JPS5889272A - Immune reaction promotor - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention] TECHNICAL FIELD The present invention relates to an immune reaction suppressant.

Recently, autoimmune diseases and rejection after single organ transplantation have occurred.

It has been known that the immune system can sometimes cause reactions that are unfavorable to the body. Traditionally, immunosuppressive drugs have been administered as a treatment for these diseases, but immunosuppressive drugs have side effects that inhibit the normal immune system and are not satisfactory. Plasma exchange therapy, in which the patient's plasma is removed by centrifugation or evaporation and exchanged with normal human plasma, has been attempted, and excellent results have been reported. There are drawbacks. On the other hand, it has recently become clear that soluble factors involved in activation and suppression of the immune system exist in patient plasma. Therefore, if these soluble factors can be selectively adsorbed and removed from plasma, it is thought that immune reactions can be controlled. The main ones of these soluble factors are thought to be relatively low molecular weight substances with molecular weights of tens of thousands or less from the particles that pass through the ultrafiltration membrane, but immunosuppressive factors and immunostimulatory factors are found in similar molecular weight fractions. Therefore, in order to suppress the immune reaction, it is necessary to selectively adsorb the immune promoting factors. As a result of intensive studies, the inventors of the present invention found that the average pore diameter (
D) is in the range of 30 to 3-oooom, and the sum of the volumes of pores with a pore diameter in the range of 0.8D to 1.2D accounts for 80% or more of the total pore volume, and The present invention was achieved by discovering that a porous body having a negative charge is most suitable for the purpose of the present invention.

The average pore diameter of the porous body in the present invention is 50 to 100
If it is less than 501, the amount of adsorption of soluble factors is extremely low (and if it is more than 10001, the selectivity decreases. A more preferable range for the average pore diameter is 5.
It is 0-5001. It is desirable that the pore size distribution of the skeleton porous material be narrow. When the average pore diameter is D, the total pore volume is the sum of the volumes of pores with pore diameters in the range of 0.8D to 1.2D. If it is less than 80%, the selectivity will decrease. The above average pore diameter is appropriately selected depending on the molecular weight of the soluble factor to be removed. Further, the porous body of the present invention must have a negative charge on its surface, and a porous body without a negative charge has low selectivity.

Examples of the porous body used in the present invention include porous glass, porous silica, and porous alumina. Porous glass is produced by melt-forming alkali borosilicate glass, then heat-treating it in a transition temperature range, and then treating with acid a fine phase-separated glass. Porous silica is produced by acid treatment of an aqueous sodium silicate solution. Further, porous alumina is produced by firing a hydrated alumina compact. Of course, as mentioned above, these porous bodies
When the average pore diameter is 3oL1oooX and the average pore diameter is D, the sum of the volumes of pores with pore diameters in the range of 0.8D to 1.2D is 8096 of the total pore volume. It is necessary to account for at least the following. Among the above-mentioned porous materials, those having acidic groups such as silanol groups, carboxyl groups, and sulfonic acid groups on the surface have a high adsorption capacity for soluble factors and hardly adsorb useful components such as sugars and vitamins. Preferably used. Therefore, among the above-mentioned materials, porous glass and porous silica are preferably used, and porous glass is most preferred since it has excellent adsorption performance and high mechanical strength.

These porous materials have a pore customer volume of 0.1α, 21 or more, and 2
It is preferably cc/f or less, and 0.5 cc/1
As mentioned above, it is particularly preferable that it is 2 cc/f or less. 0
．． Below 1 α′f, the adsorption performance is insufficient;
When CC/1 or higher, the mechanical strength becomes low. The porous body used in the present invention preferably has a particle size in the range of 4 to 270 mesh, particularly 4 to 50 mesh for whole blood, and a particle size of 4 to 50 mesh for whole blood. For those that do not contain it, those with a content of 4 to 270 mesh are preferable.

In the case of porous glass and porous silica, the surface has a negative charge due to silanol groups, but substituents such as carboxyl groups and sulfonic acid groups that suppress the negative charge can be introduced depending on the surface of the porous material. can do. The carboxyl group is
Introduction of an amino group with r-aminopropyltriethoxysilane followed by succinic anhydride or succinic acid in the presence of carbodiimide. It can be introduced by reacting with the compound. After the aminosilanization treatment, the sulfonic acid group can be introduced by treating with glutaraldehyde to introduce an aldehyde group, and then treating this with taurine. Furthermore, another method for introducing charges is a method in which the porous body is treated with a polymer having a substituent having a negative charge such as a carboxyl group or a sulfonic acid group. Examples of such polymers include polyacrylic acid, polymethacrylic acid, polystyrene sulfonic acid, and copolymers of monomers and hydrophilic monomers that are constituent units of these polymers. As a hydrophilic monomer,
Hydrophilic acrylic acid ester monomer.

Examples include hydrophilic methacrylic acid ester monomers. Particularly preferred are the following general formula (1) salient sulfuric acid ester single phosphor, general formula (8''), (W) vLr
) is a copolymer consisting of a polymerizable monomer having an epoxy group represented by:

0M2= CjRt (1) co
o-it2-on.

am1m1' == (3Ryu' (■) (However, 1. In the above general formula, R,, R1', R,' are hydrogen or methyl groups; islands have 2 carbon atoms with or without substituents.
-5 divalent alkylene group or poly(oxyalkylene) group RJ has 1 to 5 carbon atoms with or without a substituent
5 divalent alkylene group or poly(oxyalkylene) group; 14.4j is hydrogen or an alkyl group having 1 to 3 carbon atoms, and the alkyl group may further have a hydroxyl group; ) The porous body used in the present invention needs to be sterilized when used for therapeutic purposes.

Steam sterilization is particularly preferred, and a copolymer coated with a polymer containing a polymerizable monomer having an epoxy group as a structural unit is used as the edible agent.

It is recommended to crosslink and insolubilize the material by heat treatment or the like after brewing. The copolymerization ratio of the polymerizable monomer having an epoxy group is
0.1 to 10% by weight is suitable.

As a method for coating the surface of a porous body with a hydrophilic polymer, a mixture of a hydrophilic polymer or the above monomer and a polymerization initiator can be used.
A method of dissolving the solution in a suitable solvent such as methanol or ethanol and coating it on a porous body by dipping, spraying, or hot coagulation can be adopted. , the coating layer gives the porous body a suitable blood affinity.

In addition, it is necessary that the adsorption performance of the porous body is not significantly reduced.

When coated with a copolymer containing a monomer containing a Kin group as a constituent component, the
By heat treatment for 4 hours, crosslinking insoluble (8LE),
Blood from patients with autoimmune diseases such as rheumatoid arthritis, autoimmune hemolytic anemia, thrombocytopenic purpura, or rejection after organ transplantation when in contact with blood, plasma, or its low molecular weight fractions.

Plasma or its low molecular weight fraction comes to exhibit a suppressive effect on the immune response of lymphocytes. Although its mechanism of action is not clear, it is a low-molecular-weight immune-stimulating factor (gr) in patient plasma.
It is thought that the okth factor) is selectively removed.

Methods for using the immune reaction suppressor of the present invention include circulating patient blood extracorporeally and directly and continuously bringing the blood into direct and continuous contact with the immune reaction suppressor, circulating patient blood extracorporeally and subjecting it to centrifugation or centrifugation. A method of obtaining plasma and continuously contacting it with the immunoreaction suppressant, and then removing the patient's blood or plasma or a fraction thereof from the body.

There are methods such as contacting the chain immune reaction suppressant in a batch manner and then returning it to the patient.

Hereinafter, one aspect of the use of the immune reaction suppressant of the present invention will be explained in more detail using figures. Part 1 is an example of a column packed with the immune reaction suppressant of the present invention. The main body 1 is
It has an inlet 2 and an outlet 3 for blood, and a filter 4 is provided at the inlet and outlet. Moreover, the immune reaction suppressant (porous material) 5 of the present invention is filled in the darkness of the filter. Blood, plasma, or a fraction thereof enters through inlet 2.

After passing through the filter 4 and coming into contact with the porous body 5, the soluble factors are deposited on the porous body, and then exit from the column through the outlet 3.
Ru. The material of the column can be glass, polyethylene, polypropylene, polycarbonate, polystyrene, polymethyl methacrylate, etc., but polypropylene, polycarbonate, etc., which can be cocooned in an autoclave, are particularly preferred.The filter is physiologically inert and has high strength. Any porous material may be used, but polyester material is particularly preferred, and those having a mesh size of 80 to 180 mesh are preferably used.When using the porous material of the present invention to adsorb and remove soluble factors in blood, As shown in Figure 2, a system can usually be adopted in which blood is taken out of the body, then supplied to the column 1 using a pump 6, soluble factors are adsorbed and removed, and the blood is returned back into the body. Also.

In another system, as shown in Figure 6, blood is taken out of the body, blood cells and plasma are separated by a plasma separator 7, and then only the plasma is passed through a column 1, soluble factors are adsorbed and removed, and the blood cells are returned to the blood cells. It is possible to create a system that mixes with ingredients and returns them to the body. These systems can be freely selected depending on the purpose. Moreover, it can also be used by being incorporated into a system other than such an extracorporeal circulation system.

Example (1) Preparation of immune reaction suppressant Porous glass with an average pore diameter of 580X (pore diameter of 0.8D to 1.2D, proportion of pore volume 86%, pore volume 1 .39cc/7, surface area 90 m/f/, particle size 8Ω~120 mesh) was aminated by refluxing with ter-aminopropyltriethoxysilane in toluene, and further diluted with anhydrous succinic acid in dioxane. Carboxylation was carried out by reaction with acid at room temperature.

(;1 > Preparation of soluble factor solution Dialyze the patient's plasma after renal transplantation with a cellulose membrane to remove the immunosuppressant dissolved in the plasma, and filter and sterilize it with a filter (Millipore 8LG8.0.22μ). A soluble factor solution was obtained.

(ij) Soluble factor solution 0211t of soluble factor adsorption treatment (ii) and carboxylated porous glass 0 of (1) which is the immune reaction inhibitor of the present invention
．． No. 11 was contacted at 37° C. for 1 hour.

(iV) ') Preparation of lymphocyte suspension and PH reaction Heparinized peripheral blood of a healthy adult was layered on Ficoll-Paque and centrifuged at 6ΩO1 for 15 minutes to separate lymphocytes. The lymphocytes were adsorbed with ILPM 11640 medium supplemented with human type B blood and treated with 2X10' (11J) or untreated soluble factor I & (J, 025ml) and incubated at 67°C. % CO2 incubator. On the 5th day, 25 μci/ml of tritiated thymidine (1,025 ml) was added, and the incubation was carried out again for 24 hours in a C sham incubator at 37°C.
- Tritium taken up by lymphocytes was counted using a counter.

(V) Calculation of PfI membrane reaction inhibition rate The RAM response inhibition rate of soluble factors is x-100(*) In addition, the inhibition rate of PHM reaction by adsorption treatment of soluble factors is xloo (%) I asked for it as.

(■1) Results The inhibition rate of the PHM reaction by soluble factors that were not adsorbed was 74.5%. Furthermore, when soluble factors were treated with carboxylated porous glass, the inhibition rate increased to 91.6% for lymphocytes B compared to untreated cells, and to 48.1% for lymphocytes B compared to untreated cells. , lymphocyte Pl by treatment with carboxylated porous glass
It was found that the inhibitory effect of soluble factors on the IM reaction was enhanced.

[Brief explanation of drawings]

FIG. 1 is a diagram showing an example of a column filled with the immune reaction suppressant of the present invention. FIGS. 2 and 3 are examples of a system for treating soluble factors in blood or plasma using the immune reaction suppressant of the present invention. Patent Applicant Riichi Kinsha Co., Ltd. Agent Patent Attorney Honda-Pakkenbo 1 @ ↓ Written amendment to the procedure for Figure 3 (voluntary) January 22, 1980 Date and time - 156-1875216 No. 2, Invention Name of Immune Reaction Suppressant Representative Director Tsugu Okabayashi 6 Contents of Amendment (1) Name of the invention in the specification tr Immune Reaction Suppressant” (2) Amend the claims in the specification as shown in the attached sheet. (5) In the following portion of the specification, "immune reaction inhibitor" is amended to "immune reaction inhibitor". 1) Page 2, line 2 2) $99 page 63) Page 9, line 16 5) Page 9, line 20 6) Page 10, line 2 7) Page 10 4th line 8) @1o page 6th line? ) Page 10, line 10 10) Page 11, line 18 11) Page 12, lines 15-16 12) Page 14, line 15 13) Page 14, line 15 and above Attachment 2, Claims 1. The average pore diameter (D) is in the range of 30 to 1oooA, and the sum of the volumes of pores with a pore diameter in the range of 0.8D to 1.2D accounts for 80% or more of the total pore volume. immunoreaction inhibition by a porous material with a negative charge on the surface and a negative charge on the surface
Sword. 2. The immune reaction suppressing material according to claim j1g1, which has an average pore diameter (D) in the range of 50 to SOa. 5. The immune reaction suppressing material according to claim 1 or 2, wherein the porous body is selected from porous glass, porous silica, and porous alumina. 4. The immune reaction suppressing material according to claim s1 or JIZ, wherein the porous body is porous glass. 5.jlL The immune reaction suppressing material according to any one of claims jg1 to IM4, which is a substituent that is separated from the sulfonic acid group. 380

Claims (1)

[Claims] 1. The sum of the volumes of pores with an average pore diameter CD) in the range of 50 to 1000 μm and a pore diameter in the range of 0.8 D to 1.2 D is the total pore size. An immune reaction inhibitor comprising a porous material occupying 80% or more of the volume and having a negative charge on the surface. 2. The immune reaction inhibitor of claim 1, item 1, wherein the average pore diameter (D) is in the range of 50 to s'oom. 5. The porous body is porous glass, porous silica, porous The immune reaction suppressant according to claim 11111 or 2, which is selected from alumina. 4. The immune reaction inhibitor according to claim II#I, item 1 or 2, wherein the porous body is porous glass. 5. Negative charge is 1. sila f-nol group 9, carboxyl group. The immune reaction inhibitor according to any one of claims 15 to 4, which is a substituent selected from sulfonic acid groups.