WO2019211483A1

WO2019211483A1 - A composition for reduction of scar formation

Info

Publication number: WO2019211483A1
Application number: PCT/EP2019/061496
Authority: WO
Inventors: Lars Adolfsson; Olle Larm
Original assignee: Bonoss Medical Aktiebolag
Priority date: 2018-05-04
Filing date: 2019-05-06
Publication date: 2019-11-07
Also published as: EP3787593A1

Abstract

The present invention relates to a composition comprising an ionic complex of chitosan and a negatively charged polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, for use in reducing scar formation during healing of a dermal wound in a mammalian subject. The invention further relates to a method of reducing scar formation during healing of a dermal wound in a mammalian subject, by applying topically a composition comprising an ionic complex of chitosan and a negatively charged polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, to the dermal wound.

Description

A COMPOSITION FOR REDUCTION OF SCAR FORMATION

Field of the invention

The present invention relates to a composition for use in the treatment of wounds to reduce scar formation. The present invention further involves a method of reducing scar formation using said composition.

Background

Scar formation is a natural part of the wound healing process following an injury to the skin. With the exception of very minor lesions, every wound (e.g., after accident, disease, or surgery) results in some degree of scarring.The appearance of the scar after the wound has healed is affected by several factors, such as the type of wound, the area of the body affected and the healing process itself. Wound healing consists mainly of the following six phases: i) The hemostatic inflammatory phase including the coagulation cascade, complement cascade and platelet activation to stop the bleeding. ii) The cellular inflammatory phase includes phagocytosis of debris and bacteria to prevent infection, followed by an increase of macrophages, which secrete mediators to regulate the healing process. An increase in fibroblasts occurs several weeks after the injury, resulting in collagen secretion and closure of large wounds. iii) The re-epithelialization proliferative phase involves remodeling of the wound with the aid of enzymes and fibroblasts. The level of moisture in the area limits the rate of this phase. iv) The neovascularization proliferative phase involves angiogenesis and the formation of granulation tissue. v) The collagen deposition proliferative phase involves the deposit of type III collagen. vi) The maturation or remodeling phase involves the deposit of type I collagen, which leads to a reduction in the size of the scar and increased strength of the skin.

A quick rate of wound healing is not necessarily associated with a low degree of scar formation. A wound can heal slowly, with a low degree of scar formation, or quickly, with a high degree of scar formation.

Minimizing scar formation is highly desirable in most situations, whether the wound is caused by injury, a surgical procedure or disease. The most straightforward way to prevent the formation of scars is obviously to limit injury to the skin. The most common methods for minimizing scar formation during the healing process include keeping the wounded area clean throughout the healing process, keeping the wound moist using a petroleum jelly or similar ointment, and covering the wounded area with a bandage to prevent further injury or entry of bacteria. There remains a need for improved methods for reducing scar formation.

With regard to wound ointments and similar agents for the treatment of dermal wounds, i.e. disorders in the form of wounds in the epidermis arising through accidents, burns, surgical incisions or without exterior influence, for example wounds in connection with metabolic disorders, such as diabetes or acne, so called chaps or the like, the market is flooded with a plethora of different products. These products typically contain as active constituents active substances of different kinds, for example antibiotics, antiinflammatory agents etc., whose action primarily is not directed to stimulate healing but instead are directed to act as desinfectants or in some other manner perform a cleaning action.

As an alternative healing agent so called heparin-chitosan complexes (HC- Complexes) have been contemplated. When aqueous solutions of heparin and chitosan are mixed, an ionic complex is immediately formed and precipitates. The pure heparin-chitosan complex, with enough material included for biological activity, can be prepared as beads, films, sponges, gels or suspensions. HC-Complexes and their use as wound healing agents are described e.g. in the published international patent application WO 96/02260 (Larm et ai).

Heparin is a commercially available polysaccharide, which is isolated from mammalian tissue (pig mucosa or beef lung). Since its discovery in 1916 by Jay McLean, heparin has been recognized for its blood anticoagulant properties. Heparin has been used clinically for more than 50 years as a blood anticoagulant and antithrombotic agent. Heparin is present in the basophilic granules of mast cells. Today, it is extensively used in the clinic as a blood anticoagulant and/or anti-inflammatory agent.

Heparin is a glycosaminoglycan. A glycosaminoglycan (GAG) is defined as the carbohydrate portion of a proteoglycan. The proteoglycans are built up by linear chains of disaccharide units containing uronic acid units linked to D- glucose amine. These chains are in turn linked to amino functions on a simple polypeptide. Examples include, dermatan sulfates, chondroitin sulfates, heparan sulfates and heparin.

Heparin GAGs are built up by alternating D-glucosamine and uronic acid residues (L-iduronic and D-glucuronic). They are linear, highly charged (sulfated) heterogeneous polysaccharides. The polysaccharide chitin is the second most abundant organic compound in nature after cellulose. Commercially, chitin is obtained mainly from crab and shrimp shells. Chitin has a regular structure and is composed of b-1 ,4-linked N-acetyl-D-glucosamine residues. Chitosan is positively charged and is obtained by partial N-deacetylation of chitin, e.g. via treatment of chitin with a strong base.

In vivo, chitosan is degraded by lysozyme and other glycosaminodases to mono- and oligomers. A chitosan which is rich in N-acetyl-D-glucosamine residues is degraded faster in vitro, and probably also in vivo, than a chitosan with a high proportion of D-glucosamine residues.

Description of the invention

It is an object of the present invention to provide a topical composition which, when administered to a dermal wound of a subject reduces scar formation during healing of the wound.

It is an object of the present invention to provide a topical pharmaceutical composition which, when administered to a dermal wound of a patient reduces scar formation during healing of the wound.

A further object of the present invention is to provide a topical pharmaceutical composition for reduces scar formation, which provides a sustained effect, thus reducing the required frequency of administration.

The above mentioned objects, as well as further objects which will become apparent to a person skilled in the art in view of the present disclosure, are achieved by the various aspects of the present invention.

In a first aspect thereof, the present invention provides a composition comprising an ionic complex of chitosan and a negatively charged polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, for use in reducing scar formation during healing of a dermal wound in a mammalian subject.

When solutions of negatively charged polysaccharide and chitosan are mixed, an ionic complex is immediately formed and precipitates. In such a complex, chitosan protects the negatively charged polysaccharide from enzymatic degradation in vivo, and that the half-life and beneficial effect of the negatively charged polysaccharide is thereby considerably prolonged. The compositions, methods and uses for reduction of scar formation disclosed herein are applicable to mammalian subjects in general and to human subjects in particular.

The composition of the invention may preferably be applied topically to a site in need of treatment to reduce scar formation, such as for example to a surface of surgical or burn wound.

In order to achieve the reduced scar formation, the composition should preferably be applied to the wound repeatedly during the wound healing process. The inventors have found that the reduction of scar formation is most significant when the composition is applied repeatedly during certain of the wound healing phases described above in the background section.

According to some embodiments, the composition is applied to the wound at least once per day, preferably at least two times per day, more preferably at least three times per day during at least the wound healing phases i-iii, and preferably at least the wound healing phases i-v.

According to some embodiments, the composition is applied to the wound at least once per day, preferably at least two times per day, more preferably at least three times per day, for a period of at least 3 days, preferably at least 5 days, more preferably at least 7 days, more preferably at least 10 days.

According to some embodiments, the composition is applied to the wound at least once per day, preferably at least two times per day, more preferably at least three times per day, for a period of at least 14 days, preferably at least 21 days, more preferably at least 28 days.

According to some embodiments, the composition is applied to the wound at least once per day for a period of at least 3 days, preferably at least 2 times per day for a period of at least 3 days, preferably at least 3 times per day for a period of at least 3 days.

According to some embodiments, the composition is applied to the wound at least once per day for a period of at least 5 days, preferably at least 2 times per day for a period of at least 5 days, preferably at least 3 times per day for a period of at least 5 days.

According to some embodiments, the composition is applied to the wound at least once per day for a period of at least 7 days, preferably at least 2 times per day for a period of at least 7 days, preferably at least 3 times per day for a period of at least 7 days.

According to some embodiments, the composition is applied to the wound at least once per day for a period of at least 10 days, preferably at least 2 times per day for a period of at least 10 days, preferably at least 3 times per day for a period of at least 10 days.

When applied to the surface of a dermal wound, the composition forms a physical barrier covering the surface onto which it is applied. The physical barrier, also referred to as a film, helps to prevent direct contact of the surface with objects or materials that would otherwise be brought into direct contact with potentially exposed nerve endings at the surface, thereby causing discomfort or pain to the patient.

The physical barrier, further helps prevent the introduction of bacteria into the wound, thereby reducing the risk of infection.

Preventing contact of the surface of a wound with objects or materials that would otherwise be brought into direct contact may further help accelerate the natural healing process since irritation and infection of the wound can be avoided.

The composition does not merely act as a physical barrier which protects the surface of the wound. The ionic complex between chitosan and a negatively charged polysaccharide, when applied to a mucous membrane, further provides a slow release of the negatively charged polysaccharide, e.g.

heparin, to the membrane. The negatively charged polysaccharide present in the ionic complex is gradually released when the chitosan degrades. Heparin has inherent antimicrobial, antiinflammatory, pain relieving and wound healing properties. A slow release of heparin at the surface of a dermal wound is thus highly beneficial to the patient.

The use of the composition for reducing scar formation is applicable both when the reduction of scar formation is medically motivated, in order to reduce or ameliorate pain or discomfort caused by excessive scar formation, and when the reduction of scar formation is not medically motivated, but entirely for cosmetic/aesthetic reasons.

Thus in some embodiments use of the composition for reducing scar formation is a method for medical treatment of a subject in need thereof. In some embodiments, the use of the composition for reducing scar formation is a cosmetic, non-medical method with the purpose of reducing visible signs of scar formation on the skin of the subject.

In the present context the expression "dermal wound" is intended to cover any disorder in association with dermis in the form of regular wounds arising through accident, surgical incision or in another manner by exterior influence, or wounds which have arisen without exterior influence, such as acne, so called chaps or other fissures in the dermis.

The composition may be used for reducing scar formation in any type of dermal wound. However, minimizing scar formation has been found to be of particular interest in certain patient groups. One patient group of particular interest is patients undergoing aesthetic surgery, where for obvious reasons it is desired to minimize any scar formation. Another patient group of particular interest is patients undergoing a caesarean section. Yet another patient group of particular interest is burn patients, since burns will often result in extensive scar formation.

In some embodiments the dermal wound is a surgical wound. In some embodiments the dermal wound is a surgical wound formed in connection with aesthetic surgery. In some embodiments the dermal wound is a surgical wound formed in connection with a caesarean section. In some embodiments the dermal wound is a burn wound.

In the present disclosure the ionic complex is formed or made up of positively charged chitosan, electrostatically bound to negatively charged

polysaccharide.

Chitosan is a positively charged linear 1 ,4-bound polysaccharide based on b- D-glucosamine residues. Chitosan is formed by partial N-deacetylation of chitin, a polymer comprised, e.g., in crab and shrimp shells. N-deacetylation may for example be performed by treatment of the chitin with a strong base or acid and results in the conversion of acetamido groups to amine groups. In vivo, chitosan is degraded by lysozyme and other glycosaminodases to mono- and oligomers. Chitosan which is rich in N- acetyl-D-glucosamine is degraded faster in vitro, and probably also in vivo, than a chitosan with a high proportion of D-glucosamine residues. The chitosan used with the

composition of the invention may generally have a degree of deacetylation in the range of 50 to 99 %. It has been found, however, that a chitosan having a degree of deacetylation in the range of about 80 to 95 % is especially useful. Thus, in an embodiment, the chitosan used in the composition has a degree of deacetylation in the range of 50 to 99 %, preferably in the range of 80 to 95 %.

Chitosan degrades under physiological conditions in mammals to

physiologically acceptable, non-harmful and readily metabolized

subcomponents, low molecular weight endogenous carbohydrates.

The negatively charged biologically active polysaccharide may preferably be a negatively charged polysaccharide selected from the group consisting of glycosaminoglycans (GAG:s) and semisynthetic biologically active

polysaccharides, such as dextran sulfates. In embodiments, the negatively charged biologically active polysaccharide is selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate. The term“a heparin” as used herein includes native heparin as well as modified forms of heparin, provided that the specific wound healing and scar formation reducing activity is maintained. Likewise, the terms“a heparan sulfate”,“a chondroitin sulfate”,“a dermatan sulfate” and “a dextran sulfate” as used herein include heparan sulfate, chondroitin sulfate, dermatan sulfate and dextran sulfate, respectively, as well as modified forms thereof, provided that the specific wound healing and scar formation reducing activity of the complex is maintained. In some embodiments, the negatively charged biologically active

polysaccharide is a GAG. In some embodiments, the negatively charged biologically active polysaccharide is selected from the group consisting of heparin and heparan sulfate. In a preferred embodiment, the negatively charged biologically active polysaccharide is heparin.

Heparin GAG:s are built up by alternating D-glucosamine and uronic acid residues (L-iduronic and D-glucuronic). They are highly charged (sulfated) heterogeneous polysaccharides. Heparin is commercially available on the market from several manufacturers. There are three forms of heparin, unfractionated heparin, low molecular weight heparin (LMWH), and ultra low molecular weight heparin (ULMWH). Also partially modified forms of heparin can be used, provided that the specific wound healing and scar formation reducing activity is maintained.

Examples of partially modified forms of heparin that may be used in the inventive composition include, but are not limited to, heparin oxidized by either sodium periodate or lead tetraacetate followed by reduction with sodium borohydride. Heparin oxidized by sodium periodate or lead

tetraacetate followed by reduction with sodium borohydride looses its specific anticoagulative properties, but maintains its specific wound healing and scar formation reducing activity. This characteristic may be of particular interest for patients suffering from or at risk of bleeding complications. Proteoglycans are a type of glycoprotein composed of a protein core with one or more covalently attached glycosaminoglycan (GAG) chains. Proteoglycans are found in connective tissues, and they contribute to the organization and the physical properties of the extracellular matrix. In some embodiments, the negatively charged biologically active polysaccharide is a proteoglycan. The negatively charged biologically active polysaccharides and proteoglycans also degrade under physiological conditions to non-harmful and readily metabolized sub-components, such as mono- and oligomeric carbohydrates.

When aqueous solutions of heparin and chitosan are mixed, an ionic complex is immediately formed and precipitates. The ionic complex of heparin and chitosan is useful for accelerating, stimulating and/or promoting the natural healing of dermal wounds.

The ionic complex of a negatively charged biologically active polysaccharide with chitosan is advantageous in that it may be biologically degradable and form non-toxic, environmentally friendly, naturally occurring and/or readily metabolized residues upon degradation.

In some embodiments, the composition comprises a heparin-chitosan ionic complex (HC-Complex). In such an ionic complex, the weight ratio of chitosan to heparin may be from about 1 :1 to 10:1 , such as from about 1 :1 to about 5:1. Examples of more specific intervals are from about 2:1 to about 4:1 , and from about 2:1 to about 3:1. In an embodiment, the weight ratio of chitosan to heparin is about 2.5:1. The weight ratio of chitosan to heparin in the ionic complex affects the physical characteristics of the complex, in particular its rheological properties and adhesiveness. Furthermore, having an excess of heparin would entail a risk of unwanted blood-anticoagulation, due to the interaction between heparin and plasma proteins in blood. The ranges given above are to be seen as guidelines for the skilled person to find the optimal ratio based on the particular situation and administration form in which the composition is to be used.

In a preferred embodiment, the number of positive charges contributed by said chitosan are in excess over the number of negative charges contributed by the heparin in the ionic complex. With an excess of chitosan in comparison to heparin on a charge basis, the composition is immobilized at the site of treatment. This is because in general, all cells are negatively charged. In an embodiment of the composition, the charge ratio of positive charges in said chitosan to negative charges in said negatively charged polysaccharide is in the range of from 10:1 to 10:8, preferably in the range of from 10:3 to 10:6, more preferably in the range of from 10:4 to 10:5. A charge ratio in the range of from 10:4 to 10:5 is especially advantageous, since it provides very good adhesion to the surface of a wound.

The composition may preferably be formulated for topical administration.

The composition may be in the form of a suspension of particles of the complex of chitosan and heparin in a liquid medium. The liquid medium may preferably be water or water based. Depending on the concentration of the complex in the composition, the composition may be referred to as a suspension or a gel.

The total concentration of the complex of chitosan and said negatively charged polysaccharide in said composition may preferably be selected such that a film of the complex is formed when the composition is applied to a surface.

The viscosity of the composition generally increases with increasing concentration of the complex of chitosan and said negatively charged polysaccharide. The total concentration of the complex of chitosan and said negatively charged polysaccharide in said composition may preferably be selected such that the viscosity of the composition is capable of forming a film on the wound upon application.

The composition can be presented in different physical forms, for example as powders, ointments, pastes, gels, emulsions or suspensions. The composition may be in the form of a suspension of particles of the complex in a liquid medium. In a particularly preferred embodiment the composition is in a suspension form suitable for application to a wound by spraying. The liquid medium may preferably be water or water based.

Depending on the concentration of the complex in the composition, the composition may be referred to as a suspension or a gel.

The viscosity of the composition generally increases with increasing concentration of the complex. The total concentration of the complex in said composition may preferably be selected such that a suitable viscosity for applying the composition to a wound is achieved.

The total concentration of the complex in the composition may generally be in the range of from 0.1 to 5 %, preferably in the range of from 0.1 to 3 % by weight, based on the total weight of the composition. A suitable viscosity may for example be obtained when the total concentration of the complex in the composition is in the range of from 0.1 to 5 %, based on the total weight of the composition.

It has been found that a suspension form of the complex, wherein the total concentration of complex is in the range of from 0.1 to 2 % by weight, for example in the range of 0.5 to 2 % by weight, based on the total weight of the composition is especially useful for application to a wound by spraying. Thus, in an embodiment, the total concentration of the complex in the composition is in the range of from 0.1 to 2 % by weight, based on the total weight of the composition, for example in the range of 0.5 to 2 % by weight, based on the total weight of the composition.

In a more specific embodiment thereof, the present invention provides a composition suitable for topical administration by spraying, comprising a suspension of a heparin/chitosan complex (HC-Complex), wherein said chitosan has a degree of deacetylation in the range of 80 to 95 %, the charge ratio of positive charges in the chitosan to negative charges in the heparin is in the range of from 10:4 to 10:5, and the total concentration of the HC- Complex in said composition is in the range of from 0.1 to 2 % by weight, based on the total weight of the composition. Such a composition has been found by the inventors to provide a very beneficial combination of properties for treating dermal wounds in mammals. The beneficial combination of properties includes good adhesion to wound surfaces and good film forming properties allowing the formation of a physical barrier over the wound surface. The film is not only sufficiently durable and adhesive, but also provides a controlled release of heparin to the site under treatment, and is biologically degraded to physiologically acceptable degradation products.

In an embodiment, the composition consists solely of a suspension of particles of HC-Complex in water or a physiologically acceptable buffer, without further additives. This embodiment may be advantageous to reduce discomfort experienced by patient during application of the composition to a wound, which may be irritated by additional components in the composition. The complex of chitosan and heparin itself has been found not to cause pain or discomfort even in patients experiencing highly painful sores or ulcers.

In some embodiments, the composition may further comprise various pharmaceutically acceptable excipients and additives. Examples of such excipients and additives include, but are not limited to buffers, surfactants, viscosity adjusting agents, flavoring agents, antifoaming agents and

antimicrobial agents, such as anti-fungal agents and anti-bacterial agents.

In an embodiment, the composition further comprises an antimicrobial agent, such as an antifungal agent or an antibacterial agent. Examples of such agents include, but are not limited to, methyl paraben and propyl paraben.

In an embodiment, the composition further comprises an antifoaming agent. The composition may also comprise additional pharmaceutically active agents, that may further improve the healing or pain relieving effects of the composition. Examples of such additional pharmaceutically active agents include, but are not limited to, analgesic agents, anti-inflammatory agents and antibiotics.

The composition may also comprise a dermally acceptable carrier or excipient. In some embodiments, the matrix is an aqueous matrix, the carrier or the excipient containing a viscosity increasing polysaccharide, which can be selected from hemicelluloses, arabinoxylanes and glucomannanes, plant gums, for example guar gum, locust bean gum, celluloses and derivatives thereof, for example methyl cellulose, ethyl cellulose, hydroxyethyl cellulose, starch and starch derivatives, for example hydroxyethyl starch or crosslinked starch, xanthan gum, curdlan, pullulan, or dextran. Also algae

polysaccharides can be used as a constituent in the carrier or excipient.

A preferred viscosity increasing polysaccharide is a cellulose derivative, for example hydroxyethyl cellulose.

In a second aspect thereof, the present invention provides a method of reducing scar formation during healing of a dermal wound in a mammalian subject, by applying topically a composition comprising an ionic complex of chitosan and a negatively charged polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, to the dermal wound.

The term "mammalian", as used herein, includes humans unless otherwise specifically stated. The term "topical", as used herein, generally means the application of a pharmaceutical composition to body surfaces such as the skin or mucous membranes. The composition is typically applied topically to the site in need of treatment. The composition may preferably be applied in an amount sufficient to form a film, which covers the surface to be treated or protected. The composition may be applied once or a suitable number of times during administration, such that a single layer film or a film having more than one layer is formed.

In order to achieve the reduced scar formation, the composition should preferably be applied to the wound repeatedly during the wound healing process.

According to some embodiments, the composition is applied to the wound at least once per day for a period of at least 5 days, preferably at least 2 times per day for a period of at least 5 days, preferably at least 3 times per day for a period of at least 5 days. According to some embodiments, the composition is applied to the wound at least once per day for a period of at least 7 days, preferably at least 2 times per day for a period of at least 7 days, preferably at least 3 times per day for a period of at least 7 days.

The composition used in the method of the second aspect of the invention may be further defined as described above in respect of the first aspect of the invention.

The composition is suitably applied in the form of a powder, an ointment, a paste, a gel, an emulsion, or a suspension. Preferably, the composition is applied by spraying of the composition in suspension form. The type of patient and wound to be treated may be further defined as described above in respect of the first aspect of the invention.

The cause of the wound may be further defined as described above in respect of the first aspect of the invention.

In a third aspect thereof, the present invention further provides the use of a composition comprising an ionic complex of chitosan and a negatively charged polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, in the manufacture of a medicament for reducing scar formation during healing of a dermal wound in a mammalian subject. The use according to the third aspect of the invention may be further defined as described above in respect of the first aspect of the invention.

In a fourth aspect thereof, the present invention provides a method for reducing scar formation during healing of a dermal wound in a mammalian subject, by applying topically a composition comprising an ionic complex of chitosan and a negatively charged polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, to the dermal wound.

The method for reducing scar formation is applicable both when the reduction of scar formation is medically motivated, in order to reduce or ameliorate pain or discomfort caused by excessive scar formation, and when the reduction of scar formation is not medically motivated, but entirely for cosmetic/aesthetic reasons.

Thus in some embodiments the method for reducing scar formation is a method for medical treatment of a subject in need thereof.

In some embodiments, the method for reducing scar formation is a cosmetic, non-medical method with the purpose of reducing visible signs of scar formation on the skin of the subject.

In some embodiments the dermal wound is a surgical wound formed in connection with aesthetic surgery. In some embodiments the dermal wound is a surgical wound formed in connection with a caesarean section. In some embodiments the dermal wound is a burn wound. In a preferred embodiment, the dermal wound is a surgical wound formed in connection with aesthetic surgery.

The composition used in the fourth aspect of the invention may be further defined as described above in respect of the first aspect of the invention. In order to achieve the reduced scar formation, the composition should preferably be applied to the wound repeatedly during the wound healing process.

The composition used in the method of the fourth aspect of the invention may be further defined as described above in respect of the first aspect of the invention.

In a more specific embodiment thereof, method according to the fourth aspect comprises applying to the dermal wound by spraying, a suspension of a heparin/chitosan complex (HC-Complex), wherein said chitosan has a degree of deacetylation in the range of 80 to 95 %, the charge ratio of positive charges in the chitosan to negative charges in the heparin is in the range of from 10:4 to 10:5, and the total concentration of the HC-Complex in said composition is in the range of from 0.1 to 2 % by weight, based on the total weight of the composition, wherein the composition is applied to the dermal wound at least 3 times per day for a period of at least 7 days.

All features of all embodiments of all aspects of the invention can be used in any possible combination thereof, provided that such combination is not demonstrably unfeasible as determined without undue experimentation by a person having ordinary skill in the art.

The invention is not restricted to the embodiments presented above and has a scope only limited by the appended claims. EXAMPLES

EXAMPLE 1 - Preparation of low molecular chitosan by nitrous acid degradation

Chitosan (50.0 g, Primex TM 3728) was suspended in purified water (850 g) and pH was adjusted to 1.4 - 1.6 with 2 M hydrochloric acid (VWR) during mixing. When all chitosan was dissolved, a solution of sodium nitrite (167 mg, Sigma Aldrich) in purified water (10 mL) was added.

After 1 hour stirring at room temperature, pH was adjusted to 5.5 with 1 M sodium hydroxide (VWR). A solution of sodium borohydride (0.33 g, Sigma Aldrich) in purified water (10 mL) was slowly added. Stirring was continued during another 30 minutes.

Thereafter pH was adjusted to 3.0 with 2 M hydrochloric acid (VWR) and then after another 30 minutes pH was adjusted to 5.0 with 1 M sodium hydroxide (VWR).

The degraded chitosan solution was poured into a saturated sodium chloride solution (1 liter, Sigma Aldrich) and pH was adjusted to 11 - 12 with 20% sodium hydroxide solution (Scharlau) by careful mixing.

The chitosan suspension was collected on a glass filter (no. 1 ) and was washed with purified water, about 10 liter, to neutral. A few drops of the filtrate were added to a 0.1 M silver nitrate solution (1 mL, Scharlau) and a clear solution without any precipitation showed that all sodium chloride was removed.

The chitosan was added to purified water (1.3 liter) and 2 M hydrochloric acid (VWR) was added during stirring until all chitosan had dissolved and pH was 5.0. The solution was first filtered through a 5 mhh filter and then through a 0.8/0.2 miti filter. The filtered solution of chitosan hydrochloride was freeze dried.

EXAMPLE 2 - Preparation of the heparin-chitosan complex

Heparin Solution

Sodium heparin (1 .20 g, SPL) was dissolved during stirring in purified water (48.8 g) until a clear solution was obtained. The heparin solution was sterile filtered through a 0.2 pm filter.

Chitosan Solution

Chitosan hydrochloride salt (3.75 g), prepared as described in Example 1 , was dissolved in purified water (546.3 g). Mixing was continued until all chitosan had been dissolved. The pH was adjusted to 5.1 with 1 M sodium hydroxide (VWR). The solution was sterile filtered through a 0.2 pm filter.

Sterile HC-Complex

The solutions of sterile filtered heparin and chitosan solutions were mixed together under sterile conditions. Mixing was continued until a smooth complex was formed.

The heparin-chitosan complex prepared was subjected to a sterility control. The analysis was performed in accordance with European Pharmacopoeia (Ph. Eur.) 9th edition, section 2.6.1 . The results showed that the complex fulfilled the requirements for a sterile product. EXAMPLE 3 - Wound healing

The patient (female, age 71 ) suffered from a lower lip tissue wound, probably resulting from sun burn. The wound had not healed even after 4 months of treatment with Aquaphor as well as sun block when exposed to sun.

Application of the HC-Complex formulation prepared according to Example 2 was applied, mainly in spray form, 8-12 times daily on day 1 through 4 of treatment, and 4-5 times daily on day 5 through 10 of treatment.

The patient commented that the spray was easy to apply. And that the burning, painful, feeling went away almost immediately after the first application. After one day of treatment a scab had formed, which fell off one day later. On day 2 the skin was still very red but on the forth day the lip looked and felt quite normal.

After 3 months, no redness or scarring was visible, which would otherwise have been expected from this type of wound.

EXAMPLE 4 - Clinical study of reduction of scar formation Patients undergo breast augmentation performed with standard surgical treatment. Incisions are trans-axillary (underarm area), peri-areolar (from about the 4 to 8 o'clock position of the areolas), or inframammary fold (in or just above the crease below the breasts). Directly after or in connection with surgery, the incisions wounds are on one side of the body sprayed with an aqueous suspension of the HC-Complex prepared according to Example 2. The other side is sprayed with

physiological saline solution. The sides are selected at random.

The treatment is repeated three times daily for five days.

Pain relief, time to wound healing and scar formation are monitored. Wound healing and scar formation are documented by photography for 3 months.

Claims

1. A composition comprising an ionic complex of chitosan and a negatively charged polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, for use in reducing scar formation during healing of a dermal wound in a mammalian subject.

2. A composition for use according to claim 1 , wherein said chitosan has a degree of deacetylation in the range of 50 to 99 %, preferably in the range of 80 to 95 %.

3. A composition for use according to any one of the preceding claims, wherein said negatively charged polysaccharide is heparin.

4. A composition for use according to any one of the preceding claims, wherein in said composition the charge ratio of positive charges in said chitosan to negative charges in said negatively charged polysaccharide is in the range of from 10:1 to 10:8, preferably in the range of from 10:3 to 10:6, more preferably in the range of from 10:4 to 10:5.

5. A composition for use according to any one of the preceding claims, wherein the total concentration of said chitosan and said negatively charged polysaccharide in said composition is in the range of from 0.1 to 5 % by weight, preferably in the range of from 0.1 to 3 % by weight, more preferably in the range of from 0.1 to 2 % by weight, based on the total weight of the composition.

6. A composition for use according to claim 5, wherein the total concentration of said chitosan and said negatively charged polysaccharide in said composition is in the range of from 0.5 to 5 % by weight, preferably in the range of from 1 to 3 % by weight, based on the total weight of the

composition.

7. A composition for use according to any one of the preceding claims, comprising the complex of heparin and chitosan in combination with a demnally acceptable carrier or excipient.

8. A composition for use according to any one of the preceding claims, wherein the composition comprises an antimicrobial agent.

9. A composition for use according to any one of the preceding claims, wherein the composition comprises an antifoaming agent.

10. A composition for use according to any one of claims 1 -6, consisting solely of the complex of heparin and chitosan in water or a physiologically acceptable buffer, without further additives.

11. A composition for use according to any one of the preceding claims, wherein the composition is in the form of a spray.

12. A composition for use according to any one of the preceding claims, wherein the composition is applied to the wound at least once per day for a period of at least 3 days, preferably at least 2 times per day for a period of at least 3 days, preferably at least 3 times per day for a period of at least 3 days.

13. A composition for use according to any one of the preceding claims, wherein the composition is applied to the wound at least once per day for a period of at least 5 days, preferably at least 2 times per day for a period of at least 5 days, preferably at least 3 times per day for a period of at least 5 days.

14. A composition for use according to any one of the preceding claims, wherein said dermal wound is a surgical wound.

15. A composition for use according to any one of the preceding claims, wherein said dermal wound is a surgical wound formed in connection with aesthetic surgery.

16. A composition for use according to any one of the preceding claims, wherein said dermal wound is a surgical wound formed in connection with a caesarean section.

17. A composition for use according to any one of the preceding claims, wherein said dermal wound is a burn wound.

18. A method of reducing scar formation during healing of a dermal wound in a mammalian subject, by applying topically a composition

comprising an ionic complex of chitosan and a negatively charged

polysaccharide, selected from the group consisting of a heparin, a heparan sulfate, a chondroitin sulfate, a dermatan sulfate and a dextran sulfate, to the dermal wound.

19. A method according to claim 18, wherein the composition is a cosmetic, non-medical method.

20. A method according to any one of claims 18-19, wherein the composition is applied to the wound at least once per day for a period of at least 3 days, preferably at least 2 times per day for a period of at least 3 days, preferably at least 3 times per day for a period of at least 3 days.

21. A method according to claim 20, wherein the composition is applied to the dermal wound at least 3 times per day for a period of at least 7 days.

22. A method according to any one of claims 18-21 , wherein the composition is applied to the wound at least once per day for a period of at least 5 days, preferably at least 2 times per day for a period of at least 5 days, preferably at least 3 times per day for a period of at least 5 days.

23. A method according to any one of claims 18-22, wherein said composition is further defined in accordance with any one of claims 2-11.

24. A method according to any one of claims 18-23, wherein said dermal wound is further defined in accordance with any one of claims 14-17.

25. A method according to any one of claims 24, wherein said dermal wound is a surgical wound formed in connection with aesthetic surgery.