JP5927047B2 - TRPV3 activator - Google Patents

Description

  The present invention relates to a TRPV3 activator and a warming agent.

  Sensory sensation occurs when externally received stimuli are converted into electrical signals and transmitted to the brain through nerve cells. In order to convert an external stimulus into an electronic signal, the presence of a receptor that senses the external stimulus is necessary. Among such receptors, a capsaicin receptor (TRPV1) was identified in 1997 as a receptor that senses temperature sensation (Non-patent Document 1). TRPV1 was activated by thermal stimulation at 43 ° C. or more in addition to capsaicin, and thus was expected to be a temperature sensory receptor. Furthermore, since TRPV1-deficient mice show only a dull response to thermal stimulation, it has been confirmed that TRPV1 functions as a heat receptor (Non-patent Document 2).

  However, since TRPV1 activation temperature of 43 ° C. is a temperature that causes pain in the living body, and capsaicin, which is a chemical ligand, is both a warming substance and an analgesic substance, TRPV1 is a pain receptor. It is considered to function as a body (Non-patent Document 3).

  TRPV3 is an ion channel belonging to the same TRP (Transient Receptor Potential) family as TRPV1, and is a temperature receptor identified in 2002 (Non-patent Document 4). Since this receptor has an activation threshold in the range of 30 ° C., it is expected to be a receptor having a warm temperature rather than pain. Furthermore, analysis using TRPV3-deficient mice has confirmed that TRPV3 is involved in temperature acceptance in the skin (Non-Patent Document 5).

  TRPV3 is known to be activated by plant-derived components such as camphor, which is a component of hinoki cypress, and thymol and carvacrol, which are components of thyme and oregano (Non-Patent Document 6).

  Xanthorizole is a kind of sesquiterpene, which is a typical constituent of curcuma xanthoriza, a turmeric genus plant. As the pharmacological action of xanthorizole, antibacterial action (Non-Patent Documents 7 and 8), uterine contraction action (Non-Patent Document 9), apoptosis-inducing action (Non-Patent Document 10), papilloma inhibitory action (Non-Patent Document 11), An inhibitory action on toxicity induced by anticancer agents (Non-Patent Documents 12 and 13) and an inhibitory action on cancer metastasis (Non-Patent Document 14) are known.

  However, it has not been known so far that xantolizole and its analogs have TRPV3 activity action and warming action.

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  The present invention relates to the provision of a TRPV3 activator capable of imparting warm feeling and a warm feeling imparting method using the same.

  As a result of activating TRPV3, the present inventors have found that the alkylphenol derivative represented by the following formula is useful as a material for giving the user a warm feeling, that is, imparting a warm feeling. .

That is, the present invention relates to the following (1) to (4).
(1) The following formula (1):

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
A TRPV3 activator comprising an alkylphenol derivative represented by the formula:
(2) The following formula (1):

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
The warming agent which uses the alkylphenol derivative represented by these as an active ingredient.
(3) The TRPV3 activator according to (1) or the warming agent according to (2), wherein R ¹ is a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ .
(4) The following formula (1):

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
A method for imparting warmth, characterized by using an alkylphenol derivative represented by the formula:

  When the alkylphenol derivative of the present invention is blended in, for example, cosmetics, foods, pharmaceuticals or quasi drugs, the user can take it and apply it safely and simply without causing pain. It can be felt.

The TRPV3 activation of the present invention refers to activating TRPV3 which is a receptor, that is, exhibiting agonist activity with respect to the TRPV3 receptor. The TRPV3 receptor is activated by the TRPV3 receptor agonist activity, and the user perceives it as a warm feeling.
Therefore, the TRPV3 activator and warming agent of the present invention are blended in cosmetics, foods, pharmaceuticals, quasi drugs, etc., and without pain when the user takes and applies them. You can feel the warmth.

Among the alkylphenol derivatives of the present invention, the compound in which R ¹ is —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ is Xanthorrizol, while R ¹ is —CH The compound that is a (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ group is dihydroxanthorizole.
Since the alkylphenol derivative of the present invention has an asymmetric carbon atom, an optical isomer exists. In the present invention, any of R-isomer, S-isomer, and a mixture thereof may be used.
Among the alkylphenol derivatives of the present invention, from the viewpoint of TRPV3 activity, xanthorizole is preferable, and (R)-(−)-xanthorizole represented by the following formula is particularly preferable.

The alkylphenol derivatives according to the present invention are all known compounds and can be chemically synthesized according to a known method or a method analogous thereto. For example, xantolizole has been disclosed by T.W. According to the method of Ehara et al. (Chem. Pharma. Bull., 2007, 55 (9), 1361-1364.) I. It can be produced by the method of Agillar et al. (Rev. Soc. Quim. Mex., 2001, 45 (2), 56-59.).
Xanthorizole is also described in R.I. The method of Horst et al. (From Zeitschrift fuel Natureforschung, Teil B: Anorganische Chemie, Organische Chemie, Biochemie, Biophysik, Biologie 1970, 25 (9), 99c-ur. In addition, (R)-(−)-xanthorizole can be isolated by extraction, fractionation, and purification from kuryu-kon, for example, as shown in Production Example 1.

  The alkylphenol derivative obtained as described above can be further purified using a conventional purification means such as a recrystallization method or column chromatography, if necessary.

  Since the alkylphenol derivative of the present invention has a TRPV3 activating action as shown in the examples described later, the user can feel warmth when applying them. That is, the alkylphenol derivative of the present invention can be a TRPV3 activator or a warming agent, and can be used to produce a TRPV3 activator or a warming agent.

  The TRPV3 activator or warming agent may itself be a pharmaceutical, food, cosmetic, quasi-drug, or the like for activating or imparting warmth to TRPV3, or the pharmaceutical, food, cosmetic. Further, it may be a material or a preparation used by being blended in a quasi drug. The TRPV3 activator or warming agent may be one using the alkylphenol derivative of the present invention alone, or starch, protein, fiber, carbohydrate, lipid, fatty acid, vitamin, mineral, flavor, Even if it is a composition combined with additives and excipients used in pharmaceuticals, quasi drugs, cosmetics, foods, such as coloring agents, sweeteners, seasonings, preservatives, preservatives, and antioxidants Good. Also, their form is not particularly limited, and can be prepared in any form such as a solution, suspension, syrup, powder, granule, particle and the like.

  Here, dosage forms of pharmaceuticals and quasi-drugs are tablets, capsules, granules, powders, syrups, intravenous injections, intramuscular injections, suppositories, inhalants, percutaneous absorption agents, eye drops, dots Nose, poultices, poultices, ointments, lotions, creams, oral preparations (dentifrices, liquid dentifrices, mouthwashes, gum massage creams, oral ointments, gargle tablets, troches, throat lozenges, etc.) Any of these may be used, and the administration form may be any of oral administration (internal use) and parenteral administration (external use, injection).

  The food may be any of candy, gum, tablet, capsule, various beverages and the like.

  As cosmetics, any of skin external preparations, cleaning agents, makeup cosmetics, scalp and hair cosmetics, bath preparations, etc. may be used. Depending on the method of use, they may be lotions, emulsions, gels, creams, ointments. It can be provided in various dosage forms such as agents, powders and granules.

  The blending amount in the preparation when the TRPV3 activator or warming agent of the present invention is blended and used in the above pharmaceuticals, foods, cosmetics, and quasi drugs is not particularly limited as long as it has a sufficient warming effect. For internal preparations (oral medicines, foods, oral compositions, etc.), it should be 0.01 ppm or more, preferably 1 ppm or more, and 500,000 ppm or less, preferably 100,000 ppm or less. For external preparations (parenteral pharmaceuticals, cosmetics, etc.), the content is 0.01% by mass or more, preferably 0.1% by mass or more and 20% by mass or less, preferably 10% by mass or less.

  In the method of imparting a warm feeling using the alkylphenol derivative of the present invention, the TRPV3 receptor is activated by applying (taking, applying, etc.) the compound to a human (user), whereby the user feels warm. It is characterized by feeling. The method for imparting warmth is not particularly limited as long as the user can feel warmth with an alkylphenol derivative, and for example, the form in which the compound or warming agent is directly applied may be used, or other pharmaceuticals Further, it may be applied in the form of being included in cosmetics, foods, quasi drugs and the like.

With respect to the above-described embodiment, the following aspects are disclosed in the present invention.
<1> The following formula (1)

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
A TRPV3 activator comprising a 2,5-dialkylphenol derivative represented by the formula: <2> The following formula (1)

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
The warming agent which uses the 2, 5- dialkylphenol derivative represented by these as an active ingredient.
<3> The TRPV3 activator according to <1> or the warming agent according to <2>, wherein R ¹ is a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ .
<4> Formula (1) below

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
A method for imparting warmth, characterized by using a 2,5-dialkylphenol derivative represented by the formula:
<5> An internal preparation containing the TRPV3 activator of <1> or the warming agent of <2>, wherein the preparation contains an alkylphenol derivative in an amount of 1 ppm to 100,000 ppm.
<6> A preparation for external use containing the TRPV3 activator of <1> or the warming agent of <2>, wherein the preparation contains an alkylphenol derivative in an amount of 0.1% by mass to 10% by mass.

Example 1: Test sample preparation example (-)-Preparation of xanthorizole:

  1.6 vol of 50 vol% ethanol was added to 160 g of rhizome of Curcuma xanthorrhiza, and after immersion extraction at room temperature for 7 days, an extract was obtained by filtration. The extract was dried under reduced pressure to obtain an extract (16.8 g). This extract was dissolved in 50 vol% ethanol and purified using ODS preparative HPLC (60% acetonitrile-0.1% TFA aqueous solution as mobile phase) to obtain (−)-xanthorizole (121 mg). It was. This was dissolved in ethanol to prepare a test sample.

Preparation of (−)-dihydroxanthorizole:
(-)-Xanthorizole (64.2 mg) was dissolved in ethyl acetate (5 mL), 10% palladium carbon (64 mg) was added, and the mixture was stirred at room temperature in a hydrogen atmosphere for 1 hour, and then filtered through Celite. The filtrate was dried under reduced pressure and purified by silica gel chromatography to obtain (−)-dihydroxanthorizole (61.5 mg). This was dissolved in ethanol to prepare a test sample.

Example 2: Evaluation of TRPV3 activity (1) Production of stable expression strain of human TRPV3:
In order to produce a human TRPV3 stable expression HEK293 cell line, the human TRPV3 gene was cloned. The full-length human TRPV3 gene was amplified using RT-PCR method using total RNA extracted from human epidermal keratinocytes (Kurabo) as a template.

  The obtained PCR product was cloned into the entry vector pENTR-D / TOPO (Invitrogen), then subcloned into pcDNA3.2-V5 / DEST (Invitrogen), and HEK293 using Lipofectamine 2000 (Invitrogen). Cells were transduced. Transduced cells were selected by growing in DMEM medium containing 450 μg / ml G-418 (Promega). Since HEK293 cells do not express endogenous TRPV3, they can be used as a control for the TRPV3 transduced strain.

(2) Calcium imaging:
TRPV3 activity transduced into HEK293 cells was measured using fluorescent calcium imaging. First, cultured TRPV3-expressing cells were seeded (30,000 cells / well) in a 96-well plate (BD Falcon) coated with poly-D-lysine, incubated at 37 ° C. overnight, and then the culture solution was removed. Then, Fluo4-AM (2 μg / ml; manufactured by Dojindo) dissolved in Ringer's solution was added and incubated at 37 ° C. for 30 to 60 minutes. After that, the 96-well plate is set in a fluorescence plate reader (FDSS3000; manufactured by Hamamatsu Photonics), and the fluorescence image by Fluo4 when excited at an excitation wavelength of 480 nm in a state where the temperature inside the apparatus is 25 ° C. is detected wavelength. Detection was performed with a CCD camera at 520 nm. The measurement is performed every second for 4 minutes, and 15 seconds after the start of the measurement, the test sample prepared above (final concentration 100 μM) is added by a dispenser with a built-in FDSS 3000, and the activity of TRPV3 is evaluated by the change in fluorescence intensity. .

(3) Evaluation of TRPV3 activation:
In order to eliminate the influence of autofluorescence due to the test sample, TRPV3 activity was obtained by subtracting autofluorescence from the following formula. Fluorescence intensity after subtracting autofluorescence (F _sub ) = fluorescence intensity of TRPV3-expressing cells (F _V3 ) −fluorescence intensity of HEK293 cells (F _HEK ) TRPV3 activation action by xanthorizole and its derivatives Evaluation was made using the peak of the fluorescence intensity ratio later. The fluorescence intensity ratio was calculated using the following formula. Fluorescence intensity ratio = evaluated in F _sub / measurement starting F _sub each treatment 2 wells per group at each time point, and the average value was used.

(4) TRPV3 activation action of alkylphenol derivatives:
Table 1 shows the TRPV3 activation strength of each test sample. As a control, carbachol known to activate TRPV3 was used, and TRPV3 activation intensity by other test samples was calculated with TRPV3 activation by carvacrol being 1.00.

  Xantolizole and dihydroxanthrazole prepared as described above were shown to significantly activate TRPV3 compared to carvacrol.

Claims (3)

Following formula (1)

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
A TRPV3 activator comprising a 2,5-dialkylphenol derivative represented by the formula: Following formula (1)

[Wherein R ¹ represents a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ or a group —CH (CH ₃ ) CH ₂ CH ₂ CH ₂ CH (CH ₃ ) ₂ . ]
The warming agent which uses the 2, 5- dialkylphenol derivative represented by these as an active ingredient. The TRPV3 activator according to claim 1, or the warming agent according to claim 2, wherein R ¹ is a group —CH (CH ₃ ) CH ₂ CH ₂ CH═C (CH ₃ ) ₂ .