Refractory mixed proliferative and membranous lupus nephritis treated with the topoisomerase I inhibitor irinotecan as add-on therapy.

OBJECTIVE: To evaluate the safety and effects of irinotecan, an inhibitor of topoisomerase I, on refractory lupus nephritis. METHOD: A patient with refractory lupus nephritis under medication with mycophenolic acid, prednisolone, and hydroxychloroquine was treated with add-on low-dose irinotecan. Irinotecan was applied every fourth week at a dose of 50 mg/m2 for four cycles followed by 100 mg/m2 for another eight cycles. Renal function and anti-double-stranded DNA antibodies as well as blood count for evaluation of side effects were assessed during the treatment with irinotecan. RESULTS: Before starting the treatment with irinotecan, a urine protein/creatinine ratio of 1298 mg/g was determined. This declined to 613 mg/g after four cycles with 50 mg/m2 irinotecan and was further reduced to 198 mg/g when using the higher dose of irinotecan. Kidney function remained stable, with creatinine levels of 1.66 mg/dL at the beginning and 1.76 mg/dL at the end of treatment with irinotecan. Importantly, no side effects, such as diarrhoea or neutropenia, were observed during the entire course of treatment. CONCLUSION: Administration of low-dose irinotecan as add-on medication for the treatment of refractory lupus nephritis was shown to be safe. Clinical trials are needed to determine whether irinotecan can improve kidney function and the outcome of patients with refractory lupus nephritis.

Subset size, activation threshold and distribution of autoreactive MZ and FO B cells do not differ in a sex-specific manner in the NZB/W F1 murine lupus model: an experimental mouse study.

OBJECTIVES: Systemic lupus erythematosus (SLE) shows a strong sex bias, preferentially affecting females, and B cells are thought to play a pivotal role in its pathogenesis. Here, we compared the splenic B-cell compartments, their autoreactivity and activation threshold of female and male NZB/W F1, a murine lupus model reflecting the sex bias observed in patients with SLE. METHODS: Autoantibody levels and the amount of autoantibody secreting cells were determined using ELISA and ELISPOT. Flow cytometry and immunofluorescence were applied to analyse the composition of the splenic B-cell pool. Purified follicular (FO) and marginal zone (MZ) B cells were stimulated and the frequency of autoreactive cells was determined. Finally, the proliferative response of FO and MZ B cells upon stimulation was assessed using CFSE dilution and [(3)H]-Thymidin incorporation. RESULTS: Higher autoantibody titres were detected in female NZB/W F1 mice, which were mainly produced in the spleen. Analysing the composition of the splenic B-cell subsets, no differences were found prior to disease development. Autoreactive dsDNA-specific B cells were mostly found in the MZ compartment, while SmD1((83-119))-reactive cells were more evenly distributed. Equal frequencies of autoreactive B cells were found in female and malemice, and no difference in the response to polyclonal stimuli of the cells of both sexes was detected. CONCLUSIONS: No differences in the composition or functionality of splenic B cells were observed that account for the different disease course in both sexes.

CXCR3+CD4+ T cells are enriched in inflamed kidneys and urine and provide a new biomarker for acute nephritis flares in systemic lupus erythematosus patients.

OBJECTIVE: The high frequency of CD4+ T cells in interstitial infiltrates of patients with lupus nephritis suggests a contribution of these cells to local pathogenesis. The aim of this study was to examine the role of CXCR3 and the chemokine CXCL10 in recruiting these cells into the kidney and to determine whether the infiltrating T cells could be monitored in the urine to provide a reliable biomarker for acute lupus nephritis. METHODS: The frequencies of CD3+ T cells, CXCR3+ cells, and CXCL10+ cells were determined by immunohistochemical and immunofluorescence analyses of kidney sections from 18 patients with lupus nephritis. The frequency of CXCR3+CD4+ T cells was determined by flow cytometry of peripheral blood and urine from 38 patients with systemic lupus erythematosus (SLE), and the values were compared with disease activity as determined by the Systemic Lupus Erythematosus Disease Activity Index. RESULTS: In renal biopsy tissues from patients with lupus nephritis, a mean of 63% of the infiltrating cells expressed CXCR3, approximately 60% of them were T cells, and the CXCR3+ cells colocalized with CXCL10-producing cells. In biopsy tissues from SLE patients with acute nephritis, approximately 50% of the urinary CD4+ T cells were CXCR3+, as compared with 22% in the peripheral blood, and the frequency of urinary CXCR3+CD4+ T cells correlated with disease activity. Moreover, the number of urinary CD4+ T cells reflected nephritis activity, and elevation above 800 CD4+ T cells per 100 ml of urine sharply delineated active from inactive nephritis. CONCLUSION: CXCR3+ T cells are recruited into the inflamed kidneys, are enriched in the urine, and are a valuable marker of nephritis activity in SLE. They also present a potential target for future therapies.

The systemic and SmD183-119-autoantigen-specific cytokine memory of Th cells in SLE patients.

OBJECTIVES: The aim of the study was to analyse the cytokine memory of T-cells derived from systemic lupus erythematosus (SLE) patients and healthy donors enriched for autoantigen-specific T-cells by in vitro stimulation with SmD1(83-119), a common autoantigen in SLE. METHODS: Autoreactive CD3+ T-cells derived from 37 SLE patients and 14 healthy donors were enriched by repetitive ex vivo stimulation of peripheral blood mononuclear cells (PBMCs) with SmD1(83-119). For control, PBMCs were stimulated only with interleukin-2 (IL-2). After two rounds of antigenic stimulation, cultures were stimulated with PMA/ionomycin to probe the cytokine memory by intracellular cytokine staining. Frequencies of cytokine-expressing T-cells were analysed and, in SLE patients, compared with disease activities and autoantibody levels. RESULTS: Comparing the cytokine memory in the cultures, SLE patients displayed higher frequencies of tumour necrosis factor-alpha (TNF-alpha)+ T-cells than healthy donors and the frequencies correlated with disease activity. Frequencies of SmD1(83-119)-specific TNF-alpha+ T-cells and of memory T-cells expressing interferon-gamma (IFN-gamma) correlated with serum anti-dsDNA antibody levels. The frequencies of IL-10 expressing SmD1(83-119)-specific T-cells were lower among PBMCs of SLE patients. Relatively higher frequencies of IL-10+ T-cells in SLE patients correlated with low disease activities, and low anti-dsDNA and anti-SmD1(83-119) antibody concentrations in culture supernatants. CONCLUSIONS: The memory of autoreactive SmD1(83-119)-specific and unspecific stimulated peripheral Th cells for re-expression of cytokines is shifted towards more cells expressing TNF-alpha and less IL-10+ cells, when compared SLE patients with normal donors. This shift towards proinflammatory memory effector Th cells correlates with disease severity and humoral autoimmunity.

T cell cytokine imbalance towards production of IFN-gamma and IL-10 in NZB/W F1 lupus-prone mice is associated with autoantibody levels and nephritis.

OBJECTIVE: The role of T cell-derived cytokine production in lupus is poorly understood. We analysed the cytokine production of CD4(+) T cells in the NZB/W F1 mouse strain, the mouse model probably most closely resembling human systemic lupus erythematosus (SLE), and assessed whether a possible shift in the cytokines expressed is associated with age or disease activity. METHODS: We used intracellular cytokine staining and flow cytometry to determine the cytokine expression of splenic CD4(+) T cells for interferon-gamma (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha), interleukin-4 (IL-4) and IL-10. NZB/W F1 mice at different ages spanning 5 to 36 weeks were analysed, healthy Balb/cxNZW F1 (CWF1) mice were used as controls. Serum anti-double-stranded DNA (anti-dsDNA) antibody levels were determined by enzyme-linked immunosorbent assay (ELISA), and proteinuria and plasma creatinine were estimated using commercial test kits. RESULTS: The cytokine profile of CD4(+) T cells was shifted towards T-helper 1 (Th1) cells and the frequencies of Th cells expressing IFN-gamma(+) correlated with age, anti-dsDNA-immunoglobulin G (IgG) titre and proteinuria. An increased percentage of IL-10 producers correlated positively with anti-dsDNA-IgG and proteinuria, and a small gain in IL-4 producers correlated with plasma creatinine. Neither the percentage of IL-10 producers nor IL-4 producers showed a significant correlation with age. There was no significant change observed in the frequency of TNF-alpha T cells. The IFN-gamma/IL-4 ratio demonstrated an increasing shift towards a Th1-type response during disease development that was not present in healthy mouse strains. CONCLUSION: The association between the frequencies of T cells expressing IFN-gamma and IL-10 and clinical findings suggests a key role for these cells in the pathogenesis of lupus.