Updates on Clinical Trials in Systemic Lupus Erythematosus.

PURPOSE OF REVIEW: Because of the complexity of systemic lupus erythematosus (SLE), different approaches are undertaken while investigating potential therapeutic compounds to treat the disease. The purpose of this review is to summarize the results from recent clinical trials, which investigated different compounds for treating SLE. RECENT FINDINGS: Targeting B cells and type I interferons constitutes the major focus in recent clinical trials. The potential for therapeutic effects of small molecule inhibition such as JAK, Tyk, and Btk is now being investigated for treating SLE. The immunoregulation of T cell activation in SLE is studied using low-dose IL-2 and CD40 ligand inhibition. There are clinical trials that study bispecific antibodies, with binding specificities for 2 different target molecules related to T- and B-cell activation or to different aspects of B cell activation. An approach of combination treatment is also being studied. Clinical trials are underway and new treatment compounds for SLE are being anticipated.

PPP2R2D suppresses IL-2 production and Treg function.

Protein phosphatase 2A is a ubiquitously expressed serine/threonine phosphatase that comprises a scaffold, a catalytic, and multiple regulatory subunits and has been shown to be important in the expression of autoimmunity. We considered that a distinct subunit may account for the decreased production of IL-2 in people and mice with systemic autoimmunity. We show that the regulatory subunit PPP2R2D is increased in T cells from people with systemic lupus erythematosus and regulates IL-2 production. Mice lacking PPP2R2D only in T cells produce more IL-2 because the IL-2 gene and genes coding for IL-2-enhancing transcription factors remain open, while the levels of the enhancer phosphorylated CREB are high. Mice with T cell-specific PPP2R2D deficiency display less systemic autoimmunity when exposed to a TLR7 stimulator. While genes related to Treg function do not change in the absence of PPP2R2D, Tregs exhibit high suppressive function in vitro and in vivo. Because the ubiquitous expression of protein phosphatase 2A cannot permit systemic therapeutic manipulation, the identification of regulatory subunits able to control specific T cell functions opens the way for the development of novel, function-specific drugs.

Systemic lupus erythematosus favors the generation of IL-17 producing double negative T cells.

Mature double negative (DN) T cells are a population of αß T cells that lack CD4 and CD8 coreceptors and contribute to systemic lupus erythematosus (SLE). The splenic marginal zone macrophages (MZMs) are important for establishing immune tolerance, and loss of their number or function contributes to the progression of SLE. Here we show that loss of MZMs impairs the tolerogenic clearance of apoptotic cells and alters the serum cytokine profile, which in turn provokes the generation of DN T cells from self-reactive CD8+ T cells. Increased Ki67 expression, narrowed TCR V-beta repertoire usage and diluted T-cell receptor excision circles confirm that DN T cells from lupus-prone mice and patients with SLE undergo clonal proliferation and expansion in a self-antigen dependent manner, which supports the shared mechanisms for their generation. Collectively, our results provide a link between the loss of MZMs and the expansion of DN T cells, and indicate possible strategies to prevent the development of SLE.

Serine/threonine phosphatase PP2A is essential for optimal B cell function.

Protein phosphatase 2A (PP2A), a serine/threonine phosphatase, has been shown to control T cell function. We found that in vitro-activated B cells and B cells from various lupus-prone mice and patients with systemic lupus erythematosus display increased PP2A activity. To understand the contribution of PP2A to B cell function, we generated a Cd19CrePpp2r1afl/fl (flox/flox) mouse which lacks functional PP2A only in B cells. Flox/flox mice displayed reduced spontaneous germinal center formation and decreased responses to T cell-dependent and T-independent antigens, while their B cells responded poorly in vitro to stimulation with an anti-CD40 antibody or CpG in the presence of IL-4. Transcriptome and metabolome studies revealed altered nicotinamide adenine dinucleotide (NAD) and purine/pyrimidine metabolism and increased expression of purine nucleoside phosphorylase in PP2A-deficient B cells. Our results demonstrate that PP2A is required for optimal B cell function and may contribute to increased B cell activity in systemic autoimmunity.

T cell metabolism: new insights in systemic lupus erythematosus pathogenesis and therapy.

T cell subsets are critically involved in the development of systemic autoimmunity and organ inflammation in systemic lupus erythematosus (SLE). Each T cell subset function (such as effector, helper, memory or regulatory function) is dictated by distinct metabolic pathways requiring the availability of specific nutrients and intracellular enzymes. The activity of these enzymes or nutrient transporters influences the differentiation and function of T cells in autoimmune responses. Data are increasingly emerging on how metabolic processes control the function of various T cell subsets and how these metabolic processes are altered in SLE. Specifically, aberrant glycolysis, glutaminolysis, fatty acid and glycosphingolipid metabolism, mitochondrial hyperpolarization, oxidative stress and mTOR signalling underwrite the known function of T cell subsets in patients with SLE. A number of medications that are used in the care of patients with SLE affect cell metabolism, and the development of novel therapeutic approaches to control the activity of metabolic enzymes in T cell subsets represents a promising endeavour in the search for effective treatment of systemic autoimmune diseases.

PP2A enables IL-2 signaling by preserving IL-2Rß chain expression during Treg development.

Tregs require IL-2 signaling for signal transducer and activator of transcription 5 (STAT5)-mediated induction of Foxp3. While phosphatase 2A (PP2A) is a negative regulator of IL-2 production in effector T cells and Tregs do not produce IL-2, it is not known whether PP2A controls IL-2 signaling in Tregs. To address the role of PP2A in IL-2 signaling in Tregs we studied mice engineered to lack PP2A in all Foxp3-expressing cells. We report that PP2A is required to enable Foxp3 expression and to maintain sufficient numbers of Tregs in the thymus. We show for the first time that PP2A prevents the selective loss of surface IL-2Rß and preserves IL-2R signaling potency in Tregs. The loss of IL-2Rß in thymus- and spleen-derived Tregs that lack PP2A is due to increased sheddase activity. Pan-sheddase or selective A disintegrin and metalloproteinase 10 (ADAM10) inhibition, like forced expression of IL-2Rß in PP2A-deficient Tregs restored IL-2Rß expression and signaling. Thus, PP2A restrains the sheddase activity of ADAM10 in Treg cells to prevent the cleavage of IL-2Rß from the cell surface to enable competent IL-2R signaling which is essential for Tregs development and homeostasis.

Regulatory T cells in the treatment of disease.

Regulatory T (Treg) cells suppress inflammation and regulate immune system activity. In patients with systemic or organ-specific autoimmune diseases or those receiving transplanted organs, Treg cells are compromised. Approaches to strengthen Treg cell function, either by expanding them ex vivo and reinfusing them or by increasing the number or capacity of existing Treg cells, have entered clinical trials. Unlike the situation in autoimmunity, in patients with cancer, Treg cells limit the antitumour immune response and promote angiogenesis and tumour growth. Their immunosuppressive function may, in part, explain the failure of many immunotherapies in cancer. Strategies to reduce the function and/or number of Treg cells specifically in tumour sites are being investigated to promote antitumour immunity and regression. Here, we describe the current progress in modulating Treg cells in autoimmune disorders, transplantation and cancer.

Indoleamine-2,3-dioxygenase in murine and human systemic lupus erythematosus: Down-regulation by the tolerogeneic peptide hCDR1.

וֹndoleamine-2,3-dioxygenase (IDO) plays a role in immune regulation. Increased IDO activity was reported in systemic lupus erythematosus (SLE). We investigated the effects of the tolerogenic peptide hCDR1, shown to ameliorate lupus manifestations, on IDO gene expression. mRNA was prepared from splenocytes of hCDR1- treated SLE-afflicted (NZBxNZW)F1 mice, from blood samples of lupus patients, collected before and after their in vivo treatment with hCDR1 and from peripheral blood mononuclear cells (PBMC) of patients incubated with hCDR1. IDO gene expression was determined by real-time RT-PCR. hCDR1 significantly down-regulated IDO expression in SLE-affected mice and in lupus patients (treated in vivo and in vitro). No effects were observed in healthy donors or following treatment with a control peptide. Diminished IDO gene expression was associated with hCDR1 beneficial effects. Our results suggest that the hCDR1-induced FOXP3 expressing regulatory T cells in lupus are not driven by IDO but rather by other hCDR1 regulated pathways.

Precision DNA demethylation ameliorates disease in lupus-prone mice.

Defective DNA methylation in T cells leads to a series of T cell abnormalities in lupus; however, the full effect of T cell lineage-specific DNA methylation on disease expression has not been explored. Here, we show that 5-azacytidine, a DNA methyltransferase inhibitor, targeted to either CD4 or CD8 T cells in mice with established disease using a nanolipogel delivery system dramatically ameliorates lupus-related pathology through distinct mechanisms. In vivo targeted delivery of 5-azacytidine into CD4 T cells favors the expansion and function of Foxp3+ Tregs, whereas targeted delivery to CD8 T cells enhances the cytotoxicity and restrains the expansion of pathogenic TCR-αß+CD4-CD8- double-negative T cells. Our results signify the importance of cell-specific inhibition of DNA methylation in the treatment of established lupus.

The tolerogenic peptide hCDR1 immunomodulates cytokine and regulatory molecule gene expression in blood mononuclear cells of primary Sjogren's syndrome patients.

Primary Sjogren's syndrome (pSS) is an autoimmune disease characterized by lymphocytic infiltration of exocrine glands. We investigated whether the tolerogenic peptide, hCDR1, that ameliorates lupus manifestations would have beneficial effects on pSS as well. The in vitro effects of hCDR1 on gene expression of pro-inflammatory cytokines and regulatory molecules were tested in peripheral blood mononuclear cells (PBMC) of 16 pSS patients. hCDR1, but not a control peptide, significantly reduced gene expression of IL-1ß, TNF-α, MX-1 and BlyS and up-regulated immunosuppressive (TGF-ß, FOXP3) molecules in PBMC of pSS patients. hCDR1 did not affect gene expression in patients with rheumatoid arthritis and anti-phospholipid syndrome. Further, hCDR1 up-regulated the expression of Indoleamine 2,3-dioxygenase (IDO) via elevation of TGF-ß. IDO inhibition led to a significant decrease in the expression of FOXP3 which is crucial for the induction of T regulatory cells. Thus, hCDR1 is potential candidate for the specific treatment of pSS patients.

The serine/threonine protein phosphatase 2A controls autoimmunity.

Protein phosphatase 2A (PP2A) is the first serine/threonine phosphatase recognized to contribute to human and murine lupus immunopathology. PP2A expression in SLE is controlled both epigenetically and genetically, and it is increased in patients with SLE, which contributes to decreased IL-2 production, decreased CD3ζ and increased FcRγ expression on the surface of T cells, increased CREMα expression, hypomethylation of genes associated with SLE pathogenesis, and increased IL-17 production. ß regulatory subunit of PP2A regulates IL-2 deprivation-induced T cell death and is decreased in SLE patients. A mouse overexpressing PP2Ac in T cells displays peripheral granulocytosis, elevated IL-17 production, and develops glomerulonephritis when challenged. A mouse which lacks PP2Ac only in regulatory T cells develops severe autoimmunity and multiorgan inflammation because of loss of restraint on mTORC1 and inability of Foxp3+ cells to regulate conventional T cells. Targeting PP2A in T cell subsets may be therapeutic for SLE and other autoimmune diseases.

Empowering Regulatory T Cells in Autoimmunity.

Regulatory T cells (Tregs) are capable of dampening immune-mediated inflammation and avert the destructive effects of uncontrolled inflammation. Distinct molecules and pathways, including various transcription factors, phosphatases, and kinases, impact the ability of Tregs to function as negative regulators of the immune response, and are presumably amenable to therapeutic manipulation. Here, we discuss recently identified molecular networks and the therapeutic potential for treating autoimmune diseases.

Induced Sputum Analysis in Subjects With Systemic Sclerosis.

BACKGROUND: Induced sputum is a noninvasive tool, aimed at collecting cellular and soluble materials from lung airways. Induced sputum sampling analysis has been validated in chronic obstructive lung diseases as well as in various diffuse interstitial lung disorders. Our objective was to evaluate the utility of induced sputum speciments of cellular and soluble materials noninvasively sampled from the lung airways of subjects with systemic sclerosis compared with healthy controls and determine possible correlation with disease manifestations. METHODS: The study population included 20 consecutive subjects with systemic sclerosis and 16 healthy volunteers (controls). All of the participants underwent pulmonary function testing (PFT), single-breath diffusion capacity of carbon monoxide, and induced sputum cytology analysis. RESULTS: The systemic sclerosis subjects' induced sputum samples contained a higher neutrophil count (P = .09) and lower lymphocyte count (P = .045) compared with the controls. Within the systemic sclerosis population, several clinical, PFT, and induced sputum findings were positively correlated: the modified Rodnan skin score score and CD3 (P = .044), modified Rodnan skin score and CD4 (P = .04), modified Rodnan skin score and percentage neutrophils (P = .059), and total lung capacity (TLC) and percentage lymphocytes (P = .02). Induced sputum neutrophil counts were inversely associated with TLC (P = .02) and FVC (P = .02). Induced sputum samples from the subgroup of systemic sclerosis subjects with PFT restrictive pattern contained a significantly lower percentage of lymphocytes (P = .004) and lower CD4/CD8 ratio (P = .03) compared with controls. In this subgroup, TLC also was negatively correlated with the neutrophil count (P = .043), and positive correlations were found between TLC and CD3 (P = .053), disease duration and CD4/CD8 ratio (P = .033), TLC and FVC and percentage macrophages (P = .033 and P = .052, respectively), diffusion per unit of alveolar volume, and percentage macrophages (P = .032), and C-reactive protein and percentage lymphocytes, percentage macrophages, and neutrophil count (P = .02, P = .008, and P = .006, respectively). CONCLUSIONS: Induced sputum analysis of subjects with systemic sclerosis, including those with restrictive lung disease, shows changes in cellular pattern and correlation with several highly relevant clinical and PFT parameters.

The Effect of the Presence of Fibromyalgia on Common Clinical Disease Activity Indices in Patients with Psoriatic Arthritis: A Cross-sectional Study.

OBJECTIVE: To study the effect of the presence of fibromyalgia (FM) on common clinical disease activity indices in patients with psoriatic arthritis (PsA). METHODS: Seventy-three consecutive outpatients with PsA (mean age 51.7 yrs; 42 females, 57.5%) were enrolled in a prospective cross-sectional study. FM was determined according to American College of Rheumatism criteria (2010 and 1990). All patients underwent clinical evaluation of disease activity and completed the Health Assessment Questionnaire (HAQ), the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), the Dermatology Life Quality Index, and the Leeds Enthesitis Index (LEI). Disease activity was evaluated using the Composite Psoriatic Disease Activity Index (CPDAI), minimal disease activity (MDA), and the Disease Activity Index for Psoriatic Arthritis (DAPSA) scores. RESULTS: The overall prevalence of FM was 17.8% (13 patients), and all but 1 were women (12 patients, 92.3%, p = 0.005). CPDAI and DAPSA scores were significantly higher in patients with coexisting PsA and FM (9.23 ± 1.92 and 27.53 ± 19.23, respectively) than in patients with PsA only (4.25 ± 3.14 and 12.82 ± 12.71, respectively; p < 0.001 and p = 0.003). None of the patients with FM + PsA met the criteria for MDA, whereas 26 PsA-only patients did (43.3%, p = 0.003). HAQ, BASDAI, and LEI scores were significantly worse in patients with PsA and associated FM. CONCLUSION: Coexisting FM is related to worse scores on all tested measures in patients with PsA. Its influence should be taken into consideration in the treatment algorithm to avoid unnecessary upgrading of treatment.

Novel approaches to the development of targeted therapeutic agents for systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a chronic multisystem disease in which various cell types and immunological pathways are dysregulated. Current therapies for SLE are based mainly on the use of non-specific immunosuppressive drugs that cause serious side effects. There is, therefore, an unmet need for novel therapeutic means with improved efficacy and lower toxicity. Based on recent better understanding of the pathogenesis of SLE, targeted biological therapies are under different stages of development. The latter include B-cell targeted treatments, agents directed against the B lymphocyte stimulator (BLyS), inhibitors of T cell activation as well as cytokine blocking means. Out of the latter, Belimumab was the first drug approved by the FDA for the treatment of SLE patients. In addition to the non-antigen specific agents that may affect the normal immune system as well, SLE-specific therapeutic means are under development. These are synthetic peptides (e.g. pConsensus, nucleosomal peptides, P140 and hCDR1) that are sequences of conserved regions of molecules involved in the pathogenesis of lupus. The peptides are tolerogenic T-cell epitopes that immunomodulate only cell types and pathways that play a role in the pathogenesis of SLE without interfering with normal immune functions. Two of the peptides (P140 and hCDR1) were tested in clinical trials and were reported to be safe and well tolerated. Thus, synthetic peptides are attractive potential means for the specific treatment of lupus patients. In this review we discuss the various biological treatments that have been developed for lupus with a special focus on the tolerogenic peptides.

The tolerogenic peptide, hCDR1, down-regulates the expression of interferon-α in murine and human systemic lupus erythematosus.

BACKGROUND: The tolerogenic peptide, hCDR1, ameliorated manifestations of systemic lupus erythematosus (SLE) via the immunomodulation of pro-inflammatory and immunosuppressive cytokines and the induction of regulatory T cells. Because type I interferon (IFN-α) has been implicated to play a role in SLE pathogenesis, we investigated the effects of hCDR1 on IFN-α in a murine model of SLE and in human lupus. METHODOLOGY PRINCIPAL FINDINGS: (NZBxNZW)F1 mice with established SLE were treated with hCDR1 (10 weekly injections). Splenocytes were obtained for gene expression studies by real-time RT-PCR. hCDR1 down-regulated significantly IFN-α gene expression (73% inhibition compared to vehicle treated mice, p = 0.002) in association with diminished clinical manifestations. Further, hCDR1 reduced, in vitro, IFN-α gene expression in peripheral blood mononuclear cells (PBMC) of 10 lupus patients (74% inhibition compared to medium, p = 0.002) but had no significant effects on the expression levels of IFN-α in PBMC of primary anti-phospholipid syndrome patients or of healthy controls. Lupus patients were treated for 24 weeks with hCDR1 (5) or placebo (4) by weekly subcutaneous injections. Blood samples collected, before and after treatment, were frozen until mRNA isolation. A significant reduction in IFN-α was determined in hCDR1 treated patients (64.4% inhibition compared to pretreatment expression levels, p = 0.015). No inhibition was observed in the placebo treated patients. In agreement, treatment with hCDR1 resulted in a significant decrease of disease activity. IFN-α appears to play a role in the mechanism of action of hCDR1 since recombinant IFN-α diminished the immunomodulating effects of hCDR1 on IL-1ß, TGFß and FoxP3 gene expression. CONCLUSIONS SIGNIFICANCE: We reported previously that hCDR1 affected various cell types and immune pathways in correlation to disease amelioration. The present studies demonstrate that hCDR1 is also capable of down-regulating significantly (and specifically to lupus) IFN-α gene expression. Thus, hCDR1 has a potential role as a novel, disease specific treatment for lupus.

Immune recovery after cyclophosphamide treatment in multiple myeloma: implication for maintenance immunotherapy.

Multiple myeloma (MM) is a progressive B-lineage neoplasia characterized by clonal proliferation of malignant plasma cells. Increased numbers of regulatory T cells (Tregs) were determined in mouse models and in patients with MM, which correlated with disease burden. Thus, it became rational to target Tregs for treating MM. The effects of common chemotherapeutic drugs on Tregs are reviewed with a focus on cyclophosphamide (CYC). Studies indicated that selective depletion of Tregs may be accomplished following the administration of a low-dose CYC. We report that continuous nonfrequent administrations of CYC at low doses block the renewal of Tregs in MM-affected mice and enable the restoration of an efficient immune response against the tumor cells, thereby leading to prolonged survival and prevention of disease recurrence. Hence, distinctive time-schedule injections of low-dose CYC are beneficial for breaking immune tolerance against MM tumor cells.