WO2015114176A1

WO2015114176A1 - Cosmetic product with anti-skin-aging properties

Info

Publication number: WO2015114176A1
Application number: PCT/ES2014/070068
Authority: WO
Inventors: Gabriel SERRANO SANMIGUEL; Juan Manuel SERRANO NUÑEZ; Ana TORRENS TOMAS
Original assignee: Dermopartners, S.L.
Priority date: 2014-01-31
Filing date: 2014-01-31
Publication date: 2015-08-06

Abstract

The invention relates to a cosmetic product with anti-skin-aging properties, containing trepenone, tripeptide-32, L-glutamylamidoethyl imidazole, as the main active ingredients, and other active ingredients such as hyaluronic acid, adenosine, phytosphingosine, lysate of bifidobacteria and ascorbic or retinyl palmitate, some of which are liposomal.

Description

COSMETIC PRODUCT WITH PROPERTIES AGAINST

THE AGING OF THE SKIN

OBJECT OF THE INVENTION

The present invention relates to a cosmetic product that has properties to combat skin aging and that has its application in the field of beauty, cosmetics and dermatology centers.

BACKGROUND OF THE INVENTION

A multitude of different preparations to combat skin aging are known to date but none having the composition of the cosmetic product described herein.

DESCRIPTION OF THE INVENTION

The invention claimed is a new cosmetic preparation against skin aging that has a series of active ingredients, of which the most relevant are trepenone, tripeptide-32 and L-glutamyl amidoethyl imidazole, in addition to others such as hyaluronic acid, adenosine, phytosphingosine, lysate of bifidobacteria and ascorbyl palmitate or retinol palmitate and some of them are liposomes. The age we attribute to a face is a global and unconscious visual evaluation of various signs of skin appearance.

A complete anti-aging treatment should provide a youthful appearance to the face:

- Recovering the loss of tone (sagging)

- Reducing pigmentation related to the sun or redness

- Avoiding dry skin

- Avoid dilated pores

- Improve surface regularity

All these skin dysfunctions are related to the lower physiological efficiency of the cells, this leads us to a composite tissue "cells aged ", which enter the senescence phase more quickly than necessary due to the successive damage to DNA that is repaired less efficiently and also a decrease in the protection of telomeres by an enzyme known as telomerase.

In this sense, the cosmetic product described here consists of a series of active ingredients that decrease the number of cells that enter the senescence phase while promoting the repair of skin cell DNA. The most relevant active ingredients and adjuvants found in the composition of the product object of the patent are the following:

Renovage®

Tripeptide Liposomes-32

Chronocyclin®

Hyaluronic acid

Sk-lnflux®

Hyaluronic Acid Liposomes

Phytosphingosine liposomes

Adenosine Liposomes

Liposomes of bifidobacteria lysate

Filagrinol®

Each of these active ingredients has properties that contribute to the product having these properties to combat skin aging and are indicated below:

RENOVAGE ®

INCI: Teprenone and Caprilico / Caprico Triglicerido

GGA: Geranil geranone active against senescence

Improvement of tissue quality through optimal cellular interactions (cellular communication).

Rebalance of cellular functions (metabolism).

Protection against aging and stress thanks to the stabilization effect of telomeres and the maintenance of DNA (cell division). Fight against all signs of aging functionally: dehydration, age spots, and structurally: wrinkles, pores, erythrosis.

Mechanism of action:

Geranylgeranyl isoprene is a key molecule in cellular functioning, both at the level of membrane receptors, and in the functionality of intracellular proteins and their signaling.

It can be considered as a physiological facilitator.

The telomere is a repeated sequence of nucleotides that terminates the chromosomes and is partially bent over their end to protect it.

In each cycle of cell division, the chromosomes are detangled, duplicated and then recompacted. When the telomere reaches a short critical length, due to its progressive erosion after each replication, the cells enter senescence.

Telomeres can also be damaged by UV radiation and exogenous molecules.

TRIPEPTIDE LIPOSOMES-32

The properties of the active substance tripeptide-32 are: · It is a "CLOCK GEN ACTIVATOR", that is, an ingredient that activates one or more biological clock genes present in keratinocytes to produce proteins that repair DNA damage.

• It is a "DNA repair enzyme", an enzyme that is able to function to repair DNA damage.

· Enzyme capable of "Repairing" means, with respect to the skin, that the viability of keratinocytes, strength and longevity were generally improved.

• It is able to penetrate the cell and nuclear membranes to act at the level of genes. Mechanism of action:

- Healthy keratinocytes: internal natural mechanisms for the repair of lesions, or DNA mutations (Timin dimers).

- CLOCK and PER1, genes that are activated by the day to produce proteins that protect cells from damage. - As protein levels increase during the day, feedback inhibition occurs and genes "turn off" as night approaches.

- Tripeptide-32 is an activator of the CLOCK and PER1 genes which in turn are activators of keratinocytes. Activation of these genes promotes cell viability, cell longevity, inhibition of cell damage due to environmental aggressions, and repair by healthy keratinocytes of DNA damage.

CHRONOCYCLIN®

INCI: L-Glutamyl Amidoethyl Imidazole

Glutamyl Amidoethyl Imidazole GAEI is a pseudopeptide derived from histamine that restores and stimulates the skin's natural defense system and cellular metabolism, in a mechanism of action similar to cytokines. It produces in the skin an optimization of the activity of Vitamin D and microcirculation, a resynchronization of circadian rhythms and a stimulation of circadian genes.

Its effects on skin cells are as follows:

- Increase the expression of circadian genes: CLOCK AND PER 1

- Optimize cell defenses.

- Optimizes cell renewal.

- Activates the phototransformation of Vitamin D

Phytosphingosine LIPOSOMES

• Ceramides are fatty amides of sphingosine or phytosphingosine. They are located in the stratum corneum of the skin and form a substantial part of the lipid barrier of human skin, which is responsible for the aggressions of the external environment and to maintain the balance of hydration necessary for its good conservation.

• Moisturizer

• Softener. Calms with a protective effect of the epidermis. Hyaluronic Acid of High Molecular Weight

• Moisturizing substance that retains water molecules and ensures the cohesion of connective cells. Together with the mucopolysaccharides, it constitutes the fundamental substance of the dermis. • Ability to retain water in a percentage equivalent to thousands of times its weight.

• Forms a moisturizing film.

• Immediate flash effect.

SK-INFLUX ®

INCI: Ceramide NP, Phytosphingosine, Ceramide AP, Ceramide EOP

• It is a product that consists of a concentrate of skin-like lipids that restore the skin's protective barrier function.

• A concentrated mixture of different types of ceramides, cholesterol, free fatty acids and phytosphingosine that make it an ideal ingredient for personal care products with a unique restorative capacity.

• Its application translates into greater hydration and protection, also leads to less sensitive and less dry skin.

ADENOSINE LIPOSOMES

- It has an important function in biochemical processes, such as energy transfer, in the form of ATP and ADP

- It is an antioxidant and therefore it is used in cosmetics to prevent aging.

- Wrinkle effect.

FILAGRINOL®

INCI: Europalea Olea Oil, Unsaponifiable Soy Wisteria Oil,

Triticum Vulgare, Pollen Extract.

- Filagrin is a protein synthesized in the epidermis of the granular stratum. It has a key role in the keratinization process:

1. Because it promotes the organization and aggregation of the stratum keratin protein chains

2. As a consequence of its degradation, a group of water-soluble molecules that make up the natural moisturizing factor (NMF) is generated, which provides integrity and good health to the stratum corneum and, consequently, to the entire epidermis. - Among its applications are: treatment of dehydrated, sensitive, dry and reddened skin; treatment of skin spots and other forms of redness; the treatment of skin aging caused by sun damage. LIFOSOMAS USED FOR BIFIDOBACTERIES

INCI: Lysate ferment of bifida.

- Bifidobacteria lysate. It is a type of digestive bacteria, which can prevent damage induced by UV radiation in the epidermis and dermis, stimulating the enzymatic repair of the endogenous cellular system.

- It is a true DNA repair agent

- Beneficial effects on the skin:

- Decreased skin sensitivity.

- Increased resistance of the skin barrier.

- Maintenance of the concentrations of the hydration factor in the skin (urea).

PREFERRED EMBODIMENT OF THE INVENTION

As a preferred embodiment, a liposome cosmetic product is proposed, which has different cosmetic forms such as solution, serum, emulsion, suspension, etc. in which the different components can be found in the case of nourishing cream in the following proportions of active ingredient and liposomes:

ARGAN 1-2 OIL

PHANOXYETHANOL 1-2

INSAPONIFYABLE Soy GLYCINE OIL <1

INSAPONIFICABLE TRITICUM VULGARE GERMEN OIL <1

CYCLOPENTASILOXANE <1

LECITINE <1

POLISILICONA-1 1 <1

TOCOFERIL ACETATO <1

ALCOHOL <1

CYCLHEXASILOXAN <0.5

CITRONELIL METHYLROTONATE <0.5

PERFUME <0.5

LAURIL SODIUM LACTILATE <0.5

POLYSORBATE 20 <0.5

CARBOMER <0.5

XANTANA RUBBER <0.5

DISODIC EDETATE <0.5

ETILHEXI LGLICERINE <0.5

SODIUM HYDROXIDE <0.5

POLEN EXTRACT <0, 1

TREPENONE <0, 1

BIFIDOBACTERIA LISTING <0.1 LIF BIFIDA 5-6%

TRIPEPTIDE-32 <0, 1 LIP.TRIPEPTIDE 5-6%

CERAMIDE NP <0, 1

PROPILEN GLICOL <0, 1

SODIUM HIALURONATE <0, 1

AP CERAMIDE <0, 1

PHYTHOSPHINGOSINE <0, 1

CHOLESTEROL <0, 1

SODIUM CATATO <0, 1

TOCOPHEROL <0, 1

ASCORBILE PALMITATE <0, 1

ADENOSINE <0.1 LIP. ADENOSINE 1-2%

GLICERIL OLEATO <0, 1

METHYLABLES <0.01

ETILPARABEN <0.01

PROPILPARABEN <0.01

PHYTOSPHINGOSINE HCL <0.01 PHYPHOSPHINGOSINE LIP 1-2%

CITRIC ACID <0.01

CHLORIDE ACID <0.01

CERAMIDE EOP <0.01 In a second preferred embodiment, as in the case of a moisturizing serum, the different components are in the following proportions of active ingredient and liposomes:

DESCRIPTION OF THE DRAWINGS

To complement the description that is being made, and in order to help a better understanding of the characteristics of the invention, a series of figures corresponding to the results of the tests carried out on fish embryos, with this type of products.

In these figures, for illustrative purposes the following has been represented:

Figure 1.- The results against the control of the expression of the Perl gene Figure 2.- The results against the control of the expression of the ClockEst gene. Figure 3.- The results against the control of the expression of the Telomerase gene

Figure 4.- The results against the control of the expression of the Cbx5 / HP1 gene.

CLINICAL STUDIES As a justifying complement to the efficacy of the product claimed in this patent, two different clinical trials have been carried out.

On the one hand, a clinical trial has been carried out with the objective of evaluating how the cosmetic product of the present patent modifies the expression of two genes, Perl and Clock, in fish eleutheroembryos. On the other hand, an assay has been made with the objective of evaluating how the cosmetic product of the present patent modifies the expression of two genes, Telomerase and CBX5 / HP1. 1.- Modification of the Perl and Clock genes

As for the modification of the expression of the Perl and Clock genes, the product of the Clock gene regulates the biological clock through circadian rhythms and metabolism. The protein encodes a transcription factor of the basic helix-loop-helix family (bHLH) and a DNA-binding histone acetyltransferase.

The Perl gene is a member of the Period family of genes and is expressed in a circadian pattern in the suprachiasmatic nucleus of the brain, the primary pacemaker of the mammalian brain. The genes of this family encode components that regulate circadian rhythms of locomotor activity, metabolism and behavior. Circadian expression in the suprachiasmatic nucleus continues in constant darkness and a change in the light / dark cycle causes a proportional change in the gene expression in the suprachiasmatic nucleus. The specific function of this gene is not yet known.

- Process:

First, the set-up of the pairs of primers was carried out for each of the genes studied in the trial.

The quantitative PCR measurement experiment (qPCR) was done in triplicate for each of the genes, and three independent experiments (3x3) were performed, each gene being tested a total of 9 times. In each of the samples 15 embryos from the same batch collected 4 days before the test were used, and common treatments for several samples were performed simultaneously as synthesized in the diagram.

After completion of the corresponding treatment for each sample, the embryos were immediately frozen in liquid nitrogen, RNA was extracted, cDNA was synthesized and quantitative PCR (qPCR) was performed in real time. Total RNA extraction was performed using the TRIZOL reagent and subsequently treated with the DNA-free kit. The concentration of the RNAs was quantified with the Nano-Drop spectrophotometer and the cDNAs were synthesized from 1 RNA using Random Primers hexamers and Superscrip III reverse transcriptase. By means of RT-PCR it was verified that the cDNA synthesis process had worked properly. The qPCRs were subsequently performed using 1.7 ng of cDNA per reaction, with SYBrGreen Master Mix and 0.25 mM of each pair of direct and reverse oligonucleotides. The PCR conditions were: 40 cycles of denaturation at 95 ° C-15 seconds and banding at 58 ° C-30 seconds, fluorescence reading being taken in this last step. The melting curve was performed by plotting the fluorescence data against temperature (55 ° C to 95 ° C) to verify that there was only one amplification peak. Control sample without cDNA was included to verify the absence of non-specific amplification products. For each sample, the threshold cycle (Ct) of the internal control (Actin) was subtracted from the threshold amplification value of the study gene cycle (Ct; Perl or ClockEst). The mathematical method Pfaffl was used to calculate the relation of the relative gene expression related to the value of Actin (internal control). The qPCR reactions were performed in triplicate for each experiment.

- Conclusions:

The treatment of fish eleutheroembryos with the SesGen32 serum increases the expression of the Perl gene by 12 ± 3% (range 9-15%, p <0.0005) and with a 95% confidence interval between 6-17%. (Fig. 1)

The treatment of fish eleutheroembryos with the SesGen32 cream inhibits the expression of the Clock gene by 8 ± 3% (range 5-1 1%, p <0.05) and with a 95% confidence interval between 1-14% . (Fig. 2)

2.- Modification of the expression of the Telomerase and CBX5 / HP1 genes. The Telomerase (Tert) gene encodes a ribonucleoprotein polymerase that maintains the length of telomeres by adding the TTAGGG telomeric repeat. The enzyme consists of a protein component with reverse transcriptase activity, encoded by the Tert gene, and an RNA component that acts as a template for telomeric repeat. Telomeres are essential to keep the ends of the chromosomes intact and prevent their degradation. Telomerase maintains the length of telomeres in the cells in which it is expressed (mainly in embryonic cells, stem cells and gonadal cells) and prevents their shortening, which would cause progressive degradation of DNA and cellular aging. Telomerase expression plays a key role in cellular senescence, since it is normally inhibited in postnatal somatic cells, resulting in a progressive shortening of telomeres. Studies in the mouse suggest that telomerase also participates in chromosomal repair, since de novo synthesis of telomeric repeats could occur in the breaks of two strands of DNA.

The Cbx5 / HP1 gene (Chromobox homolog 5) encodes a highly conserved non-histone protein that is a member of the heterochromatin protein family. The Cbx5 protein has a single Chromo domain at the Nterminal end that can bind to histone proteins through methylated lysine residues and a "shadow" chromo domain (Chromo shadow domain, CSD) that is responsible for homodimerization and interaction with a number of non-histone proteins associated with chromatin. The protein is enriched in heterochromatin and is associated with centromers. Cbx5 is involved in the epigenetic regulation of gene expression and interacts with telomeric sequences to maintain its integrity.

Therefore, the Tert and CBX5 / HP1 genes are involved in maintaining the integrity of cell telomeres and inhibiting cell senescence.

- Process:

First, the set-up of the pairs of primers was performed for each of the medaka genes (Tert and Cbx5 / HP1) whose expression was analyzed in the assay. For this, a mixture of cDNA synthesized from mRNA of embryos of medaka of different stages of development. The primers were tested by PCR with a temperature gradient in the "annealing" phase to determine the most efficient temperature range of the primers. Three independent experiments of eleutheroembryos treatment with the cream were performed and in each experiment the expression of each gene was determined in triplicate by qPCR, each gene being tested a total of 9 times.

In each of the samples 15 embryos of 96 h from the same batch were used and the treatments were performed simultaneously. After completion of the corresponding treatment for each sample, the embryos were immediately frozen in liquid nitrogen, RNA was extracted, cDNA was synthesized and quantitative PCR (qPCR) was performed in real time. Total RNA extraction was performed using the TRIzol ® reagent and subsequently treated with the DNA-free kit to remove traces of DNA.

The concentration of the RNAs was quantified with Nano-Drop spectrophotometer and the cDNAs were synthesized from ^ g of RNA using Random Primers hexamers and Superscrip III reverse transcriptase. By means of RT-PCR it was verified that the cDNA synthesis process had worked properly. The qPCRs were subsequently performed using 1.7 ng of cDNA per reaction, with SYBrGreen Master Mix and 0.25 mM of each pair of direct and reverse oligonucleotides. The PCR conditions were: 40 cycles of denaturation at 95 ° C-15 seconds and banding at 58 ° C-30 seconds, fluorescence reading being taken in this last step. The melting curve was performed by plotting the fluorescence data against temperature (55 ° C to 95 ° C) to verify that there was only one amplification peak. Control sample without cDNA was included to verify the absence of non-specific amplification products. For each sample, the threshold cycle (Ct) of the internal control (Actin) was subtracted from the threshold amplification value of the study gene cycle (Ct; Telomerase, CBX5 / HP1). The mathematical method Pfaffl was used to calculate the relation of the relative gene expression related to the value of Actin (internal control). The qPCR reactions were performed in triplicate for each experiment.

- Conclusions: The treatment of fish eleutheroembryos with the product of the present patent increases the expression of the Telomerase gene by 26 ± 9% (range 17-35%, p <0.005) and with a 95% confidence interval between 7-44% . (Fig. 3)

The treatment of fish eleutheroembryos with the product of the present patent inhibits the expression of the Cbx5 / HP1 gene by 10 ± 4% (range 6-14%, p <0.05) and with a 95% confidence interval between 1-18% (Fig. 4)

Once the nature of the present invention has been sufficiently described, it only remains to be added that said invention may undergo certain variations in the components and percentages, as long as said alterations do not substantially vary the characteristics claimed below:

Claims

1. - Cosmetic product with properties to combat skin aging characterized by comprising: - a first active ingredient consisting of tripeptide-32 liposomes. A second active substance consisting of trepenone. a third active ingredient consisting of L-Glutamyl Amidoethyl Imidazole.

2. - Cosmetic product with properties to combat skin aging according to claim one characterized in that the total weight percentage of tripeptide-32 is <0.1% and the percentage of liposomes of tripeptide-32 is 5 -6% in the cream and 10-12% in the serum.

3. - Cosmetic product with properties to combat skin aging according to claim one, characterized in that the percentage of trepenone is <0.1%.

4. Cosmetic product with properties to combat skin aging according to claim one characterized in that the percentage of L-Glutamyl amidoethyl Imidazole is 0.2-1%

5. Cosmetic product with properties to combat skin aging according to claim one, characterized in that, in addition to trepenone, tripeptide-32 and L-Glutamyl amidoethyl Imidazole, it comprises as active ingredients:

Sodium hyaluronate.

Phytosphingosine

Adenosine liposomes

Liposomes of bifidobacteria lysate.

6. Cosmetic product with properties to combat skin aging according to claim 5, characterized in that the percentages of the active ingredients of claim 5 are as follows: Sodium hyaluronate in a percentage of total weight <0.1% in the cream and in a percentage of total weight of <0.001% and a percentage of liposomes of 1-2% in the serum.

Phytosphingosine liposomes with a percentage of phytosphingosine in total weight <0.1% and a liposome percentage of 1-2%.

Adenosine liposomes with a percentage of adenosine in total weight <0.1% and a percentage of liposomes of 1-2%.

Liposomes of bifidobacteria lysate with a percentage of bifidobacteria lysate <0.5% and a percentage of liposomes of 5-6% in the cream and 10-12% in the serum.