HK1177695A1

HK1177695A1 - Pharmaceutical composition comprising vitamin d analogue and cosolvent-surfactant mixture

Info

Publication number: HK1177695A1
Application number: HK13104898.0A
Authority: HK
Inventors: Karsten Petersson
Original assignee: Leo Pharma A/S
Priority date: 2009-12-22
Filing date: 2009-12-22
Publication date: 2013-08-30
Also published as: BR112012015447A2; JP2013515017A; AU2009357263A1; RU2500387C1; EP2515912A4; CN102770143B; AU2009357263B2; JP5548275B2; WO2011076206A1; CA2785249A1; ZA201204619B; MX2012007227A; CN102770143A; NZ601002A; EP2515912A1; IL220513A0; IL220513A

Abstract

A topical pharmaceutical composition which is an oil-in-water-oil emulsion comprising a vitamin D derivative or analogue dissolved in a mixture of a non-ionic surfactant and a lower alkanol. The topical pharmaceutical composition may be used in the treatment of dermal conditions, such as psoriasis.

Description

Pharmaceutical composition comprising vitamin D analogue and cosolvent-surfactant mixture

Technical Field

The present invention relates to a topical pharmaceutical composition for dermal administration comprising a pharmacologically active agent, a surfactant, a co-solvent and an aqueous phase.

Technical Field

Psoriasis is a chronic inflammatory skin disease that manifests as erythema, dryness, and desquamation spots caused by hyperkeratosis. The plaques are most commonly found on the elbows, knees and scalp, although more extensive damage may occur to other parts of the body, particularly the lumbosacral area. The most common treatment of mild to moderate psoriasis involves topical application of compositions containing corticosteroids as the active ingredient. Although effective, corticosteroids have the disadvantage of having many side effects such as skin atrophy, striation, acneiform rash, perioral dermatitis, overgrowth of skin fungi and bacteria, hypopigmentation of pigmented skin, and rosacea.

However, for many years, the advantageous non-steroidal treatment of psoriasis has been in topical treatment using the vitamin D analogue compound calcipotriol (by LEO Pharma) formulated in ointment compositionsOrOintments commercially available), wherein calcipotriol is present in a solution or cream composition (available from LEO Pharma)OrCream commercially available). The solvent in the ointment composition is propylene glycol, which has the advantage of increasing the penetration of the active ingredient into the skin, resulting in increased efficacy, but is also known as a skin irritant. Thus, it has been reported that the inclusion of propylene glycol in topical compositions often causes Contact Dermatitis in patients (one study reported a number of irritation responses to propylene glycol of 12.5%, see m.hannuksela et al, Contact Dermatitis 1, 1975, pp.112-116), and that the number of irritation responses increased when propylene glycol was used at high concentrations (as described in j.catanzaro and j.graham Smith, j.am.acad.dermotol.24, 1991, pp.90-95). Due to the increased penetration of calcipotriol into the skin, especially caused by the presence of propylene glycol, it has been found thatThe ointment can be used for treating psoriasisCreams are more effective but also cause skin irritation in a significant proportion of patients with psoriasis.

It is therefore an object of the present invention to provide topical compositions comprising as an active ingredient a vitamin D derivative or analogue having a vitamin D-binding activityThe ointment has skin penetrating and bioactive properties comparable to those of the ointment, but it does not contain propylene glycol as a solvent.

Summary of The Invention

Human skin, particularly the outer layer, the stratum corneum, provides an effective barrier to penetration by microbial pathogens and toxic chemicals. While this property of the skin is generally beneficial, it complicates dermal administration of drugs because a large, if not most, amount of the active ingredient administered on the skin of a patient suffering from a skin disorder cannot penetrate into the viable skin (viable) layer in which it is active. To ensure sufficient penetration of the active ingredient into the dermis and epidermis, it is generally preferred to include the active ingredient in a dissolved state, typically in the presence of a solvent in the form of an alcohol, such as ethanol, or a glycol, such as propylene glycol. Propylene glycol is a well-known penetration enhancer (penetration enhancer), i.e. a low molecular weight ingredient such as a therapeutically active ingredient that is capable of penetrating the stratum corneum and "dragging" the vehicle into the epidermis. Propylene glycol can itself cause significant skin irritation, and it can also "drag" the low molecular weight and potentially irritating ingredients of the vehicle into the epidermis, resulting in the overall irritation effect of conventional vehicles including propylene glycol. For this reason, the presence of propylene glycol as a solvent in compositions intended to treat inflammatory skin diseases may exacerbate the inflammatory response.

In the research leading to the present invention, it was an object to identify a solvent combination which more effectively dissolves sparingly soluble (sparingly soluble) compounds such as vitamin D analogues than low molecular weight alcohols or glycols when used as co-solvents on their own in admixture with aqueous phase, and which also contains a significantly lower amount of low molecular weight alcohol co-solvent. It has been surprisingly found that mixing certain surfactants with certain lower alkanols provides mixtures with exceptionally high solvency. The resulting composition, in which the individual solvent components act synergistically, results in satisfactory penetration of the vitamin D derivative or analogue thereof into the viable layer of the skin at a lower co-solvent concentration than when an alcohol or glycol alone is used as co-solvent. Furthermore, the compositions of the present invention exhibit a combination ofOintment equivalent or higher biological activity as determined in the target gene activation test described in example 4 below. Furthermore, the composition is physically more stable, and wherein the vitamin D analogue is chemically stable.

Accordingly, the present invention relates to a topical composition for skin application which is an oil-in-water-in-oil (oil-in-water-in-oil) emulsion comprising an aqueous phase containing a lipophilic phase dispersed therein, the lipophilic phase comprising:

(a) a vitamin D derivative or analogue in dissolved form;

(b) a nonionic surfactant selected from polyoxyethylene glycerides (polyoxyl glycerides), polyoxyethylene castor oil derivatives, polyoxyethylene alkyl ethers, polysorbates, or poloxamers; and

(c) a lower alkanol co-solvent;

the aqueous phase is dispersed in a pharmaceutically acceptable anhydrous lipophilic carrier or vehicle.

In another aspect, the present invention relates to a topical composition described herein for use in the prevention or treatment of a skin disease or disorder.

Brief Description of Drawings

Figures 1-4 are graphs showing the solubility of calcipotriol monohydrate contained in the co-solvent-surfactant mixture of the compositions of the present invention, compared to the solubility of calcipotriol monohydrate in the co-solvent or surfactant alone.

Figures 5a and 5b are diagrams showing penetration of a composition of the present invention into the skin.

FIG. 6 is the passage of vitamin D in human keratinocytes₃Schematic representation of the activation of the gene encoding cathelicidin. The mechanism of cathelicidin gene activation was applied in a biological assay using reconstituted human epidermis (human keratinocytes cultured to form the epidermal layer characteristic of human skin) to which a composition containing calcipotriol of the present invention was applied to activate cathelicidins, as described in detail in example 4 below.

Detailed Description

Definition of

In this context, the term "nonionic surfactant" refers to a surfactant comprising hydrophilic and hydrophobic moieties, wherein the hydrophilic moieties are uncharged, but derive their surface activity from highly polar groups such as polyoxyethylene groups. For the purposes of the present invention, the surfactant is an oil-in-water surfactant having an HLB value of from 9 to 18.

The term "lower alkanol co-solvent" means consisting essentially of C_1-6A straight or branched chain alkanol (e.g., methanol, ethanol, propanol, isopropanol, or butanol).

The term "vitamin D derivative" means vitamin D₃Such as calcitriol, or precursors of such metabolites, such as alfacalcidol.

The term "vitamin D analogue" refers to a synthetic compound comprising a vitamin D backbone and having side chain modifications and/or modifications of the backbone itself. The analogs exhibit biological activity at vitamin D receptors comparable to naturally occurring vitamin D compounds.

"calcipotriol" is a vitamin D analog having the formula:

calcipotriol has been found to exist in two crystalline forms, an anhydrous form and a monohydrate form. Calcipotriol monohydrate and its preparation are described in WO 94/15912.

The term "storage stability" means that the composition exhibits chemical and physical stability characteristics to allow storage of the composition in a refrigerator, preferably at room temperature, for a sufficient period of time such as at least 12 months, especially at least 18 months, and preferably at least 2 years, for which the composition is commercially viable.

The term "chemical stability" or "chemically stable" means that no more than 10%, preferably no more than 5%, of the vitamin D derivative or analogue degrades over the shelf-life of the product (typically 2 years). An approximation of the chemical stability at room temperature was obtained by subjecting the compounds to accelerated stability studies at 40 ℃. If less than about 10% of the material degrades after 3 months at 40 ℃, this generally corresponds to a shelf life of 2 years at room temperature. By "chemical stability", in particular with respect to calcipotriol, is meant that the calcipotriol does not significantly degrade over time in the finished pharmaceutical product to 24-epi-calcipotriol or other degradation products of calcipotriol.

The term "physical stability" or "physically stable" means that the composition retains its macroscopic and microscopic appearance during the shelf life of the product, e.g., the vitamin D derivative or analog does not precipitate from the solvent phase, or there is no visible phase separation of the solvent phase and the carrier phase.

The term "substantially anhydrous" means that the free water content of the lipophilic carrier or vehicle is no more than about 2% by weight of the carrier or vehicle, preferably no more than about 1% by weight.

The term "solvency" refers to the ability of a solvent or solvent mixture to dissolve a given substance, expressed as the amount required to achieve complete dissolution of the substance.

The term "synergistically" means that when the combination of co-solvent and surfactant is present in the aqueous phase, the solubility of the vitamin D derivative or analogue is significantly higher, in some cases several times higher, than the sum of the solubilities of the co-solvent or surfactant when the co-solvent or surfactant is added to the aqueous phase alone.

The term "skin penetration" refers to the diffusion of an active ingredient into the different layers of the skin, namely the stratum corneum, epidermis and dermis.

The term "skin penetration" refers to the flow of active ingredients through the skin into the systemic circulation, or into the receptor fluid of the Franz cell device used in the experiment in an in vitro study such as those reported in example 3 below.

The term "biological activity" refers to the activity of a vitamin D derivative or analog when applied to the skin in the compositions of the present invention. The biological activity of the composition was determined in an in vitro test measuring the activation of the target gene encoding cathelicidin in a reconstituted human epidermal model (involving cultured human keratinocytes), as described in detail in example 5 below.

Modes for carrying out the invention

In an embodiment of the invention, the composition comprises a vitamin D derivative or analogue selected from the group consisting of: calcipotriol, calcitriol, tacalcitol, maxacalcitol, paricalcitol and alfacalcidol. In a currently preferred embodiment, the composition comprises calcipotriol or calcipotriol monohydrate as the vitamin D analogue.

In the compositions of the present invention, the surfactant is typically present at a concentration of from about 0.5% to about 5%, or from about 1% to about 3%, or from about 1.2% to about 2%, such as about 1.5%, by weight of the composition.

According to the invention, the nonionic surfactant is preferably selected from polyethylene glycol 8 caprylic/capric glycerides (polyethylene glycol derivatives of a mixture of mono-, di-and triglycerides of caprylic and capric acids with an average of 8 moles of ethylene oxide) or polyethylene glycol 6 caprylic/capric glycerides (polyethylene glycol derivatives of a mixture of mono-, di-and triglycerides of caprylic and capric acids with an average of 6 moles of ethylene oxide). The nonionic surfactant is preferably polyethylene glycol 8 caprylic/capric glyceride, such as is available from gattefosse under the trade name Labrasol, or from Condea under the trade name Softigen 767.

The nonionic surfactant may also preferably be polyethylene glycol C_6-20A fatty acid glyceride selected from caprylocaproyl PEG glyceride (caproyl PEG glyceride), lauroyl PEG glyceride, linoleoyl (linoleoyl) PEG glyceride, oleoyl PEG glyceride, and stearoyl PEG glyceride; polyoxyethylene C_8-20An alkyl ether, a ketone,selected from PEG monocetyl ether, PEG monododecyl ether, PEG monooleyl ether, and PEG monostearyl ether; a polysorbate selected from polysorbate 20, 40, 60, and 80; a poloxamer selected from the group consisting of poloxamers 124, 237, 338, and 407; or a polyoxyethylene castor oil derivative, such as polyoxyethylene (polyoxyl) castor oil or hydrogenated polyoxyethylene castor oil.

As mentioned above, the composition further comprises a lower alkanol co-solvent, which may preferably be selected from methanol, ethanol, n-propanol, isopropanol, n-butanol or 2-butanol. It has been surprisingly found that the amount of lower alkanol required for complete dissolution of a vitamin D derivative or analogue in the presence of a surfactant can be significantly reduced (e.g. by a factor of 2-5) compared to the amount required when the lower alkanol alone is used as a solvent. The lower alkanol co-solvent may advantageously be present at a concentration of about 0.5 to 5%, especially about 1 to 3%, or about 2% by weight of the composition.

In a currently preferred embodiment, the co-solvent is ethanol and the non-ionic surfactant is polyethylene glycol 8 caprylic/capric glycerides, polysorbate 80 or PEG monocetyl ether, or the co-solvent is isopropanol and the non-ionic surfactant is polyoxyethylene castor oil, polyethylene glycol 8 caprylic/capric glycerides, polysorbate 80.

The ointment carrier may be of chain length C₅To C₆₀A hydrocarbon or a mixture of hydrocarbons. Common ointment carriers are petrolatum or white soft paraffin (white soft paraffin) having a peak height of about C_40-44Or petrolatum with liquid paraffin (from a peak of about C)_28-40Of different chain length hydrocarbons). While petrolatum provides occlusion of the treated skin surface, reduces the transdermal loss of water, and enhances the therapeutic efficacy of the active ingredients in the composition, it tends to have a greasy and/or tacky feel, persists for a considerable period of time after application, and is not easily spreadable. Thus, it may be preferred to use paraffins composed of lower chain length hydrocarbons, such as those with a chain length peak of C_14-16、C_18-22、C_20-22、C_20-26Paraffin wax or mixtures thereof (the hydrocarbon composition of paraffin wax has been determined by gas chromatography). Such paraffins have been found to be more cosmetically acceptable because they are less tacky and/or greasy when applied and are easier to spread. They are therefore expected to lead to improved patient compliance. Suitable paraffins of this type are known as petrolatum (petroleum jelly), produced by Sonneborn and sold under the trade name Sonnecone, such as Sonnecone CM, Sonnecone DM1, Sonnecone DM2 and Sonnecone HV. These paraffins are further disclosed and characterized in WO 2008/141078, which is hereby incorporated by reference.

To impart the desired tackiness to the composition of the invention, it may be convenient to include a lipophilic viscosity building component, such as a wax. The wax may be comprised of a high molecular weight hydrocarbon (e.g., saturated C)_35-70Alkanes) such as microcrystalline waxes. Alternatively, the wax may be a vegetable or animal wax, such as C_14-32Fatty acids and C_14-32Esters of fatty alcohols, such as beeswax. The amount of tackifying component can vary depending on the tackifying ability of the component, but generally can be in the range of about 1-20% by weight of the composition. When the viscosity-building component is a microcrystalline wax, it is typically present in an amount of about 5 to 15% by weight of the composition, for example about 10% by weight.

The composition may further comprise an epidermal thickening emollient (emollient) useful for softening psoriatic plaques. Suitable emollients for inclusion in the compositions of the present invention may be silicone waxes or volatile silicone oils, as the presence of silicone has also been found to assist in penetration of calcipotriol into the skin. It has also been found that compositions comprising silicone cause less skin irritation. Suitable silicone oils for inclusion in the compositions of the present invention may be selected from cyclomethicone (cyclomethicone), dimethicone (dimethicone). The amount of silicone oil included in the compositions of the present invention is generally from about 1 to about 10%, for example about 5%, by weight of the composition.

In thatThe presence of propylene glycol in ointments is believed to be the primary cause of skin irritation experienced by many patients. However, calcipotriol itself has been found to be mildly irritant in some patients (a.fullerton and j.cerup, br.j.dermotol.137, 1997, pp.234-240 and a.fullerton et al, br.j.dermotol.138, 1998, pp.259-265). It is therefore advantageous to include an anti-irritant compound such as glycerol, butylene glycol, sorbitol, sucrose, saccharin, menthol or niacinamide in the compositions of the present invention. Glycerol has been described as a substance capable of protecting the skin from irritants (j. bettinger et al, Dermatology 197, 1998, pp.18-24), and we have found that it reduces the release of IL-1 α in a dose-dependent manner: thus, it has been found that the presence of 15% by weight of glycerol in the calcipotriol ointment results in a significantly lower level of IL-1 α release than calcipotriol ointment comprising 10% by weight of glycerol, which in turn results in a significantly lower level of IL-1 α release than ointment comprising 5% by weight of glycerol.

However, in addition to the anti-irritant effect, glycerol was surprisingly found to be able to enhance the biological activity of calcipotriol, since the expression of cathelicidin (in the test method described in example 4 below) was found to increase with decreasing amount of glycerol in the composition (i.e. more cathelicidin was expressed when the amount of glycerol was 5% by weight compared to 10% or 15% glycerol, respectively): this suggests that a balance should be found between good anti-irritant effect and good potentiation in the case of glycerol inclusion. We have found that inclusion of about 5-10% by weight of glycerol in the compositions of the invention produces significant anti-irritant effects and that the biological activity of calcipotriol is significantly enhanced.

Calcipotriol is known to be an extremely sensitive substance to acidic conditions (pH below about 7.0 in aqueous compositions or acidic reactive substances in non-aqueous compositions) which lead to rapid degradation of calcipotriol. To ensure that the substance has sufficient chemical stability over the shelf life of the composition, it may be appropriate to include a compound capable of neutralizing acidic impurities which may be present in one or more excipients of the composition and which are detrimental to the chemical stability of calcipotriol. The acid neutralizing compound may advantageously be selected from buffering agents, such as phosphate buffers, which may be included in an amount of about 0.025-0.1% by weight of the composition. The acid neutralizing compound may also be an amine, such as triethanolamine, tromethamine, monoethanolamine or diethanolamine, which may be included in the composition in an amount of about 0.1-2% by weight.

To maintain good physical stability of the composition, especially to avoid separation of the aqueous and lipid phases therein, it may be advantageous to include a water-in-oil emulsifier having an HLB value of from 3 to 8. An example of such an emulsifier is polyoxyethylene C_8-22Alkyl ethers, such as polyoxyethylene stearyl ether, polyoxyethylene cetyl ether or polyoxyethylene lauryl ether.

The amount of water in the composition can be from about 1% to about 15% by weight of the composition, for example from about 5% to about 10% by weight.

In a specific embodiment, the composition of the invention comprises:

in another embodiment, the composition of the present invention comprises:

in yet another embodiment, the composition of the present invention comprises:

in another embodiment, the composition of the present invention comprises:

the compositions of the present invention may also contain other Ingredients commonly used in skin formulations, such as antioxidants (e.g., alpha-tocopherol), preservatives, sodium edetate, pigments, skin soothing agents (skin soothing agents), skin healing agents (skin healing agents) and skin conditioning agents such as urea, allantoin or bisabolol, see CTFA Cosmetic Ingredients Handbook, second edition, 1992.

The compositions of the present invention are useful for treating psoriasis, seborrheic psoriasis (sebopsis), palmoplantar pustulosis (pustulosis palmoplantaris), dermatitis, ichthyosis (ichthyosis), rosacea and acne and related skin disorders by topically applying an effective amount of the compositions of the present invention to a patient in need of such treatment. The method preferably comprises topically administering a therapeutically sufficient dose of the composition once or twice daily. For this purpose, the compositions according to the invention preferably contain from about 0.001 to 0.5mg/g, preferably from about 0.002 to 0.25mg/g, in particular from about 0.005 to 0.05mg/g of vitamin D derivative or analogue. The compositions of the invention can advantageously be used for maintenance therapy of these skin diseases, i.e. continuing the therapy after the visible symptoms have disappeared in order to delay the recurrence of the symptoms.

In order to provide a more effective treatment for psoriasis and other skin diseases in the acute phase, it may be desirable to include one or more other therapeutically active ingredients in the composition. Examples of such other active ingredients include, but are not limited to, anti-inflammatory drugs, such as corticosteroids, such as betamethasone and esters thereof, e.g. valerate or dipropionate, clobetasol or esters thereof, such as propionate, hydrocortisone or esters thereof, such as acetate; non-steroidal anti-inflammatory drugs such as naproxen, indomethacin, diclofenac, ibuprofen, dexibuprofen, ketoprofen, flurbiprofen, piroxicam, tenoxicam, lornoxicam or nabumetone, phosphodiesterase 4 inhibitors or p38MAP kinase inhibitors.

The invention is further illustrated by the following examples, which do not in any way limit the scope of the invention as claimed.

Example 1

Compositions of the invention

The compositions shown in tables 1a and 1b below were prepared as follows: first the surfactant (Cremophor EL, Labrasol, polysorbate 80 or cetomacrogol 1000) and the co-solvent (ethanol or isopropanol) were mixed, the calcipotriol monohydrate was dissolved in the mixture, and finally the mixture was added to an aqueous buffer solution adjusted to pH 8.0 and glycerol (when included). The resulting dispersion was then mixed with a mixture of paraffin wax, emulsifier (polyoxyethylene stearyl ether), DL-alpha-tocopherol and sodium edetate.

TABLE 1a

Compositions A to F according to the invention

TABLE 1b

Compositions G to L of the invention

TABLE 2

Compositions M and N of the invention

Compositions M and N were prepared essentially as described for compositions a-L above, but using a mixture of white petrolatum (Sonnecone DM1) and microcrystalline wax instead of white soft paraffin and, for composition N, triethanolamine was added to the aqueous phase instead of disodium hydrogen phosphate.

Example 2

Solubility of calcipotriol in solvent/surfactant mixtures in aqueous buffers

Polycetostearyl alcohol 1000 in combination with ethanol

The solubility of calcipotriol monohydrate (measured as calcipotriol) at 25 ℃ in an aqueous buffer pH7.4 containing cetomacrogol 1000 as surfactant and ethanol as co-solvent is shown in table 3 below. Table 3 shows that the combination of cetostearyl alcohol 1000 and ethanol has a small synergistic effect on the solubility of calcipotriol in aqueous phosphate buffered solutions, i.e. the observed solubility of calcipotriol in aqueous phosphate buffered solutions containing 15% of cetostearyl alcohol 1000 and 20% of ethanol is 1468 μ g/g, which is about 1.4 times the sum of the solubilities of calcipotriol in aqueous phosphate buffered solutions containing 15% of cetostearyl alcohol 1000 or 20% of ethanol (1049 μ g/g +2.33 μ g/g-1051 μ g/g), see fig. 1.

Table 3: solubility (. mu.g/g) of calcipotriol at 25 ℃ in aqueous vehicles using Polycetostearyl alcohol 1000 as surfactant and ethanol as co-solvent

Solvent	Solubility μ g/g
		Phosphate buffer ph 7.4: ethanol (80: 20)	2.33
Phosphate buffer ph 7.4: polycetostearyl alcohol 1000 (85: 15)	1049
		Phosphate buffer ph 7.4: 1000 parts of polycetol: ethanol (65: 15: 20)	1468

Labrasol and ethanol combination

The solubility of calcipotriol monohydrate (measured as calcipotriol) in an aqueous buffer pH7.4 containing Labrasol as surfactant and ethanol as co-solvent at 25 ℃ is shown in table 4 below. Table 4 shows that the combination of Labrasol and ethanol has a significant synergistic effect on the solubility of calcipotriol in aqueous phosphate buffered phosphate solutions, i.e. the observed solubility of calcipotriol in aqueous phosphate buffered phosphate solutions containing 15% Labrasol and 20% ethanol is 1059 μ g/g, which is about 3.6 times the sum of the solubilities of calcipotriol in aqueous phosphate buffered phosphate solutions containing 15% Labrasol or 20% ethanol (292 μ g/g +2.33 μ g/g-294 μ g/g), see fig. 2.

Table 4: solubility (. mu.g/g) of calcipotriol at 25 ℃ in an aqueous vehicle using Labrasol as surfactant and ethanol as co-solvent

Solvent	Solubility μ g/g
		Phosphate buffer ph 7.4: ethanol (80: 20)	2.33
Phosphate buffer ph 7.4: labrasol (85: 15)	292
		Phosphate buffer ph 7.4: labrasol: ethanol (65: 15: 20)	1059

Labrasol in combination with isopropanol

The solubility of calcipotriol monohydrate (measured as calcipotriol) in an aqueous buffer pH7.4 containing Labrasol as surfactant and isopropanol as co-solvent at 25 ℃ is shown in table 5 below. Table 5 shows that the combination of Labrasol and isopropanol has a significant synergistic effect on the solubility of calcipotriol in aqueous phosphate buffered solutions, i.e. the observed solubility of calcipotriol in aqueous phosphate buffered solutions containing 15% Labrasol and 20% isopropanol is 1508 μ g/g, which is about 4.9 times the sum of the solubilities of calcipotriol in aqueous phosphate buffered solutions containing 15% Labrasol or 20% isopropanol (292 μ g/g +17.2 μ g/g-309 μ g/g), see fig. 3.

Table 5: solubility (. mu.g/g) of calcipotriol at 25 ℃ in an aqueous vehicle using Labrasol as surfactant and isopropanol as co-solvent

Solvent	Solubility μ g/g
		Phosphate buffer ph 7.4: isopropanol (80: 20)	17.2
Phosphate buffer ph 7.4: labrasol (85: 15)	292
		Phosphate buffer ph 7.4: labrasol: isopropanol (65: 15: 20)	1508

Polysorbate 80 in combination with ethanol

The solubility of calcipotriol monohydrate (measured as calcipotriol) at 25 ℃ in an aqueous buffer pH7.4 containing polysorbate (tween) 80 as a surfactant and ethanol as a co-solvent is shown in table 6 below. Table 6 shows that the combination of polysorbate 80 and ethanol has a synergistic effect on the solubility of calcipotriol in aqueous phosphate buffered saline, i.e. the observed solubility of calcipotriol in aqueous phosphate buffered saline containing 15% polysorbate 80 and 20% ethanol is 740 μ g/g, which is about 2.0 times the sum of the solubilities of calcipotriol in aqueous phosphate buffered saline containing 15% polysorbate 80 or 20% ethanol (360 μ g/g +2.33 μ g/g-362 μ g/g), see fig. 4.

Table 6: solubility (. mu.g/g) of calcipotriol at 25 ℃ in aqueous vehicles using polysorbate 80 as surfactant and ethanol as co-solvent

Solvent	Solubility μ g/g
		Phosphate buffer ph 7.4: ethanol (80: 20)	2.33
Phosphate buffer ph 7.4: polysorbate 80 (85: 15)	360
		Phosphate buffer ph 7.4: polysorbate 80: ethanol (65: 15: 20)	740

The results shown in tables 3-6 below show that higher solubility of calcipotriol monohydrate in aqueous solution can be achieved by the synergistic effect of the combination of co-solvent and surfactant instead of using either co-solvent alone or surfactant alone. This means that a lower amount of the combination can achieve the same solubility of calcipotriol monohydrate than either solvent alone.

Example 3

Penetration study

To study the skin penetration and penetration of calcipotriol from the compositions of the present invention, skin diffusion experiments were performed. Full thickness skin from pig ears was used in this study. The ears were kept frozen at-18 ℃ prior to use. The day before the experiment, the ears were placed in a refrigerator (5. + -. 3 ℃ C.) to be thawed slowly. On the day of the experiment, the hair was removed using a veterinary shaver. The skin was cleaned of subcutaneous fat using a scalpel, and two pieces of skin were cut from each ear and mounted in Franz diffusion cells at balanced grade (balanced order).

The application has a length of 3.14cm, substantially as described in T.J.Franz, "The fine technique as a valid in a viromodel for The study of The cultural of The evolution in man", Current documents in Dermatology, 1978, J.W.H.Mall (eds.), Karger, Basel, pp.58-68²Static Franz-type diffusion cells with available diffusion area and receiving volume of 8.6-11.1 ml. The specific volume of each cell was determined and registered. A magnetic bar is placed in the receiving chamber of each cell. After the skin is mounted, each receiving chamber is filled with physiological saline (35 ℃) for hydrating the skin. The cell was placed in a thermally controlled water bath placed on a magnetic stirrer set at 400 rpm. The circulating water in the water bath was maintained at 35 + -1 deg.C so that the skin surface temperature was about 32 deg.C. After one hour, the saline was replaced with a receiving medium of 0.04M isotonic phosphate buffer pH7.4 (35 ℃) containing 4% bovine serum albumin. Sink conditions (Sink conditions) were maintained at all times during the study, i.e., the concentration of the active compound in the receiving medium was less than 10% of the solubility of the compound in the medium.

Each test compound was tested in 6 replicates (i.e., n-6)Skin penetration. At 0 hour at 4mg/cm²The expected dosage of each test composition was applied to the skin membrane. A glass spatula was used for this application and the residual amount of the composition was measured to obtain the amount of composition actually applied to the skin.

The skin penetration test was allowed to proceed for 21 hours. Samples were then collected from the following chambers:

by usingThe stratum corneum was collected by tape stripping (tape striping) 10 times (diameter 22mm, CuDerm corp. Each tape strip was applied to the test area for 5 seconds using standard pressure and removed from the test area in a gentle, continuous motion. The direction of tearing is different for each repeat of the strip. Viable (viable) epidermis and dermis are then collected from the skin in a similar manner.

A sample (1ml) of the receiving liquid remaining in the diffusion cell was collected and analyzed.

The concentration of calcipotriol in the sample was determined by LC mass spectrometry.

The results are shown in figures 5a and 5b below, which show the amount of calcipotriol in viable skin (epidermis and dermis) and in the receiving liquid in% of the administered dose. The compositions of the present invention were found to have excellent penetration and penetration characteristics, particularly those containing Labrasol as the surfactant component.

Example 4

Biological activity of the composition

As shown in fig. 6 below, cathelicidins are antimicrobial peptides expressed in human keratinocytes. Cathelicidin expression is strongly induced by skin infection or disruption of the skin barrier. In psoriasis, cathelicidin levels are elevated in the affected skin of psoriasis patients. Has already been used forIt was found that the expression of the gene coding for cathelicidin could be influenced by vitamin D₃Or vitamin D analogues such as calcipotriol are induced by binding to vitamin D receptors (see TT Wang et al, J.Immunol.173(5), 2004, pp.2909-2912; J Schauber et al, Immunology 118(4), 2006, pp.509-519; Schauber and Gallo, J.Allergy Clin Immunol 122, 2008, pp.261-266; M.Peric et al, ploS One4(7), 7.7.22.2009, e 6340). This finding has been used to develop an assay in which the uptake and biological activity of calcipotriol in human keratinocytes from a test composition is determined by measuring the induction level of the gene encoding cathelicidin.

In this test, composition A, B, C, F, G, I, J, K, L prepared as described in example 1 above was topically applied in triplicate in an amount of 10 μ l to reconstituted human epidermis consisting of layers at 0.5cm²Polycarbonate filter membrane (fromObtained from Laboratories, Nice, France) for 12 days. The tissue was treated for 2 days, then the epidermis was separated from the polycarbonate filter and snap frozen in liquid nitrogen. RNA was extracted from the cells and cDNA was synthesized by a conventional method. Real-time quantitative pcr (qpcr) was then performed using the following assay from Applied Biosystems: CAMP Hs0018038_ m1 and gapdh 99999905_ m 1. Normalizing the expression level of cathelicidin to GAPDH and comparingRelative quantification was performed by ointment comparison.

The results are shown in table 7 below.

TABLE 7

¹Relative toOintment

The results shown in table 7 show that the compositions of the invention result in higher target gene activation, i.e. they may have higher in vivo biological activity than the commercially available ointments.

Example 5

Local tolerance studies in miniature pigs

Topical tolerability of compositions B, G, M and N of example 1 was evaluated daily by skin application to mini-pigs for 4 weeks. Applications ofOintment for comparison. Animals were exposed to the test items for 8 hours per day.

The study was performed in 10 females Gttingen SPF mini-pigs. Each animal had 6 application sites and received a 250mg volume of the test formulation at each application site.

Clinical signs were recorded daily, and skin reactions at the application site were scored for erythema and edema once daily before the start of dosing, and also on the day of necropsy. Food consumption was recorded daily and body weight was recorded weekly. At the end of the treatment period, all animals were subjected to a full necropsy and skin samples were collected from histopathological examination.

The results show that no adverse treatment-related clinical signs were observed during the study, although grade 1-2 skin reactions (erythema) were observed. For erythema ratio, in addition to composition GThose observed for ointmentsAnd is less obvious. The results show the possible ratios of the compositions of the inventionThe ointment is better tolerated by human patients.

Claims

1. A topical composition for skin application which is an oil-in-water-in-oil emulsion comprising an aqueous phase containing dispersed therein a lipophilic phase comprising:

(a) calcipotriol in dissolved form or calcipotriol monohydrate;

(b) a nonionic surfactant selected from polyethylene glycol C in an amount of 0.5% to 5% by weight of the composition_6-20Fatty acid glyceride, polyoxyethylene C_8-20Alkyl ethers or polysorbates; and

(c) a lower alkanol co-solvent selected from ethanol and isopropanol in an amount of 0.5 to 5% by weight of the composition;

2. The composition according to claim 1, wherein the nonionic surfactant is present at a total concentration of 1% to 3%, or 1.2% to 2%, or 1.5% by weight of the composition.

3. A composition according to claim 1 or 2, wherein polyethylene glycol C_6-20The fatty acid glyceride is selected from caprylocaproyl PEG glyceride, lauroyl PEG glyceride, linoleoyl PEG glyceride, oleoyl PEG glyceride and stearoyl PEG glyceride; polyoxyethylene C_8-20The alkyl ether is selected from PEG monocetyl ether, PEG monododecyl ether, PEG monooleyl ether and PEG monostearyl ether; the polysorbate is selected from polysorbate 20, 40, 60 and 80.

4. A composition according to claim 1 or 2, wherein the non-ionic surfactant is polyethylene glycol 8 caprylic/capric glycerides or polyethylene glycol 6 caprylic/capric glycerides.

5. The composition according to claim 1, wherein the lower alkanol co-solvent is present at a concentration of 1-3%, or 2% by weight of the composition.

6. The composition according to claim 1, wherein the co-solvent is ethanol and the non-ionic surfactant is polyethylene glycol 8 caprylic/capric glycerides, polysorbate 80 or PEG monocetyl ether, or wherein the co-solvent is isopropanol and the non-ionic surfactant is polyethylene glycol 8 caprylic/capric glycerides or polysorbate 80.

7. The composition according to claim 1, wherein said carrier comprises at least one paraffin wax selected from the group consisting ofChain length C determined chromatographically₅To C₆₀And the peak value of the chain length is C_14-16、C_18-22、C_20-22、C_20-26、C_28-40And C_40-44Paraffin wax or mixtures thereof.

8. The composition of claim 7, further comprising a viscosity-increasing component.

9. The composition of claim 8, wherein the adhesion promoting component is a wax.

10. The composition according to claim 1, further comprising a compound capable of neutralizing acidic impurities in the composition that are detrimental to the chemical stability of calcipotriol or calcipotriol monohydrate.

11. The composition according to claim 10, wherein said compound is triethanolamine.

12. The composition according to claim 1, comprising 0.001-0.5mg/g, or 0.002-0.25mg/g, or 0.005-0.05mg/g of calcipotriol or calcipotriol monohydrate.

13. A topical composition for skin application which is an oil-in-water-in-oil emulsion comprising an aqueous phase containing dispersed therein a lipophilic phase comprising:

(a)0.003-0.008% w/w calcipotriol monohydrate in dissolved form;

(b) polyethylene glycol 8 caprylic/capric glycerides in an amount of 1-3% by weight of the composition, polyoxyethylene stearyl ethers in an amount of 3-8% by weight of the composition; and

(c) ethanol in an amount of 1-3% by weight of the composition;

the aqueous phase is dispersed in a pharmaceutically acceptable anhydrous lipophilic carrier or solvent;

wherein the composition comprises 5-10% w/w water and 80-93% w/w paraffin carrier.

14. A topical composition for skin application which is an oil-in-water-in-oil emulsion comprising an aqueous phase containing dispersed therein a lipophilic phase comprising:

(a)0.003-0.008% w/w calcipotriol monohydrate in dissolved form;

(b) polysorbate 80 in an amount of 1-3% by weight of the composition, polyoxyethylene stearyl ether in an amount of 3-8% by weight of the composition; and

(c) ethanol in an amount of 1-3% by weight of the composition;

15. A topical composition for skin application which is an oil-in-water-in-oil emulsion comprising an aqueous phase containing dispersed therein a lipophilic phase comprising:

(a)0.003-0.008% w/w calcipotriol monohydrate in dissolved form;

(b) polyethylene glycol monocetyl ether in an amount of 1-3% by weight of the composition, polyoxyethylene stearyl ether in an amount of 3-8% by weight of the composition; and

(c) ethanol in an amount of 1-3% by weight of the composition;

16. A topical composition for skin application which is an oil-in-water-in-oil emulsion comprising an aqueous phase containing dispersed therein a lipophilic phase comprising:

(a)0.003-0.008% w/w calcipotriol monohydrate in dissolved form;

(c) isopropanol in an amount of 1-3% by weight of the composition;

17. A topical composition for skin application which is an oil-in-water-in-oil emulsion comprising an aqueous phase containing dispersed therein a lipophilic phase comprising:

(a)0.003-0.008% w/w calcipotriol monohydrate in dissolved form;

(c) isopropanol in an amount of 1-3% by weight of the composition;

18. The composition according to claim 1, further comprising one or more additional therapeutically active ingredients.

19. The composition according to claim 18, wherein the further active ingredient is selected from corticosteroids, non-steroidal anti-inflammatory drugs, phosphodiesterase 4 inhibitors or p38MAP kinase inhibitors.

20. The composition according to claim 1 for use in the treatment of a skin disease or disorder.

21. The composition of claim 20, wherein the skin disease or disorder is psoriasis, palmoplantar pustulosis, dermatitis, ichthyosis, or acne.

22. The composition of claim 20, wherein the skin disease or disorder is seborrheic psoriasis or rosacea.