Systematic Approach Revealed SERPING1 Splicing-Affecting Variants to be Highly Represented in the Czech National HAE Cohort.

Hereditary angioedema due to C1 inhibitor deficiency (C1-INH-HAE) is a rare and life-threatening condition characterized by recurrent localized edema. We conducted a systematic screening of SERPING1 defects in a cohort of 207 Czech patients from 85 families with C1-INH-HAE. Our workflow involved a combined strategy of sequencing extended to UTR and deep intronic regions, advanced in silico prediction tools, and mRNA-based functional assays. This approach allowed us to detect a causal variant in all families except one and to identify a total of 56 different variants, including 5 novel variants that are likely to be causal. We further investigated the functional impact of two splicing variants, namely c.550 + 3A > C and c.686-7C > G using minigene assays and RT-PCR mRNA analysis. Notably, our cohort showed a considerably higher proportion of detected splicing variants compared to other central European populations and the LOVD database. Moreover, our findings revealed a significant association between HAE type 1 missense variants and a delayed HAE onset when compared to null variants. We also observed a significant correlation between the presence of the SERPING1 variant c.-21 T > C in the trans position to causal variants and the frequency of attacks per year, disease onset, as well as Clinical severity score. Overall, our study provides new insights into the genetic landscape of C1-INH-HAE in the Czech population, including the identification of novel variants and a better understanding of genotype-phenotype correlations. Our findings also highlight the importance of comprehensive screening strategies and functional analyses in improving the C1-INH-HAE diagnosis and management.

Myeloid lineage cells evince distinct steady-state level of certain gene groups in dependence on hereditary angioedema severity.

Hereditary angioedema (HAE) is a rare genetic disorder with variable expressivity even in carriers of the same underlying genetic defect, suggesting other genetic and epigenetic factors participate in modifying HAE severity. Recent knowledge indicates the role of immune cells in several aspects of HAE pathogenesis, which makes monocytes and macrophages candidates to mediate these effects. Here we combined a search for HAE phenotype modifying gene variants with the characterization of selected genes' mRNA levels in monocyte and macrophages in a symptom-free period. While no such gene variant was found to be associated with a more severe or milder disease, patients revealed a higher number of dysregulated genes and their expression profile was significantly altered, which was typically manifested by changes in individual gene expression or by strengthened or weakened relations in mutually co-expressed gene groups, depending on HAE severity. SERPING1 showed decreased expression in HAE-C1INH patients, but this effect was significant only in patients carrying mutations supposedly activating nonsense-mediated decay. Pro-inflammatory CXC chemokine superfamily members CXCL8, 10 and 11 were downregulated, while other genes such as FCGR1A, or long non-coding RNA NEAT1 were upregulated in patients. Co-expression within some gene groups (such as an NF-kappaB function related group) was strengthened in patients with a severe and/or mild course compared to controls. All these findings show that transcript levels in myeloid cells achieve different activation or depression levels in HAE-C1INH patients than in healthy controls and/or based on disease severity and could participate in determining the HAE phenotype.

ß-blockers and ACE inhibitors are not a risk factor for severe systemic sting reactions and adverse events during venom immunotherapy.

BACKGROUND: There is controversy whether taking ß-blockers or ACE inhibitors (ACEI) is a risk factor for more severe systemic insect sting reactions (SSR) and whether it increases the number or severity of adverse events (AE) during venom immunotherapy (VIT). METHODS: In this open, prospective, observational, multicenter trial, we recruited patients with a history of a SSR and indication for VIT. The primary objective of this study was to evaluate whether patients taking ß-blockers or ACEI show more systemic AE during VIT compared to patients without such treatment. RESULTS: In total, 1,425 patients were enrolled and VIT was performed in 1,342 patients. Of all patients included, 388 (27.2%) took antihypertensive (AHT) drugs (10.4% took ß-blockers, 11.9% ACEI, 5.0% ß-blockers and ACEI). Only 5.6% of patients under AHT treatment experienced systemic AE during VIT as compared with 7.4% of patients without these drugs (OR: 0.74, 95% CI: 0.43-1.22, p = 0.25). The severity of the initial sting reaction was not affected by the intake of ß-blockers or ACEI (OR: 1.14, 95% CI: 0.89-1.46, p = 0.29). In total, 210 (17.7%) patients were re-stung during VIT and 191 (91.0%) tolerated the sting without systemic symptoms. Of the 19 patients with VIT treatment failure, 4 took ß-blockers, none an ACEI. CONCLUSIONS: This trial provides robust evidence that taking ß-blockers or ACEI does neither increase the frequency of systemic AE during VIT nor aggravate SSR. Moreover, results suggest that these drugs do not impair effectiveness of VIT. (Funded by Medical University of Graz, Austria; Clinicaltrials.gov number, NCT04269629).

Acquired Angioedema with C1 Inhibitor Deficiency: Occurrence, Clinical Features, and Management: A Nationwide Retrospective Study in the Czech Republic Patients.

INTRODUCTION: Acquired angioedema with C1 inhibitor deficiency (AAE-C1-INH) is rare but a potentially life-threatening disease. There are no official prevalence data, nor approved therapies for this condition. OBJECTIVE: In this study, we aimed to collect and analyze clinical data on patients with AAE-C1-INH in the Czech Republic. METHODS: We have conducted a retrospective analysis of AAE-C1-INH patients from Czech referral centers for the treatment of hereditary angioedema with C1 inhibitor deficiency. The inclusion criteria involved recurrent episodes of angioedema with the first manifestation at or after the age of 40, negative family history of angioedema, and C1 inhibitor function 50% or less. RESULTS: A total of 14 patients (7 males and 7 females) met the inclusion criteria for AAE-C1-INH. The median age of the symptom onset was 59.5 years, and the median diagnosis delay was 1 year. The most common clinical manifestation was facial edema (100%) and upper airway swelling (85.7%). All patients responded to the acute attack treatment with icatibant and plasma-derived or recombinant C1 inhibitor concentrate. Lymphoid malignancy was identified in 9 patients (64%), monoclonal gammopathy of uncertain significance in 3 (21%), and in 1 patient autoimmune disease (ulcerative colitis) was considered causative (7%). We were not able to identify any underlying disease only in 1 patient (7%). In 6 of 7 patients (86%) treated for lymphoma, either a reduction in the frequency of angioedema attacks or both angioedema symptoms' disappearance and complement parameter normalization was observed. CONCLUSIONS: The prevalence of AAE-C1-INH in the Czech Republic is about 1:760,000. This rare condition occurs in approximately 8% of the patients with angioedema with C1 inhibitor deficiency. AAE-C1-INH is strongly associated with lymphoproliferative disorders, and treating these conditions may improve the control of angioedema symptoms.

Anaphylaxis - acute and long-term management.

Anaphylaxis is a serious, potentially lifethreatening condition, and all healthcare professionals should be aware of it. Prompt recognition of anaphylaxis signs and early initiation of adequate treatment are essential for successful acute management. The firstline treatment is the administration of intramuscular adrenalin, followed by other interventions. Patients should be moni tored after recovery for possible biphasic reaction. Before discharge, the individual risk of further reaction should be assessed and where appropriate an adrenalin autoinjector should be prescribed. Allergy specialist followup is essential for the identification of possible triggers and cofactors. Elimination of these factors reduces the risk of future reactions. Useful preventive measure is allergen immunotherapy, which is definitely indicated in patients with anaphylaxis induced by an insect sting.

Analysis of Sensitization Profiles in Central European Allergy Patients Focused on Animal Allergen Molecules.

INTRODUCTION: Frequently observed multiple sensitizations to several animals highlights the importance of a molecular diagnosis, distinguishing between sensitizations specific to single species and sensitizations due to cross-reactivity. OBJECTIVE: The aim of our study was to assess the usefulness of a molecular diagnosis in the description of sensitization profiles in allergy patients living in Central Europe, with a particular focus on animal-derived molecules. METHODS: The molecular diagnosis was performed using the ImmunoCAP ISAC microarray. Results of 1,255 allergy patients were subjected to statistical analysis. RESULTS: The highest sensitization rates were observed for uteroglobin Fel d 1 (31.8%) and kallikrein Can f 5 (16.4%), followed by animal lipocalins Can f 1 (13.9%), Equ c 1 (6.2%), Fel d 4 (5.3%), Can f 2 (4.2%), and Mus m 1 (4.1%). Sensitization rates to serum albumins Fel d 2, Can f 3, Equ c 3, and Bos d 6 were very low, with the highest being 3.2% to Fel d 2. Detailed subanalysis confirmed the dominant role of Fel d 1 or Can f 5 and/or Can f 1 in cat- or dog-sensitized patients, respectively. Further analysis focused on lipocalins and albumins confirmed a high rate of cosensitizations within both groups. CONCLUSION: The sensitization to animal allergen molecules is very frequent in Central Europe. The most common is sensitization to species-specific cat uteroglobin Fel d 1 and dog kallikrein Can f 5, followed by sensitizations to animal lipocalins. Our data suggest that commonly observed multiple sensitizations detected by extract approach can be explained not only by true cosensitization, but also by cross-reactivity, mainly in the frame of lipocalins. Cross-reactive serum albumins are minor sensitizers and are probably not important from this point of view.

Cross-sectional study on sensitization to mite and cockroach allergen components in allergy patients in the Central European region.

BACKGROUND: The major sources of allergens in the indoor air include house dust mites, dander derived from domestic animals and rodents, cockroach, and several fungi. Mites are the main cause of allergies in some countries with a warmer climate, but the epidemiological significance of mite and cockroach allergens in Central Europe has not been established yet. METHODS: We assessed sensitization profiles of allergy patients in a Central European region in regard to sensitization to mites and cockroach. We used molecular diagnosis by means of the microarray ISAC, and we investigated 1766 patients with clinical suspicion to an allergic disorder. 1255 of them were positive to at least one allergen component, and this group was subjected to statistical analysis. RESULTS: The sensitization to at least one mite-specific molecule (Der p 1, 2, Der f 1, 2) was observed relatively frequently in 32.7% of patients. Specific IgE to mite group 2 molecules is almost fully cross-reactive. Group 1 allergens are also cross-reactive, but in some patients, a species-specific response was observed. Relatively high rate of sensitization both to group 1 and 2 allergens in our patients indicates the greater role of co-sensitizations. Isolated sensitizations to molecules derived from glyciphagid mites Lep d 2 and/or Blo t 5 without sensitization to other mite-derived molecules were observed only exceptionally (in 0.6% of cases). True sensitization to at least one cockroach-specific molecule (Bla g 1, 2, 5) was very rare (in 0.6% of cases), and nearly all of them were co-sensitizations with other noncockroach-derived molecules. Sensitization to an inhaled tropomyosin was observed rarely in 2.2% of patients (Der p 10 in 1.9% and Bla g 7 in 1.5%). Co-sensitization of inhaled tropomyosins with the respective mite- or cockroach-specific molecules was observed only in the minority of patients suggesting the different route of sensitization being more frequent. CONCLUSIONS: The majority of patients are co-sensitized to several molecules of the respective allergen source. The knowledge of this molecular spectrum of sensitization is important for optimal diagnosis and treatment in respect to allergen content in mite extracts used for diagnostic and therapeutic purposes. In regard to the sensitization patterns of Central European patients, it is necessary to point out the importance of quantifying at least three major mite components Der f 1, Der p 1 and Der f 2 (or Der p 2).

Utility of laboratory testing for the diagnosis of Hymenoptera venom allergy.

BACKGROUND: A diagnosis of Hymenoptera venom allergy is based on clinical history and the results of skin tests and/or laboratory methods. OBJECTIVE: To analyze the utility of available laboratory tests in diagnosing Hymenoptera venom allergy. METHODS: Ninety-five patients with Hymenoptera venom allergy with a history of bee (35) or wasp (60) anaphylactic sting reaction and positive skin test with bee or wasp venom were included in this analysis. Specific immunoglobulin E (to bee venom extract, wasp venom extract, available recombinant molecules, and a basophil activation test with venom extracts were assessed in all the patients. Test sensitivity and specificity were calculated by using standard threshold values; then, receiver operating characteristic curve analysis was performed to compute optimal threshold values. Also, statistical analysis of the utility of different combinations of laboratory tests was performed. RESULTS: The optimal threshold values were revealed to be the following: 1.0 kIU/L for bee venom extract (sensitivity, 97.14%; specificity, 100%), 0.35 kIU/L for rApi m 1 (sensitivity, 68.57%; specificity, 100%), 1.22 kIU/L for wasp venom extract (sensitivity, 88.33%; specificity, 95.45%), 0.7 kIU/L for rVes v 5 (sensitivity, 86.67%; specificity, 95.45%), 1.0 kIU/L for rVes v 1 (sensitivity, 56.67%; specificity, 95.45%), 6.5% for basophil activation test with bee venom extract (sensitivity, 80%; specificity, 95.45%), and 4.5% for basophil activation test with wasp venom extract (sensitivity, 91.53%; specificity, 95.45%). The best test combinations were found to be the following: bee venom extract plus rApi m 1 (sensitivity, 97.14%; specificity, 95.45%) in bee and either wasp venom extract plus rVes v 5, or rVes v 5 plus rVes v 1 (both sensitivity, 98.33%; specificity, 95.45%) in patients with wasp venom allergy. CONCLUSION: Our analysis confirmed that currently used laboratory tests represent effective tools in diagnosing Hymenoptera venom allergy. Moreover, our probabilistic approach offered another way to interpret concrete values of laboratory test results and opened possible direction on how to optimize the laboratory diagnostic procedure.

A comprehensive analysis of middle-European molecular sensitization profiles to pollen allergens.

Molecular diagnosis of allergy and microarray technology have opened a completely new avenue of insight into sensitization profiles from both the clinical and the epidemiological point of view. We used this innovative tool in the description of sensitization patterns in pollen-sensitized patients in Middle Europe. Immunoglobulin E detection using 112 different allergenic molecules was carried out employing the ImmunoCAP ISAC microarray system. Sera from 826 patients sensitized to at least one pollen-derived molecule were subjected to analysis. The highest observed sensitization rate was 81.0% to grass-specific molecules (the most frequent being Phl p 1; 69.6%). The second most frequent sensitization was 54.8% to Betulaceae-specific molecules (Bet v 1; 54.2%). Together, grasses and Betulaceae components (and their cosensitizations with other components) comprised the vast majority of pollen sensitizations. Unexpectedly frequently observed sensitizations were those to Cupressaceae-specific molecules (14.1%), Oleaceae-specific molecules (10.8%), and the plane tree-derived molecule Pla a 2 (15.5%). The sensitization rates for all other molecules were within the expected range (Art v 1, 13.6%; Pla l 1, 9.6%; Che a 1, 8.4%; Par j 2, 0.9%; Amb a 1, 0.8%, and Sal k 1, 0.5%). Cross-reacting molecule sensitization rates were found to be 12.4% for profilins, 5.0% for polcalcins, and 6.4% for lipid transfer proteins. Molecular diagnosis of allergy gives a more precise and comprehensive insight into pollen sensitization patterns than extract-based testing, allowing a better understanding of the sensitization process and regional differences. The data presented here may help to improve the diagnostic and allergen-specific treatment procedures in the respective region.