RU2617537C1 - System for autologous serum obtaining - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to a system for production of an autologous serum with an increased content of IL-1RA and with a reduced content of pro-inflammatory cytokine, comprising a syringe with internal walls coated with a solid phase polymeric sorbent with monoclonal IL-1β antibodies fixed thereon, containing glass balls, 2.5 to 3.5 mm in diameter, occupying up to 40% of the total volume of the syringe, and at least one syringe with internal walls coated with a solid phase polymer sorbent with human IL-1β fixed thereon, nd a sterile adapter designed to move the autologous serum from a syringe with antibodies to a syringe with human IL-1β.

EFFECT: significant decrease in pro-inflammatory cytokine IL-1β concentration and increased effectiveness of the therapeutic effect of the autologous concentrated serum.

4 cl, 3 ex, 7 dwg

Description

The invention relates to medicine, namely to a system for producing autologous serum with a high content of IL-1RA, growth factors and with a reduced content of pro-inflammatory cytokines and other pro-inflammatory factors, and can be used to obtain autologous plasma preparations in various variants that are actively used in medicine for the treatment of a wide range of diseases.

Among the pro-inflammatory cytokines, IL-1 and TNF-a are currently considered the most significant.

Unfortunately, the use of drugs that block these cytokines (remikade, adalimumab) cannot be carried out routinely to treat degenerative diseases due to the large number of limitations and cost. However, the use of biotechnology to increase IL-1RA (an antagonist of interleukin 1 IL-1) in blood serum (autologous concentrated serum - ACS) and the administration of this drug intra-articularly (paravertebrally, paracendicular, etc.) give positive results in the experiment and clinic.

The main objective of the study is to find ways to stimulate the release of anti-inflammatory cytokines from blood cells in high concentrations and remove the resulting pro-inflammatory cytokines (and other factors). In this case, it is supposed to stimulate the release of cytokines with inert, easily removable components or media.

Autologous plasma preparations are a valuable complement and alternative to existing treatment approaches. No other type of injection therapy (for example, the introduction of corticosteroids or hyaluronic acid) has the severity and duration of the effect characteristic of this method.

Therapy with autologous plasma preparations involves the introduction of autologous conditioned serum (ACS) with an increased content of interleukin 1 receptor antagonist (IL-1RA) into the affected joint, place of enthesopathy, or into the inflamed tendon.

A study on back pain conducted by Bochum University (Becker et al., 2007) compared autologous plasma therapy with corticosteroids for epidural perineural administration. No specific side effects were observed.

Autologous plasma drug therapy is internationally recognized as a highly effective, innovative method of local therapy; The advantages of this method are described in specialized medical journals.

The prior art technical solutions.

A known apparatus for producing therapeutically active proteins in the blood (international application No. 20158585348, 06/18/2015), including a tube containing silica-coated granules. After collecting the blood, the tube is incubated under aseptic conditions at 37 ° C for 24 hours. After incubation, each tube is centrifuged (3500 rpm, 10 min) and the top layer, being plasma or serum, is transferred to a sterile syringe and injected back to the patient.

Also known is a system for producing an interleukin-1 receptor antagonist or a therapeutically effective protein, selected as the closest analogue to the patented solution (US No. 2010125236, 05.20.2010). Blood is obtained from the patient using a conventional syringe, and then injected into a double Luer centrifuge tube. The centrifuge tube is equipped with silanized coated balls. The tube is then incubated and centrifuged. After incubation and centrifugation of serum containing therapeutically active autologous proteins, such as IL-1RA, the resulting plasma is introduced back into the patient's body.

However, studies of recent years have shown that known methods lead to an increase in the serum of both anti-inflammatory and pro-inflammatory cytokines simultaneously. And this fact casts doubt on the unequivocal effectiveness of the use of the obtained products in the treatment of chronic and acute arthritis.

The technical result of the proposed technical solution is a significant reduction in the concentration of the pro-inflammatory cytokine IL-1β (possibly additionally TNF-α, IL-6, MMP), as a result, increasing the effectiveness of the therapeutic effect of autologous concentrated serum.

The claimed technical result is achieved by using the design of a multicomponent system to obtain autologous serum with a high content of IL-1RA and with a reduced content of pro-inflammatory cytokines, including one syringe (syringe 1), the inner walls of which are coated with a solid-phase polymer sorbent with monoclonal antibodies to IL fixed on it -1β (possibly in addition to TNF-α, IL-6), in addition to the walls, the solid-phase polymer sorbent can be located on an additional structure (sponge, mesh, systems tubes) inside a syringe containing glass balls with a diameter of 2.5 to 3.5 mm, occupying up to 40% of the total volume of the syringe, and 3-6 syringes (syringe 2), the inner walls of which are coated with a solid-phase polymer sorbent with human fixed on it IL-1β (optionally additional TNF-α, IL-6), and a sterile adapter for transferring autologous serum from a syringe with antibodies to a syringe with human IL-1β.

One three-component syringe can also be used to obtain autologous serum with a high content of IL-1RA and a reduced content of pro-inflammatory cytokines, which also provides the indicated result.

The technical result is achieved due to the antigen-antibody reaction, in which IL-1 (possibly IL-6, TNF-α), formed upon receipt of autologous serum or initially found in human blood in high concentration, binds to a specific protein (monoclonal antibody), in the course of this reaction, the monoclonal antibody – IL-1 complex remains fixed on the solid-state sorbent (possibly an additional monoclonal antibody – TNF-α, the monoclonal antibody – IL-6), and free autologous serum in the syringe but it contains a significantly reduced amount of IL-1β (possibly IL-6, TNF-α).

Syringe 1 is essentially a tube that acts as a syringe due to vacuum and a double-sided needle.

The use of an additional porous structure or various variants of the tube wall ribbing (syringe 1) increases the contact area between the plasma and the antibodies fixed on the solid phase, to a greater extent, fixing the pro-inflammatory cytokines and their lower concentration in the resulting plasma.

The volume of the syringe, the walls of which are coated with an antibody to IL-1b (possibly in addition to IL-6, TNF-α), is from 20 to 60 ml. The most likely use of monoclonal mouse antibodies obtained by hybridoma technology, it is also possible to use other types of genetically engineered monoclonal antibodies.

The volume of the syringe, the walls of which are internally coated with human IL-1β (possibly additionally IL-6, TNF-α), suitable for in vivo use to eliminate murine monoclonal antibodies, is 5-10 ml.

The sterile adapter is a polymer tube with a diameter of up to 4 mm and a length of 40 to 100 mm, which can also be coated from the inside with either monoclonal antibodies or human IL-1, depending on the need. The adapter makes it possible to make multiple passages of the plasma, thereby stimulating the binding of pro-inflammatory cytokines to fixed antibodies.

Further, the solution is illustrated by reference to the figures, which represent the following.

FIG. 1 - embodiments of the first syringe.

FIG. 2 is a top view of the syringe of FIG. 1.

FIG. 3 - embodiments of the syringe with an additional porous structure.

FIG. 4 is a top view of a syringe sector with an additional porous structure.

FIG. 5 is a flow diagram of a process for producing plasma using two syringes.

FIG. 6 is a flowchart of a plasma production process using an adapter and two syringes.

FIG. 7 is a flowchart of a plasma production process using a three-component syringe.

The system includes a syringe 1 (pos. 1), the inner walls of which are coated with a solid-phase polymer sorbent (pos. 2) with monoclonal antibodies to IL-1β fixed on it (possibly in addition to TNF-α, IL-6). In addition to the walls, the solid-phase polymer sorbent can be located on an additional structure (sponge, mesh, tube system) (item 4) inside syringe 1. Syringe 1 contains glass balls (item 3) with diameters from 2.5 to 3.5 mm, which occupy up to 40% of the total volume of the syringe, and 3-6 syringes (syringe 2), the inner walls of which are coated with a solid-phase polymer sorbent with human IL-1β fixed on it (optionally TNF-α, IL-6 is possible), and a sterile adapter (pos. 5, 7) to transfer autologous serum from a syringe with antibodies to a syringe with human IL-1β.

Obtaining autologous serum through the proposed system is as follows. Blood is obtained from the patient using a conventional syringe and injected into syringe 1 (item 1), it is possible to inject not only blood, but also other autologous fluids, plasma, and platelet-rich plasma into syringe 1. Syringe 1 is then incubated. Incubation is carried out on a rack specially designed for these purposes (with the possibility of giving the syringe 1 additional rocking movements or certain cycles of spatial position with a change in the angle of inclination to the horizon from 90 to 30 °, at a temperature of 36.5-37 ° C for 2-24 hours For incubation, for example, racks of LABMEDIX firms (http://labmedix.ru/preanalitika/miksery/programmiruemyj-rotator-vstryaxivatel-rm-1.html), VILITEK (http://vilitek.ru/products/) can be used. 173/868 /). GFL (http://deltaanalytics.ru/3025.php) and other racks with similar characteristics. Centrifugation can be carried out It is possible to add an additional incubation cycle both before and after incubation.At the end of this stage, 5–25 ml of autologous serum is formed in the syringe in the upper part, depending on the volume of blood taken.

After aseptic processing, a sterile adapter (pos. 5, 7) is inserted into syringe 1, through which the obtained serum is transferred from syringe 1 to syringe 2 (pos. 6). The movement can be carried out in several modes, with a change in the speed parameters and a possible change in the direction of flow. The most likely use of the regime at low speed, providing the transition of serum from syringe 1 to syringe 2 within 5-15 minutes.

System consisting of syringes 3 (pos. 8). The system combines in one syringe all the essential details of the plasma production process, with a high content of IL-1RA, with a low content of pro-inflammatory cytokines. The system consists of a vacuum tube that is transformed into a 5-10 ml syringe containing glass beads with a diameter of 2.5 to 3.5 mm, occupying up to 30-40% of the total volume of the syringe and piston, consisting of many capillaries coated inside with a solid phase sorbent with antibodies to IL-1 fixed on it (possibly, in addition to IL-6, TNF-α). The syringe has a piston lock in position 2 (key 10).

The claimed system can be implemented in the following processes for the production of autologous serum.

Process 1.

The process of obtaining plasma using two syringes: syringe 1 (pos. 1) and syringe 2 (pos. 6). Blood is centrifuged in syringe 1 (any modification is possible), then the resulting plasma is collected in syringe 2 using a universal adapter with double-sided needles (item 5). In the process, pro-inflammatory cytokines bind to antibodies and are fixed on the polymer (syringe 1), and antibody residues are removed in syringe 2 using IL-1.

Process 2.

Blood is centrifuged in syringe 1 (any modification is possible), then the resulting plasma is collected in syringe 2 using a universal adapter with double-sided needles (item 5). In the process, pro-inflammatory cytokines bind to antibodies and are fixed on the polymer (syringe 1), and antibody residues are removed in syringe 2 using IL-1. The process can be intensified by passing blood through an adapter (pos. 7), which is internally coated with antibodies to proinflammatory cytokines (IL-1) (it is possible to place fixed IL-1 on the inner surface of the adapter).

Process 3.

Blood is centrifuged in the syringe 3 (pos. 8) (the piston is in position 1 (pos. 9)). Then the piston is transferred from position 1 (pos. 9) to position 2 (pos. 10) using a needle, which is inserted into the test tube. The resulting plasma slowly passes through the piston capillaries, the inner surface of which is covered with monoclonal antibodies to IL-1 (possibly TNF, IL-6). Pro-inflammatory cytokines (IL-1, possibly TNF, IL-6) bind to antibodies and are fixed on the polymer. The piston moves with the needle to the latch. With the same needle, the plasma is drawn into a syringe (universal), which is injected.

Claims (4)

1. A system for producing autologous serum with an increased content of IL-1RA and with a reduced content of pro-inflammatory cytokine, including a syringe, the inner walls of which are coated with a solid-phase polymer sorbent with monoclonal antibodies to IL-1β fixed on it, containing glass beads with diameters from 2.5 to 3.5 mm, occupying up to 40% of the total volume of the syringe, and at least one syringe, the inner walls of which are coated with a solid-phase polymer sorbent with human IL-1β fixed on it, and a sterile adapter, performed with the ability to move autologous serum from a syringe with antibodies into a syringe with human IL-1β. 2. The system according to claim 1, in which monoclonal antibodies to TNF-α and / or IL-6 are additionally used. 3. The system according to claim 1, in which human TNF-α and / or IL-6 are additionally used. 4. The system according to claim 1, in which the solid-phase polymer sorbent is located on an additional structure, which is a sponge and / or mesh, and / or a system of tubes inside the first syringe.

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