Ewa Robak

Serum levels of interleukin-6 type cytokines and soluble interleukin-6 receptor in patients with rheumatoid arthritis.

We investigated the serum concentrations of interleukin-6 (IL-6) and two IL-6 family of cytokines (leukaemia inhibitory factor (LIF) and ciliary neurotrophic factor (CNTF) as well as IL-6 soluble receptor (sIL-6R) using an enzyme-linked immunosorbent assay (ELISA) in 66 patients with rheumatoid arthritis (RA) and 24 healthy controls. We examined a possible association between the serum levels of these peptides and RA activity according to the Mallya and Mace scoring system and Ritchies index. We also evaluated the correlation between the serum levels of IL-6, LIF, CNTF and sIL-6R and duration of the disease and calculated sIL-6R/IL-6 ratio in RA patients and in the control group. IL-6 and sIL-6R were detectable in all 66 patients with RA and 24 normal individuals. LIF was also found in the serum of all patients with RA and in 16 (66.7%) normal individuals. In contrast CNTF was measurable only in 15 (22.7%) patients with RA and 24 (33.3%) normal individuals. The highest IL-6 and sIL-6R levels were found in the patients with Stages 3 and 4 of RA activity and the lowest in the control group. In contrast there were no statistically significant differences between the LIF and CNTF levels in RA patients and normal individuals. We found positive correlation between IL-6 and sIL-6R concentrations and Ritchies index and a lack of such correlation with LIF and CNTF. IL-6 serum level correlated positively with the disease duration, but sIL-6R, LIF and CNTF did not. Serum sIL-6R/IL-6 ratio was significantly lower in RA patients than in healthy controls. In conclusion, an increase in the serum levels of IL-6 and sIL-6R, but not LIF and CNTF concentrations, may be useful markers for RA activity.

Tyrosine kinase inhibitors as potential drugs for B-cell lymphoid malignancies and autoimmune disorders

Introduction: In the last few years, several tyrosine kinase inhibitors (TKIs) have been synthesized and become available for preclinical studies and clinical trials. This article summarizes recent achievements in the mechanism of action, pharmacological properties, and clinical activity and toxicity, as well as the emerging role of TKIs in lymphoid malignancies, allergic diseases, and autoimmune disorders. Areas covered: A literature review was conducted of the MEDLINE database PubMed for articles in English. Publications from 2000 through January 2012 were scrutinized. The search terms used were Brutons tyrosine kinase (Btk) inhibitors, PCI-32765, GDC-0834, LFM-A13, AVL-101, AVL-292, spleen tyrosine kinase (Syk) inhibitors, R343, R406, R112, R788, fostamatinib, BAY-61-3606, C-61, piceatannol, Lyn, imatinib, nilotinib, bafetinib, dasatinib, GDC-0834, PP2, SU6656 in conjunction with lymphoid malignancy, NHL, CLL, autoimmune disease, allergic disease, asthma, and rheumatoid arthritis. Conference proceedings from the previous 5 years of the American Society of Hematology, European Hematology Association, American Society of Clinical Oncology, and ACR/ARHP Annual Scientific Meetings were searched manually. Additional relevant publications were obtained by reviewing the references from the chosen articles. Expert opinion: The use of TKIs, especially inhibitors of Btk, Syk, and Lyn, is a promising new strategy for targeted treatment of B-cell lymphoid malignancies, autoimmune disorders and allergic diseases. However, definitive data from ongoing and future clinical trials will aid in better defining the status of TKIs in the treatment of these disorders.

New anti-CD20 monoclonal antibodies for the treatment of B-cell lymphoid malignancies.

Over the last few years, new generations of anti-CD20 monoclonal antibodies (mAbs) have been developed for potential benefits over the classical, first-generation mAb rituximab. Compared with rituximab, new mAbs have enhanced antitumor activity resulting from increased complement-dependent cytotoxicity (CDC) and/or antibody-dependent cellular cytotoxicity (ADCC) and increased Fc binding affinity for the low-affinity variants of the FcγRIIIa receptor (CD16) on immune effector cells. The second-generation mAbs, which include ofatumumab, veltuzumab, and ocrelizumab, are humanized or fully human to reduce immunogenicity, but with an unmodified Fc region. Ofatumumab is a fully human anti-CD20 IgG1 mAb in clinical development for hematological malignancies and autoimmune diseases. Ofatumumab specifically recognizes an epitope encompassing both the small and large extracellular loops of CD20 molecule, and is more effective than rituximab at CDC induction and killing target cells. Veltuzumab (IMMU-106, hA20) is a humanized anti-CD20 mAb with complementarity-determining regions similar to rituximab. This antibody has enhanced binding avidities and a stronger effect on CDC compared with rituximab. Ocrelizumab is a humanized mAb with the potential for enhanced efficacy in lymphoid malignancies compared with rituximab due to increased binding affinity for the low-affinity variants of the FcγRIIIa receptor. The third-generation mAbs are also humanized mAbs, but in addition they have an engineered Fc to increase their binding affinity for the FcγRIIIa receptor. The third-generation mAbs include AME-133v, PRO131921 and GA-101. AME-133v (LY2469298) is a type I, humanized IgG1 mAb with enhanced affinity for FcγRIIIa receptor and an enhanced ADCC activity compared with rituximab. PRO131921 is a humanized anti-CD20 mAb engineered to have improved binding to FcγRIIIa and better ADCC compared with rituximab. GA-101 (RO5072759) is a fully humanized, type II, IgG1 mAb derived from humanization of the parental B-Ly1 mouse antibody and subsequent glycoengineering using GlycoMab® technology. GA-101 was designed for enhanced ADCC and superior direct cell-killing properties, in comparison with currently available type I antibodies. TRU-015 is a small modular immunopharmaceutical (SMIP) derived from key domains of an anti-CD20 antibody. TRU-015 represents a novel biological compound that retains Fc-mediated effector functions and is smaller than mAbs. In this article we review data on new anti-CD20 mAbs that are potentially useful in the treatment of lymphoid malignancies.

Vascular endothelial growth factor and its soluble receptors VEGFR-1 and VEGFR-2 in the serum of patients with systemic lupus erythematosus

We investigated the serum concentration of vascular endothelial growth factor (VEGF) and its two soluble receptors, sVEGFR-1 and sVEGFR-2, in a group of 60 patients with systemic lupus erythematosus (SLE), and 20 healthy controls, using an enzyme-linked immunosorbent assay. We examined a possible association between serum levels of these proteins and certain clinical and laboratory parameters as well as SLE activity. VEGF, sVEGFR-1 and sVEGFR-2 were detectable in all patients with SLE and in all normal individuals. The VEGF level was higher in active SLE (mean, 300.8 pg/ml) than in inactive SLE (mean, 165.9 pg/ml) (p < 0.05) or in the control group (mean, 124.7 pg/ml) (p < 0.04). The highest sVEGFR-1 concentrations were also detected in active SLE patients (mean, 42.2 pg/ml) and the lowest in inactive disease (mean, 32.0 pg/ml) (p < 0.01). In contrast, the levels of sVEGFR-2 were lower in SLE (mean, 12557.6 pg/ml) than in the control group (mean, 15025.3 pg/ml) (p < 0.05). We found a positive correlation between sVEGFR-1 concentration and the SLE activity score p = 0.375 (p < 0.004) and a negative, but statistically insignificant correlation between sVEGFR-2 and SLE activity (p = -0.190, p > 0.05). Treatment with steroids and cytotoxic agents did not influence VEGF or its soluble receptors levels. In conclusion, in SLE patients the levels of VEGF and sVEGFR-1 are higher in patients with active SLE than in inactive disease or healthy persons. In contrast, the level of sVEGFR-2 is lower in active SLE than in inactive disease. The imbalance between VEGF and its soluble receptors may be important in SLE pathogenesis.

Lymphocyctes Tgammadelta in clinically normal skin and peripheral blood of patients with systemic lupus erythematosus and their correlation with disease activity.

Human Tgammadelta lymphocytes constitute from 1 to 15% of all peripheral blood lymphocytes. Recent work has demonstrated that this population plays a major role in the pathogenesis of infectious and immune diseases. Increased numbers of gammadelta T cells have been found in affected skin from systemic sclerosis and chronic cutaneous lupus erythematosus patients. In our study, we have determined the numbers of Tgammadelta lymphocytes and their subpopulations in peripheral blood from 29 patients with systemic lupus erythematosus (SLE) and in 19 healthy volunteers using flow cytometry and specific monoclonal antibodies. The same cells in uninvolved skin from SLE patients and human controls using immunohistochemical analysis were estimated. T-Cell receptor (TCR) delta chain gene rearrangement was identified with primers for Vdelta1, Vdelta2 and Vdelta3 by the polymerase chain reaction. Statistical analysis showed a significantly decreased number of gammadelta T cells in SLE patients (26.4+/-16.9/microl) compared with the control group (55.3+/-20.6/microl (p < 0.001). The number of Vdelta2 TCR+ and Vgamma9 TCR+ subpopulations was also lower in SLE patients than in healthy persons. No statistical correlation between disease activity and the number of gammadelta T cells was demonstrated. The percentage of Tgammadelta lymphocytes in clinically normal skin from SLE patients was twice (22.0+/-9.4%) that found in the skin from healthy persons (11.1+/-5.5%) (p < 0.002). Higher percentages of the Vdelta2 TCR+ and Vgamma9 TCR+ subpopulation of lymphocytes were found in the skin from SLE patients. We have also found positive correlation between the percentage of Tgammadelta lymphocytes in skin and the activity of SLE (r=0.594, p < 0.001), and between subpopulation Vdelta3 TCR+ and disease activity (r=0.659, p< 0.001). In conclusion, the results of our studies demonstrate that, in patients with SLE, accumulation of Tgammadelta lymphocytes can be seen in clinically normal skin, and the percentage of these cells correlates with the activity of the disease.

Quality of life and satisfaction with life in SLE patients—the importance of clinical manifestations

To assess the correlation between quality of life (QoL) and satisfaction with life (SL) in SLE patients and correlate both with clinical symptoms of the disease. The study was performed in 83 patients. QoL was assessed by Short Form 36, and SL was assessed by the Satisfaction with Life Scale. Clinical manifestations presented at the time of examination were taken into consideration. SLE patients assessed their QoL and SL as rather low. Those with photosensitivity as well as neurological symptoms presented lower QoL in particular domains, while those with renal manifestation of SLE assessed their QoL as higher. Similar observations were made for SL only in relation to neurological symptoms. Moreover, our findings show that although SL is a part of QoL, both these parameters should be distinguished in order to fully assess the state of the patient.

Monoclonal antibodies in the treatment of systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by B cell hyperactivity and defective T-cell function, with production of high titer autoantibodies. In the recent years, conceptual advances and the introduction of new therapies are yielding improvements in the management of this disease. In recent years, clinical studies have been undertaken with selected monoclonal antibodies (mAbs) in the treatment of SLE. The important role of B cells in the pathogenesis of autoimmune disorders has provided a strong rationale to target B cells in SLE. Selective therapeutic depletion of B-cells became possible with the availability of the anti-CD20 antibody rituximab and anti-CD22 antibody epratuzumab. Several clinical studies confirm high activity of rituximab in SLE patients especially with lupus nephritis and neuropsychiatric involvement. Recently, several new mAbs reacting with CD20 have been developed. New mAbs directed against CD20 include fully human mAb ofatumumab (HuMax CD20), IMMU-106 (hA20) which has a >90% humanized framework and GA-101, a novel third-generation fully humanized and optimized mAb. These agents are highly cytotoxic against B-cell lymphoid cells. Proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and iterleukin-6 (IL-6) play an important role in propagating the inflammatory process responsible for tissue damage. Blocking of these cytokines by mAbs can be also a successful therapy for patients with SLE. Finally, mAb eculizumab that specifically inhibits terminal complement activation has been recently developed and investigated in the phase I single dose study in SLE. In this review, new mAbs, potentially useful in SLE are presented.

Circulating angiogenesis inhibitor endostatin and positive endothelial growth regulators in patients with systemic lupus erythematosus

Serum concentrations of three angiogenic cytokines: vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), interleukin-18 (IL-18) and antiangiogenic factor endostatin in the serum of 52 patients with systemic lupus erythematosus (SLE) and 20 healthy subjects were investigated. The possible association between serum levels of these proteins and SLE activity as well as correlation between the concentrations of angiogenic cytokines and the level of endostatin was also analyzed. VEGF and IL-18 were detectable in all SLE patients and healthy control group. bFGF was measurable in 71.2% of patients with SLE and 65% of healthy persons. Endostatin was detectable in 94.2% of SLE patients and 95% of normal subjects. The serum levels of endostatin and bFGF were not significantly different in SLE and healthy control (P > 0.05). The median concentration of VEGF was higher in active SLE (238.4pg/ml) than in inactive disease (118.1pg/ml, P < 0.05) or in control group (133.5pg/ml, P < 0.04). The median serum level of IL-18 was higher in the SLE patients (595.2pg/ml) than in the control group (252.7pg/ml) (P < 0.001). The correlations between the levels of angiogenic cytokines and endostatin with clinical features, laboratory abnormalities and also with the type of treatment were analysed. We found a positive correlation between VEGF serum concentration and SLE activity according to SLAM score (r=0.275, P < 0.05). The significant positive correlation was also found between IL-18 and endostatin (r=0.289, P < 0.04). In contrast, the correlation between bFGF and endostatin was significantly negative (r=7 0.299, P < 0.04). In conclusion, serum levels of the angiogenic and antiangiogenic factors may play an important role in SLE pathogenesis.

Estimation of SLE activity based on the serum level of chosen cytokines and superoxide radical generation

We estimated the serum levels of IL-6, TNF-alpha and IL-10, and generation of superoxide radicals, as well as their mutual dependence, in 63 SLE patients at various stages of disease activity. Our results indicate a statistically significant increase of the serum levels studied, and an increase of superoxide anion generation by granulocytes, in correlation with SLE activity. These results indicate that oxygen metabolism and the examined cytokines play an important role in pathogenesis of SLE. The assessment of these parameters can be useful in the estimation of disease activity.

Expression of Toll-Like Receptors 3, 7, and 9 in Peripheral Blood Mononuclear Cells from Patients with Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) is an autoimmune disease of unknown aetiology. The results of experimental studies point to the involvement of innate immunity receptors—toll-like receptors (TLR)—in the pathogenesis of the disease. The aim of the study was to assess the expression of TLR3, 7, and 9 in the population of peripheral blood mononuclear cells (PBMC) and in B lymphocytes (CD19+), T lymphocytes (CD4+ and CD8+) using flow cytometry. The study group included 35 patients with SLE and 15 healthy controls. The patient group presented a significantly higher percentage of TLR3- and TLR9-positive cells among all PBMCs and their subpopulations (CD3+, CD4+, CD8+, and CD19+ lymphocytes) as well as TLR7 in CD19+ B-lymphocytes, compared to the control group. There was no correlation between the expression of all studied TLRs and the disease activity according to the SLAM scale, and the degree of organ damage according to the SLICC/ACR Damage Index. However, a correlation was observed between the percentage of various TLR-positive cells and some clinical (joint lesions) and laboratory (lymphopenia, hypogammaglobulinemia, anaemia, and higher ESR) features and menopause in women. The results of the study suggest that TLR3, 7, and 9 play a role in the pathogenesis of SLE and have an impact on organ involvement in SLE.