Publications by authors named "Michael P Horn"

25 Publications

  • Page 1 of 1

Humoral and cellular responses to mRNA vaccines against SARS-CoV-2 in patients with a history of CD20 B-cell-depleting therapy (RituxiVac): an investigator-initiated, single-centre, open-label study.

Lancet Rheumatol 2021 Sep 7. Epub 2021 Sep 7.

Department of Nephrology and Hypertension, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland.

Background: B-cell-depleting therapies increase the risk of morbidity and mortality due to COVID-19. Evidence-based SARS-CoV-2 vaccination strategies for patients on B-cell-depleting therapies are scarce. We aimed to investigate humoral and cell-mediated immune responses to SARS-CoV-2 mRNA-based vaccines in patients receiving CD20-targeted B-cell-depleting agents for autoimmune disease, malignancy, or transplantation.

Methods: The RituxiVac study was an investigator-initiated, single-centre, open-label study done at the Bern University Hospital (Bern, Switzerland). Patients with a treatment history of anti-CD20-depleting agents (rituximab or ocrelizumab) and with no previous history of SARS-CoV-2 infection were enrolled between April 26 and June 30, 2021, for analysis of humoral and cell-mediated immune responses (by interferon-γ [IFNγ] release assay) at least 4 weeks after completing vaccination against SARS-CoV-2. Healthy controls without a history of SARS-CoV-2 infection were also enrolled at least 4 weeks after completing vaccination against SARS-CoV-2. All study participants received two doses of either the Pfizer-BioNTech BNT162b2 vaccine or the Moderna mRNA-1273 vaccine. The primary outcome was the proportion of patients with a history of anti-CD20 treatment who showed a humoral immune response against the SARS-CoV-2 spike protein in comparison with immunocompetent controls. Prespecified secondary endpoints were the effect of anti-CD20 therapy (including time since last treatment and cumulative dose) on humoral or cell-mediated immune responses to SARS-CoV-2 vaccination, and biomarkers of immunocompetence. This study is registered with ClinicalTrials.gov, NCT04877496.

Findings: The final study population comprised 96 patients and 29 immunocompetent controls. The median age of patients was 67 years (IQR 57-72) and of controls was 54 years (45-62), and 51 (53%) of 96 patients and 19 (66%) of 29 controls were female. The median time since last anti-CD20 treatment was 1·07 years (IQR 0·48-2·55) and the median cumulative dose of an anti-CD20 depleting agent was 2·80 g (1·50-5·00). Anti-spike IgG antibodies were detected in 47 (49%) of 96 patients 1·79 months (IQR 1·16-2·48) after the second vaccine dose compared to 29 (100%) of 29 controls 1·81 months (1·17-2·48) after the second vaccine dose (p<0·001). SARS-CoV-2-specific IFNγ release was detected in 13 (20%) of 66 patients and 21 (75%) of 28 of healthy controls (p<0·001). Only nine (14%) of 66 patients were double positive for anti-SARS-CoV-2 spike IgG and cell-mediated responses, compared with 21 (75%) of 28 healthy controls (p<0·001). Time since last anti-CD20 therapy (>7·6 months; positive predictive value 0·78), peripheral CD19 cell count (>27 cells per μL; positive predictive value 0·70), and CD4 lymphocyte count (>653 cells per μL; positive predictive value 0·71) were predictive of humoral vaccine response (area under the curve [AUC] 67% [95% CI 56-78] for time since last anti-CD20 therapy, 67% [55-80] for peripheral CD19 count, and 66% [54-79] for CD4 count).

Interpretation: This study provides further evidence of blunted humoral and cell-mediated immune responses elicited by SARS-CoV-2 mRNA vaccines in patients with a history of CD20 B-cell-depleting treatment. Lymphocyte subpopulation counts were associated with vaccine response in this highly vulnerable population. On validation, these results could help guide both the administration of SARS-CoV-2 vaccines and B-cell-depleting agents in this population.

Funding: Bern University Hospital.
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http://dx.doi.org/10.1016/S2665-9913(21)00251-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8423431PMC
September 2021

A novel functional mast cell assay for the detection of allergies.

J Allergy Clin Immunol 2021 Aug 18. Epub 2021 Aug 18.

Department of BioMedical Research, University of Bern, Bern, Switzerland; Department of Rheumatology, Immunology and Allergology, University Hospital Bern, Bern, Switzerland. Electronic address:

Background: Clinical management of allergic diseases has been hampered by the lack of safe and convenient tests to reliably identify culprit allergens and to closely follow changes in disease activity over time. Because allergy diagnosis is a complex and laborious multistep procedure, there is an urgent need for simpler but still functionally accurate ex vivo assays allowing objective diagnosis, substantiating treatment choices, and quantifying therapeutic responses.

Objective: In this study, we sought to develop a novel functional cell-based assay that relies on passive sensitization of allergic effector cells with patient serum, circumventing current limitations in allergy diagnosis.

Methods: We genetically engineered a conditional homeobox B8 (Hoxb8)-immortalized progenitor line from the bone marrow of mice that are transgenic for the human high-affinity IgE receptor (FcεRIα). These cells can be reproducibly differentiated into mature Hoxb8 mast cells within 5 days of culture in virtually unlimited numbers.

Results: We demonstrate that the established Hoxb8 mast cell assay can be used to accurately measure total IgE levels, identify culprit allergens, longitudinally monitor allergen-specific immunotherapy, and potentially determine the time point of tolerance induction upon allergen-specific immunotherapy in patients with allergy. To facilitate the analysis of large testing volumes, we demonstrate a proof-of-concept for a high-throughput screening application based on fluorescent cell barcoding using the engineered Hoxb8 mast cells.

Conclusions: Our results indicate that this novel mast cell assay could represent a valuable tool to support clinicians in the identification of IgE-mediated allergies and in the quantification of treatment efficacy as well as duration of therapeutic response.
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http://dx.doi.org/10.1016/j.jaci.2021.08.006DOI Listing
August 2021

Cutaneous involvement in anti-HMGCR positive necrotizing myopathy.

J Autoimmun 2021 Sep 28;123:102691. Epub 2021 Jul 28.

Department of Rheumatology and Immunology, University Hospital and, University of Bern, CH-3010, Bern, Switzerland; Medical Center Monbijou (MZM), CH-3011, Bern, Switzerland. Electronic address:

Objective: Anti-3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) positive immune-mediated necrotizing myopathy (IMNM) is a rare disease. It is induced by exogenous substances, most often by statins. Little is known about cutaneous manifestations of HMGCR positive IMNM and about HMGCR antibody positivity in other diseases.

Methods: The characteristics of patients with anti-HMGCR autoantibodies measured at our laboratory between January 2012 and September 2020 were studied. Characteristics of patients with IMNM were compared to those patients with positive antibodies but without muscle involvement. Associations of IMNM with other organ involvements were searched for.

Results: Of the 32 patients studied, 23 showed characteristics of IMNM, 9 did not fulfill current classification criteria but most showed signs of connective tissue diseases. Patients with IMNM were older (66 and 35 years, respectively; 0.92 (0.73-0.98); p < 0.001), had more frequent statin exposure (87% and 33%, respectively; 0.84 (0.61-0.94); p = 0.005) and higher mean peak CK (8717U/l and 329U/l, respectively; 1.0 (0.85-1.0); p < 0.001). 13/23 (56%) of IMNM patients showed cutaneous lesions; none of the patients suffered from cancer; only three IMNM patients showed drug-free complete remission. Incidence of IMNM in the catchment area of our center is at least 2.7/Mio/year.

Conclusion: Cutaneous lesions were found to be more frequent in anti-HMRCR positive IMNM than previously reported. Titer of anti-HMGCR antibodies and CK levels were significantly higher in IMNM than in other autoimmune connective tissue diseases. The data support the hypothesis of an antigen-driven response in IMNM, and suggests an activation of autoreactive B-lymphocytes in non-IMNM patients.
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http://dx.doi.org/10.1016/j.jaut.2021.102691DOI Listing
September 2021

The Release Kinetics of Eosinophil Peroxidase and Mitochondrial DNA Is Different in Association with Eosinophil Extracellular Trap Formation.

Cells 2021 02 3;10(2). Epub 2021 Feb 3.

Institute of Pharmacology, University of Bern, Inselspital, INO-F, CH-3010 Bern, Switzerland.

Eosinophils are a subset of granulocytes characterized by a high abundance of specific granules in their cytoplasm. To act as effector cells, eosinophils degranulate and form eosinophil extracellular traps (EETs), which contain double-stranded DNA (dsDNA) co-localized with granule proteins. The exact molecular mechanism of EET formation remains unknown. Although the term "EET release" has been used in scientific reports, it is unclear whether EETs are pre-formed in eosinophils and subsequently released. Moreover, although eosinophil degranulation has been extensively studied, a precise time-course of granule protein release has not been reported until now. In this study, we investigated the time-dependent release of eosinophil peroxidase (EPX) and mitochondrial DNA (mtDNA) following activation of both human and mouse eosinophils. Unexpectedly, maximal degranulation was already observed within 1 min with no further change upon complement factor 5 (C5a) stimulation of interleukin-5 (IL-5) or granulocyte/macrophage colony-stimulating factor (GM-CSF)-primed eosinophils. In contrast, bulk mtDNA release in the same eosinophil populations occurred much slower and reached maximal levels between 30 and 60 min. Although no single-cell analyses have been performed, these data suggest that the molecular pathways leading to degranulation and mtDNA release are at least partially different. Moreover, based on these data, it is likely that the association between the mtDNA scaffold and granule proteins in the process of EET formation occurs in the extracellular space.
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http://dx.doi.org/10.3390/cells10020306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7913244PMC
February 2021

Serum calprotectin as new biomarker for disease severity in idiopathic pulmonary fibrosis: a cross-sectional study in two independent cohorts.

BMJ Open Respir Res 2021 01;8(1)

Department for Pulmonary Medicine, Inselspital, Bern University Hospital, Department for BioMedical Research (DBMR), University of Bern, Bern, Switzerland

Background: Non-invasive biomarkers for the assessment of disease severity in idiopathic pulmonary fibrosis (IPF) are urgently needed. Calprotectin belongs to the S-100 proteins produced by neutrophils, which likely contribute to IPF pathogenesis. Calprotectin is a well-established biomarker in inflammatory bowel diseases. In this cross-sectional study, we aimed to establish the potential role of calprotectin as a biomarker in IPF. Specifically, we hypothesised that patients with IPF have higher serum calprotectin levels compared with healthy controls, and that calprotectin levels are associated with disease severity.

Methods: Blood samples were obtained from healthy volunteers (n=26) and from two independent IPF cohorts (derivation cohort n=26, validation cohort n=66). Serum calprotectin levels were measured with a commercial kit adapted for that purpose and compared between healthy controls and patients with IPF. Clinical parameters, including forced vital capacity, diffusing capacity for carbon monoxide (DLCO) and the Composite Physiologic Index (CPI), were correlated with calprotectin serum levels.

Results: The IPF derivation cohort showed increased serum calprotectin levels compared with healthy controls (2.47±1.67 vs 0.97±0.53 µg/mL, p<0.001). In addition, serum calprotectin levels correlated with DLCO% predicted (r=-0.53, p=0.007) and with CPI (r=0.66, p=0.007). These findings were confirmed in an independent IPF validation cohort.

Conclusion: Serum calprotectin levels are significantly increased in patients with IPF compared with healthy controls and correlate with DLCO and CPI. Calprotectin might be a potential new biomarker for disease severity in IPF.
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http://dx.doi.org/10.1136/bmjresp-2020-000827DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7813379PMC
January 2021

Evaluation of diagnostic criteria and red flags of myelin oligodendrocyte glycoprotein encephalomyelitis in a clinical routine cohort.

CNS Neurosci Ther 2021 04 13;27(4):426-438. Epub 2020 Oct 13.

Department of Neurology, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland.

Aims: Myelin oligodendrocyte glycoprotein antibodies (MOG-IgG) have been proposed to define "MOG encephalomyelitis" (MOG-EM), with published diagnostic and "red flag" criteria. We aimed to evaluate these criteria in a routine clinical setting.

Methods: We retrospectively analyzed patients with borderline/positive MOG-IgG and applied the diagnostic and red flag criteria to determine likelihood of MOG-EM diagnosis. Para-/clinical parameters were described and analyzed with chi-square test.

Results: In total, 37 patients fulfilled MOG-EM diagnostic criteria (female-to-male ratio: 1.6:1, median onset age: 28.0 years [IQR 18.5-40.5], n = 8 with pediatric onset). In 24/37, red flags were present, predominantly MOG-IgG at assay cutoff and/or MRI lesions suggestive of multiple sclerosis (MS). As proposed in the consensus criteria, these patients should rather be described as "possible" MOG-EM. Of these, we classified 13 patients as "unlikely" MOG-EM in the presence of the red flag "borderline MOG-IgG" with negative MOG-IgG retest or coincidence of ≥1 additional red flag. This group mainly consisted of patients diagnosed with MS (n = 11). Frequency of cerebrospinal fluid (CSF-)-specific oligoclonal bands (OCB) is significantly lower in definite vs possible and unlikely MOG-EM (P = .0005).

Conclusion: Evaluation of diagnostic and red flag criteria, MOG-IgG retesting (incl. change of assay), and CSF-specific OCB are relevant in clinical routine cohorts to differentiate MOG-EM from MS.
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http://dx.doi.org/10.1111/cns.13461DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7941167PMC
April 2021

Accuracy of serological testing for SARS-CoV-2 antibodies: First results of a large mixed-method evaluation study.

Allergy 2021 03 13;76(3):853-865. Epub 2020 Nov 13.

University Institute of Clinical Chemistry, Inselspital University Hospital, Bern, Switzerland.

Background: Serological immunoassays that can identify protective immunity against SARS-CoV-2 are needed to adapt quarantine measures, assess vaccination responses, and evaluate donor plasma. To date, however, the utility of such immunoassays remains unclear. In a mixed-design evaluation study, we compared the diagnostic accuracy of serological immunoassays that are based on various SARS-CoV-2 proteins and assessed the neutralizing activity of antibodies in patient sera.

Methods: Consecutive patients admitted with confirmed SARS-CoV-2 infection were prospectively followed alongside medical staff and biobank samples from winter 2018/2019. An in-house enzyme-linked immunosorbent assay utilizing recombinant receptor-binding domain (RBD) of the SARS-CoV-2 spike protein was developed and compared to three commercially available enzyme-linked immunosorbent assays (ELISAs) targeting the nucleoprotein (N), the S1 domain of the spike protein (S1), and a lateral flow immunoassay (LFI) based on full-length spike protein. Neutralization assays with live SARS-CoV-2 were performed.

Results: One thousand four hundred and seventy-seven individuals were included comprising 112 SARS-CoV-2 positives (defined as a positive real-time PCR result; prevalence 7.6%). IgG seroconversion occurred between day 0 and day 21. While the ELISAs showed sensitivities of 88.4% for RBD, 89.3% for S1, and 72.9% for N protein, the specificity was above 94% for all tests. Out of 54 SARS-CoV-2 positive individuals, 96.3% showed full neutralization of live SARS-CoV-2 at serum dilutions ≥ 1:16, while none of the 6 SARS-CoV-2-negative sera revealed neutralizing activity.

Conclusions: ELISAs targeting RBD and S1 protein of SARS-CoV-2 are promising immunoassays which shall be further evaluated in studies verifying diagnostic accuracy and protective immunity against SARS-CoV-2.
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http://dx.doi.org/10.1111/all.14608DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7537154PMC
March 2021

Conventional autoantibodies against brain antigens are not routinely detectable in serum and CSF of narcolepsy type 1 and 2 patients.

Sleep Med 2020 11 7;75:188-191. Epub 2020 Aug 7.

Department of Neurology, Inselspital, Bern University Hospital and University of Bern, Bern, Switzerland; Neurology Department, Sechenov First Moscow State Medical University, Moscow, Russia.

Narcolepsy with cataplexy (NT1) is a chronic hypothalamic disorder with a presumed immune-mediated etiology leading to a loss of hypocretin neurons. Previous studies reported conflicting results in terms of presence of auto-antibodies involved in narcolepsy pathophysiology. A total of 86 patients with primary/idiopathic narcolepsy (74 NT1, 12 NT2) and 23 control patients with excessive daytime sleepiness due to other causes were tested for the presence of a wide range of anti-neuronal antibodies in both serum and cerebrospinal fluid (CSF). Anti-neuronal antibodies were rarely found in patients with narcolepsy (n = 2) and in controls (n = 1). Our results are in line with previous reports. We can therefore support the current evidence, that conventional anti-neuronal antibodies are not routinely detected during the workup of NT1 and other CDH patients.
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http://dx.doi.org/10.1016/j.sleep.2020.08.001DOI Listing
November 2020

PR3-ANCA and panel diagnostics in pediatric inflammatory bowel disease to distinguish ulcerative colitis from Crohn's disease.

PLoS One 2018 17;13(12):e0208974. Epub 2018 Dec 17.

Division of Pediatric Gastroenterology, Hepatology and Nutrition, University Children's Hospital, Inselspital, University of Bern, Bern, Switzerland.

Background: Accurate classification of patients with inflammatory bowel disease into the subtypes ulcerative colitis (UC) and Crohn's disease (CD) is still a challenge, but important for therapy and prognosis.

Objectives: To evaluate the diagnostic utility of anti-neutrophil cytoplasmic antibodies specific for proteinase-3 (PR3-ANCA) for ulcerative colitis (UC) and the value of an antibody panel incorporating PR3-ANCA to differentiate between Crohn's disease (CD) and UC.

Study Design: In this cohort study, 122 pediatric and adolescent individuals were retrospectively included (61 IBD patients of two clinical centers, 61 non-IBD controls). All subjects had a comprehensive antibody profile done from stored sera taken close to time of diagnosis. By employing quasi-exhaustive logistic regression the best discriminative model for UC and CD,subjects was determined in a training cohort and confirmed in a validation cohort.

Results: PR3-ANCA was specifically associated with UC (odds ratio (OR), 17.6; 95% confidence interval (CI); 3.6, 87); P < .001). A four antibody-panel including PR3-ANCA had an AUC of 90.81% (95%CI; 81.93, 99.69) to distinguish between UC and CD in the training cohort. In a smaller external validation cohort, the AUC was 84.13% (95%CI; 64.21, 100) for accurate diagnosis of CD and UC.

Conclusion: PR3-ANCA is highly specific for UC. The differentiating capability of a panel, which contains PR3-ANCA and weighs broadly available antibodies, is superior and utilization of the panel can support accurate classification in the work-up of pediatric and adolescent patients with IBD patients.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0208974PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6296712PMC
May 2019

Rebound After Fingolimod and a Single Daclizumab Injection in a Patient Retrospectively Diagnosed With NMO Spectrum Disorder-MRI Apparent Diffusion Coefficient Maps in Differential Diagnosis of Demyelinating CNS Disorders.

Front Neurol 2018 27;9:782. Epub 2018 Sep 27.

Department of Neurology, Inselspital, University Hospital and University of Bern, Bern, Switzerland.

Differential diagnosis of neuromyelitis optica spectrum disorders (NMOSD) and multiple sclerosis (MS) or mimics can be challenging, especially in patients with atypical presentations and negative serostatus for aquaporin-4 antibodies (AQP4-Ab). This brief research report describes magnetic resonance imaging (MRI) findings focusing on quantitative apparent diffusion coefficient (ADC) histogram analysis as a potential tool to differentiate NMOSD from MS. Longitudinal MRI data obtained during routine clinical examinations were retrospectively analyzed in a patient with histologically determined cerebral NMOSD, a patient with an acute tumefactive MS lesion, and a patient with ischemic stroke. Histogram analyses of ADC maps were evaluated. A patient diagnosed with MS experienced a severe rebound after fingolimod withdrawal and a single daclizumab injection. Cerebral NMOSD manifestation was confirmed by brain biopsy. However, the patient did not fulfill consensus criteria for NMOSD and was AQP4-Ab negative. Comparison of ADC histogram analyses of this patient with those from a patient with MS and one with ischemic stroke revealed differential ADC characteristics: namely a more pronounced and prolonged ADC leftward shift in inflammatory than in ischemic pathology, even more accentuated in NMOSD versus MS. ADC map histograms and ADC threshold values for different conditions may be useful for differentiation of large inflammatory brain lesions and further studies are merited.
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http://dx.doi.org/10.3389/fneur.2018.00782DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6170610PMC
September 2018

Isolation of Natural Anti-FcεRIα Autoantibodies from Healthy Donors.

Methods Mol Biol 2017 ;1643:5-22

University Institute of Clinical Chemistry and Center of Laboratory Medicine, Inselspital, University Hospital of Bern, CH-3010, Bern, Switzerland.

Natural antibodies are defined as antibodies detected in a healthy individual without active immunization. These antibodies are specific for exoantigens, as well as for autoantigens, mostly without any pathogenic role. Most of the studies conducted with natural (auto-) antibodies have been performed using affinity purified antibodies from individual sera or polyclonal Ig-preparations such as Intravenous Ig (IVIg). For in-depth analysis of such autoantibodies affinity-purified Ig-preparations from healthy individuals are of no use, as they are oligoclonal or polyclonal. Thus, there is a need of human monoclonal autoantibodies. Human monoclonal autoantibodies can be produced from B cells isolated from humans; however, this requires the screening of a large number of antibodies to identify one among them specific to an antigen. Using the phage display technology we generated such autoantibodies against the alpha subunit of the high-affinity IgE receptor (FcεRIα). Here we describe the step-by-step protocol for the generation of such libraries and the isolation of autoantibodies by affinity panning.
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http://dx.doi.org/10.1007/978-1-4939-7180-0_2DOI Listing
March 2018

Rapid immunoassays for diagnosis of heparin-induced thrombocytopenia: Comparison of diagnostic accuracy, reproducibility, and costs in clinical practice.

PLoS One 2017 8;12(6):e0178289. Epub 2017 Jun 8.

Department of Haematology, Inselspital, Bern University Hospital, University of Bern, Berne, Switzerland.

Background: Immunoassays are crucial in the work-up of patients with suspected heparin-induced thrombocytopenia (HIT) and rapid tests have been recently developed. However, comparative data on diagnostic accuracy, reproducibility, and analytical costs of different immunoassays in clinical practice are limited.

Methods: Samples of 179 consecutive patients evaluated for suspected HIT in clinical practice using a polyspecific enzyme-linked immunoabsorbent assay (GTI diagnostics; ELISA) and a rapid particle gel immunoassay (PaGIA), were additionally analysed with a IgG-specific chemiluminescent immunoassay (AcuStar HIT-IgG). Presence of HIT was defined as a positive functional heparin-induced platelet aggregation test. Diagnostic accuracy was determined for low, intermediate and high thresholds as previously established (ELISA: optical density 0.4, 1.3, and 2.0 respectively; PaGIA: positive/negative, titre of 4, titre of 32; AcuStar HIT-IgG: 1.0 U/ml, 2.8, 9.4) and reproducibility was assessed by repeated measurements. Costs of test determination were calculated taking reagents, controls, and working time of technicians according to Swiss health care system into account.

Results: Data on PaGIA results were available for 171 patients (95.5%), ELISA for 144 patients (80.4%), and AcuStar HIT-IgG for 179 patients (100%). Sensitivity was above 95% for all assays at low and intermediate thresholds. Specificity increased with higher thresholds and was above 90% for all assays with intermediate and high thresholds. Specificity of AcuStar HIT-IgG (92.8%; 95% CI 87.7, 96.2) was significantly higher than PaGIA (83.0%; 95% CI 76.3, 88.5) and higher than ELISA (81.8%, 95% CI 74.2, 88.0) at low threshold (p<0.05). Reproducibility was adequate for all assays. Total costs per test were CHF 51.02 for ELISA, 117.70 for AcuStar HIT-IgG, and 83.13 for PaGIA.

Conclusions: We observed favourable diagnostic accuracy measures and a high reproducibility for PaGIA and AcuStar HIT-IgG. Implementation into 24-hours-service might improve patient care but the results must be confirmed in other settings and larger populations as well.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0178289PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5464550PMC
September 2017

Rituximab for the Treatment of Isolated Refractory Desquamative Gingivitis Due to Mucous Membrane Pemphigoid.

JAMA Dermatol 2016 12;152(12):1396-1398

Department of Dermatology, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland.

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http://dx.doi.org/10.1001/jamadermatol.2016.3434DOI Listing
December 2016

Designed ankyrin repeat proteins: a new approach to mimic complex antigens for diagnostic purposes?

PLoS One 2013 23;8(4):e60688. Epub 2013 Apr 23.

University Institute of Immunology, University of Bern, Inselspital, Bern, Switzerland.

Inhibitory antibodies directed against coagulation factor VIII (FVIII) can be found in patients with acquired and congenital hemophilia A. Such FVIII-inhibiting antibodies are routinely detected by the functional Bethesda Assay. However, this assay has a low sensitivity and shows a high inter-laboratory variability. Another method to detect antibodies recognizing FVIII is ELISA, but this test does not allow the distinction between inhibitory and non-inhibitory antibodies. Therefore, we aimed at replacing the intricate antigen FVIII by Designed Ankyrin Repeat Proteins (DARPins) mimicking the epitopes of FVIII inhibitors. As a model we used the well-described inhibitory human monoclonal anti-FVIII antibody, Bo2C11, for the selection on DARPin libraries. Two DARPins were selected binding to the antigen-binding site of Bo2C11, which mimic thus a functional epitope on FVIII. These DARPins inhibited the binding of the antibody to its antigen and restored FVIII activity as determined in the Bethesda assay. Furthermore, the specific DARPins were able to recognize the target antibody in human plasma and could therefore be used to test for the presence of Bo2C11-like antibodies in a large set of hemophilia A patients. These data suggest, that our approach might be used to isolate epitopes from different sets of anti-FVIII antibodies in order to develop an ELISA-based screening assay allowing the distinction of inhibitory and non-inhibitory anti-FVIII antibodies according to their antibody signatures.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0060688PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3634029PMC
November 2013

Preclinical in vitro and in vivo characterization of the fully human monoclonal IgM antibody KBPA101 specific for Pseudomonas aeruginosa serotype IATS-O11.

Antimicrob Agents Chemother 2010 Jun 22;54(6):2338-44. Epub 2010 Mar 22.

Kenta Biotech AG, Rehhagstrasse 79, CH-3018 Bern, Switzerland.

Pseudomonas aeruginosa infection in ventilator-associated pneumonia is a serious and often life-threatening complication in intensive care unit patients, and new treatment options are needed. We used B-cell-enriched peripheral blood lymphocytes from a volunteer immunized with a P. aeruginosa O-polysaccharide-toxin A conjugate vaccine to generate human hybridoma cell lines producing monoclonal antibodies specific for individual P. aeruginosa lipopolysaccharide serotypes. The fully human monoclonal antibody secreted by one of these lines, KBPA101, is an IgM/kappa antibody that binds P. aeruginosa of International Antigenic Typing System (IATS) serotype O11 with high avidity (5.81 x 10(7) M(-1) +/- 2.8 x 10(7) M(-1)) without cross-reacting with other serotypes. KBPA101 specifically opsonized the P. aeruginosa of IATS O11 serotype and mediated complement-dependent phagocytosis in vitro by the human monocyte-like cell line HL-60 at a very low concentration (half-maximal phagocytosis at 0.16 ng/ml). In vivo evaluation of KBPA101 demonstrated a dose-response relationship for protection against systemic infections in a murine burn wound sepsis model, where 70 to 100% of animals were protected against lethal challenges with P. aeruginosa at doses as low as 5 microg/animal. Furthermore, a high efficacy of KBPA101 in protection from local respiratory infections in an acute lung infection model in mice was demonstrated. Preclinical toxicology evaluation on human tissue, in rabbits, and in mice did not indicate any toxicity of KBPA101. Based on these preclinical findings, the first human clinical trials have been initiated.
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http://dx.doi.org/10.1128/AAC.01142-09DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2876355PMC
June 2010

Pharmacokinetics and safety profile of the human anti-Pseudomonas aeruginosa serotype O11 immunoglobulin M monoclonal antibody KBPA-101 in healthy volunteers.

Antimicrob Agents Chemother 2009 Aug 18;53(8):3442-6. Epub 2009 May 18.

Kenta Biotech, Rehhagstrasse 79, 3018 Bern, Switzerland.

KBPA-101 is a human monoclonal antibody of the immunoglobulin M isotype, which is directed against the O-polysaccharide moiety of Pseudomonas aeruginosa serotype O11. This double-blind, dose escalation study evaluated the safety and pharmacokinetics of KBPA-101 in 32 healthy volunteers aged 19 to 46 years. Each subject received a single intravenous infusion of KBPA-101 at a dose of 0.1, 0.4, 1.2, or 4 mg/kg of body weight or placebo infused over 2 h. Plasma samples for pharmacokinetic assessments were taken before infusion as well as 0.25, 0.5, 1, 2, 2.5, 4, 6, 8, 12, 24, 36, and 48 h and 4, 7, 10, and 14 days after start of dosing. Plasma concentrations of KBPA-101 were detected with mean maximum concentrations of drug in plasma of 1,877, 7,571, 24,923, and 83,197 ng/ml following doses of 0.1, 0.4, 1.2, and 4.0 mg/kg body weight, respectively. The mean elimination half-life was between 70 and 95 h. The mean volume of distribution was between 4.76 and 5.47 liters. Clearance ranged between 0.039 and 0.120 liters/h. At the highest dose of 4.0 mg/kg, plasma KBPA-101 levels were greater than 5,000 ng/ml for 14 days. KBPA-101 exhibited linear kinetics across all doses. No anti-KBPA-101 antibodies were detected after dosing in any subject. Overall, the human monoclonal antibody KBPA-101 was well tolerated over the entire dose range in healthy volunteers, and no serious adverse events have been reported.
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http://dx.doi.org/10.1128/AAC.01699-08DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2715602PMC
August 2009

Mutation screening of the medium-chain acyl-CoA dehydrogenase (MCAD) and the ornithine transcarbamylase (OTC) genes by multiplex PCR amplification and sequencing.

Clin Chem Lab Med 2009 ;47(1):56-9

Molecular Diagnostics DOLS, Inselspital, Bern University Hospital and University of Bern, Bern, Switzerland.

Background: Sequencing based mutation screening assays of genes encompassing large numbers of exons could be substantially optimized by multiplex PCR, which enables simultaneous amplification of many targets in one reaction. In the present study, a multiplex PCR protocol originally developed for fragment analysis was evaluated for sequencing based mutation screening of the ornithine transcarbamylase (OTC) and the medium-chain acyl-CoA dehydrogenase (MCAD) genes.

Methods: Single exon and multiplex PCR protocols were applied to generate PCR templates for subsequent DNA sequencing of all exons of the OTC and the MCAD genes. For each PCR protocol and using the same DNA samples, 66 OTC and 98 MCAD sequence reads were generated. The sequences derived from the two different PCR methods were compared at the level of individual signal-to-noise ratios of the four bases and the proportion of high-quality base-signals.

Results: The single exon and the multiplex PCR protocol gave qualitatively comparable results for the two genes.

Conclusions: Many existing sequencing based mutation analysis protocols may be easily optimized with the proposed method, since the multiplex PCR protocol was successfully applied without any re-design of the PCR primers and other optimization steps for generating sequencing templates for the OTC and MCAD genes, respectively.
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http://dx.doi.org/10.1515/CCLM.2009.012DOI Listing
March 2009

Antibody responses induced by long-term vaccination with an octovalent conjugate Pseudomonas aeruginosa vaccine in children with cystic fibrosis.

FEMS Immunol Med Microbiol 2006 Jul;47(2):302-8

Berna Biotech Ltd, Research Immunology, Bern, Switzerland.

We assessed the serological responses over 10 years to repeated immunization of cystic fibrosis (CF) patients with an O-polysaccharide (OPS)-toxin A conjugate vaccine against Pseudomonas aeruginosa. A retrospective analysis was performed with sera from 25 vaccinated and 25 unvaccinated children treated at the same CF centre and matched for clinical management, age and gender. Yearly immunization led to sustained elevations of serum immunoglobulin G (IgG) antibody levels to all vaccine components. Eighteen unvaccinated patients but only eight vaccinated ones developed chronic pseudomonal lung infections. Infection rapidly caused further marked elevations of polysaccharide- but not toxin A-specific serum IgG in both immunized and nonimmunized patients, indicating that protection did not depend on the quantity of IgG present. However, qualitative analyses revealed that the protective capacity of specific serum IgG antibodies was linked to high affinity and to specificity for OPS serotypes rather than for lipopolysaccharide core epitopes.
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http://dx.doi.org/10.1111/j.1574-695X.2006.00103.xDOI Listing
July 2006

Intranasal immunisation with conjugate vaccine protects mice from systemic and respiratory tract infection with Pseudomonas aeruginosa.

Vaccine 2006 May 20;24(20):4333-42. Epub 2006 Mar 20.

Berna Biotech Ltd., Rehhagstrasse 79, CH-3018 Bern, Switzerland.

We tested intranasal application of anti-Pseudomonas conjugate vaccine in mice. Comparison of immunisation via the intra-muscular versus intranasal routes showed the induction of equivalent levels of specific serum IgG and IgG subclasses antibodies if cholera toxin was used as an adjuvant. In contrast, secretion of specific mucosal IgA antibodies in the upper respiratory tract was only observed after intranasal immunisation together with adjuvant. Systemic and mucosal immunity was also established via the intranasal route when CpG-containing oligonucleotides were used as adjuvant. The functionality of intranasally induced antibodies was proven in vitro by opsonophagocytosis and in vivo using the burn-wound sepsis and intra-tracheal lung infection models. These results demonstrate the feasibility of intranasal immunisation against P. aeruginosa with conjugate vaccine.
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http://dx.doi.org/10.1016/j.vaccine.2006.03.007DOI Listing
May 2006

Expression of enterotoxigenic Escherichia coli colonization factors in Vibrio cholerae.

Vaccine 2006 May 20;24(20):4354-68. Epub 2006 Mar 20.

Berna Biotech Ltd., Department of Live Bacterial Vaccines, Rehhagstrasse 79, 3018 Bern, Switzerland.

As a first step towards a vaccine against diarrhoeal disease caused by enterotoxigenic Escherichia coli (ETEC), we have studied the expression of several ETEC antigens in the live attenuated Vibrio cholerae vaccine strain CVD 103-HgR. Colonization factors (CF) CFA/I, CS3, and CS6 were expressed at the surface of V. cholerae CVD 103-HgR. Both CFA/I and CS3 required the co-expression of a positive regulator for expression, while CS6 was expressed without regulation. Up-regulation of CF expression in V. cholerae was very efficient, so that high amounts of CFA/I and CS3 similar to those in wild-type ETEC were synthesized from chromosomally integrated CF and positive regulator loci. Increasing either the operon and/or the positive regulator gene dosage resulted in only a small increase in CFA/I and CS3 expression. In contrast, the level of expression of the non-regulated CS6 fimbriae appeared to be more dependent on gene dosage. While CF expression in wild-type ETEC is known to be tightly thermoregulated and medium dependent, it seems to be less stringent in V. cholerae. Finally, co-expression of two or three CFs in the same strain was efficient even under the control of one single regulator gene.
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http://dx.doi.org/10.1016/j.vaccine.2006.02.052DOI Listing
May 2006

Prophylaxis and therapy of Pseudomonas aeruginosa infection in cystic fibrosis and immunocompromised patients.

Vaccine 2004 Dec;22 Suppl 1:S44-8

Berna Biotech Ltd., Research Immunology, Rehhagstrasse 79, Bern CH-3018, Switzerland.

Pseudomonas aeruginosa is an opportunistic bacterium responsible for chronic lung infection in cystic fibrosis patients, as well as nosocomial infections in immunocompromised patients. An O-polysaccharide-toxin A conjugate vaccine was evaluated for prophylaxis of P. aeruginosa in cystic fibrosis patients. Vaccination proved to be useful in preventing and/or delaying infection. Fully human monoclonal antibodies (mAb) against P. aeruginosa O-polysaccharides were developed for the treatment of immunocompromised patients in whom active immunoprophylaxis is not applicable. Characterisation of the mAb revealed high antigen specificity and avidity, as well as excellent efficacy in relevant in vitro and in vivo systems, permitting future clinical evaluation.
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http://dx.doi.org/10.1016/j.vaccine.2004.08.016DOI Listing
December 2004

B cells alter the phenotype and function of follicular-homing CXCR5+ T cells.

Eur J Immunol 2004 Dec;34(12):3562-71

Theodor-Kocher Institute, University of Bern, Bern, Switzerland.

The CXC chemokine receptor (CXCR)5 is rapidly induced on activated CD4(+) T cells, allowing migration toward secondary lymphoid tissue follicles, where the CXCR5 ligand CXCL13/BCA-1 is produced. Such CXCR5(+) T cells provide efficient help for B cell immunoglobulin production and are termed follicular B helper T (T(FH)) cells. However, the molecular mechanisms by which T(FH) cells provide B cell help are unknown. Here, we demonstrate that newly generated (antigen-primed) T(FH) cells express a phenotype consistent with induction of B cell proliferation, but co-culture with primed B cells resulted in a switch to a plasma cell-inducing phenotype, characterized by loss of CD154, induction of CD70 and an increase in IL-10 production capacity. The ability to produce IL-10 could be maintained as a stable phenotype, but its secretion was strictly dependent on inducible costimulator (ICOS) signaling. Furthermore, B cells preserved a lymph node migration phenotype in proliferating T(FH) cells by preventing the loss of CC chemokine receptor (CCR)7 and the induction of CCR5. Thus, B cells directly modulate the B cell helper phenotype in T(FH) cells and actively promote their prolonged co-localization with these cells.
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http://dx.doi.org/10.1002/eji.200425478DOI Listing
December 2004

Natural anti-FcepsilonRIalpha autoantibodies may interfere with diagnostic tests for autoimmune urticaria.

J Autoimmun 2004 Feb;22(1):43-51

Institute of Immunology, Sahlihaus 1, Inselspital, CH-3010 Bern, Switzerland.

IgG autoantibodies against the alpha-chain of the high affinity IgE receptor are claimed to play a pathogenetic role in autoimmune urticaria. The best methods for detection of functional autoantibodies are currently the autologous serum skin test and the basophil histamine release assay. A simplified and feasible screening test would facilitate the diagnosis of autoimmune urticaria. Here we offer an explanation for the difficulties in establishing a screening test for autoantibodies directed against the alpha-chain of the high affinity IgE receptor in autoimmune urticaria. Identical autoantibodies in chronic urticaria patients and healthy donors belonging to the natural autoantibody repertoire were found by sequence analysis of anti-alpha-chain autoantibodies isolated by repertoire cloning from antibody libraries. These natural autoantibodies bound to the receptor and triggered histamine release but only if IgE was previously removed from the receptor. Diagnostic assays used for detection of antibodies directed against the IgE receptor may require signal comparison with and without the artificial removal of IgE, immune complexes, and complement in order to avoid false positive or negative results. After IgE removal diagnostic tests will detect natural autoantibodies against the high affinity IgE receptor regardless of whether they are pathogenic or not.
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http://dx.doi.org/10.1016/j.jaut.2003.09.007DOI Listing
February 2004

C-kit positive porcine bone marrow progenitor cells identified and enriched using recombinant stem cell factor.

J Immunol Methods 2003 Sep;280(1-2):113-23

Institute of Virology and Immunoprophylaxis, Sensemattstrasse 293, CH-3147 Mittelhäusern, Switzerland.

Porcine haematological studies have been hampered by the lack of monoclonal antibodies against porcine CD34 or CD117 expressed on haematological progenitors. The present report describes the enumeration, phenotyping and isolation of porcine haematopoietic progenitor cells expressing stem cell factor (SCF, c-kit ligand) receptor (c-kit, CD117). Recombinant porcine (rp) SCF and granulocyte-macrophage colony-stimulating factor (GM-CSF) were expressed in the mammalian HEK293 cell-based expression system. Both were biologically active and induced the proliferation of the human erythroleukemic cell line TF-1, as well as of porcine bone marrow haematopoietic cells (BMHC), in a concentration-dependent manner. The effect of rpSCF on BMHC proliferation was synergistic with rpGM-CSF. Furthermore, rpSCF had a synergistic effect on the generation of BMHC-derived dendritic cells (DC) induced by GM-CSF and TNF-alpha. RpSCF was expressed with a 6-histidine epitope, permitting both its purification and immunological detection. Binding studies with BMHC demonstrated ligation of SCF to 4-11% of BMHC. These cells represented the SWC3(low/-)SWC8- BMHC subset, with characteristics of immature proliferative progenitor BMHC. In contrast, no expression was noted on the SWC3+SWC8- monocytic, the SWC3+SWC8+ granulocytic or the SWC3-SWC8+ B cell lineage cells. Using magnetic or fluorescence-activated cell sorting, SCF-ligating BMHC were enriched for pluripotent progenitor cells. In this manner, porcine haematological studies can be pursued in a detailed manner not before possible.
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http://dx.doi.org/10.1016/s0022-1759(03)00273-4DOI Listing
September 2003

The prevalence of proteolytic antibodies against factor VIII in hemophilia A.

N Engl J Med 2002 Feb;346(9):662-7

INSERM Unité 430 and Université Pierre et Marie Curie, Hôpital Broussais, Paris, France.

Background: Factor VIII inhibitors are IgG alloantibodies that arise during replacement therapy in 25 to 50 percent of patients with severe hemophilia A. The hydrolysis of factor VIII by anti--factor VIII antibodies has been proposed as a mechanism of inactivation of factor VIII.

Methods: We purified IgG from patients with severe hemophilia A. The proteolytic activity of the antibodies was assessed by incubating the IgG with biotinylated human factor VIII and analyzing patterns of factor VIII cleavage by sodium dodecyl sulfate--polyacrylamide-gel electrophoresis and immunoblotting. The controls were normal human IgG and IgG purified from plasma of patients with hemophilia who did not have inhibitory antibodies.

Results: Significant proteolytic activity was detected in IgG from 13 of 24 inhibitor-positive patients. No hydrolytic activity was detected in control antibodies of IgG from patients without inhibitors. The rate of hydrolysis of factor VIII by purified IgG correlated positively with the factor VIII--neutralizing activity of IgG in plasma (r2=0.67, P=0.029). Principal-component analysis of migration profiles of digestion fragments demonstrated the heterogeneity of the catalytic potential of factor VIII inhibitors among patients.

Conclusions: Proteolysis is a mechanism by which IgG antibodies against factor VIII can inactivate factor VIII.
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http://dx.doi.org/10.1056/NEJMoa011979DOI Listing
February 2002
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