Clinical efficacy, safety and pharmacokinetic properties of the factor VIII concentrate Haemoctin SDH in previously treated patients with severe haemophilia A.

Clinical efficacy, safety and pharmacokinetic properties of the high-purity double-virus inactivated plasma-derived factor VIII concentrate Haemoctin SDH (pdFVIII) were evaluated in three prospective open-label uncontrolled studies in previously treated patients (PTPs) with severe haemophilia A. The pharmacokinetic properties assessed at baseline and after 3 months of treatment are in accurate accordance with published data and remain unchanged over time (study A, n = 12). Mean terminal elimination half-life was 11.8 and 11.9 h, mean incremental recovery (IU dL(-1)/IU kg(-1)) was 2.3 and 2.0, respectively. Long-term efficacy and safety, in particular the potential immunogenicity, were investigated in a total of 53 PTPs (studies A and B) treated prophylactically and on-demand, as required. PdFVIII has shown to be effective in preventing and controlling bleeding episodes; 23.5% of patients were free of bleeding events. A total of 177 haemorrhages occurred with 74.0% resolving after a single infusion, 87.6% within two infusions. 98.3% of responses reported on haemorrhages were rated as 'excellent' or 'good'. Moreover, 'excellent' haemostatic efficacy has been demonstrated in 10 surgical procedures including general and severe orthopaedic interventions (study C). No complication occurred in any surgery. Few adverse events were reported, one patient developed a high-titre FVIII inhibitor without clinical relevance. In all three studies, over 6 million units were administered in nearly 4300 infusions, approximately 94% units or infusions were given for prophylaxis and only 6% for treatment on-demand. In conclusion, pdFVIII has shown to be effective, safe and well tolerated in long-term prophylaxis and treatment on-demand as well as after minor and major surgical procedures.

Comparison of fresh frozen plasma with a standardized serum protein solution following therapeutic plasma exchange in patients with autoimmune disease: a prospective controlled clinical trial.

The aim of the study was the comparison of the influence of fresh frozen plasma (FFP) (Freiburg, Germany) and Biseko, Biotest Pharma GmbH (Dreieich, Germany), as a plasma substitute (a standardized, virus inactivated human serum protein solution) on the coagulation factors, inhibitors, proteins, and complement factors in the plasma of autoimmune disease patients following membrane plasma separation. Patients (n = 24) with autoimmune disease were randomized to receive either FFP or Biseko for membrane plasma separation therapy. During each plasma exchange, 100% of the plasma volume was replaced by the respective substitute. Plasma exchange volume was performed once daily for 3 days. Target test parameters of the coagulation system were fibrinogen, fibrinopeptide A, factor VIII (FVIIIC), von Willebrand factor antigen (vWFAg), partial thromboplastin time (PTT), thromboplastin time (Quick value), and antithrombin (AT III). The immunoglobulins were IgG, IgA, and IgM and C-reactive protein (CRP). The thrombocytes were platelet factor 4 (PF4), and complement factors were C3 and C4. Biseko was well tolerated with 1 mild adverse drug reaction (ADR) (n = 1) while FFP gave rise to ADR on 7 occasions (n = 4). Statistically significant differences in the 2 groups were observed for fibrinogen, PTT, Quick value, and AT III. From the clinical point of view, all fluctuations and differences in parameter levels remained clinically silent. The differences had no clinical consequences. Reflecting on a potential decrease in the risk of infections in comparison to FFP therapy and the lower rate of adverse drug reactions, it is possible to postulate an advantage of Biseko for plasma exchange therapy.

Improvement of virus safety of a S/D-treated factor VIII concentrate by additional dry heat treatment at 100 degrees C.

In order to increase the virus safety of a solvent/detergent-treated Factor VIII concentrate in regard to non-lipid coated viruses and to respond to the continuous discussion about reports on hepatitis A transmission by Factor VIII preparations, we have investigated the effect of a terminal dry heat treatment (30 min 100 degrees C) on HAV and various other viruses. By this treatment Hepatitis A virus was inactivated below detectable level after a few minutes (> 5.3 log10). Other RNA viruses such as the Human Immunodeficiency Virus (> 6.6 log10), bovine viral diarrhoea virus (> 6.6 log10) and vesicular stomatitis virus (> 5.8 log10) were also inactivated below detectable level. Pseudo rabies virus and reovirus Type 3 are inactivated by 5.7 and > 6.0 log10, respectively. SV40 and bovine parvo virus showed significant resistance to dry heat treatment. We conclude that the involvement of two strong virus inactivation steps, acting by different mechanisms, improves the virus safety of Factor VIII concentrates without destroying the Factor VIII activity. Moreover, the terminal 100 degrees C heat treatment for 30 min represents an effective measure to inactivate non-lipid enveloped viruses, in particular hepatitis A, which is resistant to solvent/detergent treatment.

[IgG subclass distribution in intravenous immunoglobulin products: determination of reproducibility of a method of IgG subclass determination. Ring Study of the "Plasma Blood Components" Section of the German Society of Transfusion Medicine and Immunohematology]. / IgG-Subklassenverteilung in IVIG-Produkten: Ermittlung der Reproduzierbarkeit einer Methode der IgG-Subklassenbestimmung. Ringstudie der Sektion "Plasmatische Blutbestandteile" der Deutschen Gesellschaft für Transfusionsmedizin und Immunhämatologie (DGTI).

BACKGROUND: The working group "Blood Plasma Constituents" of the DGTI has carried out a multicenter study with human immunoglobulins for intravenous administration (IVIG) and a NIBSC serum, with the designation 67/98C, since the European Pharmacopoeia IVIG monograph gives no information for the IgG subclass determination. Aim of the ring study was 1) the determination of the IgG subclass distribution with a standardized method and 2) the determination of the IgG subclass distribution in the NIBSC serum 67/98C and to test if the NIBSC serum coded 67/98C is suited as a reference serum for the IgG subclass determination. MATERIAL AND METHODS: Ten laboratories from Europe participated in the ring study. The IgG subclasses were determined by radial immunodiffusion (RID). RESULTS: The results showed good consistency between the participating laboratories and showed distinct differences in the IgG subclass composition of the tested IVIGs. The reproducibility of the method showed coefficients of variation of approximately 10%. CONCLUSION: The NIBSC serum 67/98C is suitable as reference serum for the IgG subclass determination in sera and IVIGs.

Assessment of the Fc function in human immunoglobulins for intravenous administration. Multicenter study of the section 'Blood plasma Constituents' of the German Association for Transfusion Medicine and Immunohematology (DGTI).

BACKGROUND: The working group 'Blood plasma Constituents' of the DGTI has carried out a multicenter study with human immunoglobulins for intravenous administration (IVIG). The aim of the ring study was the determination of the reproducibility of the IVIG Fc function method proposed for the European IVIG monograph. MATERIAL AND METHODS: Eight laboratories from Europe participated in this ring study. The test of Fc function was carried out according to a standardized procedure. RESULTS: The results showed good consistency between the participating laboratories. The reproducibility of the method showed coefficients of variation of 5-10%. CONCLUSIONS: All commercial IVIG preparations met the requirement of the Fc function of > or = 80% in relation to the Fc function of the standard, an immunoglobulin preparation for intramuscular (i.m.) administration.

Safety tests of the intravenous tolerance of immunoglobulins. Multicenter study of the section 'Blood Plasma Constituents' of the German Association of Transfusion Medicine and Immunohematology (DGTI).

The Working Group 'Blood Plasma Constituents' of the DGTI has carried out four multicenter trials with human immunoglobulins for intravenous application (IVIG). The ACA method elaborated here has led to relatively consistent ACA results for commercially available IVIG preparations in the participating laboratories. On their own, the in vitro tolerance tests of the IVIGs are not sufficient for safety testing, since there is no clear correlation between the ACA values and the tolerance in a rat model in which changes of blood pressure, pulse rate, ECG and respiratory resistance after administration of IVIGs are determined. The role of polymer constituents in tolerance reactions is to be regarded as substantiated, whereas the tolerance of the dimer fractions of beta-propiolactone-treated IVIGs does not differ appreciably from that of their monomer constituents. Determination of the IgG subclass composition requires standardization. The collation of the antibody specificities to the subclasses must be clarified before making inferences with regard to the clinical significance of the subclass composition of the preparations.

[Evaluating the safety with reference to intravenous tolerance of immunoglobulins]. / Sicherheitsprüfungen zur intravenösen Verträglichkeit von Immunoglobulinen.

The section 'Plasmatic Blood Constituents' of DGTI (German Society for Transfusion Medicine and Immunohematology) performed four ACA (anticomplementary activity) ring tests with human immunoglobulins for intravenous application (IVIG). The laboratories carrying out the analyses presented relatively consistent ACA results for IVIG commercial preparations that were obtained by means of the ACA method elaborated based on these tests. The tolerance trials evaluating the IVIGs solely by means of in vitro tests are not sufficient to serve as safety controls since there is no clear correlation existing between the ACA values and their compatibility in the animal model (rat).

Quality control of immunoglobulins for intravenous administration.

Since there is as yet no official European pharmacopoeia or monograph for intravenously administrable immunoglobulins (IVIGs) and the manufacturers of IVIGs use different ACA methods to test their products until today, the study group 'Blood Plasma Constituents' of the DGTI carried out a multicenter study with six IVIGs from five manufacturers. The results of this first multicenter study have not been analysed statistically, because the participants have decided to carry out a second multicenter study using a uniform ACA method.

[IgG subclass level in healthy children]. / IgG-Subklassenspiegel bein gesunden Kindern.

In 159 healthy German children aged 3 months to 16 years IgG subclass concentrations were measured by radial immunodiffusion using polyclonal antisera. IgG-4 levels were also determined by a monoclonal ELISA kit (Binding Site). Both methods for IgG-4 determination showed a significant correlation (r = 0.91); the WHO reference serum 67/97 was used for calibration. The mean range and percentiles for age are given. IgG subclass levels were correlated with the age of the children in the order IgG-2 greater than IgG-1 greater than IgG-4, but this was not the case for IgG-3. The normal ranges of IgG subclasses were in good agreement with those of previous studies. No IgG-4 was detected by RID in only 8.8% of all the children, but in all aged 7 years and above.

Virus safe plasma exchange medium containing essential human plasma proteins.

CFFP (Coagulation Factor Free Plasma from Biotest) is manufactured under standardized conditions from pooled human plasma of more than 1000 donors per lot. It is sterilized by beta-PL/UV and has not the risks of FFP to transmit viral diseases. CFFP offers desirable advantages as a plasma exchange solution over albumin and FFP as it avoids serious depletions or deviation of normal plasma constituents.

[Subcutaneous injection of desmopressin (DDAVP) for increasing factor VIII and von Willebrand factor in plasmapheresis]. / Subkutan injiziertes Desmopressin (DDAVP) zur Erhöhung des Faktors VIII und von-Willebrand-Faktors bei Plasmapherese.

Desmopressin (DDAVP) is an effective tool for increasing factor VIII (FVIII)/von Willebrand factor (vWF) in normal subjects and in patients with haemophilia A and von Willebrand's disease. There are a few studies utilizing DDAVP to increase the FVIII/vWF yield in plasmapheresis donors. In these studies, DDAVP was given either by intranasal administration or intravenous infusion. The aim of our pilot study was to investigate the subcutaneous (s.c.) injection of DDAVP in combination with double bag plasmapheresis. 8 donors with blood group A and 5 with O were given 0.4 micrograms/kg body weight DDAVP as a s.c. injection 30 min prior to the first donation. FVIII increased 3.7 and ristocetin cofactor 2.5 the initial value in the donors. In the plasma bags, a 3 (1st bag) and 3.5 (2nd bag) fold higher FVIII level was found, when compared with baseline levels. There was only a mild decrease of blood pressure, and serious adverse effects were not observed. Our data suggest that s.c. DDAVP may be more effective and especially suited for increasing FVIII/vWF yield in plasmapheresis donors.

[Clinical relevance of immunoglobulin subclasses in children with chronic pulmonary diseases]. / Klinische Relevanz der Immunglobulin-Subklassen bei Kindern mit chronisch pulmonalen Erkrankungen.

In 52 children (age 4-19.5 years; 17 female, 35 male) with chronic chest symptoms IgG-Subclass levels were measured by radial immunodiffusion techniques. Thirteen children had proven bronchiectasis, 22 chronic bronchitis and 17 steroid dependent asthma bronchiale. Cystic fibrosis, alpha-1-antitrypsin deficiency, tuberculosis and chronic foreign bodies were excluded in each patient; all of them showed normal levels of total IgG. Four children (3 with bronchiectasis, one with steroid dependent asthma bronchiale) had a complete lack if IgG4. In two children with chronic bronchitis one showed fluctuating levels of IgG2 and IgG4 and another deficiency of IgG3. All patients with an isolated IgG4 deficiency were treated with immunoglobulins. Of 3 patients with bronchiectasis one improved, two remained unchanged as shown by positive sputum cultures and of chronic chest symptoms. One patient with steroid dependent asthma bronchiale markedly improved during immunoglobulin therapy. It is concluded, that early screening of IgG subclass deficiency is indicated in all children with chronic chest symptoms.

[Inhibitory and immunological profile of therapeutic serum protein solutions]. / Inhibitorisches und immunologisches Profil therapeutischer Serumproteinlösungen.

Biseko is prepared from pooled human plasma by specific stepwise adsorption of the coagulation factors avoiding spontaneous coagulation. Biseko is manufactured from pooled plasma from more than 1000 donors. In order to ensure its hepatitis safety, the starting plasma is cold sterilized by beta-propiolactone and UV-irradiation. The inhibitory and immunological profile of the cold sterilized Biseko was compared with another commercial serum preserve and normal serum. alpha 1-antitrypsin, alpha 2-macroglobulin and antithrombin III are present in Biseko and normal serum in their biologically active forms. A certain amount of the opsonin, fibronectin, is heparin-precipitable in normal serum and has thus retained its native character, while the fibronectin in the commercial serum preserve examined is not heparin-precipitable. Biseko contains fibronectin only in trace amounts. The IgG, IgA and IgM immunoglobulin concentrations and activities in the serum preserves are equivalent to those in normal serum. One major difference between normal serum and the cold sterilized Biseko is that metabolites from the coagulation pathways are absent in Biseko. Normal serum is not suitable for therapeutic purposes because of activated enzymes formed during coagulation. The chemical analysis of the protein pattern in Biseko resembles more fresh frozen plasma without coagulation factors than normal serum.

Inactivation of hepatitis B and non-A, non-B viruses by combined use of Tween 80, beta-propiolactone, and ultraviolet irradiation.

To assess the sterilization efficacy of a combined Tween 80, beta-propiolactone and ultraviolet irradiation procedure applied to a F VIII preparation to which an estimated 10(5.9) chimpanzee infectious doses (CID50) of hepatitis B virus had been added per ml, two chimpanzees were inoculated with 10 ml each of treated and untreated preparations. The untreated preparation, which was obtained from donors with normal alanine aminotransferase (ALT) levels, induced non-A, non-B hepatitis in both recipient animals, and delayed hepatitis B infection in one of these. Neither animal receiving the treated preparation developed either type of hepatitis. When subsequently challenged with the untreated material, both of the latter animals developed non-A, non-B and hepatitis B infection, proving their susceptibility to both types of infection. It was concluded that the combined procedure inactivated an estimated 10(6.9) CID50 of hepatitis B virus and an unknown quantity of a non-A, non-B virus. The finding of non-A, non-B virus infectivity in a pooled F VIII preparation despite careful ALT screening of plasma donors emphasizes the necessity of subjecting such preparations to sterilization procedures.

Evaluation of thrombogenicity of beta-propiolactone/ultraviolet (beta-PL/UV) treated PPSB in chimpanzees.

The thrombogenicity of beta-PL/UV-treated PPSB (factor IX concentrate) was evaluated in chimpanzees. PPSB isolated from beta-propiolactone-treated and UV-irradiated plasma was injected into chimpanzees at a dose of approximately 100 units/kg body weight. An FDA licensed PPSB preparation served as the negative control, and a preparation containing activated as well as precursor clotting factors served as the positive control. 15 minutes, 1 h, 4 h, and 24 h after the PPSB application the following parameters were determined in the chimpanzee blood: factors II, VII, IX, X, VIII, fibrinogen, AT III, thrombin coagulase, Quick value, APTT and platelet count. Neither the untreated control preparation, nor the PPSB isolated from beta-propiolactone-treated and UV-irradiated plasma, showed signs of thrombogenicity in the chimpanzee model. The positive control indicated that the chimpanzee is a suitable model for the thrombogenicity testing of activated clotting factors.

Factor VIII concentrate from cold sterilized human plasma.

beta-Propiolactone (beta-PL) in combination with UV irradiation (UV) is an established method for the sterilization of factor IX concentrate or stabilized human serum. Because of the extreme sensitivity of factor VIII to beta-PL, the standard beta-PL/UV procedure cannot be used to obtain hepatitis-safe factor VIII concentrate. It has been shown in chimpanzees that from a cryoprecipitate containing hepatitis non-A, non-B viruses in addition to hepatitis B viruses a factor VIII concentrate (160 U/10 ml) can be prepared by a modified beta-PL/UV procedure, which induces neither hepatitis B nor hepatitis non-A, non-B in experimental animals; the non-sterilized original cryoprecipitate proved to be infectious.

Clinical evaluation of the hepatitis safety of a beta-propiolactone/ultraviolet treated factor IX concentrate (PPSB).

The association of viral hepatitis, type B, with the use of prothrombin complex concentrates (PPSB) has been well documented. PPSB prepared from cold-sterilized plasma (beta-propiolactone treated and UV irradiated) has been shown not to induce hepatitis in chimpanzees. 500 U of cold-sterilized PPSB were infused into 5 healthy male volunteers. Previous to, and up to 6 months after, PPSB-application in addition to careful clinical investigations, the following hepatitis parameters were determined: SGOT, SGPT, y-GT, alkaline phosphatase, total serum bilirubin, HBsAg, HBcAb and HBsAb. On the basis of all of the above parameters there was no indication of induction of viral hepatitis following the application of PPSB prepared from beta-propiolactone/UV treated plasma.

Long-term tolerance and recovery of beta-propiolactone/ultraviolet (beta PL/UV) treated PPSB in chimpanzees.

Recent experiments have shown that a preparation of PPSB (factor IX concentrate) derived form beta PL/UV treated plasma was not infectious in chimpanzees with respect to hepatitis B and non-A, non-B. To answer the question whether the beta PL/UV treatment influences the tolerance and efficacy of the PPSB-concentrate, long-term application of PPSB-Biotest was carried out in chimpanzees. After 10 applications of 25 U factor IX/kg at weekly intervals, no signs of intolerance were observed by measurement of blood pressure during i.v. application and by means of skin-testing. Determination of coagulation factor activity during the application period revealed the same factor IX recovery at the beginning and at the end of the study.