JP2003190276A - Virus and white blood cell selective removal method, removal material and remover - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method capable of removing virus and white blood cell at the same time from blood containing virus and achieving high recovery of platelet, and a material and a device for the method. <P>SOLUTION: A water insoluble carrier having the surface for capturing virus and white blood cell in blood and passing platelet is brought into contact with blood containing virus to remove the virus and the white blood cell at the same time from the blood, and the platele is recovered at high recovery rate. It is desirable that water insoluble carrier has an end hydrophilic group on the surface. <P>COPYRIGHT: (C)2003,JPO

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for simultaneously selectively removing viruses and leukocytes in blood, a selective removing material and an apparatus therefor.

[0002]

2. Description of the Related Art A conventional blood treatment system uses a plasma separation membrane or a centrifuge to separate a plasma component from a blood cell component, and then directly analyzes the plasma component with an immunoadsorbent or a low-density lipoprotein adsorbent. It was a system of contacting and adsorbing and removing unnecessary substances (for example, Patent Documents 1 to 3). In these systems, since blood cell components such as red blood cells, white blood cells, and platelets are collectively separated from plasma components, it is difficult to remove unnecessary substances such as viruses and white blood cells at the same time.
Furthermore, in these systems, since a charged material is used, platelets are activated and unnecessary substances and white blood cells cannot be removed, and at the same time, platelets cannot be collected at a high recovery rate.

Patent Document 4 describes a method of purifying blood by using a leukocyte removing material to simultaneously remove leukocytes and malignant substances such as immunoglobulin from the blood of patients with immune system diseases. However, neither the simultaneous removal of white blood cells and viruses nor the simultaneous recovery of platelets is disclosed.

Further, Patent Document 5 discloses a blood treatment apparatus and method for removing target substances such as viruses from blood without an anticoagulant by treating the blood with a carrier having a polyamine and an anticoagulant on the surface. Is disclosed,
Due to the large amount of amine on the surface, it was difficult to recover platelets sufficiently, and leukocyte removal was not sufficient.

Furthermore, as a virus removing material,
Patent Document 6 discloses a material having a cationic compound on its surface, but this publication makes no mention of removing viruses from blood. Further, Patent Document 7 discloses an agent for removing HIV and related substances, which comprises a solid substance whose surface is weakly acidic or weakly basic. This removing material has —COOH, —SO ₃ H or the like on the surface and has a surface pH of 2.5 to 6.9 or 7.4 to 1
It is characterized by being 0.5, and -CO existing on the surface
If the OH or -SO 3 _H or the like forms a salt is described as not able to remove viruses. Furthermore, in the method described in Patent Document 7, when the blood is brought into contact with the removing material, the pH of the blood changes, so that denaturation of the component protein occurs, which is not a preferable condition for blood. A problem common to these techniques is that there is a risk of blood coagulation due to denaturation of blood proteins when the removal material comes into contact with blood.

[Patent Document 1] JP-A-61-113463

[Patent Document 2] Japanese Patent Publication No. 5-50302

[Patent Document 3] Japanese Patent Publication No. 5-50303

[Patent Document 4] Japanese Patent Publication No. 5-50301

[Patent Document 5] Japanese Patent Laid-Open No. 11-267199

[Patent Document 6] Japanese Patent Laid-Open No. 3-123630

[Patent Document 7] JP-A-2-36878

[0006]

SUMMARY OF THE INVENTION An object of the present invention is to solve the above-mentioned problems of the prior art. In particular, virus and leukocytes are simultaneously removed from virus-containing blood,
Moreover, it is an object of the present invention to provide a method capable of achieving a high platelet recovery rate, a material and an apparatus therefor.

[0007]

Means for Solving the Problems As a result of intensive studies for solving the above problems, the present inventors have brought a water-insoluble carrier having a surface for removing viruses and leukocytes in blood into contact with virus-containing blood. As a result, they have found that viruses and leukocytes can be simultaneously removed from blood, and that platelets can be recovered at a high recovery rate, leading to the present invention. Furthermore, in the present invention, activated complement C3a in blood after contact is obtained.
The present invention has been made based on the finding that the virus can be efficiently removed together with leukocytes when a material having a concentration of 5 times or more the blood concentration before contact is used.

That is, the present invention comprises the step of contacting a virus- and blood-containing selective removal material comprising a water-insoluble carrier having a surface for adsorbing or removing viruses and leukocytes in blood with blood containing virus. The present invention relates to a method for simultaneously selectively removing viruses and leukocytes from blood.

Further, the present invention comprises a water-insoluble carrier having a surface for adsorbing or removing viruses and leukocytes in blood, for selectively simultaneously removing viruses and leukocytes from blood. , A platelet-passing virus and a leukocyte selective removal material.

Further, according to the present invention, the virus and leukocyte selective removing material is contained in a container having a blood introducing portion and a blood discharging portion of blood, and further the virus and leukocyte selective removing material is prevented from flowing out of the container. There is also provided a virus and leukocyte selective removal device characterized in that the device is provided.

These methods, removal materials and devices are particularly useful when the blood contains hepatitis C virus. Platelet passing virus and leukocyte selective removal material
Most preferably, the carrier surface has a terminal hydrophilic group, or a terminal hydrophilic group and a polyethylene glycol group, and additionally a terminal hydrophobic group.

[0012]

BEST MODE FOR CARRYING OUT THE INVENTION The present invention is described in detail below. In the present invention, the term "virus" includes not only free virus in blood but also protein-bound virus, infected virus present in leukocytes and the like. More specifically, the substances to be removed in the present invention are viruses in the blood, protein-bound viruses, etc., such as hepatitis A virus, B
Although any of hepatitis C virus, hepatitis C virus, HIV and the like can be removed, hepatitis C virus can be particularly well removed. The reason for this is not clear, but it is presumed that it can be removed as efficiently as leukocytes in terms of both the surface characteristics of the virus and the size of the virus. With the hepatitis C virus,
Hepatitis C virus in blood, hepatitis C virus adsorbed on immunoglobulins, etc., hepatitis C virus adsorbed on plasma proteins, etc., and lymphocytes activated by hepatitis C virus, macrophages present at inflammatory sites, White blood cells such as granulocytes can be mentioned. In particular, when the virus is removed simultaneously with a drug such as interferon, the hepatitis C virus, the hepatitis C virus adsorbed on plasma proteins, and the white blood cells infected with the hepatitis C virus are removed in the present invention in advance. , Lymphocytes infected with hepatitis C virus, autoreactive T activated by hepatitis C virus
You can list cells and so on. In particular, regarding leukocytes, it is useful to remove lymphocytes because hepatitis C virus infects lymphocytes.

In the present invention, the term blood also includes blood components such as plasma and serum. When treating blood, an anticoagulant can be added to the blood for the purpose of anticoagulating the blood. Examples of the anticoagulant include, but are not particularly limited to, compounds having anticoagulant activity, heparin,
Fusan, FOY, argatroban, citric acid and the like are mentioned as suitable examples, and heparin or fusan is preferably used.

In the present invention, removing viruses and leukocytes means removing them from blood by adsorption and / or filtration. The method of contacting the virus and leukocyte removing material with blood for removal is as follows. Any of a stationary method, a shaking method, an adsorption method by diffusion, a filtration method and the like may be used. Further, in the adsorption and filtration method, a method of flowing blood by a head or a pump is effectively used.

It has been surprisingly found that the virus and leukocyte removing material of the present invention preferably has at least a terminal hydrophilic group on the surface. As the preferred terminal hydrophilic group on the carrier surface of the virus and leukocyte removing material, a neutral functional group having no electric charge is usefully used. For example, hydroxyl group,
A hydroxyl group-containing alkyl group such as a hydroxymethyl group, a hydroxyethyl group, a hydroxypropyl group, a hydroxyisopropyl group, a hydroxybutyl group and a hydroxyisobutyl group, and a methoxypolyethylene glycol group such as a methoxydiethylene glycol and a methoxytriethylene glycol group are preferably used. Of these, a hydroxyl group, a hydroxypropyl group, a hydroxyisopropyl group and a hydroxyisobutyl group are preferably used. Further, in order to improve the passage of platelets, methoxydiethylene glycol and methoxytriethylene glycol groups are preferable. It is most preferable to use a hydroxyl group, a hydroxypropyl group, a hydroxyisopropyl group, a hydroxyisobutyl group and a methoxydiethylene glycol or a methoxytriethylene glycol group in combination in order to improve both the removal rate of viruses and leukocytes and the recovery rate of platelets. .

In the present invention, the term "terminal" means the terminal of the main chain or the terminal of the side chain. The terminal group may be directly bonded to the main chain, or may be bonded via an ester bond, an amide bond, a urethane bond or the like. In the latter case, it refers to the terminal portion that does not include these bonds.

Preferred terminal hydrophobic groups on the surface of the carrier of the virus and leukocyte removing material of the present invention are methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group,
t-butyl group, pentyl group, hexyl group, heptyl group,
An alkyl group having 1 to 30 carbon atoms, such as an octynyl group,
Examples thereof include aromatic rings such as phenyl group, and aliphatic rings such as cyclopentyl group and cyclohexyl group. From the viewpoint of platelet permeability, an alkyl group having 10 or more and less than 30 carbon atoms, an alkyl group such as a methyl group and an ethyl group, and an alkyl group having 10 or more and less than 20 carbon atoms, and a methyl group and an ethyl group are most preferable. .

In the material for selective removal of viruses and leukocytes of the present invention, the hydrophilic group is effective in adsorbing / removing hydrophilic protein-adsorbing virus and leukocytes, and also effective in improving platelet recovery ability. Further, the hydrophobic group is effective for improving the adsorptivity of viruses and viruses adsorbing hydrophobic proteins, but the virus and leukocyte removing material of the present invention removes viruses and leukocytes and maintains the passage of platelets. so,
It is also important to balance the terminal hydrophilic group and the terminal hydrophobic group. Excessive hydrophobicity is detrimental to platelet recovery.

The preferable ratio of the terminal hydrophilic group is 2% or more and less than 100%. When the abundance ratio of the terminal hydrophilic group is less than 2%, the hydrophobicity is high so that leukocytes and viruses can be adsorbed, but the passage rate of platelets is extremely lowered, which is not preferable. On the other hand, if the abundance ratio of the hydrophilic group is 100%, the adsorptivity of the virus decreases, which is not preferable. From the above viewpoints, a more preferable range is 3% or more and less than 90%, and most preferably 5% or more and less than 80%.

Further, the abundance ratio of the terminal hydrophobic group is also important, and it is useful when it is 0.1% or more and less than 70%. When the terminal hydrophobic group is 70% or more, the recovery rate of platelets decreases, which is not preferable. On the other hand, if less than 0.1%,
Since the hydrophobicity is low, the adsorptivity of the virus-adsorbing protein decreases, which is not preferable. From the above viewpoints, more preferably 1% or more and less than 60%, most preferably 1% or more and 55%.
It is less than%.

In the present invention, the abundance ratio of the terminal hydrophilic group and the terminal hydrophobic group is the ratio of the hydrophilic group and the hydrophobic group existing on the surface of the virus and leukocyte removing material, and each functional group on the surface portion where the carrier and blood can come into contact with each other. Refers to the molar ratio of groups. The abundance ratio of these terminal groups is a known solid state nuclear magnetic resonance spectrum,
It can be determined by infrared absorption spectrum, XPMS, ESCA, or the like. Further, when the surface of the carrier is modified by coating or the like, it is possible to show the abundance ratio in the coated polymer by using the molar ratio.

The surface in the present invention means a surface which can be contacted with viruses and the like, and does not include the inside of a material which cannot be contacted with viruses. And, in the present invention, having a surface that captures viruses and leukocytes in blood and allows platelets to pass is sufficient if it has a terminal hydrophilic group or the like on the carrier surface, and the surface is coated with a terminal hydrophilic group or the like. Not only in the case of providing, but also in the case of introducing a functional group into the surface of the carrier by radiation grafting or covalent bonding, or in the case where the carrier material itself has these functional groups on the surface.

The virus and leukocyte removing material may have a terminal positive group on the carrier surface in addition to the terminal hydrophilic group. The terminal positive group is usefully used particularly in terms of improving adsorption of a virus having a negative charge on the surface. When the terminal positive group is exemplified, a tertiary amino group formed by bonding a dimethylamino group, a diethylamino group, a dipropylamino group or the like to the terminal of the polymer main chain or a side chain,
Examples thereof include aromatic rings such as heterocycles. Among them, a dimethylamino group and a diethylamino group are usefully used. When the terminal is a primary or secondary amino group, the ionicity is strong and the platelet recovery rate is lowered, which is not preferable. The existence ratio of the terminal positive group is preferably less than 15%. If it exceeds 15%, the platelet recovery rate is lowered due to an excess of positive groups, which is not preferable. A preferred abundance ratio is less than 13%, and most preferably less than 11%.

The virus and leukocyte selective removing material of the present invention can be most efficiently removed leukocytes and viruses at the same time by using a material that activates the concentration of activated complement C3a five times or more before and after contact with blood. Do you get it. The virus is more likely to be adsorbed under the influence of complex formation with activated complement C3a, while the excess activated complement C3a
It was found that the adsorptivity was decreased due to the relative decrease in virus concentration due to the increase in concentration. That is, activated complement C3
When a is less than 5 times the blood concentration before contact, the ability to remove viruses and leukocytes is extremely reduced, which is not preferable. On the other hand, it is effective if the activated complement C3a is 5 times or more the blood concentration before contact, but the activated complement C3a has 10 times the blood concentration before contact.
When it is more than 00 times, it is not practical because it causes anaphylaxis etc. due to complement. Further, if the concentration change of 500 times or more occurs, the change of blood components becomes remarkable, which is not preferable. More preferred activated complement C3 before and after contact with blood
The a concentration is 7 times or more, and most preferably 10 times or more.

As a material that activates the concentration of activated complement C3a by 5 times or more before and after contact with blood, the above-mentioned material having 5 mol% or more of terminal hydrophilic groups on the surface is used. Examples of the terminal hydrophilic group-containing monomer include hydroxyalkyl methacrylates such as 2-hydroxyethyl methacrylate, 2-hydroxypropyl methacrylate, 2-hydroxyisopropyl methacrylate, 2-hydroxybutyl methacrylate, and 2-hydroxyisobutyl methacrylate as the terminal hydrophilic group monomer. Examples thereof include methoxypolyethylene glycol methacrylate such as methoxydiethylene glycol methacrylate, methoxytriethylene glycol methacrylate, and methoxytetraethylene glycol methacrylate.

For activation, a polymer obtained by copolymerizing these monomers in binary or ternary is effectively used. For example, a random copolymer (copolymerization molar ratio, HBMA / MDG / MMA) of 2-hydroxyisobutyl methacrylate (hereinafter abbreviated as HBMA), methoxydiethylene glycol methacrylate (hereinafter abbreviated as MDG) and methyl methacrylate (hereinafter abbreviated as MMA). = 5-40: 5-
30: 40-60), 2-hydroxyisopropyl methacrylate (hereinafter abbreviated as HPMA), methoxydiethylene glycol methacrylate and methyl methacrylate random copolymer (copolymerization molar ratio HPMA / MD
G / MMA = 5-40: 5-30: 40-70), HP
MA and dimethylaminoethylmethacrylate (hereinafter DM
Abbreviated) random copolymer (copolymerization molar ratio HPMA
/ DM = 80 to 93: 3 to 20) and the like. In particular,
2-hydroxyisobutyl methacrylate (hereinafter HB
MA), methoxydiethylene glycol methacrylate (hereinafter abbreviated as MDG) and methyl methacrylate (hereinafter abbreviated as MMA) random copolymer (copolymerization molar ratio, HBMA / MDG / MMA = 5-40: 5-3).
0: 40-60) is most usefully used.

Also, the material itself is effectively used when the activated complement C3a is increased to 5 times or more the blood concentration before contact. A material having a hydrophilic group on its surface is used. Examples of the material include natural polymers such as cellulose and / or its derivatives, polyesters such as polyethylene terephthalate and polybutylene terephthalate, ethylene vinyl alcohol copolymers and polyurethanes. Molecular materials are usefully used. Particularly preferred from the viewpoint of activation are polyesters such as polyethylene terephthalate and polybutylene terephthalate, ethylene vinyl alcohol copolymers, cellulose and the like. Most preferably, polyesters such as polyethylene terephthalate and polybutylene terephthalate are usefully used.

As a carrier for the material for selectively removing viruses and leukocytes used in the present invention, granular, bead-like, porous material, flat membrane,
Nonwoven fabrics and woven fabrics can be exemplified. Among them, porous bodies and non-woven fabrics are preferably used, and non-woven fabrics are most preferable because they can remove leukocytes simultaneously with viruses and have a high surface area.

The material of the carrier is not particularly limited as long as it can be surface-treated, and natural polymers such as cellulose and / or its derivatives, polyester such as polyethylene terephthalate and polybutylene terephthalate, polyethylene, polypropylene. Polymer materials such as polyolefin, polyvinylidene fluoride, polyamide, polyimide, polyurethane, polysulfone, polyacrylonitrile and the like can be exemplified.

When these non-woven fabrics have an affinity for viruses and white blood cells without surface modification, they can be used as they are. Further, when those functions cannot be exhibited without surface modification, it is preferable to perform surface modification such as coating for the purpose of exhibiting the function. In particular, those whose surface is modified by a treatment such as coating are preferably used for the purpose of adsorbing and / or removing target substances and leukocytes and improving collectability of platelets.

When the carrier is a non-woven fabric, the filament may be a monofilament or a multifilament, and may be a porous filament or an atypical filament. Further, as the non-woven fabric, a non-woven fabric having an average fiber diameter of 2.0 μm or more and less than 50 μm is preferable. If the fiber diameter becomes large, it becomes difficult to secure the surface area of the base material, which not only reduces the adsorption area of the virus but also reduces the leukocyte removal performance, which is not preferable. On the other hand, when the fiber diameter is small, clogging of the removing material is likely to occur, and it becomes difficult to collect platelets, which is not preferable. From the above viewpoints, the average fiber diameter is more preferably 2.0 μm or more and less than 30 μm, and most preferably 2.3 μm or more and less than 20 μm.

Further, in the case of a non-woven fabric, its bulk density is 0.10 g / c in order to remove leukocytes and improve the permeability of platelets.
It is also important that it is m ³ or more and less than 0.45 g / cm ³ . When the bulk density is less than 0.10 g / cm ³ , the leukocyte removal performance deteriorates, which is not preferable. On the other hand, the bulk density is 0.
When it is 45 g / cm ³ or more, platelet permeability is extremely reduced, which is not preferable. Further, more preferably above aspects 0.15 g / cm ³ or more 0.45 g / cm less than ^3, and most preferably 0.15 g / cm ³ or more 0.40 g / cm ³
Is less than.

In the case of the nonwoven fabric, the adsorption of the virus, in terms of increasing the removal and permeability of platelets in leukocytes, it is also important that the specific surface area is less than 0.010 ² / g or more 4.0 m ² / g. When the specific surface area is less than 0.01 m ² / g,
It is not preferable because the ability to remove viruses and leukocytes decreases. On the other hand, when the specific surface area is 4.0 m ² / g or more, the platelet permeability is extremely reduced, which is not preferable. Further, from the above viewpoint, it is preferably 0.02 m ² / g or more and less than 3.0 m ² / g, and most preferably 0.04 m ² / g or more and less than 2.5 m ² / g.

As a surface-modifying compound capable of removing viruses and leukocytes and recovering platelets at a high rate, a polymer compound having at least a terminal hydrophilic group in its side chain, and a high-molecular compound having a terminal hydrophilic group and a terminal hydrophobic group in its side chain at the same time. Molecular compounds are mentioned.

As examples of the monomers constituting these polymer compounds, 2-hydroxyethyl methacrylate, 2-hydroxypropyl methacrylate, 2-hydroxyisopropyl methacrylate, 2-hydroxybutyl methacrylate, and 2-hydroxyisobutyl are used as the hydrophilic terminal monomer. Examples thereof include hydroxyalkyl methacrylate such as methacrylate, methoxydiethylene glycol methacrylate, methoxytriethylene glycol methacrylate, and methoxypolyethylene glycol methacrylate such as methoxytetraethylene glycol methacrylate.

Examples of the terminal hydrophobic group monomer include alkyl methacrylate such as methyl methacrylate, ethyl methacrylate, propyl methacrylate, butyl methacrylate and octadecyl methacrylate, and aromatic methacrylate such as benzyl methacrylate and phenyl methacrylate.

The above-mentioned polymerizable functional group can be effectively used even if it is vinyl, acrylate or the like. The surface coating material used in the present invention can be effectively obtained by copolymerizing monomers in a binary or ternary manner according to the abundance ratio of hydrophilic groups and hydrophobic groups.

Particularly preferred examples of copolymers using these monomers include copolymers of methoxydiethylene glycol methacrylate, 2-hydroxyisobutyl methacrylate, and methyl methacrylate. When the terminal positive group is introduced, examples of the terminal positive group monomer include dialkylaminoalkyl methacrylates such as dimethylaminoethyl methacrylate and diethylaminoethyl methacrylate.

The polymer compounds exemplified above may be used directly or
It is effectively used by coating the surface of the substrate. Further, in addition to directly using these monomers, it can be effectively used by introducing a necessary terminal group after polymerization or copolymerization using a monomer such as glycidyl methacrylate.

As a method for imparting a hydroxyl group, a polyethylene glycol group, a hydrophobic group or a neutral group to the surface of the carrier, various known methods can be applied. Here, the term “applying” means that the carrier is present on the surface of the carrier so as not to be eluted in water or blood, and examples thereof include a graft polymerization method, a coating method,
Epoxy group, amino group, formyl group, carboxyl group,
Examples thereof include a method in which a functional group such as a hydroxyl group, an acid halide group, or a cyanogen halide group is introduced on the surface of a carrier, and then the compound having a target functional group is bonded directly or through a coupling agent or a spacer.

The virus and leukocyte selective removing material of the present invention can be preferably carried out by the blood processing apparatus of the present invention. The virus and leukocyte removal device of the present invention is a device for adsorbing and / or removing viruses, protein-bound viruses, and leukocytes from blood, and the virus and leukocyte selective removal material is a blood introduction part of blood and blood derivation. A device for selectively removing virus and leukocytes, which is contained in a container having a portion and is provided with a means for preventing the virus and leukocyte selective removal material from flowing out of the container.

As means for preventing the removal material from flowing out of the container, any means that allows all blood components to pass but does not allow the removal material to pass can be used. This can be carried out by providing a mesh or filter having a mesh finer than the material diameter of the removing material at least on the outlet side of the container. Further, when fibers such as non-woven fabric are used as the carrier, in addition to using the mesh as described above, the non-woven fabric is adhered to the upper or lower end of the container with an adhesive or sandwiched between the containers. Is possible. Particularly in the case of a cylindrical depth filter, it can be carried out by closing one end of the cylinder and adhering the outlet end to a nozzle or the like. The adhesive used is preferably one that does not weaken the adhesive force due to the influence of swelling or the like even when it is brought into contact with a liquid such as blood, and urethane and epoxy adhesives are useful, but are not limited thereto. Do not mean.

The apparatus of the present invention is particularly useful when it is connected to a blood collecting means, an anticoagulant mixing means, a blood returning means and the like through a tube. The housing of the device of the present invention is preferably made of a synthetic resin such as polypropylene, polycarbonate, polyethylene, polystyrene, polymethacrylic acid, or a metal such as glass or stainless steel.

Experimental examples and examples of the present invention will be shown below, but the present invention is not limited thereto. As an experimental example, an example of producing a polymer used for surface-modifying a carrier in the present invention will be shown.

[Experimental Example 1] A random copolymer of 2-hydroxyisobutyl methacrylate (hereinafter abbreviated as HBMA), methoxydiethylene glycol methacrylate (hereinafter abbreviated as MDG) and methyl methacrylate (hereinafter abbreviated as MMA) was used as an ordinary radical initiator. Was synthesized. Polymerization conditions were as follows: MDG monomer, HEMA monomer, and MMA monomer (molar ratio / MDG: HBMA: MMA = 30: 20: 50) per 300 ml of ethanol, and azobisvaleronitrile (V-65) 0 as an initiator. .1
Polymerization was carried out at 70 ° C. for 6 hours in the presence of g. The obtained polymerization solution was dropped into 10 L of water with stirring to precipitate a copolymer, and a water-insoluble matter was recovered. The composition ratio in the obtained copolymer was the same as the monomer charging ratio. Therefore,
In this copolymer, the abundance ratio of the terminal hydrophilic groups is 50%, and the abundance ratio of the terminal hydrophobic groups is 50%.

[0045]

[Experimental Example 2] A copolymer of 2-hydroxyisopropylmethacrylate (HPMA) and dimethylaminoethylmethacrylate (DM) was prepared in the same manner as in Experimental Example 1. The copolymer molar ratio HPMA: DM is 97: 3.
The abundance ratio of the terminal hydrophilic group was 97%, and the ratio of the terminal positive group was 3%.

[0046]

Experimental Example 3 By the same operation as in Experimental Example 1, a copolymer of 2-hydroxyethyl methacrylate (HEMA), dimethylaminoethyl methacrylate (DM) and methyl methacrylate (MMA) was prepared. The molar ratio HEMA: MMA: DM of the copolymer was 62: 30: 8, and the abundance ratio was 62% of terminal hydrophilic groups, 30% of terminal hydrophobic groups, and 8% of terminal positive groups.

[0047]

EXAMPLES Examples of the blood treatment system for hepatitis C are shown below, but the present invention is not limited thereto.

Example 1 1 g of the copolymer obtained in Experimental Example 1 was added to 70 g.
% Aqueous solution of ethanol to dissolve it to obtain a 1% coating solution. Nonwoven fabric composed of polyethylene terephthalate fibers having an average fiber diameter of 2.9 μm (Basis weight 90 g / m ² , thickness 0.40 mm, bulk density 0.24 g / cm ³ , specific surface area 0.966 m ² / g) 1 g 1% coating liquid 10 m
After soaking in 1 l, it was dried at 25 ° C. for 12 hours. 0.01 g of the obtained non-woven fabric was cut into strips, collected in a vial, and 1 mL of patient blood containing hepatitis C virus was added to it.
Shake for 2 hours at 7 ° C. After that, 100μm blood after treatment
Sample in L vial and centrifuge at 5000 rpm for 1 minute to determine the amount of hepatitis C virus in the supernatant.
It was measured as RNA. An Amplicore HCV monitor manufactured by Nippon Roche was used for measuring the amount of hepatitis C virus. In addition, the number of white blood cells and the number of platelets of the treated blood were determined using an automatic hemocytometer (SF-3000 manufactured by Sysmex Corporation). In addition, activated complement C before and after treatment
The concentration of 3a was measured by the nephelometry method, and the increase rate of the post-treatment value with respect to the pre-treatment value was determined. As a control experiment, the same operation as in Example 1 was performed without adding the removing material of the present invention.

The hepatitis C virus adsorption rate (%), leukocyte removal rate (%), and platelet recovery rate (%) were calculated by the following formulas. Virus adsorption rate (%) = [(Vd−Vc) / Vd] × 10
0 Vc: Control experiment Virus concentration in blood Vd: Adsorption experiment Virus concentration in blood Leukocyte removal rate (%) = [(Wd-Wc) / Wd] x 100 Wc: Control experiment Leukocyte concentration in blood Wd: Adsorption experiment Leukocyte concentration in blood Platelet recovery rate (%) = Pd / Pc × 100 Pc: Platelet concentration in control experiment blood Pd: Adsorption experiment Platelet concentration in blood Table 1 shows the results. Table 1 also shows the results of the control experimental example in which the removing material was not added. Even in the control experimental example, a slight decrease is observed due to the attachment of white blood cells and platelets to the container.

[0049]

Example 2 Non-woven fabric made of polypropylene fiber having an average fiber diameter of 2.5 μm (Basis weight: 60 g / m ² , thickness: 0.35)
mm, bulk density 0.12 g / cm ³ , specific surface area 1.768
The same operation as in Example 1 was carried out except that m ² / g) was used. The results are shown in Table 1.

[0050]

[Table 1]               Virus adsorption rate Leukocyte removal rate Platelet passage rate C3a concentration   Example 1 85% 80% 90% 11.6 times   Example 2 89% 82% 94% 18.5 times   Control experiment 0% 1% 98% 2.3 times

[0051]

[Comparative Example 1] Non-woven fabric made of polyethylene terephthalate fiber having an average fiber diameter of 2.9 μm (Basis weight: 90 g / m ² ,
The same operation as in Example 1 was performed except that the thickness (0.40 mm) was used as it was. The results are shown in Table 2.

[0052]

[Comparative Example 2] Non-woven fabric made of polypropylene fiber having an average fiber diameter of 2.5 μm (Basis weight: 60 g / m ² , thickness: 0.35)
mm) was used as it was, and the same operation as in Example 1 was performed. The results are shown in Table 2.

[0053]

[Table 2]             Virus adsorption rate Leukocyte removal rate Platelet passage rate C3a concentration   Comparative Example 1 58% 84% 21% 3.5 times   Comparative Example 2 63% 80% 18% 1.8 times

[0054]

Example 3 The removal material of Example 1 was cut into a circle having a diameter of 6.8 mm, and five pieces were set in a column to evaluate the hepatitis C virus adsorption rate, leukocyte removal rate, and platelet recovery rate.
Human fresh blood with ACD-A added as an anticoagulant to each column (virus amount: 100,000 cells / ml,
White blood cell count: 4,500 to 8,400 / μL, platelet count:
150,000-440,000 / μL) (Blood: AC
D-A = 8: 1) 1.5 mL was made to flow at room temperature at a constant flow rate of 0.5 mL / min using a syringe pump, and the virus concentration, leukocyte concentration and platelet concentration in blood before and after passing through the nonwoven fabric were measured. The virus adsorption rate, leukocyte removal rate and platelet recovery rate were determined in the same manner as in Example 1. The results are shown in Table 3.

[0055]

Example 4 Example 3 except that the removing material of Example 2 was used.
The same operation was performed. The results are shown in Table 3.

[0056]

[Table 3]             Virus adsorption rate Leukocyte removal rate Platelet passage rate C3a concentration     Example 3 91% 95% 70% 12.5 times     Example 4 94% 97% 75% 23.5 times

[0057]

Comparative Example 3 The same operation as in Example 3 was performed except that the material of Comparative Example 1 was used. The results are shown in Table 4.

[0058]

Comparative Example 4 The same operation as in Example 3 was performed except that the material of Comparative Example 2 was used. The results are shown in Table 4.

[0059]

[Table 4]             Virus adsorption rate Leukocyte removal rate Platelet passage rate C3a concentration     Comparative Example 3 60% 98% 8% 4.8 times     Comparative Example 4 75% 97% 6% 2.3 times

[0060]

Example 5 A removing material was obtained by the same procedure using the same non-woven fabric as in Example 1 except that the polymer produced in Experimental Example 2 was used for coating, and blood was treated in exactly the same manner as in Example 1. Hepatitis C virus adsorption rate, leukocyte removal rate, and platelet recovery rate were evaluated. The results are shown in Table 5.

[0061]

Example 6 Except that the polymer produced in Experimental Example 3 was used for coating, the same non-woven fabric as in Example 1 was used to obtain a removing material by the same procedure, and blood was treated in the same manner as in Example 1 to treat C. The hepatitis B virus adsorption rate, leukocyte removal rate, and platelet recovery rate were evaluated. The results are shown in Table 5.

[0062]

[Table 5]             Virus adsorption rate Leukocyte removal rate Platelet passage rate C3a concentration     Example 5 79% 89% 87% 19.5 times     Example 6 91% 95% 86% 22.5 times

[0063]

Example 7 The same nonwoven fabric as in Example 1 was cut into a piece having a width of 150 mm and a length of 300 mm and wound around a polyethylene cylindrical mesh having a diameter of 3.4 mm. Next, a non-woven fabric composed of polyester fibers with an average fiber diameter of 12 μm (30 g
/ M ² units) as the first prefilter, width 150 mm
Wrapped in. Further, a non-woven fabric (50 g / m ² basis weight) made of polyester fiber having an average fiber diameter of 33 μm was used as a second pre-filter and wound with a width of 150 mm. A polyethylene mesh was wound on the outside by 150 mm. The diameter of this cylinder is 3
It was 9 mm. Block both ends of this cylinder with urethane,
A cylindrical polycarbonate container having an inner diameter of 41 mm having a blood inlet and a blood outlet at the ceiling and a bottom, respectively, and a cylindrical outer peripheral surface communicating with the blood inlet of the container and an inner peripheral surface communicating with the blood outlet. It was created. Fresh bovine blood supplemented with heparin as an anticoagulant (white blood cell count:
4,500-6,400 / μL, Platelet count: 150,0
00-320,000 / μL) (heparin concentration: 100
After adding 50 ml of plasma containing hepatitis C virus to 2000 mL of 0 IU / L) (virus amount: 2500000 / l), the blood cells were allowed to flow at a constant flow rate of 50 mL / min at room temperature to remove leukocytes. Leukocyte removal device passage 20
The virus concentration, leukocyte concentration and platelet concentration in blood before and after the treatment with 00 ml were measured, and the virus adsorption rate, leukocyte removal rate and platelet recovery rate were determined in the same manner as in Example 1.
The results are shown in Table 6.

[Table 6]             Virus adsorption rate Leukocyte removal rate Platelet passage rate     Example 7 69% 93% 62%

[0064]

As is apparent from the above examples, according to the present invention, a virus and leukocyte removing material capable of selectively adsorbing and / or removing viruses and leukocytes present in blood can be provided. it can. Further, by using a blood processing device filled with the above-mentioned removal material,
It is possible to selectively remove hepatitis C virus and leukocytes in a liquid to be treated such as blood, plasma, serum and the like, and to collect platelets at a high rate.

   ─────────────────────────────────────────────────── ─── Continued front page    F-term (reference) 4C077 AA07 AA12 BB02 BB03 KK09                       KK11 LL01 LL02 LL15 LL16                       LL17 LL23 MM04 MM06 MM09                       NN02 NN15 PP02 PP09 PP10                       PP12 PP13 PP15 PP21

Claims (25)

[Claims] 1. A method for simultaneously selectively removing viruses and leukocytes from blood, which comprises virus-containing blood,
A method comprising the step of capturing viruses and leukocytes in blood and bringing them into contact with a virus and leukocyte selective removal material comprising a water-insoluble carrier having a surface through which platelets pass. 2. The method according to claim 1, wherein the virus and leukocyte selective removal material and the virus-containing blood are contacted by an adsorption method and / or a filtration method. 3. A step of mixing an anticoagulant with a virus-containing blood, wherein the virus-containing blood mixed with the anticoagulant is used as a virus and leukocyte selective removal material at 10 ml / min or more 10.
The method according to claim 1 or 2, comprising a step of contacting at a flow rate of less than 0 ml / min. 4. The method according to claim 1, wherein the leukocytes to be removed include at least lymphocytes. 5. The method according to claim 1, wherein the leukocytes to be removed include at least autoreactive T cells. 6. The method according to claim 1, wherein the virus and leukocyte selective removing material has at least a hydroxyl group and a polyethylene glycol group on the surface. 7. The material for selectively removing viruses and leukocytes,
The method according to any one of claims 1 to 6, which has the ability to activate the concentration of activated complement C3a in blood 5 times or more before and after the contact. 8. The method according to claim 1, wherein the water-insoluble carrier is fibrous. 9. The method according to claim 8, wherein the leukocyte-removing carrier is a non-woven fabric. 10. The method according to claim 1, wherein the virus is hepatitis C virus. 11. Platelet permeation for selective simultaneous removal of viruses and leukocytes from blood, characterized by comprising a water-insoluble carrier having a surface for capturing viruses and leukocytes in blood and allowing passage of platelets. Type virus and leukocyte selective removal material. 12. The platelet-permeable virus and leukocyte selective removal material according to claim 11, wherein the carrier has a terminal hydrophilic group on at least the surface thereof. 13. The terminal hydrophilic group is a hydroxyl group and / or a polyethylene glycol group, according to claim 1.
2. The platelet-permeable virus and leukocyte selective removal material according to 2. 14. The platelet-permeable virus and leukocyte selective removal material according to claim 12, wherein the carrier has a terminal hydrophobic group on at least the surface thereof. 15. The platelet-passing virus and leukocyte selective removing material according to claim 14, wherein the terminal hydrophobic group is an alkyl group. 16. The method according to claim 13, wherein the abundance ratio of the terminal hydrophilic groups on the surface of the carrier is 2% or more and less than 100% and the abundance ratio of the terminal hydrophobic groups is 0.1% or more and less than 70%. 14. The platelet-passing virus and leukocyte selective removal material according to 14. 17. The carrier surface has a terminal hydrophilic group of 5% or more and less than 30% of a methoxydiethylene glycol group, 5% or more and less than 40% of a 2-hydroxyisobutyl group, and a terminal hydrophobic group of 40% or more and less than 60% of a methyl group. The platelet-passing virus and leukocyte selective removal material according to claim 16, wherein 18. The platelet-passing virus and leukocyte selection according to any one of claims 11 to 17, wherein the concentration of activated complement C3a after contact with blood is 5 times or more that before contact with blood. Removal material. 19. The platelet-passing virus and leukocyte selective removal material according to claim 11, wherein the carrier is a non-woven fabric. 20. The carrier has an average fiber diameter of 2.0 μm or more and 5
The platelet-passing virus and leukocyte selective removal material according to claim 19, which is a non-woven fabric having a thickness of less than 0.0 μm. 21. The bulk density of the non-woven fabric is 0.10 g / cm ^3.
21. The platelet-passing virus and leukocyte selective removal material according to claim 19 or 20, which is not less than 0.45 g / cm ³ . 22. The specific surface area of the non-woven fabric is 0.01 m ² / g
22. The platelet-passing virus and leukocyte selective removal material according to any one of claims 19 to 21, wherein the material is less than 4.0 m ² / g. 23. The platelet-passing virus and leukocyte selective removal material according to any one of claims 11 to 22, wherein the virus is hepatitis C virus. 24. The virus and leukocyte selective removal material according to any one of claims 11 to 23 is contained in a container having a blood introduction part and a blood derivation part, and the virus and leukocyte selective removal material container is further included. A device for selectively removing viruses and leukocytes, which is provided with a means for preventing outflow to the outside. 25. The virus and leukocyte selective removal device according to claim 24, wherein the virus is hepatitis C virus.