The use of agonist-ligand-specific receptor type rarto increase the rate of apoptosis


(57) Abstract:

The invention relates to medicine, specifically to pharmacology and dermatology and cosmetology. A new tool to increase the rate of apoptosis in cancer cells, precancerous cells in the state and in thymocytes. As a tool used agonist-ligand-specific receptor type RAR. The invention expands the Arsenal of tools for the treatment or prevention of cancer, to reduce the effects, leading to graft rejection. In addition, the invention can be used for the prevention and treatment of skin aging. 3 S. and 9 C.p. f-crystals, 1 table.

The present invention relates to the use of certain retinoids for the preparation of pharmaceutical compositions intended to increase the rate of apoptosis. These retinoids can also be used in cosmetic compositions, which, in particular, are designed to prevent and/or fight svetoindutsirovannoi and/or age-related skin aging.

In cell death involves two mechanisms. The first, which is a classic type, is called necrosis. Morphologically necrosis is characterized by swelling of mitochondria and zieleniak. Necrosis occurs in passive or sudden manner. For example, tissue necrosis usually occurs due to the fact that the cells were subjected to physical trauma or chemical poison.

Another form of cell death called apoptosis (Kegg J. F. R. and Wyllie, A. H. , Br. J. Cancer, 265, 239, (1972)), however, in contrast to necrosis, apoptosis does not lead to any inflammatory reaction. It was reported that apoptosis can occur in different physiological States. This is a highly selective form of cell death characterized by well-marked morphological and biochemical characteristics. So, especially characteristic of the condensation of chromatin, which is associated or not with their endonuclease activity, the formation of apoptotic cells and fragmentation of deoxyribonucleic acid (DNA) in the activation of endonucleases in DNA fragments consisting of 180-200 base pairs (these fragments can be detected by electrophoresis in Agronova gel).

Apoptosis can be considered as programmed cell death, which are involved in the processes of development, differentiation and tissue repair. You can also assume that the differentiation, growth and maturation of cells closely associated with apoptosis and th is CLASS="ptx2">

In the field of medicine in many pathological conditions is modified or even unmanaged mechanism of apoptosis. So, it was reported that the intentional modulation of apoptosis, induction or suppression, it may be possible to treat a large number of diseases and, in particular, diseases associated with cell hyperproliferation, as is the case in cancer, autoimmune diseases and allergies, or, on the other hand, diseases associated with the disappearance of cells, as in the case of virus syndrome immunodeficita human (HIV), neurodegenerative diseases (disease Alzheimer) or significant lesions resulting from myocardial infarction.

It was particularly noted in Oncology that a large number of anticancer drugs, such as dexamethasone, cyclophosphamide and cisplatin, is able to induce apoptosis.

In the field of cosmetology apoptosis plays a special role in the signs of skin aging, arising mainly as a result of dysfunction of the main biological mechanisms of the skin. Consequently, it is possible to imagine that any product that induces apoptotic mechanism is a product that is acceptable for the prevention and/or control S="ptx2">

In the field of retinoids known that all-TRANS retinoic acid is a potent modulator (i.e., inhibitor or, on the other hand, stimulator, depending on the nature of the treated cells) differentiation and proliferation of many normal and transformed cell types. For example, it inhibits the differentiation of epithelial cells, such as cells of the stratum corneum of the epidermis. It also inhibits proliferation of many transformed cells, such as melanoma cells. A similar effect on the proliferation or differentiation can occur in the same cell type, as, for example, in the case of cells of the human promyelocytic HL-60; thus, it is known that proliferation of these cells inhibited by all-TRANS retinoic acid, and at the same time, induced their differentiation into granulocytes and their apoptosis.

It is known that, in General, fully TRANS-reviewa acid affects differentiation and cell proliferation via interaction with nuclearization, which is called the RAR receptors (the receptors retinova acid) and which are located in the cell nucleus. Currently identified three subtypes of RAR receptors, additional receptors interact with specific response elements (RARE) in the promoter region of genes, regulated by retinoic acid. To connect with the response elements, RARs heterodimerizes with the receptor of another type, known as RXR receptors. The natural ligand for RXRs is 9-CIS-retinoic acid. RXRs are exemplary regulatory proteins, because they interact with other members of the subgroup steroid /thyroid receptors such as the receptor for vitamin D3(VDR), the receptor for triiodthyronine (TR) u PPARs (peroxisomal receptors, activating proliferation) with the formation of heterodimers, as is the case with RARs. Moreover, RXRs can interact with specific response elements (RXRE) in the form of homodimers. The presence of these complex interactions, the existence of multiple RAR and RXR receptors, which are expressed differently depending on tissue and cell type, explains the pleiotropic effects of retinoids in virtually all cells.

In the literature already describes a large number of synthetic structural analogs of all-TRANS retinoic acid or 9-CIS-retinoic acid, usually called "retinoids". Some of these molecules are able to bind and specifically activate RARs or, on the other hand, RXR. the finally, the other analogues do not show any special election activity against these different receptors. In this regard, for example, 9-CIS-retinoic acid activates both RARs and RXRs without any selectivity to some of these receptors (nonspecific agonist-ligand), whereas all-TRANS-retinoic acid selectively activates RARs (RAR - specific agonist ligand), regardless of the subtype. In General, qualitative terms the substance (or ligand) is specific for receptors of this group (or in relation to a specific receptor in this group), when referred to a substance exhibits a stronger affinity to all receptors of this group (or to a specific receptor in this group) compared to those, which manifests itself in respect of all receptors of any other group (or, respectively, for all other receptors of the same group or another).

It was reported that 9-CIS-retinoic acid and all-TRANS-retinoic acid are modulators of apoptosis (activator or inhibitor of apoptosis, in particular, depending on cell type) and that of these two modulators 9-CIS-retinoic acid is more active, this phenomenon can explain the Noah's acid, which activates only RARs.

In connection with the foregoing, it is of interest to search for new modulators of apoptosis.

In this regard found that the agonist is a ligand that is specific for a receptor type RAR, are excellent inducers of apoptosis for a variety of cell types, and particularly in respect of thymocytes.

Thus, the present invention relates to the use of at least one agonist-ligand-specific receptor type RAR - for the preparation of pharmaceutical compositions, which is designed to increase the rate of apoptosis in at least one cell population, in which apoptosis can be induced through activation of receptor type RAR.

The invention also relates to the use in the cosmetic composition at least one agonist-ligand-specific receptor type RAR, as an inducer of apoptosis in at least one cell populations of the skin, in which apoptosis can be induced by receptor type RAR.

Thus, this composition will give the opportunity to prevent and/or fight with svetoindutsirovannoi or age-related skin aging, in particular, PU which accumulate over time.

Therefore, the invention finally relates to a cosmetic method for preventing and/or dealing with svetoindutsirovannoi age or aging skin, wherein inducing apoptosis cosmetic composition, such as described previously, is applied to the skin.

Pharmaceutical or cosmetic composition of the invention comprises a physiologically acceptable medium.

Under the agonist-ligand-specific receptor type RAR, in accordance with the present invention, understand agonist-ligand, which has the relation R dissociation constants of the ligand for the receptor type RAR to the dissociation constant of the ligand for the receptor type RAR - above or equal to 10 and which induces the differentiation of F9 cells.

So, it is known that all-TRANS retinoic acid and some of its analogues capable of inducing differentiation of mouse embryonic cells teratocarcinoma (F9 cells); therefore, they are treated as agonists for the RAR receptors. Secretion plasminogen activator, which accompanies this differentiation is an indicator of the response of biological reactions F9 cells to retinoids (Skin pharmacol., 1990; 3: PP.256-267).

The Constitution. These tests are described, in particular, in the following sources:

1. "Selective Synthetic Ligands for Nuclear Retinoic Acid Receptor Subtypes in RETINOIDS, Progress in Research and Clinical Applications, Chapter 19 (pp. 261-267), Marcel Dekker Inc., edited by Maria A. Livrea and Lester Packer;

2. "Synthetic Retinoids: Receptor selectivity values and Biological Activity in Skin Pharmacol., Basel, Karger, 1993, Volume 5, pp. 117-127;

3. "Selective Synthetic Ligands for Human Nuclear Retinoic Acid Receptors in Skin Pharmacology, 1992, Vol.5, pp. 57-65;

4. "Identification of Synthetic Retinoids with selectivity values for Human Nuclear Retinoic Acid Receptor-" in Biochemical and Biophysical Research Communications, Vol.186, No. 2, July 1992, pp. 977-983;

5. "Selective High Affinity RAR or RAR Retinoic Acid Receptor Ligands" in Mol. Pharmacol., Vol.40, pp. 556-562.

Other characteristics, aspects, objectives and advantages of this invention will become clearer from the following description, and various specific examples, which are intended to illustrate and not limit the scope of the invention.

Agonists-ligands specific for receptors of type RAR, and which may be mentioned are 6-3-(1-substituted)-4-hydroxyphenyl)-2-naphthoic acid, (E)-4-(1-hydroxy-1-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-2-propenyl)benzoic acid, 4-[(E)-2-(3-(1-of substituted)-4-hydroxyphenyl)-1-propenyl] benzoic acid, 5',5',8',8'-tetramethyl-5',6', 7',8'-tetrahydro-[2',2']binaphthalene-6-carboxylic acid, 2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naftalekete, 6-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalene-2-carbonyl) naphthalene-2-carboxylic acid, 3,7-dimethyl-7-(1,2,3,4-tetrahydro-1,4 and,9b-trimethyl-1,4-metasedimentary-8-yl)-2,4,6-heptatriene acid, 6-(1,2,3,4, -tetrahydro-1,4 and, 9b-trimethyl-1,4-MetaDimension-8-yl)-naphthalene-2-carboxylic acid, 6-[hydroxyimino-(5,5,8,8-tetramethyl-5,6,7,8 - tetrahydro-naphthalen-2-yl-methyl] naphthalene-2-carboxylic acid, 4-[(6-hydroxy-7-(1-substituted)-2-naphthyl] benzoic acid, 5-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-anthracene-2-yl)-thiophene-2-carboxylic acid, (-), -6-[hydroxy-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-methyl] -naphthalene-2-carboxylic acid, 6-(3-adamantane-1-yl-4-prop-2-ynyloxy-phenyl)-naphthalene-2-carboxylic acid, 4-[(2-oxo-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-ethoxy] -benzoic acid, 4-[2-oxo-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-acetylamino]-benzoic acid, 4-[2-fluoro-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-acetylamino]-benzoic acid, 6-[3-(1-substituted-4-(2-hydroxypropyl)phenyl]-2-naphthoic acid, 5-[3-oxo-3-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-propenyl] -thiophene-2-carboxylic acid, 6-[3-(1-substituted-4-(2,3-di-hydroxypropyl)phenyl] -2-naphthoic acid, 4-[3-hydroxy-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-1-prop� 4-[(3-(1-methylcyclohexyl)-4-hydroxyphenyl)vinyl] -benzoic acid, 4-[(E)2-(3-(1-substituted)-4-hydroxyphenyl)-vinyl]-benzoic acid, 4-[3-(1-substituted)-4-hydroxyphenylethyl] -benzoic acid, 5-[3-(1-substituted)-4-hydroxyphenylethyl] -2-thiencarbazone acid, 5-[3-(1-substituted)-4-methoxyphenylacetyl]-2-thiencarbazone acid, 4-[2-(3-tert-butyl-4-methoxyphenyl)-propenyl] benzoic acid, 4-{2-[4-methoxy-3-(1-methylcyclohexyl) phenyl]-propenyl}-benzoic acid, 6-[3- (1-substituted)-4-(3-methoxy-2-hydroxypropyl)-phenyl] -2-naphthoic acid, 2-hydroxy-4-[3-hydroxy-3-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-1-PROPYNYL] benzoic acid, 6-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yloxy)-naphthalin-2-carboxylic acid, 6-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl-sulfanyl)-naphthalene-2-carboxylic acid, 4-[2-propoxyimino-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-acetylamino] benzoic acid, 6-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl-amino)naphthalene-2-carboxylic acid, 1-methyl-4-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydropyrazin-2-yl)-1H-pyrrole-2-carboxylic acid, 2-methoxy-4-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-anthracene-2-yl)-benzoic acid, 4-[2-nonlocking-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-acetylamino] -benzoic acid, (-)-2-hydroxy-4-[3-hydroxy-3-(5,5,8,8-tetramethyl-5,6,7,8-those who on-naphthalen-2-yl)-prop-1-inyl] benzoic acid, 2-hydroxy-4-[3-hydroxy-3-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-buta-1-inyl] -benzoic acid, 6-(3-bromo-5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yloxy)-naphthalene-2-carboxylic acid, 3-[(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-2H-1-benzopyran] -7-carboxylic acid, 4-[3-(3,5-di-tert-butyl-4-hydroxy-phenyl)-prop-1-inyl] benzoic acid, 4-[3-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-prop-1-inyl]-benzoic acid, 4-[3-(5,6,7,8-tetrahydro-5,5,8,8 - tetramethyl-2-naphthyl)-1-PROPYNYL] -salicylic acid, 4[{3-(1-substituted)-4-(2-hydroxyethyl)-phenyl}ethinyl]-benzoic acid and 4-[{3-(1-substituted)-4-(3-hydroxy-propyl)phenyl}ethinyl] -benzoic acid.

In the present invention, it is preferable to use the agonist-ligand-specific receptor type RAR, which have the relation R greater than or equal to 50. These compounds include: 6-3-(1-substituted)-4-hydroxyphenyl)-2-naphthoic acid, 6-(5,5,8,3-tetramethyl-5,6,7,8-tetrahydronaphthalen-2-carbonyl)-naphthalene-2-carboxylic acid, 6-(1,2,3,4-tetrahydro-1,4 and, 9b-trimethyl-1,4 - metasedimentary-8-yl)-naphthalene-2-carboxylic acid, 6-[hydroxyimino-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-methyl] -naphthalene-2-carboxylic acid, 5-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-anthracene-2-yl)-type the acid, 6-[3-(1-substituted-4-(2-hydroxypropyl)-phenyl] -2-naphthoic acid, 6-[3- (1-substituted-4-(2,3-dihydroxypropyl)-phenyl]-2-naphthoic acid, 4-[3-(1-substituted)-4-hydroxyphenylethyl]-benzoic acid, 5-[3-(1-substituted)-4-hydroxyphenylethyl] -2-thiencarbazone acid, 5-[3-(1-substituted)-4-methoxyphenylacetyl] -2-thiencarbazone acid, 6-[3-(1-substituted)-4-(3-methoxy-2-hydroxypropyl)-phenyl] -2-naphthoic acid, 1-methyl-4-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydropyrazin-2-yl)-1H-pyrrole-2-carboxylic acid, (-)-2-hydroxy-4-[3-hydroxy-3-(5,5,8,8-tetramethyl-5,6,7,8 - tetrahydronaphthalen-2-yl)-prop-1-inyl]-benzoic acid and 2-hydroxy-4-[3-hydroxy-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-buta-1-inyl]-benzoic acid.

Especially preferred 6-3-(1-substituted)-4-hydroxyphenyl)-2-naphthoic acid.

Thus, it is possible to use a pharmaceutical composition comprising the agonist-ligand-specific receptor type RAR, in those cases where it is necessary to increase the rate of apoptosis. Naturally, this effect will be achieved only in cell populations, in which apoptosis may be the induction by activation of receptor type RAR and therefore, especially in those where there are receptors type RAR-t to occur mainly in two cases. The first case refers to diseases or disorders associated with inadequate rate of apoptosis. The second case relates to the treatment required at the time of transplantation in order to suppress effects, leading to rejection of the transplanted organ. Thus, it is possible to regulate the increase of immunotolerance in respect of the transplanted organ by reducing the immune response of T cells by inducing their rate of apoptosis.

Diseases or disorders associated with inadequate rate of apoptosis and which can be noted, constitute violations arising from precancerous conditions or cancer, which may be the result of proliferation of certain cell populations, autoimmune diseases, allergic or inflammatory reaction in which the number of cells, causing damage is too high, or some viral infection, in which proteins of the virus have anti-apoptotic effect.

Thus, in the case of autoimmune diseases special mention should be made insulin-dependent diabetes, chronic active hepatitis, rheumatoid arthritis, pemphigus, multiple Sak precancerous condition. In the case of cancer it should be called lymphoma, carcinoma, such as cancerous lesions, endocrinology sphere and gormonozavisimye tumors, such as cancerous lesions of the ovary. From the above-mentioned viral infections especially it is necessary to transfer the herpes viruses, adenoviruses and variola viruses (tospovirus). In a number of allergies and inflammatory reactions that may be mentioned include contact eczema, atopic eczema, asthma and urticaria.

In accordance with the present invention the composition can be entered enteral, parenteral, local and intraocular way. Preferably the pharmaceutical composition is Packed in a form acceptable to the systemic route of administration (injection or infusion).

With the introduction of enteral through composition, in particular pharmaceutical composition, may be in the form of tablets, hard gelatin capsules, coated tablets, syrups, suspensions, solutions, powders, granules, emulsions, microspheres or nanospheres or lipid or polymeric vesicles, in which it is possible to achieve a controlled release. With the introduction of parenteral composition may be in the form of solutions or suspensions for infusion or injection.

The local introduction of a pharmaceutical or cosmetic composition according to the invention is intended for treating the skin and mucous membranes and may be in the form of ointments, creams, jelly, lipstick, powders, absorbing buffers, solutions, gels, sprays, lotions or suspensions. It can also be in the form of microspheres or nanospheres or lipid or polymeric vesicles or polymeric patches or hydrogels, in which it is possible to achieve a controlled release. The song, which is administered by local, may be either anhydrous or in aqueous form.

When introduced into the eye, the composition is mainly in the form of eye drops.

The agonist-ligand-specific receptor type RAR-use for cutaneous or intraocular injection in a concentration that is typically between about 0.001% and 10% by weight, preferably between 0.1 and 1% by weight of the composition.

Finally, the present invention relates to a method for preventing and/or dealing with svetoindutsirovannoi or age-related skin aging, characterized in that the cosmetic HDMI is epicly for receptor type RAR, in a cosmetically acceptable medium, applied to the skin.

Naturally, the composition previously described, may further include an inert or even pharmacodynamic active additives or combinations of these additives, in particular: wetting agents; depigmenting substances such as hydroquinone, azelaic acid, caffeic acid or kojic acid, softening means; moisturizing agents such as glycerol, PEG 400, thiomorpholine and its derivatives or urea; substances dandruff or against acne, such as S-carboxymethylcysteine, S-benzyl group probably facilitates and their salts or derivatives, or it with benzoyl; antifungal agents, such as ketoconazole or polymethylene-4,5-isothiazoline-3-ones; antibacterial agents; carotenoids and, in particular, -carotene; agents against psoriasis, such as anthralin and its derivatives; and, finally, eicosa-5,8,11,14-tetralogia or eicosa-5,8,11-TRINOVA acid and their esters and amides.

These compositions can also contain substances that improve the taste, preservatives such as esters of parahydroxybenzoic acid, stabilizers, substances that regulate the humidity, substances that regulate pH, substances for modifying the osmotic pressure, emulsifiers, is 2">

Naturally, the person skilled in the art it is clear that the connection(I), add(s) in these compositions is selected/chosen such that the advantages of the present invention do not change or change negligible when added.

Below are some examples, which are intended to illustrate the present invention but not to limit its scope.

Example 1

This experiment demonstrates the effectiveness of agonist-ligand in vivo, specific for RAR-, as an inducer of apoptosis.

Used male NMRI mice at 4 weeks of age (LATI , Hungary). In order to induce apoptosis in the thymus, these male mice administered a single injection of either 0.5 mg of dexamethasone, or 0.5 mg 6-3-(1-substituted)-4-hydroxyphenyl)-2-naphthoic acid, dissolved in a mixture of 0.8 ml of saline and 0.2 ml of ethanol.

Dexamethasone is a well-known inducer of apoptosis. So, there is involution of the thymus: i.e., a decrease of approximately 75% of the weight of the thymus 48 hours after the above processing. A similar effect is observed with the introduction of 6-3-(1-substituted)-4-hydroxyphenyl)-2-naphthoic acid, in this case, is in a similar experiment, through various time intervals after treatment was selected samples of the thymus in untreated and treated animals. After washing and homogenization was determined by the activity of transglutaminase on the inclusion of [3H] putrescine N,N'-dimethylbutan. The activity of the transglutaminase is calculated nolah [3H] putrescine incorporated into protein per hour.

It was reported that tissue transglutaminase is one of the elements involved in the development of apoptosis (Piacentini M. et al. (1994). Apoptosis: The Molecular Basis of Apoptosis in Disase. Curr. Comm. in Cell Mol. Biol., 8 (Tomei L. D. and Cope, T. O. ed) pp.143-165). Thus, observed that the above involution of the thymus occurs as a result of processing mouse agonist-ligand, specific RAR, and is a phenomenon which accompanies and is proportional to the activity of tissue transglutaminase.

Moreover, such an involution of the thymus correlate with the advent of DNA-fragments that define the analysis electrophoresis Agronova gel of DNA isolated from the thymus of treated animals (detailed methodology is presented in example 2).

Example 2

This example shows the effectiveness of in vitro agonist-ligand-specific receptor type RAR, as the inductor AP is pensii of thymocytes were prepared from thymus raw 4-week-old male NMRI mice (supplied LATI , Hungary). As the medium used medium RPMI 1640, Sigma, enriched with fetal calf serum, Gibco, 2 mm glutamine, 100 IU of penicillin and 100 μg streptomycin/ml Then the thymocytes were washed and diluted to a final concentration of 107cells/ml before incubation at 37oWith in a humidified incubator in an atmosphere with 5% CO2and 95% air. Cell death was measured by the absorption Trypanosoma blue.

Qualitative and quantitative analysis of DNA

The thymocytes were incubated in 24 holes with various test substances at different concentrations. After 6 hours incubation, 0.8 ml of cell suspensions were literally by adding 0.7 ml of lyse buffer containing 0.5% (volume/volume) Triton X-100, 10 mm Tris, 20 mm EDTA, pH 8.0 before centrifugation at 13,000 g for 15 minutes.

Quantitative analysis of DNA: DNA that is located in nagadoches fluid (fragments) and sediment (intact chromatin) was besieged by an equivalent amount of 10% trichloroacetic acid, resuspendable in 5% trichloroacetic acid and then was quantified using diphenylamine reagent (Burton K. (1956), Biochem. J., 62, 315-322).

Qualitative analysis of DNA: parallel nadosadocnuu liquid was treated with ethanol, including the future 10 mm Tris, 1 mm EDTA, pH 8.0, and these solutions are then processed RNA Asai; then was extracted with an equal volume of phenol and chloroform/isoamyl alcohol (24/1), after which DNA was besieged in ethanol before electrophoresis for 3 hours at 60 v 1.8% Agronova gel. The DNA fragments were then shown a UV light after staining the gel ethidium bromide. The resulting gels was a picture of the "ladder" of DNA fragments consisting of 180-200 base pairs and which were typical for the induction of apoptosis. During the whole experience fragmentation correlated with the number of dead cells, giving a positive reaction with Trifanova blue.

The results of the quantitative analysis presented in the table.

The percentage of DNA fragments in this table corresponds to the difference between the percentage of DNA fragments obtained in the treated thymocytes, and the percentage of DNA fragments obtained in untreated thymocytes (basic rate of apoptosis for these thymocytes).

Data show that the percentage of occurrence of DNA fragments increases with R. Thus, the effect of inducing apoptosis increases with the specificity of the ligand in relation to the RAR.p

1. Prime induction of apoptosis in, at least one population of cancer cells or cells in precancerous condition, or in thymocytes.

2. Application under item 1, wherein the agonist is a ligand to the receptor type RAR - represents 6-3-(1-substituted)-4-hydroxyphenyl)-2-naphthoic acid, (E)-4-(1-hydroxy-1-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-2-propenyl)benzoic acid, 4-[(E)-2-(3-(1-of substituted)-4-hydroxyphenyl)-1-propenyl]benzoic acid.

3. Application under item 1 or 2, wherein the agonist is a ligand to the receptor type RAR has the relation R above or equal to 50.

4. Use one of the PP.1-3, wherein the agonist is a ligand to the receptor type RAR - represents 6-3-(1-substituted)-4-hydroxyphenyl)-2-naphthoic acid.

5. Use one of the PP.1-4 for the treatment of diseases or disorders associated with insufficient speed of apoptosis.

6. Application under item 5, wherein the diseases or disorders associated with insufficient speed apoptosis are cancer or precancerous conditions.

7. Application under item 6, characterized in that cancers are cancer of endocrine diseases and cancerous lesions of the ovary.

9. The use of at least one agonist is a ligand that is specific for receptors of the retinoic acid type RAR, in cosmetic compositions as an inducer of apoptosis in at least one population of skin cells, in which apoptosis can be induced through activation of receptor type RAR.

10. Application under item 9, wherein the agonist is a ligand specific for the receptor type RAR, selected from the compounds listed in paragraph 2.

11. Cosmetic method for preventing and/or dealing with svetoindutsirovannoi age or ageing of the skin, characterized in that a cosmetic composition, inducing apoptosis and comprising a cosmetically acceptable medium, at least one agonist is a ligand that is specific for receptors of the retinoic acid type RAR-applied to the skin.

12. The method according to p. 11, wherein the agonist is a ligand specific for the receptor type RAR, selected from the compounds listed in paragraph 2.2


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