Mycophenolate mofetil as induction and maintenance therapy for lupus nephritis: rationale and protocol for the randomized, controlled Aspreva Lupus Management Study (ALMS).

The Phase III Aspreva Lupus Management Study (ALMS) will investigate mycophenolate mofetil (MMF) therapy for lupus nephritis (LN). Eligibility criteria include: 12-75 years of age; diagnosis of systemic lupus erythematosus according to revised American College of Rheumatology criteria; and biopsy-demonstrated LN (Class III-V). Randomized patients will receive open-label induction therapy with MMF or cyclophosphamide in combination with corticosteroids for 24 weeks. The primary efficacy endpoint is treatment response [decreased proteinuria and stabilized (within 25% of baseline) or improved serum creatinine level]. Patients achieving response or complete remission (normalization of all parameters) will be rerandomized to double-blind, placebo-controlled maintenance treatment with MMF or azathioprine, both plus corticosteroids. The maintenance phase primary endpoint is time to treatment failure. To detect a 15% rate improvement in the MMF group compared with cyclophosphamide, and to provide 90% power, a total of 358 patients will be required for the induction phase. On the basis of a projected 278 rerandomized patients, the maintenance phase will have 90% power to detect a difference between treatment groups assuming azathioprine and MMF three-year failure rates of 59.5% and 40.7%, respectively. Aspreva Lupus Management Study may provide invaluable comparative data on the efficacy and safety of MMF as LN induction and maintenance therapy.

Block and tackle: CTLA4Ig takes on lupus.

Blockade of antigen nonspecific costimulatory signals is a promising approach for the treatment of autoimmune diseases including systemic lupus erythematosus (SLE). CTLA4Ig, an antagonist of the CD28/B7 costimulatory interaction, effectively prevents SLE onset in several murine models and, when used in combination with cyclophosphamide, can induce remission of active SLE nephritis. In this review we describe the known mechanisms of action of CTLA4Ig both in normal immunity and in autoimmune disease models and address issues about its activity that still need to be resolved. We discuss the preclinical use of CTLA4Ig in murine SLE models and the rationale for a clinical trial in SLE patients.

Treatment of autoimmune diseases by inhibition of T-cell costimulation.

Abstract Advances in our understanding of the mechanisms involved in immune activation and immune tolerance have laid the foundation for the development of new strategies for treating autoimmune diseases. In particular, the dissection of the two-signal process of T-cell activation has identified distinct targets that may provide a means of blocking pathological autoimmune responses without causing sustained blockade of protective immune responses. These strategies have shown great promise in animal models for autoimmune diseases, and they are currently the focus of clinical investigation in several autoimmune diseases of humans.

The immune tolerance network and rheumatic disease: immune tolerance comes to the clinic.

The development of effective, new, biologically based therapies for RA has created real excitement and justifiable optimism in recent years among rheumatologists and among patients with rheumatic diseases. Recent advances in our understanding of the mechanisms of immune activation and immune tolerance provide further cause for optimism. Against this background, the establishment of the ITN is an important step. However, significant hurdles remain to be cleared. First, despite dramatic scientific progress, restoration of immune tolerance in the face of an established autoimmune response is still an elusive goal, even in the laboratory. Not only does the ITN face this fundamental scientific challenge, but it also faces daunting practical and political challenges. For example, can the ITN influence the research agenda of the pharmaceutical and biotechnology industries? This question and other important questions will only be answered as the ITN matures. Autoimmune disease, although individually uncommon, affects more than 2% of Americans. The rheumatologist is especially aware of the devastating potential of autoimmune diseases. If the ITN succeeds in linking basic research into the mechanisms of autoimmunity with clinical trials of promising new therapies, it can be expected to play a critical role in advancing the practice of clinical rheumatology.

Phase I clinical trial of a monoclonal antibody against CD40-ligand (IDEC-131) in patients with systemic lupus erythematosus.

OBJECTIVE: To investigate the safety and pharmacology of a humanized monoclonal antibody against CD40-ligand (IDEC-131) in patients with systemic lupus erythematosus (SLE). METHODS: Cohorts of 3 to 5 patients with symptomatic lupus each received 0.05, 0.25, 1.0, 5.0, or 15.0 mg/kg of IDEC-131 as a single intravenous infusion. Patients were followed for 3 months to evaluate toxicity and pharmacokinetics. RESULTS: This phase I, single dose, dose-escalating study was conducted in 23 patients at a single institution. All patients experienced at least 1 adverse event (AE) during a 3 month followup period, although 58 AE in 17 patients were considered possibly or probably related or of unknown relationship to treatment. No dose relationship in the distribution of AE was apparent. No infusion related cytokine-release syndrome was observed; no infusions were interrupted, and all patients completed treatment. Eight mild (grade 1 or 2) infections were reported in 8 patients. All infections were considered unrelated to drug administration and all resolved uneventfully. No patient developed detectable antibodies to IDEC-131. Flow cytometry revealed no apparent treatment related depletion of lymphocyte subsets. Pharmacokinetic analysis indicated that the maximum serum concentration and the area under the concentration curve of IDEC-131 were proportional to the dose administered. At doses between 1.0 and 15.0 mg/kg, the serum half-life ranged from 299 to 320 h. Efficacy was not formally evaluated in this single dose study. CONCLUSION: IDEC-131 (humanized Mab against CD40L) administered in a single intravenous infusion at doses of 0.05-15.0 mg/kg is safe and well tolerated in patients with SLE.

Cutting edge: reversal of murine lupus nephritis with CTLA4Ig and cyclophosphamide.

Cyclophosphamide (CTX) prevents progression of nephritis and prolongs survival in (NZB x NZW)F(1) (B/W) mice and is used to treat humans with lupus nephritis. To compare the efficacy of CTLA4Ig with CTX and determine whether there is an incremental benefit to combining CTLA4Ig with CTX, we treated B/W mice with CTX, CTLA4Ig, or both agents. In mice with mild renal disease, treatment delayed the onset of proteinuria and prolonged survival in all groups. In mice with advanced renal disease, treatment with both agents reduced proteinuria in 71% of mice, whereas mice treated with either agent alone had no such improvement. Survival was also markedly improved among mice treated with both agents. Thus, combination treatment with CTX and CTLA4Ig is more effective than either agent alone in reducing renal disease and prolonging survival of B/W mice with advanced nephritis. This striking reversal of proteinuria is unprecedented in animal models of SLE.

Nephritogenic cytokines and disease in MRL-Fas(lpr) kidneys are dependent on multiple T-cell subsets.

BACKGROUND: Renal parenchymal cells produce cytokines, colony-stimulating factor-1 (CSF-1), granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor-alpha (TNF-alpha), which recruit autoreactive T cells and, in turn, elicit renal injury in MRL-Fas(lpr) mice. METHODS: To determine whether select T-cell populations regulate intrarenal nephritogenic cytokines (CSF-1, GM-CSF, and TNF-alpha) and renal disease, we compared MRL-Fas(lpr) mice that are genetically deficient in T-cell receptor (TCR) alpha beta T cells, CD4 T cells, and major histocompatibility complex class I (MHC class I), lacking CD8 and double negative (DN) T cells, with wild-type mice. To identify the T cells instrumental in downstream (effector) events, we delivered CSF-1 or GM-CSF into the kidney via gene transfer in these select T-cell-deficient and wild-type strains. RESULTS: Intrarenal CSF-1, GM-CSF, and TNF-alpha were absent or dramatically reduced in TCR alpha beta, CD4, and class I-deficient MRL-Fas(lpr) strains as compared with wild-type mice. In addition, the decrease in CSF-1, GM-CSF, and TNF-alpha was associated with a reduced kidney leukocytic infiltrates and spontaneous autoimmune nephritis. Intrarenal ex vivo retroviral gene transfer of CSF-1 and GM-CSF failed to elicit nephritis in these T-cell-deficient MRL strains (TCR alpha beta, CD4, CD8/DN) as compared with wild-type mice. CONCLUSIONS: Multiple T-cell populations initiate renal disease by increasing intrarenal nephritogenic cytokines, CSF-1, GM-CSF, and TNF-alpha. CSF-1 and GM-CSF recruit additional CD4 and CD8 and DN T cells, which augment downstream events, resulting in progressive autoimmune renal disease. We suggest that MRL-Fas(lpr) kidney disease is driven by a T-cell amplification feedback loop dependent on multiple T-cell populations.

The effect on immunoglobulin glycosylation of altering in vivo production of immunoglobulin G.

The effect on murine immunoglobulin G (IgG) glycosylation of altering IgG production in vivo was assessed in interleukin (IL)-6 transgenic and CD4 knockout mice. C57BL/6 mice carrying the IL-6 transgene showed increased levels of circulating IgG. This was associated with decreased levels of galactose on the IgG oligosaccharides. No decrease in beta4-galactosyltransferase mRNA or in enzyme activity was seen in IL-6 transgenic mice. MRL-lpr/lpr mice normally have elevated levels of circulating IgG, again accompanied by decreased levels of IgG galactose. Disruption of the CD4 gene in MRL-lpr/lpr mice led to a substantial decrease in the concentration of circulating IgG, but IgG galactose levels remained low. Thus, an enforced decrease in IgG levels in the lymphoproliferative MRL-lpr/lpr mice did not alter the percentage of agalactosyl IgG in these mice, suggesting that agalactosyl IgG production is not simply caused by excessive IgG synthesis leading to an insufficient transit time in the trans-Golgi, but rather to a molecular defect in the interaction between galactosyltransferase and the immunoglobulin heavy chain.

Effects of anti-B7 monoclonal antibodies on humoral immune responses.

The costimulatory interaction between CD28 on T cells and B7-related molecules on antigen presenting cells plays an important role in a broad range of functions of the immune system, including protective immunity, tolerance induction, allograft rejection, and the development of autoimmune diseases. Monoclonal antibodies to B7-1 and B7-2 have been used in vivo to examine the mechanisms underlying these processes and to evaluate costimulation antagonism as an approach to treatment of chronic autoimmune diseases. To determine whether anti-B7 mAb might elicit, or inhibit, a host immune response that could influence the effects of these antibodies in vivo, we assessed the immune response to rat anti-B7-1 and anti-B7-2 mAb in healthy (BALB/c) mice and in lupus-prone NZB/NZW F1(B/W) mice. In BALB/c mice, low doses (1-10 microg) of mAb to B7-1 and mAb to B7-2 elicited brisk immune responses that occurred earlier and were significantly greater than the immune response to an isotype-matched control rat mAb to ovalbumin. In contrast, at higher doses (100-500 microg), both anti-B7 mAb, but not the control mAb, blocked the mouse anti-rat response. No such blockade occurred in B/W mice, who generated a significant mouse anti-rat response even at very high doses of anti-B7 mAb (1,000-4,000 microg). Blockade of the immune response to the anti-B7 mAb in BALB/c mice apparently did not reflect generalized immune suppression, because high doses of these mAb had little, if any effect on the humoral immune response to another antigen. These findings indicate that: (1) mAb to B7-1 and B7-2 can elicit either a potent immune response or no immune response at all depending upon the dose administered; (2) blockade of the immune response to anti-B7 mAb may be more difficult in the setting of autoimmunity; and (3) neither anti-B7-1 nor anti-B7-2 causes generalized suppression of humoral immunity.

Enhanced lymphoproliferation and diminished autoimmunity in CD4-deficient MRL/lpr mice.

MRL/lpr mice spontaneously develop an autoimmune disease with features of systemic lupus erythematosus. They also develop a lymphoproliferative disorder characterized by a massive accumulation of double-negative (DN) T cells that lack both CD4 and CD8. To clarify the role of CD4 in autoimmunity and lymphoproliferation in these mice, CD4-deficient MRL/lpr mice were generated. CD4-deficient MRL/lpr mice developed massive expansion of DN T cells in the blood, spleen, and lymph nodes, which significantly exceeded the degree of lymphoproliferation in CD4-expressing control MRL/lpr mice. Despite this lymphoproliferation, CD4-deficient MRL/lpr mice produced little, if any, antibodies to double-stranded DNA, and they had prolonged survival relative to CD4-expressing littermates. However, they eventually developed moderately severe nephritis, characterized by immunoglobulin and complement deposition in glomeruli, vasculitis, and renal infiltration by CD8+ T cells. These findings indicate that (1) lymphoproliferation in MRL/lpr mice does not require the expression of CD4; (2) autoantibody production in MRL/lpr mice is dependent on the expression of CD4 and not on the accumulation of DN T cells; and (3) the development of nephritis in MRL/lpr mice involves both CD4-dependent and CD4-independent mechanisms.

Opportunities for future biological therapy in SLE.

The development of monoclonal antibodies and the emergence of recombinant DNA technology has made it possible to identify and selectively inhibit distinct cell subsets, surface molecules and secreted products that contribute to normal and pathological immune responses. These advances have helped to clarify the mechanisms that promote autoimmune diseases. As a result, it is now possible to contemplate rational strategies for the treatment of these diseases. Some of these strategies are designed to influence the cell surface interactions that determine whether potentially autoreactive T cells become activated or tolerant following antigen stimulation. Other strategies are designed to augment or inhibit distinct cytokines that regulate autoimmunity. All of these strategies have shown promise in animal models for systemic lupus erythematosus, and they may soon be translated into effective new therapies for people.

Long-term inhibition of murine lupus by brief simultaneous blockade of the B7/CD28 and CD40/gp39 costimulation pathways.

Murine lupus in NZB/NZW F1 (B/W) mice can be retarded by sustained administration of CTLA4Ig and by brief treatment early in life with mAb that block CD40/gp39 interactions. We sought to determine whether brief therapy with CTLA4Ig could provide sustained benefit in B/W mice and whether a synergistic effect could be derived by blockade of both the B7/CD28 and the CD40/gp39 pathways. We found that a short course of CTLA4Ig at the onset of disease produced only short-term benefit. However, when CTLA4Ig was combined with anti-gp39, there was long-lasting inhibition of autoantibody production and renal disease. Ten months after the 2-wk course of therapy, 70% of these mice were alive, compared with only 18% and 0% of those that received only anti-gp39 or CTLA4Ig, respectively. These findings demonstrate that brief simultaneous blockade of the B7/CD28 and CD40/gp39 costimulation pathways can produce benefit that lasts long after treatment has been discontinued.

The CD28-B7 costimulatory pathway and its role in autoimmune disease.

The activation of naive CD4+ T cells requires two discrete signals: a signal delivered by the T cell receptor following recognition of antigen and an accessory signal transduced when costimulatory receptors interact with their ligands. Particularly important in the development of an immune response to foreign antigens is the T cell molecule CD28, which delivers a potent costimulus when engaged by ligands, B7-1 and B7-2, on antigen-presenting cells. It is interesting that blockade of B7 molecules, which disrupts interactions with CD28 and prevents delivery of the CD28 costimulus, also alters the immune responses to self antigens and prevents the development of clinical disease in murine models of systemic and organ-specific autoimmunity. Herein we review the roles of CD28 and its B7 ligands in the pathogenesis of autoimmunity, discuss efforts to treat animal models of autoimmunity by modifying the CD28 signal, and consider the mechanisms by which manipulation of the CD28 signal alters the course of experimental autoimmune disease.

Blind T-cell homeostasis in CD4-deficient mice.

Recently, it has been proposed that normal T-cell count is maintained by a homeostatic mechanism which is "blind" to the distinction between CD4+ T cells and CD8+ T cells. Interest in this blind homeostasis hypothesis (BHH) stems in part from its implications regarding the pathogenesis and treatment of HIV infection. In this report, BHH was tested in CD4-deficient mice. We found that as predicted by BHH, despite the absence of CD4+ T cells, CD4-deficient mice maintain normal absolute T-cell counts in the blood and spleen primarily through a marked increase in CD8+ T cells. These findings provide strong new support for BHH.

Interleukin 6 promotes murine lupus in NZB/NZW F1 mice.

To investigate the role of IL-6 in systemic lupus erythematosus (SLE), we selectively inhibited IL-6 in lupus-prone NZB/NZW F1(B/W) mice by chronic administration of a rat mAb to mouse IL-6. Anti-IL-6 alone elicited an anti-rat response that blocked its biologic effects. To circumvent this problem, we rendered B/W mice tolerant to the rat mAb by administration of anti-CD4 concurrent with the first dose of anti-IL-6. Thereafter, the mice received weekly injections of anti-IL-6 alone. There were two control groups: one group received the tolerizing regimen of anti-CD4 along with a control rat IgG1 mAb (GL113) instead of anti-IL-6; the other control group received PBS. Mice that received anti-CD4 were tolerant to the rat mAb for 6 mo. Throughout this period, treatment with anti-IL-6 prevented production of anti-dsDNA, significantly reduced proteinuria, and prolonged life. Mice that received anti-IL-6 without anti-CD4 developed an immune response to the rat mAb and then developed anti-dsDNA antibodies, proteinuria, and mortality comparable with control mice. These findings establish that IL-6 promotes autoimmunity in B/W mice. They further indicate that, although mAb to IL-6 can suppress murine lupus, the development of host immunity to the mAb abrogates its beneficial effects. Finally, this is the first study to demonstrate that a brief course of anti-CD4 can induce tolerance to another therapeutic mAb, in this case an anti-cytokine mAb.

Treatment of murine lupus with monoclonal antibodies to lymphocyte function-associated antigen-1: dose-dependent inhibition of autoantibody production and blockade of the immune response to therapy.

Monoclonal antibodies (mAb) to lymphocyte function-associated antigen-1 (LFA-1) have been used successfully in vivo to inhibit immune responses and to block inflammatory reactions. To determine whether these effects of anti-LFA-1 could retard autoimmune disease, we treated lupus-prone NZB/NZW F1 (B/W) mice with a rat mAb to LFA-1 (anti-CD11a). Mice received high-dose therapy (500 micrograms twice weekly), low-dose therapy (40 micrograms thrice weekly), or phosphate-buffered saline from age 5 months to age 10 months. Treatment with high doses of anti-CD11a suppressed both the immune response to the rat mAb and the production of autoantibodies to double-stranded DNA. In contrast, treatment with low doses of anti-CD11a elicited an immune response to the rat mAb and did not suppress autoantibody production. The immunosuppressive effects of high doses of anti-CD11a were not due to target cell depletion. In fact, treatment induced a marked lymphocytosis which involved all lymphocyte subsets equally. Despite inhibiting autoantibody production, high-dose therapy had only modest effects on longevity.

Treatment of murine lupus with CTLA4Ig.

The interaction of B7-related molecules on antigen-presenting cells with CD28 or CTLA-4 antigens on T cells provides a second signal for T cell activation. Selection inhibition of the B7-CD28 or B7-CTLA-4 interactions produces antigen-specific T cell unresponsiveness in vitro and suppresses immune function in vivo. To determine whether selective inhibition of the B7-CD28 or B7-CTLA-4 interactions could suppress spontaneous autoimmune disease, a B7-binding protein was generated by genetic fusion of the extracellular domain of murine CTLA-4 to the Fc portion of a mouse immunoglobulin G2a monoclonal antibody (muCTLA4Ig). In lupus-prone NZB/NZW filial generation (F1) mice, treatment with muCTLA4Ig blocked autoantibody production and prolonged life, even when treatment was delayed until the most advanced stage of clinical illness. These findings suggest a possible role for human CTLA4Ig in the treatment of autoimmune diseases in humans.