Deep Phenotyping of Urinary Leukocytes by Mass Cytometry Reveals a Leukocyte Signature for Early and Non-Invasive Prediction of Response to Treatment in Active Lupus Nephritis.

Non-invasive biomarkers are necessary for diagnosis and monitoring disease activity in lupus nephritis (LN) to circumvent risks and limitations of renal biopsies. To identify new non-invasive cellular biomarkers in the urine sediment of LN patients, which may reflect kidney inflammation and can be used to predict treatment outcome, we performed in-depth urinary immune cell profiling by mass cytometry. We established a mass cytometric workflow to comparatively analyze the cellular composition of urine and peripheral blood (PB) in 13 patients with systemic lupus erythematosus (SLE) with active, biopsy-proven proliferative LN. Clinical and laboratory data were collected at the time of sampling and 6 months after induction of therapy in order to evaluate the clinical response of each patient. Six patients with different acute inflammatory renal diseases were included as comparison group. Leukocyte phenotypes and composition differed significantly between urine and paired PB samples. In urine, neutrophils and monocytes/macrophages were identified as the most prominent cell populations comprising together about 30%-83% of nucleated cells, while T and B lymphocytes, eosinophils, and natural killer (NK) cells were detectable at frequencies of <10% each. The majority of urinary T cells showed phenotypical characteristics of activated effector memory T cells (EM) as indicated by the co-expression of CD38 and CD69 - a phenotype that was not detectable in PB. Kidney inflammation was also reflected by tissue-imprinted macrophages, which phenotypically differed from PB monocytes by an increased expression of HLA-DR and CD11c. The presence of activated urinary T cells and macrophages could be used for differential diagnosis of proliferative LN forms and other renal pathologies. Most interestingly, the amount of EM in the urine sediment could be used as a biomarker to stratify LN patients in terms of response to induction therapy. Deep immunophenotypic profiling of urinary cells in LN allowed us to identify a signature of activated T cells and macrophages, which appear to reflect leukocytic infiltrates in the kidney. This explorative study has not only confirmed but also extended the knowledge about urinary cells as a future non-invasive biomarker platform for diagnosis and precision medicine in inflammatory renal diseases.

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.

Observational study and brief analysis of diagnostic criteria in relapsing polychondritis.

Observational study of patients with relapsing polychondritis (RPC) and brief evaluation of widely used diagnostic criteria. A retrospective analysis of 18 patients with RPC treated in the past 15 years at the Charté-Universitätsmedizin Berlin was performed. Three different diagnostic criteria were applied to our cohort. Sensitivities of diagnostic criteria of McAdam et al., Damiani and Levine and Michet et al. were calculated as well as the 5- and 10-year survival. Analysis of diagnostic criteria revealed a sensitivity of 88.9% using Damiani and Levine criteria, 66.7% for Michet et al. and 50% for McAdam et al., respectively. Modifying the criteria of Michet et al. increases the sensitivity to 88.9%. The 5- and 10-year survival were 100 and 90.9%, respectively. Current diagnostic criteria in RPC should be reappraised covering the diversity of clinical findings with the aim to improve clinical care and research in RPC.