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• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61M—DEVICES FOR INTRODUCING MEDIA INTO, OR ONTO, THE BODY; DEVICES FOR TRANSDUCING BODY MEDIA OR FOR TAKING MEDIA FROM THE BODY; DEVICES FOR PRODUCING OR ENDING SLEEP OR STUPOR
  • A61M1/00—Suction or pumping devices for medical purposes; Devices for carrying-off, for treatment of, or for carrying-over, body-liquids; Drainage systems
  • A61M1/36—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits
  • A61M1/3679—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits by absorption
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
  • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
  • A61K35/14—Blood; Artificial blood
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
  • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
  • A61K35/14—Blood; Artificial blood
  • A61K35/16—Blood plasma; Blood serum
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
  • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
  • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
  • B01J20/26—Synthetic macromolecular compounds
  • B01J20/262—Synthetic macromolecular compounds obtained otherwise than by reactions only involving carbon to carbon unsaturated bonds, e.g. obtained by polycondensation
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
  • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
  • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
  • B01J20/26—Synthetic macromolecular compounds
  • B01J20/265—Synthetic macromolecular compounds modified or post-treated polymers
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
  • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
  • B01J20/28—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties
  • B01J20/28014—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties characterised by their form
  • B01J20/28047—Gels
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
  • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
  • B01J20/30—Processes for preparing, regenerating, or reactivating
  • B01J20/3071—Washing or leaching
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
  • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
  • B01J20/30—Processes for preparing, regenerating, or reactivating
  • B01J20/3085—Chemical treatments not covered by groups B01J20/3007 - B01J20/3078
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
  • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
  • B01J20/30—Processes for preparing, regenerating, or reactivating
  • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
  • B01J20/3202—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the carrier, support or substrate used for impregnation or coating
  • B01J20/3206—Organic carriers, supports or substrates
  • B01J20/3208—Polymeric carriers, supports or substrates
  • B01J20/3212—Polymeric carriers, supports or substrates consisting of a polymer obtained by reactions otherwise than involving only carbon to carbon unsaturated bonds
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
  • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
  • B01J20/30—Processes for preparing, regenerating, or reactivating
  • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
  • B01J20/3231—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the coating or impregnating layer
  • B01J20/3242—Layers with a functional group, e.g. an affinity material, a ligand, a reactant or a complexing group
  • B01J20/3268—Macromolecular compounds
  • B01J20/3272—Polymers obtained by reactions otherwise than involving only carbon to carbon unsaturated bonds
  • B01J20/3274—Proteins, nucleic acids, polysaccharides, antibodies or antigens

Definitions

• the present invention concerns an immunosorbent useful for reducing or removing rheumatoid factors from whole blood or blood plasma.
• Rheumatoid factors are autoantibodies having speci- ficity to immunoglobulin G (IgG). These antibodies may be of both IgM and IgG class and are present i.a. in the blood of patients with rheumatoid arthritis and other autoimmune connective tissue diseases. Rheumatoid factors are believed to have importance for the formation of immune complexes causing e.g. arthritis and vasculitis (H.E. Jasin et al., Clinical Immunobiology b ⁇ _, p. 365, Ed. F.H. Bach, 1975). Jasin et al. describes a plurality of analytical methods for RF, in particular IgM RF, based on the binding of RF to IgG, in particular for use in differential diagnosis of rheumatic diseases.
• IgG was used for the mentioned immunoabsorptions of isolated RF because the binding constant of RF for heat- aggregated IgG is relatively high (10 1/mole), whill ⁇ e the binding constant for native IgG is very low (102 - - 11..11 xx 1100 4 1/mole) (D. Dissanayake: Immunology 32, 309, 1977).
• the art comprises pro- Waits for unspecific reduction of the RF concentration in blood or plasma based on plasmapheresis and for un ⁇ specific IgG removal by immunoabsorption on protein A Sepharose.
• IgG anti Factor IX antibodies of high avidity, such as IgG anti Factor IX, by specific immunoabsorption.
• the possibility of removing RF by specific absorption on aggregated IgG is mentioned in the DE-A 27 25 608. However, as mentioned, it is not demonstrated in the specification that such removal is feasible in practice.
• the object of the present invention is to provide an immunosorbent enabling such a specific removal or reduc ⁇ tion of RF.
• the invention is based on the surprising finding that coupling of substantially monomeric IgG (native IgG or chemically or enzymatically modified IgG) to a matrix of a porous material provides an immuno ⁇ sorbent which allows effective specific removal of both IgM RF and IgG RF from plasma and whole blood without simultaneous removal of normal IgG and IgM.
• CRF-IgG3 d RF i.e. about 50?o' of RF in plasma is bound to IgG.
• the immunosorbent of the invention is accordingly charac ⁇ terized by comprising a matrix of a porous material to which substantially monomeric IgG is coupled.
• the invention also concerns a process for reducing or removing RF from blood or blood plasma, said process being characterized by using as immunosorbent a matrix of a porous material to which substantially monomeric IgG is coupled.
• the IgG used for the coupling with a view to removal of RF is monomeric, preferably native human IgG. This is surprising because if the skilled person would think of examining the possibility of effectively removing RF by using IgG as antigen by immunoabsorption, he would expect on the basis of the above-mentioned literature that heat-aggregated IgG would be the preferred possibility because of the considerably higher binding constant. As appears from the following tests and discus ⁇ sion the monomeric IgG is far superior.
• intravenous injection of purified IgG can activate the complement system by binding comple- ment Clq.
• Chemical reduction of IgG e.g. by dithiothreitol or mercaptoethanol and subsequent alkylation e.g. by means of iodoacetamide or enzymatic modification e.g. by means of pepsin or plasmin to change the IgG conformation, thus reducing the Clq binding, is well-known, cf. the DE-C2 23 11 333,
• IgG from other animal species such as horse, sheep, rabbit and monkey, also react with RF (R.M. Pope: J. Lab. Clin. Med. 9J_, 842, 1981) and such IgG, too, can therefore be used in the immunosorbent.
• the matrix in the immunosorbent is a porous material, such as a gel, preferably an open gel with an exclusion molecular weight exceeding 10 kD, in particular exceeding 10 kD.
• the matrix must be compatible with an extra- corporal blood circulation.
• Preferred gels are agarose gels, in particular the so-called Sepharose gels, preferably Sepharose CL-2B or
• Sepharose CL-4B but also gels of acrylic polymers can be used.
• the present IgM RF is determined by the Rose-Waaler titre.
• a Rose-Waaler titre below 40 corresponds to normal plasma. It will thus be seen that all the gels result in a reduction with respect to the starting titre of 384, and that the best effect is obtained with "Sepharose CL-2B". It is remarkable that the absorption is specific, so that normal IgG and IgM are not also removed by the absorption. For purposes of comparison, the effect of the absorption with protein A Sepharose is shown. This gel reduces the IgG content in the plasma significantly, and the bound IgG is capable of binding some rheumatoid factor. The protein A Sepharose, however, does not absorb RF as effectively as IgG coupled directly to Sepharose CL-2B.
• IgM RF In the literature mentioned previously, the authors have concentrated on IgM RF. IgG RF can be shown in a predominant part of patients with juvenile rheumatoid arthritis (Florin-Christensen et al.: Ann. Theum. Dis. (1974) 33, 32), and the efficiency of the present process with respect to removal of IgG RF is therefore of great practical importance.
• Polymeric IgG was isolated from heat-aggregated IgG by gel filtration on Sepharose CL-4B.
• the surprisingly effective removal of RF with monomeric IgG coupled to Sepharose in spite of the low binding constant of rheumatoid factors to monomeric IgG may be due to several reasons.
• the coupling of the monomeric IgG may cause the individual IgG molecules to change their conformation, so that the binding constant to RF is increased.
• the coupling may also have caused the IgG molecules to be seated so closely that an RF immuno ⁇ globulin is bound to more than one IgG molecule.
• IgG Sepharose CL-2B Eight ml of native human IgG (100 mg/ml in buffer A) are immediately added and incubated with the gel for 2 hours.
• IgG Sepharose CL-2B is washed with 40 ml of buffer A and incubated overnight after washing with 40 ml 1 M glycine in buffer A.
• IgG Sepharose CL-2B is washed with 40 ml of buffer A, 40 ml of 0.1 M glycine/HCl, pH 2.5, 50 ml of PBS (50 mM Na phosphate, 0.15 M NaCl, pH 7.35) and 100 ml of 0.9?,. NaCl, 0.5 unit heparin/ml.
• the immunosorbent prepared contains 20 mg of bound IgG/ml.
• IgG in a concentration of 50 mg/ml was incubated with between 0 and 20 mM dithiothreitol (DTT) for 30 minutes at room temperature. Then iodoacetamide was added in a concent-ration corresponding to the double of the DTT concentration. After 30 minutes at room temperature DTT and iodoacetamide were removed from the IgG solu ⁇ tion by dialysis against buffer A and then coupled to Sepharose CL-2B, as described in example 1. 15 mg of IgG/ml og gel were coupled. Then 1 ml of gel was incubated with 2 ml of plasma in which the RW titre is 1280. With the procedure described in example lb the results listed in Table 4 were achieved.
• DTT dithiothreitol

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• 1985
  • 1985-03-13 EP EP85901390A patent/EP0215775A1/de not_active Withdrawn
  • 1985-03-13 WO PCT/DK1985/000026 patent/WO1986005397A1/en not_active Ceased

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