WO1999064440A1 - Procede de production d'un interferon alpha par immunoadsorption et filtration d'excretion de virus - Google Patents

Procede de production d'un interferon alpha par immunoadsorption et filtration d'excretion de virus Download PDF

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Publication number
WO1999064440A1
WO1999064440A1 PCT/FI1999/000506 FI9900506W WO9964440A1 WO 1999064440 A1 WO1999064440 A1 WO 1999064440A1 FI 9900506 W FI9900506 W FI 9900506W WO 9964440 A1 WO9964440 A1 WO 9964440A1
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interferon
ifn
virus
solvent
subtypes
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PCT/FI1999/000506
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English (en)
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Hannele TÖLÖ
Jaakko Parkkinen
Hanna-Leena Kauppinen
Gunnar Alm
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Suomen Punainen Risti Veripalvelu
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Priority to AU47835/99A priority Critical patent/AU4783599A/en
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/52—Cytokines; Lymphokines; Interferons
    • C07K14/555—Interferons [IFN]
    • C07K14/56—IFN-alpha
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
    • A01K2217/00—Genetically modified animals
    • A01K2217/05—Animals comprising random inserted nucleic acids (transgenic)

Definitions

  • the present invention relates to the manufacture of ⁇ -interferon drugs from human or animal cell culture or from transgenic animals.
  • the invention relates to the manufacture of a highly purified and virus-safe ⁇ -interferon solution, which contains major natural ⁇ -interferon subtypes.
  • ⁇ -Interferons are important drugs with antiproliferative, antiviral and immunomodulatory effects.
  • Human leukocytes are known to produce several IFN- ⁇ subtypes in culture when induced by Sendai virus (Nyman et al., Biochem. J. 329, 295- 302, 1998).
  • Human lymphoblastoid cells produce similarly several IFN- ⁇ subtypes after Sendai virus induction (Zoon et al, J. Biol. Chem. 267, 15210-15216, 1992).
  • Both leukocyte and lymphoblastoid IFN- ⁇ (hereinafter called "multicomponent ⁇ -interferon”) are used as drugs in the treatment of neoplastic and viral diseases.
  • IFN-a drugs are also produced by recombinant DNA technology in bacteria.
  • the recombinant bacterial products contain a single IFN-a subtype, which differs structurally from the corresponding natural subtypes.
  • Recombinant bacterial products may induce antibodies against IFN-a in patients, which may result in the disappearance of the therapeutic response.
  • Multicomponent IFN-a products have restored the therapeutic response in these patients, and multicomponent products may have also other therapeutic benefits in comparison with recombinant bacterial products (Farrell, Hepatology 26, 96S-100S, 1997).
  • the therapeutic benefits of multicomponent IFN-a are evidently based on the native structure of the IFN-a subtypes in these products. It is also possible to produce native IFN-a subtypes with recombinant DNA technology in cultured animal cells or transgenic animals, which, unlike the recombinant bacterial products, would contain the correct structure even with respect to posttranslational modifications.
  • IFN- ⁇ An important goal in the purification of IFN- ⁇ from human or animal cell culture is to include all major natural subtypes in the finished product. This must take place in a reproducible way in order to have a constant subtype composition in the product, which is a prerequisite for the constant quality of the drug. Further, it is equally important in the purification of IFN- ⁇ from human or animal cell culture or from transgenic animals that viruses present in the starting materials are inactivated and removed. Leukocytes, cell lines, serum and its fractions and animals used in the production of natural IFN- ⁇ subtypes may harbour viruses and other infectious agents of diverse physicochemical properties, which cannot be effectively inactivated by a single virus inactivation treatment.
  • Blood-borne viruses potentially present in serum fractions and leukocytes include enveloped viruses, such as Hi-viruses, hepatitis C and B viruses, and also non-enveloped viruses resistant to many physicochemical treatments, such as parvovirus B19.
  • Lymphoblastoid cell lines may harbour e.g. retroviruses.
  • US Patent No. 5,391,713 describes a purification process for leukocyte interferon comprising an immunoaffinity chromatography step, in which a polyclonal goat antibody against IFN- ⁇ is used.
  • the purification process comprises precipitation steps with thiocyanate solution and aqueous ethanol, but no other specific virus inactivation or removal steps.
  • the use of polyclonal antibodies in the purification may be problematic since the specificity of the antibodies obtained in different immunization times may change and result in changes in the subtype composition and impurity profile of the drug purified with different antibody batches.
  • US Patent No. 5,503,828 concerns a purification process of leukocyte interferon comprising immunoaffinity chromatography with a monoclonal antibody NK2.
  • the use of this monoclonal antibody results in a composition, which consists of at least 50 % of the IFN- ⁇ subtypes ⁇ 2 and ⁇ 8, and additional subtypes from the group oc4, ⁇ 7, ⁇ lO, ⁇ l6, ⁇ l7 and ⁇ 21.
  • at least subtypes IFN- ⁇ 1, IFN- ⁇ 21 and IFN- ⁇ 14 are lost. Of these subtypes, IFN- ⁇ 1 is the most abundant natural subtype (Nyman et al., Biochem. J. 329, 295-302, 1998).
  • the purification process contains low pH treatment for at least 2 days but no other, specific virus inactivation or removal steps.
  • the current methods for the purification and manufacture of leukocyte interferon do not comprise several specific virus inactivation or removal steps with different mode of action and they do not allow for inactivation of viruses resistant to e.g. low pH or aqueous ethanol and thiocyanate.
  • the invention aims at providing a novel method for producing a higly purified, virus-safe multicomponent IFN- ⁇ solution from crude interferon preparations.
  • a manufacturing process for multi- component IFN- ⁇ comprising multiple virus elimination steps.
  • the present invention is based on the finding that by subjecting an intermediate proteineous composition of an IFN- ⁇ isolation process to immunoadsorption chromatography with at least two monoclonal antibodies, which have complementary subtype specificities, it is possible not only to separate reproducibly all major IFN- ⁇ subtypes but also effectively to remove the most persistant infectious agents from IFN- ⁇ , including non-enveloped viruses and physico- chemically resistant infectious agents, such as those causing transmissible spongiform encephalopathies ("prions"). More specifically, the present invention is characterized by what is stated in the characterizing part of claim 1.
  • the invention provides considerable advantages.
  • by combining immuno- adsorption chromatography with a preceding solvent/detergent treatment, and with a subsequent treatment at low pH and filtration with a virus removal filter it is possible to achieve four efficacious virus elimination steps, of which at least two eliminate of all types of viruses and infectious agents potentially present in the cell cultures.
  • the present process has the capacity to purify extensively all major natural IFN- ⁇ subtypes with a good yield and continuously give rise to a product with a constant subtype composition.
  • Figure 1 shows in the form of a flow diagram of a preferred embodiment of the present process for producing highly purified, virus-safe IFN- ⁇ drug containing all major IFN- ⁇ subtypes
  • Figure 2 shows a typical reverse phase-HPLC profile of the purified leukocyte IFN- ⁇ drug substance.
  • the present invention describes a method for producing highly purified ⁇ -interferon product.
  • the process generally comprises the steps of removing the cells from a leukocyte culture, concentrating the crude interferon thus obtained, filtering and subjecting the crude interferon concentrate to solvent/detergent treatment (in the following also abbreviated S D treatment), binding IFN- ⁇ from the S/D treated crude interferon concentrate with at least one monoclonal antibody in immunoadsorption chromatography, eluting IFN- ⁇ from the column and keeping it at low pH at least for 16 h, neutralizing the eluate and separating residues of the mouse antibodies by gel filtration and finally subjecting the purified drug substance thus obtained to virus filtration and sterile filtration in a formulation suitable for the finished product.
  • solvent/detergent treatment in the following also abbreviated S D treatment
  • binding IFN- ⁇ from the S/D treated crude interferon concentrate with at least one monoclonal antibody in immunoadsorption chromatography
  • Viral safety of the product is confirmed by providing at least two effective virus elimination steps for all viruses representing diverse physicochemical properties.
  • S/D treatment inactivates effectively all enveloped viruses.
  • immunoadsorption chromatography with the excessive washing steps removes all types of viruses.
  • incubation at low pH effectively inactivates most viruses, except some acid-resistant viruses.
  • virus filtration removes all viruses, including even the smallest non- enveloped viruses.
  • other physicochemically resistant infectious agents such as "prions" are also effectively removed in the immunoadsorption chromatography and virus filtration steps.
  • the process comprises at least two effective steps against all types of physicochemically resistant infectious agents, and even four steps for the most important blood-derived viruses, Hi-viruses and hepatitis B and C viruses, which carry a lipid envelope and are effectively inactivated in S/D and low pH treatments and removed in immunoadsorption chromatography and virus filtration.
  • Figure 1 An exemplifying embodiment of the manufacturing process is depicted in Figure 1, which illustrates the process starting from a leukocyte culture.
  • the first step of the preferred embodiment comprises producing a crude interferon in leukocyte culture, in particular by Sendai virus induction (Cantell et al., Methods Enzymol. 78, 29-38, 1981).
  • the starting material can equally well be crude interferon obtained in human lymphoblastoid cell culture by Sendai virus induction (Mizrahi et al., Methods Enzymol. 78, 54-66, 1981).
  • the cells are removed from the culture by decantation, microfiltration and/or by centrifugation.
  • the filtrate or supernatant is concentrated by ultrafiltration to provide a crude IFN concentrate, filtered and treated with S/D reagents in order to decompose the lipid enveloped viruses.
  • S/D treatments are disclosed in the art, in particular in US Patents Nos. 4,540,573, 4,764,369 and 4,820,805, which are herewith incorporated by reference.
  • the organic solvent is preferably selected from dialkylphosphates and trialkylphosphates having alkyl groups which contain 1 to 10 carbon atoms.
  • trialkylphosphates are tri-(n-butyl)phosphate, tri-(t-butyl)phosphate, tri-(n-hexyl)phosphate and tri-(2- ethylhexyl)phosphate.
  • the concentration of the solvent is in the range of 0.001 % to 10 %, preferably about 0.01 % to about 1 %, a typical concentration of tri-(n-butyl)-phosphate being about 0.3 % (calculated from the weight of the solvent).
  • the solvent can be used together with a non-toxic detergent, which is capable of enhancing the effectiveness of the solvent.
  • non-ionic surfactants e.g. oxyethylated alkyl phenols, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene alcohols and sodium deoxycholate.
  • the amount of the detergent is in the range of 0.001 to 10 %, preferably about 0.01 to 1.5 %.
  • the detergent comprises 1 % polysorbate 80 (Tween 80), 1
  • Triton X-100 % Triton X-100 or 0.2 % sodium deoxycholate.
  • S/D substances may be used even at higher concentrations, such as 2 % tri-(n-butyl)phosphate without a detergent.
  • the treatment time may be 4-50 h, and the temperature 24-30 °C.
  • Protein impurities, DNA, possible residual viruses and S/D chemicals are removed by binding IFN- ⁇ to an immunoadsorbent consisting of monoclonal antibodies coupled to a solid matrix.
  • an immunoadsorbent consisting of monoclonal antibodies coupled to a solid matrix.
  • monoclonal antibodies with complementary subtype specificities should be used. It is possible to implement the invention using only one monoclonal antibody.
  • Monoclonal antibodies can be produced by using purified leukocyte interferon as an immunogen.
  • the antibody producing cell clones are selected by testing the ability of the antibodies to bind leukocyte interferon in immunoadsorption (Example 1).
  • the monoclonal antibodies should together bind at least 90 % of interferon activity present in crude interferon, and the yield of interferon activity eluted with low pH should be at least 80 % of the bound activity.
  • the antibodies should have complementary binding specificities to IFN- ⁇ subtypes so that they bind together all major IFN- ⁇ subtypes. Particularly, the antibodies should bind at least the IFN- ⁇ subtypes ⁇ l, ⁇ 2, ⁇ 8, ⁇ lO, ⁇ l4, ⁇ l7, and ⁇ 21. It is preferred to have at least one antibody which binds subtype ⁇ l and one which binds subtype ⁇ 2.
  • the immunoadsorbent column is washed extensively and the bound IFN- ⁇ is eluted with low pH buffer (pH 2 to 2.5).
  • the elution buffer may contain albumin or other stabilizers, such as a non-ionic detergent, to stabilize IFN- ⁇ .
  • the eluate is incubated at low pH at least for 16 h to inactivate possible residual viruses.
  • the eluate is neutralized by adding alkaline buffer or sodium hydroxide.
  • the neutralized eluate is concentrated by ultrafiltration.
  • IFN- ⁇ can be bound to an ion-exchange chromatography gel.
  • Mouse IgG and its fragments leaking from the immunoadsorbent and possible albumin stabilizer are removed by gel filtration or ion-exhange chromatography.
  • the IFN- ⁇ containing fractions are pooled and the purified IFN- ⁇ drug substance thus obtained is stored frozen at -70 °C.
  • the IFN- ⁇ drug substance is stabilized with a suitable stabilizer, such as albumin or a non- ionic detergent, and filtered with a virus removal filter with high retentive capacity even for the smallest non-enveloped viruses.
  • virus filters capable of reducing the concentration of model viruses, preferably model viruses having a size of 20 to 40 nm, in the spiking tests with at least 4 log, are considered to have a "high virus retentive capacity". It is particularly important that the filters used have such capacity also in relation to small non-enveloped viruses.
  • the theoretical pore sizes of the virus removal filters can be set at about 10 to 40 nm, preferably about 10 to 20 nm.
  • the yield of IFN- ⁇ in virus filtration and the capacity of the filter are clearly higher when a non-ionic detergent, such as polysorbate 80, is used as a stabilizer instead of albumin, as indicated in Example 4.
  • a non-ionic detergent such as polysorbate 80
  • Two virus removal filters can be used sequentially, which further improves virus retentive capacity.
  • Virus filtration may be also performed after sterile filtration of the formulated bulk solution. Additional stabilizers may be added to the bulk solution after virus filtration.
  • IFN- ⁇ drug substance it is also possible to formulate the IFN- ⁇ drug substance to a suitable pharmaceutical composition before the virus filtration step, as indicated in Figure 1.
  • the process according to the present invention effectively removes all types of impurities present in leukocyte and lymphoblastoid cell cultures, as indicated by more than million fold removal of DNA and about million fold removal of ovalbumin, which is the major heterologous protein derived from Sendai virus preparation used in the induction of IFN- ⁇ production in the cells (Example 2).
  • the level of mouse IgG leaking from the immunoadsorbent is typically below 10 ng/3 mill. IU of purified IFN- ⁇ .
  • the present process is economical containing only two chromatography steps, and it gives good yield of the purified drug substance, on an average about 60 % (calculated from the immunochemical concentration of IFN- ⁇ ) as shown in Example 2.
  • IFN- ⁇ concentration was measured by a time-resolved fluoroimmunoassay (FIA) on microtitre plates.
  • FSA fluoroimmunoassay
  • the IgG fraction of a bovine antiserum against human leukocyte IFN- ⁇ was used in capturing and a mixture of two Eu-labelled mouse IgG monoclonal antibodies to IFN- ⁇ for detection.
  • the monoclonal antibodies were the same as used in the purification of IFN- ⁇ . The details of the assay have been described elsewhere (R ⁇ nnblom et al., APMIS 105, 531-536, 1997).
  • IFN- ⁇ concentration was expressed as IU/ml using a laboratory standard, which was calibrated by the virus plaque reduction assay against the International Reference Preparation of Interferon, Human Leukocyte 69/19 (NIBCS, U.K.).
  • the antiviral activity of the IFN was determined by a virus plaque reduction assay in 35 mm petri dishes using Human Epithelial 2 (HEp2) cells challenged with Vesicular stomatitis virus (VSV).
  • the IFN- ⁇ samples, control and standard were diluted serially at 0.25 log intervals to concentration of 0.3 - 3 IU/ml in Eagle's Minimum Essential Medium (EMEM) supplemented with fetal calf serum (FCS) 7 % and aureomycin 0.004 %.
  • EMEM Eagle's Minimum Essential Medium
  • FCS fetal calf serum
  • aureomycin 0.004 % The samples were assayed as triplicates at four dilutions in at least two assay series.
  • Total protein concentration was measured according to Lowry using human albumin as a standard (Total Protein Standard, Finnish Red Cross Blood Transfusion Service, Helsinki,
  • Mouse IgG concentration was measured by enzyme immunoassay (EIA) on microtiter plates.
  • EIA enzyme immunoassay
  • Polyclonal goat anti-mouse IgG, (Fc) F(ab') 2 fragment was used in capturing and polyclonal peroxidase-conjugated goat anti -mouse IgG (H+L) (Jackson Immuno Research Laboratories, U.S.A.) for detection.
  • Mouse myeloma IgG (Cappel, Belgium) was used as a standard.
  • Ovalbumin concentration was measured by a time-resolved fluoroimmunoassay (FIA) in microtitre plates.
  • FIA fluoroimmunoassay
  • the samples were treated with sodium dodecyl sulphate and proteinase K at pH 11 and the concentration of DNA was measured by the Threshold Total DNA Kit (Molecular Devices, U.S.A. ).
  • IFN- ⁇ subtypes were separated by reverse phase high performance liquid chromatography (RP-HPLC).
  • IFN- ⁇ sample c. 1 mill. IU in 400 ⁇ l
  • RP-HPLC reverse phase high performance liquid chromatography
  • IFN- ⁇ sample c. 1 mill. IU in 400 ⁇ l
  • a Delta-PakTM HP1 C4 column Waters, U.S.A.
  • IFN- ⁇ samples containing human albumin were first run through a Superdex 75 HR 10/30 column (Pharmacia Biotech, Uppsala, Sweden) in 50 mmol/1 Tris buffer, pH 7.5, containing 0.15 mol/1 NaCl to remove albumin from IFN- ⁇ .
  • IFN- ⁇ used for immunization was partially purified leukocyte interferon (Finnferon-alpha, Finnish Red Cross Blood Transfusion Service), which had been further purified by immunoadsorption using bovine polyclonal anti-IFN- ⁇ antibodies coupled to CNBr-Sepharose (Pharmacia Biotech, Sweden). Bound IFN- ⁇ was eluted at low pH and stabilized with sodium dodecyl sulphate (SDS). The immunization procedure consisted of giving one intraperitoneal injection of 5 mill.
  • the fusion partner cell line was a non-secreting mouse myeloma cell line
  • Fusing agent was polyethylene glycol 1500 and the medium was DMEM supplemented with HT and 10 % FCS.
  • mice were subcloned and tested for the production of mouse monoclonal antibodies against human IFN- ⁇ in enzyme immuno assay using the purified human leukocyte IFN- ⁇ as an antigen.
  • strongly positive monoclonal antibodies were selected for further testing.
  • the monoclonal antibodies were incubated with human leukocyte interferon.
  • Rabbit anti- mouse IgG antibodies coupled to Sepharose 4B gel (Pharmacia Biotech) were added and after incubation, the gel with bound mouse monoclonal antibodies and IFN- ⁇ was separated by centrifugation. The supernatant was determined for interferon activity.
  • Antibodies which bound at least 70 % of interferon activity were selected for further testing in combination of two different antibodies, and antibodies which as a mixture bound at least 90 % of the activity were considered as suitable candidates for the immunoadsorption chromatography step. Further testing was carried out by coupling the monoclonal antibodies to CNBr-Sepharose 6B (Pharmacia Biotech) and testing their binding properties in immunoadsorption. The selection criterium was that two monoclonal antibodies should together bind at least 90 % of interferon activity of crude leukocyte interferon and that the yield of the activity eluted with low pH should be at least 80 % of the bound activity.
  • the two mouse IgG monoclonal antibodies selected for the purification process bind together all major leukocyte IFN- ⁇ subtypes (IFN- ⁇ 1, IFN- ⁇ 2, IFN- ⁇ 8, IFN- ⁇ lO, IFN- ⁇ l4, IFN- ⁇ l7, and IFN- ⁇ 21). Neither of them binds beta- or omega-interferon.
  • IFN- ⁇ 1, IFN- ⁇ 2, IFN- ⁇ 8, IFN- ⁇ lO, IFN- ⁇ l4, IFN- ⁇ l7, and IFN- ⁇ 21 Neither of them binds beta- or omega-interferon.
  • a typical RP-HPLC chromatogram of the product purified by using these two monoclonal antibodies in immunoadsorption is shown in Figure 2.
  • This example was performed by following the steps from crude interferon to purified drug substance in the scheme of Figure 1.
  • the cells were removed from the leukocyte culture first by decanting and by microfiltration using 0.3 ⁇ m membranes.
  • the filtrate was concentrated 20-fold by ultrafiltration using 10 kD membranes. If not used immediately the concentrate was stored frozen at -40 °C. After thawing, the concentrate was filtered through 1.2 ⁇ m and 0.22 ⁇ m filters, and the clarified concentrate was treated with 0.3 % tri(n-butyl) ⁇ hosphate and 1 % polysorbate 80 for 16 h at 26 °C.
  • the concentrate was applied to an immunoadsorbent column containing the two monoclonal antibodies coupled (4 mg/ml of each) to CNBr-Sepharose 4FF gel (Pharmacia Biotech).
  • the load was 30-40 mill.
  • the immunoadsorbent column was washed with 40 column volumes of 0.011 mol/1 sodium phosphate buffer, pH 7, 0.14 mol/1 NaCl (PBS) containing 0.1 % polysorbate 80 followed by 10 column volumes of PBS without polysorbate 80.
  • the bound IFN- ⁇ was eluted with pH 2-buffer.
  • Human albumin 0:2 g/1 (Finnish Red Cross Blood Transfusion Service) was added to pH 2-buffer to stabilize IFN- ⁇ . After incubation for 18 - 20 h at pH 2, the eluate was neutralized by adding slowly 0.5 mol/1 NaOH and concentrated about 30-fold by ultrafiltration using 10 kD membranes. The concentrated eluate was applied to a Superdex 75 column (Pharmacia Biotech) equlibrated with PBS. The IFN- ⁇ containing fractions were pooled and stored frozen at -70 °C. The yield of IFN- ⁇ in 11 purification batches is shown in Table 1.
  • the purified drug substance may contain albumin up to 20 % of total protein.
  • Sensitive immunochemical assays have been used for the measurement of main heterologous protein impurities which are ovalbumin and mouse IgG. The amount of ovalbumin per a standard dose of 3 mill.
  • IU IFN- ⁇ has been 0.1-0.3 ng and that of mouse IgG 1-9 ng in the drug substance batches.
  • the purification of IFN- ⁇ has been c. million fold in relation to ovalbumin (Table 2).
  • the amount of the main process-derived impurity, tri(n-butyl)phosphate has been less than 0.2 ⁇ g per 3 mill.
  • IU of IFN- ⁇ in three drug substance batches analyzed.
  • the amount of DNA was measured in two drug substance batches and found to be less than 60 pg per 3 mill.
  • the purification of IFN- ⁇ was more than million fold in relation to DNA (Table 2). Table 2. Purification of IFN- ⁇ in relation to ovalbumin and DNA in the purification process
  • Figure 2 shows a typical RP-HPLC chromatogram of the purified drug substance obtained by using the process described in Example 2.
  • the RP-HPLC chromatogram was devided into seven peak groups as indicated by vertical lines in Figure 2.
  • the comparison of 11 drug substance batches indicates that the relative peak areas and thus the subtype composition remained very consistent in different batches (Table 3).
  • This example was performed by formulating the purified drug substance and subjecting it to virus filtration and sterile filtration according to the scheme in Figure 1.
  • the purified IFN- ⁇ drug substance was diluted to concentration of c. 4 mill. IU/ml with PBS containing 1 mg/ml albumin.
  • the formulated solution was prefiltered through 0.1 ⁇ m particle filter and thereafter with Planova 15N filter (Asahi Chemical Industry Co, Japan).
  • Planova filtration was carried out at room temperature using constant pressure (0.9 bar).
  • the IFN- ⁇ yield in the Planova filtration was c. 90 %.
  • the virus filtered IFN- ⁇ solution was filtered with a 0.1 or 0.22 ⁇ m sterile filter and filled aseptically in the final containers.

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Abstract

La présente invention concerne un procédé de production d'un interféron alpha à partir de préparations d'interféron brutes provenant d'une culture cellulaire humaine ou animale ou d'animaux transgéniques. Le procédé consiste à placer une composition traitée au solvant/détergent au contact d'au moins deux anticorps IgG monoclonaux d'une souris présentant des spécificités de sous-types complémentaires, dans une étape d'immunoadsorption. Les sous-types de l'interféron alpha liés par les anticorps IgG monoclonaux sont élués et l'éluat obtenu est purifié et filtré avec un filtre d'excrétion de virus. Le procédé de l'invention permet d'éliminer effectivement tous les types d'impuretés présents dans les produits d'interféron bruts et donne un interféron à constituants multiples très pur et exempt de virus contenant des sous-types principaux de l'interféron alpha.
PCT/FI1999/000506 1998-06-10 1999-06-09 Procede de production d'un interferon alpha par immunoadsorption et filtration d'excretion de virus WO1999064440A1 (fr)

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AU47835/99A AU4783599A (en) 1998-06-10 1999-06-09 Alpha-interferon manufacturing process using immunoadsorption and virus removal filtration

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FI981338 1998-06-10
FI981338A FI105319B (fi) 1998-06-10 1998-06-10 Menetelmä multikomponentti-alfa-interferonin valmistamiseksi

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2006085092A2 (fr) * 2005-02-12 2006-08-17 Viranative Ab Procede et utilisations d'anticorps dans la purification d'interferon
GB2460283A (en) * 2008-03-13 2009-11-25 Academia Sinica Method of changing a globular protein structure into a fibrillar protein structure.
US8268974B2 (en) 2003-05-29 2012-09-18 Academia Sinica Process to produce fibrillar proteins
US8357652B2 (en) 2009-11-20 2013-01-22 Academia Sinica Anti-tumor fibrillar human serum albumin methods and compositions

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1981002899A1 (fr) * 1980-04-11 1981-10-15 D Secher Anticorps monoclonal
EP0152345A2 (fr) * 1984-02-07 1985-08-21 INTERFERON SCIENCES, INC a Delaware Corporation Excipients pour l'administration d'interféron
WO1990015817A1 (fr) * 1989-06-20 1990-12-27 Bionative Ab Procede de purification d'interferon

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1981002899A1 (fr) * 1980-04-11 1981-10-15 D Secher Anticorps monoclonal
EP0152345A2 (fr) * 1984-02-07 1985-08-21 INTERFERON SCIENCES, INC a Delaware Corporation Excipients pour l'administration d'interféron
WO1990015817A1 (fr) * 1989-06-20 1990-12-27 Bionative Ab Procede de purification d'interferon

Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US8268974B2 (en) 2003-05-29 2012-09-18 Academia Sinica Process to produce fibrillar proteins
WO2006085092A2 (fr) * 2005-02-12 2006-08-17 Viranative Ab Procede et utilisations d'anticorps dans la purification d'interferon
WO2006085092A3 (fr) * 2005-02-12 2006-10-12 Viragen Inc Procede et utilisations d'anticorps dans la purification d'interferon
US7745147B2 (en) 2005-02-12 2010-06-29 Viranative Ab Methods and uses of antibodies in the purification of interferon
GB2460283A (en) * 2008-03-13 2009-11-25 Academia Sinica Method of changing a globular protein structure into a fibrillar protein structure.
GB2460283B (en) * 2008-03-13 2010-04-21 Academia Sinica Method of producing fibrillar proteins
US8357652B2 (en) 2009-11-20 2013-01-22 Academia Sinica Anti-tumor fibrillar human serum albumin methods and compositions
US9226951B2 (en) 2009-11-20 2016-01-05 Academia Sinica Anti-tumor fibrillar human serum albumin methods and compositions

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FI105319B (fi) 2000-07-31
FI981338A (fi) 1999-12-11
FI981338A0 (fi) 1998-06-10

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