Background: Lupus nephritis is a leading cause of morbidity and mortality for lupus patients. Although current immunosuppressive regimens have dramatically improved its prognosis they are not uniformly effective and are associated with significant toxicities. Elucidation of the pathogenesis of lupus nephritis is of paramount importance to improve the currently available therapeutic regimens. Production of pathogenic autoantibodies by B lymphocytes to a limited number of nuclear and cytoplasmic antigens and formation of immune complexes are the hallmarks of the disease. Autoantibody production is antigen driven and T cell dependent. In addition to producing autoantibodies, B cells are one of the most effective autoantigen presenting cells and thus may play a more proximal role in the generation of autoimmune processes. Although absolute B lymphocyte numbers are low or normal in patients with SLE, they have an activated phenotype. Little is known, however about the role of different subsets of B cells and their relation to disease activity. Tissue injury in lupus is mediated predominantly by deposition of immune complexes (IC) in tissues such as kidneys. Most pathogenic immunecomplexes contain DNA and anti-DNA antibodies. It has long been postulated that bacterial DNA contributes to induction or perpetuation of SLE. Bacterial DNA contains CpG motifs that activate lymphocytes, macrophages and NK cells. CpG motifs derived from bacterial DNA can boost the production of autoantibodies in animal models and CpG rich DNA is present in the anti-dsDNA/DNA complexes found in lupus sera. The most effective immunosuppressive treatments for lupus nephritis are based on cyclophosphamide. A significant proportion of patients, especially African-Americans, do not respond appropriately to this drug, however. The toxicities associated with cyclophosphamide can be severe, therefore distinguishing between these two groups before starting therapy would be highly desirable. Genetic polymorphisms in several key enzymes of cyclophosphamide metabolism have been described recently and identifying those that may be related to the different responses of individual patients would be helpful in making treatment decisions and spare the risk of toxicity from patients who are unlikely to respond to cyclophosphamide. Objectives: 1. To analyze B lymphocyte subsets and identify subsets that may be related to changes in disease activity 2. To analyze the response of lupus patients to oligodeoxynucleotides containing CpG-motifs. 3. To study the pharmacogenomics of cyclophosphamide therapy. Results: 1. B cell subsets in SLE. In collaboration with the Autoimmunity Branch, we characterized the abnormal B cell pattern of lupus patients and we identified B cell subsets that are not normally found in the peripheral blood. The functional characterization of these cells is in progress. 2. Response of lupus patients to oligodeoxynucleotides containing CpG-motifs: In collaboration with the FDA, we compared the response to CpG motifs of PBMC from lupus patients and normal controls. We found that upregulation of cell surface co-stimulatory molecules and the secretion of interferon-a and IL-6 in response to CpG ODN was reduced in monocytes from SLE patients. Similarly, CpG induced interferon-? secretion of natural killer cells was also reduced. In contrast, the IgM and IL-10 response of B cells to CpG ODN was normal. 3. Pharmacogenomics: we initiated a study to identify genomic markers of cyclophosphamide markers. Several genetic polymorphisms were identified that could influence response to cyclophosphamide. We also identified patients who have been good responders and those who were resistant to or poor responder to cyclophosphamide treatment. The genetic polymorphisms of the candidate genes will be compared between these two groups. Lay Summary: Involvement of the kidneys (nephritis) is the main cause of morbidity and mortality in patients with systemic lupus erythematosus (SLE). In this so called autoimmune disease, the body turns against itself producing antibodies directed to molecules found in body tissues (antigens). In lupus nephritis, complexes of antigens with antibodies (immune complexes) deposit in the kidney and cause inflammation leading to kidney failure if left unchecked. Antibodies are produced by a certain type of immune cells called B lymphocytes. Although patients with active SLE usually have lower number of B lymphocytes in their blood they are more active then in healthy people. We have identified some abnormal B lymphocytes in the blood of patients with active lupus patients that are not found in healthy people. In another study we have shown that immune cells from lupus patients respond differently to synthetic DNA that looks like bacterial DNA then healthy people. We started to explore the genomic basis of why some lupus patient respond well to cyclophosphamide (the most effective therapy for lupus nephritis) while others do not. Finally, we monitor a group of over 300 lupus patients that have participated in studies on the treatment of lupus nephritis in an effort to better understand the course of the disease and the long-term side effects of treatment.
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