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Summary Background B cells produce alloantibodies and activate alloreactive T cells, negatively affecting kidney transplant survival. By contrast, regulatory B cells are associated with transplant tolerance. Immunotherapies are needed that inhibit B-cell effector function, including antibody secretion, while sparing regulators and minimising infection risk. B lymphocyte stimulator (BLyS) is a cytokine that promotes B-cell activation and has not previously been targeted in kidney transplant recipients. We aimed to determine the safety and activity of an anti-BLyS antibody, belimumab, in addition to standard-of-care immunosuppression in adult kidney transplant recipients. We used an experimental medicine study design with multiple secondary and exploratory endpoints to gain further insight into the effect of belimumab on the generation of de-novo IgG and on the regulatory B-cell compartment. Methods We undertook a double-blind, randomised, placebo-controlled phase 2 trial of belimumab, in addition to standard-of-care immunosuppression (basiliximab, mycophenolate mofetil, tacrolimus, and prednisolone) at two centres, Addenbrooke's Hospital, Cambridge, UK, and Guy's and St Thomas' Hospital, London, UK. Participants were eligible if they were aged 18–75 years and receiving a kidney transplant and were planned to receive standard-of-care immunosuppression. Participants were randomly assigned (1:1) to receive either intravenous belimumab 10 mg per kg bodyweight or placebo, given at day 0, 14, and 28, and then every 4 weeks for a total of seven infusions. The co-primary endpoints were safety and change in the concentration of naive B cells from baseline to week 24, both of which were analysed in all patients who received a transplant and at least one dose of drug or placebo (the modified intention-to-treat [mITT] population). This trial has been completed and is registered with ClinicalTrials.gov, NCT01536379, and EudraCT, 2011–006215–56. Findings Between Sept 13, 2013, and Feb 8, 2015, of 303 patients assessed for eligibility, 28 kidney transplant recipients were randomly assigned to receive belimumab (n=14) or placebo (n=14). 25 patients (12 [86%] patients assigned to the belimumab group and 13 [93%] patients assigned to the placebo group) received a transplant and were included in the mITT population. We observed similar proportions of adverse events in the belimumab and placebo groups, including serious infections (one [8%] of 12 in the belimumab group and five [38%] of 13 in the placebo group during the 6-month on-treatment phase; and none in the belimumab group and two [15%] in the placebo group during the 6-month follow-up). In the on-treatment phase, one patient in the placebo group died because of fatal myocardial infarction and acute cardiac failure. The co-primary endpoint of a reduction in naive B cells from baseline to week 24 was not met. Treatment with belimumab did not significantly reduce the number of naive B cells from baseline to week 24 (adjusted mean difference between the belimumab and placebo treatment groups −34·4 cells per μL, 95% CI −109·5 to 40·7). Interpretation Belimumab might be a useful adjunct to standard-of-care immunosuppression in renal transplantation, with no major increased risk of infection and potential beneficial effects on humoral alloimmunity. Funding GlaxoSmithKline.

Background Reactive oxygen species (ROS), generated via the phagocyte NADPH oxidase cytochrome b558, are essential for effective immune responses to common and serious pathogens. The phagocyte NADPH oxidase is a multisubunit protein complex and deficiency of either the membrane bound or cytoplasmic components leads to chronic granulomatous disease, a serious and often fatal illness characterised by recurrent infections and autoimmunity. Moreover, abnormal generation of ROS has been implicated in the pathogenesis of multigenic autoimmune diseases such as systemic lupus erythematosus. Eros (essential for reactive oxygen species), encoded by bc017643 , is a novel transmembrane protein that is highly expressed in the immune system and highly conserved in evolution but has no previously identified function. Eros is an orthologue of the plant protein Ycf4, necessary for expression of proteins of the photosynthetic photosystem 1 complex, an NADPH oxio-reductase complex. We elucidated its role in infection in mice. Methods ROS are essential for host defence against the serious bacterial pathogen Salmonella enterica serovar Typhimurium. We screened individual knockout mice (Wellcome Trust Knockout mouse project) for susceptibility to salmonella infection. Having identified mice deficient in Eros as being highly susceptible to salmonella, we used ex-vivo approaches including reactive oxygen burst assays and western blot, to characterise their defect further. Findings We found that Eros was essential for host defence to infection. Eros was crucial for generating reactive oxygen species through regulation of the essential NADPH oxidase components, gp91 and p22. Eros-deficient mice expressed almost no gp91 and p22 in neutrophils and macrophages secondary to accelerated degradation in the absence of Eros. As a result Eros-deficient mice died rapidly after infection with salmonella or listeria. Eros also regulated the ROS-dependent formation of neutrophil extracellular traps and melanoma metastases. Interpretation We have found a a key role for Eros in regulating host defence. The finding that Eros-deficient mice lack gp91 and p22 at the protein, though not mRNA, level shows how these key components of the reatcive oxygen burst are protected from degradation and furthers our understanding of reactive oxygen burst biology. Eros is highly conserved between mouse and man so it is likely that it also has a crucial role in human immunity. Funding Wellcome Trust, Academy of Medical Sciences starter grant.

Background Calprotectin, an endogenous toll-like receptor 4 (TLR 4) agonist that is expressed in neutrophils, monocytes, and infiltrating macrophages, promotes endothelial activation and transcription of pro-inflammatory cytokines. We investigated calprotectin in renal biopsy samples and serum of patients with anti-neutrophil cytoplasmic antibody associated vasculitis (AAV) and in mice deficient in calprotectin (cal−/−), and we assessed the interaction of calprotectin with macrophages and endothelial cells in vitro. Methods We examined renal biopsy samples with immunohistochemistry. Serum calprotectin levels were measured with ELISA, and cell surface expression with flow cytometry. Accelerated nephrotoxic nephritis experiments were performed on both wild-type (WT) and cal−/− mice. Macrophages were isolated from WT, TLR4−/−, and cal−/− mice, and kidney endothelial cells from WT mice, and stimulated with calprotectin and supernatants harvested. Phagocytosis with opsonised beads was compared between WT and cal−/− macrophages. Findings Patients with active AAV glomerular lesions demonstrated the most calprotectin positivity in renal biopsy samples, sclerotic lesions the least (p Interpretation Serum calprotectin is a potential biomarker in AAV, and may predict relapse. Calprotectin contributes to pathogenesis by promoting leukocyte and endothelial cell activation in a positive feedback loop. Funding UK Medical Research Council.