WO2007137733A2

WO2007137733A2 - Wound dressing comprising polyacrylates and the use thereof

Info

Publication number: WO2007137733A2
Application number: PCT/EP2007/004517
Authority: WO
Inventors: Hans Smola
Original assignee: Paul Hartmann Ag
Priority date: 2006-05-26
Filing date: 2007-05-22
Publication date: 2007-12-06
Also published as: WO2007137733A3; DE102006024748A1

Abstract

The invention relates to wound dressings and to the use of metalloprotease inhibitors during the treatment of the wound. Said metalloprotease inhibitors can especially be used in the granulation phase of the wound healing process.

Description

Title: Wound dressing with polyacrylates and their use

description

The invention relates to protease inhibitors, in particular for wound treatment, and to a wound dressing and their use in modern wound treatment.

The activity of an enzyme can be influenced in many different ways. Thus, for example, the activity of the enzyme can be increased or decreased by influencing the enzyme protein itself, the coenzyme or the substrate. This influence can occur not only by inhibitors or activators, but also by external factors such as temperature, pH, ionic strength or the polarity of the solvent. An influence of the latter factors is nonspecific by modification of the enzyme structure via surface effects, e.g. Interactions with charged groups or disturbances of the hydration shell. The active center is not specifically influenced.

Enzyme inhibition (inhibition) is understood to mean the negative influence on the enzyme activity or the reduction of the catalytic activity of an enzyme or coenzyme by specific or nonspecific inhibitors or inhibitors. In this case, the inhibition can be reversible or irreversible.

An irreversible enzyme inhibition is usually the result of a covalent linkage of the inhibitor with the active site of the enzyme. These irreversible inhibitors are substances which are first recognized by the enzyme as a substrate and taken up in the active center. There, the inhibitors enter into a solid covalent bond with amino acid residues of the active site, thereby permanently blocking it. A known irreversible inhibitor is the antibiotic penicillin, which can irreversibly turn off an enzyme of bacterial cell wall synthesis.

The so-called reversible enzyme inhibition is basically reversible and plays a crucial role in the fine regulation of metabolism in living organisms. Reversible inhibition is further distinguished into competitive inhibition and non-competitive inhibition. In competitive inhibition, the substrate competes with the inhibitor for binding to the active site of the enzyme. Because the substrate and the Inhibitor can not simultaneously bind to the enzyme and the inhibitor, in contrast to the substrate is not enzymatically implemented, the measurable enzyme activity is reduced. As the concentration of the inhibitor increases, increasing displacement of the substrate occurs, resulting in a decrease in enzyme activity. Increasing the substrate concentration reverses this process and allows for increased substrate conversion. In the case of non-competitive inhibition, the inhibitor outside the active site binds to both the free enzyme and the enzyme / substrate complex. The binding of the inhibitor does not lead to the inhibition of substrate binding, but to a conformational change of the enzyme, which is thereby inactivated. The binding of the inhibitor and the inactivation of the enzyme take place independently of the presence of substrate.

Proteases (peptidases, peptide hydrolases) are enzymes that catalyze the hydrolytic cleavage of a peptide bond in proteins and peptides (proteolysis). Proteases thus belong systematically to the group of hydrolases. Proteases are subdivided with respect to the cleavage site in the substrate. Thus, the cleavage of peptide bonds in the interior of peptides or proteins is catalyzed by endopeptidases (proteinases), whereas peptide bonds at the end of a peptide or protein molecule are cleaved by exopeptidases (formerly peptidases).

Another distinction of proteases is made with regard to their active sites for catalysis responsible groups. For example, a) serine proteases, b) cysteine proteases, c) aspartate proteases and d) metallo proteases are distinguished from one another. For example, the group of metallo proteases (metallo-peptidases) have metal ions in their active center involved in the catalytic mechanism of proteolysis. These metal ions, especially divalent metal cations such as magnesium, zinc, calcium, iron and the like are also considered coenzyme. According to the distinguishing feature described above, there are furthermore a) metallo-endopeptidases (metalloproteinases) and b) metallo-exopeptidases.

Matrix metallo-proteases (MMPs) - also called matrixins - belong to a family of proteases that have been defined by structural homologies. Their enzymatic activity is dependent on metal ions in the active center. Metallo proteases were first identified by their role in tissue remodeling, particularly by degradation of the extracellular matrix. Systematically, this group of proteases belongs to the metallo-endopeptidases (metalloproteinases). Among the matrix metalloproteinases include other collagenases, gelatinases, stromeolysins, matrilysin and others. For a review and classification see Parks, Matrix Metalloproteinases, Biology of Extracellular Matrix Series, Ed. Mecham, RP, Academic Press, Inc., San Diego, Calif. (1998) and in Table 1 below. Synonyms and, where appropriate, the numbering of the enzymes according to the Enzyme Commision (EC) are given in Table 1. The matrix metalloproteinases are synthesized and secreted as inactive precursors. Complex mechanisms that have not yet been finally clarified in detail will convert them into the active form. All metallo proteases can be inhibited by ethylenediaminetetraacetic acid (EDTA).

Table 1: Matrix metallo-proteases (MMP)

- A -

Serine proteases have an active L-serine residue essential for catalysis, which can be inhibited by diisopropylfluorophosphate. These proteases are also assigned a crucial function in wound healing. The group of serine proteases includes, for example, chromotrypsin, elastase, kallikrein, plasmin, trypsin, thrombin, etc. Most known serine proteases have, in addition to the L-serine residue in their active center, the residues of the amino acids L-histidine and L-histidine. Asparagine on. All three amino acid residues participate in the protolysis of a cascade of reactions in which a proton of the L-serine residue is transferred to the substrate. Insofar as a serine-dependent mechanism is found in proteinases, one also speaks of serine proteinases.

Matrix metalloproteases are believed to play a crucial role in wound healing. For example, keratinocytes require matrix metallo proteases to migrate across the provisional matrix for epithelial wound closure (Pilcher et al., 1997, Beare et al., 2003, Mirastschijski et al., 2004). These metallo proteases are formed in cells as an inactive pro form, secreted from the cell, and activated in the extracellular milieu. They are characterized by structural similarities and require divalent ions in the active site for proteolytic activity. Only after activation and in the presence of, for example, Ca ²⁺ or Zn ²⁺ ions can they degrade the respective substrates (review in Nagase and Woessner 1999).

In more recent work in the field of wound treatment, it has been found that pathological wound healing associates excess protease activity in wound secretion (Trengove et al., 1999, Wysocki et al., 1993, Weckroth et al., 1996). It was also shown experimentally that increased MMP expression in the skin markedly retards wound healing (Di Colandrea et al., 1998). Furthermore, an excess of protease activity has been shown to decrease growth factors (Tengrove et al., 2003). It has also been shown that in chronic wounds, the extracellular matrix (Wysocki et al., 1990), growth factors (Duckworth et al., 2004, Lauer et al., 2000) and their receptors are degraded, so that in the wound area Relative deficit arises on these substances. Thus, excessive protease activity, particularly excessive metallo-protease activity, hampers wound healing to an extraordinary degree. Attempts to pharmacologically reduce this protease activity by specific inhibitors have not been successful so far.

Therefore, it is an object of the present invention to provide an improved agent for the treatment of chronic wounds. In addition, should be a means be provided, which influences the pathological condition of a wound so that a normal, natural wound healing process can take place. In addition, a method for the treatment of chronic wounds and a use for the production of an agent for the treatment of chronic wounds to be provided.

Surprisingly, this object is achieved by metallo-protease inhibitors from the group of polyacrylates. In particular, it has been shown that these polyacrylates inhibit metallo-proteases via diffusible mechanisms as well as by direct binding compartmentalize and thus escape the wound exudate or the wound. Thus, the present invention also relates to the use of a metallo-protease inhibitor from the group of polyacrylates for the preparation of an agent for the treatment of chronic wounds. In particular, the use of polyacrylates for the inhibition of metallo-proteases in chronic wounds and the use of polyacrylates for the preparation of an agent for the inhibition of metallo-proteases in chronic wounds is the subject of the invention.

In addition, it has been shown that metallo-proteases are bound or compartmentalized by polyacrylates so that these metallo-proteases can be removed with the polyacrylates from a wound fluid or a wound. Thus, by means of the polyacrylates, an excess of metallo proteases in chronic wounds can be trapped so that a natural healing process can take place. Thus, the use of polyacrylates for inhibition by compartmentalization of metallo-proteases in chronic wounds and the use of polyacrylates for the preparation of an agent for inhibition by compartmentalization of metallo-proteases in chronic wounds is the subject of the present invention.

Chronic wounds can thus be treated with a topical agent in a particularly preferred manner, since it enables the metalloproteases bound to the polyacrylates to be removed together with the polyacrylates. Thus, the use of polyacrylates in topical agents for the inhibition of metallo-proteases in chronic wounds is the subject of the present invention.

The healing of wounds is based on the ability of the skin to regenerate epithelia as well as connective and supporting tissues. It is characterized as a complex event of interdisciplinary cell activities that progressively drive the healing process. Thus, regardless of the type of wound, three essential ones in the literature Healing phases of a wound described. These include the inflammatory or exudative phase for hemostasis and wound cleansing (phase 1, cleansing phase), the proliferative phase for building granulation tissue (phase 2, granulation phase) and the differentiation phase for epithelialization and scarring (phase 3, epithelialization phase). An excess of metallo-proteases is particularly troublesome in the granulation phase of wound healing, since due to the disturbed equilibrium of metallo-proteases to newly formed connective tissue, insufficient tissue build-up can take place. The tissue structure is critically dependent on the stabilization of newly synthesized extracellular matrix molecules that are assembled into a three-dimensional tissue, followed by angiogenesis and proliferation of cells into the wound area in parallel and in further steps.

It has now been found that polyacrylates can be used in particular in the proliferative phase of wound healing. In this phase, any inflammations have subsided or not present so far that a granulation and thus a

Cell growth could take place, provided that there is no excess of metallo proteases. This excess can now be trapped by means of polyacrylates. Thus, the tissue structure is promoted by the elimination of the wound healing disturbing mechanisms, which is clinically noticeable in an improvement of the wound state and the reduction of wounds by decreasing the wound volume and / or the size of the wound. Thus, the use of polyacrylates to inhibit metalloproteases for tissue engineering and / or regulation of tissue engineering in chronic wounds and the use of polyacrylates for the preparation of an agent for inhibiting metallo proteases for tissue engineering and / or for regulating tissue engineering in chronic Wounds also the subject of the present invention.

In the context of the present invention, a polyacrylate is understood as meaning a synthetic polymer comprising as monomer (M 1) acrylic acid (2-propenoic acid, CH ₂ = CHCO ₂ H) and / or a salt thereof, which has a monomer content of more than 70 Wt .-% acrylic acid and / or a salt thereof (based on the total weight of the polyacrylate). In particular, polyacrylates according to the invention have a monomer content of more than 80% by weight of acrylic acid and / or a salt thereof and very particularly preferably more than 95% by weight of acrylic acid and / or a salt thereof, based on the total weight of the polyacrylate. In this case, the polyacrylate may be present as homopolymer, copolymer or block polymer. If the polyacrylate is present as a copolymer or block polymer, the monomer content of the monomer M1 is in each case the polymer at more than 70%, in particular more than 80% and most preferably more than 95% based on the total weight of the polyacrylate. In these copolymeric polyacrylates or block polymer polyacrylates in addition to the monomer M1 as comonomers M2 in particular α.ß-unsaturated ethers (vinyl ethers), α, ß-unsaturated carboxylic acids or α, ß-unsaturated carboxylic acid esters (vinyl esters) may be included. Of the comonomers M2 of the α, β-unsaturated carboxylic acids, especially methacrylic acid (2-methylpropenoic acid), ethacrylic acid (2-ethylpropenoic acid), crotonic acid (2-butenoic acid), sorbic acid (frans-2,4-hexadienoic acid), maleic acid (c / s-2-butenedioic acid) or fumaric acid (frans-2-butenedioic acid). In a particularly preferred form of the invention, however, it can also be provided that the polyacrylate of a) a

Homopolymer of acrylic acid and / or b) a copolymer of i) acrylic acid and a salt of acrylic acid, ii) of methacrylic acid and a salt of methacrylic acid or iii) of acrylic acid and methacrylic acid and salts thereof. Furthermore, however, it can also be provided that the polyacrylate is a mixture of different polyacrylates.

In particular, the α, β-unsaturated carboxylic acids and the acrylic acid may be present in neutralized form as a salt, in unneutralized form as the free acid or in mixtures thereof. In particular, polyacrylates composed of acrylic acid and salts of acrylic acid have been found to be particularly effective. In particular, alkali metal or alkaline earth metal salts should be emphasized. In particular, polyacrylates consisting of homopolymers and / or of copolymers comprising as monomers acrylic acid and / or sodium or potassium acrylate have been found to be particularly effective.

Furthermore, it has surprisingly been found that the present object is achieved in a special way to be emphasized by metallo-protease inhibitors from the group of crosslinked and / or crosslinked polyacrylates. These polyacrylates preferably comprise a) a homopolymer which consists of the monomers M1 and is crosslinked and / or cross-linked by means of a crosslinker, and / or b) a copolymer which consists of the monomers M1 and M3, the monomer M1 being acrylic acid and / or or a salt thereof and the monomer M3 is selected from the group of crosslinkers. That is, these polyacrylates include a post-crosslinked polyacrylate and / or a polyacrylate copolymerized from acrylic acid and / or a salt thereof and a crosslinker. This also includes the use of a polyacrylate which comprises a) a homopolymer consisting of the monomers M1 crosslinked and / or cross-linked by means of a crosslinker, and / or b) a copolymer consisting of the monomers M1 and M3, wherein the monomer M1 is acrylic acid and / or a salt thereof and the monomer M3 is selected from the group of crosslinkers, for the preparation of a means for the treatment of chronic wounds subject of the present invention.

In particular, it has been found that crosslinked and / or crosslinked polyacrylates containing as crosslinking agents compounds V1 which have at least two ethylenically unsaturated groups within one molecule, or compounds V2 which have at least two functional groups which are functionalized with acrylic acid functional groups or / and a salt thereof in a condensation reaction, in an addition reaction or in a ring-opening reaction, or compounds V3 having at least one ethylenically unsaturated group and at least one functional group functional with functional groups of the acrylic acid and / or one of its salts and / or the α, β-unsaturated comonomers can react in a condensation reaction, in an addition reaction or in a ring-opening reaction, are particularly effective. Crosslinking of the polymers by the free-radical polymerization of the ethylenically unsaturated groups of the crosslinker molecule with the monoethylenically unsaturated monomers acrylic acid and / or one of its salts and / or one of the α, β-unsaturated comonomers is achieved by the compounds V1, while in the compounds V2 crosslinking of the polymers is achieved by condensation reaction of the functional groups with the functional groups of the acrylic acid and / or one of its salts or of an α, β-unsaturated comonomer. In the case of the compounds V3, crosslinking of the polymer accordingly takes place both by free-radical polymerization of the ethylenically unsaturated group and by condensation reaction between the functional group of the crosslinker and the functional groups of the monomers.

Preferred compounds V1 are polyacrylic acid esters or polymethacrylic acid esters obtained, for example, by the reaction of a polyol, such as, for example, ethylene glycol (1,2-ethanediol), propylene glycol (1,2-propanediol), trimethylolpropane (2-ethyl-2-hydroxymethyl-1, 3 propanediol), 1,6-hexanediol, glycerol (1,2,3-propanetriol), pentaerythritol (2,2-bis (hydroxymethyl) propane-1,3-diol), polyethylene glycol (HO- (CH ₂ -CH ₂ - O) _n -H where n = 2 to 20) where n = 1 to 20), polypropylene glycol (HO- (CH (CH ₃ ) -CH ₂ -O) _n -H where n = 2 to 20), an aminoalcohol, a polyalkylenepolyamine, such as diethylenetriamine or triethylenetetraamine, or an alkoxylated polyol with acrylic acid or Methacrylic acid are recovered. Particularly preferably, the crosslinked polyacrylate may be a polyacrylate which is crosslinked by means of a compound V1 which is a di-, tri- or tetraester of polyacrylic acid or polymethacrylic acid which is obtained by reacting an alkoxylated polyol, in particular an ethoxylated polyol. in particular ethoxylated ethylene glycol, ethoxylated propylene glycol, ethoxylated trimethylolpropane, ethoxylated 1,6-hexanediol or ethoxylated glycerol, having an average number of ethylene oxide units n per hydroxy group of n = 1 to 10 can be obtained with acrylic acid or methacrylic acid. Further compounds V1 are polyvinyl compounds, polyallyl compounds, polymethylallyl compounds, acrylic acid esters or methacrylic acid esters of a monovinyl compound, acrylate or

Methacrylic acid esters of a monoallyl compound or monomethylallyl compound, preferably monoallyl compounds or monomethylallyl compounds of a polyol or an aminoalcohol. In this context, reference is made to DE 195 43 366, DE 195 43 368.

Surprisingly, the above object is also achieved by metallo-protease inhibitors from the group of polyacrylate superabsorbers. For the purposes of the present invention, a polyacrylate superabsorber is understood as meaning a polyacrylate having the abovementioned features and capable of carrying at least 15 times its own weight of physiological saline solution (0.9% strength NaCl solution,

Water). Thus, the present invention also relates to the use of a metallo-protease inhibitor from the group of polyacrylate superabsorbents for the preparation of an agent for the treatment of chronic wounds.

Superabsorbents have long been known in modern wound care. However, these are used only as a wound fluid absorbing medium. When these superabsorbents are used in this manner, only the fluid balance of the wound or wound bed is regulated. Surprisingly, it has therefore been found that polyacrylate superabsorbents are able to inhibit metallo proteases and thus actively promote wound healing. In particular, it has also been found that polyacrylate superabsorbents, in particular crosslinked polyacrylate superabsorbents, inactivate matrix metalloproteases. Thus, the problem is solved in a preferred manner of matrix metallo-protease inhibitors. Particularly preferred are matrix metallo-protease inhibitors selected from the group of polyacrylate superabsorbents and in particular from the group of crosslinked polyacrylate superabsorbents. In either case, the polyacrylate superabsorbent may be in the form of fibers, particles or gels. In this case, the polyacrylate superabsorber may be in dry or already swollen form. In swollen form, the polymer is present in gel-like, discrete fibers or particles. In this case, an inventive metallo-protease inhibitor from the group of polyacrylate superabsorbents can be present in the form of particles which are mixed with water, physiological saline solution or Ringer's solution, the particles already being present in swollen form.

Moreover, polyacrylate superabsorbents and in particular crosslinked polyacrylate superabsorbents which are present as particles and have a particle size distribution whose particle fraction between 850 μm and 300 μm has at least 55%, in particular at least 65% and very particularly at least 70%, have proven to be particularly preferred. is (measured according to EDANA 420.2-02). Further preferred are crosslinked and / or crosslinked polyacrylate superabsorbents. In particular, superabsorbent polyacrylates are also preferred whose particle fraction of particles smaller than 850 μm is at least 85%, particularly at least 90% and very particularly at least 95%, based on all particles (measured according to EDANA 420.2-02).

Surprisingly, the above-mentioned object is achieved, in particular, by matrix metalloproteinase inhibitors from the group of the polyacrylates, in particular the crosslinked and / or crosslinked polyacrylates. Polyacrylates here are polyacrylates with the characteristics described above. Thus, the present invention also relates to the use of a matrix metallo-protease inhibitor from the group of polyacrylates, in particular the crosslinked and / or crosslinked polyacrylates for the preparation of an agent for the treatment of chronic wounds. In particular, the use of polyacrylates for the inhibition of matrix metallo-proteases in chronic wounds and the use of polyacrylates for the preparation of an agent for the inhibition of matrix metallo-proteases in chronic wounds are the subject of the invention.

In a further advantageous use, the polyacrylates are used for inhibition by compartmentalization of matrix metalloproteases in chronic wounds and for the production of an agent for inhibition by compartmentalization of matrix metalloproteases in chronic wounds. In addition, these polyacrylates may preferably be used for inhibiting matrix metalloproteases for tissue engineering and / or for regulating the tissue structure in chronic wounds and for producing an agent for inhibiting metalloproteases for tissue engineering and / or regulation of the tissue Tissue construction can be used in chronic wounds.

Chronic wounds can be defined as wounds whose healing process deviates from normal wound healing in one or all stages of wound healing. Thus, from acute, normal healing wounds, e.g. wound infection causes a chronic wound characterized by a delayed rate of healing. The transition from an acute to a chronic wound can take place at any stage of wound healing. Clinically, chronic wounds are defined as wounds that require more than 6-8 weeks to heal, although this definition does not cover all clinical pictures correctly. Chronic wounds are more a diagnosis based on the clinical experience of the medical staff.

Chronic wounds occur in particular due to mechanical stress (pressure sores, pressure ulcers, pressure ulcers), venous insufficiency (ulcus cruris venosum, venous ulcers), atherosclerotic vascular change (ulcus cruris arteriosum, arterial ulcers), neuropathic changes (diabetic foot syndrome, neuropathic Ulzera), but also as a result of autoimmune diseases, tumors (exulsive tumors) or radiation damage in tumor therapy.

The pressure ulcer is defined as by external (long-term) pressure with

Compression of vessels and local ischaemia-induced trophic disturbance of tissues (especially skin and subcutaneous tissue) with necrosis, maceration, possibly infection. Decubital ulcers are especially associated with bed rest, especially at body sites where the skin is in direct contact with the bone, but also e.g. under badly fitting dentures and too narrow plaster casts.

The decubitus ulcer is divided into the following stages. Deco Stages Stage II, Stage III and Stage IV are known as chronic wounds in particular:

- Pressure ulcers - Level I: This is a persistent, circumscribed reddening of the skin, which persists even with relief. The redness is sharply circumscribed and may be hardened or overheated. The skin is still intact.

- Pressure ulcers - Stage II: In this phase, blistering and abrasion of the skin occurs and thus partial loss of the skin. The epidermis to the parts of the dermis is damaged. At this stage there is a superficial wound or flat ulcer. - Pressure ulcers - Level IM: At this advanced stage, a loss of all layers of the skin is already observed. In addition, there are subcutaneous damage To observe tissue and possibly necrosis, which can reach down to the underlying muscle tissue. Experience has shown that it is necessary to delineate the necrotic tissue until the entire extent of tissue damage becomes apparent. The pressure ulcer III shows clinically as an open, deep ulcer. - Pressure ulcers - Stage IV: At this most critical stage, there is a loss of all

Skin layers with extensive destruction, tissue necrosis or damage to muscles, bones or supporting structures (tendons, joint capsules) are recorded. The pressure ulcer IV shows clinically as a large, open and deep ulcer.

Thus, the present invention also relates to the use of a metalloproteinase inhibitor, in particular a matrix metallo-protease inhibitor, from the group of the polyacrylates, in particular the crosslinked and / or crosslinked polyacrylates, for the treatment of and for the preparation of an agent for treatment of decubitus ulcer, venous leg ulcer, leg ulcer or diabetic foot syndrome. In this case, the tissue structure is promoted by the elimination of the wound healing disturbing mechanisms, which is clinically noticeable in an improvement of the wound state and the reduction of wounds.

In a further advantageous use, the polyacrylates are used for inhibition by compartmentalization of matrix metalloproteases and for the preparation of an agent for inhibition by compartmentalization of matrix metalloproteases in pressure ulcers, venous leg ulcers, leg ulcers or diabetic foot syndrome. In addition, these polyacrylates may preferably be used for inhibiting metalloproteases and / or matrix metalloproteases for tissue engineering and / or for regulating the tissue structure in these diseases.

According to a further aspect of the present invention, a wound dressing comprising a metalloproteinase inhibitor, in particular a matrix metalloproteinase inhibitor selected from the group of the polyacrylates, in particular the crosslinked and / or crosslinked polyacrylates, is also an object of the present invention , In this case, all polyacrylates of the type described above can be used. It is provided in particular that the inhibitor in solid form as particles or in pre-swollen, gel-form on a carrier material, in particular a fiber material or is introduced. Thus, the use of a wound dressing comprising a carrier material, in particular a fiber material, and a metallo-protease inhibitor, in particular a matrix metallo-protease inhibitor selected from the group of polyacrylates, in particular the crosslinked and / or cross-linked polyacrylates, for the preparation of a Agent for the treatment of chronic wounds Subject of the present invention. In addition, a wound dressing comprising a carrier material, in particular a fiber material, and at least one polyacrylate, in particular a crosslinked and / or crosslinked polyacrylate, in the form of particles, fibers or a gel for inhibiting matrix metalloproteases in chronic wounds is the subject of the present invention ,

As has been shown, metallo-proteases, in particular matrix metallo-proteases, are bound or compartmentalized by the polyacrylate, which is an essential constituent of the wound dressing, and which may be present in particular in the form of particles, fibers or a gel. This has the advantage that an excess of metalloproteases, in particular of matrix metalloproteases, can be brought to a reduced amount required for natural wound healing by the wound dressing. These bound metallo proteases are no longer available as inhibitors of wound healing. By changing the dressing, these metallo-proteases can be permanently removed. Thus, a wound dressing according to the invention comprises at least one polyacrylate as metalloproteinase inhibitor, wherein the metallo proteases are bound and / or compartmentalized by the polyacrylate.

In particular, non-wound-adhesive wound dressings comprising a metalloproteinase inhibitor, in particular a matrix metalloproteinase inhibitor selected from the group of polyacrylates, are also subject matter of the present invention. These wound dressings are particularly supportive of wound healing in the granulation phase of wound healing since, on the one hand, due to the content of metalloproteinase inhibitor, they absorb an excess of metalloproteases, in particular by compartmentalization or binding, and on the other hand, they do not damage newly grown tissue during a dressing change. These non-wound adhesive dressings include as

Wound contact layer, in particular a non-wound-adhesive knitted fabric, knitted fabric or nonwoven, which furthermore preferably consists of a hydrophobic fiber material. In particular, the wound contact layer may be a knit, knit or fabric of a hydrophobic polyethylene, polypropylene, polyester or viscose polymeric material. The design as a knitted fabric or fabric, the

Wound contact layer are stretched or deformed in one or more directions, without that she contracts or aligns herself again. In addition, the surface of such a wound contact layer conforms precisely to the surface of the skin or wound to be treated.

Alternatively, the wound dressing may also comprise a polyacrylate, a crosslinked and / or crosslinked polyacrylate, or a polyacrylate superabsorbent and a hydrophilic fiber material. In particular, water-insoluble fibers of cellulose, in particular largely delignified industrial pulp fibers, in particular wood pulp fibers, in particular a fiber length of <5 mm, can be used as the hydrophilic fiber material. The fibrous material may also contain hydrophilic fibrous material of regenerated cellulose, carboxymethylcellulose, carboxyethylcellulose, hydroxymethylcellulose or hydroxyethylcellulose. It may also be contemplated to provide a fiber blend of cellulosic, regenerated cellulose, carboxymethylcellulose, carboxyethylcellulose, hydroxymethylcellulose or hydroxyethylcellulose fibers and fibers of polyethylene, polypropylene or polyester. In a very particularly preferred embodiment, the wound dressing comprises a mixture of cellulose fibers, polypropylene fibers and a particulate, crosslinked polyacrylate superabsorber.

A wound dressing according to the invention is particularly suitable for the treatment of comparatively deep chronic wounds. For example, you can treat deep dermal ulcers, which are very often very wet. In addition, dry wounds such as dry leg ulcers can be treated with an existing wound dressing. In this case, the wound dressing shows its ability to supply liquid to the wound and at the same time to ensure that undesired substances, deposits and necroses are gently debrided. Supported is the

Wound healing process by semi-occlusive closure using a secondary wound dressing such as a film dressing to prevent unwanted contamination. Other categories of wounds to which the dressing may be applied include, but are not limited to, bedsores stage II, III, IV (ulcer), ulcus cruris (leg ulcer, open leg), diabetic foot syndrome, especially chronic forms of these wounds. Thus, the present invention also relates to the use of a wound dressing comprising a matrix metallo-protease inhibitor from the group of polyacrylate superabsorbents for the production of a wound healing agent, in particular for the treatment of bedsores stage II, III, IV (pressure ulcer) or ulcus cruris (Lower leg ulcer, open leg) or diabetic foot syndrome, especially chronic forms of these wounds. In the following, the effectiveness of the polyacrylates will be illustrated by means of a few tests. The following relationships are explained in more detail with the figures:

FIG. 1: Inhibition of MMP activity in wound exudate by a polyacrylate FIG. 2: Inhibition of MMP activity by direct binding to a polyacrylate FIG. 3: Inhibition of MMP activity without direct binding to a polyacrylate

Wound exudate is treated in the following as representative of biological fluids including excrements. Wound exudate with high protease activity was recovered from patients with chronic wounds and incubated with liquid pre-activated polyacrylates for 2 hours at room temperature.

In the following example, the preactivation was carried out by mixing 0.2 g of crosslinked polyacrylate (Favor Z3034, Stockhausen, Krefeld - Germany) with 36.94 ml of Ringer's solution (8.6 g of NaCl, 0.3 g of KCl, 0.33 g CaCl ₂ • 2 H ₂ O ad 1000 ml water), wherein the mixture for 24 hours in a sealed vessel is allowed to stand. 100 μg wound fluid protein in a chronic wound (ulcus cruris) with increased metalloproteinase activity was diluted with Ringer's solution to 150 μl and mixed with the same volume of pre-activated polyacrylate, whereby the mass of the pre-activated polyacrylate superabsorber can be approximately equated to the volume

(Note: Wound fluid contains different concentrations of protein depending on the patient.) For each test, the protein content in each wound fluid (WF) was determined and an aliquot of 100 μg protein was taken and made up to 150 μl with Ringer's solution). After incubation, the supernatant is purified by means of an enzyme substrate ((Gly-PrO-HyP ^ _5- Gly-PrO-LyS (MCa) -gly-PrO-PrO-GliIVaI-VaI-Gly-Glu-LyS (DnP) -gly-glu -GIn- (Gly-Pro-Hyp) ₅ -NH ₂ ) * Hyp = 4-hydroxy-L-proline) for metallo proteases and fluorescence photometric measurement for the presence of gelatinase enzyme activity against a standard or appropriate controls. For this purpose, a commercially available kit InnoZyme® Gelatinase Activity Assay Kit (Catalog No. CAB003, Calbiochem, San Diego - USA) was used. Contrary to the instructions of the manufacturer, the activation of the enzyme by APMA (p-aminophenylmercuric acetate) was dispensed with in order to detect the existing MMP activity. The modified protocol was: - 90 μl sample (wound fluid, polyacrylate superabsorber or EDTA pretreated wound fluid) or standard (standard was with activation buffer containing APMA) or control consisting of activation buffer only, was treated with 10 μl of the final substrate solution (see Regulation of the manufacturer). - The mixture was incubated at 37 ⁰ C for 24 hours.

The fluorescence was measured at a wavelength of 320 nm (excitation) and 405 nm (emission) and the values were plotted against the standard.

FIG. 1 shows the inhibition of MMP activity in the wound exudate by a polyacrylate. In this case, 100 μg of wound exudate protein (Wound Fluid) were filled up with Ringer's solution to 150 μl and incubated with 150 μg of Ringer's solution pre-activated polyacrylate for 2 hours. Thereafter, the supernatant was spiked with the MMP peptide substrate, and the enzyme activity was measured fluorescence photometrically after incubation for 24 hours. As biological controls wound fluid without polyacrylate treatment (set at 100%) and wound fluid whose MMP activity was blocked by addition of EDTA (10 mM final concentration) were used as a negative control.

Here, the polyacrylate-treated wound fluids were measured against untreated wound fluid of the same patient (control), which was set as 100% enzyme activity. The activity of the polyacrylate-treated wound fluids, when measured, reached values well below the values of the control. Inhibition with 10 mM EDTA, a strong inhibitor of the metalloproteases, was only able to inhibit the residual activity of the polyacrylate-treated wound fluids by a few percentage points (FIG. 1). These experiments were reproduced with wound fluid from different patients.

It was then analyzed whether MMP activity from wound fluid binds to polyacrylate. Wound secretion of chronic wounds was incubated with polyacrylate for 2 hours analogously to the above experiments. Thereafter, the supernatant was removed and mixed with the same volume 2x SDS-GeI sample buffer (see below), boiled at 95 ⁰ C for 10 minutes in a water bath and then stored until its use at - 20 ⁰ C. The polyacrylate superabsorbent thus treated was incubated 3x with an excess (5 w / w) wash buffer (10g Bovine Serum Albumin (Sigma A-2153), 8.6g NaCl, 0.3g KCl, 0.33g CaCl ₂ - 2 H ₂ O ad 1000 ml of water). The thus treated polyacrylate was mixed with 1 volume (v / w) 2x SDS-gel sample buffer, boiled at 95 ⁰ C for 10 minutes in a water bath and then stored at -20 ⁰ C or directly subjected to gelatin zymography. For this purpose, an aliquot of the wound fluid, the polyacrylate-treated supernatant and the polyacrylate-bound proteins were applied to a gelatin-containing SDS gel and separated by electrophoresis. This technique is extremely sensitive and is based on a separation of the proteases in an SDS gel, which simultaneously contains a protease substrate (gelatin).

The gel is composed of (all chemicals Fa. Sigma-Aldrich Chemie GmbH, 89555 Steinheim - Germany):

- 3.3 ml gelatin solution (gelatin, porcine skin (CAS No 9000-70-8), 3 mg / ml H ₂ O),

0.85 ml of distilled H ₂ O,

- 2.5 ml of 1.5M Tris (tris (hydroxymethyl) aminomethane) /

HCl / 0.4% SDS solution (sodium dodecyl sulfate solution), pH 8.8, - 3.35 ml 30% acrylamide / 0.8% bisacrylamide solution,

50 μl (10%, w / v) ammonium persulfate and

- 5 μl of TEMED (N, N, N'-tetramethylenediamine); the Obergel corresponds

- 3.075 ml of distilled H ₂ O, - 0.625 ml of 0.5M Tris (tris (hydroxymethyl) aminomethane) /

HCl / 0.4% SDS solution (sodium dodecyl sulfate solution), pH 6.8,

0.75 ml of 30% acrylamide / 0.8% bisacrylamide solution,

50 μl (10%, w / v) ammonium persulfate and

- 5 μl of TEMED (N, N, N ', Λ /' - tetramethylenediamine)

The gels can be used as so-called minigels or in other apparatuses as conventional zymograms for the separation of the protein mixture. The person skilled in this apparatus and methods are known.

The sample application buffer may be any protein application buffer that is not allowed to receive reducing substances. After separation, the proteases are renatured (wash for 2x15 min in 2.5% Triton-X-100 in H ₂ O, then wash for ₂ x 15 min in 50 mM Tris / HCl pH 7.4, 5 mM CaCl ₂ ) and incubated for 24-48 hr in the last wash buffer. The renatured metallo proteases degrade the gelatin substrate in their immediate vicinity. If these zymograms are stained with a protein dye (Coomassie CAS No. 6104-59-2, Sigma-Aldrich Chemie GmbH, 89555 Steinheim - Germany), followed by decolorization according to standard protocols, the undegraded gelatin turns to homogenous blue in the gel (shown in the figure Figure 2 gray / black). Only in places where proteolytic activity is and the gelatin has been degraded, see clear, transparent bands, so-called "feeding bands" (in the figure Fig. 2 white on a gray / black background shown) Pattern and the molecular weight range determined by a protein marker (not shown here), it was possible to determine the type of enzyme. The method described here and variations thereof (eg Herron et al., J Biol Chem. 1986 261: 2814-8) are familiar to the expert.

In these experiments a clear binding of the MMP to the polyacrylate superabsorber was observed. As a result, depletion of MMP activity in the polyacrylate-treated wound fluid was noted. The binding and immobilization on the polyacrylate can be considered as high affinity since it was retained despite washing steps. The results are shown in Fig. 2, wherein in the first column (1) the MMP-2 and MMP-9 activity is shown in the untreated wound fluid. In the second and third columns, the activity of the wound fluid of the same wound treated with the above-mentioned polyacrylate is shown. In the third column (3), the activities of the MMP bound to the preactivated polyacrylate are plotted (the MMPs were removed from the polyacrylate by heating in the loading buffer) whereas in the second column (2) the activity of the MMP remaining in the supernatant is shown.

Finally, it was investigated whether polyacrylate can inhibit MMP activity even without direct contact. For this purpose, wound fluid with high MMP activity in a gelatin zymography gel was applied three times side by side with a marker and separated by electrophoresis. The gelatin zymogram was cut into three parts (each containing marker and wound fluid protein) and the first part was renatured and developed according to the standard procedure. The second part was treated identically except that the development buffer (50 mM Tris / HCl pH 7.4, 5 mM CaCl ₂ ) was added during development with Ringer's solution pre-activated polyacrylate (15 g

Ringer solution pre-activated polyacrylate / 30 ml development buffer). The third part was developed in the presence of 50 mM EDTA (final concentration) in development buffer. The three zymogram sections were stained together and the MMP activity was assessed on the clear bands, with the zymogram (1) the positive control, the activity of MMP-2 and MMP-9 without treatment with the above polyacrylate, zymogram (2) the activity the MMP from polyacrylate-treated wound exudate (the portion not bound by the polyacrylate) and the zymogram (3) shows the activity of MMPs from EDTA-treated wound fluid (see Figure 3). Figure 3 shows a clear inhibition of MMP activity in the Ringer's solution pre-activated polyacrylate gel portion. EDTA, a known inhibitor of MMP activity, has, as expected, completely blocked MMP activity in these experiments. The experiments clearly demonstrate that polyacrylates strongly and effectively reduce MMP activity directly by compartmentalization but also by inhibition over a distance and without direct contact. In the biological context, the ratio of granulation tissue degradation due to excessive MMP activity in chronic wounds and granulation tissue buildup in wound healing shifts in favor of construction and improved healing.

The inhibition of matrix metalloproteases resulting in the healing of chronic wounds has been demonstrated in clinical studies. In this study, three representative patients suffering from an open ulcer (ulcus cruris) were treated with a polyacrylate in the form of particles containing wound dressing (TenderWet activ - Fa Hartmann). The wounds are classified as chronic wounds. During the treatment, a dressing change was performed daily for a period of 4 weeks. After 24 hours the wound dressing was changed and the used bandages stored at -80 ° C for late examination. To examine each wound dressing, 300 mg polyacrylate particles were taken from each wound dressing, incubated with 225 μl sample buffer (see above) and boiled at 95 ^° C. for 5 minutes in a water bath. A serial dilution series of each sample (each wound dressing) was performed. By way of example, FIG. 4 shows the result of one of these investigations, wherein A represents a dilution series of a sample containing polyacrylate from a wound dressing used on the first day of treatment and B represents a dilution series of a sample containing polyacrylate from a wound dressing used on the twelfth day of treatment. The Coomassie staining indicates the total protein content of the sample and the zymography shows the matrix metalloprotease activity contained therein. The research methods Coommassie and Zymography are the same as described above. This in vivo study shows that the metalloproteases MMP-2 or MMP-9 are bound by polyacrylate. Due to the compartmentalization of the metalloproteases and the daily dressing change, the wounds were deprived of an excess of metallo-proteases hindering wound healing. The accompanying clinical finding of the treatment is that the treatment with a polyacrylate-containing wound dressing can transform a chronic wound into a normal wound healing course. Clinically, this was shown by a reduction in the defect volume (wound depth and / or wound size).

Claims

claims

1. Use of polyacrylates for the preparation of an agent for the inhibition of metallo proteases in chronic wounds.

2. Use of polyacrylates according to claim 1 for the preparation of a topical agent for the inhibition of metallo proteases in chronic wounds.

3. Use of polyacrylates according to claim 1 or 2 for the preparation of an agent for inhibition by compartmentalization of metallo-proteases in chronic

Wounds.

4. Use of polyacrylates according to at least one of the preceding claims for the preparation of an agent for the inhibition of metallo-proteases for tissue engineering and / or for the regulation of tissue structure in chronic wounds.

5. Use according to at least one of the preceding claims, characterized in that the polyacrylate is a crosslinked and / or crosslinked polyacrylate.

6. Use according to at least one of the preceding claims, characterized in that the polyacrylate i. comprises a homopolymer which consists of the monomers M1 and is crosslinked and / or cross-linked by means of a crosslinker and / or ii. a copolymer consisting of the monomers M1 and M3, wherein the monomer M1 is acrylic acid and / or a salt thereof and the monomer M3 is selected from the group of crosslinkers.

7. Use according to at least one of the preceding claims, characterized in that the polyacrylate is in the form of particles, fibers or a gel.

8. Use according to at least one of the preceding claims, characterized in that the polyacrylate is a polyacrylate superabsorber, which is in the form of particles, fibers or a gel.

9. Use according to at least one of the preceding claims, characterized in that the agent is a wound dressing.

10. Wound dressing comprising a metallo-protease inhibitor selected from the group of polyacrylates.

11. Wound dressing according to claim 10, characterized in that the polyacrylate binds and / or compartmentalizes metallo proteases.

12. Wound dressing according to claim 10 or 11, characterized in that the polyacrylate is a crosslinked and / or crosslinked polyacrylate.

13. Wound dressing according to at least one of the preceding claims 10 to 12, characterized in that the polyacrylate i. comprises a homopolymer which consists of the monomers M1 and is crosslinked and / or cross-linked by means of a crosslinker and / or ii. a copolymer consisting of the monomers M1 and M3, wherein the monomer M1 is acrylic acid and / or a salt thereof and the monomer M3 is selected from the group of crosslinkers.

14. Wound dressing according to at least one of the preceding claims 10 to 13, characterized in that the polyacrylate is in the form of particles, fibers or a gel.

15. Wound dressing according to at least one of the preceding claims 10 to 14, characterized in that the polyacrylate is a polyacrylate superabsorber, which is in the form of particles, fibers or a gel.

16. Wound dressing according to at least one of the preceding claims 10 to 15, characterized in that the wound dressing further comprises a carrier material, in particular a fiber material.

17. Use of a wound dressing according to any one of claims 10 to 16, for the preparation of an agent for the treatment of chronic wounds.