ObjectivesTo characterise the clinical features, immune manifestations and molecular mechanisms in a recently described autoinflammatory disease caused by mutations in TRNT1, a tRNA processing enzyme, and to explore the use of cytokine inhibitors in suppressing the inflammatory phenotype.MethodsWe studied nine patients with biallelic mutations in TRNT1 and the syndrome of congenital sideroblastic anaemia with immunodeficiency, fevers and developmental delay (SIFD). Genetic studies included whole exome sequencing (WES) and candidate gene screening. Patients’ primary cells were used for deep RNA and tRNA sequencing, cytokine profiling, immunophenotyping, immunoblotting and electron microscopy (EM).ResultsWe identified eight mutations in these nine patients, three of which have not been previously associated with SIFD. Three patients died in early childhood. Inflammatory cytokines, mainly interleukin (IL)-6, interferon gamma (IFN-γ) and IFN-induced cytokines were elevated in the serum, whereas tumour necrosis factor (TNF) and IL-1β were present in tissue biopsies of patients with active inflammatory disease. Deep tRNA sequencing of patients’ fibroblasts showed significant deficiency of mature cytosolic tRNAs. EM of bone marrow and skin biopsy samples revealed striking abnormalities across all cell types and a mix of necrotic and normal-appearing cells. By immunoprecipitation, we found evidence for dysregulation in protein clearance pathways. In 4/4 patients, treatment with a TNF inhibitor suppressed inflammation, reduced the need for blood transfusions and improved growth.ConclusionsMutations of TRNT1 lead to a severe and often fatal syndrome, linking protein homeostasis and autoinflammation. Molecular diagnosis in early life will be crucial for initiating anti-TNF therapy, which might prevent some of the severe disease consequences.

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Purpose Secondary bacterial infections are a common complication of influenza. Innate immune host defenses appear to be impaired following influenza, leading to susceptibility to subsequent bacterial infections. Alternatively activated macrophages (AAM) in the lungs may play a critical role in eliciting the hypersusceptibility to secondary bacterial pneumonia. Methods C57BL6 mice were challenged with sublethal doses of the mouse-adapted A/PR/8/34 (PR8) influenza virus or saline and allowed to recover. At complete recovery (day 14), mice were re-challenged with sublethal doses of Streptococcus pneumoniae serotype 3 (Sp3). Results PR8-recovered mice developed a rapidly fatal pulmonary infection to a 100-fold sublethal pneumococcal challenge, whereas PR8-naive mice demonstrated no mortality or illness. The cytokines which induce AAM (IL-4 and IL-13) and the expression of genes associated with AAM (Arginase-1, FIZZ1, and YM1) were elevated after PR8 infection. Flow cytometry suggests that alveolar macrophages demonstrate the AAM-phenotype, as indicated by MGL-1 and MHCII expression, in response to PR8 infection. Recovery from PR8 was associated with blunted cytokine responses to TLR ligands. Conclusions The mechanisms of immune regulation during recovery from influenza are being elucidated. We provide evidence that pulmonary AAM are induced during influenza infection and may contribute to the elicitation of hypersusceptibility to a secondary bacterial infection.

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BACKGROUND Sarilumab (anti-interleukin-6 receptor-alpha; monoclonal antibody) may attenuate the inflammatory response in Covid-19. METHODS We performed an adaptive, phase 2/3, randomized, double-blind, placebo-controlled trial of intravenous sarilumab 200 mg or 400 mg in adults hospitalized with Covid-19. The phase 3 primary analysis population (cohort 1) was patients with critical Covid-19 receiving mechanical ventilation (MV) randomized to sarilumab 400 mg or placebo. The primary end point for phase 3 was the proportion of patients with ≥1-point improvement in clinical status from baseline to day 22. RESULTS Four-hundred fifty-seven (457) and 1365 patients were randomized and treated in phases 2 and 3, respectively. Among phase 3 critical patients receiving MV (n=289; 34.3% on corticosteroids), the proportion with ≥1-point improvement in clinical status (alive not receiving MV) at day 22 was 43.2% in sarilumab 400 mg and 35.5% in placebo (risk difference [RD] +7.5%; 95% confidence interval [CI], − 7.4 to 21.3; P=0.3261), representing a relative risk improvement of 21.7%. Day 29 all-cause mortality was 36.4% in sarilumab 400 mg versus 41.9% in placebo (RD − 5.5%; 95% CI, −20.2 to 8.7; relative risk reduction 13.3%). In post hoc analyses pooling phase 2 and 3 critical patients receiving MV, the hazard ratio (HR) for death in sarilumab 400 mg compared with placebo was 0.76 (95% CI, 0.51 to 1.13) overall, improving to 0.49 (95% CI, 0.25 to 0.94) in patients receiving corticosteroids at baseline. CONCLUSION In hospitalized patients with Covid-19 receiving MV, numerical benefits with sarilumab did not achieve statistical significance, but benefit may be greater in patients receiving corticosteroids. A larger study is required to confirm this observed numerical benefit.

Background Open-label platform trials and a prospective meta-analysis suggest efficacy of anti–IL-6R therapies in hospitalized patients with COVID-19 receiving corticosteroids. This study evaluated the efficacy and safety of sarilumab, an anti–IL-6R monoclonal antibody, in the treatment of hospitalized patients with COVID-19. Methods In this adaptive, phase 2/3, randomized, double-blind, placebo-controlled trial, adults hospitalized with COVID-19 (ClinicalTrials.gov: NCT04315298) received intravenous sarilumab or placebo. The phase 3 primary analysis population included patients with critical COVID-19 receiving mechanical ventilation randomized to sarilumab 400 mg or placebo. The primary outcome was proportion of patients with ≥1-point improvement in clinical status from baseline to day 22. Results There were 457 and 1365 patients randomized and treated in phases 2 and 3, respectively. In phase 3, patients with critical COVID-19 receiving mechanical ventilation (n = 298; 28.2% on corticosteroids), the proportion with ≥1-point improvement in clinical status (alive, not receiving mechanical ventilation) at day 22 was 43.2% in sarilumab and 35.5% in placebo (risk difference +7.5%; 95% CI, –7.4 to 21.3; P = .3261), a relative risk improvement of 21.7%. In post-hoc analyses pooling phase 2 and 3 critical patients receiving mechanical ventilation, the hazard ratio for death in sarilumab versus placebo was 0.76 (95% CI, .51–1.13) overall and 0.49 (95% CI, .25–.94) in patients receiving corticosteroids at baseline. Conclusions This study did not establish the efficacy of sarilumab in hospitalized patients with severe/critical COVID-19. Post-hoc analyses were consistent with other studies that found a benefit of sarilumab in patients receiving corticosteroids.

Purpose of reviewNext-generation sequencing is revolutionizing the molecular taxonomy of human disease. Recent studies of patients with unexplained autoinflammatory disorders reveal germline genetic mutations that target important regulators of innate immunity.Recent findingsWhole-exome analyses of previously undiagnosed patients have catalyzed the recognition of two new disease genes. First, a phenotypic spectrum, including livedo racemosa, fever with early-onset stroke, polyarteritis nodosa, and Sneddon syndrome, is caused by loss-of-function mutations in cat eye syndrome chromosome region, candidate 1 (CECR1), encoding adenosine deaminase 2. Adenosine deaminase 2 is a secreted protein expressed primarily in myeloid cells, and a regulator of macrophage differentiation and endothelial development. Disease-associated mutations impair anti-inflammatory M2 macrophage differentiation. Second, patients presenting with cold-induced urticaria, granulomatous rash, autoantibodies, and common variable immunodeficiency, or with blistering skin lesions, bronchiolitis, enterocolitis, ocular inflammation, and mild immunodeficiency harbor distinct mutations in phospholipase Cγ2, encoding a signaling molecule expressed in natural killer cells, mast cells, and B lymphocytes. These mutations inhibit the function of a phospholipase Cγ2 autoinhibitory domain, causing increased or constitutive signaling.SummaryThese findings underscore the power of next-generation sequencing, demonstrating how the primary deficiency of key molecular regulators or even regulatory motifs may lead to autoinflammation, and suggesting a possible role for cat eye syndrome chromosome region, candidate 1 and phospholipase Cγ2 in common diseases.

Background Two thirds of the 1700 patients seen at our NIH clinic for autoinflammatory diseases do not have a genetic diagnosis. Whole exome sequencing permits analysis of most of the protein coding regions of the human genome. Methods With the use of whole exome sequencing and candidate gene screening, we identified five children from four unrelated families, who had unexplained autoinflammatory disease and shared mutations in one common gene. One family from Saudi Arabia was consanguineous with two affected daughters. The second family of mixed Czech and British background had one affected boy. The third and fourth families were of mixed European ancestry from the United States and each family had one affected daughter. We performed additional experiments in patients samples including flow cytometry, immunophenotyping, cytokine profiling, mitochondria related function and ribosomal assembly assays. Protein function was studied with morpholino knockdowns in zebrafish embryos. Results All patients carried missense recessive mutations in one common gene, the TRNT1 (tRNA Nucleotidyl Transferase, CCA-Adding, 1), on chromosome 3. The two affected Saudi Arabian sisters were homozygous for a p. H215R missense mutation, while the other three children were compound heterozygous for a missense mutation, p. I223T or p. R99W, and one shared mutation p. D163V. The p. H215R mutation was not found in any public database neither in 1061 Arab control DNA samples. From the three Caucasian mutations, the p. R99W was novel whereas the p. I223T and p. D163V were found at a very low allele frequency (<0.001) at the NHLBI exomedatabase. All mutations affect highly conserved amino acid residues and are predicted to be damaging to the protein function. All children had recurrent episodes of high fevers with negative sepsis work up that occurred in association with microcytic anaemia, and a spectrum of multisystem features. Neurologic involvement ranged from mild developmental delay to nystagmus, hypotonia, optic nerve atrophy, and sensorineural hearing loss. Other variables manifestations include dysmorphic features, musculoskeletal and gastrointestinal symptoms, B cell immunodeficiency and hypogammaglobulinemia. Studies performed so far, point towards a maturation defect of the B cell lineage in the bone marrow, as a possible cause of the observed immunodeficiency. Preliminary data from cytokine analysis in two patients have shown elevated levels of the proinflammatory cytokines interleukin 6 and type 1interferon, suggesting possible therapeutic targets. Knockdown of the zebrafish TRNT1 homologue caused hydrocephaly, defects in tail development, anaemia and a reduction in the number of hair cells present in the lateral line, that has function resembling human inner ear. Conclusion The CCA-adding TRNT1 enzyme catalyses the addition of the CCA terminus to the 3 prime end of all tRNAs precursors, a step that is essential for tRNA aminoacylation and protein synthesis. The discovery that missense mutations in this essential and ubiqu...