Inborn error of WAS presenting with SARS-CoV-2-related multisystem inflammatory syndrome in children (preprint)

Multisystem inflammatory syndrome in children (MIS-C) has been reported in patients with inborn errors of immunity (IEI), providing insights into disease pathogenesis. Here, we present the first case of MIS-C in a child affected by Wiskott-Aldrich syndrome (WAS) gene mutation, elucidating underlying predisposing factors and the involved inflammatory pathways. Genetic analysis revealed a frameshift truncating variant in the WAS gene, resulting in WAS protein expression between mild and severe forms, despite a clinical phenotype resembling X-linked thrombocytopenia (XLT). IL-1β secretion by LPS-stimulated peripheral blood mononuclear cells from patient during MIS-C was lower compared to healthy subjects but increased during follow-up. Conversely, the percentage ASC (apoptosis-associated speck-like protein containing a CARD) specks in the patient's circulating monocytes during the acute phase was higher than in healthy subjects. The type I interferon (IFN) score during MIS-C was normal, in contrast to the raised IFN signature measured far from the acute event. This case confirms the association of IEI with MIS-C, potentially linked to delayed immune responses to SARS-CoV-2. The XLT phenotype underlies a subclinical immunodysregulation involving the NLRP3 inflammasome and the type-I IFN response.

Adaptive humoral immune response and cellular immune status in cancer patients and patients under immunosuppression vaccinated against SARS-CoV-2.

BACKGROUND: Patients with cancer and autoimmune diseases are at higher risk of severe COVID-19. They may not develop protective immune responses following vaccination. We investigated patients' cellular and humoral immune response after two COVID-19 vaccine doses. RESEARCH DESIGN AND METHODS: Subjects were stratified into subgroups according to therapy and grade of immunosuppression at time of vaccination. RESULTS: Antibody titers were compared to healthy controls. 32/122 (26%) did not develop detectable antibody titers. Of these, 22 (66.6%) had active therapy. Patients showed significant lower antibody titers compared to controls (median 790 vs. 3923 AU/mL, p = 0.026). Patients with active therapy had significant lower antibody titers compared to those without (median 302 vs. 3952 U/L P < 0.001). B-cell count was lower in the group without antibody titers (median 29.97 vs. 152.8; p = 0.002). 100% of patients under anti-CD20 therapy had no detectable antibody titer, followed by anti-TNF (66%), BTK inhibitors (50%), ruxolitinib (35.5%), TKI (14.2%), and lenalidomide (12.5%). Anti-CD20 therapy, ruxolitinib, BTK inhibitors, and anti-CD38 therapy presented significant lower antibody titers compared to controls. CONCLUSIONS: Patients undergoing therapy for cancer or autoimmune diseases are at higher risk of insufficient humoral immune response following COVID-19 vaccination. Furthermore, alterations in the B-cell compartment correlate with lower antibody titers.

Autoantibodies against Progranulin and IL-1 receptor antagonist due to immunogenic posttranslational isoforms contribute to hyperinflammation in critically ill COVID-19 (preprint)

Hyperinflammation is frequently observed in patients with severe COVID-19. Inadequate and defective IFN type I responses against SARS-CoV-2, associated with autoantibodies in a proportion of patients, lead to severe courses of disease. In addition, hyperactive responses of the humoral immune system have been described. In the current study we investigated a possible role of neutralizing autoantibodies against anti-inflammatory mediators. Plasma from adult patients with severe and critical COVID-19 was screened by ELISA for antibodies against PGRN, IL-1-Ra, IL-10, IL-18BP, IL-22BP, IL-36-Ra, CD40, IFN-α2, IFN-γ, IFN-ω and serpinB1. Autoantibodies were characterized and the antigens were analyzed for immunogenic alterations. In a discovery cohort with severe to critical COVID-19 high titers of PGRN-autoantibodies were detected in 11 of 30 (36.7%), and of IL-1-Ra-autoantibodies in 14 of 30 (46.7%) patients. In a validation cohort of 64 patients with critical COVID-19 high-titer PGRN-Abs were detected in 25 (39%) and IL-1-Ra-Abs in 32 of 64 patients (50%). PGRN-Abs and IL-1-Ra-Abs belonged to IgM and several IgG subclasses. In separate cohorts with non-critical COVID-19, PGRN-Abs and IL-1-Ra-Abs were detected in low frequency (i.e. in < 5% of patients) and at low titers. Neither PGRN- nor IL-1-Ra-Abs were found in 40 healthy controls vaccinated against SARS-CoV-2 or 188 unvaccinated healthy controls. PGRN-Abs were not cross-reactive against SARS-CoV-2 structural proteins nor against IL-1-Ra. Plasma levels of both free PGRN and free IL-1-Ra were significantly decreased in autoantibody-positive patients compared to Ab-negative and non-COVID-19 controls. In vitro PGRN-Abs from patients functionally reduced PGRN-dependent inhibition of TNF-α signaling, and IL-1-Ra-Abs from patients reduced IL-1-Ra- or anakinra-dependent inhibition of IL-1ß signaling. The pSer81 hyperphosphorylated PGRN isoform was exclusively detected in patients with high-titer PGRN-Abs; likewise, a hyperphosphorylated IL-1-Ra isoform was only found in patients with high-titer IL-1-Ra-Abs. Thr111 was identified as the hyperphophorylated amino acid of IL-1-Ra. In longitudinally collected samples hyperphosphorylated isoforms of both PGRN and IL-1-Ra emerged transiently, and preceded the appearance of autoantibodies. In hospitalized patients, the presence of IL-1-Ra-Abs or IL-1-Ra-Abs in combination with PGRN-Abs was associated with a higher morbidity and mortality.To conclude, neutralizing autoantibodies to IL-1-Ra and PGRN occur in a significant portion of patients with critical COVID-19, with a concomitant decrease in circulating free PGRN and IL-1-Ra, indicative of a misdirected, proinflammatory autoimmune response. The break of self-tolerance is likely caused by atypical hyperphosphorylated isoforms of both antigens, whose appearances precede autoantibody induction. Our data suggest that these immunogenic secondary modifications are induced by the SARS-CoV-2-infection itself or the inflammatory environment evoked by the infection and predispose for a critical course of COVID-19.

Autoantibodies against IL-1-receptor-antagonist in multisystem inflammatory syndrome in children (preprint)

Multisystem inflammatory syndrome in children (MIS-C or PIMS) is a rare but serious complication after an infection with SARS-CoV-2. A possible involvement of pathogenetically relevant autoantibodies has been discussed. Recently neutralizing autoantibodies against anti-inflammatory receptor antagonists progranulin (PGRN) and IL-1-receptor antagonist (IL-1-Ra) were discovered in adult patients with critical COVID-19. Plasma of an index case with severe PIMS/MIS-C was analyzed for autoantibodies against IL-1-Ra and PGRN. The study was extended by a case series of 12 additional patients. In addition to ELISA for of antibodies, IL-1-Ra plasma levels were determined and IL-1-Ra was analyzed by Western-blot and isoelectric focusing. Functional activity of the autoantibodies was examined in vitro with IL-1{beta} reporter assays. Antibodies against IL-1-Ra could be detected in 10 of 13 (76.9%) patients with PIMS/MIS-C, but not in controls. In contrast to critical COVID-19 in adults, no IL-1-Ra antibodies of the IgM class were detected in PIMS/MIS-C. IL-1-Ra-antibodies exclusively belonged to IgG1. No antibodies directed against PGRN were detected. Western blots and ELISA showed a concomitant reduction of free IL-1-Ra plasma levels in the presence of IL-1-Ra-antibodies. The antibodies inhibited IL-1-Ra function in IL-1{beta} reporter cell assays. Notably, an additional, hyperphosphorylated, transiently occurring atypical isoform of IL-1-Ra was observed in all IL-1-Ra autoantibody-positive patients. To conclude, IL-1-Ra autoantibodies were observed in high frequency in children with PIMS/MIS-C. They may represent a diagnostic and pathophysiologically relevant marker for PIMS/MIS-C. Their generation is likely to be triggered by an atypical, hyperphosphorylated isoform of IL-1-Ra.

Autoantibodies against Progranulin and IL-1 receptor antagonist in critically ill COVID-19 (preprint)

INTRODUCTION: Hyperinflammation is frequently observed in patients with severe COVID-19. Inadequate and defective IFN type I responses against SARS-CoV-2, caused by autoantibodies in a proportion of patients, lead to severe courses. In addition, hyperactive responses of the humoral immune system have been described so far. RATIONALE: In the current study we investigated a possible role of neutralizing autoantibodies against anti-inflammatory mediators. Plasma from patients with severe and critical COVID-19 was screened by ELISA for antibodies against PGRN, IL-10, IL-18BP, IL-22BP and IL-1-RA. Autoantibodies were characterized and the antigens were analyzed for immunogenic alterations. RESULTS: PGRN-autoantibodies were detected with high titers in 11 of 30 (36.7%), and IL 1-RA-autoantibodies in 14 of 30 (46.7%) patients of a discovery cohort with severe to critical COVID-19. In a validation cohort of 41 patients with critical COVID-19 high-titered PGRN-Abs were detected in 12 (29.3%) and IL-1-RA-Abs in 19 of 41 patients (46.2%). PGRN-Abs and IL-1-RA-Abs belonged to IgM and several IgG subclasses. In separate cohorts with non-critical COVID-19, PGRN-Abs and IL-1-RA-Abs were detected significantly less frequently and at low titers. Neither PGRN- nor IL-1-RA-Abs were found in 40 healthy controls vaccinated against SARS-CoV-2. PGRN-Abs were not cross-reactive against SARS-CoV-2 structural proteins or against IL-1-RA. Plasma levels of both free PGRN and IL-1-RA were significantly decreased in autoantibody-positive patients compared to Ab-negative and non-COVID controls. Functionally, PGRN-Abs from patients reduced PGRN-dependent inhibition of TNF- signaling in vitro. The pSer81 hyperphosphorylated PGRN isoform was exclusively detected in patients with high-titer PGRN-Abs; likewise, a yet unidentified hyperphosphorylated IL-1-RA isoform was only found in patients with high-titer IL-1-RA-Abs. No autoantibodies against IL-10, IL-18BP or IL-22BP were found. CONCLUSION: To conclude, neutralizing autoantibodies to IL-1-RA and PGRN occur in a significant proportion of patients with critical COVID-19, with a concomitant decrease in circulating PGRN and IL-1-RA, which is indicative of a misdirected, proinflammatory autoimmune response. The break of self-tolerance is likely caused by atypical isoforms of both antigens due to hyperphosphorylation. It remains to be determined whether these secondary modifications are induced by the SARS-CoV-2-infection itself, or are preexisting and predispose for a critical course.