JP4182682B2 - Carcinoembryonic antigen adsorbent and extracorporeal circulation column - Google Patents

Description

【００３９】
表１の比較例１では極性基やイオン性基のリガンドを持たないため吸着性がないことが分かる。実施例４は比表面積が大きいため、吸着性が実施例５よりも吸着能が高い。これらの結果から、吸着には極性基もしくは親水性基が必要であることが分かる。また、イオン性基の中の４級アンモニウム基では、窒素原子１個当たりの炭素数が８以下のものは吸着能が小さいことが分かる。
【００４０】
【発明の効果】
本発明により、免疫抑制性物質を効率よく吸着除去することが可能となり、癌の患者の治療に有用である。[0001]
BACKGROUND OF THE INVENTION
The present invention relates to an adsorbent and an extracorporeal circulation column for removing carcinoembryonic antigen, which is thought to cause a significant decrease in immunity and promote the formation of cancer if it is excessively present in blood.
[0002]
[Prior art]
Even today, with the development of medicine, cancer remains one of the leading causes of death for Japanese people. The cause is that there are cancer cells that cannot be removed by surgery in patients, and anticancer drug treatment and radiation therapy have been performed to remove them. However, since these also damage normal cells, cancer cells cannot be completely removed while maintaining the patient's life. On the other hand, recently, cell therapy has been actively tried to improve the immunity of patients and eliminate cancer with their own white blood cells. The most promising is dendritic cell infusion therapy in which a patient's dendritic cells are stimulated with cancer antigens outside the body and then returned to the patient to induce and treat cancer-specific killer cells (CTLs). However, in many cases, CTL can be induced from the blood of healthy animals, but cannot be induced from cancer-bearing animals at the end of cancer. In animal experiments, even if good results are obtained, there is almost no visible effect in clinical practice. The reason for this is thought to be because immunosuppressive substances produced by cancer cells are present in the blood of the patient. One of the typical immunosuppressive substances is carcinoembryonic antigen, which is thought to increase abnormally with the progression of digestive organ cancer and specific lung cancer, and to help cancer cell growth. Since human carcinoembryonic antigen is a protein having a molecular weight of about 200,000, it is a substance that is difficult to adsorb and remove with a separation membrane or a conventional adsorbent. Therefore, no adsorbent is known that can efficiently remove abnormally increased carcinoembryonic antigen in the blood of cancer patients. The removal of these immunosuppressive substances is theoretically effective in plasma exchange, but has an unavoidable inherent disadvantage that there is a risk of being infected with an unexpected pathogen from a donor.
[0003]
[Problems to be solved by the invention]
The present invention is generally ubiquitous in view of the problems of the prior art, and selectively adsorbs immunosuppressive substances such as carcinoembryonic antigen directly from body fluids with high efficiency and safely. An object of the present invention is to provide a carcinoembryonic antigen-adsorbing material capable of extracorporeal circulation, which is useful for cancer treatment.
[0004]
[Means for Solving the Problems]
In order to achieve the above object, the medical carcinoembryonic antigen adsorbent and extracorporeal circulation column of the present invention have the following constitutions.
[0005]
(1) A carcinoembryonic antigen adsorbent obtained by binding a polar group or an ionic group to a water-insoluble carrier.
[0006]
(2) The carcinoembryonic antigen-adsorbing material according to (1), wherein the specific surface area of the water-insoluble carrier is 0.1 square meter or more per gram.
[0007]
(3) The carcinoembryonic antigen-adsorbing material according to (1) or (2), wherein the polar group is an amide group.
[0008]
(4) The carcinoembryonic antigen-adsorbing material according to (1) or (2), wherein the ionic group is an amine residue.
[0009]
(5) The carcinoembryonic antigen adsorbent according to any one of (1) to (4), wherein the ionic group is a quaternary ammonium residue having 12 or more carbon atoms per nitrogen atom.
[0010]
(6) The carcinoembryonic antigen adsorbent according to any one of (1) to (5), wherein the water-insoluble carrier is a polysulfone polymer.
[0011]
(7) The carcinoembryonic antigen-adsorbing material according to any one of (1) to (6), wherein the water-insoluble carrier is poly (aromatic vinyl compound).
[0012]
(8) The carcinoembryonic antigen-adsorbing material according to any one of (1) to (7), wherein the water-insoluble carrier is molded into a film, a fiber, a granular material, or an assembly thereof.
[0013]
(9) The carcinoembryonic antigen-adsorbing material according to any one of (1) to (7), wherein the water-insoluble carrier is coated on the surface of any one of a membrane, a fiber, and a granular material.
[0014]
(10) An extracorporeal circulation column packed with the carcinoembryonic antigen adsorbent according to any one of (1) to (9).
[0015]
(11) An extracorporeal circulation column for cancer treatment packed with the carcinoembryonic antigen adsorbent according to any one of (1) to (9).
[0016]
DETAILED DESCRIPTION OF THE INVENTION
Subsequently, the present invention will be described in more detail.
[0017]
The water-insoluble carrier used in the present invention is not particularly limited as long as it is insoluble in water and can fix a polar group or an ionic group. Poly (aromatic vinyl compound) represented by polystyrene, poly (p-phenylene ether) sulfone) and _{- {(p-C 6 H} 4) -C (CH 3) 2 - (p-C 6 H 4) -O- (p-C 6 H 4) -SO 2 - (p-C 6 H ₄ ) Polar groups or ions of polysulfone polymers represented by —O—} _n — (hereinafter abbreviated as “Udel polysulfone”), polyetherimide, polyimide, polyamide, polyether, polyphenylene sulfide, etc. Although the thing etc. which can fix | immobilize a sex group can be used, it is not limited to these. Reactive functional groups for immobilizing polar groups or ionic groups include halomethyl groups, haloacetyl groups, haloacetamidomethyl groups, active halogen groups such as alkyl halide groups, epoxide groups, carboxyl groups, isocyanate groups, Examples thereof include thioisocyanic acid groups and acid anhydride groups. In particular, active halogen groups, especially haloacetyl groups, are easy to produce, have moderately high reactivity, and are capable of fixing functional groups. It can be carried out under mild conditions, and the covalent bond formed at this time is preferable because it is chemically stable. More specific examples include chloroacetamidomethyl polystyrene, chloracetamidomethylated Udel polysulfone, chloroacetamidomethylated polyetherimide, and the like. Furthermore, when these polymers are soluble in an organic solvent, there is an advantage that they are easy to mold.
[0018]
In the present invention, the polar group means an amide group, a urea group, an ester group, an ether group, or the like. Specific examples of the amide group include chloroacetamidomethyl group, butyroylamidomethyl group, lauroylamidomethyl group, polypeptide, cyclic polypeptide and the like. In particular, a cyclic polypeptide represented by polymyxin B is preferable.
[0019]
The ionic group referred to in the present invention means an amino group, carboxyl group, sulfonic acid group or the like that can dissociate into ions in water. Specific examples of amino groups include laurylamine, polyethyleneimine, polymyxin B and the like, which are obtained by reacting primary and secondary amines with chloroacetamidomethyl group, N, N-dimethyloctylamine, N, N-dimethyllaurylamine, etc. A product obtained by reacting a tertiary amine having 10 or more carbon atoms with a chloroacetamidomethyl group can be raised.
[0020]
The bond density of polar groups or ionic groups in the present invention varies depending on the chemical structure and use of the water-insoluble carrier, but if it is too small, its function tends not to be expressed. Since the physical strength of the resin tends to deteriorate and the function as an adsorbent tends to decrease, the density is 0.0001 to 1.0 mol, more preferably 0.001 to 1.0 mol, per repeating unit of the water-insoluble carrier. Is good.
[0021]
The specific surface area in the present invention means the surface area per gram of the adsorbent. Since carcinoembryonic antigen is a large molecule having a molecular weight of 150,000 daltons or more, the adsorbent must have a large surface area. The specific surface area of the adsorbent of the present invention is preferably 0.1 square meters or more per gram of the adsorbent, more preferably 1 square meters or more. However, since it cannot be infinitely large, there is a practical limit, and 100 square meters or less is preferable. This specific surface area can be determined by a nitrogen gas adsorption method (BET method). As a method of increasing the specific surface area, there are a method of forming a fiber or making a molded product porous. Specifically, in order to make a carrier having a specific surface area of 0.1 square meter per gram, in the case of fiberizing polyethylene terephthalate, a fiber having a diameter of about 30 micrometers may be made.
[0022]
The adsorbent of the present invention is formed by molding a water-insoluble carrier bonded with a hydrophilic amine residue into a film, fiber, granule or an assembly thereof so that the specific surface area is 0.1 square meter or more per gram, Alternatively, a water-insoluble carrier bonded with a hydrophilic amine residue may be coated on a substrate of a film, fiber, or granule having a specific surface area of 0.1 square meter or more per gram, or the specific surface area per gram. It can be obtained by bonding a hydrophilic amine to a molded article such as a water-insoluble carrier film or fiber of 0.1 square meter or more.
[0023]
For the production of the adsorbent of the present invention, a heterogeneous reaction method in which a water-insoluble carrier solution is brought into contact with a molded product of a water-insoluble carrier, and a water-insoluble carrier solution and a hydrophilic amine solution are mixed and reacted. And a homogeneous reaction method of molding. As an example of a heterogeneous reaction method, a molded product such as a fiber or hollow fiber of chloroacetamidomethylated polysulfone is immersed in an isopropanol solution such as polyalkyleneimine and reacted at a temperature of 0 to 100 ° C. To be achieved. An example of the method by homogeneous reaction is achieved by adding the corresponding polyamine to a solution of chloroacetamidomethylated polysulfone and reacting at a temperature of 0 to 100 ° C. The amount is not particularly limited, but it is desirable to use at least 1 mole per mole of haloacetamidomethyl groups to obtain a soluble polymer. In particular, in the case of polyamine, it is preferable to use a large excess of hydrophilic amine in order to obtain a soluble carrier.
[0024]
As the reaction solvent, a solvent having a high polarity such as water, methanol, ethanol, isopropanol, dimethyl sulfoxide, N, N-dimethylformamide (DMF) has an advantage that the reaction proceeds faster. The heterogeneous reaction is not particularly limited as long as it is a solvent in which the hydrophilic amine is soluble. When the reaction is carried out in a homogeneous system, a solvent in which both the water-insoluble carrier and the hydrophilic amine are dissolved, specifically tetrahydrofuran, dimethyl sulfoxide, N, N-dimethylformamide, N, N-dimethylacetamide, N-methylpyrrolidone Etc. are preferably used. In addition, a method of surface-treating the molded product is also possible. For this purpose, a solvent that dissolves the hydrophilic amine without dissolving the insoluble carrier, such as water, methanol, or ethanol, is preferably used.
[0025]
When the adsorbent polymer of the present invention is coated on the surface of a molded product such as polyester fiber, nylon fiber, polyphenylene sulfide fiber, etc., there is an advantage that a high-order molded product having a large surface area can be obtained easily and inexpensively. Coating can be easily achieved by immersing a nylon knitted fabric or woven fabric in the adsorbent of the present invention dissolved in a low-boiling solvent such as methylene chloride or tetrahydrofuran and evaporating the solvent. Further, a wet coating method in which a solution dissolved in a solvent such as N, N-dimethylformamide is put into a poor solvent for a polymer such as water can also be used. The polymer of the molded article to be coated may be any polyamide, polyurethane, polyimide, polysulfone, polyvinyl chloride, polyester, etc., as long as it has good adhesion to the adsorbent polymer of the present invention, and there is no particular limitation on the type. An amide polymer such as nylon or polyetherimide is preferably used because of its particularly good adhesion.
[0026]
In the above molding and coating on the base material, it is also preferable to use a hollow fiber as a fiber to be employed in the form of the molded product or the base material. In this case, since an adsorbent having a filtration function can be produced, there is an advantage that an immunosuppressive substance can be removed while being used as an artificial dialyzer or a plasma separator.
[0027]
The adsorbent of the present invention is used for the treatment of extracorporeal circulation of diseases such as cancer in a packed state for the purpose of removing immunosuppressive proteins such as carcinoembryonic antigen in blood. This column can be produced by packing the adsorbent of the present invention in the form of cotton, cylinder, or felt into a cylindrical column having an appropriate size so that the void volume is about 200 mL or less. The adsorbent of the present invention can also be used for the purpose of removing carcinoembryonic antigen from serum and plasma.
[0028]
【Example】
Hereinafter, the present invention will be described more specifically by experimental examples.
[0029]
In addition, the evaluation method in a present Example followed the following.
1. Analysis of components in blood The concentration of carcinoembryonic antigen in serum was determined using CEA Test Kit TM-201 from Hope Laboratory. The total protein concentration was determined by the Burette method.
2. Nitrogen gas adsorption method (BET method)
About each measurement sample, the adsorption isotherm of nitrogen gas 77K was measured after deaeration pretreatment at 100 ° C. using a high-precision fully automatic gas adsorption apparatus “BELSORP36” manufactured by Nippon Bell Co., Ltd. The specific surface area was determined by applying the BET multilayer adsorption theory to the isotherm.
[0030]
(Production of water-insoluble carrier (raw yarn 1))
Thirty-six sea-island composite fibers, each of which is made of a core-sheath composite, were obtained using the following components under the spinning conditions of a spinning speed of 800 m / min and a draw ratio of 3 times.
Island core component; Polypropylene island sheath component: 90% polystyrene, 10% polypropylene
Sea component; Polyethylene terephthalate composite ratio obtained by copolymerization of 3% of 5-sodium sulfoisophthalic acid; Core: sheath: sea = 40: 40: 20.
This sea component was dissolved in a hot caustic soda aqueous solution to obtain a raw yarn 1 having a diameter of 4 μm as a core-sheath type polypropylene reinforced polystyrene fiber.
[0031]
(Example 1)
6 g of paraformaldehyde was dissolved in a mixed solution of 1200 mL of nitrobenzene and 780 mL of sulfuric acid at 20 ° C., cooled to 0 ° C., and 152 g of N-methylol-α-chloroacetamide was added and dissolved at 5 ° C. or lower. 20 g of the raw yarn 1 prepared above was immersed in this, and allowed to stand at room temperature for 2 hours. Thereafter, the fiber was taken out and placed in a large excess of cold methanol for washing. The fiber was thoroughly washed with methanol, washed with water, and dried to obtain 30.0 g of α-chloroacetamidomethylated polystyrene fiber (Example 1).
[0032]
(Example 2: Immobilization of hydrophobic quaternary ammonium group by heterogeneous reaction)
5 g of Example 1 was immersed in a solution of 50 g of N, N-dimethyllaurylamine and 8 g of potassium iodide in 360 mL of DMF and heated in a bath at 85 ° C. for 3 hours. The fiber was washed with isopropanol, immersed in a 1 mol / L saline solution, washed with water, and dried under vacuum to obtain 9.3 g of dimethyllauryl ammoniumated fiber (Example 2).
(Example 3: Immobilization of polymyxin B by heterogeneous reaction)
10 g of Example 1 was immersed in a solution of 1 mg of a polymyxin B sulfate aqueous solution having a concentration of 0.2 mg / mL, 1M sodium hydroxide solution was added, the pH was set to 10, and the mixture was shaken at room temperature for 24 hours. Dilute hydrochloric acid was added to the reaction mixture to adjust the pH to 4, followed by washing with water and further vacuum drying to obtain 10 g of polymyxin B-immobilized fiber (Example 3). As a result of amino acid analysis of this fiber, the amount of polymyxin B bound was 10 mg per 1 g of fiber.
[0033]
(Examples 4 and 5: Synthesis and coating of hydrophilic amine-bonded polymer)
After cooling a mixed solution of 16 mL of nitrobenzene and 32 mL of sulfuric acid to 0 ° C., 4.2 g of N-methylol-α-chloroacetamide was added and dissolved, and this was dissolved in 3 L of nitrobenzene solution of Eudelpolysulfone P3500 at 10 ° C. 300 g / 3 L) with good stirring. Furthermore, it stirred at room temperature for 3 hours. The reaction mixture was then placed in a large excess of cold methanol to precipitate the polymer. The precipitate was washed well with methanol, dried, and reprecipitated from dimethylformamide / methanol to obtain 303 g of α-chloroacetamidomethylated polysulfone (substitution rate: 0.05; polymer A).
[0034]
A solution obtained by dissolving 60 g of polyethyleneimine (average molecular weight 10,000: Wako Pure Chemical Industries, Ltd.) in 300 mL of DMF and 300 mL of DMF solution containing 30 g of Polymer A were mixed and stirred at room temperature for 48 hours. The reaction mixture was added to a large excess of saturated brine, and the precipitated polymer was collected by filtration. The polymer was washed with water, dried, and reprecipitated from dimethylformamide / methanol to prepare 27 g of N-alkylated polyalkylenimine-immobilized polysulfone (Polymer B).
[0035]
In a solution of 250 mL of methylene chloride containing 5 g of polymer B, 20 g of polyethylene terephthalate fiber cotton having a single yarn diameter of 3.5 μm is dipped. After 20 hours, the cotton is taken out, drained and air-dried, and 21 g of coated cotton. (Example 4) was obtained. In addition, 10 g of polyethylene terephthalate fiber having a single yarn diameter of 100 μm was immersed in 250 mL of methylene chloride containing 5 g of the polymer B, and after 20 hours, the cotton was taken out, drained and air-dried, and 10.4 g Coated cotton (Example 5) was obtained.
[0036]
(Example 6: Fixation of hydrophilic quaternary ammonium group by heterogeneous reaction) 5 g of Example 1 was immersed in a solution of 50 g of N, N-dimethylhexylamine and 8 g of potassium iodide in 360 mL of DMF. Heated in a bath at 3 ° C. for 3 hours. The fiber was immersed in a 1 mol / L saline solution, then washed with water and vacuum dried to obtain 7.3 g of dimethylhexylammonium-containing fiber (Comparative Example of Example 6).
[0037]
(Evaluation of adsorption capacity)
Recombinant human carcinoembryonic antigen (Chemicon International; CEA) was added to human serum (Cosmo Bio; 812-10) and shaken at 37 ° C. for 1 h to prepare CEA model serum (89 ng / mL concentration) did. 50 mg of the adsorbent was added to 1 mL of this serum, and after shaking for 4 hours at 37 ° C., the CEA concentration in the supernatant was measured, and the results shown in Table 1 were obtained. As Comparative Example 1, cotton that had not been fixed to polyethylene terephthalate fiber having a diameter of 3.5 μm used for preparation of Example 4 was used.
[0038]
[Table 1]

[0039]
It can be seen that Comparative Example 1 in Table 1 has no adsorptivity because it does not have a polar group or ionic group ligand. Since Example 4 has a large specific surface area, the adsorptivity is higher than that of Example 5. From these results, it is understood that a polar group or a hydrophilic group is necessary for the adsorption. Further, it can be seen that the quaternary ammonium group in the ionic group has a small adsorptivity when the number of carbon atoms per nitrogen atom is 8 or less.
[0040]
【The invention's effect】
According to the present invention, an immunosuppressive substance can be efficiently adsorbed and removed, which is useful for treatment of cancer patients.

Claims (11)

An amide group or an ionic group is bonded to a water-insoluble carrier, and the density of the amide group or the ionic group is 0.0001 to 1.0 mol per repeating unit of the water-insoluble carrier, A carcinoembryonic antigen adsorbent, wherein the specific surface area of the carrier is 0.1 to 100 square meters per gram . The ionic groups are amino groups, according to claim 1 Symbol placement carcinoembryonic antigen adsorbent. The carcinoembryonic antigen adsorbing material according to claim 2, wherein the amino group is obtained by reacting polymyxin B, N, N-dimethyllaurylamine or N, N-dimethylhexylamine with a chloroacetamidomethyl group. The ionic group has a carbon number per one nitrogen atom is a quaternary ammonium beam group is 12 or more, claim 1 carcinoembryonic antigen adsorbent according to any one claim of 3. The water-insoluble carrier is a polysulfone polymer, according to claim 1 carcinoembryonic antigen adsorbent according to any one claim of 4. The water-insoluble carrier is a poly (aromatic vinyl compounds), according to claim 1 carcinoembryonic antigen adsorbent according to any one claim of 4. The water-insoluble carrier film, fibers, granules or, characterized in that these are molded into assembly, according to claim 1 carcinoembryonic antigen adsorbent according to any one claim of 6. The water-insoluble carrier, film, fiber, yielding coated on either surface of the granules according to claim 1 carcinoembryonic antigen adsorbent according to any one claim of 6. The amide group is one or more amide groups selected from the group consisting of a chloroacetamidomethyl group, a butyroylamidomethyl group, a lauroylamidomethyl group, a polypeptide and a cyclic polypeptide. The carcinoembryonic antigen adsorbent according to one item. An extracorporeal circulation column packed with the carcinoembryonic antigen adsorbent according to any one of claims 1 to 9. Cancer therapy for extracorporeal circulation column carcinoembryonic antigen adsorbent is filled as claimed in any one of claims 1 to 9.