WO2011061155A1

WO2011061155A1 - Antifungal formulations and their use

Info

Publication number: WO2011061155A1
Application number: PCT/EP2010/067501
Authority: WO
Inventors: Jerome Aubert
Original assignee: Bayer Consumer Care Ag
Priority date: 2009-11-17
Filing date: 2010-11-15
Publication date: 2011-05-26

Abstract

The inventions relates to a film forming formulation comprising bifonazole, a process for its preparation and its use in the treatment of fungal infections or infections related to fungal infections of the skin.

Description

Antifungal Formulations and their Use

The invention relates to a topical drug formulation comprising bifonazole and its use for the treatment of fungal infections, such as e.g. Athlete's Foot.

Bifonazole is an antimycotic agent and was first synthesized at Bayer Pharmaceuticals in 1974. Its systematic name is l -[phenyl(4-phenylphenyl)methyl]- lH-imidazole (formula I) and it forms colorless, odorless, and tasteless crystals.

Bifonazole is a broad-spectrum imidazole antimycotic with excellent in vitro activity against dermatophytes, yeasts, moulds, and dimorphic fungi among those Trichophyton spp., Epidermophyton spp., Microsporum spp., Candida spp., Malassezia spp., Trichosporon spp., Cryptococcus neoformans, Aspergillus spp., Histoplasma spp., Coccidioides immitis, Paracoccidoides brasiliensis, Geotrichum candidum and Blastomyces dermatitidis,, and certain bacteria e.g. Corynebacterium spp., Streptococcus species, Staphylococcus spp.

Bifonazole can only be used as a topical antimycotic due to its fast metabolism, generating metabolites without antimycotic activity when administered orally. The substance is minimally absorbed after topical application of 1 % formulations. Concentrations in blood serum usually remain below 5 ng/mL, even after daily application over two to three weeks of a skin area of about 200 cm². The topical applications are well tolerated. Skin irritations and allergic reactions are rarely observed and seem to be mainly due to the galenic formulation i.e. its excipients and the skin disease to which it is applied (e.g., fatty preparations on acute, inflammative lesions).

Bifonazole is available in several galenic formulations, in general in a concentration of 1 % as (spray) solutions, powders, gels, ointments, and cream preparations indicated for the treatment of superficial fungal infections. For example, fungal infections caused by dermatophytes result in infections of the skin, scalp, and nails due to their ability to utilize keratin. Dermatophytes colonize the keratin tissues and cause fungal infections, e.g. known as tinea and ringworm. The organisms are transmitted by either direct contact with infected host (human or animal) or by direct or indirect contact with infected exfoliated skin in shared cosmetic items, textiles or wet rooms.

Depending on the species, the organism may be viable in the environment for up to 15 months. There is an increased susceptibility to infection when there is a pre-existing injury to the skin, such as recently healed scars or burns, or when the skin is submitted to physical and mechanical strain, excessive temperature and humidity.

Athlete's Foot (tinea pedis, tinea pedis interdigitalis) is a one of the cutaneous fungal infection caused by dermatophytes that results in scaling, flaking, and itch of affected foot areas, especially the skin between the toes. Dermatophytes may also cause infections on other areas of the body, i.e. under toenails (onychomycosis or tinea unguium), on the skin of the body (tinea corporis), limps, hands (tinea manuum, tinea manuum interdigitalis) and finger nails (onychomycosis or tinea unguium), on the groin (tinea cruris, tinea inguinalis), on the scalp and hair (tinea capitis) as well as in the face.

Other fungal infections may be caused by yeasts or moulds. These include pityriasis versicolor by Malassezia furfur or superficial skin infections with Candida species. Some infections caused by certain bacteria such as Corynebacterium minutissimum resulting in an infection called erythrasma, may be also associated with acne, and can be positively influenced by antifungal treatment.

The time line for cure may be in general long, often up to four weeks or longer. Topical treatment with bifonazole 1% formulations is currently recommended to last for four weeks. However, because itching associated with the infection subsides quickly, patients may not complete the courses of therapy as prescribed. Furthermore, the galenic formulations of bifonazole 1% currently available are to be applied once daily, however, they are often affected by the everyday life of the patient, e.g. can be easily sweated off or washed off when the patient pursues sporting activities or has to perform physical work. These formulations are recommended to be applied once daily for four weeks.

Film-forming preparations are generally known in the art.

WO2007/031753 describes film forming formulations comprising a pharmaceutical, a solvent, a film forming agent and a propellant. Bifonazole is only mentioned as one possible active compound in a list of antifungals that were just one class of active ingredients among others such as antipruritics, anaesthetics, antihistamines, corticosteroids, psoriasis and acne agents, dermatological drugs, antibacterials, antivirals, antiperspirants, and transdermal drugs.

US 2003/0152611 discloses pharmaceutical compositions for transdermal administration comprising a cellulosic polymer matrix, an NSAID, an absorption promoter, water and a solvent forming matrix. No antifungal agents are mentioned.

US-A-6325990 provides lipophilic vitamins etc. in the absence of water and in the presence of adhesive polysiloxane, an absorption promoter and a volatile solvent, sprayable from an aerosol can. Antimycotic agents with ketoconazole as example are mentioned, however, no information on pharmacological activity is provided.

WO 00/045795 provides medicinal spray compositions comprising a medicament in a volatile vehicle and one or more film-forming polymers. Antifungal agents are not mentioned.

It is therefore the objective of the present invention to provide a new bifonazole formulation for the treatment of fungal infections of the skin that has improved properties with respect to wear while still showing good stability, has an improved release profile into the stratum corneum with good activity on pathogenic organisms and more convenient application while being economically favourable.

The present invention is thus directed to a monophasic and saturated film forming formulation comprising bifonazole, a solvent and at least one cosolvent, a film forming agent, a propellant, and optionally at least one further excipient and/or fragrance, said film forming formulation for the treatment of fungal infections of the skin and its use for the preparation of a medicament for the treatment of fungal infections of the skin.

The formulation according to the invention was surprisingly found to have improved wear properties when compared to bifonazole formulation of the state of the art, i.e. the film is partially water resistant and resistant to friction and still ensures enhanced bifonazole delivery even when the skin is exposed to a short shower, sweat or to humid ambient conditions. Furthermore, the film formulation according to the invention is capable of delivering approximately twice the amount of bifonazole into the stratum corneum in comparison with a solution spray formulation and eight times the amount of a standard cream formulation as proved by in vitro experiments. The film formulation according to the invention while causing high bifonazole concentrations in the stratum corneum surprisingly does not lead to enhanced levels of bifonazole in the receptor medium representing the blood stream in the in vitro experiment. The film formulation when used for the preparation of a medicament in the treatment of fungal infections of the skin show enhanced activity against the pathogenic organisms and lead to reduced treatment times. The solvent according to the invention is capable of dissolving the bifonazole, the film forming agent, the propellant, and optionally at least one further excipient and/or fragrance to yield a monophasic solution. Its main function is to evaporate along with the propellant as the film forming formulation is applied and the film is formed on the skin in the concentrated remainder of the film forming formulation. According to the inventive formulation the solvent can be selected from the group of ethanol, isopropanol, aqueous methanol, aqueous ethanol and aqueous isopropyl alcohol (containing approx. 3 % water), acetone, ethyl acetate, or methylene chloride. Ethanol and isopropanol are preferred solvents. Ethanol is particularly preferred, and it is a particular advantage of the present invention that considerably smaller amounts of ethanol need to be used for the dissolution of bifonazole, by comparison with the ethanolic spray formulation of the art.

The amount of solvent chosen for the present invention lies between 10 and 40% w/w of the formulation, preferably between 12 and 35%, most preferably between 15 and 30%.

The at least one cosolvent is present in a smaller amount as compared to the solvent. It is also capable of dissolving the major ingredients of the film forming formulation, and its presence influences the properties of the resulting film forming formulation favourably, e.g. as a penetration enhancer, as a possible plasticizer or by influencing the properties in a less distinct way such as the overall tackiness or the regularity of the film surface of the film forming formulation. The at least one cosolvent according to the inventive formulation may be an even better solvent for bifonazole than ethanol and can be selected from the group of benzyl alcohol, propylene glycol, polyethylene glycol, propylene carbonate, dimethyl sulphoxide, glycerin, isopropyl myristate, tetraglycol, cyclomethicone, N-methyl-2 pyrrolidone, Transcutol®, which is diethylene glycol monoethyl ether or Arlasolve®, which is dimethy isosorbide, and oleic acid. Especially preferred is the use of benzyl alcohol, isopropyl myristate, Transcutol® or Arlasolve®, alone or as mixture. The amounts of the cosolvent are chosen to be between 0,05 % to 7% each, preferably between 1 and 4%, most preferably between 2,5 and 3%.

In another embodiment, benzyl alcohol is present in an amount of 0,05 to 5 %, preferably 0,92 to 4 %, most preferably 1 to 3%.

In another embodiment, Transcutol® is present in an amount of 0,05% to 5%, preferably 0,25 to 3%, most preferably in an amount of 0,75 to 1,5%.

Arlasolve® is preferably present in an amount of 0,05% to 5%, preferably 0,25 to 3%, most preferably in an amount of 1%. Isopropyl myristate is present in an amount of 0,1 % to 5, preferably between 1 and 4%, most preferably between 2,5 and 3%.

The film-forming agent may suitably be a polymer approved for topical administration, such as polyvinyl pyrrolidone (PVP) or polyvinyl alcohol (PVA), acrylic polymers or copolymers, methacrylate polymers and copolymers, poly (vinyl acetate), and cellulose based polymers and co-polymers. Especially preferred are polyacrylates or methacrylates, especially preferred Eudragit® RS PO which is defined to be a copolymer of acrylic and methacrylic acid esters with a low content in quaternary ammonium groups. The ammonium groups are present as salts and make the polymers permeable. The average molecular weight is approx. 150,000. The film forming agent may be present in an amount of 1 to 10%, preferably 5 to 8%, most preferably in an amount of 6-7%.

The propellant may be a non-combustible hydro fluoroalkane HFA, such as HFA 134a or HFA 227 a, respectably. Preferably, HFA 134a is used. The amounts of the propellant are chosen to be between 50 and 90 %, preferably between 55 and 75%, most preferably between 55 and 65%.

In another embodiment of the invention, optionally, at least one further excipient is present which may have a further favourable effect on the film forming formulation. These effects may lie in the field of consumer needs such as aesthetic requirements (gloss, colour, etc.) or may add to the performance of the film forming formulation such as e.g. further anti-nucleating agents to further stabilize the resulting supersaturated system. The film forming formulation of the present invention can thus be further combined with any other suitable additive or pharmaceutically or cosmetically acceptable excipient. Such additives include any of the substances already mentioned, as well as any of those used conventionally, such as those described in Remington: The Science and Practice of Pharmacy (Gennaro and Gennaro, eds, 20th edition, Lippincott Williams & Wilkins, 2000); Theory and Practice of Industrial Pharmacy (Lachman et al., eds., 3rd edition, Lippincott Williams & Wilkins, 1986); Encyclopedia of Pharmaceutical Technology (Swarbrick and Boylan, eds., 2nd edition, Marcel Dekker, 2002). These can be referred to herein as "further excipient" to indicate they are combined with the other ingredients and can be administered safely to a subject for therapeutic purposes.

In a preferred embodiment, fragrances may also be present in the film forming formulation, they are known by a person skilled in the art and are chosen from plant extracts and essential oils like eucalyptus, lemon, orange, cinnamon, mint, tangerine, grapefruit, spearmint, peppermint, clove, or rosemary oil, or synthetic fragrances such as vanilline, especially preferred is eucalyptus oil or mint oil in an amount of 0,1 to 1,2%, preferably 0,2 to 0,5%, especially preferred is eucalyptus oil. In another embodiment of the film forming formulation the solvent is present in an amount of 10 to 40% and the at least one cosolvent is present in an amount of 0,05 to 7% each, the film forming agent is present in an amount of 1-10%, the propellant is present in an amount of 50 to 90%, and optionally fragrance is present in an amount of 0,1 to 1,2%.

In a further embodiment of the invention the solvent is present in an amount of 12 to 35% and the cosolvent is present in an amount of 1 to 4% each, the film forming agent is present in an amount of 5- 8%, the propellant is present in an amount of 55 to 75%, and optionally fragrance is present in an amount of 0,1 to 1,2%.

In a preferred embodiment of the inventive film formulation comprising bifonazole, the solvent is ethanol or isopropyl alcohol in an amount of 15 to 30% and the cosolvent is benzyl alcohol, isopropyl myristate, Transcutol®, Arlasolve® or a mixture thereof in an amount of 0,05 to 7% each, the film forming agent is polyvinyl pyrrolidone (PVP), polyvinyl alcohol (PVA), acrylic polymer or copolymer, methacrylate polymer or copolymer, poly (vinyl acetate), cellulose based polymers or co-polymer, or Eudragit® RS PO in an amount of 6-7%, the propellant is HFA 134a or HFA 227a in an amount of 55 to 75%, and optionally eucalyptus oil , menthol or limonene is chosen in an amount of 0,2 to 0,5% as a fragrance.

The formulations of the invention are capable of forming a film on topical administration, typically to the skin. In particular, the majority of the propellant component of the formulation will normally evaporate almost immediately, thereby concentrating the remainder of the formulation. The film forming agent may be such as to form a film after the evaporation of a portion of the solvent.

It will be appreciated that the formulation will be saturated with bifonazole and be monophasicjmder conditions of use. In this respect, these requirements relate to the formulation immediately prior to dispensing, such as when in an aerosol canister.

A preferred film formulation according to the invention provides a saturated, monophasic solution of bifonazole in the formulation and upon film formation - when topically administrered - exhibits passive diffusion fluxes greater than those predicted by Fick's law. The term 'monophasic' is used to indicate that the formulation does not contain undissolved bifonazole, and also that there is only the one liquid phase, and not a colloid or micro-colloid, for example. There is only one phase, and that phase is liquid.

Bifonazole should be present in a saturating amount in the formulation In this respect, it will be appreciated that a formulation held at a higher temperature will require greater amounts of bifonazole in order to be saturated, for most solvents. In this regard, the monophasic requirement remains important, but the saturation may be determined by whether the formulation, when applied to a test membrane such as disclosed in WO2007/031753 A2, transcends Fick's law or only provides a flux at or below that predicted by Fick's law. In the meaning of the invention a substantially saturated film formulation is defined, wherein at least 80% of that amount of the bifonazole needed to achieve saturation is present. This amount is preferably at least 90%, and more preferably 95%. At the temperature of use, it is preferred that the formulation be as close to saturated as possible, while remaining monophasic. Supersaturated solutions are also included, but these are generally less preferred, as they are not generally stable, and have short shelf-lives before ceasing to be monophasic.

It is preferred that the amount of bifonazole present be as close to full saturation as possible, but many monophasic solutions are not stable at such high concentrations. In such cases, the addition of the at least one further excipient that functions as antinucleating agents, such as PVA when PVP is used as the film- forming ag ent or methyl c e llulose , ethyl c ellulos e, hydroxyalkylce llulose s , such as hydroxypropylmethylcellulose and hydroxypropylcellulose, glycol esters, polyacrylic acid, and derivatives thereof, may be advantageous, as may a slight drop in saturation, down as far as 80%, which is considered to be a saturating amount for the purposes of the present invention.

A preferred film formulation according to the invention provides a saturated and monophasic solution of bifonazole in the formulation, which is stable and allows for shelf-lives of up to three years, preferably up to 2,5 years, most preferably up to 2 years without degradation of the active compound or deterioration of the release profile of the resulting film.

One of the advantages of the present invention lies in the combined high saturation levels and the use of propellant. The propellant is typically a highly volatile liquid with a low boiling point, such that it can force the formulation from a dispenser. Evaporation is almost instantaneous and the boiling during transfer from the dispenser to the site of administration has the effect of causing the evaporation of a substantial amount of the solvent, which are as defined below, but is typically ethanol or isopropyl alcohol. Thus, the solvent is preferably a volatile solvent, and the vigorous decompression of the propellant causes the disruption and loss of solvent by evaporation. This loss can be up to 50% and even higher. The effect of the loss of solvent is to drive the remaining solution towards supersaturation. It is for this reason that saturation levels of at least 80% are necessary, as levels much below this tend to result in saturated solutions rather than supersaturated solutions, and little advantage is to be seen. At 80% and above, levels of supersaturation of up to 2.5 times saturation may be achieved as calculated in WO2007/031753 A2, with the concomitant ability to drive permeation across the stratum corneum. Lower levels of saturation require greater loss of solvent before supersaturation is achieved. Furthermore, the use of the propellant and its effect on the enhanced solvent evaporation leads to the fast formation of the film on the skin enabling the patient to soon resume his daylife activities after administration of the film forming formulation.

It was surprisingly found that the presence of benzyl alcohol and transcutol in the film forming formulation led to an increased amount of bifonazole in the stratum corneum when compared to film formulation without either of the compounds or only comprising one of them. The effect of the two compounds was observed to be more than additional as can be seen in Fig. 1. A large amount of bifonazole delivered from the inventive film formulation into the stratum corneum leads to an improved pharmacological activity on the pathogenic organisms as present therein. It is preferred that benzyl alkohol and Transcutol® are present in amount of 0,05 to 5% and 0,05 to 5%, respectively, preferably in an amount of 0,92 to 4% and 0,25 to 3%, especially preferred benzyl alcohol in an amount of 1 to 3 % and Transcutol® in an amount of 0,75 to 1,5 %,.

In another embodiment of the invention, isopropyl myristate is present as a cosolvent in the film forming formulation. These formulations surprisingly exhibit improved wear properties with respect to tackiness without affecting the film's water and friction resistancy and advantageously can be administered more easily by the patient. Isopropyl myristate is favourably present in an amount of 0,1 to 5%, preferably 1 to 4%, most preferably 2,5 to 3%.

In another embodiment of the invention, a second antifungal agent may be present in the film forming formulation on the prerequisite that the film forming formulation will still be monophasic and saturated_^,. The second antifungal agent may be selected from the group consisting of terbinafine, naftifine, butenafine, clotrimazole, econazole, isoconazole, ketoconazole, miconazole, oxiconazole, sertaconazole, sulconazole, tioconazole, itraconazole, nystatin, caspofungk^ tolnaftate, terconazole, amorolfine, ciclopirox and undecylenic acid-in particular terbinafine-, and topically acceptable salts of any of said compounds.

In another embodiment of the invention, additionally to bifonazole or bifonazole and the second antifungal agent another local anaestethic e.g. lidocaine or xylocaine or anti-itching drug is selected from the group consisting of benzydamide, crotamicron, strontium and its salts, corticosteroids e.g. hydrocortisone and topically acceptable salts thereof. A further object of the present invention is a process for the preparation of the film forming formulation, comprising the steps of

1. Preparation of a bulk premix comprising bifonazole, a solvent and at least one cosolvent, a film forming agent, and optionally at least one further excipient and/or fragrance by thorough mixing

2. Addition of the propellant directly into the film forming formulation.

In another embodiment of the invention, bifonazole is dissolved by careful mixing with the solvent, at least one cosolvent, and optionally at least one further excipient and/or fragrance in a suitable vessel. The film forming agent is added slowly to the vessel with constant stirring, until a monophasic viscous solution is formed. The bulk solution is filled on aerosol filling equipment into the suitable packaging means, preferably canisters, more preferably aluminium canisters, suitable closure caps, e.g. valves, are crimped on and the propellant is added to yield a monophasic and saturated solution. A typical formulation preparation process is described in Example 1.

Bifonazole is readily soluble in the stated solvents and at least one cosolvent while all other excipients are completely miscible with each other. The risk of any loss of solvent is mitigated by manufacture in an enclosed environment. A homogenous solution is ensured by final mixing and subsequently confirmed by analysis of the bifonazole content in the finished formulation as filled in the packaging means. The aerosol filling and propellant charging is performed on standard equipment such as a dosing system Flexicon and crimping system Pamasol.

A further aspect of the present invention is the film forming formulation comprising bifonazole, a solvent and at least one cosolvent, a film forming agent, a propellant, and optionally at least one further excipient and/or fragrance as defined above for the treatment of fungal infections and infections related to fungal infections of the skin and the use of the film forming formulation comprising bifonazole, a solvent and at least one cosolvent, a film forming agent, a propellant, and optionally at least one further excipient and/or fragrance as defined above for the preparation of a medicament for the treatment of fungal infections and infections related to fungal infections of the skin. In a preferred embodiment, the film formulation itself will be used as the medicament without further modification.

The film formulation or a medicament comprising the film formulation is capable of delivering approximately twice the amount of bifonazole into the stratum corneum in comparison with a solution spray formulation and eight times the amount of a standard cream formulation as proved by in vitro experiments making it especially suitable for the treatment of the above mentioned infections. It was surprisingly found that although high bifonazole concentrations in the stratum corneum are achieved by the inventive film formulation or a medicament comprising the film formulation, this does not lead to enhanced levels of bifonazole in the receptor department representing the blood stream in the in vitro experiment although penetration enhancers are present. Consequently, the inventive film formulation or a medicament comprising the film formulation ensures an excellent topical performance without causing a systemic strain on the patient.

Fungal infections according to the invention are understood to be infections caused by fungi or infections related to fungal infections. Fungal infections or infections related to fungal infection of the skin are those that are caused by dermatophytes, yeasts, moulds, and dimorphic fungi among those Trichophyton spp., Epidermophyton spp., Microsporum spp., Candida spp., Malassezia spp., Trichosporon spp., Cryptococcus neoformans, Aspergillus spp., Histoplasma spp., Coccidioides immitis, Paracoccidoides brasiliensis, Geotrichum candidum and Blastomyces dermatitidis„ and certain bacteria e.g. Cory neb acterium spp., Streptococcus species, Staphylococcus spp.

Especially important are cutaneous fungal infections caused by dermatophytes such as Athlete's Foot (tinea pedis, tinea pedis interdigitalis) that results in scaling, flaking, and itch of affected foot areas, especially the skin between the toes. Further infections comprise also those on other areas of the body, i.e. under toenails (onychomycosis or tinea unguium), on the skin of the body (tinea corporis), limps, hands (tinea manuum, tinea manuum interdigitalis) and finger nails (onychomycosis or tinea unguium), on the groin (tinea cruris, tinea inguinalis), on the scalp and hair (tinea capitis) as well as in the face.

Other fungal infections according to the invention may be caused by yeasts or moulds. These include pityriasis versicolor by Malassezia furfur or superficial skin infections with Candida species, e.g. Candida albicans, Candida glabatra, Candida tropicalis, Candida parapsilosis, Candida krusei or Candida neoformans.

Further infections caused by certain bacteria such as Corynebacterium minutissimum resulting in an infection called erythrasma, may be also associated with acne, and can be positively influenced by the inventive antifungal treatment.

The film formulation when used for the preparation of a medicament in the treatment of fungal infections of the skin show enhanced activity against the pathogenic organisms and lead to reduced treatment times.

The daily dosage of the bifonazole comprising film formulation or medicament may depend on various factors, such as sex, age, weight and individual condition of the patient and the type and gravity of the infection. The film formulation used in the prepared medicament may be applied once, twice or three times daily, preferred is the application twice daily, especially preferred once daily.

The administration of bifonazole via the inventive film forming formulation to the skin avoids the problem of hepatic first-pass metabolism, improves patient compliance due to improved wear and administrating properties, and reduced treatment times from up to 4 to 6 weeks.

A further aspect of the present invention is a method of treating a patient with a fungal infection or an infection related to a fungal infection of the skin, nail, and/or mucous membrane by administering the film forming formulation or the medicament comprising the film forming formulation at least once daily onto the affected area of the body.

The inventive film forming formulation when used for the preparation of a medicament can be applied for the treatment of mammals, especially humans but can also be applied to animals, including, but not limited to dogs, cats, cows, sheep, pigs, poultry and others.

The formulation according to the invention has the advantage of having improved wear properties when compared to bifonazole formulation of the state of the art, i.e. the film is water resistant and resistant to friction and still ensures bifonazole delivery even when the skin is exposed to a short shower, sweat or to humid ambient conditions. Furthermore, the film formulation according to the invention is capable of delivering approximately twice the amount of bifonazole into the stratum corneum in comparison with a solution spray formulation and eight times the amount of a standard cream formulation as proved by in vitro experiments. The film formulation according to the invention while causing high bifonazole concentrations in the stratum corneum does not lead to enhanced levels of bifonazole in the receptor compartment or receptor medium representing the blood stream in the in vitro experiment. The film formulation when used for the preparation of a medicament in the treatment of fungal infections of the skin show enhanced activity against the pathogenic organisms and lead to reduced treatment times compared to ca. four weeks and exhibit excellent stability both with regard to the active compound as well as with regard to release profiles of the resulting films.

Figures:

Fig. 1 shows the dependence of the permeation of bifonazole into the stratum corneum from the concentration of benzyl alcohol and transcutol in the film forming formulation as disclosed e.g. in Example A1-A3, A5-A8 and A13-A16.

Fig. 2 shows the concentrations of bifonazole in the different skin layers for several bifonazole formulations.

Fig. 3 shows the total amounts of bifonazole ( μg/cm², mean ± standard deviation) detected in the skin compartments and in the receptor fluid.

Fig. 4 shows the concentrations of bifonazole in the different skin layers for further bifonazole formulations.

Fig. 5 shows the total amounts of bifonazole μ g/cm², mean ± standard deviation) detected in the skin compartments and in the receptor fluid.

Fig. 6 shows the concentrations of bifonazole in the different skin layers for further bifonazole formulations in comparison with the Canesten® Extra solution spray formulation.

Fig. 7 shows the total amounts of bifonazole μ g/cm², mean ± standard deviation) detected in the skin compartments and in the receptor fluid in comparison with the Canesten® Extra solution spray formulation.

Fig. 8 shows the comparative in- vitro release profile of T = 0 months samples versus a T = 6 months samples stored at accelerated conditions 40°C/75%RH.

Examples

The invention is illustrated by the following examples without limiting it in its scope to said examples.

In Fig. 1 the dependence of the permeation of bifonazole into the stratum corneum from the concentration of benzyl alcohol and transcutol in the film forming formulation as disclosed in e.g. Examples A1-A3, A5-A8 and A13-A16 is shown.

In Fig. 2 the results of the in vitro investigation of the permeation behaviour of absorbed bifonazole ^g/cm², mean ± standard deviation) after 24 h in different formulations are shown. A9 absorbes within 24 h ca. twice the amount of bifonazole in the stratum corneum (full horny layer) as compared to the Canesten® Extra solution spray formulation and about 8 times as much as is absorbed from the Canesten ® Extra cream formulation.

In Fig. 3 the total amounts of bifonazole ^g/cm², mean ± standard deviation) detected in the skin compartments and in the receptor fluid. Although there are Arlasolve® and benzyl alcohol that are known to act as penetration enhancers present in the formulation A9, no enhanced level of bifonazole is detected in the receptor fluid representing the blood serum in the in vitro experiment.

In Fig. 4 and 6 the results of the in vitro investigation of the permeation behaviour of absorbed bifonazole ^g/cm², mean ± standard deviation) after 24 h in further formulations are shown.

Fig. 5 and 7 show the total amounts of bifonazole ^g/cm², mean ± standard deviation) detected in the skin compartments and in the receptor fluid for these respective formulations.

Fig. 8 shows the comparative in- vitro release profile of T— 0 months samples versus a T— 6 months samples stored at accelerated conditions 40°C/75%RH. This graph shows that despite of these accelerated conditions, the release profile remains remarkably similar to the initial value. The method used hereby is a Frantz cell set up with an artificial silicone membrane. This formulation is very stable, both from a physical-chemical point of view, and from a drug release performance point of view.

Example 1 : Preparation of the exemplary film forming formulation A15 :

The preparation of a bulk premix (150 kg) and the subsequent addition of the propellant is described in the present example. The bulk premix refers to the ingredients of the film forming formulation without the propellant (Table 1 ). Table 2 shows the relation between bulk and propellant in the final formulation in the packaging means. Table 3 summarizes the qualitative and quantitative composition of an exemplary final formulation.

Preparation:

The ethanol, isopropyl myristate, benzyl alcohol, transcutol and eucalyptus oil were introduced into a vessel and stirred for 10 min until a homogeneous clear solution was obtained. Bifonazole was slowly added under stirring and the mixture was continuously stirred for 20 min until again a homogeneous clear solution appeared. The Eudragit RS PO was added slowly under stirring to the bifonazole solution and the mixture was stirred for 2 h until, through visual control, a homogeneous and monophasic_^solution was obtained. The required amounts of bulk premix were then added to aluminium canisters after they were purged with nitrogen. The premix was filled in as fast as possible in an enclosed environment and the valve was immediately applied to the top of the canisters. The propellant gas HFA 134 a was then filled in to yield a monophasic and saturated solution and the canisters were weight checked and checked for tightness.

Euca yptus oi Menthol oil, ***Limonene

Investigation of bifonazole permeation into the stratum corneum

The in vi7ro_percutaneous absorption of bifonazole through human skin was investigated.

The general approach was to investigate the percutaneous absorption of bifonazole, incorporated at a concentration of 1% in two different formulations (film forming formulations prepared according to Example 1 and Table 4), a state of the art cream formulation of bifonazole (Canesten® EXTRA cream, manufactured by Bayer Vital (Germany, c.f. table 5) and a state of the art bifonazole solution spray (Canesten® EXTRA spray, manufactured by Bayer Vital (Germany), c.f. table 6) after a single topical application to human skin.

Table 6: Bifonazole Solution Spray Composition

Composition Amount (g)

Drug Substance

Bifonazole 1.00

Excipients

Ethanol (96 per cent) 30.0

Isopropyl myristate 52.6

Weight 83.6

The percutaneous absorption through human skin was measured quantitatively in vitro, on human dermatomed skin biopsies mounted in Franz™ diffusion cells type (LARASPIRAL, Dijon, France) with an effective diffusion area of 2.02 cm². The dermal part of the skin was in contact with a saline solution comprising a phosphate buffer in which the substance absorbed through the skin was assayed over time. At the end of the 24 hours of diffusion, the receptor fluid i.e. the saline solution comprising the phosphate buffer was sampled. Then, bifonazole was assayed in the washing liquids and in the different skin structures: stratum corneum (horny layer [strips]), viable epidermis and dermis. The mass balance of the study was calculated.

One Caucasian skin biopsy was obtained from a bank of human skin from cosmetic surgery from 48 years old individual, frozen from January 2008. The skin biopsy was cleared of adhering subcutaneous fat with a scalpel. Then, each skin was sliced with a dermatome at a theoretical constant thickness of about 250/500 μπι using a Brown™ dermatome (Zimmer, 25460 ETUPES, France). Quality control of the skin thickness was performed for each skin sample before being mounted on a diffusion cell using a specific device (thickness control system with a reading to 1/100 mm - Ets G. Boutillon - 21300 Chenove, France). Skin samples were then randomized and mounted in the diffusion cells without being subject to any other treatment.

The integrity of the skin barrier and the water-tightness of the experimental model were verified for each diffusion cell before application of the studied products, by measurement of the transepidermal water loss (TEWL). The measurement was performed directly on the epidermal compartment using an evaporimeter (Tewameter™, Courage & Khazaka, Koln Germany). The probe was left in place on the skin surface for 2 minutes; the transepidermal water loss was then recorded for one minute. Any cell presenting an abnormal transepidermal water loss, compared to the other cells used for the test and to data from the literature (Rougier A., TEWL and transcutaneous absorption, in Bioengineering of the skin: « water and the stratum corneum », Ed. P. Eisner, E. Berardesca, H.I. Maibach, CRC Press, p: 103-113, 1994), for the same anatomical site, was noted.

The skin biopsy was mounted in horizontal position between the two parts of the cell demarcating two compartments, one on each side of the sample: firstly, an "epidermal" compartment consisting of a glass cylinder with a surface of 2.02 cm², accurately defined, applied to the upper side of the skin. And secondly, a "dermal" compartment with the receptor fluid applied to the lower side of the tegument, consisting of a 4.03 mL fixed volume compartment, with a sampling port for analysis.

The two compartments were held in place by a horseshoe clamp, and water-tightness was provided by the thickness of the skin maintained between them. The "dermal" receptor compartment was filled with the receptor fluid made of phosphate buffer (pH 7.4) saline solution containing 0.1 % sodium azide as a preservative and 2% of Brij 98 (polyoxyethylene (20) oleyl ether), as a non ionic solubilizer. This solution allows the dissolution of bifonazole and maintains "sink" conditions, The "dermal" compartment was immersed in a water bath in order to maintain the epidermal surface temperature at 32 ± 2° C. Homogeneous distribution of the temperature of the "dermal" compartment was maintained by a magnetic stirring bar, with the diffusion cell mounted on a stirring device. Samples of the fluid contained in the "dermal" compartment (receptor fluid) were collected from the sampling port during the study and at the end of the diffusion time.

During the contact duration of the product with the skin, the upper portion, i.e. the "epidermal" compartment, was open to the air, exposing the epidermis to the ambient conditions of the laboratory environment.

The diffusion cells were prepared on the day of the application of the formulations. After control of the quality of the skin samples, approximately 10 mg (amount exactly known) of the formulations (corresponding to 5 mg of formulation/cm²) were applied on the entire surface of the epidermis circumscribed by the cylinder. Because of their differing viscosities, the formulations were accurately weighed. As mentioned before, the skin surface was left opened in contact with the atmosphere of the laboratory.

The contact between the skin and the formulations was maintained during 24 hours after the application and at the end of the 24 hours of diffusion, the total fluid contained in the receptor compartment (receptor fluid) was collected. Then, the surface of the biopsy and the glass donor part of the cell were washed to remove the excess of formulation on the skin surface by washing three times with one half cotton bud soaked in hydro-alcoholic solution (50/50,v/v) followed by one half dry cotton bud.

The residual product in the half cotton buds and on the glass donor part of the cell delimiting the area of application was extracted by contact during 72 hours in the mobile phase. The extraction solution for bifonazole was named "washing liquids".

Then the cells were dismantled, and the different skin layers were separated. The horny layer was first removed by tape stripping with a 3M™ (Magic™ Tape Scotch) adhesive tape applied on the skin during a few seconds under a constant controlled pressure (45 g/cm²). Two strips were collected and the first one was put in a polypropylene vial "SI" while the other one was put in another polypropylene vial "S2". Then, viable epidermis was separated from the dermis using forceps. The dermis was cut and separated from the adjacent compartment to the application site.

Before being assayed, all samples (washing liquids, horny layer [strips], viable epidermis, dermis, adjacent compartment to the application site and receptor fluid) were stored at + 4° C. The analysis of bifonazole in all these samples allows calculating the mass balance of the experiment that is an ultimate quality control.

The amounts found in the different samples are exemplarily summarized in table 8-12 and represented in Fig. 2-7.

Investigation of film forming formulation properties

1. General methods

To screen for consumer acceptable formulations, a range of characterization experiments were performed. Preliminary screening of the formulations was performed, formulations that produced uneven or tacky films were not characterized further.

1.1 Film aesthetics

The formulations were assessed by actuating one metered dose onto a Petri-dish and allowing the film to dry for approximately 30 min. The shape of the film was visually examined and uneven film forming formulations were rejected. The tackiness of the film was assessed by depressing a gloved finger into the film. Formulations that formed a tacky film were rejected. Excised full thickness porcine skin samples were used to determine the properties of the film. The formulation was actuated on to a skin sample and the roundness and smoothness of the film formed was assessed visually.

1.2 Drying time

A single actuation of each tested spray as prepared according to Example 1 and Table 4 was actuated onto a weigh boat and the weight loss was recorded at 10 second intervals until the weight reached plateau. Each tested formulation was actuated once on a weigh boat, left to dry over 5 min and weighed. Cotton wool was applied on the film and held down with a 100 g weight for 30 s. The cotton was subsequently removed and the weight boat weighed again. The tackiness was assessed based on the weight gain from the cotton wool on the film.

1.3 Film flexibility

Excised full thickness porcine skin samples were used to determine the flexibility of the film. The formulation was actuated on to a skin sample and the film was allowed to dry over 5 min. The skin sample containing the dry film was folded and the film was considered to be flexible when no visual cracks were observed on the film.

1.4 Resistance to water

For this test, the formulation was actuated in the middle of a Petri dish and the film formed was allowed to dry over 5 min. Following this, the film was immersed in 20 ml of water. After 1 h, the water was removed to observe if the film was still intact. If the film demonstrated resistance to water, the water was replaced and the same procedure repeated after 2, 4 and 24 h.

In a non-standardized consumer test, the film forming formulation was administered onto the skin of a person, preferably the foot, before taking a shower. Standard shampoo was applied and after towel drying the respective skin section was assessed.

7.5 Resistance to rubbing

Excised full thickness porcine samples were used to determine the resistance of each film to rubbing. The formulation was actuated onto a skin sample and the film allowed to dry over 15 min. The skin was placed onto the holder of a brushing device. The dried film was rubbed with the brush for 10 s. The integrity of the film was checked visually. If the film was demonstrated to be intact, the rubbing was repeated until the film was impaired.

2. Results

Formulations A9, Al l, A17, A18, A19 were selected and placebos were prepared for comparative assessment.

2.1 Film aesthetics

All formulations were shown to produce symmetrical and smooth films

2.2 Drying time

In general, all films dried within 1 minute from application and were found to be non-tacky when dry Amongst the formulations tested, the longest drying time (1 to 2 min) was observed for formulation A19. Higher tackiness was also observed for A19 in comparison to the other formulations.

2.3 Film flexibility

None of the films tested were found to exhibit any cracks when applied to the skin. In view of these results it can be concluded that all the prepared formulations reveal good flexibility.

2.4 Resistance to water

All films tested demonstrated a resistance to water after being immersed for a period of 24 h. In a non- standardized consumer test, the film was demonstrated to still be present after a shower and towel drying, visually detectable as a glossy section on the skin.

2.5 Resistance to rubbing

All films tested remained intact when exposed to mechanical force.

Claims

Patent Claims

1. Process for preparing a monophasic and saturated film forming formulation comprising bifonazole, a solvent and at least one cosolvent, a film forming agent, a propellant, and optionally at least one further excipient and/or fragrance comprising the steps of preparing a bulk premix and subsequently adding the propellant.

2. Process according to claim 1 whereby the bifonazole is dissolved in the solvent, the at least one cosolvent, and optionally at least one further excipient and/or fragrance, the film forming agent is then added and the propellant is added thereafter.

3. Film forming formulation comprising bifonazole, a solvent and at least one cosolvent, a film forming agent, a propellant, and optionally at least one further excipient and/or fragrance for the treatment of fungal infections or infections relating to fungal infections of the skin.

4. Use of a film forming formulation comprising bifonazole, a solvent and at least one cosolvent, a film forming agent, a propellant, and optionally at least one further excipient and/or fragrance for the preparation of a medicament for the treatment of fungal infections or infections relating to fungal infections of the skin.

5. Film forming formulation of claim3 or its use of claim 4, whereby the solvent is present in an amount of 10 to 40% and the at least one cosolvent is present in an amount of 0,05 to 7% each, the film forming agent is present in an amount of 1-10%, the propellant is present in an amount of 50 to 90%, and optionally fragrance is present in an amount of 0,1 to 1,2%.

6. Film forming formulation or its use of claim 5, whereby the solvent is present in an amount of 12 to 35% and the cosolvent is present in an amount of 1 to 4% each, the film forming agent is present in an amount of 5-8%, the propellant is present in an amount of 55 to 75%, and optionally fragrance is present in an amount of 0,1 to 1,2%.

7. Film forming formulation or its use according to any of the preceding claims, whereby the solvent is ethanol or isopropyl alcohol in an amount of 15 to 30% and the cosolvent is benzyl alcohol, isopropyl myristate, Transcutol®, Arlasolve® or mixtures thereof in an amount of 0,05 to 7% each, the film forming agent is polyvinyl pyrrolidone (PVP), polyvinyl alcohol (PVA), acrylic polymer or copolymer, methacrylate polymer or copolymer, poly (vinyl acetate), cellulose based polymers or co-polymer, or Eudragit® RS PO in an amount of 6-7%, the propellant is HFA 134a or HFA 227a in an amount of 55 to 75%, and optionally eucalyptus oil, menthol or limonene is chosen in an amount of 0,2 to 0,5% as a fragrance.

8. Film forming formulation or its use according to any of the preceding claims, whereby the formulation is monophasic and bifonazole is present at at least 80%, preferably 90%, most preferably 95% saturation, under conditions of use.

9. Film forming formulation or its use according to any of the preceding claims, whereby the formulation comprises benzyl alcohol.

10. Film forming formulation or its use according to any of the preceding claims, whereby the formulation comprises benzyl alcohol and Transcutol®.

11. Film forming formulation or its use according to any of the preceding claims, whereby the formulation comprises benzyl alcohol and Arlasolve®.

12. Film forming formulation or its use according to any of the preceding claims, whereby isopropyl myristate is present in the formulation.

13. Film forming formulation or its use according to any of the preceding claims, whereby the fungal infections or infections related to fungal infection of the skin are caused by dermatophytes, yeasts, moulds, and dimorphic fungi among those Trichophyton spp., Epidermophyton spp., Microsporum spp., Candida spp., Malassezia spp., Trichosporon spp., Cryptococcus neoformans, Aspergillus spp., Histoplasma spp., Coccidioides immitis, Paracoccidoides brasiliensis, Geotrichum candidum and Blastomyces dermatitidis„ and certain bacteria e.g. Corynebacterium spp., Streptococcus species, Staphylococcus spp.

14. Film forming formulation or its use according to any of the preceding claims, whereby the fungal infections are athlete's foot, tinea pedis, tinea pedis interdigitalis, onychomycosis, tinea unguium, tinea corporis, tinea manuum, tinea manuum interdigitalis, tinea cruris, tinea inguinalis, tinea capitis, seborrheic dermatitis, ringworm or erythrasma.

15. Film forming formulation or its use according to any of the preceding claims, whereby the treatment duration is reduced.