WO1998043606A1

WO1998043606A1 - Slimming cosmetic compositions

Info

Publication number: WO1998043606A1
Application number: PCT/FR1998/000438
Authority: WO
Inventors: Daniel Greff
Original assignee: Sederma S.A.
Priority date: 1997-04-02
Filing date: 1998-03-05
Publication date: 1998-10-08
Also published as: FR2761602B1; AU6839198A; FR2761602A1

Abstract

The invention concerns peptide sequences having lipolytic activity which can be used in slimming cosmetic products for topical application. Said activity is reinforced when the peptides are chemically modified to increase lipophilicity. Said peptides can be obtained by synthesis, biotechnology or controlled hydrolysis of plant proteins.

Description

TITLE: Slimming cosmetic compositions

The cosmetic industry is constantly looking for new active ingredients with lipolytic activities to integrate them into so-called slimming products. Many substances (pure molecules such as xanthine derivatives; complex mixtures such as certain plant extracts, and others) are proposed and used. The market nevertheless demands new and ever more active products.

Apart from adrenergic substances (adrenaline and the like) whose lipolysis-stimulating activity in adipocytes is well known but whose use in cosmetic products is prohibited, a class of substances of a different nature has been discovered for which stimulation lipolysis of triglycerides in human and / or animal adipocytes has been demonstrated: these substances are small peptides of a hormonal nature. In fact, glucagon, secretin, the peptide called PACAP (Pituitary Adenylate Cyclase Activating Polypeptide), VIP (the Vasointestinal Peptide), PHI,

PHM, helodermine, exendin are gastric peptides involved in the regulation of digestion, glucose metabolism, transformation of chemical energy (glycolysis, lipolysis, intestinal absorption, etc.). These fairly large peptides (20 or more amino acids) have a certain sequence homology, some key amino acids always being in the same positions in the N terminal part of the peptide. They thus constitute a kind of family of molecules.

Glucagon: His Ser Gin Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser Arg Arg Ala Gin Asp Phe ... Secretin: His Ser Asp Gly Thr Phe Thr Ser Glu Leu Ser Arg Leu Arg Glu Gly

Ala Arg Leu Gin Arg Leu ... PACAP: His Ser Asp Gly Ile Phe Thr Asp Ser Tyr Ser Arg Tyr Arg Lys Gin

Met Ala Val Lys Lys Tyr ... VIP: His Ser Asp Ala Val Phe Thr Asp Asn Tyr Thr Arg Leu Arg Lys Gin Met Ala Val Lys Lys Tyr ...

PHI: His Ala Asp Gly Val Phe Thr Ser Asp Tyr Ser Arg Leu Leu Gly Gin

Ile Ser Ala Lys Lys Tyr ... PHM: His Ala Asp Gly Val Phe Thr Ser Asp Phe Ser Lys Leu Leu Gly Gin

Ile Ser Ala Lys Lys Tyr ... Hélodermine: His Ser Asp Ala Ile Phe Thr Glu Glu Tyr Ser Lys Leu Leu Ala Lys Leu Ala Leu Gin Lys Tyr ...

Exendine: His Ser Asp Gly Thr Phe Thr Ser Asp Leu Ser Lys Gin Met Glu Glu

Glu Ala Val Arg Leu Phe ... Their peptide size makes their synthesis on an industrial scale very difficult, and therefore incompatible with the economic requirements of the target market. The object of this patent is the discovery that shorter peptide sequences - and therefore more easily achievable at the level of industrial synthesis - as well as some of their chemically modified analogues are also active in the in vitro and in vivo stimulation of lipolysis . Contrary to all expectations, this activity can be obtained in vivo topically, and therefore by a cosmetic treatment.

For example, the His-Ser-GIn-Gly-Thr-Phe sequence (fragment 1-6) of glucagon, has 66% of the lipolytic activity of the complete hormone. To make it more active topically, it is advantageous to make it lipophilic by grafting a fatty acid of more or less long chain (lipoyl, mirystyl, palmityl, stearyl ...) on the N-terminal residue and / or to esterify the carboxyl group of the peptide. In this way, the various fragments of the Glucagon, PACAP, VIP etc peptides, characterized in that they contain at least the sequence of general form: rV-NH-His-Serfou Ala) -Gln (or Asp) -Gly (or Ala) -Thr (or Ile or Val-Phe-OR ² where P. = H or a linear or branched, saturated or unsaturated, hydroxylated or sulfur or unsulfurized alkyl chain containing 1 to 24 carbon atoms, preferably 1 2 to 1 8 carbon atoms and R ² = H, or an alkyl chain from C1 to C24, preferably either C1 to C3, or C14 to C18 or OR ² = NR ³ R ⁴ with R ³ and R ⁴ being independently one of l 'other = H or an alkyl chain of

1 to 1 2 carbon atoms, preferably 1 to 3 carbon atoms, have a lipolytic activity by the topical route usable in cosmetics and dermopharmacy. Two examples illustrate this activity: Human adipocytes (obtained from pieces of skin removed during plastic surgery) are suspended in a survival medium. The basic lipolytic metabolism is followed by the dosage of glycerol and fatty acids released over time. Different concentrations of NH ₂ -His-Ser-Gln-Gly-Thr-Phe-OH are then added to the medium. After a more or less long incubation time, there is an increase in lipolytic metabolism by an increase in the amount of glycerol and of fatty acids released in the medium. A positive stimulation control consists of glucagon 1-29 (whole hormone). The results obtained are as follows:

Glycerol release: Glucagon at 10 ^"7 M / I 1 1 x10 ^{" 5} nmol / cell / hr

Glucagon- [1 -6] at 10 ⁷ M / l 6x10 ^"5 nmol / cell / hr In this model we tested a number of peptides derived from the basic sequence. For example, to increase the lifetime of the peptide (with respect to proteolytic enzymes) we substituted glycine (or alanine in the case of VIP peptide and Helodermine) in position 4 by D-alanine or D-serine. To increase the affinity of peptide for the skin, we attached an aliphatic chain (going from C1 to C24) either at the N-terminal end (amide bond), or at the C-terminal end (ester bond) of the peptide. some of these are:

Increase over baseline (glucagon [1-6]).

We therefore see that it is possible to modulate the lipolytic activity of the main sequence as required (solubility, duration of action, epidermal affinity, etc.). Other variants can be envisaged by those skilled in the art while remaining within the scope of the invention.

Under the same experimental conditions, it is also possible to follow the variation in the concentration of cAMP (cyclic adenosine monophosphate) in the cell medium. In fact, the activation of lipolysis in adipocytes often, if not always, involves the stimulation of the activity of adenylate cyclase as a result of which the high concentration of cAMP leads to prolonged activation of triglycerides lipases. The peptides studied have activities stimulating adenylate cyclase, which are not linearly linked to lipolytic activity, which would indicate that different mechanisms of activation of lipolysis intervene according to the peptide. Some peptides are highly lipolytic without significantly stimulating adenylate cyclase, others have both activities to a high degree.

The joint use of certain peptides which are the subject of this patent application can prove to be particularly advantageous. An in vivo test was carried out on 15 women aged 35 to 62 years. At the start of the study, the perimeter of the thighs is measured using a centimeter; the thickness of the adipose layer is determined using the ultrasound technique (ultrasound). After 4 weeks of twice daily treatment with gels (a placebo gel and a gel containing the two peptides N-Acetyl-His-Ser-GIn-Gly-Thr- Phe and N-Palmitoyl-His-Ser-GIn-D-ALa- Thr-Phe respectively at 10 ^-5 and 10 ^'6 M / l), the same parameters are measured. There is a favorable development in the perimeter and the thickness of the adipose layer on the thighs treated with the preparation containing the lipolytic peptides: -13% and -1 5% respectively for each of the parameters measured), while the decrease in these values on thighs treated with placebo is not significant. The gel used is that of Example No. 1 p.5).

The peptides which are the subject of this patent can be obtained either by conventional chemical synthesis (in solid phase or in homogeneous liquid phase), or by enzymatic synthesis (Kullman et al., J. Biol. Chem. 1,980, 255, 8234) from constituent amino acids or their derivatives. The peptides can also be obtained by fermentation of a strain of bacteria modified or not by genetic engineering to produce the sequences indicated on pages 1 and 2 and their fragments or by extraction of proteins of animal or vegetable origin, preferably vegetable, followed by a controlled hydrolysis which releases the peptide fragments of medium size in question, with the stipulation that the fragments released must contain at least the sequence NH ₂ -His-Ser (or

Ala) -Gln (or Asp) -Gly (or Ala) -Thr (or Ile or Val) -Phe-OH in the N-terminal part.

Many proteins found in plants are likely to contain these sequences within their structure. The controlled hydrolysis makes it possible to release these peptide fragments. It is possible, but not necessary to carry out the invention, to extract either the proteins concerned first and then to hydrolyze them, or to carry out the hydrolysis first on a crude extract and to purify the peptide fragments then . We can also use the hydrolyzed extract without extracting the peptide fragments which interest us, however making sure to have stopped the hydrolytic enzymatic reaction in time and to measure the presence of these peptides by appropriate analytical means (tracing by radioactivity , immunofluorescence or immunoprecipitation with specific antibodies etc.). Other simpler or more complex processes leading to products cheaper or purer are easily conceivable by those skilled in the art knowing the trade of extraction and purification of proteins and peptides. By way of example illustrating the invention, a few representative but non-limiting cosmetic formulas are cited:

Example 1: Slimming gel

Carbopol ^R 1 342 0.3

Propylene glycol 2

Glycerin 1

White petrolatum 1 .5

Cylomethicone 6

Sipol C16C18S3 0.5

Lubrajel MS 10

Triethanolamine 0.3

N-Acetyl-His-Ser-GIn-Gly-Thr-Phe 0.0001

N-Palmitoyl-His-Ser-GIn-D-ALa-Thr-Phe 0.00005

Water, preservatives, perfume qs 100 g.

The placebo gel used for the test does not contain | have both peptides.

Example 2: Slimming cream

B Brriijj ^RR 772211 2 2..44

Brij ^R 72 2.6

ArlamoIR E 8.0

Beeswax 0.5

Abil ^R ZP 2434 3.0

Propylene glycol 3.0

Carbopol ^R 941 0.25

0.25 triethanolamine

N-Lipoyl-His-Ser-Gln i-ALa-Thr-Phe 0.00001

Caffeine 1 .0

Water, preservatives, perfumes qs 100 g

Example 3: alcoholic lotion

Ethanol 5.0

Propylene glycol 2.0

Abil ^R B8851 0.5

EumulginR L 0.6

His-Ala-Asp-Gly-Val-Phe-OMe 0.0002 water, preservatives, perfume qs 100 g.

The lipolytic peptides that are the subject of this patent can be used regardless of their origin (synthesis, biotechnological fermentation, extraction) in any dosage form used in cosmetics: O / W and W / O emulsions, milks, lotions, gels, ointments, body oils, shampoos, soaps, sticks, sprays, powders, without this list being exhaustive.

It is possible to use the lipolytic peptides in the form of a solution, dispersion or emulsion or encapsulated in cosmetic vectors such as macro-, micro- or nanocapsules, liposomes or chylomicrons, or included in macro- , micro- or nanoparticles, or in micro-sponges, or adsorbed on powdery organic polymers, talcs, bentonites and other mineral supports.

The concentration of use of these lipolytic peptides can vary between 0.000001 and 1% (w / w), more particularly between 0.00001 and 0.1% and preferably between 0.0001 and 0.01%, in the finished product.

Peptides of a lipolytic nature can be combined in cosmetic compositions with any other ingredient usually used in cosmetics: extraction and / or synthetic lipids, gelling and viscosifying polymers, surfactants and emulsifiers, water-soluble or liposoluble active ingredients, extracts of other plants, tissue extracts, marine extracts, etc.

The combination with other lipolysis stimulating agents such as caffeine, theophylline, xanthine derivatives in general is particularly advantageous for carrying out the invention.

Claims

1. Cosmetic compositions for slimming treatment, characterized in that they contain one or more peptides of the following structure: R ¹ -NH-His-Ser (or Ala) -Gln (or Asp) -Gly (or Ala or Ser) -Thr ( or Ile or Val) -Phe-0-R ² where R ¹ = H or a linear or branched alkyl chain, saturated or unsaturated, hydroxylated or sulfurized or not containing 1 to 24 carbon atoms, preferably 1 2 to 18 carbon atoms and R ² = H, or an alkyl chain from C1 to C24, preferably either C1 to C3 or C14 to C18 or OR ² = NR ³ R ⁴ with R ³ and R ⁴ being independently of each other = H or an alkyl chain of 1 to 1 2 carbon atoms, preferably 1 to 3 carbon atoms.

2. Cosmetic compositions for the slimming treatment according to claim 1, characterized in that the glycine in position 4 is replaced by D-Alanine or D-Serine.

3. Cosmetic compositions for the slimming treatment according to claims 1 and 2 characterized in that the peptides are of chemical or enzymatic synthesis origin or obtained by fermentation or extraction of plants.

4. Cosmetic compositions for the slimming treatment according to claims 1 and 2 characterized in that the peptides are obtained by conventional peptide synthesis in homogeneous or heterogeneous phase or by enzymatic synthesis from the constituent amino acids or their derivatives.

5. Cosmetic compositions for the slimming treatment according to claims 1 and 2 characterized in that the peptides are obtained by extraction of plant proteins, followed by enzymatic hydrolysis so as to generate peptide fragments of medium size, part of the fragments released must contain at least the NH ₂ -His-Ser (or Ala) -Gln (or Asp) -Gly (or Ala) -Thr (or Ile or Val) -Phe-OH sequence in the N-terminal part.

6. Cosmetic compositions for the slimming treatment according to any one of claims 1 to 5, characterized in that the concentration of use of these peptides of lipolytic nature varies between 0.000001 and 1% (w / w), more particularly between 0.00001 and 0.1% and preferably between 0.0001 and 0.01% in the finished product.

7. Cosmetic compositions for the slimming treatment according to any one of Claims 1 to 6, characterized in that the lipolytic peptides are used in the form of a solution, dispersion or emulsion or encapsulated in macro-, micro- or nanocapsules, liposomes or chylomicrons, or included in macro-, micro- or nanoparticles, or in micro-sponges, or adsorbed on powdery organic polymers, talcs, bentonites and other mineral supports.

8. Cosmetic compositions for the slimming treatment according to any one of claims 1 to 7, characterized in that the lipolytic peptides are used in any galenic form used in cosmetics or dermopharmacy, namely O / W and W / O emulsions, milks, lotions, gels, ointments, body oils, shampoos, soaps, sticks, sprays, powders. 9) Cosmetic compositions for the slimming treatment according to any one of claims 1 to 8, characterized in that the lipolytic peptides are combined in the cosmetic compositions with any other ingredient usually used in cosmetics: extraction lipids and / or synthesis, gelling and viscosifying polymers, surfactants and emulsifiers, water- or liposoluble active ingredients, plant extracts, tissue extracts, marine extracts, caffeine, theophylline, xanthine derivatives and other lipolytic agents.