Real-world, long-term effectiveness of allergy immunotherapy in allergic rhinitis: Subgroup analyses of the REACT study.

BACKGROUND: Randomized controlled trials have demonstrated the efficacy of allergy immunotherapy (AIT) in allergic rhinitis (AR) and the disease-modifying effects of the SQ grass sublingual immunotherapy (SLIT) tablet. OBJECTIVE: We sought to assess real-world, long-term effectiveness and safety across AIT subgroups: route of administration, therapeutic allergen, persistence to AIT, and SQ grass SLIT tablet. METHODS: The primary outcome of AR prescriptions from a retrospective cohort study (REAl-world effeCtiveness in allergy immunoTherapy; 2007-2017) was assessed across prespecified AIT subgroups in subjects with AR with and without AIT prescriptions (controls). Safety was assessed as anaphylaxis for 2 days or less of the first AIT prescription. Subgroup follow-up continued until samples were fewer than 200 subjects. RESULTS: Subcutaneous immunotherapy (SCIT) and SLIT tablets showed similarly greater reductions in AR prescriptions than controls (SCIT vs SLIT tablets: year 3, P = .15; year 5, P = .43). Comparably greater reductions in AR prescriptions were observed for grass- and house dust mite-specific AIT than for controls, but significantly smaller reductions were observed for tree-specific AIT (tree vs house dust mite, and vs grass: years 3 and 5, P < .0001). Persistence to AIT was associated with greater reductions in AR prescriptions versus nonpersistence (persistence vs nonpersistence: year 3, P = .09; year 5, P = .006). SQ grass SLIT tablet showed sustained reductions versus controls for up to 7 years (year 3, P = .002; year 5, P = .03). Rates of anaphylactic shock were low (0.000%-0.092%), with no events for SQ SLIT tablets. CONCLUSIONS: These results demonstrate real-world, long-term effectiveness of AIT, complement disease-modifying effects observed in SQ grass SLIT-tablet randomized controlled trials, and highlight the importance of using newer evidence-based AIT products for tree pollen AR.

Impaired mucus clearance exacerbates allergen-induced type 2 airway inflammation in juvenile mice.

BACKGROUND: Type 2 airway inflammation plays a central role in the pathogenesis of allergen-induced asthma, but the underlying mechanisms remain poorly understood. Recently, we demonstrated that reduced mucociliary clearance, a characteristic feature of asthma, produces spontaneous type 2 airway inflammation in juvenile ß-epithelial Na+ channel (Scnn1b)-transgenic (Tg) mice. OBJECTIVE: We sought to determine the role of impaired mucus clearance in the pathogenesis of allergen-induced type 2 airway inflammation and identify cellular sources of the signature cytokine IL-13. METHODS: We challenged juvenile Scnn1b-Tg and wild-type mice with Aspergillus fumigatus and house dust mite allergen and compared the effects on airway eosinophilia, type 2 cytokine levels, goblet cell metaplasia, and airway hyperresponsiveness. Furthermore, we determined cellular sources of IL-13 and effects of genetic deletion of the key type 2 signal-transducing molecule signal transducer and activator of transcription 6 (STAT6) and evaluated the effects of therapeutic improvement of mucus clearance. RESULTS: Reduced mucociliary allergen clearance exacerbated Stat6-dependent secretion of type 2 cytokines, airway eosinophilia, and airway hyperresponsiveness in juvenile Scnn1b-Tg mice. IL-13 levels were increased in airway epithelial cells, macrophages, type 2 innate lymphoid cells, and TH2 cells along with increased Il33 expression in the airway epithelium of Scnn1b-Tg mice. Treatment with the epithelial Na+ channel blocker amiloride, improving airway surface hydration and mucus clearance, reduced allergen-induced inflammation in Scnn1b-Tg mice. CONCLUSION: Our data support that impaired clearance of inhaled allergens triggering IL-13 production by multiple cell types in the airways plays an important role in the pathogenesis of type 2 airway inflammation and suggests therapeutic improvement of mucociliary clearance as a novel treatment strategy for children with allergen-induced asthma.

Regulatory T-cell impairment in cystic fibrosis patients with chronic pseudomonas infection.

RATIONALE: Patients with cystic fibrosis (CF) lung disease have chronic airway inflammation driven by disrupted balance of T-cell (Th17 and Th2) responses. Regulatory T cells (Tregs) dampen T-cell activation, but their role in CF is incompletely understood. OBJECTIVES: To characterize numbers, function, and clinical impact of Tregs in CF lung disease. METHODS: Tregs were quantified in peripheral blood and airway samples from patients with CF and from lung disease control patients without CF and healthy control subjects. The role of Pseudomonas aeruginosa and CF transmembrane conductance regulator (CFTR) in Treg regulation was analyzed by using in vitro and murine in vivo models. MEASUREMENTS AND MAIN RESULTS: Tregs were decreased in peripheral blood and airways of patients with CF compared with healthy controls or lung disease patients without CF and correlated positively with lung function parameters. Patients with CF with chronic P. aeruginosa infection had lower Tregs compared with patients with CF without P. aeruginosa infection. Genetic knockout, pharmacological inhibition, and P. aeruginosa infection studies showed that both P. aeruginosa and CFTR contributed to Treg dysregulation in CF. Functionally, Tregs from patients with CF or from Cftr(-/-) mice were impaired in suppressing conventional T cells, an effect that was enhanced by P. aeruginosa infection. The loss of Tregs in CF affected memory, but not naive Tregs, and manifested gradually with disease progression. CONCLUSIONS: Patients with CF who have chronic P. aeruginosa infection show an age-dependent, quantitative, and qualitative impairment of Tregs. Modulation of Tregs represents a novel strategy to rebalance T-cell responses, dampen inflammation, and ultimately improve outcomes for patients with infective CF lung disease.

Hypoxic epithelial necrosis triggers neutrophilic inflammation via IL-1 receptor signaling in cystic fibrosis lung disease.

RATIONALE: In many organs, hypoxic cell death triggers sterile neutrophilic inflammation via IL-1R signaling. Although hypoxia is common in airways from patients with cystic fibrosis (CF), its role in neutrophilic inflammation remains unknown. We recently demonstrated that hypoxic epithelial necrosis caused by airway mucus obstruction precedes neutrophilic inflammation in Scnn1b-transgenic (Scnn1b-Tg) mice with CF-like lung disease. OBJECTIVES: To determine the role of epithelial necrosis and IL-1R signaling in the development of neutrophilic airway inflammation, mucus obstruction, and structural lung damage in CF lung disease. METHODS: We used genetic deletion and pharmacologic inhibition of IL-1R in Scnn1b-Tg mice and determined effects on airway epithelial necrosis; levels of IL-1α, keratinocyte chemoattractant, and neutrophils in bronchoalveolar lavage; and mortality, mucus obstruction, and structural lung damage. Furthermore, we analyzed lung tissues from 21 patients with CF and chronic obstructive pulmonary disease and 19 control subjects for the presence of epithelial necrosis. MEASUREMENTS AND MAIN RESULTS: Lack of IL-1R had no effect on epithelial necrosis and elevated IL-1α, but abrogated airway neutrophilia and reduced mortality, mucus obstruction, and emphysema in Scnn1b-Tg mice. Treatment of adult Scnn1b-Tg mice with the IL-1R antagonist anakinra had protective effects on neutrophilic inflammation and emphysema. Numbers of necrotic airway epithelial cells were elevated and correlated with mucus obstruction in patients with CF and chronic obstructive pulmonary disease. CONCLUSIONS: Our results support an important role of hypoxic epithelial necrosis in the pathogenesis of neutrophilic inflammation independent of bacterial infection and suggest IL-1R as a novel target for antiinflammatory therapy in CF and potentially other mucoobstructive airway diseases.

Improved survival in osteosarcoma patients with atypical low vascularization.

BACKGROUND: Osteosarcoma is considered a highly vascularized bone tumor with early metastatic dissemination through intratumoral blood vessels mostly into the lung. Novel targets for therapy such as tumor vascularization are highly warranted since little progress has been achieved in the last 30 years. However, proof of relevance for vascularization as a major prognostic parameter has been hampered by tumor heterogeneity, difficulty in detecting microvessels by immunohistochemistry, and small study cohorts. Most recently, we demonstrated that highly standardized whole-slide imaging could overcome these limitations (Kunz et al., PloS One 9(3):e90727, 2014). In this study, we applied this method to a multicenter cohort of 131 osteosarcoma patients to test osteosarcoma vascularization as a prognostic determinant. METHODS: Computer-assisted whole-slide analysis, together with enzymatic epitope retrieval, was used for CD31-based microvessel quantification in 131 pretreatment formalin-fixed and paraffin-embedded biopsies from three bone tumor centers. Kaplan-Meier-estimated survival and chemoresponse were determined and multivariate analysis was performed. Conventional hot-spot-based microvessel density (MVD) determination was compared with whole-slide imaging. RESULTS: We detected high estimated overall (p ≤ 0.008) and relapse-free (p ≤ 0.004) survival in 25 % of osteosarcoma patients with low osteosarcoma vascularization in contrast to other patient groups. Furthermore, all patients with low osteosarcoma vascularization showed a good response to neoadjuvant chemotherapy. Comparison of conventional MVD determination with whole-slide imaging suggests false high quantification or even exclusion of samples with low osteosarcoma vascularization due to difficult CD31 detection in previous studies. CONCLUSION: Low intratumoral vascularization at the time of diagnosis is a strong predictor for prolonged survival and good response to neoadjuvant chemotherapy in osteosarcoma.

Long-term health care resource and cost savings with allergy immunotherapy: REACT study results.

Background: Allergy immunotherapy (AIT) can be administered as subcutaneous immunotherapy (SCIT) injections in the clinic or as sublingual immunotherapy (SLIT) tablets at home after initiation under medical supervision. To achieve long-term, sustained effects, a 3-year treatment duration is recommended. Objective: Our aim was to assess the association of AIT (SCIT and SLIT tablets) with long-term health care resource use (HRU) and costs in subjects with allergic rhinitis. Methods: REACT was a retrospective propensity score-matched cohort study using claims data from a German health insurance database (2007-2017), with up to 9 years of follow-up after AIT initiation. HRU and costs were evaluated for hospitalizations, ambulatory care visits, and prescriptions, in subjects who received AIT versus in matched controls with allergic rhinitis who had not received AIT, as well as for SCIT and SLIT tablets. Results: Across all 9 years, the subjects who received AIT had a significantly lower incidence of hospitalization than the controls did. Generally, proportions of subjects with ambulatory care visits and hospitalizations were lower, and length of hospitalization was shorter, for those receiving SLIT tablets than those who received SCIT. Total costs were significantly higher with AIT versus for the controls during the treatment period (years 1 to 3), driven by prescriptions and ambulatory care visits, but they were lower in years 4 to 9. During years 1 to 3, prescription costs were generally higher for SLIT tablets than for SCIT, whereas ambulatory care costs were numerically lower. In most years, hospitalization costs were numerically lower for SLIT tablets than for SCIT. Conclusion: Initial higher HRU and costs of AIT during the expected treatment period are offset in the long term. At-home administration of SLIT tablets may further reduce ambulatory care costs.

Long-term real-world effectiveness of allergy immunotherapy in patients with allergic rhinitis and asthma: Results from the REACT study, a retrospective cohort study.

BACKGROUND: Allergen immunotherapy (AIT) is the only causal treatment for respiratory allergy. Long-term real-life effectiveness of AIT remains to be demonstrated beyond the evidence from randomised controlled trials (RCTs). METHODS: REACT (Real world effectiveness in allergy immunotherapy) is a retrospective cohort study using claims data between 2007 and 2017. Study eligibility was a confirmed diagnosis of allergic rhinitis (AR), with or without asthma, and AIT. To ensure comparable groups, AIT-treated subjects were propensity score matched 1:1 with control subjects, using characteristic and potential confounding variables. Outcomes were analysed as within (pre vs post AIT) and between (AIT vs control) group differences across 9 years of follow-up (ClinicalTrial.gov: NCT04125888). FINDINGS: 46,024 AIT-treated subjects were matched with control subjects and 14,614 were included in the pre-existing asthma cohort. AIT-treated subjects were 29·5 (16·3) years and 53% were male. Compared to pre-index year, AIT was consistently associated with greater reductions compared to control subjects in AR and asthma prescriptions, including both asthma controller and reliever prescriptions. Additionally, the AIT group had significantly greater likelihood of stepping down asthma treatment (P <0·0001). In addition to the reduction in asthma treatment in the AIT group, a greater reduction in severe asthma exacerbations was demonstrated (P<0·05). Reductions in pneumonia with antibiotic prescriptions, hospitalisations, and duration of inpatients stays were all in favour of AIT. INTERPRETATION: The study extends the existing RCT evidence for AIT by demonstrating longer-term and sustained effectiveness of AIT in the real world. Additionally, in patients with concurrent asthma, AIT was associated with reduced likelihood of asthma exacerbations and pneumonia. FUNDING: The study was funded by ALK A/S.

Stressors faced by healthcare professionals and coping strategies during the early stage of the COVID-19 pandemic in Germany.

BACKGROUND: The COVID-19 pandemic has exerted great pressure on national health systems, which have aimed to ensure comprehensive healthcare at all times. Healthcare professionals working with COVID-19 patients are on the frontline and thereby confronted with enormous demands. Although early reports exist on the psychological impact of the pandemic on frontline medical staff working in Asia, little is known about its impact on healthcare professionals in other countries and across various work sectors. The present cross-sectional, online survey sought to investigate common work stressors among healthcare professionals, their psychological stress as well as coping resources during the pandemic. METHODS: A sample of 575 healthcare professionals (57% male) in three different sectors (hospital, prehospital emergency care, and outpatient service) reported their experiences concerning work and private stressors, psychological stress, and coping strategies between April 17, 2020 and June 5, 2020. To capture pandemic-specific answers, most of the items were adapted or newly developed. Exploratory factor analyses (EFA) were conducted to detect underlying latent factors relating to COVID-specific work stressors. In a next step, the effects of these latent stressors across various work sectors on psychological stress (perceived stress, fatigue, and mood) were examined by means of structural equation models (SEM). To add lived experience to the findings, responses to open-ended questions about healthcare professionals' stressors, effective crisis measures and prevention, and individual coping strategies were coded inductively, and emergent themes were identified. RESULTS: The EFA revealed that the examined work stressors can be grouped into four latent factors: "fear of transmission", "interference of workload with private life", "uncertainty/lack of knowledge", and "concerns about the team". The SEM results showed that "interference of workload with private life" represented the pivotal predictor of psychological stress. "Concerns about the team" had stress-reducing effects. The latent stressors had an equal effect on psychological stress across work sectors. On average, psychological stress levels were moderate, yet differed significantly between sectors (all p < .001); the outpatient group experienced reduced calmness and more stress than the other two sectors, while the prehospital group reported lower fatigue than the other two sectors. The prehospital group reported significantly higher concerns about the team than the hospital group (p < .001). In their reports, healthcare professionals highlighted regulations such as social distancing and the use of compulsory masks, training, experience and knowledge exchange, and social support as effective coping strategies during the pandemic. The hospital group mainly mentioned organizational measures such as visiting bans as effective crisis measures, whereas the prehospital sector most frequently named governmental measures such as contact restrictions. CONCLUSION: The study demonstrated the need for sector-specific crisis measures to effectively address the specific work stressors faced by the outpatient sector in particular. The results on pandemic-specific work stressors reveal that healthcare professionals might benefit from coping strategies that facilitate the utilization of social support. At the workplace, team commitment and knowledge exchange might buffer against adverse psychological stress responses. Schedules during pandemics should give healthcare workers the opportunity to interact with families and friends in ways that facilitate social support outside work. Future studies should investigate cross-sector stressors using a longitudinal design to identify both sector- and time-specific measures. Ultimately, an international comparison of stressors and measures in different sectors of healthcare systems is desirable.

Regulatory T cells and tolerance induction.

Transplantation tolerance is the ultimate goal of organ transplantation. Natural regulatory T cells (Treg) have been expected to reach clinical applications for tolerance induction because their initial description. More than ten yr later, Treg have started moving from experimental animal models into clinical applications. Although the molecular mechanism of contact-dependent inhibition remains to be unraveled, alterations of Treg numbers have been shown in several human diseases: Whereas several autoimmune diseases have been reported to be associated with decreased Treg numbers, Treg are frequently accumulated in solid tumors and hematologic malignancies. Monitoring of Treg numbers could be instrumental in identifying patients with risk of graft failure and might help minimizing immunosuppressive therapy in transplant recipients. Molecular mechanisms of Treg proliferation and Treg elimination such as CD95 ligand (CD95L)-mediated apoptosis are currently explored for their clinical usability as therapeutical targets. Immunosuppressive drugs might modulate the number of Tregs. Expansion of the Treg numbers in vivo or in vitro resembles a novel therapeutical strategy to reach partial or even operational tolerance after organ transplantation.

Death receptor signaling and its function in the immune system.

Death receptors belong to the TNF (tumor necrosis factor)/NGF (nerve growth factor) receptor superfamily. Signaling via death receptors plays a distinct role, e.g. in the immune system, where it contributes to regulation of the adaptive immune response in various ways, most notably by triggering activation-induced cell death (AICD) of T cells. Thus, dysregulation of death receptor signaling, either allowing too much or too little apoptosis, can lead to autoimmune disorders and also impacts on tumorigenesis or other diseases. In this chapter we address components, molecular mechanisms and regulation of death receptor signaling with particular focus on CD95 (APO-1, Fas). We discuss the role of death receptor-mediated AICD in regulation of the adaptive immune response against foreign and self antigens in comparison to cytokine deprivation-mediated death by neglect. Finally, the contribution of dysregulated death receptor/ligand systems to autoimmune diseases such as diabetes, multiple sclerosis and Hashimoto's thyroiditis is discussed.

Personalized Medicine in Allergic Asthma: At the Crossroads of Allergen Immunotherapy and "Biologicals".

Major allergic disease can be viewed as clinical syndromes rather than discrete disease entities. Emerging evidence indicates that allergic asthma includes several disease phenotypes. Immunological deviation toward high T helper cell type 2 cytokine levels has been demonstrated for a subgroup of pediatric asthma patients, and now, several novel monoclonal antibodies have been approved for treatment of this subgroup as a stratified approach of "personalized" medicine in allergy. Introduction of component-based IgE testing before allergen immunotherapy (AIT), i.e., testing for IgE cross-reactivity before initiation of AIT, has also brought stratified medicine into allergy therapy. Improved responder criteria, which identify treatment-responders previous to therapy, might foster this stratification and even individualized AIT might have an impact for tailor-made therapy in the future. Furthermore, combining antibody-based treatment with AIT could help to establish more rapid AIT protocols even for allergens with a high risk of anaphylactic reactions. Efforts to advance such "personalized" medicine in pediatric allergy might be challenged by several issues including high costs for the health-care system, increasing complexity of allergy therapy, the need for physician allergy expertise, and furthermore ethical considerations and data safety issues.

Similar sensitivity of regulatory T cells towards CD95L-mediated apoptosis in patients with multiple sclerosis and healthy individuals.

Impaired suppressive function of CD4(+)CD25(high) regulatory T cells (T(reg)) has been reported as a novel pathogenetic mechanism in Multiple sclerosis (MS). We addressed if high apoptosis sensitivity of MS-T(reg) could explain this functional T(reg) defect. T(reg) from treatment-naïve MS patients showed high sensitivity towards CD95Ligand-mediated apoptosis and exhibited enhanced cell death to IL-2 and TCR-signal deprivation. Since susceptibility of T(reg) to cell death was similar in MS patients and healthy controls, this cannot explain the inhibitory dysfunction of T(reg) associated with MS. Furthermore, as cell death is not enhanced, therapeutic expansion of MS-T(reg)in vitro should be a reasonable and novel therapeutic option.

CD8+/FOXP3+-ratio in osteosarcoma microenvironment separates survivors from non-survivors: a multicenter validated retrospective study.

Osteosarcoma is the most common primary bone tumor characterized by juvenile onset, tumor heterogeneity, and early pulmonary metastasis. Therapeutic improvement stagnates since more than two decades. Unlike major malignancies, biomarkers as prognostic factors at time of diagnosis are missing. Disease rareness hampers study recruitment of patient numbers sufficient to outweigh tumor heterogeneity. Here, we analyzed in a multicenter cohort the osteosarcoma microenvironment to reduce effects of tumor cell heterogeneity. We hypothesized that quantitative ratios of intratumoral CD8+T-cells to FOXP3+T-cells (CD8+/FOXP3+-ratios) provide strong prognostic information when analyzed by whole-slide imaging in diagnostic biopsies. We followed recommendations-for-tumor-marker-prognostic-studies (REMARK). From 150 included cases, patients with complete treatment were identified and assigned to the discovery (diagnosis before 2004) or the validation cohort (diagnosis 2004-2012). Highly standardized immunohistochemistry of CD8+ and FOXP3+, which was validated by methylation-specific gene analysis, was performed followed by whole-slide analysis and clinical outcome correlations. We observed improved estimated survival in patients with CD8+/FOXP3+-ratios above the median (3.08) compared to patients with lower CD8+/FOXP3+-ratios (p = 0.000001). No patients with a CD8+/FOXP3+-ratio above the third quartile died within the observation period (median follow-up 69 mo). Multivariate analysis demonstrated independence from current prognostic factors including metastasis and response to neoadjuvant chemotherapy. Data from an independent validation cohort confirmed improved survival (p = 0.001) in patients with CD8+/FOXP3+-ratios above 3.08. Multivariate analysis proofed that this observation was also independent from prognostic factors at diagnosis within the validation cohort. Intratumoral CD8+/FOXP3+-ratio in pretreatment biopsies separates patients with prolonged survival from non-survivors in osteosarcoma.

Osteosarcoma microenvironment: whole-slide imaging and optimized antigen detection overcome major limitations in immunohistochemical quantification.

BACKGROUND: In osteosarcoma survival rates could not be improved over the last 30 years. Novel biomarkers are warranted to allow risk stratification of patients for more individual treatment following initial diagnosis. Although previous studies of the tumor microenvironment have identified promising candidates, novel biomarkers have not been translated into routine histopathology. Substantial difficulties regarding immunohistochemical detection and quantification of antigens in decalcified and heterogeneous osteosarcoma might largely explain this translational short-coming. Furthermore, we hypothesized that conventional hot spot analysis is often not representative for the whole section when applied to heterogeneous tissues like osteosarcoma. We aimed to overcome these difficulties for major biomarkers of the immunovascular microenvironment. METHODS: Immunohistochemistry was systematically optimized for cell surface (CD31, CD8) and intracellular antigens (FOXP3) including evaluation of 200 different antigen retrieval conditions. Distribution patterns of these antigens were analyzed in formalin-fixed and paraffin-embedded samples from 120 high-grade central osteosarcoma biopsies and computer-assisted whole-slide analysis was compared with conventional quantification methods including hot spot analysis. RESULTS: More than 96% of osteosarcoma samples were positive for all antigens after optimization of immunohistochemistry. In contrast, standard immunohistochemistry retrieved false negative results in 35-65% of decalcified osteosarcoma specimens. Standard hot spot analysis was applicable for homogeneous distributed FOXP3+ and CD8+ cells. However, heterogeneous distribution of vascular CD31 did not allow reliable quantification with hot spot analysis in 85% of all samples. Computer-assisted whole-slide analysis of total CD31- immunoreactive area proved as the most appropriate quantification method. CONCLUSION: Standard staining and quantification procedures are not applicable in decalcified formalin-fixed and paraffin-embedded samples for major parameters of the immunovascular microenvironment in osteosarcoma. Whole-slide imaging and optimized antigen retrieval overcome these limitations.