Safety Aspects of Immunoadsorption in IgG Removal Using a Single‐Use,Multiple‐pass Protein A Immunoadsorber (LIGASORB): Clinical Investigation in Healthy Volunteers

Therapeutic apheresis and immunoadsorption are used to deplete efficiently pathogenic autoantibodies in crises in several acute autoimmune driven diseases. This prospective, non‐comparative cohort study was conducted at a single study center under standardized conditions in 10 healthy volunteers. Efficient immunoglobulin G (IgG) removal (?86% versus baseline) was achieved after 3 apheresis treatments on 3 consecutive days. The treatments were well tolerated. Safety laboratory parameters did not show unexpected or pathological changes. The effects were transient, with most parameters exhibiting complete recovery between treatments. Minimal complement activation and moderate transient fibrinogen depletion were observed. Immunoadsorption with LIGASORB® provides a safe and effective treatment alternative to TPE in acute episodes of peripheral neurological diseases mediated by pathogenic IgG autoantibodies.     

A case report of successful treatment with immunoadsorption onto protein A in mixed connective tissue disease in childhood

An 11-year-old male patient suffering mixed connective tissue disease with life-threatening pulmonary arterial hypertension, progressive heart failure (New York Heart Association class III-IV), skin ulcers, Raynaud's phenomenon and arthritis, showing no improvement after intensive immunosuppressive therapy or high dose steroids, was treated with immunoadsorption onto protein A. With a combined therapy of low-dose cortisone and bosentan and 22 sessions of immunoadsorption, his condition improved significantly and he continues in clinical remission. At the time of writing no further immunosuppressive therapy or immunoadsorption had been necessary. The patient is now 15 years old and healthy with an age-based constitution comparable to the normal population.     

HLA class II influences humoral autoimmunity in patients with type 2 autoimmune hepatitis

BACKGROUND/AIMS: Type 2 autoimmune hepatitis (AIH) is characterized by the presence of anti-liver kidney microsome (anti-LKM-1) and/or anti-liver cytosol type 1 (anti-LC1) autoantibodies. However, the correlation between these autoantibodies and the genetic background has not been studied. METHODS: Frequencies of HLA class II alleles were compared between the 60 Caucasian children with type 2 AIH and 313 control subjects. The anti-LKM1 antibody reactivity directed against antigenic sites of CYP2D6 was analysed by ELISA. RESULTS: HLA-DQB1 *0201 allele was found to be the primary genetic determinant of susceptibility to type 2 AIH by conferring the highest odd-ratio (OR = 6.4). HLA-DRB1 *03 allele was significantly increased (P < 0.0001) among patients with both anti-LKM1 and anti-LC1 autoantibodies as well as in those with only anti-LC1(+) compared to those with anti-LKM1(+) alone. In contrast, HLA-DRB1 *07 allele was significantly associated (P < 0.0001) with anti-LKM1(+) alone compared to groups with both anti-LKM and anti-LC1 or with LC1+ alone. Children with the DRB1 *07 allele develop anti-LKM1 autoantibodies having a more restricted specificity (2 epitopes) than to those having HLA-DRB1 *03 allele (5 epitopes). CONCLUSIONS: The HLA-DR locus is involved in autoantibody expression, while the DQ locus appears to be a critical determinant for the development of type 2 AIH.     

Increased ethyl mercury load in protein a immunoadsorption

Immunoadsorption is an adsorption technique for extracorporeal removal of circulating autoantibodies in autoimmune diseases. To prevent microbial growth during storage, the protein A columns are primed with thiomersal, which contains toxic ethyl mercury, which may be released during the procedure and potentially begin to accumulate and become toxic. To reduce the thiomersal-related mercury release during immunoadsorption treatment, we introduced a modified rinsing solution containing N-acetylcysteine, which is an avid mercury scavenger. Thirteen patients received 17 protein A immunoadsorption treatments and their venous blood samples were collected immediately before and after each session. The total blood mercury levels were measured by atomic absorption spectrometry, and the ethyl mercury levels by atomic fluorescence spectrometry. Following the manufacturer's recommendations, we used 600 mg of N-acetylcysteine to rinse the mercury from protein-loaded columns before each immunoadsorption treatment. After immunoadsorption, the ethyl mercury levels increased from 0.148 +/- 0.402 ng/g to 2.026 +/- 1.944 ng/g (P < 0.001), and the total blood mercury levels increased from 2.447 +/- 3.065 ng/g to 20.437 +/- 28.603 ng/g (P = 0.02). The post-treatment values of total blood mercury exceeded the upper safety level of 5 ng/g in all 17 immunoadsorption treatments, but no patient developed clinical signs of mercury toxicity. The results of our study showed an increase in total blood mercury and ethyl mercury levels during the immunoadsorption treatments, suggesting mercury release from thiomersal-primed columns despite the addition of N-acetylcysteine to the rinsing solution.     

PentraSorb C‐Reactive Protein: Characterization of the Selective C‐Reactive Protein Adsorber Resin

C‐reactive protein (CRP) is well known as a general marker of inflammation. It furthermore represents a reliable risk factor for cardiac events and mediates tissue damage in acute myocardial infarction (AMI). It has been demonstrated that selective CRP depletion by extracorporeal apheresis in a porcine AMI model had beneficial effects on the infarcted area and the cardiac output. We therefore developed a novel adsorber for CRP apheresis from human plasma (PentraSorb CRP). It is intended for use in the clinic as therapy for patients suffering from AMI or other acute inflammatory diseases with elevated CRP plasma levels. The PentraSorb resin specifically bound CRP from human blood plasma and almost no other proteins as determined via Sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS‐PAGE). The resin further efficiently and selectively depleted CRP from plasma with low as well as high CRP concentrations (10–100 mg/L) at different flow rates, ranging from 17 to 40 mL/min. The resin was regenerable for up to 200 times without losing its CRP binding capacity or affecting biocompatibility. The depletion of CRP from plasma was comparable between the utilized small‐scale column (0.5 mL resin) and the PentraSorb CRP adsorber (20 mL resin volume). The established features can therefore be applied to the clinical setting. In summary, PentraSorb CRP provides a novel, specific, and efficient CRP‐binding resin that could be used in apheresis therapy for patients suffering from inflammatory diseases such as AMI, stroke, acute pancreatitis, and Crohn's disease.     

Targeting anti‐beta‐1‐adrenergic receptor antibodies for dilated cardiomyopathy

Anti‐beta‐1‐adrenergic receptor antibodies (anti‐β₁AR Abs) have long been implicated in the pathogenesis of dilated cardiomyopathy (DCM). It is believed that these autoantibodies bind to and constitutively stimulate the β₁AR to promote pathological cardiac remodelling and β₁AR desensitization and downregulation. The prevalence of anti‐β₁AR Abs in patients with DCM ranges from 26% to 60%, and the presence of these autoantibodies correlates with a poor prognosis. Several small studies have shown improvements in functional status, haemodynamics, and biomarkers of heart failure upon removal or neutralization of these antibodies from the sera of affected patients. Traditionally, removal of anti‐β₁AR Abs required immunoadsorption therapy with apheresis columns directed against human immunoglobulins (Igs) and subsequent i.v. Ig infusion, thereby essentially performing a plasma exchange transfusion. However, recent advances have allowed the development of small peptides and nucleotide sequences that specifically target and neutralize anti‐β₁AR Abs, providing a hopeful avenue for future drug development to treat DCM. Herein, we briefly review the clinical literature of therapy directed against anti‐β₁AR Abs and highlight the opportunity for further research and development in this area.     

Correcting immune imbalance: the use of Prosorba column treatment for immune disorders

The treatment of selected refractory autoimmune diseases has been complemented by the use of Protein A (Prosorba column) immunoadsorption. US Food and Drug Administration-approved clinical applications include idiopathic thrombocytopenia purpura (ITP) and rheumatoid arthritis (RA). Other common off label uses include thrombotic thrombocytopenic purpura (TTP) and hemolytic uremic syndrome (HUS). Less common experimental uses in diseases in which efficacy has been reported include autoimmune CNS syndromes, peripheral neuropathies, autoimmune pancytopenia, hemolytic anemia and solid organ transplant rejection. Prosorba column treatment is generally well tolerated but a small proportion of treated patients experience chills, fever, tremor, hypotension and rash. The mechanism of action suggested for the efficacy of the column is the restoration of normal immune balance and normal tolerance. Observations in ITP has suggested that column treatment stimulates a rise in anti-idiotype antibody directed against antiplatelet antibodies, effecting a decrease in pathogenic antiplatelet antibodies and immune complexes.     

Background and Indications for Protein A‐Based Extracorporeal Immunoadsorption

Abstract: Protein A (SPA), a major cell wall component of Staphylococcus aureus, has occupied numerous investigators from its discovery in the late fifties. Its availability and avid binding to human immunoglobulins have led to extensive usage for diagnostic and research purposes. Today, SPA‐based extracorporeal immunoadsorption relies on two rather different systems, namely, SPA‐silica (Prosorba), and SPA‐Sepharose (Immunosorba). Both systems are approved by the Food and Drug Administration for the core indications of rheumatoid arthritis and idiopathic thrombocytopenic purpura (SPA‐silica) or hemophilia with inhibitors (SPA‐Sepharose). Off label indications include immune disorders with a conceivable connection between autoantibody titers and disease activity, like forms of glomerulonephritis, systemic lupus erythematodes, myasthenia, and the Guillain‐Barré syndrome as well as alloantibody formation in the context of e.g., transplantation. This review summarizes historical developments and important properties of SPA. Indications for extracorporeal therapy are discussed on the basis of available information and personal experience.     

Technical basis for immunoadsorption

Immunoadsorption is a new extracorporeal treatment for immunologicaly‐mediated diseases. This review deals with the primary plasma separation for immunoadsorption, the various immunoadsorbers and the technique of immunoadsorption itself. Primary plasma separation can be performed either by plasma membrane filtration or by centrifugation depending on the quantity of processed plasma. Currently, there are non‐selective, semi‐selective and highly selective adsorbers commercially available. Adsorbers can be subdivided into single‐use and reusable devices. The choice of the adsorber is dependent on the clinical situation of the patient. An adapted anticoagulant regimen must be used to avoid complications in the patient and to obtain optimal adsorber performance. Thus, immunoadsorption is an effective and safe extracorporal treatment in severely ill patients suffering from autoimmune diseases.     

Immunoglobulin Binding Properties of the Prosorba Immunadsorption Column in Treatment of Rheumatoid Arthritis

Abstract: Studies of the humoral effects of the Prosorba column were conducted in conjunction with the Phase 3 trial of Prosorba versus sham therapy for rheumatoid arthritis (RA). When perfused with normal human plasma in vitro, Prosorba bound predominantly IgG with a maximal capacity of approximately 462 g of Ig per Prosorba column, equal to about 1.5% of circulating IgG. Prosorba treatment did not alter the concentrations of albumin, IgG, IgM, and IgA in 3 RA patients, except for a small dilutional effect. Kinetic studies demonstrated that Prosorba removed IgG > IgM, IgA, and IgM rheumatoid factor (RF) during the initial moments of apheresis and almost exclusively IgM RF after 15 min. No net protein removal occurred at 60 min. Mean values of circulating immune complexes (CICs) were not significantly decreased by 12 weekly treatments. Complement was activated by the apheresis system upstream of the Prosorba column without changing C3 or C4 levels. We conclude that the Prosorba mechanism of action in RA is not bulk removal of Ig, but might involve modification of the CIC repertoire and could include, but not be limited to, effects related to complement activation.     

Remission of demyelinating polyneuropathy with immunoadsorption, low dose corticosteroids and anti-CD20 monoclonal antibody

Demyelinating polyneuropathy with anti-myelin associated glycoprotein (anti-MAG) antibodies is an immune mediated disorder characterized by proximal and distal symmetric weakness. Electrophysiological measurements depict features characteristic for demyelination, including prolonged distal latency, retarded conduction velocity, delayed or absent F-waves, and, rarely, partial conduction block. We report on a 65-year-old patient who was diagnosed with demyelinating polyneuropathy and anti-MAG antibodies five years before admission. Despite immunosuppressive agents and extracorporeal therapy (plasmapheresis) the disease progressed as assessed by clinical symptoms and neurological tests. Laboratory results showed an increase of serum immunoglobulin M and anti-MAG antibodies over time. Because of progressive disease we decided to treat the patient with immunoadsorption followed by application of the anti-CD20 antibody, rituximab. Six cycles of selective immunoadsorption were performed over three-weekly intervals with a repetitively used column (Globaffin); each cycle consisted of four consecutive daily treatments. Starting at cycle 4 the anti-CD20 antibody rituximab was administered with 375 mg/m(2) after immunoadsorption. The pretreatment anti-MAG antibody level of 10,000 U/mL, indicating disease activity, initially increased during treatment to a maximum of 30,559 U/mL. However, after completion of the six cycles, the anti-MAG level had decreased to 2348 U/mL; 16 months after the last immunoadsorption cycle the anti-MAG level had increased to 4134 U/mL, while the conduction velocity and compound motor action potentials remained stabile. Immunoadsorption in combination with a monoclonal anti-CD20 antibody in patients with demyelinating polyneuropathy with anti-MAG is effective and can be used an alternative treatment option in patients with progressive disease.     

Increased frequency of organ-specific cardiac antibodies in healthy relatives of patients with dilated cardiomyopathy: Evidence for autoimmunity in polish families

Background and hypothesis: Autoantibodies represent markers of autoimmune involvement and are found with increased frequency in patients and their symptom-free relatives at risk compared with normal controls. Cardiac-specific autoantibodies, detected by immunofluorescence, were found in 20% of symptom-free relatives of patients with dilated cardiomyopathy (DCM) from England and Italy. The role of autoimmunity may vary in DCM patients from Poland due to ethnic differences in genetic susceptibility to autoimmune disease. Methods: We assessed the frequency of the organ-specific cardiac autoantibodies in 162 symptom-free relatives of DCM patients [85 male, mean (SD) age 27 (18) years] and 80 control subjects from Poland. Familial DCM (>1 affected member) was present in 4 families, nonfamilial DCM in the remaining 24 pedigrees. We performe Results: The frequency of cardiac-specific autoantibodies was higher among patients with documented DCM (probands and relatives) (50%) and their symptom-free relatives (38%) than in unrelated normal subjects (10%;p=0.0001). In 24 (86%) of the pedigrees studied, autoantibodies were found in the proband and/or in at least one family member and tended to be more common in familial than in nonfamilial DCM (50 vs. 35%, p=NS). Echocardiographic indices of left ventricular size and function were similar in relatives with and without detectable antibodies.d antibody screening and noninva-sive cardiological assessment in the whole group. Conclusions: The presence of cardiac-specific autoantibodies in symptom-free relatives of DCM patients provides evidence for autoimmunity in the majority (86%) of our pedigrees, including both familial and nonfamilial forms of DCM.     

Serum anti-p53 antibodies do not occur in patients with heart failure due to idiopathic dilated and ischemic cardiomyopathies

BACKGROUND: P53 is a key protein which controls cell cycle arrest and apoptosis in response to DNA damage. Auto-antibodies against p53 have been detected in some cancer patients and also in patients with autoimmune diseases. In these patients, the main cause of anti-p53 antibody occurrence was considered to be increased intracellular p53 protein in cancer cells and autoreactive lymphocytes, respectively. Intracellular p53 also increases with cardiomyocyte apoptosis during heart failure and autoreactive lymphocytes play a role in the course of idiopathic dilated cardiomyopathy (IDC) and ischemic cardiomyopathy (ICM). Based on these observations, we hypothesized that anti-p53 antibody response may also occur in patients with heart failure due to ICM and IDC. AIM: The aim of this study was to evaluate anti-p53 antibodies in the serum of patients with heart failure due to IDC and ICM. METHODS: 70 eligible patients with heart failure and severe left ventricular systolic dysfunction (mean fractional shortening 12.03 +/- 3.93%) were included in the study. The aetiology of heart failure was IDC in 26 patients and ICM in 44 patients, according to the angiographic and echocardiographic findings. RESULTS: Anti-p53 antibodies were not detected in any of the patients. CONCLUSION: Anti-p53 antibodies do not occur in patients with heart failure due to IDC and ICM, possible explanations are discussed in the text.     

Immunoadsorption as a Tool for the Immunomodulation of the Humoral and Cellular Immune System in Autoimmune Disease

Immunoadsorption onto staphylococcal protein A is a newly developed semiselective extracorporeal adsorption technique for immunoglobulins applied in patients suffering from severe autoimmune disease. Its effect on the humoral and cellular immune system was investigated using standard immunological assays. The elimination capacity for total IgG and IgG subclasses 1, 2, and 4 was more than 90% but for subclass IgG3 varied between 30 and 90%. Autoantibodies, e.g., anti-dsDNA, anti-glomerular basement membrane (anti-GBM), anti-cardiolipin, and anti-human leukocyte antigen (anti-HLA) antibodies, were eliminated in comparable amounts. The affinity of protein A for circulating immune complexes (CIC) was 300 times greater than for soluble IgG. HLA-II expression on monocytes and T lymphocytes was reduced over time during repeated IAs (IA). The number of activated T lymphocytes declined while the percentage of naive T cells increased. A diminished CD4/CD8 ratio normalized during IA treatment. These results indicate that IA actively modulates the humoral as well as the cellular immune system in addition to its immunoglobulin reducing effect.     

Ig-Therasorb Immunoadsorption for Selective Removal of Human Immunoglobulins in Diseases Associated with Pathogenic Antibodies of All Classes and IgG Subclasses,Immune Complexes,and Fragments of Immunoglobulins

Abstract: Therasorb immunoadsorption (IA), by selectively eliminating pathogenic substances from the circulation, allows for successful therapy of previously insufficiently treatable diseases. Molecules (specific polyclonal sheep antibodies) coupled to a matrix (Sepharose CL 4B) selectively bind plasma components in extracorporeal circulation. This procedure has been established in the treatment of various diseases. Examples are familial hypercholesterolemia (LDL-Therasorb) and selected autoimmune diseases (Ig-Therasorb). Ig-Therasorb IA has been performed in a variety of clinical indications, primarily in the treatment of autoimmune diseases. In most cases, Ig-Therasorb IA has been applied in patients who have failed to respond to conventional therapy with a high rate of clinical improvement. In defined groups of patients with autoimmune diseases and alloantibodies, immunoadsorption can now be considered an established therapeutic means. The fast and efficient removal of immunoglobulins obviously exceeds the efficiency of conventional plasma exchange by far. Autoimmune diseases could be induced by balanced and nonbalanced immunity. The importance of autoantibodies remains unclear, but the efficacy of Ig-Therasorb IA suggests a key role for them. In addition to the established indications for removal of immunoglobulins, there may be a number of promising new indications.