WO2002009793A1

WO2002009793A1 - Device for treating immune diseases

Info

Publication number: WO2002009793A1
Application number: PCT/DE2001/002884
Authority: WO
Inventors: Torsten Matthias
Original assignee: Aesku.Lab Diagnostika
Priority date: 2000-07-27
Filing date: 2001-07-27
Publication date: 2002-02-07
Also published as: AU2001278407A1; DE10193004D2; WO2002009793A8; DE10036742A1

Abstract

The invention relates to a device for treating immune diseases, comprising an incoming line for blood and/or aqueous components thereof, which can be connected to a blood withdrawal site on the human body, an outgoing line which can be connected to a blood re-entry site on the human body and a support which is situated between the incoming line and the outgoing line and which has a surface. The device is characterised in that the surface of the support has immobilised antigens which selectively bind aggressive immunoglobulins. The immobilised antigens, which are preferably not in serum, are especially bound to the support by a non-covalent bond.

Description

description

DEVICE FOR TREATING IMMUNE DISEASES

The invention relates to a device for the treatment of immune diseases and the use of substances to which autoantibodies bind, such as e.g. of antigens, for the production of an agent for the treatment of immune diseases and a kit containing such agents.

The immune system's task is to ward off attacks on the organism and to fight foreign objects that have penetrated. It must not only differentiate between dangerous and harmless foreign bodies, but also be able to recognize the body's own substances. Dangerous attacks on the organism are, for example, the penetration of microorganisms, whereas the penetration of food antigens into the bloodstream when eating or the inhalation of bee pollen is harmless. In the defense, the destruction of foreign cell material is e.g. in the case of a parasitic infestation, as desirable as the destruction of the body's own malignant cells, whereas the attack on healthy body's own tissue, such as in autoimmune diseases, is undesirable. This means that the immune system has regulatory processes that prevent destructive self-reactivity. This ability is called "tolerance". If this tolerance is lost, immune diseases develop.

Immune diseases based on an unwanted immune response or response are playing an increasingly common role. For example, it is known that the frequency of immune reactions, which are directed against harmless foreign bodies such as grass pollen and animal hair, but also against food components, increases and leads to sensitization and thus to an allergy of an organism, which lead to serious diseases if sufficiently exposed. Another Re unwanted immune disease is the rejection reaction of the host versus graft type or in organ transplants, which are recognized as foreign by the body's immune system and are attacked and destroyed accordingly like a bacterium.

Another important undesirable malfunction of the immune system are the so-called autoimmune diseases. The body's own tissue or body's own cell substances or cells are wrongly regarded as foreign and are combated accordingly. The development of such autoimmune diseases is diverse. They can be triggered, for example, by intracellular substances entering the bloodstream as a result of injuries. Since these substances were previously hidden or enclosed in a cell, their antigens or epitopes could not be known to the immune system and could not be recognized as the body's own. They are therefore considered foreign to the body. For this reason, they are able to trigger an immune response. An example of such an autoimmune disease is sympathetic ophthalmia, in which a traumatic event releases an endogenous substance or an antigen that is normally enclosed in the eye.

Another cause for the development of autoimmune diseases is, for example, that antigens recognized by the immune system as the body's own, so-called "own" antigens, are changed by chemical, physical or biological effects and are therefore no longer regarded as "own" but as "foreign" In this way, they become immunogenic and can trigger an immune attack on your own body. It is known, for example, that some chemicals bind to the body's own proteins and thus make them immunogenic, as can be seen, for example, in contact dermatitis. In so-called sun allergy, skin proteins are changed by ultraviolet light so that they are no longer recognized by the immune system and trigger an immune response, making the patient allergic.

Finally, there is also the possibility that foreign antigens, for example of bacteria and viruses, trigger an immune reaction that produces antibodies that happen to cross-react with normal body's own antigens. That's the way it is For example, it is known that cross-reactions can occur between the streptococcal M protein and heart muscle proteins. Another example of this is the encephalitis triggered in rare cases by a rabies vaccination, in which the autoimmune cross-reaction is triggered by a vaccine that is not pure and contaminated with animal brain tissue. Finally, autoantibodies can also be the result of mutations in the formation of antibodies in immunocompetent cells. The development of autoimmune diseases and the failure of the body's immune tolerance is not yet fully understood, especially since genetic factors also play a role in autoimmune diseases.

A variety of attempts have been made to treat such immune responses. For example, for the treatment of allergies, attempts are made to prevent the histamine release from mast cells.

To prevent rejection of organ and tissue transplants, immunosuppressive substances are usually used, such as anti-inflammatory glucocorticoids, e.g. B. cortisone, non-steroidal anti-inflammatory drugs (NSAIDs), which intervene in the prostaglandin metabolism, immunosuppressive cytochins such as interleukin 10, antimetabolites such as metotrexate, cytostatics such as cyclophosphamide or inhibitors of inflammation-stimulating cytokines such as cyclosporin A or mycophenolinol, also mycophenolunol. Attempts have also been made to administer polyclonal antibodies for the treatment of autoimmune diseases in order to prevent the uptake of undesired antigens by antigen presenting cells (APC) and the presentation of these antigens to T cells. It is also possible to trap pathogenic autoantibodies using polyclonal antibodies.

Finally, attempts have already been made to contain the cellular immune reactions in autoimmune diseases by using anti-MHC antibodies to inhibit MHC detection or by using anti-TCR and anti-CD4 antibodies to inhibit T cell activation. Vaccination is carried out with peptides that are specific for the T cell clones involved in the disease. These competitively displace the pathogenic peptides on the MHC molecule and thus lead to a reduction in disease activity in the animal model. Finally, attempts have been made to remove more or less all immunoglobulin, ie all types of immunoglobulins, non-specifically in order to reduce rejection reactions. However, this has the disadvantage that the patient treated in this way no longer has any immune protection. This is particularly risky since the patient receives immunosuppressive substances anyway, especially in the case of transplants and autoimmune diseases. A patient treated in this way is exposed to virtually any attack by infectious microorganisms without protection.

Attempts have already been made to overcome the disadvantages described above by means of a patient-specific immune absorber. This should be used on patients who suffer from diseases that are caused by a disregulation of the immune system. For example, WO-97/14964 describes a procedure according to which the immune complexes circulating in it are isolated from the patient's blood using conventional methods and these are separated into their antigen and antibody components by methods known per se. The antigens and antibodies thus obtained are then covalently coupled individually or as a mixture to suitable carriers and used as an immune absorber for precisely those patients from whom the immune complexes have been isolated. In this way, specifically acting immune absorbers for patients suffering from autoimmune diseases are to be produced.

However, it has been shown that this approach for a variety of important life-threatening diseases, such as. B. Goodpasture syndrome, myasthenia gravis and / or Wegner's granulomatosis and a number of autoimmune asthma and heart diseases or celiac disease does not work. This is because WO97 / 14964 assumes that the immunocomplexes isolated from the blood are also used to obtain the agent which causes disease or causes disease. It has now been shown that this is usually not correct, so that in many cases treatment with absorbers produced in this way fails.

In addition, the manufacture of such patient-specific immune absorbers is extremely time consuming. Considering that in many critical cases, such as Wegner's granulomatosis and / or Goodpasture syndrome, the patient often die in 1 to 2 days, this procedure is not applicable. In addition, as already mentioned above, in a large number of cases the antigens which cause the disease, ie which trigger the autoimmune disease, are not obtained. Antigens that cause severe allergies such as e.g. B. trigger celiac disease.

In addition, even with the methods described there, it is not possible to prevent the development of immune diseases.

The aim of the invention is therefore to overcome the disadvantage of the prior art and to provide a means and a device with which undesirable immune diseases can be treated without completely suppressing the patient's immune system. The agent should be quick and inexpensive to manufacture, easy to store and quick to assemble, ready to use, and moreover, it should also be quickly adaptable to the patient's immunological needs on site. The invention is also intended to treat particularly fatal immune diseases such as sprue, celiac disease, Goodpasture syndrome, myasthenia gravis, Wegner's granulomatosis, asthma and heart diseases, IgE-mediated allergies, pemphigus vulgaris, autoimmune hepatitis, Rasmussen's encephalitis, Schnitz / or enable chronic urticaria. The invention also aims to preventively prevent the development of immune diseases and / or to reduce or to avoid the consequential damage of such diseases.

This goal is achieved by the features defined in the claims.

It was namely found according to the invention that misdirected immune reactions and the development of tolerance can be improved by selectively removing the autoaggressive immunoglobulins. According to the invention, in particular those B cells which form such immunoglobulins and their immunoglobulins or antibody-presenting precursors or precursor cells are also removed.

The invention thus relates to a device for the treatment of immune diseases which has substances immobilized on a solid phase which bind gressive immunoglobulins, preferably selectively. Such substances are, in particular, antigens which trigger the immune disease. The blood or its aqueous constituents are washed around the carrier, the pathogenic immunoglobulins being removed from the blood. Those immunocompetent cells and precursor cells which form the aggressive antibodies, in particular B cells, are preferably also removed. This effect on which the invention is based is all the more surprising because it was to be expected that the exposure of antigens to the immune system of the blood should increase the immune response, ie the formation and post-production of immunoglobulins directed against it (so-called booster Effect). However, it has surprisingly been found according to the invention that the aggressive immune response can be reduced or even down-regulated in a targeted manner if the undesired aggressive antibodies or immunoglobulins are selectively removed. This selective removal is preferably achieved according to the invention by means of an antigen directed against the aggressive antibodies, against which the aggressive immunoglobulin is directed. For the treatment of transplant rejections, these are in particular the foreign HLA or MHC antigens. For the treatment of autoimmune diseases, these are the antigens which trigger autoimmunity. B. in Wegner's granulomatosis, proteinase 3. The antigens bound in the device according to the invention are immobilized on a solid phase by methods known to the person skilled in the art. Such immobilization reactions are well known to the person skilled in the art from the production of diagnostic test kits such as ELISA or from affinity chromatography. The immunoglobulin-binding substances can be bound to the solid phase either directly or indirectly. They are preferably connected to one another by means of a non-covalent coupling consisting of at least 2 complementary coupling members. The coupling member attached to the immobilized agent is referred to as the anchor and the complementary coupling member bound to the solid phase is referred to as the counter anchor. Examples include only immobilization using cyanogen bromide or fixation using glutaraldehyde (direct binding), and biotinylation and the formation of avidin or streptavidin complexes.

According to the invention, the antigens are bound to the surface or solid phase by means of a spacer, ie an arm or spacer. According to the invention preferred spacers allow a distance of the antigen from the solid surface or the solid phase of preferably at least 5 A, in particular at least 10 Å, with at least 15 A and in particular at least 20 Å being particularly preferred. Spacer arms which allow a distance of at least 28 A, in particular at least 30 A, are very particularly preferred. The spacers are expediently constructed from a long carbon chain, but they are preferably constructed from repetitive units of smaller hydrocarbon chains, which in particular have heteroatoms. Preferred repetitive units are in particular C ₂ -C ₈ units, with C to C _{6 being} particularly preferred. The individual units are preferably linked to one another via the heteroatoms. Preferred heteroatoms are nitrogen, oxygen and sulfur atoms. The repetitive units are preferably connected to one another via amino acid functions.

The spacer arms preferably have different reactive groups at their two ends, one group of which reacts with the antigen to be bound, ie to be immobilized, and the other binds to the surface and, where appropriate, reacts there with it. Preferred functional groups which react with the antigens are, in particular, N-hydroxysuccinimide, sulfo-N-hydroxysuccinimide groups which react, for example, with a primary amine to form an amide group with the antigen or maleimide groups, for example with a free sulfhydryl group of an antigen to form a thioether bridge. Other reactive groups are haloacetyl, especially an iodoacetyl group, which also reacts with sulfhydryl to form a thioether bridge. Other important reactive reagents are, for example, hydrazide groups, which react with an oxidized carbohydrate group, for example an aldehyde group to form a hydrazone bond, further important groups are, for example, photoactivatable azido groups, for example aromatic azido groups, which contain nucleic acids and protein carbohydrates react with a primary amine to form a ring expansion. Finally, amino groups are used which react to form a free carboxylic acid or carboxy group to form an amido bond. Many such reagents are commercially available. In the case of the aforementioned functional reagents, the reagents containing N-hydroxysuccinimide and the maleimide containing reagents and the amino groups and the photoactivatable azido groups are preferred.

In a further preferred embodiment according to the invention, the spacer arm has an anchor group which, in particular non-covalently, couples with a counter anchor immobilized on the solid surface. The anchor on the spacer arm is preferably arranged at the point opposite the binding point of the spacer with the antigen. Anchors preferred according to the invention are biotin, thioredoxin, and / or polyhistidine tags, in particular those with 4 to 6 histidine residues adjacent to one another. Preferred counter anchors are avidin and in particular streptavidin as counter anchor to biotin, a chelato-immobilized metal ion, in particular nickel and / or cobalt, expediently as counter anchor to polyhistidine tags, and an antibody directed against the anchor. Such antibodies are easy to produce and in some cases also as monoclonal antibodies e.g. from ACC-2207 (antipentahistidyl antibody) commercially available from Quiagen. Such anchors and spacers containing reactive groups can be readily produced by a person skilled in the art and are also commercially available, for example. For example, Pierce, Rockford, IL, 61105, USA sells them under the name EZ-Link ™ and under the name Sulfo-LC-SPDP, LC-SPDP and SPDP. Fissile crosslinking reagents are also commercially available under this name.

The procedure according to the invention makes it possible to provide the surface of the solid phase with the counter-anchor first, even under reaction-hostile protein and nuclein conditions. Since it is not necessary to pay attention to biologically compatible reaction conditions, very high densities of counter anchors can also be achieved on the surface of the solid phase, as was previously not possible with the direct immobilization of antigens. After completion of the solid phase thus provided with the counter anchor, it can be loaded with the antigen under physiological conditions using its anchor day. In this way it is possible to achieve loading densities of> 0.01 pmol / cm ² > 0.1 pmol / cm ² and in particular> 1 pmol / cm ² . Even densities of> 5 pmol / cm ² can be easily achieved. If columns are used, densities of> 10 nmol / cm ³ , in particular> 20 nmol / cm ³ and beyond, can be produced, for example, in the case of beads with a diameter of 40-90 μm. When using the streptavidin The biotinylin system, which can bind up to four biotinylin anchors, enables exceptionally high loading densities. In this way, small, highly effective immune absorbers can be produced.

This has the advantage that in the treatment, i.e. H. in the case of extracorporeal apheesis, there is only a small volume of blood outside the body in the absorber, which avoids the risk of anemia in the already severely weakened patient. Furthermore, it is possible to use these small absorbers to produce aerial devices which are so small that they can be worn on the patient's body during the treatment. The patient is therefore not tied to a hospital or other medical care facility, but can remain at home during the treatment period. It is also possible to carry out long-term treatments and long-term treatments, during which the patient remains mobile and can move around the home completely unhindered. With more or less already healed patients, such a mobile or transportable device even enables them to pursue their usual occupations and needs.

Finally, the procedure according to the invention also makes it possible to produce the required immune absorbers on site, ie. H. For example, in a clinic or other therapy center to manufacture yourself if necessary. Since it is no longer necessary to store the entire required devices, but only the non-specific solid phases provided with the anchor, i. H. Columns, dialysis tubes, hollow fibers etc. as well as the respective antigens provided with the spacer arm and anchor, it is possible, if necessary, to immediately produce a specific immune absorber for the particular disease required by producing a solution of the antigen provided with spacer and anchor and in contact with the surface of the solid phase. Since the respective antigens can be stored for a long time, for example in the lyophilized state, no large, voluminous storage is necessary in order to provide a large number of absorbers for various diseases.

In addition, it is also possible to rapidly combine combinatorial columns or absorbers in patients who have different autoimmune diseases at the same time to produce, in which the absorber is loaded with a combination of different antigens. This enables patients with heterogeneous autoantibodies, such as those with simultaneous occurrence of lupus erythematosus and simultaneous antiphospholipid syndrome, to be treated inexpensively and effectively in one step using DS-DNA, β-2-glycoprotein and cardiolipin as a pre-made antigen.

Another advantage is that it is designed by providing a general prefabricated absorber that can be quickly prepared for a variety of different diseases.

The procedure according to the invention thus makes it possible to bind the antigens in their native physiological conformation without being sterically hindered by the immobilization on the surface, as a result of which almost all epitopes of the antigen are accessible to the antibodies to be removed or to the cells producing them , The anchor systems used according to the invention bind the antigen to the absorber with a high affinity, thereby preventing detachment or bleeding. Furthermore, the binding of the antigen according to the invention to the column enables washing of the bound autoagressive substances after use, for. B. by using high salt solutions and / or changes in pH. In this way, the respective immune absorbers can be used multiple times, i. H. be recycled.

According to the invention, carrier materials are all substances to which antigens or components thereof can be bound. Usual carrier materials to which the antigens are bound are also known from the manufacture of diagnostic test kits and for the manufacture of dialysis tubes. In principle, they are not subject to any restrictions, they may only be non-toxic and not themselves be allergic. Suitable carrier materials are, for example, silica gels as used for packed columns, latex particles, in particular non-allergenic latex particles, glass beads, gel matrices and hollow body matrices such as filter materials and dialysis tubes. In principle, particles made of polystyrenes, polyvinyl alcohols, Sepharose etc. can also be used. The solid phases preferably have a large surface area around which the blood serum or an aqueous solution of components thereof flows, so that a maximum contact area between antigens and the immunoglobulins circulating in the blood is created. According to the invention, the term antigens means all substances against which aggressive antibodies are directed. These are usually proteins, nucleic acids, and complexes of these two, possibly also with membrane components. This includes not only the complete antigens, but also parts or fragments of them that have bindable epitopes, such as peptides, oligopeptides and oligonucleotides, which can possibly also be produced artificially. Preferred antigens to be immobilized according to the invention are double-stranded DNA, single-stranded DNA, nucleosomes, centromeres (kinetochore antigen), Jo-1 (histidyl synthetase), SCL-70 (topoisomerase I), PCNA (proliferating cell nuclear antigen) , SS-B / La, SS-A (Ro), U1-RNP, SN antigen (small nuclear antigen), histones, PR3 (proteinase 3), F _c fragment of immunoglobulin, citrulline, phospholipids such as cardiolipin, ANA , ribosomal protein, RA 33, PM-Scl, ANCA, GBM (basement membrane of the kidney glomeruli), acetylcholine receptors, insulin receptors, antibody-sensitized platelets. Other antigens are, for example, antigens of sympathetic ophthalmia and B5 for Behcet's disease, B 27 for ankylosing spondylitis, Reiter's disease, and acute anterior uveitis, B 35 for subacute thyroiditis, Cw 6 for psoriasis vulgaris, DR 3 and possibly DR 4 for dermatitis herpetiformis, celiac disease, and Graves' disease, type I diabetes mellitus, myasthenia gravis, systemic lupus erythematosus (SLE), idiopathic membranous nephropathy; DR 2 for narcolepsy and multiple sclerosis, DR 4 for rheumatoid arthritis, and DR 5 for Hashimoto's thyroiditis and pernicious anemia and DRw8 for juvenile chronic arthritis; as well as the class of the spectrins, in particular α- and β-fodrin. Insoluble membrane-bound antigens are particularly preferred according to the invention.

Using the procedure according to the invention, it is also possible to immobilize the respective antigens which trigger the autoaggressive reaction. It has been shown that these do not occur in the serum, particularly in the case of particularly fatal diseases. For example, with the procedure according to the invention, the respective main antigens, such as the C-terminal end of the α-3 chain of the Isolate type IV collagen of the glomerular basement membrane separately and provide it with the appropriate spacer arm having an anchor. As already mentioned, this can also be done recombinantly. In principle, it is also possible to introduce membrane-bound antigens of this type into phospholipid vesicle membranes and then to bind them to the surface of the absorber in the manner described above. Such important antigens that are identical for most affected patients are in particular transglutaminase and gliadin for celiac disease or sprue, and the aforementioned III collagen, in particular its C-terminal end for Goodpasture syndrome. Further special antigens to be used according to the invention are, for example, the acetylcholine receptor and the β-2-adrenergic receptor for the treatment of myasthenia gravis and the proteinase 3 which occurs in the α-granules of the neutrophilic granulocytes for the treatment of Wegner's granulomatosis. The use of β-adrenergic receptors, in particular β1 and β2 receptors, has proven to be particularly suitable for the treatment of asthma and heart diseases. Furthermore, IgE receptors can be used according to the invention as antigens for severe cases of IgE-mediated allergies. Another antigen of the desmosome, in particular desmocholine, which is special and not present in the serum and which is used for the treatment of pemphigus vulgaris. Further special antigens to be used according to the invention are the asialoglycoprotein receptor for autoimmune hepatis, the glutamate receptor for Rasmussen encephalitis, the interleukin-1-α receptor for Schnitzler's syndrome, and the IgE receptor for chronic urticaria.

According to the invention, not only are the autoantibodies involved in the formation of tissue-damaging immune complexes removed, but also the removal of such autoantibodies which can cause tissue damage per se. The application thus affects more diseases and can also be used as a preventive measure that removes the autoantibodies before the formation of immune complexes. In addition, not only real autoantibodies, but also other pathogenic antibodies can be removed, such as B. Allergy Antibodies.

Such a device according to the invention is now connected to a blood collection point arranged on a patient via a feed line analogous to dialysis. the blood being drawn from this and passed over the device according to the invention. According to the invention, not only the immunoglobulins which cause illness, but also these producing cells and precursors thereof are preferably bound to the immobilized antigens and the blood is freed of them. After this "dialysis or filtration", this "liberated" or "cleaned" blood is returned to the patient via a drain by means of a blood return point. The removal, filtration and recycling are preferably carried out continuously. Blood sampling or blood return point are devices such as are known, for example, from dialysis or from blood donation.

With the device according to the invention it is not only possible to remove immunoglobulins, but it is also possible to selectively remove immunocompetent cells, which are responsible for the excessive immune reaction, without depriving the patient of his remaining protection, which is necessary for sufficient immunity to pathogens , For example, according to the invention it is also possible to immobilize the immobilized antigens on fine iron particles and to fetch the immunoglobulins or cells carrying such immunoglobulins bound to them from the blood or serum using a magnet (immunomagnetic separation, IMS). Such particles are available, for example, from Bekton-Dickensen under the name Dynabeads. Another possibility is, for example, to provide the antigens with a cytometrically recognizable marker, in particular fluorescent marker, and then to remove corresponding unwanted immune cells by means of flow cytometry. For the treatment of allergies, for example, it is particularly desirable to remove IgE mast cell complexes from the blood.

Although in a patient treated in accordance with the invention in this way the immune tolerance is brought back into equilibrium solely by removing the aggressive immunoglobulins, whereby the patient is primarily cured, in individual cases it may prove to be useful additionally to immunosuppressive substances such as glycorticoids or corresponding antimetabolites over a period of time To administer cytostatics etc. However, it has been shown that the high dose that would otherwise normally have to be applied for the treatment of immune diseases is never necessary for this. The invention thus also relates to a therapy kit which device according to the invention or the agent produced according to the invention together with an immunosuppressant.

According to the invention, it is also possible to connect a plurality of devices having different antigens in series and to remove a number of different aggressive immunoglobulins in this way. This can be done, for example, by filling the antigens immobilized on particles in columns and connecting several columns coated with different antigens in series. In principle, this is also possible with appropriate dialysis tubes or with filter matrices or hollow fibers.

The diseases to be treated with the device according to the invention are in particular all allergic and autoimmune diseases in which antibodies play a role relevant to the disease, i.e. have a pathogenic effect. These include, in particular, systemic lupus ery-thematodes (SLE), Sjogren's syndrome, scleroderma, polymyositis, dermatomyositis and mixed collagen disease (MCTD) as well as rheumatoid arthritis, primary vasculitis, and antiphospholipid syndrome, gypsum , Polychondritis, uveitis, Behcet's disease and especially the Goodpasture syndrome (AK against renal basal membrane), as well as myasthenia (myasthenia gravis), multiple sclerosis, cardiomyopathy, heart failure and pemphigus vulgaris.

The invention will be briefly explained using the following example.

To treat Wegner's granulomatosis, purified proteinase 3 (PR 3) is concentrated to about 10 mg / ml by means of ultrafiltration over a 10 kDa membrane (made of polyether sulfone or regenerated cellulose). The concentration is estimated by means of the absorbance at 280 nm by dilution of 1: 100 in PBS, using pure PBS as the basic value. The measured extinction value is to be multiplied by 100, a value of 1 corresponding to approximately 1 mg / l protein.

The solution thus concentrated is buffered on a 5 ml hightrap desalination column (Pharmacia, Sweden) in PBS, with a maximum of 1 ml sample per run being charged. The protein solution thus obtained is contained with a 10 mg / ml A sulfo-NHS-LC-LC-biotin (Pierce, USA) in water was mixed with 10 mM PR3 (M = 670 daltons), namely at least 110 μg sulfo-NHS-LC-LC-biotin per mg of PR3. Since this reagent is very sensitive to hydrolysis, it should be processed as quickly as possible. The reagent is added to the protein in about 4-fold excess and incubated on ice for two hours. Unreacted reagent is separated by means of gel filtration, as previously described for the buffering. The biotinylated antigen obtained in this way can be stored with 0.1% sodium azide at 4 ° C. and soon processed further or lyophilized for longer storage.

The PR3 antigen thus obtained, which was provided with a biotin anchor, was placed on a column containing polymethacrylate beads, which are coated with monomeric avidin. Such avidin-coated polymethacrylate beads are available, for example, under the name Softlink ™ from Promega, Wl, USA. For this purpose, the material is incubated for 15 minutes at room temperature with 5 mM biotin in order to suppress non-specific bindings. Then the column was eluted with 10% acetic acid and then with PBS. The biotinylated PR3 antigen was then loaded onto the column and then washed.

The column thus obtained was charged with 500 ml of plasma from a Wegner patient. The antibody titer of the plasma showed an OD of 1.9 in an anti-PR3 ELISA test from Aesku.lab Diagnostika before application to the column. After passing through this column (10 ml column) it still showed an OD of 0.15. This means that with a single pass on such a tiny column, 92.1% of the disease-causing agent has already been removed.

Claims

claims:

1. An apparatus for the treatment of immune diseases comprising a supply line for blood and / or aqueous components thereof which can be connected to a blood sampling point arranged on a human body, a lead line which can be connected to a blood return point arranged on the human body and a surface which is arranged between supply line and discharge line Carrier, characterized in that the surface of the carrier has immobilized antigens which trigger the immune disease and which selectively bind aggressive immunoglobulins, the antigens being bound to the surface by means of a spacer.

2. Device according to claim 1, characterized in that the antigens are bound to the surface by means of a spacer.

3. Device according to claim 1, characterized in that the antigen is bound to the surface by means of a non-covalent bond comprising an anchor and a complementary counter anchor.

4. Device according to one of the preceding claims, characterized in that the spacer by means of a reactive group comprising an N-hydroxysuccinimide, sulfo-N-hydroxysuccinimide, maleimide, haloacetyl, hydrazide, an amino group and / or a photoactivatable azido group is bound to the antigen.

5. Device according to one of the preceding claims, characterized in that the anchor is biotin, thioredoxin, and / or a polyhistidine tag.

6. Device according to one of the preceding claims, characterized in that the complementary counter anchor is avidin, streptavidin, a chelato-immobilized metal ion and / or an antibody directed against the anchor.

7. Device according to one of the preceding claims, characterized in that the spacer comprises a chain of at least 5 A length, which may have heteroatoms in the carbon backbone.

8. Device according to one of the preceding claims, characterized in that the C chain consists of repetitive units consisting of C ₂ - C _8. , In particular C ₄ - C ₆ groups, preferably of aminohexane groups, hexanesulfide and / or oxyhexane groups consist.

9. Device according to one of the preceding claims, characterized in that the column has an antigen density of greater than 0.01 pmol / cm ² solid phase surface.

10. Device according to one of the preceding claims, characterized in that the immune disease sprue, celiac disease, Goodpasture syndrome, myasthenia gravis, Wegner's granulomatosis, asthma and heart diseases, IgE-mediated allergies, pemphigus vulgaris, an autoimmune hepatitis, a rasmussen encephalitis , Schnitzlers syndrome and / or chronic urticaria and the antigen is transglutaminase, type IV collagen of the glomerular basement membrane, acetylcholine receptor, proteinase 3 of the alpha granules of the neutrophils, beta-andrenergic receptors, in particular beta-2 and beta-1 receptors , IgE receptors, desmocholine, in particular desmosomes, asialoglycoprotein receptor, glutamate receptor, interleukin-1-alpha receptor, IgE receptor.

11. Device according to one of the preceding claims, characterized in that the carrier comprises a dialysis tube, a filter matrix, a hollow fiber, silica gel particles, glass beads, latex particles, polyacrylate and polymethacrylate particles and / or polystyrene particles.

12. Device according to one of the preceding claims, characterized in that it comprises one or more carriers connected in series.

13. Device according to one of the preceding claims, characterized in that the antigen comprises kidney glomeruli, single-stranded DNA, double-stranded DNA, proteinase 3, acetyl receptors, spectrins, nucleosomes, histones, nuclear proteins and antigens of the inside of the eye.

14. Device according to one of the preceding claims, characterized in that it contains immobilized proteases which digest the bound immunoglobulins.

15. Device according to one of the preceding claims, characterized in that it contains a metering device for metering additives.

16. Device according to one of the preceding claims, characterized in that the additives are immunosuppressants.

17. Use of antigens for the preparation of an agent for the treatment and prevention of aggressive immune diseases according to one of claims 1-16, in which the antigen is brought into contact with the blood, serum or components thereof of a patient to be treated, the antigen with the aggressive immunoglobulins react and the reaction product is removed.

18. Use according to claim 17, characterized in that the immune disease is an autoimmune disease, a transplant rejection and / or an allergy.

19. Therapy kit comprising a device according to one of claims 1-16 and / or an agent produced according to one of claims 17-18 and an immunosuppressive drug.