Interferon-Driven Immune Dysregulation in Common Variable Immunodeficiency-Associated Villous Atrophy and Norovirus Infection.

PURPOSE: About 15% of patients with common variable immunodeficiency (CVID) develop a small intestinal enteropathy, which resembles celiac disease with regard to histopathology but evolves from a distinct, poorly defined pathogenesis that has been linked in some cases to chronic norovirus (NV) infection. Interferon-driven inflammation is a prominent feature of CVID enteropathy, but it remains unknown how NV infection may contribute. METHODS: Duodenal biopsies of CVID patients, stratified according to the presence of villous atrophy (VA), IgA plasma cells (PCs), and chronic NV infection, were investigated by flow cytometry, multi-epitope-ligand cartography, bulk RNA-sequencing, and RT-qPCR of genes of interest. RESULTS: VA development was connected to the lack of intestinal (IgA+) PC, a T helper 1/T helper 17 cell imbalance, and increased recruitment of granzyme+CD8+ T cells and pro-inflammatory macrophages to the affected site. A mixed interferon type I/III and II signature occurred already in the absence of histopathological changes and increased with the severity of the disease and in the absence of (IgA+) PCs. Chronic NV infection exacerbated this signature when compared to stage-matched NV-negative samples. CONCLUSIONS: Our study suggests that increased IFN signaling and T-cell cytotoxicity are present already in mild and are aggravated in severe stages (VA) of CVID enteropathy. NV infection preempts local high IFN-driven inflammation, usually only seen in VA, at milder disease stages. Thus, revealing the impact of different drivers of the pathological mixed IFN type I/III and II signature may allow for more targeted treatment strategies in CVID enteropathy and supports the goal of viral elimination.

Evaluation of Laboratory and Sonographic Parameters for Detection of Portal Hypertension in Patients with Common Variable Immunodeficiency.

Timely detection of portal hypertension as a manifestation in a subgroup of patients with common variable immunodeficiency (CVID) represents a challenge since it is usually not associated with liver cirrhosis. To identify relevant markers for portal hypertension, we evaluated clinical history, laboratory parameters, and abdominal ultrasound including liver elastography and biomarkers of extracellular matrix formation. Twenty seven (6%) of 479 CVID patients presented with clinically significant portal hypertension as defined by either the presence of esophageal varices or ascites. This manifestation occurred late during the course of the disease (11.8 years after first diagnosis of CVID) and was typically part of a multiorgan disease and associated with a high mortality (11/27 patients died during follow up). The strongest association with portal hypertension was found for splenomegaly with a longitudinal diameter of > 16 cm. Similarly, most patients presented with a liver stiffness measurement (LSM) of above 6.5 kPa, and a LSM above 20 kPa was always indicative of manifest portal hypertension. Additionally, many laboratory parameters including Pro-C4 were significantly altered in patients with portal hypertension without clearly increasing the discriminatory power to detect non-cirrhotic portal hypertension in CVID. Our data suggest that a spleen size above 16 cm and an elevated liver stiffness above 6.5 kPa should prompt further evaluation of portal hypertension and its sequelae, but earlier and better liquid biomarkers of this serious secondary complication in CVID are needed.

Cytokine adsorption in patients with severe COVID-19 pneumonia requiring extracorporeal membrane oxygenation: protocol for a randomised, controlled, open-label intervention, multicentre trial.

INTRODUCTION: Veno-venous extracorporeal membrane oxygenation (V-V ECMO) is a last resort treatment option in patients with severe COVID-19 related acute respiratory distress syndrome (ARDS). Mortality in these critically ill patients is high. Elevated interleukin-6 (IL-6) levels in these severe courses are associated with poor outcome. Extracorporeal cytokine adsorption is an approach to lower elevated IL-6 levels. However, there is no randomised controlled data on the efficacy of cytokine adsorption and its effect on patient outcome in severe COVID-19 related ARDS requiring V-V ECMO support. METHODS AND ANALYSIS: We here report the protocol of a 1:1 randomised, controlled, parallel group, open-label intervention, superiority multicentre trial to evaluate the effect of extracorporeal cytokine adsorption using the CytoSorb device in severe COVID-19 related ARDS treated with V-V ECMO. We hypothesise that extracorporeal cytokine adsorption in these patients is effectively reducing IL-6 levels by 75% or more after 72 hours as compared with the baseline measurement and also reducing time to successful V-V ECMO explantation. We plan to include a total of 80 patients at nine centres in Germany. ETHICS AND DISSEMINATION: The protocol of this study was approved by the ethical committee of the University of Freiburg as the leading institution (EK 285/20). Additional votes will be obtained at all participating centres. TRIAL REGISTRATION NUMBERS: NCT04385771 and DRKS 00021248.

Nuclear antigen-reactive CD4+ T cells expand in active systemic lupus erythematosus, produce effector cytokines, and invade the kidneys.

Systemic lupus erythematosus is a systemic and chronic autoimmune disease characterized by loss of tolerance towards nuclear antigens with autoreactive CD4+ T cells implicated in disease pathogenesis. However, very little is known about their receptor specificity since the detection of human autoantigen specific CD4+ T cells has been extremely challenging. Here we present an analysis of CD4+ T cells reactive to nuclear antigens using two complementary methods: T cell libraries and antigen-reactive T cell enrichment. The frequencies of nuclear antigen specific CD4+ T cells correlated with disease severity. These autoreactive T cells produce effector cytokines such as interferon-γ, interleukin-17, and interleukin-10. Compared to blood, these cells were enriched in the urine of patients with active lupus nephritis, suggesting an infiltration of the inflamed kidneys. Thus, these previously unrecognized characteristics support a role for nuclear antigen-specific CD4+ T cells in systemic lupus erythematosus.

Abatacept Use Is Associated with Steroid Dose Reduction and Improvement in Fatigue and CD4-Dysregulation in CVID Patients with Interstitial Lung Disease.

BACKGROUND: Interstitial lung disease (ILD) represents a severe clinical manifestation of systemic immune dysregulation in patients with common variable immunodeficiency (CVID). Its treatment often requires systemic immunosuppression beyond corticosteroids. OBJECTIVE: To assess the safety and efficacy of abatacept in patients with CVID and ILD. METHODS: Ten patients with confirmed diagnosis of CVID and ILD were included in a single-center, prospective, open-label, nonrandomized trial. Abatacept was administered subcutaneously at a dose of 125 mg/wk for 12 months. RESULTS: Abatacept was a safe treatment for ILD in CVID except for 1 case of bronchopulmonary aspergillosis. One additional patient terminated the trial prematurely because of recurrent bronchitis. Five of 8 patients treated per protocol benefited from the treatment according to American Thoracic Society/European Respiratory Society criteria. The primary end point of the study was met because single breath diffusing capacity of the lung for carbon monoxide was stable (62.5%) or improved (37.5%) in all patients treated per protocol. Although nodules (71%) and ground-glass opacities (57%) improved in most patients, other computed tomography pathologies were less responsive. Quality of life improved in 87.5% and fatigue in 57% of patients. Abatacept treatment was associated with significant improvement in CD4 T-cell dysregulation, signified by a decrease in serum soluble IL-2 receptor levels and of proliferating Ki67+ CD4 T cells, and a recovery of total lymphocytes, CD4+ T cells, and naive CD4 T cells. CONCLUSIONS: Abatacept may represent a treatment option for CVID-associated ILD. This pilot study demonstrated a good safety profile, steroid-sparing effect, positive immune modulation, and overall positive treatment response especially in quality of life. Larger controlled studies are needed to confirm these findings.

IL-2 Therapy Diminishes Renal Inflammation and the Activity of Kidney-Infiltrating CD4+ T Cells in Murine Lupus Nephritis.

An acquired deficiency of interleukin-2 (IL-2) and related disturbances in regulatory T cell (Treg) homeostasis play an important role in the pathogenesis of systemic lupus erythematosus (SLE). Low-dose IL-2 therapy was shown to restore Treg homeostasis in patients with active SLE and its clinical efficacy is currently evaluated in clinical trials. Lupus nephritis (LN), a challenging organ manifestation in SLE, is characterized by the infiltration of pathogenic CD4+ T cells into the inflamed kidney. However, the role of the Treg-IL-2 axis in the pathogenesis of LN and the mode of action of IL-2 therapy in the inflamed kidneys are still poorly understood. Using the (NZB × NZW) F1 mouse model of SLE we studied whether intrarenal Treg are affected by a shortage of IL-2 in comparison with lymphatic organs and whether and how intrarenal T cells and renal inflammation can be influenced by IL-2 therapy. We found that intrarenal Treg show phenotypic signs that are reminiscent of IL-2 deprivation in parallel to a progressive hyperactivity of intrarenal conventional CD4+ T cells (Tcon). Short-term IL-2 treatment of mice with active LN induced an expansion the intrarenal Treg population whereas long-term IL-2 treatment reduced the activity and proliferation of intrarenal Tcon, which was accompanied by a clinical and histological amelioration of LN. The association of these immune pathologies with IL-2 deficiency and their reversibility by IL-2 therapy provides important rationales for an IL-2-based immunotherapy of LN.

Evaluating laboratory criteria for combined immunodeficiency in adult patients diagnosed with common variable immunodeficiency.

Some patients diagnosed with common variable immunodeficiency (CVID) actually suffer from combined immunodeficiency (CID) and therefore may require a different, CID-adapted treatment. Several CD4 T-cell-based criteria have been proposed in the past to identify patients with CID within the cohort of adult CVID patients. In this monocentric study, we used retrospective immunological and clinical data of 238 CVID patients to compare four different proposals of how to define CID among CVID patients. We demonstrate that none of the current definitions sufficiently separates CID from CVID patients and that the relative reduction of naïve CD4 T cells <10% has the highest sensitivity of all tested markers for patients with clinical complications often associated with CID. Thus, a very low percentage of naïve CD4 T cells in any adult CVID patient should raise suspicion, but is not sufficient to define CID among CVID patients.

Low-dose interleukin-2 therapy in refractory systemic lupus erythematosus: an investigator-initiated, single-centre phase 1 and 2a clinical trial.

BACKGROUND: An acquired deficiency of interleukin-2 (IL-2) and related defects in regulatory T cell homeostasis are thought to play a crucial role in the pathogenesis of systemic lupus erythematosus. We hypothesised that reconstitution of regulatory T-cell homoeostasis with low doses of IL-2 would be beneficial to patients with systemic lupus erythematosus. METHODS: In this uncontrolled, phase 1 and 2a trial done in the Department of Rheumatology and Clinical Immunology at Charité-University Medicine Berlin (Berlin, Germany), we assessed the safety and tolerability of low-dose recombinant human IL-2 (aldesleukin) and its effects on regulatory T cells. We recruited patients aged 18-75 years with a confirmed diagnosis of systemic lupus erythematosus and moderate-to-severe disease activity despite previous treatment with at least two conventional therapies. Patients were given four cycles of low-dose aldesleukin daily for 5 days followed by a 9-16 day rest. The primary endpoints were safety and the number of patients who achieved at least a 100% increase in the proportion of CD25hi-expressing cells among circulating CD3 + CD4 + FOXP3 + CD127lo regulatory T cells at day 62 (ie, after four treatment cycles). Secondary endpoints included disease activity as measured by the Safety of Estrogens in Lupus National Assessment-Systemic Lupus Erythematosus Disease Activity Index (SELENA-SLEDAI) and the British Isles Lupus Assessment Group (BILAG) score, disease flares as measured by the SLEDAI flare index, auto-antibody and complement concentrations at day 62. Exploratory endpoints included various cellular and immunological parameters. The trial is registered with WHO/ICTRP, number DRKS00004858. FINDINGS: Between March 31, 2014, and May 27, 2016, 13 patients were screened, of whom ten met eligibility criteria and were enrolled in the trial. Two additional patients were treated between April 1, 2013, and March 11, 2014, in a compassionate use setting. Eleven (92%) of the 12 patients achieved the primary endpoint. 159 adverse events were recorded, 75 (47%) of which were treatment related. Most treatment-related adverse events were transient and mild to moderate (grade 1-2). The most common adverse event was injection-site reaction (20%). No serious adverse events occurred during the treatment period. In ten (83%) of 12 patients, SELENA-SLEDAI scores were lower at day 62 than at baseline, and no severe disease flares were observed during the treatment period. Decreased disease activity correlated with the magnitude of increase in the proportion of activated regulatory T cells. IL-2 treatment resulted in a preferential proliferation of regulatory T cells that retained suppressive capacity. We observed decreases in cells that are involved in the regulation of germinal-centre reactions. INTERPRETATION: Low-dose IL-2 therapy is safe and well tolerated and selectively promotes the expansion of functional regulatory T cells in patients with moderate-to-severe systemic lupus erythematosus. Low-dose IL-2 treatment might also be beneficial in reducing disease activity, although larger trials are needed to address efficacy. FUNDING: German Research Foundation.

[Chronic granulomatous disease as a rare differential diagnosis of inflammatory bowel disease]. / Septische Granulomatose als seltene Differenzialdiagnose einer chronisch-entzündlichen Darmerkrankung.

BACKGROUND: Chronic granulomatous disease is a rare disease with a prevalence of approximately 150 cases in Germany. An intestinal manifestation that mimics chronic inflammatory bowel disease (IBD) has only been described in a few cases. As a result of a deficient superoxide-synthesis, frequent and recurrent infections caused by rare pathogens have been described. We present the case of a 28-year old patient who has been diagnosed with IBD at the age of 2 years. He showed recurrent liver abscesses and the picture of a chronic IBD. METHODS: Clinical and laboratory data was obtained and endoscopic, radiologic and histologic examinations, tests for granulocytic functions as well as a genetic analysis were performed. Literature of the PubMed database and recent literature were analyzed. CASE: Under immunosuppressive therapy, with TNF -blocker Adalimumab followed by therapy with integrin-receptor antagonist Vedolizumab, the patient developed recurrent abscesses of the liver. Those were the result of infection with a sensitive Staphylococcus aureus strain. Colonoscopy showed stenosis of the rectum and some inflammatory activity. Intestinal symptoms were unresponsive to all therapies for IBD. Furthermore, there was a presence of active acne and recurrent liver abscesses due to bacteria not typical for intestinal infections. Consequently, we considered a granulocyte dysfunction as the underlying cause. Diagnosis of a chronic granulomatous disease was confirmed by flow cytometry and oxidative burst test. Genetic analysis showed a homozygote mutation of the p47phox (NCF1) gene located on chromosome 7, which represents the most common autosomal recessive form with 20 - 25 % of cases. RESULTS: In light of recent literature, this case report shows that chronic granulomatous disease should be considered as a differential diagnosis to therapy refractory IBD. This is the case, especially in young patients, when recurrent bacterial lesions caused by intestine-atypical pathogens appear.

T+ NK+ IL-2 Receptor γ Chain Mutation: a Challenging Diagnosis of Atypical Severe Combined Immunodeficiency.

PURPOSE: All reported patients with hypomorphic X-linked severe combined immunodeficiency (X-SCID) due to c.664C>T (p.R222C) mutations in the gene (IL2RG) encoding the common γ chain (γc) have presented with opportunistic infections within the first year of life, despite the presence of nearly normal NK and T cell numbers. Reporting five children of one extended family with hemizygous mutations in IL2RG, we explore potential diagnostic clues and extend our comprehension of the functional impact of this mutation. METHODS: Whole exome sequencing (WES); detailed immune phenotyping; cytokine-induced STAT phosphorylation; B, T, and NK cell activation; and quantification of sjTRECs in five Arab children with c.664C>T (p.R222C) IL2RG mutation. RESULTS: The mean age at clinical presentation with respiratory tract infection or diarrhea was 6.8 (range: 2-12) months. None of the children presented with opportunistic infections. Diagnostic clues were early onset in the first year of life, and a suggestive family history associated with reduced naïve CD4 T cells and absent switched memory B cells. Number and phenotype of NK cells and innate-like lymphocytes were normal. The diagnosis was made by WES and corroborated by absent STAT phosphorylation and reduced functional response after IL-2 and IL-21 stimulation. Four patients underwent successful hematopoietic stem cell transplantation. CONCLUSIONS: As early diagnosis and treatment are important, a high index of suspicion in the diagnosis of c.664C>T (p.R222C) X-SCID is needed. This requires prompt genetic testing by next generation sequencing in order to avoid unnecessary delays in the definite diagnosis since immunological work up may not be discriminating. Assays directly testing cytokine signaling or cytokine-dependent functions are helpful in confirming the functional impact of the identified hypomorphic variants.

Low-dose interleukin-2 selectively corrects regulatory T cell defects in patients with systemic lupus erythematosus.

OBJECTIVES: Defects in regulatory T cell (Treg) biology have been associated with human systemic autoimmune diseases, such as systemic lupus erythematosus (SLE). However, the origin of such Treg defects and their significance in the pathogenesis and treatment of SLE are still poorly understood. METHODS: Peripheral blood mononuclear cells (PBMC) from 61 patients with SLE and 52 healthy donors and in vitro IL-2 stimulated PBMC were characterised by multicolour flow cytometry. Five patients with refractory SLE were treated daily with subcutaneous injections of 1.5 million IU of human IL-2 (aldesleukin) for five consecutive days, and PBMC were analysed by flow cytometry. RESULTS: Patients with SLE develop a progressive homeostatic dysbalance between Treg and conventional CD4+ T cells in correlation with disease activity and in parallel display a substantial reduction of CD25 expression on Treg. These Treg defects resemble hallmarks of IL-2 deficiency and lead to a markedly reduced availability of functionally and metabolically active Treg. In vitro experiments revealed that lack of IL-2 production by CD4+ T cells accounts for the loss of CD25 expression in SLE Treg, which could be selectively reversed by stimulation with low doses of IL-2. Accordingly, treatment of patients with SLE with a low-dose IL-2 regimen selectively corrected Treg defects also in vivo and strongly expanded the Treg population. CONCLUSIONS: Treg defects in patients with SLE are associated with IL-2 deficiency, and can be corrected with low doses of IL-2. The restoration of endogenous mechanisms of immune tolerance by low-dose IL-2 therapy, thus, proposes a selective biological treatment strategy, which directly addresses the pathophysiology in SLE.