HK1111599B - Pharmaceutical compositions for the treatment of cellulite - Google Patents

Description

Pharmaceutical composition for treating cellulite

The invention relates to a pharmaceutical preparation for the local or mesodermal treatment of cellulite (cellulite), which contains a PDE3 inhibitor as active ingredient, optionally together with other substances having anti-cellulite activity.

Background

Cellulite, or "edema-fibrosclerotic panniculopathy", is a condition that affects the subcutaneous tissue, the tissue located below the dermis, primarily of adipose nature. Cellulite affects almost exclusively women, and approximately 80-85% of the adult female population suffers from this disease. Subcutaneous adipose tissue accounts for 25% of women's body weight, and serves 3 basic functions: a) it provides physical and mechanical protection; b) their release of lipids and proteins involved in lipid metabolism; and c) it exerts endocrine and paracrine actions.

Statistically, this defect affects most white women. This problem is much less common in women of other ethnic groups. The most susceptible to this condition are the "mediterranean" women, generally due to their richer supply of estrogen.

Even slim women tend to exhibit more pronounced fat accumulation in the thighs.

Some factors have adverse effects, causing local changes that affect the microcirculation of the fatty substance. Ultimately, this leads to anatomical and functional impairment of the tissue vasculature, creating problems affecting the subcutaneous tissue and the layer immediately above it, the dermis.

Cellulite is caused by the degeneration of the microcirculation of adipose tissue and the subsequent alteration of its most important metabolic functions.

The visible consequences of this tissue degeneration are increased volume of adipocytes, fluid retention in the interstitial spaces and fluid stagnation.

Cellulite may have genetic (familial predisposition), physical, hormonal and vascular causes, often exacerbated by: sedentary lifestyle, stress, liver disease, poor diet, intestinal disorders, or disorders characterized by significant fluid retention.

Hormonal imbalances (affecting ovarian, pituitary and thyroid hormones) are responsible for cellulite; due to the action of estrogen and its effect on the microcirculation, women are susceptible to this condition, especially in adolescence, pregnancy and pre-menopausal stages, when ovarian hormone activity is at a peak.

Susceptibility to cellulite is therefore primarily hormone based rather than genetic factors.

The structural features of the subcutaneous connective tissue have a marked amphipathic difference, which predisposes women to the formation of the irregular prominence of the adipose tissue characteristic of dermal cellulite. It has been proposed that: the predominant anti-lipolytic activity of subcutaneous adipose tissue (alpha 2-adrenergic-dependent receptors) in women compared to men promotes increased deposition of fat in the thighs and thus the appearance of cellulite (Rosenbaum et al, plastics and Reconstructive surgery, 101 (7): 1934 1939, 1998).

Furthermore, the lipolytic response of adipocytes originating from the hip, hip or thigh sites to catecholamines is lower than that of adipocytes originating from visceral adipose tissue (Lafontan and Berlan, TIPS, 24: 276-.

Currently, cellulite is generally treated as follows:

-physical therapy

In addition to massage techniques, techniques such as electron lipolysis and more recently laser therapy, ion electrophoresis, ultrasonic therapy, and ozone therapy are now widely used; however, none of these techniques fundamentally solves the problem.

-dietary supplements

There are numerous dietary supplements (inorganic salts, especially potassium, vitamins, "fat-burning agents" or diuretic plant extracts, intestinal regulators and bioflavonoids) on the market, claiming to increase metabolism, improve circulation, protect cells from damage and reduce fat absorption; no effective clinical trials are known to support the efficacy of these dietary supplements in the treatment of cellulite.

Pharmacologically active products

According to a study published in European J.of Dermatology (10(8) 596-.

Other widely used compounds are:

a) aminophylline, due to its ability to increase cAMP and lipolysis; both advantageous and disadvantageous findings regarding their anti-cellulite activity have been published.

b) Levothyroxine, which utilizes thyroid hormones to increase the ability of metabolism. In view of the high dose of levothyroxine systemic absorption may occur, followed by a cardiac stimulatory effect and interference with the thyroid gland, which is particularly harmful for hyperthyroid patients.

c) Aescin, due to its vasoprotective heparinoid ability.

Mesoderm therapy (mesotherapy)

This technique involves the local intradermal injection of drugs, usually administered by the systemic route. This method allows a small amount of product to be injected directly into the cellulite region; this technique is therefore non-systemic and not very invasive. Long-term therapeutic effects can be obtained using mesodermal therapy because the absorption of drugs at the skin level is slow. The compounds currently used in mesodermal therapy for cellulite treatment are coenzyme a, phosphatidylcholine, aminophylline, aescine and homeopathic (homeopathic) products.

Adipocytes are a cell that easily changes in size: lipogenesis increases in volume and lipolysis decreases in volume.

Lipogenesis is caused by LPL (lipoprotein lipase): adipocytes adjacent to capillaries synthesize and release LPL, which hydrolyzes Triglycerides (TG) present in Very Low Density Lipoproteins (VLDL) and kilomicron. Glycerol and fatty acids are released, captured by adipocytes, and esterified to triglycerides.

Lipolysis is caused by Hormone Sensitive Lipase (HSL): this enzyme hydrolyzes TG in adipocytes to free fatty acids and glycerol. The active enzyme is phosphorylated by cAMP-dependent protein kinase a.

cAMP synthesis is dependent on two opposing enzyme systems:

d) adenylate cyclase, which converts ATP to cAMP. It is negatively regulated by α 2-adrenergic receptors and positively regulated by β adrenergic receptors.

e) Phosphodiesterase, which breaks down cAMP into AMP inert to HSL. This activity is inhibited by caffeine, theophylline and aminophylline.

In addition, adipocytes secrete a variety of factors, such as leptin (satiety factor), angiogenic factor (angiotensinogen), prostaglandin PGE₂(anti-lipolytic Properties) and PGI₂(cell differentiation), lysophosphatidic acid (cell proliferation) and steroids.

Treatment strategies

The development of the pharmacological basis for cellulite treatment has gone through a series of stages. In the eighties, xanthine (caffeine) was used to inhibit Phosphodiesterase (PDE). In the nineties, plant extracts (flavonoids and saponins) with drainage and anti-edema activity were sought to address this problem by improving venous lymphatic insufficiency (venolymphatic insufficiency). More recently, attempts have been made to reconstitute connective tissue with extracellular matrix and degrading enzyme components.

However, the most promising therapeutic approach appears to be one designed to increase adipocyte metabolism and lipolysis.

Lipolysis is initiated by: a) beta-adrenergic receptor stimulation; b) adenosine or alpha₂-adrenergic receptor inhibition; c) phosphodiesterase inhibition.

A series of clinical trials have evaluated topical application of beta-adrenergic agonists (isoproterenol), alpha₂The effects of adrenergic antagonists (yohimbine) and phosphodiesterase inhibitors (aminophylline). The results show that: local fat reduction can be achieved by pharmacological means without dieting or exercise (Greenway et al, Obes res, 3: 561S-568S, 1995).

Stimulation of β -adrenergic receptors increases cAMP concentrations in adipocytes, thus stimulating lipolysis. Another method to increase cAMP is to prevent its degradation by inhibiting phosphodiesterase.

The most common PDE inhibitors (caffeine, theophylline and aminophylline) are not satisfactory because of their low specificity and poor local absorbability. PDE3, particularly PDE3B, is present in human adipocytes; it therefore appears necessary to selectively inhibit these subtypes of enzymes in order to obtain an effect that is confined within adipose tissue.

Patents filed on this subject:

EP692250 relates to the use of flavones to improve microcirculation, and GB1588501, FR2797765, EP1261310 and EP1259221 relate to cosmetic use of xanthine-activated lipases.

From the information given above, it can be seen that lipolytic activity is necessary but not sufficient for the treatment of cellulite. The treatments that have been used to date, which often use phosphodiesterase inhibitors such as xanthines, have not proven to be entirely satisfactory, and there is a particular need for new, effective treatments for this multifactorial, change in the form of subcutaneous fat.

A group of aryldihydropyridazinones and aryldimethylpyrazolones which are selective PDE3B inhibitors and are potentially useful in the treatment of obesity are described in Bioorganic & Medicinal Chemistry Letters, (2003), 13, 3983-.

Disclosure of Invention

It has now been found that PDE3 (phosphodiesterase-3) inhibitors, in particular inhibitors of the PDE3B isoform, which is predominantly expressed in human adipose tissue, are surprisingly effective against cellulite.

Accordingly, the present invention provides the use of a PDE3, preferably a PDE3B inhibitor, for the manufacture of a pharmaceutical composition for the local or mesodermal therapy of cellulite.

Among the PDE3 inhibitors, the compounds anagrelide (anagrelide), cilostazol (cilostazol), pimobendan (pimobendan), milrinone (milrinone), amrinone (amrinone), olprinone, enoximone (enoximone), cyclohexylquinamide (cilostamide), vesnarinone (vesnarinone), troquinazine (trequinsin) and pharmaceutically acceptable salts thereof are preferred. Particular preference is given to milrinone, troquinazine and cyclohexolamide.

Another group of preferred PDE3 inhibitors according to the invention are the compounds described in Bioorganic & medicinal chemistry Letters, (2003), 13, 3983-:

1)6- [4- (2-benzyl-3-oxo-cyclohex-1-enylamino) -phenyl ] -5-methyl-4, 5-dihydro-2-H-pyridazin-3-one (Compound 8a)

2)3- {2- [4- (4, 4-dimethyl-5-oxo-4, 5-dihydro-1-H-pyrazol-3-yl) -2, 3-difluoro-phenylamino ] -6-oxo-cyclohex-1-enylmethyl } -benzonitrile (Compound 18n)

3)5- {4- [2- (2, 6-dichloro-benzyl) -3-oxo-cyclohex-1-enylamino ] -2-fluoro-phenyl } -4, 4-dimethyl-2, 4-dihydro-pyrazol-3-one (Compound 18h)

4)5- [4- (2-benzyl-3-oxo-cyclohex-1-enylamino) -phenyl ] -4, 4-dimethyl-2, 4-dihydro-pyrazol-3-one (Compound 18a)

5)5- {4- [2- (3-Nitro-benzyl) -3-oxo-cyclohex-1-enylamino ] -2-fluoro-phenyl } -4, 4-dimethyl-2, 4-dihydro-pyrazol-3-one (Compound 18f)

6)6- [4- (2-benzyl-3-oxo-cyclohex-1-enylamino) -2-fluoro-phenyl ] -5-methyl-4, 5-dihydro-2-H-pyridazin-3-one (Compound 14)

The term "topical" as used herein means designed for or related to topical application and action. The topical compositions are preferably in the form of creams, non-oily creams, ointments, oily-non-oily formulations (oil-non-oily formulations), gels, spray-gels and patches. For mesodermal therapy, the composition should be in a form suitable for topical intradermal injection, preferably in the form of an injection solution.

For topical forms, the concentration of the active ingredient is 0.1 to 3%, preferably 1 to 2%, based on the total weight of the composition, and for injectable forms for mesodermal therapy, the concentration of the active ingredient is 0.1 to 1%, based on the total weight of the composition. In addition to PDE3 inhibitors, the compositions according to the invention may contain compounds, mixtures of compounds or extracts active on the microcirculation, preferably saponins or flavones or extracts containing them. Most preferred are extracts of Ginkgo biloba (Ginkgo biloba), Arnica (arnica), pineapple (ananas), Angelica sinensis (Angelica siniensis), Centella asiatica (Centella asiatica), and aescin.

The compound, extract or mixture of substances active on microcirculation is contained in the composition at a concentration of 0.1-4%.

The composition according to the invention may further contain pharmaceutically acceptable excipients, such as adjuvants, in particular water or alcohol (ethanol), vitamins, in particular tocopherol, dexpanthenol or retinol palmitate, thickeners, preservatives, protective colloids, humectants, fragrances, electrolytes, wetting agents, gelling agents, skin penetration enhancers, polymers or copolymers, emulsifiers, emulsion stabilizers and other pharmaceutically acceptable excipients.

Preferred preservatives are hypoallergenic substances, such as ethanol or benzyl alcohol.

Topical formulations may contain unsaturated derivatives of oleic acid, such as 10-trans-12-cis-linoleic acid.

Particularly suitable gelling agents are carbomers, more preferably carbomer 940, polyacrylamide, isoparaffin-polyethylene glycol monododecylether-7, xanthan gum, carrageenan, gum arabic, guar gum, agar gel, alginates and methylhydroxycelluloses, carboxymethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, hydroxyethylcellulose, ethylcellulose, polyacrylates, polyvinyl alcohol, polyvinylpyrrolidone, colloidal silica. Urea and panthenol are examples of wetting agents according to the present invention.

Techniques for preparing the Pharmaceutical compositions of the present invention are known to those skilled in the art and are described, for example, in Remington's Pharmaceutical Sciences, XVIII edition, mack publishing co.

The above pharmaceutical composition comprising a PDE3 inhibitor is conveniently used in the topical treatment of cellulite in humans, preferably in women.

The following examples illustrate the invention in more detail.

Examples

Example 1 non-oily cream (% by weight)

Active ingredients:

amrinone 1

Aescin 2

Excipient:

glyceryl monostearate 8

Polyethylene glycol cetostearyl alcohol ether

(Macrogol cetostearyl ether) 2.5

Liquid paraffin 2

White vaseline 2

Isopropyl myristate 4

Myristyl alcohol 3

P-hydroxybenzoate 0.3

Purified water in proper amount of 100g

Example 1.2

Active ingredients:

milrinone 1

Excipient:

cetostearyl alcohol 4.5

Glyceryl monostearate 8.0

Liquid paraffin 2

White vaseline 2

Dimethicone 0.30

Isopropyl myristate 1

Myristyl alcohol 3

Proper amount of essential oil

Purified water in proper amount of 100g

Example 1.3

Active ingredients:

quinazine 2

Excipient:

oleic acid 5.0

Stearic acid polyethylene glycol ester 40

(Macrogol stearate40) 9.0

Cetostearyl alcohol 6.0

Butyl hydroxy anisole 0.02

Tromethamine 0.1

Dimethicone 0.3

Carbopol 9800.3

Propylene glycol 20.0

Sodium sulfite 0.1

Proper amount of essential oil

Purified water in proper amount of 100g

Example 2: hydroalcoholic gel (Hydroalcoholic gel) (% by weight)

Active ingredients:

milrinone 2

Excipient:

carbomer 1.5

Ethanol 96 degree EP 40ml

Proper amount of essential oil

Proper amount of triethanolamine for adjusting pH

Purified water in proper amount of 100g

Example 3: lipophilic cream (% by weight)

Active ingredients:

amrinone 1

Excipient:

polyglyceryl diisostearate (3) ester 4

Oleic acid glyceride 2

Beeswax 7

Dioctyl ether 10

Hexyldecanol/hexyldecanol laurate 10

85% Glycerol 5

Magnesium sulfate heptahydrate 1

P-hydroxybenzoate 0.1

Proper amount of essential oil

Purified water in proper amount of 100g

Example 4: non-oily cream (% by weight)

Active ingredients:

3- {2- [4- (4, 4-dimethyl-5-oxo-4, 5-dihydro-1-H-pyrazol-3-yl) -2, 3-difluoro-phenylamino ] -6-oxo-cyclohex-1-enylmethyl } -benzonitrile: 1

Excipient:

oleic acid 5.0

Stearic acid polyglycol ester 409.0

Cetostearyl alcohol 6.0

Butyl hydroxy anisole 0.02

Tromethamine 0.1

Dimethicone 0.3

Carbopol 9800.3

Propylene glycol 20.0

Sodium sulfite 0.1

Purified water in proper amount of 100g

The following active ingredients can be made into non-oil creams according to example 4:

6- [4- (2-benzyl-3-oxo-cyclohex-1-enylamino) -phenyl ] -5-methyl-4, 5-dihydro-2-H-pyridazin-3-one;

5- {4- [2- (2, 6-dichloro-benzyl) -3-oxo-cyclohex-1-enylamino ] -2-fluoro-phenyl } -4, 4-dimethyl-2, 4-dihydro-pyrazol-3-one;

5- [4- (2-benzyl-3-oxo-cyclohex-1-enylamino) -phenyl ] -4, 4-dimethyl-2, 4-dihydro-pyrazol-3-one;

5- {4- [2- (3-nitro-benzyl) -3-oxo-cyclohex-1-enylamino ] -2-fluoro-phenyl } -4, 4-dimethyl-2, 4-dihydro-pyrazol-3-one;

6- [4- (2-benzyl-3-oxo-cyclohex-1-enylamino) -2-fluoro-phenyl ] -5-methyl-4, 5-dihydro-2-H-pyridazin-3-one.

Example 5: injection solution for mesoderm therapy (5 ml/bottle) (dosage in mg/ml)

Active ingredients:

milrinone 5

Excipient:

sodium pyrosulfite and lactic acid 0.2-0.5

Adding appropriate amount of purified water to 5ml

Example 6: efficacy test

Cellulite is a difficult problem with multi-factorial etiology in which adipogenesis plays a key role. Therefore, in the initial stage of screening for potential anti-cellulite drugs, in vitro measurement of lipolytic activity is an essential element, although it is generally insufficient for the determination of the final drug candidate. In vitro testing of lipolytic activity must therefore be combined with testing to assess in vivo efficacy.

In vitro assay

● determination of extracellular Glycerol produced by lipolysis of triglyceride in adipocytes

In vitro lipolysis assays were performed using human adipocytes, which are primary cultures derived from preadipocytes (Promocell), the progenitor cells of which were obtained by inducing differentiation. Actual adipocyte differentiation was determined by measuring glycerol-3-phosphate dehydrogenase, an enzyme expressed in mature adipocytes but not in preadipocytes, and measuring intracellular levels of aggregated lipids using oil red o (sigma).

Preincubating PDE3 inhibitor with adipocytes at different concentrations for 15 min; 10nM isoproterenol, which induces a weak stimulation of lipolysis (25% maximal response is obtained with 1. mu.M isoproterenol), is then added. After 4 hours the medium was removed and the amount of intracellular lipids accumulated in each well was determined using oil red O.

Lipolytic activity was assessed by measuring glycerol released into the extracellular matrix following hydrolysis of triglycerides.

In this test, a formulation with a PDE3 inhibitor, comprising a selective PDE3 inhibitor according to the invention, proved to have comparable or better effects than a similar formulation containing a non-selective PDE inhibitor (caffeine, theophylline).

Activity of certain PDE inhibitors on Glycerol Release stimulated by 10nM isoproterenol in cultured human adipocytes

● measurement of cAMP production in adipocytes

cAMP is an intracellular messenger, the level of which depends on its synthesis (adenylate cyclase activity) and decomposition (phosphodiesterase activity). Adipocytes were transferred to saline solution, to which a lipolytic agent (adenylate cyclase activator), an anti-lipolytic agent (phosphodiesterase activator) and a PDE3 inhibitor were added. After a suitable period of incubation, the incubation was interrupted and the cAMP formed within the cells was extracted. Cell extracts were lyophilized and then reconstituted and assayed using a special commercially available colorimetric Enzyme Immunoassay (EIA) (Amersham, Cayman). cAMP levels were determined according to the manufacturer's instructions. In this test, the formulations with the PDE3 antagonist according to the invention proved to be more effective than similar formulations containing non-selective PDE inhibitors (caffeine, theophylline).

In vivo assay

10 healthy adult women with pronounced cellulite on the upper thigh were examined.

Each patient served as its own control to evaluate efficacy and safety. A cream containing the active ingredient (1% milrinone) was applied to the exterior of one thigh in a random fashion, while a cream base (no active ingredient) was applied to the same area of the other thigh.

Using about 2-3cm cream therapy, massage for 2-3 minutes until absorbed, repeated twice daily for 2 months.

During the trial, the patient did not exercise the program and was not subject to any dietary restrictions.

The circumference of each thigh, two thirds of which is between the knee and greater trochanter, is measured periodically until the end of the treatment. The circumference of the drug-treated thigh was shortened by 2.9 + -0.7 cm, which ranged from 1.3-4.7cm, compared to the untreated thigh.

At the end of the observation period, PDE3 was measured in the blood and the presence of compound was not detected.

Claims (12)

1. Use of a phosphodiesterase-3B inhibitor selected from the group consisting of: anagrelide, cilostazol, pimobendan, milrinone, amrinone, enoximone, troquinazine.

2. Use according to claim 1, wherein the phosphodiesterase-3B inhibitor is milrinone or troquinazine.

3. Use according to claim 1, wherein the topical pharmaceutical composition is in the form of a gel, ointment or patch.

4. Use according to claim 3, wherein the topical pharmaceutical composition is in the form of a spray gel or cream.

5. Use according to claim 1, wherein the topical pharmaceutical composition is in a form suitable for topical intradermal injection or mesodermal therapy.

6. Use according to claim 5, wherein the composition is in the form of an injection solution.

7. Use according to claim 1, wherein the amount of phosphodiesterase-3B inhibitor is from 0.1 to 3% by weight.

8. Use according to claim 7, wherein said amount is 1 to 2% by weight.

9. Use according to claim 7, wherein said amount is 0.1 to 1% by weight.

10. Use according to claim 1, wherein the composition further comprises a compound selected from the group consisting of saponins and flavones, or an extract active on the microcirculation selected from the group consisting of arnica or ginkgo extracts.

11. Use according to claim 10, wherein the compound is aescin.

12. Use according to claim 10 or 11, wherein the compound or extract active on the microcirculation is present in a concentration of 0.1 to 4% by weight.