Introduction: Exposure to respiratory viruses is a significant cause of morbidity and affects virus-specific antibody levels. Little is known about determinants associated with immune response to these viruses. We aimed to investigate the determinants of respiratory syncytial virus (RSV) and rhinovirus (RV) specific IgG responses in both children and adults. Methods: The study is based on the EGEA cohort, composed of 530 samples of children in EGEA1 (1991-95) and 1241 samples of adults in EGEA2 (2003-07). Cumulative RV-specific IgG responses (species A, B and C) and IgG responses to RSV-G protein were measured by micro-array technology. Multiple linear mixed models (random effect to account for familial dependence) were performed to assess associations between age, sex, body mass index (BMI), tobacco smoke exposure and season of blood sampling with RSV and RV-specific IgG levels. Results: In children (11.1±2.8 years old, 57% of boys), higher RV-specific IgG levels were associated with older age (only for RV-B), female sex and lower BMI, while only older age was associated with higher RSV-specific IgG levels. In adults (43.5±16.7 years old, 48% of men), younger age, female sex, lower BMI, active smoking and all seasons except summer were associated with higher RV-specific IgG levels. Older age, active smoking and all seasons except summer were associated with higher RSV-specific IgG levels. Conclusion: Personal and seasonal determinants of RSV- and RV-specific IgG levels seem to vary according to the respiratory virus type and between children and adults, suggesting different patterns of responses along the life course.

Background: Respiratory birch pollen allergy and associated oral allergy syndrome affect more than 150 million people. IgE cross-sensitization to major birch pollen allergen Bet v 1 and pathogenesis-related (PR10) plant food allergens is responsible for the pollen-food allergy syndrome. Methods: We designed a recombinant protein, AB-PreS, consisting of non-allergenic peptides derived from the IgE binding sites of Bet v 1 and the cross-reactive apple allergen, Mal d 1, fused to the PreS domain of HBV surface protein as immunological carrier. AB-PreS was expressed in E. coli and purified by chromatography. The allergenic activity of AB-PreS was tested using sera and basophils from birch pollen patients allergic. The protective effect of AB-PreS was assessed by inhibition ELISA test using sera allergic patients and from immunized rabbits. Results: IgE-binding experiments and basophil activation test revealed the hypoallergenic nature of AB-PreS. IgG antibodies induced by 5 injections with AB-PreS inhibited allergic patients’ IgE binding to Bet v 1 and Mal d 1 better than did IgG induced by up to 30 injections of six licensed birch pollen allergen extract-based vaccines. Additionally, immunization with AB-PreS induced HBV-specific antibodies potentially protecting the infection. Conclusion: The recombinant AB-PreS-based vaccine is hypoallergenic, safe and superior to currently registered allergen extract-based vaccines for the treatment of the birch pollen food allergy syndrome.

Background The vast majority of SARS-CoV-2 vaccines which are licensed or under development focus on the spike (S) protein and its receptor binding domain (RBD). However, S and RBD from SARS-CoV-2 variants of concerns show considerable sequence variations and repeated injections for boosting specific immunity are necessary. Aim of this study was to develop and characterize a SARS-CoV-2 vaccine targeting the highly conserved nucleocapsid (N) protein. Methods Recombinant N protein was expressed in Escherichia coli, purified to homogeneity by chromatography and characterized by SDS-PAGE, immunoblotting, mass spectrometry, dynamic light scattering and differential scanning calorimetry. The N protein vaccine was obtained by formulation of recombinant N as squalane-based emulsion and used to immunize Balb/c mice, NOD scid gamma (NSG) mice engrafted with human PBMC, rabbits and marmoset monkeys to study safety as well as antibody and cellular immunity using ELISA for antibodies, measurement of N-specific Th1 and Th2 cytokine secretion and carboxyfluorescein succinimidyl ester (CFSE) dilution assays for CD4 + and CD8 + T cell responses. The protective effect of the vaccine was studied in SARS-CoV-2-infected Syrian hamsters. Results Immunization of mice, rabbits and Syrian hamsters with the recombinant N protein-based vaccine formulated as squalane-based emulsion (Convacell®) induced sustainable N-specific IgG responses and a N-specific mixed Th1/Th2 cytokine response. In marmoset monkeys a N-specific CD4 + as well as CD8 + T cell response was observed. Vaccinated and then infected Syrian hamsters showed reduced lung histopathology, reduced virus was detected in lung tissue, lung weight relative to the body was not increased after challenge and body weight was regained faster than in non-vaccinated animals. Repeated dose toxicity studies in mice and rabbits showed that Convacell® was well tolerated and safe. Conclusions Convacell® induced a SARS-CoV-2-specific protective immune response in Syrian hamsters. It is a new vaccine targeting the nucleocapsid protein of SARS-CoV-2 and thus may augment the armamentarium of vaccines for COVID-19.Now published:  Vaccines 2023, 11(4), 874; https://doi.org/10.3390/vaccines11040874

BACKGROUND Severe acute respiratory syndrome corona virus (SARS-CoV-2) infection frequently causes severe and prolonged disease but only few specific treatments are available. We aimed to investigate safety and efficacy of a SARS-CoV-2-specific siRNA-peptide dendrimer formulation (MIR 19 ®) targeting a conserved sequence in known SARS-CoV-2 variants for treatment of COVID-19. METHODS We conducted an open-label, randomized controlled multicenter phase II trial (NCT05184127) evaluating safety and efficacy of inhaled MIR 19 ® (3.7mg and 11.1 mg/day: groups 1 and 2, respectively) in comparison with standard etiotropic drug treatment (group 3) in patients hospitalized with moderate COVID-19. The primary endpoint was the time to clinical improvement according to predefined criteria within 14 days of randomization. RESULTS Patients from group1 had a significantly reduced (median 6 days (95% confidence interval [CI]: 5-7, HR 1.75, P=0.0005) time to clinical improvement compared to patients from group 3 (8 days (95% CI: 7-10). Normalized oxygen saturation (SpO 2>94%) occurred quicker in the group 1 (median 5 days (95% CI: 4–5, HR 1.59, P=0.0033) than in the group 3 (6 days, 95% CI: 5–8). Treatment with MIR 19® was well tolerated and safe. CONCLUSIONS MIR 19 ®, a SARS-CoV-2-specific siRNA-peptide dendrimer formulation is safe and significantly reduces time to clinical improvement in hospitalized moderate COVID-19 patients compared to standard therapy in a randomized controlled trial. MIR 19 ® treatment targets a sequence which is identical in all SARS-CoV-2 variants known so far and hence should be applicable for all of them.

Cellular and humoral responses to fourth SARS-CoV-2 vaccination in a real life cohort of patients with cancer To the Editor,Patients with cancer are at increased risk of adverse outcomes when infected with SARS-CoV-2 and show an impeded humoral and cellular immune response to vaccination (1). A fourth vaccination increased the humoral immunity against SARS-CoV-2 including Omicron sublineages better than Tixagevimab and Cilgavimab (2). However, data on effects of a fourth SARS-CoV-2 vaccination on cellular immunity, particularly in relation to antibody responses, are scarce (3).Methods : To analyze specific cellular immunity after fourth immunization, SARS-CoV-2 specific CD4+ / CD8+ T-cell responses were prospectively measured in 7 patients with histologically confirmed neoplastic disease before and at the next clinical visit after fourth vaccination against the SARS-CoV-2 spike protein (S) and the receptor binding domain (RBD). Moreover, IgG against S and RBD of Omicron (BA.4) and Hu-1, respectively were assessed. A >1.1-fold increase of antigen-specific proliferated cells and antibody levels compared to baseline was defined as a vaccine response. Assays were performed as described previously (4). This study was approved by the Ethics Committee of the Medical University of Vienna (vote 1427/2022) and performed according to the Declaration of Helsinki and its amendments. Informed consent was obtained from all included participants. Descriptive statistical analysis was performed using GraphPad Prism, Version 9.4.1 (San Diego, California, USA).Results : Six patients with solid tumors and one immunocompetent patient with CNS lymphoma (median age [range] 64 years [45-78], 7 men) were prospectively included and received a fourth vaccination (one mRNA-1273 and six BNT162b2). Of these patients, 6 patients were undergoing active anti-neoplastic therapy. The baseline blood sampling was performed in median 7 months (range 5-9 months) after the third vaccine dose, while the follow-up blood sampling was done in median 21 days (range: 19-30 days) after the fourth vaccination (Table 1) .Overall, clear signs of response on either humoral, cellular, or combined humoral and cellular levels were observed in 6/7 patients. However, a striking intra- and interpatient heterogeneity of immune response patterns was evident (Figure 1 ). Only 2/7 patients (patients 4 and 6) responded with combined increases in S and RBD specific CD4+ and CD8+ T-cell proliferation. All other patients showed inconsistent increases in T-cell activity with low vaccination responses in at least one T-cell subpopulation. Additionally, humoral response did not consistently coincide with cellular vaccine responses: patients 4 and 6, who had no or only a mild (e.g. IgG against S 0.97-fold change and 2.23-fold change, respectively) increase in antibody levels had a pronounced cellular vaccine response (e.g. CD4 against S 4-fold change and 134-fold change, respectively). Interestingly, patient 5 increased antibody levels against S without corresponding CD4+ responses. Moreover, patients with distinct antibody increases only showed mediocre vaccine responses on cellular level (Patients 1, 2 and 7). One patient (patient 3), showed severely impeded humoral and cellular vaccine responses to the fourth vaccination applied 433 days after administration of the last B-cell targeting treatment (Rituximab).Conclusions : The most important limitation of this prospective study is its small sample size and the lack of a control group. However, we observed high intra- and interpatient heterogeneity with clear indications of humoral, cellular, or combined response to fourth vaccine in most patients under active treatment. Of note, our observation indicates long-lasting impairment of specific immune responses after a fourth vaccine on both humoral and cellular levels as long as 36 months after last rituximab administration. These findings highlight the need for reliable identification of and development of management strategies for SARS-CoV-2 vaccine non-responders among patients receiving anti-cancer therapies.References :1. Mairhofer M, Kausche L, Kaltenbrunner S, Ghanem R, Stegemann M, Klein K et al. Humoral and cellular immune responses in SARS-CoV-2 mRNA-vaccinated patients with cancer. Cancer Cell. 2021;39 :1171–1172.2. Mair MJ, Mitterer M, Gattinger P, Berger JM, Valenta R, Fong D et al. Inhibition of SARS-CoV-2 Omicron BA.1 and BA.4 Variants After Fourth Vaccination or Tixagevimab and Cilgavimab Administration in Patients With Cancer. JAMA Oncol Published Online First: September 2022. doi:10.1001/jamaoncol.2022.42263. Debie Y, Van Audenaerde JRM, Vandamme T, Croes L, Teuwen L-A, Verbruggen L et al. Humoral and Cellular Immune Responses against SARS-CoV-2 after Third Dose BNT162b2 following Double-Dose Vaccination with BNT162b2 versus ChAdOx1 in Patients with Cancer. Clin Cancer Res 2022;:OF1–OF12.4. Gattinger P, Niespodziana K, Stiasny K, Sahanic S, Tulaeva I, Borochova K et al. Neutralization of SARS-CoV-2 requires antibodies against conformational receptor-binding domain epitopes. Allergy2022;77 :230–242.

Allergic sensitization without detectable specific serum IgE

Immunoglobulin E (IgE)-mediated allergy is the most common hypersensitivity disease affecting more than 30% of the population. In genetically-predisposed subjects exposure to minute quantities of allergens leads to the production of IgE antibodies which is termed allergic sensitization and mainly occurs in early childhood. Allergen-specific IgE then binds to the high (FcRI) and low affinity receptors (FcRII, also called CD23) for IgE on effector cells and antigen-presenting cells, respectively. Subsequent and repeated allergen exposure increases allergen-specific IgE levels and, by receptor cross-linking, triggers immediate release of inflammatory mediators from mast cells and basophils whereas IgE-facilitated allergen presentation perpetuates T cell-mediated allergic inflammation. Due to engagement of receptors which are highly selective for IgE even tiny amounts of allergens can induce massive inflammation. Naturally occurring allergen-specific IgG and IgA antibodies usually recognize different epitopes on allergens compared to IgE, and do not efficiently interfere with allergen-induced inflammation. However IgG and IgA antibodies to these important IgE epitopes can be induced by allergen-specific immunotherapy or by passive immunization. These will lead to competition with IgE for binding with the allergen and prevent allergic responses. Similarly, anti-IgE treatment does the same by preventing IgE from binding to its receptor on mastcells and basophils. Here we review the complex interplay of allergen-specific IgE, IgG and IgA and the corresponding cell receptors in allergic diseases and its relevance for diagnosis, treatment and prevention of allergy.

Background. First vaccines for prevention of Coronavirus disease 2019 (COVID-19) are becoming available but there is a huge and unmet need for specific forms of treatment. In this study we aimed to evaluate the potent anti-SARS-CoV-2 effect of siRNA both in vitro and in vivo. Methods. To identify most effective molecule out of a panel of 15 in silico designed siRNAs, an in vitro screening system based on vectors expressing SARS-CoV-2 genes fused with the firefly luciferase reporter gene and SARS-CoV-2-infected cells was used. The most potent siRNA, siR-7, was modified by Locked nucleic acids (LNAs) to obtain siR-7-EM with increased stability and was formulated with the peptide dendrimer KK-46 for enhancing cellular uptake to allow topical application by inhalation of the final formulation - siR-7-EM/KK-46. Using the Syrian Hamster model for SARS-CoV-2 infection the antiviral capacity of siR-7-EM/KK-46 complex was evaluated. Results. We identified the siRNA, siR-7, targeting SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) as the most efficient siRNA inhibiting viral replication in vitro. Moreover, we have shown that LNA-modification and complexation with the designed peptide dendrimer enhanced the antiviral capacity of siR-7 in vitro. We demonstrated significant reduction of virus titer and total lung inflammation in the animals exposed by inhalation of siR-7-EM/KK-46 in vivo. Conclusions. Thus, we developed a therapeutic strategy for COVID-19 based on inhalation of a modified siRNA-peptide dendrimer formulation.

Background: Multiplex tests allow for measurement of allergen-specific IgE responses to multiple allergen extracts and components and have several advantages for large cohort studies. Due to significant methodological differences, test systems are difficult to integrate in meta-analyses/systematic reviews since there is a lack of datasets with direct comparison. We aimed to create models for statistical integration of allergen-specific IgE to peanut/tree nut allergens from three IgE-test platforms. Methods: Plasma from Canadian and Austrian children with peanut/tree nut sensitization and a cohort of sensitized, high-risk, pre-school asthmatics (total n=166) were measured with three R&D multiplex IgE test platforms: Allergy Explorer, ALEX (Macro Array Dx), MeDALL-chip (Mechanisms of Development of Allergy) (Thermo Fisher), and EUROLINE (EUROIMMUN). Skin prick test (n=51) and ImmunoCAP (n=62) results for extracts were available in a subset. Regression models (Multivariate Adaptive Regression Splines, local polynomial regression) were applied if >30% of samples were positive to the allergen. Intra-test correlations between PR-10 and nsLTP allergens were assessed. Results: Using two regression methods, we demonstrated the ability to model allergen-specific relationships with acceptable measures of fit (r2=94-56%) for peanut and tree nut sIgE testing at the extract and component-level, in order from highest to lowest: Ara h 2, Ara h 6, Jug r 1, Ana o 3, Ara h 1, Jug r 2, Cor a 9. Conclusion: Our models support the notion that conversion is reasonably possible between sIgE multiplex platforms for allergen extracts and components and may provide options to aggregate data for future meta-analysis.