Drug – bio-affecting and body treating compositions – Extract – body fluid – or cellular material of undetermined... – Blood
Reexamination Certificate
1995-03-31
2002-02-12
Sayala, Chhaya D. (Department: 1761)
Drug, bio-affecting and body treating compositions
Extract, body fluid, or cellular material of undetermined...
Blood
C435S236000, C435S238000, C435S002000, C514S802000, C530S304000
Reexamination Certificate
active
06346277
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The invention relates to a process for the preparation of a plasma product which is virtually free from active viruses or of a concentrate of blood coagulation factors II, VII, IX and X which is virtually free from such viruses, by heating in the presence of stabilizers, and to a product of these factors which is thereby prepared. Such products can be used for the treatment of blood coagulation disorders.
2. Description of the Prior Art
Coagulation of blood is a complex process which proceeds in stages and is triggered of by various physiological and pathological causes, its course depending on about 20 promoting or inhibiting factors. Disorders in blood coagulation, some of which manifest themselves as diseases, occur as a result of a reduction or increase in these blood coagulation factors.
Concentrates of factors II, VII, IX and X are suitable for the treatment of various congenital or acquired disorders in the synthesis of these factors. The treatment of patients with concentrates of these factors has hitherto been associated with the risk of transmission of viruses and, in particular, hepatitis.
Albumin is hepatitis-safe if it is heated to 60° C. in aqueous solution and in the presence of stabilizers (Gellis, S. S. et al., J. Clin. Invest. (1948) 27, 239). It is therefore to be assumed that a concentrate of factors II, VII, IX and X heated in the presence of suitable stabilizers is likewise hepatitis-safe.
German Offenlegungsschrift 2,916,711 describes a process for the heat-stabilization of other coagulation factors in aqueous solution by addition of an amino acid and a mono- or oligo-saccharide or sugar-alcohol.
However, inactivation of factors II, VII, IX and X during heating cannot be prevented in this way. The observation that factors II and VII can be protected from thermal inactivation in aqueous solution by chelating agents (German Patent 3,043,857 A1) and factors IX and X can be protected by calcium (German Patent 3,045,153 A1). was therefore an advance. However, neither process allows the preparation of a product of all four factors in one operating process, as is desirable, since all four factors are reduced in cases of vitamin K deficiency or under therapy with oral anti-coagulants and must therefore be replaced at the same time.
SUMMARY OF THE INVENTION
Accordingly, there was the object of discovering a process for stabilizing an aqueous solution containing factors II, VII, IX and X from thermal inactivation.
Surprisingly, it has now been found that an aqueous solution containing all four factors II, VII, IX and X can be protected From the adverse consequences of heat treatment by addition of calcium ions and a chelating agent. If appropriate, antithrombin III (AT) and heparin can be added in order to prevent activation of factors II and VII. The preconditions were thus provided for preparing these four factors, with a high purity, and yield arid containing no active viruses, in one operation.
The chelating agent is added in addition to the citrate already present.
It is surprising that a mixture of calcium ions and chelating agent effects the desired protection of all factors II, VII, IX and X from toss in activity by heating, although, according to German Patent 3,043,857, a chelating agent is advantageous for protecting factors II and VII, whitst calcium ions are used for the same purpose for factors IX and X (German Patent 3,045,153), and chelating agents also form complexes with calcium ions. In each case only the two corresponding factors should therefore be protected from degradation by heat, and not all four, depending on whether the chelating agent or the calcium ions are present in excess.
DESCRIPTION OF PREFERRED EMBODIMENTS
The invention thus relates to a process for the preparation of a virtually virus-free and hepatitis-safe product of blood coagulation factors II, VII, IX and X by heating an aqueous solutions if appropriate in the presence of an amino acid and/or a saccharide or sugar-alcohol, which comprises heating the solution in the presence of calcium ions and a chelating agent and, if appropriate, antithrombin III and/or heparin.
Such a solution can be a solution containing these factors or a concentrate containing these, as well as plasma or a plasma or placenta fraction.
The optimum concentration of calcium ions is in the range from 1 to 50 mmol/l, preferably 25 to 50 mmol/l.
Examples of suitable salts which supply calcium ions are the chloride, acetate or nitrate, and all water-soluble calcium salts of sugar-acids, such as gluconic acid or lactonic acid. The chloride and acetate are preferably used.
Examples of suitable chelating agents are: ethylenediamino-tetraacetic acid (EDTA), ethylene glycol bis-(2-aminoethyl ether)-tetraacetic acid (EGTA), diaminocyclohexane-tetraacetic acid (CDTA), diaminopropane-tetraacetic acid and nitrilotriacetic acid, and, in particular, soluble alkali metal salts thereof.
Aliphatic aza-tri- or -tetra-carboxylic acids with 6-20 carbon atoms and 1 or 2 nitrogen atoms and soluble alkali metal salts thereof and, in particular, the sodium salts of ethylenediaminotetraacetic acid (EDTA) or ethylene glycol bis-(2-amino-ethyl ether)-tetraacetic acid (EGTA) are preferably used. The optimum concentration of the chelating agents is 1 to 20 mmol/l, preferably 5 mmol/l.
Mixtures of 25 mmol/l of calcium with 5 mmol/l of EDTA have proved particularly suitable.
Antithrombin III is employed in a concentration of 0.05-2 units/ml, based on the activity of 1 ml of citrated mixed plasma (1 unit), and together with heparin in concentrations of 0.5-2 ) USP units/ml, preferably 0.2 unit of antithrombin III with 2 USP units of heparin.
In the presence of a mixture of calcium ions and a chelating agent and, if appropriate, the antithrombin III-heparin complex, the aqueous solution of the coagulation factors can be heated until, according to the current state of knowledge, transmission of viruses, and in particular hepatitis pathogens, can be virtually excluded. This particularly applies if the pasteurization is combined with adsorption and precipitation processes, in which the active compound remains in the supernatant liquor and the hepatitis viruses can be separated off together with the insoluble precipitate. A product which has been kept at about 60° C. in aqueous solution for at least 10 hours is at present regarded as virtually hepatitis-safe, especially if the starting material is human tissue fluid in which hepatitis viruses have not been detected by a third generation test.
A particularly preferred embodiment of the invention comprises adding to a solution containing all four factors, preferably plasma or a plasma or placenta fraction, 0.2-2 units/ml of antithrombin III, 2-20 USP units/ml of heparin, 25-50 mmol/l of calcium ions and 1-20 mmol/l of EDTA, 1-3 mol/l of at least one of the amino acids glycine, alpha- or beta-alanine, lysine, leucine, valine, asparagine, serine, hydroxyprotine, protine, glutamine or alpha-, beta- or gamma-aminobutyric acid, but preferably glycine, and 20 to 60 g/100 g of solution of a mono- or oligo-saccharide or sugar-alcohol, preferably 1 to 3 mol/l of glycine and 20 to 60 g/100 g of solution of sucrose, heating the mixture to a temperature of 30° C. to 100° C., preferably 60° C. to 100° C. and keeping it at this temperature for 1 minute to 48 hours, preferably 8-12 hours, the shortest time being matched with the highest temperature and vice versa.
A pH value of 6 to 8 is maintained. A virtually virus-free product of factors II, VII, IX and X is obtained in this manner.
Depending on the solubility of the calcium salt, the amino acid or the carbohydrate, the corresponding concentrations of 0.3 and 3.0 mol/l and 60 g/100 g can be extended to higher concentrations if the calcium salt, the aminoacid or the carbohydrate have a correspondingly higher solubility at the desired temperature. The temperature treatment can also be carried out in several successive steps.
A hepatitis-safe product is achieved with the preferably used combination of a
Heimburger Norbert
Kumpe Gerhardt
Preis Hans Martin
Wormsbächer Wilfried
Aventis Behring GmbH
Finnegan Henderson Farabow Garrett & Dunner L.L.P.
Sayala Chhaya D.
LandOfFree
Process for the pasteurization of plasma or concentrates of... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Process for the pasteurization of plasma or concentrates of..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Process for the pasteurization of plasma or concentrates of... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2967691