KR20150030682A - Hyaluronic acid (HA) inhibitory biomaterials containing anti-inflammatory and anti-blood clotting components - Google Patents

Abstract

The present invention relates to an adhesion preventing agent using hyaluronic acid (HA) as a main ingredient, wherein the HA contains anti-inflammatory and anticoagulant components so as to prevent adhesion. More specifically, an immunity reaction is rarely performed in the human body so that the same is safe. HA approved by the FDA is used as a main ingredient, and an existing adhesion controlling function effectively solves the limit of ingredients.

Description

Hyaluronic acid (HA) adhesion inhibition material containing anti-inflammatory and anti-blood clotting ingredients {omitted}

The present invention relates to a hyaluronic acid gel adhesion inhibitor for greatly lowering the adhesion rate and a method for producing the same.

More particularly, the present invention relates to an anti-adhesion agent containing ingredients for inhibiting inflammation and blood coagulation caused by adhesion of a hyaluronic acid gel used for preventing adhesion between tissues after surgery, and a method for producing the same will be.

Tissue adhesion is a common complication after surgery. It is usually seen during the healing process of inflammation. When it carries out granulation or scar formation, it tangles with each other, or a large amount of fibrin is precipitated. Tissue adhesion occurs after various operations, especially when pelvic adhesion occurs, it causes chronic pain, sexual dysfunction, and infertility. In spinal surgery, if adhesion occurs, the nerve is squeezed to remove adhesions. There is a need for surgery. This coincidence is a big burden on the economy. According to the statistics of 2006, about 69,000 out of 100,000 cases of the 100,000 cases and about 51,000 cases out of 100,000 cases of 100,000 cases of hysterectomy are reported as the frequency And there is a growing interest in preventing adhesion.

Complications following abdominal surgery are known to occur in 49-74% of small intestine obstruction, 15-20% in infertility, 20-50% in chronic pelvic pain, and 19% in perforation in follow-up surgery.

The main factors that cause tissue adhesion are inflammation reaction after surgery, wound due to coarse surgery, foreign body reaction by surgical instruments, excessive bleeding, tissue reaction of suture material. The incidence of postoperative adhesions is only 55-95%. Patients with abdominal surgery suffer from persistent discomfort, intermittent abdominal pain, nausea and vomiting, and dyspepsia. In women, adherence of uterine tissue during pelvic surgery may cause infertility, ectopic pregnancy, chronic pain, and delayed recovery.

Because tissue adhesion is the cause of surgery itself, surgical interventions should be artificially interrupted to prevent adhesion. Several methods have been used to prevent adhesion from the past. These methods include minimizing wounds during surgery, incorporating physical barriers, activating tissue plasminogen activator (TPA) to prevent the use of anti-inflammatory drugs or fibrin formation have.

Although the materials developed so far show a high possibility of prevention of adhesion, only about 50 to 70% of the effect is obtained in clinical studies. Thus, no method has been reported yet that exhibits perfect adhesion prevention function.

Currently, research and development are being conducted to prevent adhesion in various directions. Particularly, in order to inhibit the action of inflammatory reaction, which is one of the main causes of adhesion, it is generally applied to an anti-inflammatory analgesic drug or to an artificial barrier material Lt; RTI ID = 0.0 > inflammatory < / RTI > Attempts to inhibit adhesion include fibrinolytic agents that break down fibrils that connect tissues and tissues, antifibrotic agents that inhibit fibrin growth and prevent adhesion, Antibiotics to inhibit inflammation caused by bacterial infections to prevent adhesion, anticoagulants to prevent adhesion due to the formation of fiber bands during hemorrhagic blood clotting, anti-inflammatory drugs to inhibit inflammation, which is one of the main causes of adhesion Drugs and the like have been used. However, they show some disadvantages in spite of good anti-adhesion effect. The release of fibrinolysis is limited due to its toxicity. In the case of fibrosis inhibitors or antibiotics, it should be continuously injected into the body constantly. In particular, antibiotics may cause resistance problems. Anticoagulants can cause side effects such as dermatitis, urticaria, hair loss, fever, gastrointestinal disturbances, and leukopenia. Excessive use of anticoagulants may not stop the bleeding. Despite these drawbacks, it seems that it is necessary to develop an anti-adhesion agent containing anti-inflammatory and anti-blood coagulation components as an approach to inhibit the adhesion phenomenon more effectively and safely in a short period of time.

First, in early in vivo studies, inhibition of anti-inflammatory drug adhesion was not effective, as demonstrated by in vitro studies. To prevent the adhesion by injecting an anti-inflammatory agent into the surgical site, or to improve the anti-adhesion efficiency by containing an artificial barrier drug. Injection of an anti-inflammatory agent into the surgical site is effective when it is necessary to stay in the in situ for a long period of time. However, since it is a solution for 1 to 3 hours and the sample itself is a solution, it is easily absorbed into the body. Reported. In order to increase the solubility of hydrophobic drugs, surfactants, which are not directly required to prevent adhesion, may be used. However, when the surfactant is used alone, the surfactant is easily absorbed in the body, Is not continuously released.

Accordingly, anti-adhesion agents developed to effectively release anti-inflammatory drugs are attracting attention in order to prevent adhesion. In addition to acting as a physical barrier, it also tries to block adhesion more efficiently by inhibiting the progression of inflammation, which is the main cause of adhesion, by releasing the anti-inflammatory component continuously.

There are drugs that inhibit the action of thrombin which forms fibrin in the blood and inhibit the activation of blood coagulation factors and block adhesion formation because blood clotting phenomenon or thrombus can induce ischemia or fibrin deposition in the surgical site. Heparin, low molecular weight heparin, oral anticoagulants, ancrod, danaparoid, enoxaparin, repirudin, and warfarin.

A variety of materials have been studied to be used as physical barriers in the development of adhesion inhibitors including water soluble polymers such as dextran, hyaluronic acid (HA), carboxymethylcellulose (CMC), polyethylene glycol poly (lactic acid), PLA), poly (glycolic acid), PGA), polylactic acid-glycol (polyglycolic acid) Poly (lactic-co-glycolic acid), PLGA) and the like. Here, hyaluronic acid is composed of D-glucuronic acid and D-N-acetylglucosamine, and is a disaccharide polymer in which β-1,4 and β-1,3 glucosidic bonds are alternately bonded. Interceed ™ (Johnson & Johnson Medical, Inc. Arlinton, TX, USA), HA and CMC crosslinked film Seprafilm ™ (Genzyme Corp. Cambridge, MA) used as biodegradable Oxidized Regenerated Cellulose , USA), Guardix ™ (Korea BioResin development, Hanmi Pharmaceutical sales), which is a mixture of HA and CMC, are commercialized and widely used. Pluronic F127 adhesion inhibitors have the advantage of ease of use, which is a sol form at room temperature and turns into a gel form at body temperature, but has drawbacks that it should only be used on dry, bleeding-free surfaces.

In addition, the anti-adhesion agent made of PLA or PGA, which is a biodegradable synthetic polymer, has a long biodegradation period of 4 weeks or more and a high mechanical strength. However, since it is a hydrophobic material, It is known that an acidic decomposition product may be produced by hydrolysis in the reaction mixture, resulting in an inflammatory reaction which causes adhesion.

Seprafilm (TM) was prepared by blending HA and CMC in a ratio of 2: 1 and adding 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride as a crosslinking activator. carbodiimide hydrochloride, EDC) to form an N-acylurea pendant. The process is limited in that the derivatization proceeds simultaneously during the production of the film to produce a uniform and transparent film. In addition, since the film is entirely derivatized in the same way, the decomposition characteristics in vivo indicate that the derivative moiety composed of HA is easily decomposed (about 66% decomposition), and the CMC derivative portion (about 33% It has remained for a long time. In addition, since fine pores are formed in the process of removing impurities from the produced film, they are easily broken in a dry state and thus are not easy to handle. Moreover, the film is curled when it comes in contact with moisture, and thus the effect varies depending on the application site, and thus the effect can not be proved.

On the other hand, hyaluronic acid (HA), which is highly biocompatible, has been shown to be effective in reducing the formation of intraperitoneal adhesions by smoothing the intestinal surface when used after surgery. The Guardix ™, the first commercialized as an anti-adhesion agent mixed with CMC and HA, is a gel type and can be applied to various surgical sites and has convenience of operation. However, since HA which is absorbed very quickly in vivo is used without modification The effect of preventing adhesion is deteriorating.

Accordingly, the present inventors have developed a safe and excellent biocompatibility hyaluronic acid gel adhesion inhibitor containing ingredients capable of inhibiting inflammation and blood coagulation in order to overcome the problems of low adhesion inhibition rate of conventional anti-adhesion agents.

A basic object of the present invention is to provide a safe hyaluronic acid gel adhesion inhibitor having an effective adhesion preventing function.

Unlike the conventional anti-adhesion agents, the anti-adhesion agent of the present invention has a technical feature that enhances the pharmacological function of the anti-inflammatory agent and the blood coagulation agent, which act as a major factor in the adhesion process.

The anti-adhesion agent of the present invention has a technical feature that the anti-inflammatory and anti-blood coagulation components contained are released at an appropriate rate in parallel with the dissolution of the hyaluronic acid gel.

The present invention minimizes or prevents the adhesion process occurring after surgery, and inhibits the formation of adhesion from immediately after injecting an anti-adhesion agent immediately after the surgical procedure. Particularly, it can be used not only as a physical barrier but also as an agent for enhancing pharmacological and physiological adhesion- The present invention relates to a composition for preventing adhesion, which overcomes the limitations of the prior art by using an inhibitor.

Hyaluronic acid is a linear polymeric polysaccharide in which β-D-N-acetylglucosamine and β-D-glucuronic acid are alternately linked. Have no species and organ specificity, and have excellent biocompatibility and very high viscoelasticity even when they are transplanted or injected into a living body. With this feature, hyaluronic acid can be used for a variety of purposes such as arthritis treatment inserts, gel / film for preventing adhesion, drug delivery system, wrinkle treatment agent, and molding aid.

The use of hyaluronic acid alone as an anti-adhesion agent has a relatively short residence time in the living body due to biodegradability due to excellent biocompatibility of hyaluronic acid, so that the effect of preventing adhesion is weak and the water solubility is high. There is a problem that it is diffused from the surface and flows down, which is a problem in reality application. The anti-inflammatory and anti-blood coagulation inhibitors of the present invention are useful as anti-inflammatory and anti-blood coagulation inhibitors for the prevention and treatment of inflammation and blood coagulation, which is a key factor of 1 to 7 days after surgery, So as to solve the problems of the conventional anti-adhesion agents and to exhibit excellent anti-adhesion properties.

It is preferable that 1 < T < 28, and 1 < T < 21 is more preferable. The number of days of shape retention means the date on which the original shape remained in vivo transplantation. That is, even if the debris of the medical material for preventing adhesion is confirmed, if the debris forms are not the original shape but are in the form of separated fragments, the shape retaining days have elapsed. If the number of days of shape retention is one day or less, it tends to be difficult to obtain sufficient adhesion prevention effect. On the other hand, if the number of days of shape maintenance is more than 28 days, there is a tendency that the possibility of harmful remnants due to living body tends to increase. When the coalesced tissue is formed, a network of fibrin is formed on the wound as a preliminary step. Next, fibroblasts invade the formed network, and collagen fibers are produced, and coalesced tissue begins to form. This series of phenomena starts from day 1 after surgery and becomes extremely active in about 7 days. Therefore, it is preferable that the form of the medical material for preventing adhesion is maintained for one day or more when fibrin network formation or production of collagen fibers is started. On the other hand, if the number of days of shape maintenance is too long, a problem such as formation of capsules may occur.

In addition, the present invention provides an anti-adhesion agent characterized by a hyaluronic acid: anti-inflammatory component: anti-blood clot component = 100: 0.01 to 5: 0.01 to 5 weight ratio. When the ratio of the anti-inflammatory component to the anti-blood coagulation component to hyaluronic acid is less than 0.01 or more than 5, it is difficult to satisfy the appropriate viscosity, adhesiveness and coating property as an anti-adhesion agent.

The present invention can maximize its effect as a gel-type adhesion inhibitor based on hyaluronic acid having an appropriate viscosity. However, the scope of the present invention is not limited thereto, and various kinds of preparations can be applied.

It is possible to provide an anti-adhesion agent that effectively lowers various adhesion phenomena occurring in a medical process by providing a gel-type anti-adhesion agent having excellent anti-inflammation and anti-blood coagulation components and having excellent safety and biocompatibility.

Figure 1 is a photograph of hyaluronic acid (HA) microparticles containing anti-inflammatory and anti-blood clotting components
Fig. 2 is a graph showing the release of the anti-inflammatory component and anti-blood coagulation component contained in the hyaluronic acid gel into the sustained release form
FIG. 3 is a graph showing the degradation rate of hyaluronic acid (HA) gel containing anti-inflammatory component and anti-blood coagulation component

Many anti-adhesion agents using hyaluronic acid (HA) are already known, but the anti-adhesion efficiency is usually about 50%. Accordingly, the present invention has been completed by including anti-inflammatory components and anti-blood coagulation components in order to provide a more effective anti-adhesion agent. It is preferable, but not limited, to mix 0.01 to 5 wt% of each component.

Antiinflammatory components may include, but are not limited to, the following:

But are not limited to, glucocorticin, nabumetone, naproxen, nimesulide, dexamethasone, dexibupropene, diclofenac, diflunisal, rhodozoleck, ronoxocam, loxoprofen, meclofenanic acid, mefenamic acid, meloxicam, But are not limited to, fumarate, fumarate, bromelain, salicylic acid imidazole, celatio peptidase, celecoxib, sulindac, synoxycam, acethethyne, aceclofenac, aspirin, But are not limited to, emolpazone, etodolac, benzidamin hydrochloride, propacetamol hydrochloride, oxaprofen, ibuprofen, ibuproxam, indomethacin, zetofrofen, ketorolac, ketoprofen, clonixin, Fumaric acid, fumaric acid, fumaric acid, fumaric acid, fumaric acid, fumaric acid, fumaric acid, fumaric acid, fumaric acid, fumaric acid, camptothecin sodium, tolpenamic acid, thiaproic acid, fenoprofen, From the group consisting of Rajinobuta zone and Piroxicam More than one can be selected.

Anti-blood clotting components may include, but are not limited to, the following: One or more members selected from the group consisting of heparin, low molecular weight heparin, oral anticoagulants, anchor rod, danaparoid, enoxaparin, repirudin, and warfarin.

The hyaluronic acid (HA) used in the present invention can be used to prepare a crosslinked gel compound through a coupling reaction. The coupling reaction can be carried out using a conventional coupling agent such as 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). The binding reaction of hyaluronic acid (HA) can be carried out using an ordinary binder under acidic conditions of pH 4 to 5. [ In the present invention, such a crosslinked product may be used separately, but it may be directly used as a crosslinked product gel without a separate separation step.

<Preparation Example> Preparation of hyaluronic acid gel containing anti-inflammatory components and anti-blood clotting components

In the present invention, a hyaluronic acid gel to be used for adhesion prevention was prepared by the following process.

0.32 g of sodium hyaluronate (10 ⁶ Da; Shiseido Co., Ltd. Japan) was dissolved in 8 ml of a sodium hydroxide solution at a concentration of 0.05 mol / l at room temperature to prepare a 4.0 wt% aqueous solution.

When sodium hyaluronate was completely dissolved, 0.4 ml of a sodium hydroxide solution at a concentration of 10 mol / l was added to the aqueous solution of hyaluronic acid to increase the degree of alkalinity, and the solution was connected to a micro-bead manufacturing apparatus. The pressure of the compressed air was adjusted to 5 psi, and a hyaluronic acid aqueous solution was flowed at a rate of 0.2 ml / min using a syringe pump to form a fine droplet of hyaluronic acid aqueous solution.

The fine droplets were dropped in a mixed solution of 0.4 ml of divinylsulfone (≥98.0%; Sigma Aldrich, Germany) and 40 ml of isobutyl alcohol, which had been prepared at a rotation rate of 140 to 160 per minute, at room temperature for 24 hours And the reaction was continued to prepare hyaluronic acid microspheres. The solution of divinyl sulfone / isobutyl alcohol in fine beads was removed by soaking in 50 ml of ethanol solution.

After 30 minutes, the second ethanol was again washed with the new ethanol solution, and after 30 minutes, the third ethanol washing was performed with the new ethanol solution. After the third ethanol washing was completed, the obtained hyaluronic acid microspheres were placed in a vacuum oven and vacuum dried at 60 DEG C and 20 mmHg vacuum. The dried hyaluronic acid microspheres were rehydrated in 80 ml of phosphate buffer and immersed for 2 hours to remove unreacted divinyl sulfone remaining in the microspheres.

The hyaluronic acid microspheres were immersed in 40 ml of distilled water in which an appropriate amount of anti-inflammatory components and anti-blood coagulation components were dissolved and stirred at a rotation speed of 140 to 160 for 5 hours or more. Thereafter, the micro-beads of hyaluronic acid were collected by filtration, placed in a vacuum oven, and vacuum dried at a degree of vacuum of 10 mmHg at 20 mmHg.

&Lt; Test Example 1 > Test for release of anti-inflammatory and anti-blood clotting components

The release behavior of each component was evaluated using a hyaluronic acid gel containing anti-inflammatory and anti-blood clotting components. The hyaluronic acid gel was prepared by the method described in the previous example. The concentration of the anti-inflammatory components and anti-blood coagulation components was maintained for 1 week and the amount of the effluent was quantified by an HPLC apparatus.

As a result, as shown in FIG. 2, it was confirmed that anti-inflammatory components and anti-blood clotting components were released in the hyaluronic acid gel.

These results indicate that hyaluronic acid gel containing anti-inflammatory components and anti-blood clotting components can inhibit ischemia by reducing inflammation and blood clotting immediately after application to the surgical site, It can be reinforced.

&Lt; Test Example 2 >

In order to investigate the degree of decomposition of hyaluronic acid gel adhesion inhibitor containing anti-inflammatory components and anti-blood clotting components prepared in the above-mentioned production example, the following method was used. The weight of the dried hyaluronic acid gel was measured. The dried hyaluronic acid gel was immersed in PBS (Phosphate buffered saline) at 37 ° C and the change in weight with time was measured. Before the measurement, the water on the surface of the hyaluronic acid gel was removed using a filter paper and the weight was measured. The decomposition rate was calculated using the following equation, and the results are shown in FIG. From FIG. 3, it can be seen that the degradation of the hyaluronic acid gel (tr HA) containing the anti-inflammatory component and the anti-blood coagulation component is slower than the pure hyaluronic acid gel. However, both gels show decomposition half-life around 5 days.

Decomposition rate (%) = weight of gel when swollen / weight of gel when dried × 100

Claims (7)

A composition comprising a hyaluronic acid gel containing an anti-inflammatory component and an anti-blood coagulation component for preventing adhesion. The composition of claim 1, wherein the anti-inflammatory component is selected from the group consisting of glutamate, naproxen, naproxen, nimesulide, dexamethasone, dexibuprofen, diclofenac, diflunisal, lona zolac, But are not limited to, fumaric acid, fumaric acid, mefenamic acid, mefenamic acid, mefenamic acid, meloxicam, monoflumate, benorylate, bromelain, salicylic acid imidazole, ceratio peptidase, cerecoxib, sulindac, Wherein the compound is selected from the group consisting of aspirin, alminopropene, amnepax sodium, emolpazone, etodolac, benzodamine hydrochloride, propacetamol hydrochloride, oxaprofen, ibuprofen, ibuproxam, indomethacin, But are not limited to, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, antioxidants, Fluorobipropen, flupenamic acid, pyrazinobutane, and &lt; RTI ID = 0.0 &gt; Wherein the hyaluronic acid gel composition is at least one selected from the group consisting of paclitaxel. 3. The method of claim 2,
Wherein the anti-inflammatory component is present in an amount from about 0.01% (w / w) to about 5.0% (w / w) of the total composition. The method according to claim 1,
Wherein the anti-blood coagulation component is an anti-blood coagulant selected from the group consisting of heparin, low molecular weight heparin, anchor rod, dinaparoid, enoxaparin, repirudin and wararin. 5. The method of claim 4,
Wherein the anti-blood coagulation component is present in an amount from about 0.01% (w / w) to about 5.0% (w / w) of the total composition. The method according to claim 1,
Wherein the hyaluronic acid polymer has an average molecular weight of greater than 1,000,000 Da. The anti-adhesion composition according to any one of claims 1, 2, and 4, wherein the release period of the anti-inflammatory component and the anti-

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