Proteins factor VIII

Produce recombinant proteins (Factor VIII, FIBsAg, insulin)... 

This ignorance led to alarm as the first cases appeared in haemophiliacs, who had received the blood protein factor VIII. This was not only a serious health concern but also a major commercial problem, since worldwide sales of blood products was estimated to be around two billion US dollars. For haemophiliacs, the situation was dire. Because of the large quantity of blood plasma that had to be processed in order to obtain factor VIII, and the large number of injections they needed each year, it was estimated that each patient could be exposed to the blood of up to three million donors. At this time, the manufacturers of factor VIII did not use heat-treatment procedures, as they did from 1987, but nonetheless, the causative agent appeared to be pretty rugged. The appearance of the disease in haemophiliacs rendered the name GRID inappropriate for the condition, and this was now replaced by the term Acquired Immune Deficiency Syndrome or AIDS. 

Vitamin Bn, see also under Animal protein factor, VIII, 42 IX, 1-26 activity, IX, 2... 

Increasingly, plasma proteins that were once measured exclusively by their catalytic activities and expressed in terms of enzyme units are now being measured by immunoassay techniques, and expressed in terms of concentration (e.g., micrograms per liter). When the results from these very different forms of analysis are compared, there is often little correlation. In many cases, this reflects the fact that proteins can exist in multiple structural forms, differing in conformational properties, in terms of proteolytic cleavage, or in terms of specific forms of post-translational modification (e.g., phosphorylation). While these forms may retain some conserved epitopes, and thus have common immunological properties, structural differences often have a profound effect on their activities. For example, it has been found that the blood protein factor VIII, which is one of the key proteins in the blood clotting cascade, loses its catalytic activity in hemophilia while... 

Factor VIII Coagulation proteins hemophilia A treatment 300 3 X 10- ... 

Immunoaffinity chromatography utilizes the high specificity of antigen—antibody interactions to achieve a separation. The procedure typically involves the binding, to a soHd phase, of a mouse monoclonal antibody which reacts either directly with the protein to be purified or with a closely associated protein which itself binds the product protein. The former approach has been appHed in the preparation of Factor VIII (43) and Factor IX (61) concentrates. The latter method has been used in the preparation of Factor VIII (42) by immobilization of a monoclonal antibody to von WiHebrand factor [109319-16-6] (62), a protein to which Factor VIII binds noncovalenfly. Further purification is necessary downstream of the immunoaffinity step to remove... 

Factor VIII, immunoglobulin, and albumin are all held as protein precipitates, the first as cryoprecipitate and the others as the Cohn fractions FI + II + III (or FII + III) and FIV + V (or FV), respectively (Table 7, Fig. 2). Similarly, Fractions FIVj + FIV can provide an intermediate product for the preparation of antithrombin III and a-1-proteinase inhibitor. This abiUty to reduce plasma to a number of compact, stable, intermediate products, together with the bacteriacidal properties of cold-ethanol, are the principal reasons these methods are stiU used industrially. 

Fig. 6. Share of U.S. market occupied by human albumin/plasma protein fraction, (—) Factor VIII concentrate, (-) intravenous immunoglobulin...

Antihemophilic factor [9001-28-9] (AHF) is a protein found in normal plasma that is necessary for clot formation. It is needed for transformation of prothrombin to thrombin. Administration of AHF by injection or infusion can temporarily correct the coagulation defect present in patients with hemophilia. Antihemophilic factor VIII (Alpha Therapeutic) has been approved by the FDA as replacement therapy in patients with hemophilia B to prevent bleeding episodes, and also during surgery to correct defective hemostasis (178). 

Hemophilia A and B are coagulation disorders that result from defects in the genes encoding for plasma coagulation proteins. Hemophilia A (classic hemophilia) is caused by the deficiency of factor VIII, and hemophilia B (Christmas disease) is caused by the deficiency of factor IX. The incidences of hemophilia A and B are estimated at 1 in 5000 and 1 in 30,000 male births, respectively. Both types of hemophilia are evenly distributed across all ethnic and racial groups.1... 

Because it is a small protein, the factor IX molecule passes into the intravascular and the extravascular spaces. Therefore, the volume of distribution of recombinant factor IX is twice that of factor VIII. Consequently, 1 unit of factor IX administered per kilogram of body weight yields a 1% rise in the plasma factor IX level (0.01 unit/mL, or 1 IU/dL). Thus 1750 units of factor IX provides an incremental increase of 50% of normal (0.5 unit/mL, or 50 IU/dL), that is, 1750 units/70 kg x 0.01 unit/mL (1 IU/dL).12 Additionally, the recovery rate with recombinant factor IX is 20% lower than that with the plasma-derived products. Therefore, initial and subsequent maintenance doses should be adjusted accordingly. 

High purity (50-1000 factor VIII units/mg protein)... 

Using the same logic, the PTO has granted patents, for example, for pure cultures of specific microorganisms and for medically important proteins (e.g. Factor VIII purified from blood (Chapter 12) and EPO purified from urine (Chapter 10)). 

Many small proteins, in particular those that function extracellularly (e.g. insulin, GH and various cytokines) are quite stable and may be fractionated on a variety of HPLC columns without significant denaturation or decrease in bioactivity. Preparative HPLC is used in industrial-scale purification of insulin and of IL2. In contrast, many larger proteins (e.g. blood factor VIII) are relatively labile, and loss of activity due to protein denaturation may be observed upon high-pressure fractionation. 

The initial steps of the intrinsic pathway are somewhat more complicated. This system requires the presence of clotting factors VIII, IX, XI and XII, all of which, except for factor VIII, are endo-acting proteases. As in the case of the extrinsic pathway, the intrinsic pathway is triggered upon exposure of the clotting factors to proteins present on the surface of body tissue exposed by vascular injury. These protein binding/activation sites probably include collagen. 

Hypercoagulable states include malignancy activated protein C resistance deficiency of protein C, protein S, or antithrombin factor VIII or XI excess antiphospholipid antibodies and other situations. Estrogens and selective estrogen receptor modulators have been linked to venous thrombosis, perhaps due in part to increased serum clotting factor concentrations. Although a thrombus can form in any part of the venous circulation, the majority of thrombi begin in the lower extremities. Once formed, a venous... 

Different mutations in the disease-causing locus may cause more or less severe expression. For example, mis-sense mutations in the factor VIII gene tend to produce less severe hemophQia than do nonsense mutations, which result in a truncated protein product and little, if any, expression of factor VIII. The presence of different mutations, or alleles, in the same locus is termed allelic heterc eneity. 

Along with the production of insulin, many other medical uses have been achieved for recombinant DNA. This includes the production of erythropoetin, a hormone used to stimulate production of red blood cells in anemic people tissue plasminogen activator, an enzyme that dissolves blood clots in heart attack victims and antihemophilic human factor VIII, used to prevent and control bleeding for hemophiliacs. These three important genetically engineered proteins were all cloned in hamster cell cultures. 

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