WO1991016058A1

WO1991016058A1 - Composition and method for topical treatment of damaged or diseased tissue

Info

Publication number: WO1991016058A1
Application number: PCT/US1991/002912
Authority: WO
Inventors: R. Martin Emanuele; Robert L. Hunter
Original assignee: Cytrx Corporation
Priority date: 1990-04-26
Filing date: 1991-04-25
Publication date: 1991-10-31
Also published as: JPH05506861A; PT97504A; IE911403A1; CN1058341A; IL97976A0; CA2081340A1; BR9106377A; EP0526578A4; EP0526578A1; AU647484B2; AU7869291A; JPH0776175B2

Abstract

In accordance with the present invention, a composition and method is provided for topically treating damaged or diseaded tissue. The present invention comprises an aqueous gel or a pharmaceutically acceptable topical carrier containing an effective amount of an acid mucopolysaccharide. The composition containing the aqueous gel is liquid at room temperature and a gel at body temperature, thereby providing a composition that can be easily applied to the damaged or diseaded tissue. After topical application to the tissue, the composition according to the present invention provides a covering over the tissue that protects the tissue and accelerates healing.

Description

COMPOSITION AND METHOD FOR TOPICAL TREATMENT OF DAMAGED OR DISEASED TISSUE

Technical Field

The present invention relates to a method and composition for the topical treatment of damaged or diseased tissue. More particularly, the present invention is a composition and method for effectively treating damaged or diseased skin and tissue and promoting the healing of the damaged or diseased skin and tissue.

Background of the Invention

As used herein, the term "topical treatment" means the treatment of the surface of the body of an animal or human or an internal surface of the human or animal that has been externalized either during surgery or by traumatic injury. The term "damaged tissue" means tissue damaged by surgery, ischemia, burns, toxins, trauma or other noxious insult. The term "diseased tissue" means tissue that is infected with a microorganism such as a yeast, bacteria or virus or a combination of organisms. The infection can be independent of or as a result of damage to the tissue. The term "burns" means tissue injuries caused by thermal, electrical, ionizing or solar radiation or chemical agents.

Tissue damage caused by either surgery, ischemia, burns, toxins, trauma or other noxious insults illicits similar physiological responses. Injury to the skin, resulting in destruction of the epidermis and on occasion dermal layers of the organ, exposes the underlying tissue to the external environment and causes blood loss, plasma loss, and an increased risk of infection. Tissue damage caused by a variety of topical insults results in similar physiological responses and requires the same care and precautions. Bums are tissue injuries caused by thermal, electrical, ionizing or solar radiation, or chemical agents. The common mechanism of burns is denaturation of protein, resulting in cell injury or death. The effect is in accord with the type, duration, and intensity of action of the particular agent causing the burn. Because such agents impinge on the organism from its environment, tissues in direct contact with this environment, e.g. skin, are damaged most quickly. In electrical burns, because most of the resistance to electric current is at the point of the contact of the skin with the conductor, these burns usually involve skin and subjacent tissues. These burns may be of almost any size and depth. Resultant slough is usually greater than the original regions would indicate. Chemical burns may be due to strong acids and akalies, phenols, cresol, mustard gas, lewisite, phosphorus and the like. All produce areas of necrosis which may extend slowly for several hours. Radiation burns may result from ionizing (radioactivity) or, more commonly, from solar radiation. Bums of this type result in skin changes ranging from mild erythema to more pronounced swelling with blistering or ulceration.

The first local evidence of a bum is dilation of capillaries and small vessels with increased capillary permeability. The resultant plasma loss under the epidermis produces edema. Later, evidence of cellular injury can be seen histologically as swollen or pyknotic nuclei with coagulation of cytoplasm. In addition, collagen fibriles lose their distinctness.

In spontaneous healing, dead tissue is sloughed off as new epithelium begins to cover the injured area. In superficial burns, regeneration occurs rapidly from uninjured epidermal elements, hair, follicles, and sweat glands; little scarring results unless infection occurs. With deep burns (destruction of the epidermis and much of the dermis), reepithelization starts from the edges of the wound or from the remains of scattered integumentary organ. This process is slow, and granulation tissue forms in excess amounts before being covered by the epithelium. Unless treated as soon as possible by skin grafting, such wounds generally contract, and disfiguring or disabling scars result.

Systemic effects of burns may quickly endanger the patient's life. Primary or neurogenic shock consists of sudden collapse from generalized vasodilation, presumably on a reflex basis from pain, fright, and anxiety; it is rarely fatal. Secondary shock develops insidiously, or rapidly if the burn is extensive. Increased capillary permeability resulting from damage to vessel walls allows large amounts of fluid to exude into the wound area and from the burn surface. This fluid consists of water, plasma, crystalloides, and about two-thirds of the plasma protein. The fluid loss is at the expense of the plasma.

When the epidermis is broken, evasion of bacteria may occur.

Dead tissue, warmth, and moisture provide ideal conditions for bacterial growth. The type of pathogen depends partly on the location of the burn; nasalpharyngeal inhabitants such as streptococci and staphylococci predominate in upper body burns, while cold form bacteria and clostridia are often important in lower body burns. What is needed is a method of covering the burn and protecting the burned area from the environment Burns are usually classified into three types. In first degree burns, damage is limited to the outer layer of the epidermis with erythema, increased warmth, tenderness, and pain. Edema, but not vesiculatipn,_, usually occurs. In second degree bums, damage extends through the epidermis and involves the dermis, but not sufficiently to interfere with rapid regeneration of the epithelium. Vesicles, blebes, or bullae form. In third degree or full thickness burns, destruction of both the epidermis and dermis is seen. Vesiculation is often absent and severe pain is unusual after the acute initial pain, because of the destruction of the nerve endings. The surface may be charred, coagulated, or white and lifeless (as from skulls), and sometimes is insensitive to pin prick. A burn may show varying degrees of damage in different areas. Frequently, it is difficult to distinguish between second and third degree bums until areas of third degree depth demarcate.

Conventional therapy for all burns includes relief of pain, strict asepsis, and care of the wound, prevention or relief of shock, control of infection, correction of enema, and maintenance of nutrition. Local treatment of chemical burns differs according to the causative agent. Following treatment, these wounds should be cared for similarly to thermal bu s of comparable size and extent.

Conventional treatment for first and second degree burns involves cleaning the burned area gently with cold or warm water and hexachlorophene or green soap solution. The burned areas are then rinsed with sterile isotonic saline. All dirt, grease, and other contaminants on broken epidermis should be removed. Unbroken blebes are usually left intact. For open or exposure treatment, the wounds are allowed to dry. For the closed type of treatment, sterile fine mesh absorbent gauze is applied. For burned limbs, a pressure dressing is often used consisting of additional layers of sterile absorbent gauze covered by abdominal pads of mechanics' waste, all enclosed by firmly applied absorbent gauze or by elastic bandages. If the bums are uncomplicated by signs of infection, the pressure dressings are left in place until the burn has healed. The burn will heal better if immobilized and elevated because these measures decrease the flow of lymph and limit the spread of infection.

For third degree burns, which are characterized by deep destruction of the skin so that the reepithelization must take place largely or entirely from the edges of the wound, conventional treatment is identical with the previous outline. However, to avoid slow healing and disfiguring scars, burns of more than a few square centimeters require skin grafting. In small areas of third degree depth, this can be done within a few days of the injury by excision of the burned area. This is followed by application of split thickness grafts. For bu s involving greater than 15% of the body surface area, the fluid requirements of the patient need to be immediately addressed. The surface wounds of patients with extensive third degree bums are usually treated with an 0.5% solution of silver nitrate or mafenide cream.

Some physicians, however, prefer open treatment. Its advantages are simplicity, better control of hyperpyrexia, and easier ambulation.

Special hospital ventilation is required to prevent bacterial contamination. This therapy is less suitable for bums of the hand and foot than for circumferential burns of the trunk. Open treatment does not reduce pain or loss of exudate. It probably should be regarded at present as a complimentary form of local therapy well suited to the treatment of bums of the face, neck, side of the trunk, or proximal part of the extremity. In general, after cleansing and debridement of the burned area, the patient is placed on surgically clean or sterile sheets in a room of proper temperature and humidity with a cradle over the bed. In 48 to 72 hours, the protein-rich exudate forms a crust over the burned areas. Fluid, antibody, electrolyte and colloid therapy, grafting, and other forms of treatment are conducted as in closed therapy.

The treatment of burns using polyoxyethylene- polyoxypropylene block copolymers is described in U.S. Patent No. 4,879,109. This patent describes use of the block copolymers either alone or in combination with other compounds, to increase the flow of biologic fluids to damaged tissue. To be effective, the composition must be introduced into the circulatory system because the copolymer works by interfering with the pathologic hydrophobic interactions in blood and other biological fluids.

The treatment of bums using an aqueous gel with silver ions therein is described in U.S. Patent No. 3,689,575. This patent describes a polyoxyethylene-polyoxypropylene block copolymer which has the unique property of being liquid at low temperatures and forming a gel at higher temperatures. Silver ions are added to the gel and the composition is used to treat bums. The composition described in the '575 patent utilizes conventional silver ion treatment of burns by dissolving the silver ions in a copolymer matrix.

The preparation of an aqueous gel is described in U.S. Patent Nos. 3,867,533; 3,748,276 and 3,740,421, all of which are incorporated herein by reference. U.S Patent No. 3,867,533 describes the addition of numerous water-insoluble pharmaceutically or cosmetically active organic ingredients. U.S. Patent No. 3,748,276 describes the addition of conventional antimicrobial agents to the gel. However, none of the aforementioned patents disclose the admixture of acid mucopolysaccharides with the copolymer for the purpose of preparing a therapeutically effective topical composition.

The treatment of soft tissue wounds using an aqueous mixture of fibrillar collagen, heparin and undegranulated platelets or platelet releasate is described in U.S. Patent No. 4,760,131. This patent indicates that collagen/glycosaminoglycan suspensions have only a minimal effect on the rate or extent of wound healing. For the composition to be effective in treating soft tissue wounds, a platelet fraction must be added. In several articles by Saliba, et al., humans with burns were treated with a combination of intravenous, subcutaneous and topical application of large doses of heparin. Because the heparin was dissolved in an aqueous liquid, the liquid had to be applied constantly either by spraying through a needle onto the burn area or by dripping the solution onto the burn. Blisters had to be ruptured and the heparin injected into the blister by needle. In addition, the reports disclose that the topical treatment must be accompanied by the administration of large doses of intravenous heparin for several days. The effectiveness of the heparin therapy is limited in that the large doses required for therapeutic effect result in compromise of the blood coagulation system and an increased risk for hemorrhage. (See Saliba, M.J., "Heparin Efficacy in Bums:

II Human thermal bum treatment with large doses of topical and parenteral heparin" Clin. Aviat. and Aero. Med., 41 (11), pgs. 1302-1306 (1970); Saliba, M.J., et al, Large bums in humans, treatment with heparin," JAMA, 225(3), pgs. 261-269 (1973))

Articles by Rudolph and Tauschel disclose that heparin, in combination with allantoin and dexpanthenol, is effective in reducing the anti- inflammatory reactions of experimentally induced erythemas. The authors state that comparative testing of a heparin cream without allantoin and dexpanthenol had shown that heparin in cream does not differ fundamentally in effect from the cream base without heparin. (See Tauschel, H.D., et al., "Investigations of the Percutaneous Activity of a Combination of Heparin, Allantoin and

Dexpanthenol in a Specific Ointment Base. Anti-Inflammatory Effect of UV Erythema in the Guinea Pig," Arzneim.-Forxch./Drug Res. 32 (H)(9), pgs. 1096-1100 (1982) and Rudolph, C, et al., "Investigations into the Percutaneous Activity of a Combination of Heparin, Allantoin, Dexpanthenol in a Specific Ointment Base. Antiallergic/Anti-Inflammatory Effect in the PCA

Test in Rats," Arzeim.-Forsch.IDmg Res. 34 (11)12), pgs. 1766-8 (1984))

In a report by Raake, (Raake, W., "Comparison of the Anti- inflammatory Effect of Mucopolysaccharide Ointments with Ointments Containing Heparin in the UV Erythema Test," Arzenim.-ForschlDrug Res. 34 (I)(4), pgs. 449-451 (1984)) the anti-inflammatory effects of heparin are investigated. The Raake reference did not disclose any studies wherein heparin was used to treat wounds nor did the reference disclose the use of heparin with poloxamers.

What is needed is a treatment for bums and other topical injuries that will cover the damaged area and protect the damaged area from the environment. The treatment should inhibit inflammatory processes, provide relief from pain and accelerate the healing process without compromise of endogenous physiologic mechanisms such as blood coagulation. In addition, the treatment should be easily applied and should be effective over long periods of time.

Summary of the Invention

The present invention provides a composition and method for the topical treatment of damaged tissue. The composition of the present invention comprises a solution of polyoxyethylene-polyoxypropylene block copolymer and an effective concentration of an acid mucopolysaccharide. The polyoxyethylene-polyoxypropylene block copolymer has the following general formula:

HO(C H O) (C H O) (C H O) H 2 4 V 3 6 a 2 4 b wherein a is an integer such that the hydrophobe represented by C3H6O has a molecular weight of at least 2250 daltons and b is an integer such that the hydrophile portion represented by C2H4O constitutes from about 10 to

90 percent by weight of the copolymer, with a preferable range of hydrophile portion of between 50% and 90%.

According to the present invention, the copolymer is and an acid mucopolysaccharide are dissolved in an aqueous solution. It is preferable that the acid mucopolysaccharide have little or no anticoagulant activity or to be of sufficient molecular size or charge so as not to be absorbed in the ciruculation to avoid the toxicity exhibited by many acid mucopolysaccharides. The composition is then topically applied to damaged tissue. This allows manifestation of the therapeutic effects of the acid mucopolysaccharide at concentrations producing minimal or no compromise to the blood coagulation system. Because the preferred copolymer is a liquid at room temperature or below, and is a gel at body temperature, the aqueous gel composition forms a protective gel over the damaged tissue. The solution which comprises the present invention unexpectedly accelerates healing of the damaged or diseased tissue while at the same time keeping the damaged tissue moist and protected from contamination by microorganisms. It is believed that the copolymer forms a matrix which slows the diffusion of mucopolysaccharide to the damaged or diseased tissue. Thus, the gelled copolymer with the acid mucopolysaccharide admixed therein provides a continuous and constant bathing of the area with the mucopolysaccharide over a long period of time. Because the copolymer gel is an aqueous gel, there is also the added advantage of the gel keeping the diseased or damaged tissue moist without the inconvenience of having to bandage the tissue.

Other pharmaceutically active agents can be added to the copolymer/acid mucopolysaccharide admixture as deemed necessary. For example, conventional antimicrobial agents could be added to the copolymer/glycosaminoglycan admixture to protect the damaged tissue from infection. Growth factors, such as human growth hormone, tissue derived growth factor, epidermal growth factor, platelet derived growth factor, fibroblast growth factor, and/or nerve growth factor may be added to the copolymer/acid mucopolysaccharide admixture to promote the regrowth of healthy tissue. This would be particularly important in treating bums.

The composition of the present invention further comprises a pharmaceutically acceptable topical carrier admixed with an acid mucopolysaccharide. The pharmaceutically acceptable topical carrier may be any suitable, commercially available ointment, cerate or salve. Other pharmaceutically active agents can be added to the carrier. The composition is administered topically to damaged or diseased tissue. Accordingly, it is an object of the present invention to provide a composition and method that is effective in topically treating damaged or diseased tissue.

It is another object of the present invention to provide a composition and method that is effective in treating bums. It is another object of the present invention to provide a composition and method that is effective in treating tissue damaged by trauma.

It is yet another object of the present invention to provide a composition and method that accelerates the healing process.

It is another object of the present invention to provide a composition and method that forms an aqueous gel over the damaged or diseased tissue thereby keeping the damaged or diseased tissue moist.

It is another object of the present invention to provide a composition and method that contains an anesthetic and will inhibit pain when applied to the damaged or diseased tissue. It is another object of the present invention to provide a composition and method that will cover damaged or diseased tissue with a protective layer that is transparent thereby allowing observation of the damaged area during the healing process.

These and other objects, features and advantages of the present invention will become apparent after a review of the following detailed description of the disclosed embodiments and the appended claims. Detailed Description of the Disclosed Embodiments

In accordance with the present invention, an aqueous gel composition is provided that is effective in topically treating damaged or diseased tissue. The aqueous gel composition of the present invention comprises an admixture of an acid mucopolysaccharide and a polyoxyethylene- polyoxypropylene block copolymer that has the physical property of being a liquid at room temperature or below and a gel at body temperature.

The polyoxyethylene-polyoxypropylene block copolymer has the following general formula:

HO(C H O) (C H O) (C H O) H 2 4 V 3 6 2 4 'b wherein a is an integer such that the hydrophobe represented by C3H6O has a molecular weight of at least 2250 daltons and b is an integer such that the hydrophile portion represented by C2H4O constitutes from about 50 to 90 percent by weight of the copolymer. The polyoxyethylene-polyoxypropylene block copolymer preferably has a hydrophobe represented by C3H6O with a molecular weight between approximately 2250 and 6000 daltons with the most preferable molecular weight of the C3H6O of approximately 4000 daltons.

The surface active copolymer blocks are formed by condensation of ethylene oxide and propylene oxide at elevated temperature and pressure in the presence of a basic catalyst. There is some statistical variation in the number of monomer units which combine to form a polymer chain in each copolymer. The molecular weights given are approximations of the average weight of copolymer molecule in each preparation. It is to be understood that the blocks of propylene oxide and ethylene oxide do not have to be pure. Small amounts of other materials can be admixed so long as the overall physical chemical properties are not substantially changed. A more detailed discussion of the preparation of these products is found in U.S. Patent No. 2,674,619, which is incorporated herein by reference. Preparation of the copolymer portion of the present invention is described in U. S. Patent Nos. 3,925,241 and 3,867,533, both of which are incorporated herein by reference. Illustrative block copolymers of the following general formula:

HO(C H O) (C H O) (C H O) H 2 4 3 6 a 2 4 'b which may be employed in the preparation of the gels of the present invention are presented in Table I. Table I

Hy rop obe ase s t e ( 3 6 )_a por on o the copolymer molecule.

** Hydrophile base is the (C2H .O)b portion of the copolymer molecule.

Not all of the block copolymers of the formula: HO(C H O) (C H O) (C H O) H

2 4 b 3 6 a 2 4 b may be employed in the present invention. Because of the nature of aqueous solutions of these block copolymers, three variables affect the formation of the gels. Therefore, it is necessary to recognize certain minima for the three variables. These variables are:

( 1 ) the weight percent concentration of block copolymers in the gel ;

(2) the molecular weight of the hydrophobe portion (C3H6θ)a; and

(3) the weight percent of the hydrophile portion (C2H4θ)b of the copolymer.

These minima define a minimum weight percent concentration of the block copolymer with a specific hydrophobe having a minimum weight percent of ethylene oxide that is necessary to form a gel. Thus, at the minimum concentration with a specific molecular weight hydrophobe, a minimum weight percent of ethylene oxide is required before a specific block copolymer will form a gel in an aqueous solution. Examples of minimum weight percent concentrations with specific molecular weight hydrophobes are set out in Table II. Table II

At least a 40 percent weight concentration of the block copolymer having a hydrophobe of at least 2,250 molecular weight with at least about 50 weight percent of ethylene oxide condensed therewith- will be necessary to form a gel in an aqueous solution. In all cases, the block copolymers above the minima indicated in Table I will form gels in aqueous solutions up to 90 weight percent concentration and higher. Above 90 weight percent concentration, however, the gels tend to become indistinguishable from the starting block copolymer itself. It is to be understood that the molecular weight of the hydrophobe may be other than those illustrated in Table I. Thus, for example, if a hydrophobe of about 2500 molecular weight is used, it is recognized that a gel may be formed from the block copolymer at a concentration of 40 weight percent in an aqueous solution where about 45 weight percent of ethylene oxide is present in the block copolymer.

From the information presented in Tables I and II, it can be seen that the following provisions must be maintained to prepare gel compositions in accordance with the present invention:

1. When a in the following formula HO(C H O) (C H O) (C H O) H 2 4 V 3 6 V 2 4 'b is an integer such that the average molecular weight of the hydrophobe is about 2250 daltons, then the ethylene oxide content is from 50 to 90 weight percent of the copolymer, the total average molecular weight of the copolymer is from 4600 daltons to 10,750 daltons and the gel composition comprises from 40 to 50 weight percent of the copolymer.

2. When a in the following formula

HO(C H O) (C H O) (C H O) H

2 4 b 3 6 a 2 4 'b is an integer such that the average molecular weight of the hydrophobe is about 2750 daltons, then the ethylene oxide content is from 45 to

90 weight percent of the copolymer, the total average molecular weight of the copolymer is from 4910 daltons to 13500 daltons and the gel composition comprises from 40 to 50 weight percent of the copolymer.

3. When a in the following formula: HO(C H O) (C H O) (C H O) H ^v 2 4 V 3 6 V 2 4 'b is an integer such that the average molecular weight of the hydrophobe is about 3250 daltons, then the ethylene oxide content is from 35 to 90 weight percent of the copolymer, the total average molecular weight of the copolymer is from 4910 daltons to 15,510 daltons, and the gel composition comprises from 30 to 50 weight percent of the copolymer,

4. When a in the following formula:

HO(C H O) (C H O) (C H O) H

2 4 b 3 6 a 2 4 b is an integer such that the average molecular weight of the hydrophobe is about 4000 daltons, then the ethylene oxide content is from 35 to 90 weight percent of the copolymer, the total average molecular weight of the copolymer is from 6150 daltons to 20,000 daltons and the gel composition comprises from 30 to 50 weight percent of the copolymer, with the further proviso that when a in the formula is an integer such that the average molecular weight of the hydrophobe is about 4000 daltons, the ethylene oxide content is from 70 to 90 weight percent, the total average molecular weight of the block polymer is from 16,000 daltons to 20,000 daltons and the gel composition comprises from 15 to 50 weight percent of the copolymer. The preferred polyoxyethylene-polyoxypropylene copolymer is poloxamer 407 which has a chemical formula of α-hydro- omega- hydroxy- poly(oxyethylene)ιoi- poly(oxypropylene)56 - poly(oxyethylene)χoi. The poloxamer 407 can also be represented by the following formula: HO(C H O) (C H O) (C H O) H ^x 2 4 3 6 V 2 4 'b wherein the molecular weight of the hydrophobe (C3H6O) is approximately

4000 daltons and the total molecular weight of the compound is approximately 13,500 daltons.

Many different acid mucopolysaccharides are effective in the present invention. The acid mucopolysaccharides, also known as glycosaminoglycans, that are admixed or dissolved with the copolymer generally consist of recurring disaccharide units, each of which contains a derivative of an aminohexose, usually D-glucosamine or D-galactosamine. At least one of the two sugars in the recurring disaccharide unit of acid mucopolysaccharides contains an acidic group having a negative charge at pH 7, the negative group being either a carboxylate or sulfate group. An example of an acidic hexose is D-glucuronate, derived from D-glucose by oxidation of the θ carbon atom to a carboxylate group.

Acid mucopolysaccharides are thus heteropolysaccharides because they consist of at least two kinds of monosaccharides in alternating sequence which vary in such characteristics as chain length and charge density.

The prefix muco- refers to the fact that these polysaccharides were first isolated from mucin, the slippery, lubricating proteoglycan of mucous secretions.

The acid mucopolysaccharide hyaluronic acid derived from the intercellular cement of animal tissues contains many alternating units of D- glucuronic acid and N-acetyl-D-glucosamine. Hyaluronic acid forms highly viscous, jellylike solutions. Hyaluronic acid is often found combined with other mucopolysaccharides.

Chondroitin, a major polysaccharide of cartilage proteoglycans, contains alternating units of D-glucuronic acid and N-acetyl-D-galactosamine. Chondroitin may be regarded as the parent material for other widely distributed polysaccharides, such as chondroitin sulfate A and chondroitin sulfate C, which differ only in the position in which sulfate is esterified to the galactosamine moiety.

Another important mucopolysaccharide, long termed chondroitin sulfates is dermatan sulfate (α-L-iduronosyl-(l-3)-β-D-N-acytylgalactosamine

4-sulfate). Dermatan consists of repeating disaccharide units of iduronosyl and acetylgalactosamine. It can be isolated from various tissues including blood vessel walls and skin. Dermatan sulfate catalyses the thrombin-heparin cofactor II interaction. Despite the antithrombotic effects of dermatan sulfate, dermatan compounds generally are weak anticoagulants.

The acid polysaccharide heparin is generated by certain types of cells that are especially abundant in the lining of arterial blood vessels. Heparin, like other mucopolysaccharides, is an anionic carbohydrate chain synthesized as part of a proteoglycan. The basic building blocks of heparin are alternating units of uronic acid and N-acetylglucosamine with the predominant repeat disaccharide unit being L-iduronic acid and 2-amino-2-deoxy-D-glucose. The L-iduronic acid arises from the epimerization of D-glucuronic acid at position 5 during its biosynthesis and, in general, increases in the L-iduronic acid residues parallels the increase in sulfation of the heparin. If the epimerization and the sulfation reactions went to completion, there would be 3 sulfate per disaccharide consisting of an N-sulfate and a 6-O-sulfate on the aminosugar and a 2-O-sulfate on the L-iduronic acid. There are approximately 2.2 sulfate residues per disaccharide in standard porcine mucosal heparin, suggesting that there is a significant number of unmodified glucuronic acid residues in heparin. Incomplete conversion of glucuronic acid to iduronic acid is essential for the anticoagulant function of heparin.

Heparin exhibits strong anticoagulant activity. Low molecular weight heparins have a mean molecular weight of less than approximately 6000 daltons show much stronger anticoagulant activity than higher molecular weight heparins. The preferred acid mucopolysaccharides that are used in the present invention have little or no anticoagulant activity.

Heparans are another group of acid mucopolysaccharides. Chemically, heparans have the same carbohydrate backbone as heparin but differ in their sulfate content and distribution of charged groups. Heparans and other acid mucopolysaccharides exhibit significantly less anticoagulant properties compared to heparin.

It has been found that strongly anionic acid mucopolysaccharides are generally preferred when practicing the present invention. "Strongly anionic" means a molecule with a high negative charge density. The strongly anionic acid mucopolysaccharides have a greater therapeutic effect when compared to weakly anionic acid mucopolysaccharides.

Strongly anionic acid mucopolysaccharides can be prepared by any method capable of distinguishing molecules with a higher negative charge density from those of lower charge density. Electrophoresis or ion exchange chromatography are two such methods and are well known by those of ordinary skill in the art. Another method would simply utilize the anionic mucopolysaccharide as the free base rather than the sodium, calcium or lithium salt as they are commonly manufactured.

The size of the acid mucopolysaccharide can vary greatly. For example, the molecular weight of the mucopolysaccharide that is used in the topical treatment of the present invention can be from between 3000 daltons to

3,000,000 daltons. The preferable average molecular weight of the acid mucopolysaccharides is between approximately 5000 and 50,000 daltons. The most preferable molecular weight of the acid mucopolysaccharides is between approximately 10,000 and 20,000 daltons. It is to be understood that the molecular weight of the acid mucopolysaccharide is not a critical factor in the present invention, and that any one preparation of acid mucopolysaccharide will contain a population of molecules that can vary greatly in size. This variance in size of molecules does not necessarily reduce the effectiveness of the present invention. The present invention utilizes acid mucopolysaccharides of higher molecular weight to minimize systemic absorption and restrict the biologic actions to the locally applied area. (See Emanuele, et al., "The Effect of

Molecular Weight on the Bioavailability of Heparin", Thrombosis Research, Vol. 48, pgs. 591-596, (1987) which is incorporated herein by reference) Thus, an acid mucopolysaccharide can be used in the present invention where the acid mucopolysaccharide is of sufficient molecular size such that it is not absorbed systemically.

The most preferred acid mucopolysaccharide for use in the present invention is dermatan sulfate. The average molecular weight of dermatan sulfate is estimated to be approximately 30,000 daltons depending on its source and method of preparation. The concentration of dermatan sulfate in the composition of the present invention is preferably between approximately 10 mg per ml and 100 mg per ml. The more preferred concentration of dermatan sulfate is between 25 mg per ml and 75 mg per ml with a most preferred concentration of dermatan sulfate of approximately 50mg per ml.

Another preferred acid mucopolysaccharide for use in the present invention has a high molecular weight. The preferred average molecular weight of the heparin is approximately 15,000 daltons. The concentration of heparin in the composition of the present invention is preferably between approximately 10 mg per ml and 100 mg per ml. The more preferred concentration of heparin is between 20 mg per ml and 50 mg per ml with a most preferred concentration of heparin of approximately 25 mg per ml. Each mg of heparin is expected to contain approximately 100 units of activity.

Some of the commonly known acid mucopolysaccharides that can be used with the present invention include, but are not limited to, heparin, heparan sulfate, heparinoids, dermatan sulfate, pentosan polysulfate, chondroitin sulfate and hyaluronic acid. In accordance with the present invention, the acid mucopolysaccharides may also be admixed with pharmaceutically acceptable topical carriers. The carrier may comprise any commercially available base including but not limited to waxes, petrolatum, and glycerides. More specifically, the acid mucopolysaccharide may be admixed to commercially available salves, ointments or cerates containing a variety of pharmaceutically active agents.

The compositions prepared in accordance with the present invention comprise at least the following ingredients: (1) a pharmaceutically effective concentration of a acid mucopolysaccharide and (2) an aqueous gel comprising, based on 100 parts by weight, (a) from 15 to 50 parts, preferably from 15 to 25 parts, of a polyoxyethylene-polyoxypropylene- block copolymer and (b) from 50 to 85 parts, preferably from about 75 to 85 parts, of water.

The method of the present invention comprises the steps of topically administering to a human or animal with damaged or diseased tissue a composition comprising an admixture of a polyoxyethylene-polyoxypropylene block copolymer that has the physical property of being a liquid at room temperature or below and a gel at body temperature, and an acid mucopolysaccharide. The composition is described hereinabove. The composition of the present invention can be administered by dabbing, dripping, pouring, spraying or painting the composition onto the damaged or diseased tissue. In addition, the composition of the present invention comprising the block copolymer and acid mucopolysaccharide can be administered by inserting the damaged or diseased tissue into a solution of the composition thereby allowing the composition to gel onto the tissue.

Preferably, the composition of the present invention should be administered so that a gel layer forms over the damaged or diseased tissue thereby allowing the damaged or diseased tissue to be protected from the environment

In addition, because the acid mucopolysaccharide is dispersed throughout the gel, the acid mucopolysaccharide will be in constant contact with the damaged or diseased tissue thereby accelerating the healing of the tissue. It is to be understood that the topical administration of the composition of the present invention includes administration to the surface of the body as well as administration to tissue that is exposed through surgery or trauma. The composition and method of the present invention is particularly effective in treating bums. These bums can be chemical bums, thermal bums, electrical bums or radioactive bums. The present invention is also effective in treating other types of tissue damage such as traumatic damage, including but not limited to, compound fractures, cuts, abrasions or damage due to infection with bacteria, fungi or other microorganisms. It may be advantageous to add additional pharmaceutically active agents to the composition of the present invention depending upon the type of injury that is being treated. For example, if one is treating a serious bu according to the present invention, antimicrobial agents may be added to the composition of the present invention to retard infection. The antimicrobial agent may be an antibiotic, antifungal agent or a mixture thereof. Representative antimicrobial agents that can be used in practicing the present invention include, but are not limited to, penicillins, cephalosporins, bacitracins, aminoglycosides and polypeptide antibiotics and the bacteriastatic compounds including, but not limited to, chloramphenicol, tetracyclines, macrolides, sulfonamides and aminosalicylic acid. Antifungal agents that can be used in practicing the present invention include, but are not limited to, nystatin, amphotericin B and griseofuvin.

In addition, silver ions can be used in practicing the present invention. The silver salts that can be employed in the preparation of the gels of the present invention are those silver salts which will preferably dissolve in water at a minimum concentration of 0.1% by weight. Representative silver salts include silver nitrate, silver acetate, silver sulfate, and silver lactate. The amount of silver salt that will produce a beneficial effect is between about 0.1% and 1.0% by weight of silver salt based on the weight of the water in the gel. Growth factors such as human growth hormone, tissue derived growth factor, epidermal growth factor, platelet derived growth factor, fibroblast growth factor, and/or nerve growth factor may optionally be added to the composition of the present invention, either singly or in combination, to enhance the growth and development of the damaged or diseased tissue. In addition, anti-inflammatory agents may be added to reduce inflammation in the damaged or diseased tissue.

The composition of the present invention can optionally contain anesthetics to alleviate pain when applied to the damaged or diseased tissue. Representative anesthetics that can be employed in the present invention include, but are not limited to, lidocaine and procaine. This invention is further illustrated by the following examples, which are not to be construed in any way as imposing limitations upon the scope thereof. On the contrary, it is to be clearly understood that resort may be had to various other embodiments, modifications, and equivalents thereof which, after reading the description herein, may suggest themselves to those skilled in the art without departing from the spirit of the present invention and/or the scope of the appended claims.

Example I A topically-applicable gel composition that is effective for treating damaged or diseased tissue is prepared using the following ingredients:

Parts Ingredients

10% Copolymer 2% Dermatan Sulfate

78% Water

The copolymer in this example has the following general formula:

HO(C H O) (C H O) (C H O) H

2 4 ' b 3 6 'a 2 4 'b wherein the molecular weight of the hydrophobe (C3H6O) is approximately 4000 daltons and the total molecular weight of the compound is approximately 13,500 daltons.

Example II

The composition of Example I is administered to a patient with a second degree bum on his arm. The composition is placed in a syringe and is cooled to approximately 20°C. The composition is slowly applied to the surface of the bum, allowing the liquid composition to gel on the bum. Enough of the composition is added to form a gel that is approximately 0.2 cm in depth.

Example III

A topically-applicable gel composition that is effective for treating damaged or diseased tissue is prepared using the following ingredients: Parts Ingredients

20% Copolymer

4% Heparin

0.5% Gentamicin sulfate

75.5% Water

The copolymer in this example has the following general formula:

HO(C H O) (C H O) (C H O) H 2 4 b 3 6 a 2 4 'b wherein the molecular weight of the hydrophobe (C3H6O) is approximately

Example IV The composition of Example HI is administered to a patient with a first degree bum on his leg. The composition is placed in a syringe and is cooled to approximately 20°C. The composition is slowly applied to the surface of the bum, allowing the liquid composition to gel on the bum. Enough of the composition is added to form a gel that is approximately 0.2 cm in depth.

Example V A topically-applicable gel composition that is effective for treating damaged or diseased tissue is prepared using the following ingredients:

Parts . Ingredients

20% Copolymer

1% Dermatan sulfate

0.5% Gentamicin sulfate

78.5% Water

The copolymer in this example has the following general formula:

HO(C H O) (C H O) (C H O) H

2 4 V 3 6 a 2 4 'b wherein the molecular weight of the hydrophobe (C3H5O) is approximately 4000 daltons and the total molecular weight of the compound is approximately

13,500 daltons. Example VI

The composition of Example V is administered to a patient with a third degree bum on his torso. The composition is placed in a syringe and is cooled to approximately 20°C. The composition is slowly applied by spraying the composition onto the surface of the bum, allowing the liquid composition to gel on the bum. Enough of the composition is added to form a gel that is approximately 0.2 cm in depth.

Example VII A topically-applicable gel composition that is effective for treating damaged or diseased tissue is prepared using the following ingredients:

Parts Ingredients

20% Copolymer l% mg/ml Heparin

1% by weight Lidocaine

78% Water

The copolymer in this example has the following general formula:

HO(C H O) (C H O) (C H O) H

2 4 3 6 a 2 4 ⁷b wherein the molecular weight of the hydrophobe (C3H6O) is approximately 4000 daltons and the total molecular weight of the compound is approximately 13,500 daltons.

Example VIII

The composition of Example VII is administered to a patient with a second degree sunburn on his back. The composition is placed in a syringe at room temperature. The composition is slowly applied by dripping the composition onto the surface of the bum, allowing the liquid composition to gel on the bum. Enough of the composition is added to form a gel that is approximately 0.1 cm in depth.

Example IX A topically-applicable gel composition which is effective in treating bums is prepared from the following ingredients: Parts Ingredients

20% Copolymer

5% Dermatan

75% Saline

The copolymer (poloxamer 407, BASF Corporation, Parsippany, NJ)) in this example has the following general formula: HO(C H O) (C H O) (C H O) H 2 4 V 3 6 a 2 4 b wherein the molecular weight of the hydrophobe (C3H6O) is approximately 4000 daltons and the total molecular weight of the compound is approximately

13,500 daltons.

The effects of the topically-applicable gel composition in the treatment of thermally injured tissue were studied in a guinea pig model^' of split thickness burns. Deeply anesthetised, hairless guinea pigs were subjected to thermal tissue injury by placing a 5 cm^, 80g metal probe (heated to 80°C) on their back for exactly 5 seconds. Thirty minutes post bum, animals were either left untreated (control), treated with the 20% gel of poloxamer 407 without dermatan or treated with 20% poloxamer 407 and 5% dermatan sulfate (Scientific Protein Labs, Waunakee, WS). Treated animals received 0.5 ml applications of the appropriate test article at 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 24, 28,

32 and 48 hours post burn. All animals were assessed for wound area, skin thickness and erythema at various intervals post bum. Histologic sections demonstrated that reduction in skin thickness correlated with a reduction in new collagen formation (scar) in the superficial dermis. The effect of the topically-applicable gel composition on wound contracture is shown in Table III. Bum wound measurements are made by determining the distance between points tattooed at the wound periphery immediately post bu . The data clearly shows that the combination of heparin and copolymer had a significant effect on wound contracture at 24 hours and 72 hours. Table III Area of Lesion

The effects of the topically-applicable gel composition on skin thickness in the bum model are shown in Table IV. The poloxamer/dermatan treated animals had a less skin thickness when compared to either poloxamer only treated animals or control animals. This was especially true at one day post bum.

Table IV Skin Thickness

The effects of the topically-applicable gel composition on erythema in the bum model are shown in Table V. Erythema was scored as 0 for little or no erythema to 3 for maximum erythema. As can be seen in Table V, the animals that were treated with the poloxamer/dermatan combination had less erythema than did the animals that were treated with poloxamer only or were left untreated.

Table V Erythema (0 to 3)

It should be understood, of course, that the foregoing relates only to a preferred embodiment of the present invention and that numerous modifications or alterations may be made therein without departing from the spirit and the scope of the invention as set forth in the appended claims.

Claims

1. A composition for topically treating damaged or diseased tissue comprising a solution of: a. a polyoxyethylene-polyoxypropylene block copolymer having the following formula:

HO(C H O) (C H O) (C H O) H 2 4 b 3 6 a 2 4 b wherein a is an integer such that the hydrophobe represented by C3H6O has a molecular weight of between 2250 and 6000 daltons and b is an integer such that the hydrophile portion represented by C2H4O constitutes from about 50 to 90 percent by weight of the copolymer, and b . an effective amount of dermatan sulfate.

2. The composition of Claim 1, wherein the hydrophobe represented by C3H6O has an average molecular weight of between approximately 2750 and 4000 daltons.

3. The composition of Claim 2, wherein the hydrophobe represented by C3H6O has an average molecular weight of approximately 4000 daltons.

4. The composition of Claim 1, further comprising an antimicrobial compound.

5. The composition of Claim 4, wherein the antimicrobial compound is an antibacterial compound.

6. The composition of Claim 4, wherein the antimicrobial compound is an antifungal compound.

7. The composition of Claim 1 , further comprising an anti- inflammatory agent.

8. The composition of Claim 1, further comprising a growth factor.

9. The composition of Claim 1, further comprising an anesthetic.

10. A method of treating damaged tissue in a human or animal comprising the step of topically administering to the human or animal with diseased or damaged tissue an aqueous solution containing: a. a polyoxyethylene-polyoxypropylene block copolymer having the following formula:

HO(C H O) (C H O) (C H O) H ^v 2 4 V 3 6 a 2 4 'b wherein a is an integer such that the hydrophobe represented by C3H6O has a molecular weight of between 2250 and 6000 daltons and b is an integer such that the hydrophile portion represented by C2H4O constitutes from about 50 to 90 percent by weight of the copolymer, and b . an effective amount of an acid mucopolysaccharide.

11. The method of Claim 10, wherein the acid mucopolysaccharide is selected from the group consisting of heparin, heparan sulfate, heparinoids, dermatan sulfate, pentosan polysulfate, chondroitan sulfate and hyaluronic acid. -

12. The method of Claim 11 , wherein the acid mucopolysaccharide is heparin.

13. The method of Claim 11, wherein the acid mucopolysaccharide is dermatan sulfate.

14. The method of Claim 10, wherein the acid mucopolysaccharide is has minimal anticoagulant activity.

15. The method of Claim 10, wherein the hydrophobe represented by C3H6O has an average molecular weight of between approximately 2750 and 4000 daltons.

16. The method of Claim 15 wherein the hydrophobe represented by C3H6O has an average molecular weight of approximately 4000 daltons.

17. The method of Claim 10, wherein the composition further comprises an antimicrobial compound.

18. The method of Claim 17, wherein the antimicrobial compound is an antibacterial compound.

19. The method of Claim 17, wherein the antimicrobial compound is an antifungal compound.

20. The method of Claim 10, wherein the composition further comprises an anti-inflammatory agent.

21. The method of Claim 10, wherein the composition further comprises an anesthetic.

22. A method of treating burns in a human or animal comprising the step of topically administering to the human or animal with the bu a composition comprising a. a polyoxyethylene-polyoxypropylene block copolymer having the following formula:

HO(C H O) (C H O) (C H O) H 2 4 V 3 6 a 2 4 'b wherein a is an integer such that the hydrophobe represented by C3H6O has a molecular weight of between 2250 and 6000 daltons and b is an integer such that the hydrophile portion represented by C2H4O constitutes from about 50 to 90 percent by weight of the copolymer, and b . an effective amount of an acid mucopolysaccharide.

23. The method of Claim 22, wherein the acid mucopolysaccharide is selected from the group consisting of heparin, heparin sulfate, heparinoids, low molecular weight heparin, dermatan sulfate, pentosan polysulfate, chondroitan sulfate and hyaluronic acid.

24. The method of Claim 23, wherein the acid mucopolysaccharide is heparin.

25. The method of Claim 27, wherein the acid mucopolysaccharide is dermatan sulfate.

26. The method of Claim 22, wherein the acid mucopolysaccharide is has minimal anticoagulant activity.

27. The method of Claim 22, wherein the hydrophobe represented by C3H6O has an average molecular weight of between approximately 2750 and 4000 daltons.

28. The method of Claim 27, wherein the hydrophobe represented by C3H6O has an average molecular weight of approximately 4000 daltons.

29. The method of Claim 22, wherein the composition further comprises an antimicrobial compound.

30. The method of Claim 29, wherein the antimicrobial compound is an antibacterial compound.

31. The method of Claim 29, wherein the antimicrobial compound is an antifungal compound.

32. The method of Claim 22, wherein the composition further comprises an anti-inflammatory agent.

33. The method of Claim 22, further comprising an anesthetic.