhibited patients' IgE binding to all 4 major allergens of grass pollen, as well as allergen-induced basophil activation. CONCLUSION: A recombinant hypoallergenic grass pollen allergy vaccine (BM32) consisting of 4 recombinant PreS-fused grass pollen allergen peptides was developed for safe immunotherapy of grass pollen allergy.

A nonallergenic birch pollen allergy vaccine consisting of hepatitis PreS-fused Bet v 1 peptides focuses blocking IgG toward IgE epitopes and shifts immune responses to a tolerogenic and Th1 phenotype.

Allergen-specific immunotherapy is the only allergen-specific and disease-modifying treatment for allergy. The construction and characterization of a vaccine for birch pollen allergy is reported. Two nonallergenic peptides, PA and PB, derived from the IgE-reactive areas of the major birch pollen allergen Bet v 1 were fused to the hepatitis B surface protein, PreS, in four recombinant fusion proteins containing different numbers and combinations of the peptides. Fusion proteins expressed in Escherichia coli and purified to homogeneity showed a lack of IgE reactivity and allergenic activity when tested with sera and basophils from patients allergic to birch pollen. Compared to Bet v 1 allergen, peptides PA and PB showed reduced T cell activation in PBMCs from allergic patients, whereas PreS fusion proteins induced less IL-5 and more IL-10 and IFN-γ. Immunization of rabbits with the fusion proteins, in particular with a PreS fusion protein 2PAPB-PreS, containing two copies of each peptide, induced high levels of IgG Abs against the major IgE-reactive site on Bet v 1 and related allergens. These IgG Abs inhibited allergic patients' IgE binding to Bet v 1 better than did IgG induced by immunization with complete Bet v 1. Furthermore, 2PAPB-PreS-induced IgG inhibited Bet v 1-induced basophil activation in allergic patients and CD23-facilitated allergen presentation. Our study exemplifies novel beneficial features for a PreS carrier-based peptide vaccine for birch pollen, which, in addition to the established reduction in allergenic activity, include the enhanced focusing of blocking Ab responses toward IgE epitopes, immunomodulatory activity, and reduction of CD23-facilitated allergen presentation.

An Observational Study to Determine the Real-Life Effectiveness of MP-AzeFlu® in Austrian Patients with Persistent Allergic Rhinitis.

BACKGROUND: Many patients with allergic rhinitis (AR) have moderate-to-severe persistent AR. Meda Pharma's AzeFlu (MP-AzeFlu®) is an intranasal AR treatment comprising a novel formulation of azelastine hydrochloride and fluticasone propionate in a single device. METHODS: This prospective observational study of 214 adults and adolescents in Austria with moderate-to-severe persistent AR assessed the effectiveness of MP-AzeFlu (one spray/nostril twice daily; daily doses: azelastine hydrochloride 548 µg; and fluticasone propionate 200 µg) for AR control in clinical practice using the visual analog scale. Symptom severity was reported on days 0, 1, 3, 7, 14, 21, 28, 35, and 42. Patient demographics, AR phenotype, allergen sensitization, symptomatology, AR treatments in the previous year, and the reason for the MP-AzeFlu prescription were recorded. RESULTS: MP-AzeFlu treatment was associated with a rapid and statistically significant reduction in the visual analog scale score from baseline to each timepoint measured, including day 1 (all p < 0.0001). Mean (standard deviation) visual analog scale score was 53.5 mm (26.3) at baseline, 25.3 mm (21.0) on day 28, and 19.6 mm (17.4) on day 42, a mean overall reduction from baseline of 41.4 (23.9) mm for completers. Results were consistent irrespective of patient age, gender, severity, or traditional AR phenotype. Prior to MP-AzeFlu prescription, congestion was considered the most bothersome symptom. The majority of patients reported using at least two AR therapies in the past year, including oral antihistamines, intranasal corticosteroids, and intranasal antihistamines. CONCLUSIONS: Many patients in Austria live with uncontrolled persistent AR despite treatment. MP-AzeFlu provides effective and rapid control of persistent AR in a real-world Austrian setting.

Mapping of conformational IgE epitopes with peptide-specific monoclonal antibodies reveals simultaneous binding of different IgE antibodies to a surface patch on the major birch pollen allergen, Bet v 1.

Allergic inflammation is based on the cross-linking of mast cell and basophil-bound IgE Abs and requires at least two binding sites for IgE on allergens, which are difficult to characterize because they are often conformational in nature. We studied the IgE recognition of birch pollen allergen Bet v 1, a major allergen for >100 million allergic patients. Monoclonal and polyclonal Abs raised against Bet v 1-derived peptides were used to compete with allergic patients' IgE binding to Bet v 1 to search for sequences involved in IgE recognition. Strong inhibitions of patients' IgE binding to Bet v 1 (52-75%) were obtained with mAbs specific for two peptides comprising aa 29-58 (P2) and aa 73-103 (P6) of Bet v 1. As determined by surface plasmon resonance, mAb2 specific for P2 and mAb12 specific for P6 showed high affinity, but only polyclonal rabbit anti-P2 and anti-P6 Abs or a combination of mAbs inhibited allergen-induced basophil degranulation. Thus, P2 and P6 define a surface patch on the Bet v 1 allergen, which allows simultaneous binding of several different IgE Abs required for efficient basophil and mast cell activation. This finding explains the high allergenic activity of the Bet v 1 allergen. The approach of using peptide-specific Abs for the mapping of conformational IgE epitopes on allergens may be generally applicable. It may allow discriminating highly allergenic from less allergenic allergen molecules and facilitate the rational design of active and passive allergen-specific immunotherapy strategies.

Glucocorticosteroids rescue basophils from dasatinib-augmented immunoglobulin E-mediated histamine release.

BACKGROUND: Dasatinib is a multikinase inhibitor active against several tyrosine kinases including ABL, KIT, Lyn and Btk. Apart from its known antileukemic activity, the drug produces several side effects including edemas and pleural effusions, which are supposedly triggered by activated immune cells. Effusion formation can be treated effectively by glucocorticosteroids. We have recently shown that low concentrations of dasatinib (<0.1 µM) promote IgE-dependent secretion of histamine in basophils, especially in allergic individuals. In the current study, we asked whether glucocorticosteroids inhibit dasatinib-induced activation of basophils. METHODS: Basophils were preincubated with dexamethasone, prednisolone and hydrocortisone for 24 h, and were then exposed to an anti-IgE antibody (normal basophils) or the allergens Bet v 1 and Phl p 5 (allergic patients) with or without low concentrations of dasatinib (0.025 µM). After incubation, basophils were examined for histamine release and expression of CD63 and CD203c. RESULTS: All three glucocorticosteroids were found to counteract IgE-dependent and dasatinib-enhanced histamine release in basophils in nonallergic and allergic individuals. In addition, glucocorticosteroids were found to inhibit anti-IgE-induced upregulation of CD63 and CD203c in the presence or absence of dasatinib. The inhibitory effects of glucocorticosteroids were dose-dependent (effective range: 1-10 µM) and seen in all donors examined. CONCLUSIONS: Glucocorticosteroids rescue IgE receptor cross-linked basophils from additional costimulatory effects of low-dose dasatinib which may have clinical implications in dasatinib-treated patients.

Recombinant allergens: what does the future hold?

This year we are celebrating not only the centenary of allergen-specific immunotherapy but also the 10-year anniversary of the first administration of recombinant allergen-based vaccines to allergic patients. By using recombinant DNA technology, defined and safe allergy vaccines can be produced that allow us to overcome many, if not all, of the problems associated with the use of natural allergen extracts, such as insufficient quality, allergenic activity, and poor immunogenicity. Here we provide an update of clinical studies with recombinant allergen-based vaccines, showing that some of these vaccines have undergone successful clinical evaluation up to phase III studies. Furthermore, we introduce a strategy for allergen-specific immunotherapy based on recombinant fusion proteins consisting of viral carrier proteins and allergen-derived peptides without allergenic activity, which holds the promise of being free of side effects and eventually being useful for prophylactic vaccination.

Expression of a major plant allergen as membrane-anchored and secreted protein in human cells with preserved T cell and B cell epitopes.

BACKGROUND: Expression of allergens in human cells is a prerequisite for the development of antigen-specific cell therapy in IgE-mediated allergy. We developed a strategy how the clinically relevant major grass pollen allergen Phl p 5 can be efficiently secreted or expressed on the surface of human cells with preserved allergenic activity. METHODS: The cDNA of Phl p 5 was fused to a leader peptide with or without a transmembrane domain and both constructs were ligated into a mammalian expression vector. Transfection of these plasmids into human cells resulted in a membrane-anchored or secreted version of Phl p 5, respectively, as determined by ELISA or flow cytometric analysis. RESULTS: Both the secreted and membrane-anchored Phl p 5 proteins bound IgE from allergic patients in an immunoblot assay and induced specific histamine release and CD203c upregulation in basophils of grass pollen-allergic patients. Proliferation of peripheral blood mononuclear cells from Phl p 5-allergic individuals was induced upon stimulation with both variants of Phl p 5 expressed in human cells similar to recombinant Phl p 5. CONCLUSIONS: Secreted and membrane-anchored Phl p 5 expressed in human cells preserved B cell as well as T cell epitopes and may be used to develop and test various cell-based strategies for allergen-specific immunomodulation and to delineate the tolerance mechanisms involved therein.

TRICOP - A Real-world effectiveness study with a single-inhaler extrafine triple therapy over 52 weeks in Austrian patients with COPD.

OBJECTIVE: Evidence of the efficacy of single-inhaler triple therapy in COPD patients inferred from RCTs has not been assessed in a real-world setting in Austria. In this non-interventional study (NIS) tolerability and effectiveness of extrafine beclometasone-dipropionate, formoterol-fumarate and glycopyrronium (Trimbow® 87/5/9 µg) was evaluated in COPD patients. METHODS: A prospective NIS was conducted over 52 weeks in 24 sites in Austria. Eligible COPD patients had an indication for treatment with single-inhaler BDP/FF/G. In this study tolerability, lung function, exacerbation rate, symptom scores and CAT scores were recorded. RESULTS: 265 patients with moderate to very severe airflow limitation (GOLD Grade 2-4: 96.2%) and persistent symptoms (GOLD B: 62.3%, GOLD D: 34%) according to the 2018 GOLD Report were included. After 52 weeks, a significant improvement was detected in lung function (FEV1, FEV1% predicted and FVC; p < 0.001) and symptoms (cough, sputum and shortness of breath; p < 0.001). A clinically relevant improvement in CAT score observed at 12 weeks persisted after 52 weeks in GOLD B and GOLD D patients (p < 0.001), paralleled by a significant reduction of moderate and severe exacerbations by 57.4% and 27.3%, respectively (p < 0.001). After 52 weeks, 93.7% of the patients continued the treatment. Of 21 adverse events reported 16 were non-serious, five were serious, none were deemed drug related. CONCLUSIONS: The present results support the tolerability and effectiveness of extrafine BDP/FF/G in COPD patients in a real-world setting, showing an improvement in lung function, symptom control and a significant reduction in exacerbations.

Dupilumab rapidly improves asthma control in predominantly anti-IL5/IL5R pretreated Austrian real-life severe asthmatics.

Dupilumab is a monoclonal antibody against the IL-4 receptor alpha which has shown efficacy in T2 high severe asthmatics in phase 3 randomized controlled trials. The purpose of this real-life study is to demonstrate the real-life effectiveness of dupilumab in Austrian severe asthma patients. We retrospectively analyzed all patients receiving dupilumab at our severe asthma clinic. Thirteen patients have so far received dupilumab at our center. The primary outcome, asthma control questionnaire 6-item scale at 2 weeks, improved by 0.57 points (p = .014), which is statistically and clinically significant. Similarly, the asthma control test at 4 weeks improved by 3.91 points (p = .024), also statistically and clinically significant. Improvements in forced expiratory volume in 1 s at 2 weeks were neither statistically, nor clinically significant. Improvements at 4 weeks (+220 ml, p = .041), and 3 months (+229 ml, p = .006), were statistically significant and clinically borderline significant. No severe adverse events or hypereosinophilia were observed. No adverse events led to treatment discontinuation. Most patients (85%) had previously received monoclonal antibody treatment for severe asthma. Previous monoclonal antibody treatment had been discontinued in these patients due to a lack of clinical response. Dupilumab is effective and safe in Austrian real-life severe asthmatics. It provides a possible treatment strategy for T2 high severe asthmatics who do not qualify for anti-immunoglobulin E or anti-IL5/IL5R monoclonal antibody treatments or do not adequately respond to these.

COVID-19 in two severe asthmatics receiving benralizumab: busting the eosinophilia myth.

Experience with very mild #COVID19 disease courses in two severe eosinophilic asthmatics with complete eosinophil depletion due to benralizumab treatment counters the recent theories that eosinophilia is protective in COVID-19 infections https://bit.ly/3cnEFvg.

Treatment response according to small airway phenotypes: a real-life observational study.

OBJECTIVE: Scant clinical data are available on the effects of current treatments for asthma on different subgroups of patients with this disease. We conducted a prospective, noninterventional, multicenter real-life study in adult patients with persistent asthma, and we specifically analyzed the effects of treatment with extrafine beclometasone dipropionate/formoterol (BDP/F) in asthma patients categorized by phenotypes related to small airways (i.e. smoking habits, disease duration, and air trapping). METHODS: Patients received BDP/F as a fixed combination (100/6 µg), administered in 1-2 inhalations twice daily over a period of 12 weeks. Peak expiratory flow (PEF), forced expiratory volume in 1 second (FEV1), forced vital capacity (FVC), number of asthma attacks, asthma control, and severity of asthma symptoms were evaluated in the overall population and in different subgroups at three different time points. RESULTS: Overall, 213 patients were enrolled. In the overall population the treatment resulted in a significant increase in the proportion of well controlled patients (from 6.1% to 66.3%; p<0.001), and a reduction of uncontrolled subjects (70.3% versus 10.0%; p<0.001). BDP/F was also associated with a reduction in asthma attacks and an improvement of symptoms. These results were confirmed in specific subgroups of patients identified as small airway phenotypes: smokers, elderly patients, those with long duration of disease and air trapping. CONCLUSIONS: This real-life observational study indicates that extrafine BDP/F in a fixed combination improves asthma control and symptoms in the overall population as well as specific subgroups of patients.