38 results about "Coagulation Factor IX" patented technology

The invention relates to a preparation method of a high-purity human coagulation factor IX, which comprises the following steps: melting refrigerated plasma, and carrying out low-temperature centrifugation; adsorbing with a DEAE (diethylaminoethanol) Sephadex A-50 gel to remove the coagulation factor IX in the cold-glue plasma; removing impure proteins in the solution by using polyethyleneglycol; carrying out S / D virus inactivation; carrying out anion exchange column chromatography to obtain a purified coagulation factor IX solution; passing through a heparin affinity column for further chromatography to obtain a high-purity coagulation factor IX solution; carrying out ultrafiltration, dialysis and concentration, and adding arginine hydrochloride and glycinate as protective agents; filtering through a 20nm filter element to remove viruses; carrying out freeze-drying; and carrying out dry heat virus inactivation. The protein protective agents are added during the gel adsorption, column chromatography and ultrafiltration dialysis, thereby lowering the activation probability of the FIX product thrombin and enhancing the qualification rate of the product. The technique has high product yield; the FIX specific activity can reach 150 IU / mg or so which is much higher than that of the traditional product; and by performing the three-step virus inactivation, the product is safe and reliable to use.

Protein replacement therapy for patients with hemophilia or other inherited protein deficiencies is often complicated by pathogenic antibody responses, including antibodies that neutralize the therapeutic protein or that predispose to potentially life-threatening anaphylactic reactions by formation of IgE. Using murine hemophilia B as a model, we have developed a prophylactic protocol against such responses that is non-invasive and does not include immune suppression or genetic manipulation of the patient's cells. Oral delivery of coagulation factor IX (F. IX) expressed in chloroplasts, bioencapsulated in plant cells, effectively blocked formation of inhibitory antibodies in protein replacement therapy. Inhibitor titers were mostly undetectable and up to 100-fold lower in treated mice when compared to controls. Moreover, this treatment eliminated fatal anaphylactic reactions that occurred after 4 to 6 exposures to intravenous F. IX protein. While only 20-25% of control animals survived after 6-8 F. IX doses, 90-95% of tolerized mice survived 12 injections without signs of allergy or anaphylaxis. This high-responder strain of hemophilia B mice represents the first hemophilic animal model to study anaphylactic reactions. The plant material was effective over a range of oral antigen doses (equivalent to 5-80 μg recombinant F.IX / kg), and controlled inhibitor formation and anaphylaxis long-term, up to 7 months. Oral antigen administration caused a deviant immune response that suppressed formation of IgE and inhibitory antibodies. This cost-effective and efficient approach to oral delivery of protein antigens to the gut should be applicable to several genetic diseases that are prone to pathogenic antibody responses during treatment.

Disclosed is a fusion protein comprising blood coagulation factor IX (FIX) and transferrin. The fusion protein exhibits improved specific FIX activity, as compared to native FIX, and can be useful in the treatment of FIX deficiency-associated diseases.

The present invention discloses a preparation method of a highly purified human blood coagulation factor IX. The human blood coagulation factor IX is prepared by performing anion exchange chromatography, cation exchange chromatography, heparin affinity chromatography to the material containing human blood coagulation factor IX (taken from human blood serum or recombined cell culture medium), which includes the period of passivating virus through S / D treatment (Solvent / Detergent treatment) or removing virus through nanofiltration. The highly purified safe coagulation factor IX preparation having a specific activity of above 150 IU / mg, with substantially undoped proteins can be prepared by the preparation method of the invention.

The invention provides a method for efficiently extracting and purifying a blood coagulation factor IX and a blood coagulation factor X. The method provided by the invention comprises the following steps: collecting blood, treating, and collecting blood plasma; and extracting an IX crude extract and an X crude extract from the blood plasma, and purifying. The method is characterized in that the purifying step is to use FIX / FX-bp-Sepharose 4B to purify the blood coagulation factor IX crude extract and the blood coagulation factor X crude extract respectively through the affinity chromatography; and the affinity ligand FIX / FX-bp is selected from ACF I, ACF II or AHP. The purified blood coagulation factor IX can be directly used as a medicament and the purified blood coagulation factor X canbe directly used as a reagent.

Monoclonal antibodies directed against coagulation factor IX and their use in inhibiting thrombosis are disclosed.

A method of reducing pathogen content in a biologically derived composition by irradiating it with at least one high-intensity short-duration pulse of broad-spectrum incoherent polychromatic light. The biomolecule of interest retains biological activity in the resulting treated composition. Biologically derived compositions, such as serum, plasma or other blood products, including insulin, transferrin, heparin, collagen, coagulation factor VIII and / or coagulation factor IX, or containing monoclonal antibodies, or genetically engineered cell lines, etc. The content of pathogens, such as viruses, bacteria, pyrogens, fungi and / or prions present in the resulting protein composition is reduced by irradiation with broad spectrum pulsed light. A single pulse of broad spectrum light significantly reduces the amount of HIV-1, SV40, canine parvovirus or bovine viral diarrhea virus in a biologically derived composition.

The invention discloses a method for simultaneously separating and purifying coagulation factors IX, X and VII from human plasma, which comprises the following steps: performing centrifugal impurity removal, gel adsorption and ultrafiltration concentration to prepare a prothrombin compound; separating the coagulation factors VII and a mixed solution containing IX and II through an anion exchange resin column; separating the coagulation factor II and IX from the mixed solution containing IX and II by affinity chromatography. According to the method, by combining anion exchange chromatography with heparin affinity chromatography, the separation and preparation of three coagulation factors II, VII and IX at the same time can be achieved; the method has the advantages of high raw material utilization, simple operation and short time consumption; meanwhile, by detecting the electric signals in the chromatography process, the corresponding coagulation factors are accurately collected, the purity of the coagulation factors is effectively improved, and the economic benefit is improved.

Monoclonal antibodies directed against coagulation factor IX and their use in inhibiting thrombosis are disclosed.

The invention discloses a pharmaceutical composition of hemocoagulase. The pharmaceutical composition is characterized in that the pharmaceutical composition is prepared from 0.00005 to 0.0005 percentof a hemocoagulase composition and the balance of a pharmaceutically acceptable carrier. The composition is applied to preparation of a medicine for treating Haemophilia B (coagulation factor IX deficiency). Active components in the hemocoagulase comprise hemocoagulase, i.e., a thrombin-like enzyme (batroxobin), and a phospholipid dependent coagulation factor X activating agent (FXA). A bleedingstopping process of the hemocoagulase does not depend on a coagulation factor IX and is mainly used for fibrin and an activated coagulation factor X, and has the effect of treating bleeding of the haemophilia B. After the hemocoagulase is used for treating, platelet aggregation can be remarkably enhanced, and the formation of blood clots or fibrin clots in a patient with the haemophilia B is recovered.

The present invention relates to Factor IX polypeptides conjugated to heparosan (HEP) polymers, methods for the manufacture thereof and uses of such conjugates. The resultant conjugates may be used - for example - in the treatment or prevention of bleeding disorders such as haemophilia B.

The present invention relates to expression constructs and recombinant adeno-associated virus (rAAV) vectors that express coagulation factor IX, viral particles and pharmaceutical compositions comprising said rAAV vectors, and their use for the treatment or prevention of hemophilia B.

The invention relates to the field of medical biology, discloses a single-domain antibody specific to a coagulation factor IX (FIX), particularly discloses a FIX binding molecule derived from the single-domain antibody, and especially discloses application in FIX detection and FIX-poor plasma preparation.

The invention relates to a new, more potent, coagulation factor IX (FIX) expression cassette for gene therapy of haemophilia B (HB). Disclosed is a vector for expressing factor IX protein, the vector comprising a promoter, a nucleotide sequence encoding for a functional factor IX protein and an intron sequence, wherein the intron sequence is positioned between exon 1 and exon 2 of the nucleotide sequence encoding for a functional factor IX protein, and wherein the intron sequence has at least 80% identity to the sequence of SEQ ID NO. 1 as disclosed herein.

An objective of the present invention is to provide an effective pharmaceutical composition or a dosage regimen for preventing and / or treating bleeding, a disease accompanying bleeding, or a disease caused by bleeding. The inventors discovered that by administering a pharmaceutical composition comprising a bispecific antigen-binding molecule that recognizes (a) blood coagulation factor IX and / or activated blood coagulation factor IX and (b) blood coagulation factor X and / or activated blood coagulation factor X according to a given dosage regimen, bleeding, a disease accompanying bleeding, or a disease caused by bleeding can be prevented and / or treated more effectively.

An objective of the present invention is to provide an effective pharmaceutical composition or a dosage regimen for preventing and / or treating bleeding, a disease accompanying bleeding, or a disease caused by bleeding. The inventors discovered that by administering a pharmaceutical composition comprising a bispecific antigen-binding molecule that recognizes (a) blood coagulation factor IX and / or activated blood coagulation factor IX and (b) blood coagulation factor X and / or activated blood coagulation factor X according to a given dosage regimen, bleeding, a disease accompanying bleeding, or a disease caused by bleeding can be prevented and / or treated more effectively.

The invention relates to a preparation method of a clinical blood coagulation inspection preparation and particularly relates to a preparation method of a blood coagulation factor IX quality control product. The preparation method comprises the following steps: carrying out affinity chromatography on the mixed blood plasma of multiple persons by using an anti-human blood coagulation factor IX monoclonal antibody immunoaffinity chromatography column, removing a blood coagulation factor IX in the mixed blood plasma of multiple persons to obtain a blood plasma in shortage of the blood coagulation factor IX; mixing the mixed blood plasma of multiple persons with the blood plasma in shortage of the blood coagulation factor IX according to a certain proportion to prepare a blood coagulation factor IX quality control product with the content of the blood coagulation factor IX at different concentration levels, adding a freeze-drying protective additive, carrying out sub-packaging, and carrying out freeze drying so as to obtain the blood coagulation factor IX quality control product. According to the blood coagulation factor IX quality control product prepared by the preparation method, the uniformity, the stability and the stability of freeze-dried aquatic product subjected to re-melting are good, and the quality control product can replace an imported product to be used for quality control on detection of blood coagulation factor IX, so that the reduction of the detection cost is facilitated and the capability of detecting the blood coagulation factor IX in China can be promoted.

The invention discloses a method for simultaneously separating and purifying coagulation factors IX, X and VII from human plasma, which comprises the following steps: performing centrifugal impurity removal, gel adsorption and ultrafiltration concentration to prepare a prothrombin compound; separating the coagulation factors VII and a mixed solution containing IX and II through an anion exchange resin column; separating the coagulation factor II and IX from the mixed solution containing IX and II by affinity chromatography. According to the method, by combining anion exchange chromatography with heparin affinity chromatography, the separation and preparation of three coagulation factors II, VII and IX at the same time can be achieved; the method has the advantages of high raw material utilization, simple operation and short time consumption; meanwhile, by detecting the electric signals in the chromatography process, the corresponding coagulation factors are accurately collected, the purity of the coagulation factors is effectively improved, and the economic benefit is improved.

The invention allows substantial improvements in stability of coagulation Factor IX in aqueous compositions. An aqueous composition sealed in a non-glass container comprising Factor IX in a buffer and calcium ions is provided, together with methods of stabilizing an aqueous Factor IX composition comprising storing said composition in a non-glass container for at least 7 days.

The invention provides application of a urine coagulation factor IX and a polypeptide fragment of the urine coagulation factor IX, and particularly relates to application of the urine coagulation factor IX and the polypeptide fragment of the urine coagulation factor IX in preparation of preparations for burn diagnosis, differential diagnosis, burn area and degree evaluation, treatment effect evaluation, monitoring, prognosis evaluation, mechanism research and the like. Burns are common important wounds in daily life, about 5000-100000 people in every one million people are burnt every year, according to statistics of World Health Organization, more than 300,000 people die from burn patients globally every year, and the treatment survival rate of serious burns is still at a lower level. Researches prove that compared with healthy people (a normal control group), the expression of the urine blood coagulation factor IX and the polypeptide fragment thereof in burn patients is increased, and the content of the urine blood coagulation factor IX and the polypeptide fragment thereof is gradually increased along with the aggravation of the burn degree. The method can be used for various purposes of application detection of burn patients. The advantages of noninvasive acquisition, large-scale repeated sampling and convenient preservation of a urine sample are exerted, and the urine blood coagulation factor IX and the polypeptide fragment thereof are detected by utilizing the urine sample.

The invention discloses a high-activity recombinant human coagulation factor IX fusion protein and a preparation method thereof. The high-activity recombinant human coagulation factor IX fusion protein comprises a promoter, a human coagulation factor IX protein gene, a human immunoglobulin IgG1 Fc fragment gene and a terminator. A carrier is used to transfer a human FIX-Fc fusion protein gene expression kit to mammalian genome to obtain a transgenic animal; human FIX-Fc fusion expression protein is attained from milk of the transgenic animal. According to the method herein, the human FIX expressed in the milk of the animal may reach 6 IU / ml in activity and 3000 IU / mg in average specific activity which is about 12 times as high as that of natural FIX, and is applicable to the production ofhuman coagulation factor IX.

The invention relates to the field of medicine and biology, and discloses a single-domain antibody against blood coagulation factor IX (FIX). Specifically, the present invention discloses a FIX-binding molecule derived from the single-domain antibody and its use, especially for detecting FIX and preparing FIX-deficient plasma.

The present invention provides a method for improving or controlling the plasma half-life and / or bio-availability of blood coagulation factor IX (FIX), the method comprising modifying the GLA domain. Examples of such modifications include: (i) non-covalent bonding of a GLA-domain-recognizing antibody or an antibody fragment thereof to the GLA domain; (ii) reduced number of Gla residues in the GLA domain, in comparison to that of a native FIX; (iii) either or both of deletion of one or more glutamic acid residues in the GLA domain and substitution of one or more glutamic acid residues in the GLA domain with another amino acid; and (iv) deletion of a part or all of the GLA domain. The present invention also provides a FIX with improved pharmacokinetics which carries such modifications, a pharmaceutical composition containing the FIX as an active ingredient, a method for producing the FIX, and such.