Pathological mutations reveal the key role of the cytosolic iRhom2 N-terminus for phosphorylation-independent 14-3-3 interaction and ADAM17 binding, stability, and activity.

The protease ADAM17 plays an important role in inflammation and cancer and is regulated by iRhom2. Mutations in the cytosolic N-terminus of human iRhom2 cause tylosis with oesophageal cancer (TOC). In mice, partial deletion of the N-terminus results in a curly hair phenotype (cub). These pathological consequences are consistent with our findings that iRhom2 is highly expressed in keratinocytes and in oesophageal cancer. Cub and TOC are associated with hyperactivation of ADAM17-dependent EGFR signalling. However, the underlying molecular mechanisms are not understood. We have identified a non-canonical, phosphorylation-independent 14-3-3 interaction site that encompasses all known TOC mutations. Disruption of this site dysregulates ADAM17 activity. The larger cub deletion also includes the TOC site and thus also dysregulated ADAM17 activity. The cub deletion, but not the TOC mutation, also causes severe reductions in stimulated shedding, binding, and stability of ADAM17, demonstrating the presence of additional regulatory sites in the N-terminus of iRhom2. Overall, this study contrasts the TOC and cub mutations, illustrates their different molecular consequences, and reveals important key functions of the iRhom2 N-terminus in regulating ADAM17.

Trajectories and predictive significance of inflammatory parameters for clinical outcome in COVID-19 patients treated with tocilizumab.

PURPOSE: The IL-6 receptor inhibitor tocilizumab reduces mortality and morbidity in severe cases of COVID-19 through its effects on hyperinflammation and was approved as adjuvant therapy. Since tocilizumab changes the levels of inflammatory markers, we aimed to describe these changes in patients treated with tocilizumab, analyse their value in predicting death and bacterial superinfection and determine their influence on mortality rates. METHODS: A retrospective analysis of 76 patients who were treated with tocilizumab for severe COVID-19 in 2020 and 2021 was conducted. Inflammatory markers (IL-6, C-reactive protein (CRP), procalcitonin) were documented before and up to seven days after tocilizumab administration. RESULTS: The overall mortality was 25% and 53.8% in patients who required invasive respiratory support. Deceased patients had higher baseline IL-6 (p = 0.026) and peak IL-6 levels after tocilizumab vs those who survived (p < 0.0001). A peak IL-6 value > 1000 pg/dl after tocilizumab administration was a good predictor of mortality (AUC = 0.812). Of the deceased patients 41.1% had a renewed CRP increase after an initial decrease following tocilizumab administration, compared to 7.1% of the surviving patients (p = 0.0011). Documented bacterial superinfections were observed in 35.5% (27/76) of patients, of whom 48.1% (13/27) died. CONCLUSION: CRP-decline and IL-6 increase after tocilizumab treatment occurs regularly. An increase of IL-6 levels exceeding tenfold of baseline IL-6 levels, an absolute peak of 1000 pg/ml or a renewed increase of CRP are associated with higher mortality. Suppressed CRP synthesis can impede the diagnosis of bacterial superinfections, thus increasing the risk for complications.

Rapid Selection of Sotrovimab Escape Variants in Severe Acute Respiratory Syndrome Coronavirus 2 Omicron-Infected Immunocompromised Patients.

BACKGROUND: Monoclonal antibodies (mAbs) that target severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are predominantly less effective against Omicron variants. Immunocompromised patients often experience prolonged viral shedding, resulting in an increased risk of viral escape. METHODS: In an observational, prospective cohort, 57 patients infected with Omicron variants who received sotrovimab alone or in combination with remdesivir were followed. The study end points were a decrease in SARS-CoV-2 RNA <106 copies/mL in nasopharyngeal swabs at day 21 and the emergence of escape mutations at days 7, 14, and 21 after sotrovimab administration. All SARS-CoV-2 samples were analyzed using whole-genome sequencing. Individual variants within the quasispecies were subsequently quantified and further characterized using a pseudovirus neutralization assay. RESULTS: The majority of patients (43 of 57, 75.4%) were immunodeficient, predominantly due to immunosuppression after organ transplantation or hematologic malignancies. Infections by Omicron/BA.1 comprised 82.5%, while 17.5% were infected by Omicron/BA.2. Twenty-one days after sotrovimab administration, 12 of 43 (27.9%) immunodeficient patients had prolonged viral shedding compared with 1 of 14 (7.1%) immunocompetent patients (P = .011). Viral spike protein mutations, some specific for Omicron (e.g., P337S and/or E340D/V), emerged in 14 of 43 (32.6%) immunodeficient patients, substantially reducing sensitivity to sotrovimab in a pseudovirus neutralization assay. Combination therapy with remdesivir significantly reduced emergence of escape variants. CONCLUSIONS: Immunocompromised patients face a considerable risk of prolonged viral shedding and emergence of escape mutations after early therapy with sotrovimab. These findings underscore the importance of careful monitoring and the need for dedicated clinical trials in this patient population.

Early combination therapy of COVID-19 in high-risk patients.

PURPOSE: Prolonged shedding of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been observed in immunocompromised hosts. Early monotherapy with direct-acting antivirals or monoclonal antibodies, as recommended by the international guidelines, does not prevent this with certainty. Dual therapies may therefore have a synergistic effect. METHODS: This retrospective, multicentre study compared treatment strategies for corona virus disease-19 (COVID-19) with combinations of nirmatrelvir/ritonavir, remdesivir, molnupiravir, and/ or mABs during the Omicron surge. Co-primary endpoints were prolonged viral shedding (≥ 106 copies/ml at day 21 after treatment initiation) and days with SARS-CoV-2 viral load ≥ 106 copies/ml. Therapeutic strategies and risk groups were compared using odds ratios and Fisher's tests or Kaplan-Meier analysis and long-rank tests. Multivariable regression analysis was performed. RESULTS: 144 patients were included with a median duration of SARS-CoV-2 viral load ≥ 106 copies/ml of 8.0 days (IQR 6.0-15.3). Underlying haematological malignancies (HM) (p = 0.03) and treatment initiation later than five days after diagnosis (p < 0.01) were significantly associated with longer viral shedding. Prolonged viral shedding was observed in 14.6% (n = 21/144), particularly in patients with underlying HM (OR 3.5; 95% CI 1.2-9.9; p = 0.02). Clinical courses of COVID-19 were mild to moderate with only few adverse effects potentially related to combination treatment. CONCLUSION: Early combination treatment of COVID-19 effectively prevented prolonged viral shedding in 85.6% of cases. Considering the rapid viral clearance rates and low toxicity, individualized dual therapy approaches may be beneficial in high-risk patients.

Status update on iRhom and ADAM17: It's still complicated.

Several membrane-bound proteins with a single transmembrane domain are subjected to limited proteolysis at the cell surface. This cleavage leads to the release of their biologically active ectodomains, which can trigger different signalling pathways. In many cases, this ectodomain shedding is mediated by members of the family of a disintegrins and metalloproteinases (ADAMs). ADAM17 in particular is responsible for the cleavage of several proinflammatory mediators, growth factors, receptors and adhesion molecules. Due to its direct involvement in the release of these signalling molecules, ADAM17 can be positively and negatively involved in various physiological processes as well as in inflammatory, fibrotic and malignant pathologies. This central role of ADAM17 in a variety of processes requires strict multi-level regulation, including phosphorylation, various conformational changes and endogenous inhibitors. Recent research has shown that an early, crucial control mechanism is interaction with certain adapter proteins identified as iRhom1 and iRhom2, which are pseudoproteases of the rhomboid superfamily. Thus, iRhoms have also a decisive influence on physiological and pathophysiological signalling processes regulated by ADAM17. Their characteristic gene expression profiles, the specific consequences of gene knockouts and finally the occurrence of disease-associated mutations suggest that iRhom1 and iRhom2 undergo different gene regulation in order to fulfil their function in different cell types and are therefore only partially redundant. Therefore, there is not only interest in ADAM17, but also in iRhoms as therapeutic targets. However, to exploit the therapeutic potential, the regulation of ADAM17 activity and in particular its interaction with iRhoms must be well understood.