WO2015097513A1

WO2015097513A1 - Topical products for aminoguanidines

Info

Publication number: WO2015097513A1
Application number: PCT/IB2014/002761
Authority: WO
Inventors: Ake Lindahl
Original assignee: Anamar Derma Ab
Priority date: 2013-12-13
Filing date: 2014-12-12
Publication date: 2015-07-02

Abstract

The present invention generally relates to the use of topical formulations to deliver aminoguanidine compounds to the skin for the treatment of diseases such as dermal inflammatory conditions.

Description

TOPICAL PRODUCTS FOR AMINOGUANIDINES

RELATED APPLICATION

The present application claims priority to U.S. Application Serial No. 61/915,815, filed December 13, 2013, the disclosure of which is incorporated by reference in its entirety.

FIELD OF THE INVENTION

The present invention generally relates to the use of topical formulations to deliver substances, certain aminoguanidines, to the skin for the treatment of diseases such as dermal inflammatory conditions.

BACKGROUND OF THE INVENTION

Adsorption and transportation of drugs through the skin can be affected by a number of variables, including skin physiology, properties of the drug and formulation of the delivery system. Topical dermatological products (anti-inflammatory agents, local anesthetics, sunscreen, etc.) usually target one or more of the layers of the skin and exist in a plethora of different formulations depending on the application (aqueous solutions, lotions, pastes, powders, ointments, foams, creams and gels, etc.). The rate-limiting barrier for percutaneous drug adsorption occurs through the stratum corneum of the skin, which is the outermost layer of the skin followed by the viable epidermis and dermis. Once the drugs have passed through the stratum corneum, the passage of deeper skin layers occurs quickly.

The stratum corneum has a low water content (20%) and a high lipid (40%) and protein (40%) content, making it tough for hydrophilic and charged molecules to pass through this layer. Hydration of the stratum corneum increases the passage of drugs through the skin layer by formation of intra- and intercellular channels and pathways. This type of hydration can be performed by skin occlusion by using non-volatile hydrocarbon bases, such as Vaseline ®. Skin occlusion prevents moisture from escaping from the skin, and thus, moisture may accumulate between the skin and the ointment layer. Certain penetration enhancers, such as alcohol, may also be utilized.

For most dermal inflammatory conditions, the site of action is the dermis, and to deliver the drug to the dermis, it must pass the stratum corneum and epidermis. Atopic dermatitis (AD) is one of the most frequent chronic inflammatory diseases, affecting approximately 1-3 % of the adult population and 25 % of children. AD is not a life- threatening illness; nevertheless, there is a substantial financial impact on the health care system. Additionally, there is a large impact on the patients' physical and psychosocial wellbeing together with an indirect cost for the health care system due to, e.g., loss of working days. There is a large unmet medical need due to lack of efficacy and limiting side effects of current treatments, i.e., corticosteroids and macrolactam immunosuppressives. From a pharmacological point of view, the use of aminoguanidines with serotonin inhibiting effect, could be of large medical value in several inflammatory skin diseases. An issue associated with aminoguanidines is that they are not soluble in water unless they are converted to a salt, and salts are charged, and therefore, do not penetrate the skin. The uncharged

aminoguanidines exist in water only at high pH, which is irritating to the skin itself or when dissolved in a water-free environment.

SUMMARY OF THE INVENTION

A topical formulation that will deliver certain aminoguanidine compounds to the skin without using high pH is presented herein.

A surprisingly high solubility of certain aminoguanidines using certain polar solvents has been discovered. More specifically, alcohols and polar esters are groups of solvents that are preferred to use as solvents for aminoguanidines, in particular diols and esters of lactic acid and citric acid. In some embodiments, the solvents are selected from propylene glycol, dipropylene glycol and triethyl citrate, or a mixture thereof. It is therefore an embodiment of the invention to utilize propylene glycol, dipropylene glycol and triethyl citrate, or a mixture of the aforementioned solvents, with high enough solubility for certain aminoguanidines to create a medically relevant penetration of these aminoguanidines into the skin from a topical formulation.

The solvents or mixtures of the solvents are however not suitable for dermal use on their own. The viscosity is low and the topical application will run off after applying it to the skin. If viscosity-enhancing compounds are introduced, the properties of the formulation will change and the skin penetration (bioavailability) will decrease. It was surprisingly found that this problem could be solved using an emulsion that does not contain water. In another embodiment of the invention, an emulsion that contains a continuous lipid phase and an inner phase consisting of polar solvent(s) with certain aminoguanidines in solution is therefore used. When applied to the skin, the emulsion may break and the polar solvent phase may have contact with the skin. During this process, the continuous phase lipids will provide occlusion to decrease water evaporation from the site of application and thus increase hydration of the stratum corneum.

In some aspects, the aminoguanidines of the invention include, but are not limited to, compounds of general fo mula I:

In one embodiment, an aminoguanidine of the invention is Compound 1 :

In another embodiment, an aminoguanidine of the invention is Compound 2:

In yet another embodiment, an aminoguanidine of the invention is Compound 3 :

Other embodiments and advantages will be more fully apparent from the following disclosure.

BRIEF DESCRIPTION OF THE DRAWINGS

The following drawings are provided to illustrate various aspects of the present inventive concept and are not intended to limit the scope of the present invention unless specified herein. Figure 1 presents dermal concentration of Compound 1 administered either in the form of a 1% cream or as a 2.77 % solution or 2.77 % suspension.

DETAILED DESCRIPTION

The foregoing and other aspects of the present invention will now be described in more detail with respect to the description and methodologies provided herein. It should be appreciated that the invention may be embodied in different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art.

The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. As used in the description of the embodiments of the invention and the appended claims, the singular forms "a," "an" and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise. Also, as used herein, "and/or" refers to and encompasses any and all possible combinations of one or more of the associated listed items. Furthermore, the term "about," as used herein when referring to a measurable value such as an amount of a compound, dose, time, temperature, and the like, is meant to encompass, in addition to the measurable value, variations of 20%, 10%, 5%, 1 %, 0.5%, or even 0.1% of the specified amount. When a range is employed (e.g., a range from x to y) it is it meant that the measurable value is a range from about x to about y, or any range therein, such as about xi to about yi, etc. It will be further understood that the terms "comprises" and/or "comprising," when used in this specification, specify the presence of stated features, integers, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and/or groups thereof. Percentages refer to weight percent, and also refer to the total formulation unless otherwise specified. Further, unless otherwise defined, all terms, including technical and scientific terms used in the description, have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

All patents, patent applications and publications referred to herein are incorporated by reference in their entirety. In the event of conflicting terminology, the present specification is controlling. Further, the embodiments described in one aspect of the present invention are not limited to the aspect described. The embodiments may also be applied to a different aspect of the invention as long as the embodiments do not prevent these aspects of the invention from operating for its intended purpose.

Embodiments of the present invention relate to topical formulations used to deliver certain aminoguanidines to alleviate dermal inflammatory conditions. Examples of inflammatory conditions that would benefit from treatment with aminoguanidines include, but are not limited to, dermatitis and psoriasis and in particular pruritus/itch. Other conditions involving inflammation caused by infections of various origins could also benefit from aminoguanidine treatment.

The importance of serotonin (5-HT) and its receptors are receiving increased scientific attention outside the CNS (central nervous system)-field. A number of studies show the important role of 5-HT in the pathogenesis and symptoms of dermatitis. 5-HT itself has a proinflammatory role as well as a direct role in pruritus. For example, 5-HT has been shown to cause an increase in vascular permeability, which contributes to the edema formation in dermatitis. The 5-HT responding cells of importance in dermatitis (dendritic cells, monocytes, mast cells, keratinocytes, fibroblasts) have all been shown to express 5-HT₂ receptors.

Targeting 5-HT₂ receptors is a new concept for the treatment of dermatitis. A suppression of inflammatory edema formation and a reduction of an itch-response have been demonstrated in vivo using the "old" 5-HT₂ receptor antagonist ketanserin (Hong et al. Pain. 2006 Sep; 124 (1- 2):27-33).

A new type of 5-HT₂ receptor antagonist, certain types of aminoguanidines, may act as anti-inflammatory compounds both in vitro and in vivo, supporting the concept of using serotonin antagonists for the treatment of dermatitis.

Therefore, more specifically, the invention relates to the use of aminoguanidines including, but not limited to, the following:

Compounds of the general formula I:

I including stereoisomers and tautomers, salts, esters or prodrugs thereof, wherein R¹ - R⁵ are independently chosen from the group comprising H, C_1-5-alkyl, OH, Ci_-5-alkoxy, SMe, SEt, CF₃, OCF₃, F, CI, Br, I, CN, S(0)Me, S(0)₂Me, S(0)₂NMe₂, NMe₂, NHC(0)H, NHC(0)Me, CONMe₂, optionally including two adjacent groups, R¹ - R², R² - R^3, R³ - R⁴, or R⁴ - R⁵, representing OCH₂0, OCH₂CH₂0 or SCH₂CH₂S;

X is non-existing or is chosen from the group comprising CH₂, (CH₂)₂, (CH₂)₃, CH=CH, CH₂CH=CH, and CH=CHCH₂;

Y is non-existing or is chosen from the group comprising CH₂, (CH₂)₂, (CH₂)₃, CH₂)₄, and (CH₂)₅;

A is chosen from the group comprising H, Ph, and 5-6 membered hetero-aromatic rings containing 1-3 hetero-atoms independently chosen from the group comprising N, O, and S; B is chosen from the group comprising H, Me, Et, Ph, benzyl, and OH,

or pharmaceutically acceptable salts of the compounds of the general formula I.

Specific aminoguandine compounds include, but are not limited to, for example, the following:

Compound 1 - (E>2-(2-chloro-3,4-dimethoxybenzylidene>N- methylhydrazinecarboximidamide

Compound 2 - (E)-2-(2-chloro-3,4-dimethoxybenzylideneVN- phenylhydrazinecarboximidamide

Compound 3 - (E - -(4-methoxvbenzvlidene -N-phenethylhydrazinecarboximidamide

These compounds can be used to treat dermal inflammatory conditions, such as dermatitis and pruritus. A particular route of administration of these aminoguanidines is through local topical application of the aminoguanidines formulated into a carrier system. These local topically applied formulations may provide an effective concentration of the aminoguanidines at the local site of action. It is therefore a primary object of the invention to provide a suitable topical formulation with a carrier system for the non-limiting examples of aminoguanidines. WO 99/26613 describes aminoguanidine compositions for treating skin proliferative diseases, such as psoriasis. Methods and compositions are described to formulate aminoguanidines into pharmaceutically acceptable carrier systems for topical application or oral administration. The aminoguanidines are formulated into various ointments and creams, including VASELINE®, NIVEA® cream, paraffin and Unguentum Emulsificans. Preferably in WO 99/26613, the active compound used is aminoguanidine or an acidic salt thereof, for example, bicarbonate, a hemisulphate or a hydrochloride salt.

By using topical applications, the active components (in the present invention, certain aminoguanidines) must be transferred through the skin to the local site of action. This poses several issues that need to be addressed in order to transfer an effective concentration of the active components into the skin. First of all, the outermost layer of the skin, the stratum corneum, is a very effective barrier for molecules to pass due to its hydrophobic character and high lipid and protein content. Thus, the rate-limiting step for the transfer of active components into the skin occurs through this skin layer. Second, the aminoguanidines exemplified by, but not limited to, Compound 1 , 2 and 3 in the present invention, are not soluble in water unless they are converted to a salt and salts are charged and difficult to transfer through the skin. The uncharged aminoguanidines exemplified above exist only at high pH (which is irritating to the skin itself) or when dissolved in a water-free environment. Therefore, there is a need to find a topical formulation that provides: 1) a suitable chemical formulation that allows for solving uncharged aminoguanidines (exemplified by, but not limited to, Compound 1, 2 and 3) without using a high pH (which is irritating to the skin), and 2) a carrier system for the transport of active components (in the present invention

exemplified by, but not limited to, the aminoguanidines Compound 1, 2 and 3) through the stratum corneum and into the deeper layers of the skin. Pharmaceutically acceptable carrier systems refer to those that are suitable for application to a subject, such as a human, without undue side effects such as toxicity or irritation to the skin. Undue side effects are those that render the formulation unsuitable for application to a subject at least because the harm from the side effects outweighs the benefits of the formulation. A surprisingly high solubility of amino guanidines (exemplified by, but not limited to, Compound 1, 2 and 3) was found using a set of polar solvents as exemplified below. We have found that alcohols and polar esters are solvents that are preferred as good solvents for the aminoguanidines. Suitable polar solvents are diols represented, but not limited to, propylene glycol, hexanediol and dipropylene glycol, and esters represented by polar citric and lactic acid esters. Other suitable solvents include esters such as propylene carbonate and ethers such as dimethyl isosorbide (dimethyl ether of an anhydride of an isomer of sorbitol, also usable as a penetration enhancer). It has been found that, in some embodiments, solvents can be mixed to generate higher solubility than if a single solvent is used. The aminoguanidines with suitable pharmacological properties have different properties with respect to lipophilicity. Having access to the teaching of this patent application a person skilled in the art can generate an optimal solvent mixture using the suitable solvents.

In some embodiments, the solvents are selected from propylene glycol, dipropylene glycol and triethyl citrate or mixtures of two or all three. As non-limiting examples, the solvents for Compound 1 are propylene glycol and dipropylene glycol while Compound 2 and Compound 3 may be better dissolved in mixtures of propylene glycol and triethyl citrate. Another aspect of the solvent property is the stability of the active components in the solvent. If the active component degrades in the solvent the use and shelf-life of any product will be limited. Solvents that do not degrade the active compound include propylene glycol, dipropyleneglycol and triethyl citrate. It is therefore an embodiment of the invention to use propylene glycol, dipropyleneglycol and triethyl citrate or mixtures thereof to provide solvents for aminoguanidines (exemplified by, but not limited to, Compound 1, 2 and 3). WO 99/26613 does not disclose the use of the preferred solvents disclosed in the present invention that allows solvation, and in particular, efficient solvation, of certain aminoguanidines (exemplified, but not limited to, Compound 1 , 2 and 3 in the present invention), without using high pH, i.e. pH > about 6.

The solution of the active component in the polar solvents is not well suited for application on skin. The solutions are low viscous and will generally run off after

administration. However using the invented solvent system, several suitable formulation types can be generated. Non-limiting examples of such single phase formulations are gels, which are solutions supplied with thickening agents preventing the formulation to run off after application. Other suitable formulation types are ointment bases.

Another embodiment of the invention is the formation of an emulsion with the polar solvents in oil (p/o). Emulsions, containing water, are the most common forms for topical administration and are well recognized by patients. However, attempts to make a conventional emulsion containing the active components, have failed due to the poor solubility of the uncharged active in water-phase or in the oil phase. The emulsion used was an oil-in-water emulsion containing approximately 30 % lipids. See WO 99/26613 for details. The ingredients were cetanol, adeps lanae, polysorbate 60, glycerol, dimeticone,

isopropylmyristate, sorbitane stearate, sorbic acid and water. In order to maintain the high solubility of the active components, attempts were made to manufacture emulsions that do not contain water. Surprisingly, a suitable emulsion system could be formulated using polar solvents in a polar phase and lipids in the oil phase.

Another embodiment of the invention is therefore to use an emulsion that contains a continuous lipid phase and an inner phase consisting of polar solvent(s) with aminoguanidine in solution. When applied to the skin, the emulsion will break and the polar solvent phase will be in contact with the skin. During this process, the continuous phase, lipids, will provide occlusion to decrease water evaporation from the site of application and thus increase hydration of the stratum corneum. WO 99/26613 does not disclose the use of an emulsion that contains a continuous lipid phase and an inner phase consisting of polar solvent(s) with certain aminoguanidines in solution.

The discontinuous phase in the present invention contains the active components exemplified, but not limited to, the mentioned aminoguanidines (Compound 1 , 2 and 3) and solvents for the active components, such as propylene glycol, dipropylene glycol or triethyl citrate or mixtures of two or three of these solvents. In some embodiments, the discontinuous phase constitutes 20 to 90 % by weight of the topical formulation, in some embodiments about 30 to about 85 % and in some embodiments about 35 to about 70% by weight of the topical formulation. Furthermore, additional compounds normally used in skin products, can also be included. Examples of such compounds include, but are not limited to, chemical stabilisers, penetration enhancers, compounds giving texture to the formulation, pH modifiers, physical or chemical sun blocking agents, antimicrobial agents, antioxidants, etc.). In some embodiments, the discontinuous phase will also contain one or more surfactants. By water- free or "anhydrous" we mean that no water is added to the formulation (unless water is a constituent of the components added to the formulation, e.g. in hydrated form(s) or as in the case of lactic acid containing about 10 % of water). That is, no direct addition of water to a formulation of the present invention. However, those skilled in the art will recognize that water may be physically and/or chemically absorbed by a formulation and/or by one or more ingredients in a formulation at any time during the preparation, storage, and/or use of a formulation of the present invention (i.e., indirect addition of water to the formulation). In some embodiments, the term "anhydrous" means that a formulation has a water content of less than 5% by weight of the formulation or any range therein. A formulation of the present invention may have a water content of less than about 5, 4.5, 4, 3.5, 3, 2.5, 2, 1.5, 1, 0.5%, by weight of the composition. Water content may be measured by methods known to those of skill in the art, such as, but not limited to, Karl Fischer titration.

HLB is defined as Hydrophile/Lipophile Balance and is measured as the balance based on the molecular weight of the hydrophile (water loving) part and the lipophile (oil loving) part. Suitable surfactants in the polar phase are those that contribute to a total HLB value of less than 9 and more than 4. Since the compositions do not contain water, non-ionic surfactant are preferred. Non-limiting examples of suitable surfactants for the polar phase are esters or ethers of polyoxyethylene and saturated fatty acids. Esters of oligosaccharides may be included as well. In one embodiment of the invention, other polar compounds are added, with the purpose of increasing the polar character of the discontinuous phase. Non-limiting examples of such compounds include glycerol and polyethylene glycol. Therefore, another embodiment include the use of a highly polar compound such as glycerol, which can be added up to about 60% by weight of the topical formulation in order to improve the physical stability of the emulsion. The highly polar compounds, exemplified by but no limited to glycerol, di and oligosaccharides and polyethylene glycol, do not precipitate the active when added and can be regarded as a polarising diluents.

The continuous phase may contain nonpolar solvents or lipids and emulsifiers.

Suitable components are di and triglycerides or mixtures thereof, short chain monoglycerides (C8 to C12), fatty acid esters exemplified, but not limited to, .cetyl esters, isopropyl esters of fatty acids esters such as isoproylmyriststate or isopropylpalmitate. In one embodiment of the invention, the continuous phase contains a surfactant or a mixture of surfactants in order to facilitate the formation of an emulsion. Suitable surfactants or mixtures of surfactants have a hydrophile/lipophile balance of less than 9 and more than 4. In one embodiment, the emulsifier shall contain saturated fatty acid esters exemplified but not limited to stearic, palmitic, myristic and lauric acids. As for the polar phase, the non-ionic surfactants are better suited. Examples of non-ionic surfactants are esters of fatty acids or mono or diglycerides of fatty acids with oligosaccharides, propandiol, glycerol or polyoxyethylene. In another embodiment of the invention the continuous phase may contain thickening agent suitable for use in lipids. Furthermore the continuous phase may contain thickening agents in the form of waxes, exemplified, but not limited to, microcrystalline wax, carnuba wax, cerosine wax and cetyl esters wax. Other suitable types of thickening agents for lipids include, for example, polymeric thickeners such as ethylene vinyl alcohol copolymers and ethylene acrylic acid copolymers. Other examples are lipophilic fused silica particles and polyethene or polyethylene particles. The continuous phase may also contain fatty acids for the purpose of generating a slightly acidic pH in contact with moist on the skin. Further useful ingredients that can be incorporated are skin penetration enhancers exemplified, but not limited to, unsaturated fatty acid alcohols, esters further exemplified by derivatives of oleic acid and linoleic acid. Yet another type of ingredients that can be incorporated into the continuous phase are stabilising agents for the compounds exemplified by butylhydroxy anisol and butylhydroxyl toluene.

The concentration of the active component in the formulation should be selected in such way that the desired pharmacological effect is achieved without causing adverse effects. This concentration is individual for each aminoguanidine depending on its ability to pass the stratum corneum and its binding properties at the site of action as would be understood by one of ordinary skill in the art. In some embodiments, the concentration of the active components of certain aminoguanidines (exemplified, but not limited to, Compound 1 , 2 and 3 in the present invention) ranges from about 0.01 to about 10%, in some embodiments about 0.1 to about 10 %, in some embodiments about 0.1 to about 5%, in some embodiments about 0.5 to about 5 % by weight of the topical formulation. These concentration levels relate to certain dosage levels of the active components corresponding to a one, two, three or four times daily topical application of the formulation.

In another embodiment of the invention, a penetration enhancer can be introduced. Non-limiting examples of such enhancers include oleyl alcohol, oleic acid and urea. In another embodiment of the invention, the active component is presented in a gel formulation. The gel formulation constitutes a single phase solution of the active compound(s) in the preferred solvents, propylene glycol, triethylcitrate and dipropyleneglycol. The gel formulation may also contain thickening agents in particle form, exemplified by hydrophilic fused silica particles, or in dissolved form exemplified by macrogols, metacrylates/acrylates and polyoxamers. Other useful components such as acidifyers, penetration enhancers, stabilisers and antimicrobial agents may be added. The compositions above are suitable for use in topical formulations exemplified but not limited to lotions, creams, ointments, foams.

A non-limiting description of a formulation is an emulsion comprising, consisting, or consisting essentially of a continuous phase, a solution of an oil, paraffine oil and

capric/caprylic monoglycerides, a thickening agent, hydrophobic fused silica, a wax, microcrystalline wax and an emulsifier and a discontinuous phase comprising, consisting, or consisting essentially of a solution of the active ingredient, including but not limited to Compound 1, dissolved in propylenglycol, glycerol and lactic acid. In some embodiments, the formulation can comprise, consist, or consist essentially of about 20 to about 90 % of the continuous lipid phase, in some embodiments about 30 to about 85 % and in some

embodiments about 35 to about 70 % of the entire formulation. Based on the total weight of the formulation the continuous phase should preferably contain an emulsifier or a combination of emulsifiers giving an mutual HLB value of about 4 to about 9 and in some embodiments · about 4.5 to about 7. The emulsifier should be present, in some embodiments, at an amount of about 1 to about 10 %. The oil should, in some embodiments make up about 20 to about 80 % of the formulation, in some embodiments about 30 to about 60 % and in some embodiments about 35 to about 50 % of the formulation. The wax should in some embodiments be present in an amount of about 0.5 to about 5% of the formulation, and in some embodiments about 1 to about 3 % of the formulation, while the thickening agent can in some embodiments be present in an amount of about 1 to about 5 % of the formulation and in some embodiments at about 2 to about 4 % of the formulation.

In some embodiments, the formulation can comprise, consist, or consist essentially of about 10 to about 80 % of the discontinuous phase, in some embodiments about 15 to about 70 % and in some embodiments about 30 to about 65 % of the formulation. The active ingredient, an aminoguanidine, represented but not limited to Compounds 1 , 2 and 3 should in some embodiments be present in an amount of about 0.1 to about 10% of the entire formulation. In some embodiments, the content should be about 0.5 to about 5 %. In general, the amount can be evaluated from pharmaceutical and/or pharmacological point of view as understood by one of ordinary skill in the art. In some embodiments, the content of a polar solvent should be about 4 to about 32 %, in some embodiments about 6 to about 28 % and in some embodiments from about 12 to about 26% of the entire formulation. The polar compound(s) exemplified by the agents glycerol and polyethylenel glycol should in some embodiments be present in an amount of about 6 to about 50 %, in some embodiments from about 10 to about 42 % and in some embodiments from about 20 to about 40 % of the entire formulation.

In some embodiments, the formulation consists of about 50 % of each of the polar and lipid phases. The lipid phase contains, for example, about 3 % of emulsifier, about 42 % of oil, about 1.7 % of wax and about 3.5 % of fused silica. The polar phase contains, for example, of about 1 % of amino guanidine, about 20 % of propyleneglycol and about 30 % of glycerol. Lactic acid is present at about 0.6 %.

The topical formulations described herein may be used to treat a dermatological condition. In particular, the topical formulations of the present invention may be used to treat dermal inflammatory conditions. In certain embodiments, the dermal inflammatory condition is dermatitis, psoriasis, pruritus and/or eczema. The topical formulation may be applied to the skin at a concentration sufficient to treat a dermatological condition. As such, in some embodiments, a topical formulation described herein may decrease, eliminate or prevent the dermal inflammatory condition, suppress inflammatory edema formation and/or reduce an itch-response. Such effects may be evaluated clinically, objectively and/or subjectively by a health care professional, a treatment subject or an observer. Additionally, embodiments of the present invention may also treat the dermatological condition without significant side effects.

The product is intended for treatment of skin or open body cavities in man or in animals.

In some embodiments, the skin may be detached from a subject or may be a sample of skin, a skin graft or artificial skin.

In some embodiments, the present invention provides a kit including an

aminoguanidine compound and a pharmaceutically acceptable carrier system packaged together to provide an anhydrous topical formulation. The kit may include other items useful in the handling, preparation and use of the formulation as well as instructions for use of the same.

The present invention will now be described with reference to the following examples. It should be appreciated that these examples are for the purpose of illustrating aspects of the present invention, and do not limit the scope of the invention as defined by the claims.

Examples

In this series of experiments, the technical features of the compositions are

demonstrated. The assay method referenced is a HPLC UV method for determination of the active compounds in creams, solutions and suspensions.

Solubility testing

Solubility testing was performed in Eppendorf low bind test tubes by adding a surplus of the test compound into the solvent followed by stirring by a magnetic bar or by rocking the test tube for 1 hour. The suspension was centrifuged and the supernatant was analysed for the test compound. The compounds were analyzed using high pressure liquid chromatography

(HPLC). The HPLC system was equipped with a CI 8 reversed phase column, a gradient pump and a UV detector. Elution was carried out using a linear gradient of an ammonium acetate buffer and acetonitrile. The compounds were monitored at 280 nm.

The solubility of three aminoguanidines (Compound 1, 2 and 3) was tested in selected solvents.

Compound 1 - ((E)-2-(2=chloro-3,4-dimethoxybenzylidene)-N- methylhydrazinecarboximidamide)

Compound 2 - ((E)-2-(2-chloro-3,4-dimethoxybenzylidene)-N- phenylhydrazinecarboximidamide)

Compound 3 - (( -2-(4-methoxybenzylidene)-N-phenethylhydrazinecarboximidamide)

In vitro testing

The in vitro skin penetration was studied in Bronaugh cells for 3 hours, 6 hours and 24 hours, respectively. Bronaugh cells refer to cells having two compartments separated by a skin membrane where the formulation is charged on the upside and receptor fluid is pumped through the lower compartment of the cell. The pump speed was 1.2 ml/h, which means that about 29 ml is used after 24 hours and 3, 6 and 24 hours fractions are collected by an auto- sampler. Several membranes have been tested, frozen pig ear skin and fresh human and pig ear skin. We have decided to use fresh pig ear skin due to the easy access and similarity with human fresh skin. Irrespective of the skin type used, the skin was dermatomed to 0.5 to 1 mm in thickness. A dermatome refers to equipment by which you can plane the skin. This is done from inside which means that the stratum corneum and epidermis are intact. A receptor solution was selected in the experiment described in example 1 , consisting of phosphate buffered saline at pH 7.4 with 2 % of a surfactant, PEG-20 oleyl ether, added. With the inclusion of the surfactant, the solubility of the active compounds increases and is sufficient for penetration studies in an aqueous environment.

The temperature of the water bath was set to 34.5°C giving a cell temperature of 32.0°C. About 50 μΐ, corresponding to about 50 mg of each formulation was charged using a plastic Pasteur pipette.

After the termination of the experiment, the donor compartment was washed and the wash fluid was collected for analysis. The cells were opened and the skin membranes split into dermis and stratum corneum/epidermis. he separated tissue was homogenized and analysed for content of aminoguanidine.

Example 1.

The solubility of the aminoguanidines in water-based solvents was low. In phosphate buffer at pH 7.4, the solubility was less than 0.5 mg/ml and when solubilising agents and/or surfactants were used, the solubility was increased to about 1 mg/ml. Details are found in Table 1.

Table 1. Results solubility in PBS buffer, -1% active, stirrer

Actives PBS PBS+2% surfactant*

ε/mi με/ml

Compound 1 547 942

Compound 2 5.7 443

Compound 3 55 1240

The surfactant is PEG-20 oleyl ether.

Example 2.

Solubility of aminoguanidines in polar solvents was tested. The solubility of one aminoguanidine, Compound 1, in the form of its hydrochloride salt or base form were tested in dipropyleneglycol, triethyl citrate, propylene carbonate and acetonitrile by adding a surplus active and stirring for 1 hour followed by centrifugation and analysis by HPLC-UV. The results are presented in Table 2. Table 2. Results of solubility testing of salt, hydrochloride and base of Compound 1.

The uncharged base form of the aminoguanidine, here exemplified by compound 1 base, is soluble in the polar solvents. In comparison, the salt form, the hydrochloride, has a solubility of < 10 mg/g.

Example 3.

In this example, we tested the solubility at the 1 % level of 3 aminoguanidines with different properties. In general, the polar solvents were effective jn dissolving the aminoguanidines while less polar ones such as isopropylmyristate were less effective.

Table 3. Results of solubility testing % (w/w) - 1% active weighed in, rocked for lh

Example 4.

Solubility testing of Compound 1, Compound 2 and Compound 3 was performed on the 5 % level and triethyl citrate was superior in dissolving Compound 3 while

dipropyleneglycol was the most effective solvent for Compound 1. Table 4. Results of solubility testing 5 - % (w/w) - 5% active charged, stirrer lh

Example S. Solubility testing of mixtures

Table 5. Results of solubility of Compound 1 with solvent mixtures % (w/w) 5% active charged, stirrer 2h

Table 6. Results of solubility of Compound 2 and Compound 3 in solvent mixtures % (w/w) 5% active charged, stirrer 2h

Example 6. Formulation of cream

In order to generate a formulation acceptable by the subject to be treated, a polar solvent in oil emulsions were manufactured. Physical stability was tested by storage at ambient temperature and in a refrigerator. The compositions below were manufactured by separate mixing and heating of phase A and B to 75°C followed by mixing of phase A and B, homogenisation for 5 minutes and cooling to ambient temperature. In this experiment we have tested the use of a nonaqeous cream composition different solvent compositions, propylene glycol/glycerol

Table 7. Change of propylene glycol/glycerol ratio

Raw material 6:1 6:2 6:3

Phase A

Cithrol™DPHS 3.00 3.00 3.00

Crodamol™ 10.50 10.50 10.50

GTCC

Paraffin oil 31.17 31.17 31.17 Macrocrystalline 1.71 1.71 1.71

wax

Aerosil R972 3.45 3.45 3.45

Phase B

Compound 1 1.00 0.7 0.7

Propylene glycol 43.68 19.35 19.35

Glycerol 4.40 29.03 29.03

Lactic acid 0.59 0.59 0.59

Tween 20 0.50 0.50 0.50

Total % (w/w) 100 100 100

Separation RT Yes No No

Separation Cool Yes No No

The products containing higher amounts of glycerol, 6:2 and 6:3, did not separate over night in RT or in refrigerator.

Example 7. Formulation of nonaqueous emulsions

In order to generate a formulation preferred and acceptable by the subject to be treated, a polar solvent in oil emulsions were manufactured. Physical stability was tested by centrifugation and storage at ambient temperature and in a refrigerator. The compositions below were manufactured by separate mixing and heating of phase A and B to 75°C followed by mixing of phase A and B, homogenisation for 5 minutes and cooling to ambient temperature.

Table 8. Compositions of formulations studied.

Ingredients 7.1 7.2

Batch number

Phase A

Cithrol™ DPHS 3.01 3.01

Crodamol™ GTCC 10.53 10.53

Paraffin oil 31.29 33.01

Microcrystalline Wax 1.72 0

Aerosil R972 3.45 3.45

Phase B

Compound 1 1.00 1.00

Propylene glycol 19.16 19.16

Glycerol 29.25 29.25

Lactic acid (90%) 0.59 0.59

Total 100.00 100.00 The two formulations were exposed to 8 hours of centrifugation, from 500 up to 3000 rpm without separation. The formulations were also exposed to slow stirring at <25 rpm for 48 hours without separation.

Example 8. Formulations using multiple emulsifiers

In this example we have investigated the effect of the chemical nature of the emulsifier(s) and the HLB of the emulsifier(s) on the emulsion stability. We have selected the following emulsifiers:

Table 9. Tested emulsifiers

Emulsifier HLB Chemical name

Chithrol™DPHS-SO-(MV) 5.5 PEG 30 dipolyhydroxystearate

Span 40-PW-(MV) 6.7 Sorbitan monopalmitate

Span 60 4.7 Sorbitan monostearate

Span 83-LQ-(MV) 3.7 Sorbitan sesquioleate

Poly(ethylene glycol) dioleate 7.5 Poly(ethylene glycol) dioleate

Span™ 20-LQ-(MV) 9.0 Sorbitane monolaurate

Myrj S8-SO-(MV) 1 1.1 Polyoxyethylene (8) stearate

Span 83, sorbitan sequioleate and polyethyleneglycoldioleate both contain usaturated acids, while the rest are aliphatic. The formulations were manufactured as in example 6 and the physical stability was tested.

Table 10. Compositions

Propylene glycol 19.16 19.16 19.16 19.16 19.16 19.16 19.16

Glycerol 29.25 29.25 29.25 29.25 29.25 29.25 29.25

Lactic acid 0.59 0.59 0.59 0.59 0.59 0.59 0.59

HLB (theoretical) 5.5 5.7 5.6 9 11 6.7 4.7

Total % (w/w) 100.0 100.0 100.0 100.0 100.0 100.0 100.0

Separation No No Yes Yes Yes No No

The compositions above were manufactured by separate mixing and heating of phase A and B to 75°C followed by mixing of phase A and B₅ homogenisation for 5 minutes and cooling to ambient temperature.

The formulations were tested for separation immediately after manufacture. The result was that preparations 7:3, 7:4 and 7:5 separated quickly. The preparations 7:3 and 7:4 include surfactants with unsaturated fatty acid chains while 7:4 and 7:5 have high HLB values. The rest of the formulations were stable. Suitable emulsifiers for this type of formulation should, alone or in combination give rise to an HLB of more than 4 and less than 9 and have saturated fatty acid chains.

Example 9. In vitro skin penetration of solutions of benzylideneguanidines.

An in vitro study was performed, using frozen pig ear skin as membrane. The amount of active component penetrating through the skin was low. However, in the dermis, the site of the intended receptors, effective concentrations of the active component were found.

Table 11. Average concentrations in pig ear dermis after 24 hours exposure using frozen skin

Cone, dermis

Formulation

(μβ/β)

1% Compound 1 in triethyl citrate (n=5) 16.2

1% Compound 1 in dipropylene glycol (n= =5) 7.5

5% Compound 1 in dipropylene glycol (n= =2) 294

5% Compound 1 in dipropylene glycol (n= =4) 76

2.8% Compound 1 in propylene glycol (n= =5) 161

The target concentration in cell suspensions is less than 1 μg/g. In this experiment we used frozen pig ear skin which tends to give high tissue concentrations.

Example 10. dermal concentrations of Compound 1 in live pig after 1 week of treatment.

4 domestic pigs were exposed to Compound 1 cream on the back for 6 hours per day during 7 days. After 6 hours the cream was washed off. After sacrifice of the animals skin samples were analyzed for Compound 1 using the method described earlier. Two creams were tested, one containing 0.5 % of Compound 1 and one 1.2 % (Table 12). Four animals, two male and two female, were included in the study and the results are listed in Table 13.

Table 12. Composition of formulations used in the test (Example 10).

Raw material 0.50% 1.20%

A

Cithrol™DPHS 8.94 7.00

Crodamoi™ GTCC 10.33 10.33

Paraffin oil 24.17 24.17

Microcrystalline Wax 1.71 1.71

Aerosil R972 3.45 3.45

B

Compound 1 0.50 1.20

Propylene glycol 43.10 42.40

Glycerol 4.40 4.40

Lactic acid 0.59 0.59

Tween 20 2.71 4.65

BHA 0.10 0.10

Total % (w/w) 100.00 100.00

Table 13. Concentration of Compound 1 in dermis from pig.

Animal no./ Compound 1 Dermis weight Tissue Average ^g/ ) Species/ Dose (re) (g) Concentration

S2-2M 0.5 % 0.900 0.0832 10.81

S1-3M 1.2 % 1.325 0.0912 14.52 12.14

S2-3M 1.2 % 1.502 0.154 9.75

S1-5F 0.5 % 0.507 0.1035 4.90 5.62

S2-5F 0.5 % 0.487 0.0769 6.33

S1-6F1.2 % 2.172 0.1813 11.98 16.25

S2-6F 1.2 % 2.525 0.123 20.53

Treatment with a cream containing 1.2 % generates dermal concentrations of 9.75 to 20.53 l^g/g_> which is well above the concentration required for pharmacological effect, 0.3 μΒ/g.

Example 11. In vitro penetration of Compound 1 into dermis from two formulations containing different amount of solvents.

The penetration of Compound 1 was tested in fresh pig ear skin from two formulations using different solvent systems. The compositions below were manufactured by separate mixing and heating of phase A and B to 75°C followed by mixing of phase A and B, homogenisation for 5 minutes and cooling to ambient temperature.

Table 14.Formulations studied.

Table 15. Concentration of Compound 1 in dermis after 24 hours of exposure ^g/g)

Product Concentration in dermis ^g/g) Standard deviation ^g/g)

13 A 1.2% 11.52 5.55

13 B 1.0 % 14.39 2.63

13 B 1.0 % half dose 5.72 2.28

It should be noted that very little Compound 1 has passed the membrane in these experiments. This means that very little exposure to systemic concentrations is expected from topical administration, i.e., about 0.1 % of the dose.

Example 12. Penetration of Compound 1 into dermis from the invented formulation compared with a solution and a suspension.

In this example, the in vitro skin penetration from three formulations was studied, one t formulation of interest, onesuspension and one solution of compound 1. In table 16, the compositions are presented.

Table 16. Formulations to be studied.

Paraffin oil 31.29 - -

Macrocrystalline Wax 1.72 - -

Aerosil R972 3.45 -

Phase B ,„_{. .} ^■ ,' _t

Compound 1 1.00 2.77 2.77

Propylene glycol 19.16 - 97.23

Glycerol 29.25 - -

Lactic acid (90%) 0.59 - -

Petrolatum, white (USP) - 97.23 -

Total 100.00 100.00 100.00

Manufacturing of the invented cream was done by heating phase A and B separately to 75 °C and mixing using a homogenizer. Cooling to ambient temperature was performed and the product was packed in polyethylene tubes. The solution was made by dissolving Compound 1 in the solvent and the suspension by mixing Compound 1 in solid form in petroleum jelly. The in vitro penetration experiment was performed according to the earlier description and the results are presented in figure 1. It is interesting to note that the dermal content of compound 1 is higher when administered in the form of the invented cream than if administered as a solution or a suspension.

Claims

THAT WHICH IS CLAIMED IS:

1. A topical formulation comprising an aminoguanidine compound in a pharmaceutically acceptable carrier system, wherein the topical formulation is an anhydrous formulation.

2. The topical formulation of claim 1, wherein the pharmaceutically acceptable carrier system comprises a continuous phase and a discontinuous phase.

3. The topical formulation of claim 2, wherein the continuous phase comprises lipids, nonpolar solvents, fatty acids, thickening agents, stabilising agents and/or emulsifiers.

4. The topical formulation of claim 2 or 3, wherein the discontinuous phase comprises about 20 to about 90 % of the topical formulation, more preferably about 30 to about 85 % and most preferably about 35 to about 70% of the topical formulation.

5. The topical formulation of any of claims 2-4, wherein the discontinuous phase comprises the aminoguanidine compound, polar solvents and/or surfactants.

6. The topical formulation of any one of claims 1-5, wherein the aminoguanidine compound has the following structure of formula I:

I and stereoisomers, tautomers and pharmaceutically acceptable salts, esters or prodrugs thereof, wherein:

R'-R⁵ are independently H, Q.s-alkyl, OH, Ci.₅-alkoxy, SMe, SEt, CF₃, OCF₃, F, CI, Br, I, CN, S(0)Me, S(0)₂Me, S(0)₂NMe₂, NMe₂, NHC(0)H, NHC(0)Me, or CONMe₂, optionally including two adjacent groups, R -R², R²-R³, R³-R⁴, or R⁴-R⁵, representing OCH₂0, OCH₂CH₂0 or SCH₂CH₂S;

X is absent or CH₂, (CH₂)₂, (CH₂)₃, CH=CH, CH₂CH=CH or CH=CHCH₂;

Y is absent or CH₂₎ (CH₂)₂, (CH₂)₃, CH₂)₄ or (CH₂)₅;

A is H, Ph or a 5-6 membered hetero-aromatic ring containing 1-3 hetero-atoms

independently N, O or S; and

B is H, Me, Et, Ph, benzyl or OH.

7. The topical formulation of any one of claims 1-6, wherein the aminoguanidine compound has the following structure:

8. The topical formulation of claim 7, wherein the topical formulation further comprises propylene glycol and/or dipropyleneglycol.

9. The topical formulation of any one of claims 1-6, wherein the aminoguanidine compound has the following structure:

10. The topical formulation of claim 9, wherein the topical formulation further comprises propylene glycol and/or triethyl citrate.

11. The topical formulation of any one of claims 1-6, wherein the aminoguanidine compound has the following structure:

12. The topical formulation of claim 1 1 , wherein the topical formulation further comprises propylene glycol and/or triethyl citrate.

13. The topical formulation of any one of claims 1-12, wherein the topical formulation comprises penetration enhancers, pH modifiers, sun blocking agents,

antimicrobial agents and/or antioxidants.

14. The topical formulation of any one of claims 1-13, wherein the topical formulation is a gel, ointment, lotion, emulsion, cream or foam.

15. A kit comprising the topical formulation of any one of claims 1-14, and optional instructions for use of the same.

16. A method of treating a dermatological condition comprising applying the topical formulation of any of claims 1-14 to the skin of a subject to treat a dermatological condition.

17. The method of claim 16, wherein the dermatological condition is dermatitis, psoriasis, pruritus and/or eczema.

18. A method of delivering an aminoguanidine compound to the skin of a subject, the method comprising administering the topical formulation of any one of claims 1-14 to the skin of the subject.