Efficacy, safety, tolerability and pharmacokinetics of a novel human immune globulin subcutaneous, 20%: a Phase 2/3 study in Europe in patients with primary immunodeficiencies.

A highly concentrated (20%) immunoglobulin (Ig)G preparation for subcutaneous administration (IGSC 20%), would offer a new option for antibody replacement therapy in patients with primary immunodeficiency diseases (PIDD). The efficacy, safety, tolerability and pharmacokinetics of IGSC 20% were evaluated in a prospective trial in Europe in 49 patients with PIDD aged 2-67 years. Over a median of 358 days, patients received 2349 IGSC 20% infusions at monthly doses equivalent to those administered for previous intravenous or subcutaneous IgG treatment. The rate of validated acute bacterial infections (VASBIs) was significantly lower than 1 per year (0·022/patient-year, P < 0·0001); the rate of all infections was 4·38/patient-year. Median trough IgG concentrations were ≥ 8 g/l. There was no serious adverse event (AE) deemed related to IGSC 20% treatment; related non-serious AEs occurred at a rate of 0·101 event/infusion. The incidence of local related AEs was 0·069 event/infusion (0·036 event/infusion, when excluding a 13-year-old patient who reported 79 of 162 total related local AEs). The incidence of related systemic AEs was 0·032 event/infusion. Most related AEs were mild, none were severe. For 64·6% of patients and in 94·8% of IGSC 20% infusions, no local related AE occurred. The median infusion duration was 0·95 (range = 0·3-4·1) h using mainly one to two administration sites [median = 2 sites (range = 1-5)]. Almost all infusions (99·8%) were administered without interruption/stopping or rate reduction. These results demonstrate that IGSC 20% provides an effective and well-tolerated therapy for patients previously on intravenous or subcutaneous treatment, without the need for dose adjustment.

Rapid subcutaneous immunoglobulin administration every second week results in high and stable serum immunoglobulin G levels in patients with primary antibody deficiencies.

Subcutaneous immunoglobulin G (SCIG) infusions as life-long replacement therapy in patients with primary antibody deficiences (PAD) is being applied increasingly. However, only a few published pharmacokinetic studies are available for this route of administration. Therefore, the pharmacokinetics of a 16% immunoglobulin G (IgG) preparation intended for subcutaneous use were investigated in patients with common variable immunodeficiency and X-linked agammaglobulinaemia. SCIG infusions (200 mg/kg body weight) were administered to 12 adult patients every 14 days for 24 weeks (total of 144 infusions). Pharmacokinetic parameters were determined based on serum IgG trough levels and antibody levels against tetanus. The median half-life of the total serum IgG and for the tetanus antibodies was 40.6 and 23.3 days respectively. Median in vivo recovery of serum IgG and tetanus immunoglobulins were 36% and 46% respectively. Median, preinfusion serum IgG trough levels per patient were high without major variations between infusions and ranged from 7.24 to 7.86 g/l. Safety, in terms of adverse events including systemic adverse reactions and local tissue reactions at infusions sites, was monitored throughout the study. Six mild, local tissue reactions were observed during the study in one patient. No systemic adverse reactions related to the study drug were observed and no serious other adverse event occurred during the study. It is concluded that the bi-weekly SCIG therapy was well tolerated in the study and that it results in high and stable serum IgG levels, offering an alternative therapy regimen to patients suffering from PAD.

Prospective open-label study of pharmacokinetics, efficacy and safety of a new 10% liquid intravenous immunoglobulin in patients with hypo- or agammaglobulinemia.

BACKGROUND AND OBJECTIVES: The aim of this study was to evaluate the pharmacokinetics, efficacy and safety of a newly developed 10% liquid immunoglobulin preparation in patients with primary immunodeficiency diseases. This new preparation for intravenous use includes three dedicated virus clearance steps in its manufacturing process to ensure a high margin of viral safety. MATERIALS AND METHODS: This was a prospective, open-label, non-controlled, multicentre study. Twenty-two subjects with primary immunodeficiency were treated initially with three infusions of a licensed intravenous immunoglobulin to standardize the immunoglobulin G (IgG) replacement therapy of all subjects to the same intravenous product. A total of nine infusions of the new 10% liquid preparation were subsequently administered. RESULTS: The median terminal half-life of total IgG following administration of the new preparation was 30.1 days. Median terminal half-lives for IgG subclasses IgG(1), IgG(2), IgG(3) and IgG(4) were 28.3, 31.3, 20.9 and 24.2 days, respectively. The median total serum IgG steady-state trough level was 8.51 g/l. No severe infection episodes started after initiation of treatment with the new preparation. The median rate of mild or moderate infection episodes was 0.48 per month. A total of 194 infusions with the new 10% liquid immunoglobulin preparation were administered. The mean dose per infusion was 0.41 g/kg body weight and the maximum infusion rates recorded were 8 ml/kg/h. Adverse experiences were mostly mild and unrelated to the study drugs. Only 4% of infusions with the new product were followed by one or more related adverse experiences. CONCLUSION: The new 10% liquid immunoglobulin preparation was well tolerated and shown to have an excellent pharmacokinetic, efficacy and safety profile. The liquid formulation provides convenience to patients and healthcare professionals.

Tetanus antibody levels of female volunteers after injection with solvent/detergent-treated human tetanus immunoglobulin (Tetabulin S/D).

BACKGROUND AND OBJECTIVES: The aim of this study was to analyse the increase of antitetanus titre in volunteers following injection with human tetanus immunoglobulin (HTI). MATERIALS AND METHODS: Twelve females with tetanus antibody titres of < or = 0.05 international units (IU)/ml were injected with 500 IU of human tetanus immunoglobulin (Tetabulin S/D). The tetanus antibody titres were determined before injection, and after 30 h, 48 h and 4 days. RESULTS: A fast and sustained increase of protective tetanus antibody levels was observed in 10 of 12 volunteers. No adverse events related to the study drug were reported. CONCLUSIONS: HTI confers rapid and effective immunity to tetanus.

Pharmacokinetics of human tick-borne encephalitis virus antibody levels after injection with human tick-borne encephalitis immunoglobulin, solvent/detergent treated, FSME-BULIN S/D in healthy volunteers.

This study assessed tick-borne encephalitis virus (TBEV) neutralizing antibody levels after injection of FSME-BULIN S/D (human tick-borne encephalitis immunoglobulin; 0.2 ml/kg body weight) in healthy volunteers. After screening of 18 volunteers for TBEV antibody titers, 12 healthy volunteers with TBEV antibody titers < 5 were entered into the pharmacokinetic part of the study. TBEV antibody titers were analyzed before injection and after 24 h, 48 h, 3 d, 4 d and 8 d. Vital signs, adverse events and laboratory tests for safety were analyzed after intramuscular injection with the immunoglobulin at 4 sites in the gluteal muscles. Injection with 0.2 ml/kg of FSME-BULIN S/D induced a fast increase in, and sustained titers of, neutralizing antibody levels against TBEV. The injections were well tolerated and the safety profile of the product was fully acceptable.

Humoral and cellular immunity induced by antigens adjuvanted with colloidal iron hydroxide.

The immunopotentiating activities of colloidal iron hydroxide, a novel, experimental mineral adjuvant, and of aluminium hydroxide. the licensed adjuvant for human vaccines, were compared. Our studies revealed that colloidal iron hydroxide and aluminium hydroxide behaved comparably with respect to supporting induction of an antibody response to tetanus toxoid. Furthermore, mice immunized with both, the experimental vaccine (tick-borne encephalitis virus (TBEV) antigen adsorbed to colloidal iron hydroxide) or with a commercially available TBEV vaccine (adjuvanted with aluminium hydroxide), developed long-lasting antibody responses which protected the animals from TBEV infection even one year after vaccination. The use of colloidal iron hydroxide as adjuvant had the additional advantage to reproducibly support induction of HIV-1 envelope-specific cytotoxic T lymphocytes (CTL), when used as adjuvant for a HIV-1 env-carrying recombinant fowlpox virus and being applied via the subcutaneous route. Aluminium hydroxide was much less active in this respect. Non-adjuvanted recombinant fowlpox elicited CTLs only when given intravenously or intraperitoneally, vaccination routes considered not to be suitable for routine use in humans. Further studies to evaluate the use of colloidal iron as possible alternative and/or supplement for routinely used mineral adjuvants may therefore be warranted.

Adjuvant/carrier activity of inactivated tick-borne encephalitis virus.

Virus-like or virus-derived particles have been reported to increase the immunogenicity of foreign antigens. In this study formaldehyde-inactivated tick-borne encephalitis virus (TBEV), a potent immunogen in humans, was tested for possible adjuvant/carrier function. The results of our study revealed that substantial antibody titers against very low doses of tetanus toxoid could be obtained when mice were immunized with the antigen covalently coupled to TBEV (using N-succinimidyl-3-(2-pyridyldithio)propionate, a heterobifunctional, cleavable crosslinker containing a disulfide bridge). In contrast, only moderate anti-tetanus toxoid titers were induced by immunizations with a simple mixture of low dose tetanus toxoid and TBEV or when the disulfide bridge of the crosslinker used to couple tetanus toxoid to TBEV was cleaved prior to immunization. The antibody response to TBEV, on the other hand, was not influenced by its linkage to tetanus toxoid. Comparable anti-TBEV titers were obtained following immunization of the animals with either the TBEV-tetanus toxoid conjugate or the mixture of tetanus toxoid and TBEV. Prior application of a TBEV vaccine did not change the antibody response against tetanus toxoid and thus carrier-induced epitopic suppression could be ruled out. The abovementioned adjuvant/carrier properties of TBEV might make it a suitable candidate for use in bi- or multivalent vaccines containing weak immunogens.

Method for the isolation of biologically active monomeric immunoglobulin A from a plasma fraction.

A purification method for immunoglobulin A (IgA) yielding monomeric IgA with a purity of over 97% has been developed. This procedure uses ethanol-precipitated plasma (Cohn fraction III precipitate) as the starting material and includes heparin-Sepharose adsorption, dextran sulfate and ammonium sulfate precipitation, hydroxyapatite chromatography, batch adsorption by an anion-exchange matrix and gel permeation. Additional protein G Sepharose treatment leads to an IgA preparation of greater than 99% purity. The isolated IgA presented with an IgA subclass distribution, equivalent to IgA in unfractionated plasma, and was biologically active, as was shown by its ability to down-modulate Haemophilus influenzae-b-induced IL-6 secretion of human monocytes.

Affinity binding of distinct functional fibronectin domains to immobilized metal chelates.

Fibronectin has been found to bind metal ions. Using metal chelate chromatography and limited proteolysis to generate the distinct functional domains of fibronectin we set out to address the metal binding sites to well-defined regions of fibronectin. The results show that the affinity binding of fibronectin to Co2+ is mediated exclusively via the collagen binding domain of the molecule, whereas fibronectin will bind to Zn2+, Ni2+, and Cu2+ by metal binding sites located in two, three, and four well-defined regions of fibronectin, respectively. Fe2+ and Mn2+ chelates did not bind any of the isolated fibronectin domains. Combined metal chelate affinity chromatography opens a possibility to isolate particular fibronectin domains.

Separation of polysaccharide-specific human immunoglobulin G subclasses using a protein A superose column with a pH gradient elution system.

Protein A Superose was employed to separate affinity-purified anticarbohydrate antibodies according to immunoglobulin G (IgG) subclass. Separation was achieved with a novel buffer system (disodium phosphate-sodium acetate-sodium chloride-glycine), which allowed the generation of a linear pH gradient from pH 8 to 3. Protein A-bound anti-carbohydrate antibodies were eluted as three peaks, two of them mainly containing IgG2 and one consisting of highly enriched IgG1. The enriched antibody preparations retained their functional activity. This separation procedure can be considered as an alternative to the preparation of IgG subclasses with subclass-specific monoclonal antibodies and could be employed whenever contamination with immune complexes has to be avoided.

IgG subclass determination in human sera with commercially available reagents: comparison of different assay systems.

IgG subclass determinations are of increasing importance for the diagnosis of humoral immunodeficiencies. The search for a method which is accurate, reliable and suitable for the clinical routine, while utilizing commercially available reagents, was the aim of this study. Different assay systems for determination of IgG subclasses were compared. Radial immunodiffusion with polyclonal antisera (RIDpoly) proved to be a reliable method for subclass determination in individual human sera. Sera deficient in one or several IgG subclasses as well as several myeloma proteins were readily and reliably detected. The RID method with monoclonal antibodies (RIDmono) yielded results comparable to those obtained with RIDpoly in the IgG1 and IgG2 determination. Differences between RIDpoly and RIDmono were observed, however, in the determination of IgG3 and IgG4. These discrepancies were shown to be due to differences in the calibration of the standards as given by the manufacturers, and not to different recognition of distinct allotypes. The results of an enzyme immunoassay (EIA) using commercially available IgG reagents without further purification did not compare satisfactorily with the results of the RIDpoly method. These discrepancies, however, were assay-inherent rather than monoclonal reagent-inherent.

IgG subclasses and antibody response to pneumococcal capsular polysaccharides in children with severe sinopulmonary infections and asthma.

A prospective open study was carried out on 30 pediatric patients with most severe chest disease whose serum immunoglobulin levels were normal. The patients entered into the study had had two or more documented episodes of pneumonia, and/or six episodes of bronchitis with fever within a year, and/or severe asthma (steroid-dependent), and/or hospitalization for chest disease for more than 30 days within the year preceding the study. Eleven patients had sinopulmonary infections, 19 had asthma. Twenty patients had low levels of one or two IgG subclasses: 11 were deficient in IgG3, three in IgG4, three in IgG3 + IgG4, and three in IgG2 + IgG4. Patients with low IgG subclass levels were distributed throughout the different clinical entities. These children had significantly longer periods of hospitalization than the patients in whom all IgG subclasses were within the normal range. They suffered more often from sinopulmonary infections. Asthmatic children with low levels of an IgG subclass reported more days with wheezing and needed more steroids than the children without subclass deficiencies.

Determination of cyclosporine by a competitive binding assay with cyclophilin.

A competitive protein-binding assay for cyclosporine based on use of the intracellular cyclosporine-binding protein cyclophilin (CYP) was used to measure cyclosporin A (CsA) and its bioactive metabolites in whole blood. CYP from cytoplasmic extracts or erythrocyte lysates was applied in the binding assay with use of [3H]CsA as tracer and charcoal adsorption for separating bound from free tracer. Binding affinities of various CsA analogs and metabolites correlated well with their reported in vitro immunosuppressive activities. The assay detected as little CsA as 50 micrograms/L (1 g = 0.832 mmol of CsA), analytical recovery was greater than 80%, and CVs were less than 8% for intra-assay and less than 11% for interassay precision in the range of 150-1000 micrograms/L. We used this assay to measure CsA concentrations in blood and compared the results with those measured by HPLC or by CsA-specific (monoclonal) and CsA-nonspecific (polyclonal) radioimmunoassays. Binding assay results were, in nearly all cases, less than those measured by the nonspecific RIA and frequently greater than 20% above the values determined by the CsA-specific assays. Individual patients had pronounced differences in the relative proportions of CsA, CYP-binding (bioactive) metabolites, and cross-reacting CsA metabolites. Because the presence of bioactive metabolites may considerably contribute to the immunosuppressive activity of CsA, we consider the binding assay clinically useful for measuring CsA in biological fluids.

Increased Fc-receptor-mediated clearance via reticuloendothelial system in patients with nephrotic syndrome.

Reticuloendothelial phagocytic cell function, which is an important participant in host defense, was investigated by studying the Fc-receptor-mediated clearance of IgG-anti-RhD-sensitized radiolabelled autologous erythrocytes in patients with nephrotic syndrome known to have an increased risk of infectious complications due to hypogammaglobulinemia. Patients with serum IgG concentrations less than 500 mg/dl had a significantly accelerated Fc-receptor-mediated clearance (t1/2 = 52 +/- 3.6 min) compared to patients with IgG levels greater than 500 mg/dl (t1/2 = 93 +/- 17 min) or healthy controls (t1/2 = 100 +/- 7.5 min). Serum IgG concentrations correlated significantly with Fc-receptor-mediated clearance (t1/2: r = 0.70; p = 0.035) in patients with nephrotic syndrome. These results indicate that Fc-receptor-mediated clearance by the reticuloendothelial system is enhanced in patients with nephrotic syndrome with low serum IgG concentrations and may, at least in part, compensate for reduced opsonic antibody concentration.

Multiple infusions of human intravenous immunoglobulin in chimpanzees do not lead to immune elimination.

Administration of human i.v. immunoglobulins was shown to lead to a permanent increase in IgG1 and IgG2 levels in chimpanzees. Half-lives of human IgG1 and IgG2 in chimpanzees were comparable to those found in humans, and no signs of immune elimination were observed. Furthermore, long-term treatment of chimpanzees had no effect on the percentage of immunoregulatory T cells (CD2+, CD4+ and CD8+ T cells) as determined by FACS analysis. In addition, serum IgM levels in chimpanzees were found to be comparable to those in humans, whereas the chimpanzees' IgG levels are slightly elevated due to higher concentrations of IgG2 and, in particular, IgG4.

Serum immunoglobulin G subclasses in patients during acute hepatitis A.

IgG subclasses were measured in sera of 47 patients with acute hepatitis A during the course of the disease. IgG1 and IgG3 serum levels were found to be elevated, whereas IgG2 and IgG4 subclass concentrations did not differ from that found in healthy control individuals. These findings indicate that, similar to the specific antiviral antibody, the polyclonal increase of serum concentrations of IgG in acute hepatitis A is not equally distributed to all IgG subclasses but is restricted to IgG1 and IgG3.

[Antibody deficiency in obstructive lung diseases]. / Antikörperdefizienz bei obstruktiven Lungenerkrankungen.

Severe chronic chest disease is the most serious complication in patients with antibody deficiency syndromes. IgG-subclass-deficiencies were a frequent finding in patients with obstructive lung disease and in patients with sinopulmonary infections. In the patient population referred to us for immunological investigation recurrent infection of the upper and lower respiratory tract was the most common reason to suspect undue susceptibility to infection. In 1034 pediatric patients analyzed 299 were found to be deficient in one of several IgG subclasses. In a group of 30 children, all of whom had severe lung disease and normal concentrations of serum IgG, IgA and IgM, airway-obstruction has been diagnosed in 19. 20 of the 30 patients had IgG-subclass-deficiency. The large percentage of IgG-subclass-deficiencies in this group of patients indicates that immunological disregulation is likely to contribute to the pathogenesis of chronic lung disease.