Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/0012—Galenical forms characterised by the site of application
  • A61K9/0014—Skin, i.e. galenical aspects of topical compositions
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
  • A61K31/425—Thiazoles
  • A61K31/426—1,3-Thiazoles
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
  • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
  • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
  • A61K47/10—Alcohols; Phenols; Salts thereof, e.g. glycerol; Polyethylene glycols [PEG]; Poloxamers; PEG/POE alkyl ethers
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
  • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
  • A61K47/20—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing sulfur, e.g. dimethyl sulfoxide [DMSO], docusate, sodium lauryl sulfate or aminosulfonic acids
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
  • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
  • A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
  • A61K47/38—Cellulose; Derivatives thereof

Definitions

• the present invention relates to a composition for topical and/or transdermal administration comprising mirabegron or a salt thereof.
• Drugs that increase fat loss belong for the most part to the class of anorectic drugs. These are largely derived from amphetamines and suppress the appetite. Another possibility to support weight reduction is the use of lipase inhibitors. However, these substances require oral or injectable administration. In the case of systemic administration, however, serious side effects must be expected, since these compounds largely interfere with the metabolism of neurotransmitters in the brain. A topical application essentially reduces systemic absorption and respective side effects.
• WO 2007/011743 Al discloses compositions for treating regional fat deposits comprising long -acting beta- 2 adrenergic receptor agonists salmeterol or formoterol and a compound that reduces desensitization of the target tissue to the long-acting P2 -adrenergic receptor agonist, for example, glucocorticosteroids and/or ketotifen.
• the composition for example, may be administered by injection and/or transdermally.
• pharmaceutically active compounds often show no or only suboptimal clinical efficacy upon topical administration because of poor penetration through the skin.
• EP 3 206 678 Bl discloses a pharmaceutical composition for topical administration comprising a prodrug for an agonist and/or an antagonist for an adrenergic receptor, wherein the prodrug is an ester, for or use in a therapeutic method of shaping a mammalian body by modulation of subcutaneous fat tissue.
• a prodrug for an agonist and/or an antagonist for an adrenergic receptor, wherein the prodrug is an ester, for or use in a therapeutic method of shaping a mammalian body by modulation of subcutaneous fat tissue.
• P3 -adrenoceptor receptor agonists (“beta-agonist”) is mirabegron.
• a permeation of the prodrug provides a transport route via the skin, but the availability of the active ingredient depends on the activity of enzymes which are capable of releasing the agonist from the prodrug, and whether a therapeutic effect can be achieved that way is unknown.
• Mirabegron is a prescription medicine from the group of p3-sympathomimetics. It has been approved in the EU for the treatment of overactive bladder since 2012. Its physicochemical properties, however, are adversary to a dermal application and include a relatively high molecular weight and melting point, it is insoluble in water (water solubility 82 pg/mL) and only limited soluble in lipids. Thus, dissolving high amounts of mirabegron in water is impossible and aqueous mixtures with typically used polar cosolvents retain low solubility for mirabegron. Further, the solubility in lipids is limited and the application of products with high lipid content is unpleasant and reduces patient compliance
• the object of the present invention is therefore to provide a composition allowing a topical or transdermal administration of mirabegron.
• the object is achieved by the composition according to claim 1.
• the object further is achieved by the composition for use according to claims 10 to 14.
• Advantageous embodiments are the subject of the dependent claims. The embodiments may be combined freely unless the context clearly indicates otherwise.
• compositions for topical and/or transdermal administration comprising mirabegron or a salt thereof
• the composition comprises: a) mirabegron or a salt thereof in a range from > 0.01 wt.-% to ⁇ 25 wt.-%; and b) a solvent selected from:
• DMSO dimethyl sulfoxide
• dimethyl sulfoxide allows for a transdermal administration of mirabegron.
• DMSO facilitated skin permeation and an increased active delivery period for mirabegron was observed.
• a mixture of dimethyl sulfoxide with alcohol or glycol ether could provide effective transdermal delivery of mirabegron.
• Pure dimethyl sulfoxide and mixtures thereof enabled sufficient maintenance of dissolved mirabegron on skin while showing high permeation.
• solutions or mixtures without dimethyl sulfoxide either lacked solubility, permeability, were too volatile or were toxicologically inacceptable.
• mirabegron As used herein, the term “mirabegron” (INN), formerly known as YM 178, refers to a p3-adrenergic receptor agonist (“beta-agonist”) of the following formula (1): Mirabegron is commercially available, in Europa under the tradename Betmiga®. Mirabegron is commercially used as non-salt free base. Also salts, preferably pharmaceutically acceptable salts, of mirabegron are usable. The term “pharmaceutically acceptable salts” refers to salts prepared from pharmaceutically acceptable non-toxic bases or acids.
• Preferred salts include ammonium, calcium, magnesium, potassium and sodium salts as well as hydrochloride, sulfate, succinate, acetate, fumarate, tartrate, besilate and mesylate salts.
• mirabegron complexes such as lauryl sulfate, octyl sulfate, myristyl sulfate, and cetyl sulfate salts/complexes are known and are usable.
• the term “topical” administration refers to that a substance, such as mirabegron, is applied to a particular place on or in the body, in particular to body surfaces such as skin or mucous membranes.
• transdermal administration refers to that a substance, such as mirabegron, upon administration permeates the upper skin layers and distributes in the subcutaneous layers, namely dermis and subcutis.
• % are weight% (wt.-%).
• the total amount of all components of the composition does not exceed 100 wt.-%.
• given wt.-% refer to the total weight of the composition.
• 100 g of a 5% mirabegron solution in DMSO/ethanol (50/50, w/w) refers to 5 g mirabegron and 47.5 g of each of DMSO and ethanol.
• the ratio of the solvents in particular the weight by weight ratio (w/w) of DMSO to a further solvent od a mixture of further solvents.
• the weight by weight ratio of DMSO to a further solvent may be in a range of 9.5:0.5 to 0.5:9.5, depending on the nature of the further solvent.
• the ratio of DMSO to the further solvent given as 10/90 (w/w) or 50/50 (w/w), defines the solubility of mirabegron in the formulation, irrespective of the presence of excipients such as gelation agents or stabilizers.
• diethylene glycol monoethyl ether which is commercially available under the tradename Transcutol®, provided high solubility when used without dimethyl sulfoxide but insufficient permeation of mirabegron.
• Other solvents such as ethanol provided high solubility and permeation, but no effect of mirabegron on lipolysis was observed. It is assumed that such a lack of efficacy of mirabegron may be due to evaporation of the solvent and precipitation of mirabegron.
• polar aprotic solvents such as N,N-Dimethylformamide and N-methyl pyrrolidone are toxicologically critical.
• the composition comprises mirabegron as active ingridient.
• the composition comprises mirabegron or a salt thereof in a range from > 0.01 wt.-% to ⁇ 25 wt.-%, preferred in a range from > 0.01 wt.-% to ⁇ 20 wt.-%; preferably in a range from > 0.1 wt.-% to ⁇ 10 wt.-%, more preferably in a range from > 0.5 wt.-% to ⁇ 5 wt.-%, based on the total weight of the composition.
• Such concentrations provided good solubility and effects of mirabegron.
• a further aspect refers to a composition for topical and/or transdermal administration comprising mirabegron or a salt thereof, wherein the composition provides a transdermal flux of mirabegron of at least 5 pg/cm 2 over 24 hours as measured in a Franz Diffusion Cell with an orifice of 1 cm 2 at 32°C according to USP (725).
• the flux is at least 10 pg/cm 2 over 24 hours, preferably at least 20 pg/cm 2 over 24 hours, more preferably at least 30 pg/cm 2 over 24 hours, even more preferably at least 50 pg/cm 2 over 24 hours, more preferably 100 pg/cm 2 over 24 hours measured according to USP (725).
• a method for the treatment of obesity or lipedema, or for providing lipolysis comprises the step of administering to a subject in need thereof, for example a mammal such as a human, a therapeutically effective amount of a composition as described herein.
• Figure 4 the amount of mirabegron permeated per cm 2 of murine skin over 24 h from a solution of 3 wt.-% mirabegron in a mixture of ethanol and isopropyl myristate (90/10, m/m).
• Figure 5 the amount of mirabegron permeated per cm 2 of murine skin over 24 h from a solution of 10 wt.-% mirabegron in a mixture of (50/50, m/m) Transcutol® (TC) and DMSO and of 3 wt.-% mirabegron in aquaeous mixtures of 40, 50 or 75 wt.-% Transcutol® and 60, 50 or 25 wt.-% water, respectively.
• TC Transcutol®
• DMSO 3 wt.-% mirabegron in aquaeous mixtures of 40, 50 or 75 wt.-% Transcutol® and 60, 50 or 25 wt.-% water, respectively.
• Solubility was tested by adding mirabegron in increasing incremental amounts of 10 mg to 5 m of the respective solvent under stirring and subsequent sonification of solution/suspension for 5 min at ambient temperature in each step. Complete dissolution was assumed if a clear, increasingly yellow solution was obtained. It was observed that polar aprotic solvents, short chain alcohols, glycols, and polytheylene glycols were suitable solvents for dissolving mirabegron.
• DMSO dimethyl sulfoxide
• NMP N-methylpyrrolidone
• Transcutol® Gattefosse SAS
• a scheme of a Franz Diffusion Cell is shown in Figure 1.
• a donor compartment (2) and a receptor compartment (4) are attached to each other using clamp attachment lug (6). Between donor compartment (2) and receptor compartment (4) a membrane (8) is mounted.
• the receptor compartment (4) has a sampling side arm (10).
• the formulation (3) to be determined is applied to the donor compartment (2).
• a receiver fluid (5) is contained in the receptor compartment (4).
• the Franz Diffusion Cell allows the measurement of the amount of mirabegron diffused across a membrane, such as abdominal human skin or mouse skin, in an acceptor medium.
• Mouse skin or excised human abdominal skin received from plastic surgery was used as membranes. Mouse skin was removed in full thickness from underlying tissue and was left unshaved.
• Human abdominal skin membranes were prepared by removing subcutaneous fatty tissue with a scalpel and then normalising to 400 pm thickness with a dermatome (Aesculap GA 643).
• a Franz Diffusion Cell as defined in the United States Pharmacopeia (USP (725)) with an orifice area of 1.00 cm 2 (PermeGear) was used at 32°C. 8 m of a solution of 6 wt.-% Brij®98 (Acres Organics) in phosphate-buffered saline (PBS) pH 7.4 (NaCl 137 mmol/L (Merck, Darmstadt, Germany), KC1 2.7 mmol/L (Merck, Darmstadt, Germany), Na 2 HPO 4 10 mmol/L (Sigma- Aldrich, Steinbach, Germany), KH2PO4 1.8 mmol/L (Carl Roth, Düsseldorf, Germany) was used as receiver fluid in the receptor compartment.
• PBS phosphate-buffered saline
• a solution of 10 wt.-% mirabegron in a mixture of 50 wt.-% DMSO and 50 wt.-% Transcutol® (Gattefosse SAS) or of 3 wt.-% mirabegron in aqueous mixtures of 40, 50 or 75 wt.-% Transcutol® and 60, 50 or 25 wt.-% water, respectively, were prepared and tested using human skin. Samples were analysed taken after 24 hours. Figure 5 shows the amount of mirabegron permeated per cm 2 of human skin over 24 h.
• solutions of 3 wt.-% mirabegron in PEG 200 (Sigma- Aldrich, Steinbach, Germany), PEG 400 (Sigma-Aldrich), propylene glycole, water, Transcutol®, ethanol/gycerol (50/50) or ethanol/gycerol/isopropyl myristate (45/45/10) showed permeation of below 3 pg/cm 2 over human skin after 24 hours. This showed that solutions without dimethyl sulfoxide lacked permeability.
• mice were treated for 10 days with either DMSO placebo or 5 wt.-% mirabegron in dimethyl sulfoxide to both flanks of the animals in the respective experimental group. After 10 days mice were sacrificed and subcutaneous fat cells analysed. Fixing of freshly isolated tissue was performed by incubation in PBS containing 4% paraformaldehyde (PF A) for no more than 16 h and washed twice 5 min each with ice-cold PBS.
• PF A paraformaldehyde
• Samples were then dehydrated in ethanol with increasing concentrations: 50 %, 70 %, 95 %, and 100 % (3 times, 20 min for each concentration).
• samples were defatted by 3 successive incubation steps in xylol for 10 min each. Samples were further incubated twice in paraffin at 60°C for 1 hour, followed by overnight incubation at 60°C.
• cassettes the processed samples were then embedded in a warmed liquid paraffin solution and subsequently solidified for storage at room temperature. Sections of 5 pm thickness were taken with a microtome (Microm) for each staining, and the stained sections were then dried on histological slides for 24 hours at 40°C.
• Microm microtome
• Figure 6 shows histological sections after haematoxylin and eosin staining of murine subcutaneous fat cells after 10 days of treatment with DMSO as control in Figure 6a) and after 10 days of treatment with 5 wt.-% mirabegron in DMSO in Figure 6b). It can be taken from a comparison of Figures 6 a) and b) that the size of fat vacuoles in white adipose tissue of the mirabegron-treated group was significantly reduced compared to the control group.
• Figure 7 shows histological sections after haematoxylin and eosin staining of human subcutaneous fat cells after two days treatment of human abdominal skin ex vivo with a mixture of 90 wt.-% ethanol and 10 wt.-% isopropyl myristate as control in Figure 7a) and after two days treatment with 3 wt.-% mirabegron in a mixture of 90 wt.-% ethanol and 10 wt.-% isopropyl myristate in Figure 7b). It can be taken from a comparison of Figures 7 a) and b) that the size of fat vacuoles in white adipose tissue of treated specimen was not significantly reduced compared to the control group. This shows that while a mixture of 90 wt.-% ethanol and 10 wt.-% isopropyl myristate provides permeation (as shown in Table 2), no effect was seen.
• dimethyl sulfoxide and mixtures comprising dimethyl sulfoxide not only provided solubility, permeability, and toxicological acceptability, but also provided for a sufficient active delivery period for mirabegron to provide the desired effects upon transdermal administration in vivo.
• thermogenesis markers after application of mirabegron in vivo
• thermogenesis markers after transdermal administration was tested in vivo in C57BL/6 (25gr) mice.
• Wild type (WT) C57BL/6 mice were split into groups of eight and treated with mirabegron gels (verum) or placebo, respectively. Mice were shaved on the flanks with an electric razor. 50 mg of gel were applied once a day to each flank for 10 d. 24 h after the last application of gel the animals were sacrificed and inguinal white adipose tissue (WATi) was removed and analyzed for browning marker expression by qPCR, western blot and histologically after haematoxylin and eosin, as well as UCP1 antibody staining. For histological ananlysis, tissue samples were fixed in 4% paraformaldehyde solution and embedded in paraffin. Sections of 5 pm thickness were stained with hematoxylin and eosin (H&E) and with UCP1 specific antibodies, respectively.
• H&E hematoxylin and eosin
• RNA and real-time polymerase chain reaction tissues were homogenized with Trizol (ThermoFisher) for mRNA extraction and treated with chloroform with subsequent precipitation by addition of propan-2-ol and washing with 75% EtOH. Concentration of isolated RNA was spectrophotometrically analyzed with a Nanodrop 200 (ThermoFisher Scientific). Synthesis of cDNA was conducted with a ProtoScript II First Strand cDNA Synthesis Kit (New England Biolabs) according to the manufacturer's instructions. Analysis of amplification was performed by staining with SYBR Green Master Mix (ThermoFisher) and measurement on a QuantStudio 5 qPCR system (ThermoFisher). The 2 -ACT method was used to calculate the relative mRNA expression of genes normalized to the housekeeping gene HRPT for murine samples.
• proteins were isolated using RIPA lysis buffer containing 50 mM Tris pH 7.5, 150 mM sodium chloride, 1% NP-4O, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate, 0.1 mM EDTA, 0. 1 mM EGTA, 1 mM Na 3 VO 4 , 10 mM NaF, containing a protease inhibitor cocktail (Roche).
• the protein content was quantified by Bradford assay. Proteins were separated by 12% SDS-PAGE and transferred to a nitrocellulose membrane.
• primary antibodies of UCP1 Sigma- Aldrich
• Calnexin and secondary anti-rabbit antibodies were used.
• Amersham ECL Western Blotting reagent was used to visualize proteins and detection was performed with an hnageQuant LAS 4000 mini (GE Healthcare). Quantification was conducted using Image J software.

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• 2023
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