Pharmacokinetics of two pasteurized factor VIII concentrates by different and multicenter assays of factor VIII activity.

We assessed the pharmacokinetic characteristics of a new high-purity pasteurized FVIII concentrate in comparison with an intermediate purity pasteurized concentrate, produced by the same manufacturer. The study was designed as a cross-over single-dose pharmacokinetic investigation in 8 non-bleeding patients with severe hemophilia A. All patients were given 25 IU/kg of each of the two concentrates, with an interval of at least one week between the two administrations. Decay curves were assessed by collecting 10 serial blood samples over 36 hours following the end of infusion. The concentration of Factor VIII in blood samples was determined in triplicate in three different laboratories using each of the following assay methods: a one-stage clotting assay, a two-stage clotting assay, and a two-stage chromogenic-peptide substrate assay. All pharmacokinetic parameters were calculated by model-independent methods. The two products were found to differ significantly both in the clearance, which was on average 13.8% lower for Haemate P, and in the in-vivo recovery, which was 11.7% lower for Factor VIII:C P on the average. In comparison with previous pharmacokinetic data obtained from other heated Factor VIII concentrates, the clearance of Haemate P was found to be significantly slower, while the half-life of both products was longer. No differences were observed in the Vd-area. These findings indicate that the purification procedures to which both products are subjected do not increase the in-vivo rate of plasma disappearance of Factor VIII.

Reproducibility of one-stage, two-stage and chromogenic assays of factor VIII activity: a multi-center study.

This study was aimed at assessing the reproducibility of Factor VIII assays between different laboratories using the same reagents. A total of 176 post-dose plasma samples were obtained from 8 Italian subjects with hemophilia-A treated with a single dose of Factor VIII concentrates. Three laboratories (in FRG, Italy, and Sweden) participated in the study. Frozen aliquots of each sample were dispatched to each of the laboratories, where the aliquots were assayed using the same one-stage, two-stage and chromogenic methods. The one-stage and the chromogenic methods were well reproducible between the three centers: pairwise correlation analyses yielded r-values ranging from 0.88 to 0.91 for the one-stage method and from 0.91 to 0.96 for the chromogenic method. The agreement between these two assays was less evident in samples with activity below 200 IU/L in which the one-stage gave, on average, higher Factor VIII concentrations than those provided by the chromogenic method. The two-stage method was not well reproducible, and the pairwise r-values ranged from 0.48 to 0.73. Our study emphasises the need to develop multi-center quality control programs to verify the reproducibility of Factor VIII assays.

Some characteristics of aggregates of IgG and plasma proteins in heat-treated factor VIII concentrates.

Eight batches of commercial heat-treated and one untreated factor VIII concentrate from 6 producers were analyzed for their content of IgG, IgG subclasses, IgG aggregates and the presence of other plasma proteins combined with the IgG as well as for anticomplement activity. Methods used were thin-layer gel filtration, immuno-gel filtration, spot immuno-precipitate assay in a double antibody version and an agarose plate haemolysis inhibition assay of complement fixation. The IgG content varied from 0.1-6.90 g/l. In all preparations IgG existed as monomers and aggregates. Associated with the IgG were also found, at significantly increased amounts compared to normal serum and intravenous immunoglobulin, one to four of the following plasma proteins; fibronectin, fibrinogen, von Willebrand factor antigen, Clq, albumin and IgA. Three batches from two producers had high anticomplementary activity, presumably caused by the IgG aggregates. Two of these deviated strikingly from normal human serum pools in percent distribution of IgG subclasses. It is hypothesized that these aggregates can induce side effects or cause immunological aberrations.

On the unreliability of one-stage factor VIII:C clotting assays after infusion of factor VIII concentrates.

A multicentre study was undertaken in order to determine the reliability of the methods of assay of F VIII:C and the position of the peak value obtained after infusion of F VIII:C concentrates. Blood samples were drawn before and 10, 30 and 60 min after injection of F VIII concentrates in six haemophiliacs in one of the centres. Coded, frozen plasma samples were dispatched to the laboratories of the other four centres. F VIII:C was determined by different one-stage methods using the same international standard but with different activators. The results of the different laboratories differed widely and no agreement was reached on the existence of a lag period. To reach a valid conclusion not only the same sample has to be analysed, as in this study, but also the same laboratory technique has to be used by all participating investigators. To reach agreement on in vivo recovery and on elimination curves for different F VIII concentrates multicentre studies must be based on reliable methods of assay.

Immunoglobulin levels in haemophiliacs at HIV seroconversion and during follow up.

Immunoglobulins and aminotransferases were followed in 66 haemophilia patients and 13 von Willebrand patients over a six-year period. The results were correlated to HIV serology and lymphocyte subsets. Elevated IgG levels were found in 29/53 patients with haemophilia A, 2/13 with haemophilia B and in 0/13 with von Willebrand's disease. Elevated IgA and IgM levels were seen in 20% and 27% of the patients respectively, with a distribution similar to the elevated IgG levels, except that elevated IgA and IgM levels were also seen in 4/13 patients with von Willebrand's disease. Patients with HIV antibodies had significantly higher immunoglobulin levels than seronegative patients, and this elevation occurred in connection with seroconversion in the majority of the former. The IgG levels could not be correlated to the T4 cell count, but there has been a trend to less clinical symptoms related to HIV infection among those with stable IgG levels during the past few years. No correlation was found between elevated IgG levels and the aminotransferase levels, nor was any correlation found with the amount of blood coagulation factor concentrate given to the patients. The elevation of immunoglobulins observed in our haemophiliacs is multifactorial, but HIV infection is maybe the most important mechanism. The longitudinal IgG pattern may contribute to the prediction of the clinical outcome of this infection.

Complexes of IgG and plasma proteins in factor VIII preparations--a possible cause of adverse reactions.

IgG in factor VIII preparations was studied as a possible cause of adverse reactions following infusion of hemophiliacs. Thin-layer immuno-gel filtration analysis of nine products from seven firms demonstrated aggregated and monomeric IgG at highly variable amounts and proportions. These factor VIII products contained from 20 to 700 mg IgG per 1000 IU of factor VIII; IgG-fibrinogen complexes were demonstrated by double-antibody testing. A relationship is suggested between aggregated IgG and/or IgG complexes in factor VIII concentrates and adverse clinical reactions. They may also partially explain abnormalities and aberrations of the immune system previously reported in hemophiliacs.

Antenatal prophylaxis of Rh immunization with 250 micrograms anti-D immunoglobulin.

Anti-D immunoglobulin (250 micrograms) was given i.m. to 830 Rh-negative primigravidae and multigravidae round the 32nd to 34th week of gestation. The multigravidae had previously been treated with anti-D immunoglobulin post partum or after abortion and had been followed up serologically after 8 months. Five hundred and twenty-nine of the women delivered Rh-positive infants and received another injection of anti-D immunoglobulin (250 micrograms) within 72 hours of delivery. At the serological follow-up 8 months after delivery 2 women (0.4%) had weak anti-D antibodies by the papain technique. No anti-D could be detected in these 2 women 14 and 20 months, respectively, after delivery. In a previously performed postnatal clinical study with 250 micrograms anti-D immunoglobulin the failure rate was 1.6% (10 out of 645 women). Thus, antenatal prophylaxis significantly (p less than 0.05) reduced the incidence of Rh immunization. The haemoglobin and bilirubin levels in cord blood and capillary blood did not differ in the Rh-positive and Rh-negative infants. Three primiparae (0.2%) had anti-D antibodies at the time of the antenatal injection before delivery. Thus, the antenatal regimen of gestation did not give full protection from Rh immunization. It is suggested that an antenatal injection at the 28th week of gestation would have been more effective, as Rh sensitization during pregnancy has been reported to be more frequent from the 29th week of pregnancy. Since the introduction of postnatal anti-D prophylaxis the Rh immunization occurring during pregnancy accounts for most of the observed failures (2, 4, 5, 14, 20). The efficacy and safety of antenatal injection of anti-D immunoglobulin has therefore been investigated (5, 6, 7). At the Växjö Hospital in Sweden, where the incidence of anti-D antibodies was 1.6% (15), 250 micrograms of anti-D immunoglobulin were given at about 32-34 weeks of gestation from March, 1973, to December, 1977. The results are presented here.

Recovery and turnover rate of hepatitis B immunoglobulin in volunteers.

Hepatitis B Immunoglobulin with an antibody content of 2 300 IU was injected i.m. into five healthy female volunteers without Hepatitis B markers in serum (HBsAg, anti-HBs, anti-HBc). The uptake of anti-HBs after i.m. injection was calculated by measuring the concentration of antibody in plasma. An international standard (WHO) containing 100 IU/ml was used as reference. The serum concentration of anti-HBs was analysed on day 1, 3, 5, 7, 9, 11, 13, 28 and 56 after the single i.m. injection. The peak serum concentrations after one single injection were observed between day 5 and 11. The mean recovery rate was calculated according to a compartment model where the plasma volume was estimated to be 45 ml/kg body weight. The mean recovery rate expressed as percentage of injected dose was 19.2% with a range of 14.2-28.2%. The mean biological half life of anti-HBs was 21.7 days. The recovery rate of Hepatitis B Immunoglobulin injected i.m. seems to be somewhat lower than expected but is within the same range as i.e. Ig anti-D (Eklund et al, BMJ 1982, 284: 854-855). The peak serum concentration of anti-HBs was reached comparatively late after the injection. This finding may in part explain the discouraging protection of Hepatitis B Immunoglobulin in the post-exposure situation. Intravenous administration of anti-HBs with an immediate peak concentration followed by vaccination will possibly be a better alternative for post-exposure prophylaxis against Hepatitis B.

Turnover rate of anti-D IgG injected during pregnancy.

Anti-D IgG was injected into 15 Rh-negative women in the 28th week of gestation and into three non-pregnant women. The uptake of anti-D after the intramuscular injections was calculated by measuring the concentration of antibody in the plasma with an autoanalyser. The biological half life and the catabolic rate of anti-D IgG were calculated according to a compartmental model. The recovery in vivo of anti-D was an average 24% in the non-pregnant women and 21% in the pregnant women. The half life of anti-D were 24 and 21 days, respectively. With a dose of 125 micrograms the plasma anti-D concentration was less than 1 ng/ml at about 10 weeks after the injection. With double the dose the concentration at delivery was at least 1 ng/ml. Although 250 micrograms of anti-D IgG seems to be effective when given in the 28th weeks of gestation, the great individual variations in uptake and recovery rates will lead to occasional cases of Rh-immunisation during pregnancy despite all routine regimens.

Acute hepatitis non-A, non-B following administration of factor VIII concentrates.

A retrospective survey on clinical hepatitis in patients with bleeding disorders was performed. Nine episodes of hepatitis non-A, non-B occurred in 8 out of 20 patients (40%) with mild hemophilia A or von Willebrand's disease, who had been treated with commercial factor VIII concentrates. Only two episodes of hepatitis B occurred during the study period. The non-A, non-B attack rate after the first treatment was 40% with factor VIII concentrate obtained from large plasma pools (= 2,000 donors) including professional plasma donors as compared to 8% after treatment with factor VIII concentrate obtained from smaller (100-250 donors) plasma pools from Scandinavian donors.

Special considerations with regard to the dosage of tranexamic acid in patients with chronic renal diseases.

Tranexamic acid is a potent antifibrinolytic drug frequently used in the treatment of haematuria and a number of other haemorrhagic conditions. Since it is eliminated mainly in the urine, the drug accumulates in patients with uraemia. The excretion of tranexamic acid in patients with renal failure has been investigated and dosage recommendations are given for tranexamic acid therapy in cases of renal failure.

Intravenous gamma-globulin infusions in patients with hypo-gamma-globulinemia: prevention of adverse reactions with corticosteroids.

A gamma-globulin preparation for intravenous use was given to 11 patients with antibody deficiency syndromes. Most of them had reacted earlier to intramuscular injections of normal gamma-globulin. The gamma-globulin employed produced adverse reactions, often quite severe in 8 of 9 intravenous infusions in three patients. After premedication with hydrocortisone, side effects appeared on 18 of 48 occasions in 10 patients, but only twice were they so severe that the infusions had to be interrupted. Thus, it seems that hydrocortisone diminishes the risk of side effects. The intravenously administered gamma-globulin seemed to be just as effective as the preparations for intramuscular use, and no severe infections appeared during the period of observation. There was no indication that the single hydrocortisone injections, usually 200 mg, increased the risk of contracting infections, but still such medication should be used with great caution in antibody-deficient patients.

Hepatitis B immune globulin in prevention of hepatitis B among hospital staff members.

In a double-blind trial of hospital staff members at risk of contracting hepatitis B, 110 subjects received hepatitis B immune globulin or immune serum globulin prophylactically. An additional 125 individuals working in the same locations were unwilling to participate in the trial and received no prophylaxis. During the study period of 28 months, 13 cases of clinical hepatitis B occurred in the untreated group, whereas only three cases occurred among the subjects who received prophylaxis.

Diffusion of tranexamic acid to the joint.

Tranexamic acid (Cyklokapron, Kabi, Stockholm) in a dose of 10 mg per kg body weight was given i.v. to 17 patients at various intervals before operation on the knee joint, in order to elucidate the diffusion of the drug to the joint fluid and the synovial membrane. The acid diffused rapidly to both the above tissues, and in the joint fluid it reached the same concentration as in the serum. The biologic half-time in the joint fluid was about 3 hours. In the treatment of joint bleedings in hemophiliacs and in association with intra-articular operations on such patients tranexamic acid is a suitable supplement to conventional substitution therapy.

Rh-prophylaxis with immunoglobulin anti-D administered during pregnancy and after delivery.

During 1968-1973 510 Rh-negative, non-Rh-immunized primi- and multigravidae giving birth to Rh-positive infants, regardless of the ABO constellation, received 250 mug immunoglobulin anti-D post partum. In serological follow-ups using the indirect Coombs' test and the papain method more than six months after childbirth, Rh-antibodies were detected in two cases with papain but not at all with the indirect Coombs' test. Seventy women gave birth to Rh-positive babies in subsequent pregnancies. Twenty-six of these non-Rh-immunized women were given an intramuscular injection of 250 mug anti-D five to ten weeks before delivery and were included in a series comprising a total of 131 Rh-negative women who had received immunoglobulin anti-D in the estimated 32nd-34th week of pregnancy. Bilirubin and haemoglobin were determined in the neonates on cord blood and serum. The infants whose mothers had received immunoglobulin anti-D during pregnancy showed no signs of haematological abnormalities related to the administration of immunoglobulin anti-D. The determination of cord blood haemoglobin or bilirubin and serum bilirubin revealed no significant difference between Rh-positive infants and Rh-negative ones. Antibodies by the papain method were detected 41 of the women at the time of delivery (22 Rh-positive babies and 19 Rh-negative ones). There was no correlation between the time at which immunoglobulin anti-D was administered and the detection of antibodies at the time of delivery, regardless of the Rh group of the infant. The indirect Coombs' test was positive only in ten of the 48 Rh-negative infants examined. 250 mug immunoglobulin anti-D provides effective post partum prophylaxis, and the same dose administered to Rh-negative pregnant women prior to delivery did not cause any detectable haemolytic damage to the fetus.

Incidence of viral hepatitis after administration of factor IX concentrates.

A high frequency of viral hepatitis has been reported after treatment with the human factor IX concentrate 'Konyne'. Clinical trials with 'Konyne' and a similar factor IX concentrate, called 'Preconativ', was started in Sweden 1969. During the first 2 years, 26 patients were treated with either one or both preparations. Nine patients developed viral hepatitis within 6 months after treatment. 'Preconativ' alone was introduced on the Swedish market in 1971. During the period 1971-1974, another 26 hemophiliacs were treated but only two cases of hepatitis have occurred. Selection of donors and screening for hepatitis B surface antigen in donor blood used for the manufacturing of 'Preconativ', might be contributing factors to this low hepatitis incidence.

Hepatitis B immune serum globulin and standard gamma globulin in prevention of hepatitis B infection among hospital staff: a preliminary report.

In May 1973 a controlled double-blind clinical trail with prophylactic injects of hepatitis B immune serum globulin (antibody titer by passive hemagglutination 1:355,000) and standard gamma globulin (1:100) was started in Sahlgren's Hospital, Göteborg, Sweden. The annual attack rate of clinical hepatitis B in the three departments studied had been 5 to 8 per cent during recent years. A total of 118 members of the hospital staff were prophylactically treated while 125 staff members were unwilling to participate and received no prophylactic treatment. During the first 20 months of study nine cases of clinical hepatitis B with jaundice occurred within the untreated group (7.2 per cent) while three cases (2.5 per cent) were observed in prophylactically treated individuals. After decoding it was found that 60 individuals had received specific hepatitis B immune serum globulin while 58 had received standard gamma globulin. Two of the three clinical cases of hepatitis B occurred within the standard gamma globulin group. Both groups included two individuals with transient antigenemia only and the standard gamma globulin group also included four individuals with antibody seroconversion.