JP5854674B2 - Dipeptidyl peptidase IV inhibitor - Google Patents

Description

  The present invention relates to dipeptidyl peptidase IV inhibitors.

  Dipeptidyl peptidase IV (hereinafter also referred to as “DDPIV”) is one of serine proteases and has an enzyme activity of recognizing the second proline or alanine from the N-terminus and cleaving the C-terminal side. DDPIV is expressed on epithelial and endothelial cells of tissues such as kidney, liver, intestinal tract, and placenta, and on the cell surface of T cells, and is considered to be involved in various physiological phenomena through its enzyme activity and the like.

  As a substrate for DDPIV, there is a hormone called incretin. Incretin is a general term for hormones secreted from the intestine by stimulation of nutrients and promoting insulin secretion from pancreatic β cells in a blood glucose-dependent manner, and GLP-1 and GIP are known. These incretins not only promote blood glucose-dependent insulin secretion, but also suppress glucagon secretion from α cells, lower blood pressure, suppress gastric emptying, and suppress food intake by acting on the hypothalamus. (See Non-Patent Document 1).

  However, since incretin is degraded by DDPIV, it is known that, for example, the half-life of GLP-1 in vivo is about 1.5 minutes. Therefore, if the enzyme activity of DDPIV can be inhibited, the half-life of incretin in the living body can be extended. As a result, type 2 diabetes, obesity, hypertension, insulin resistance can be achieved through the action of incretin. It is expected to be useful for such treatment.

  DPPIV is identical to CD26, which is one of T cell activation markers, and is known to control its activity using many immunoregulatory peptides as substrates. Therefore, it is considered that the immune response can be controlled by controlling the activity of DDPIV. In fact, in animal models such as autoimmune diseases such as rheumatoid arthritis and rejection due to transplantation, DDPIV inhibitors are known to function as immunosuppressive agents, and are used for the treatment of autoimmune diseases and transplant rejection. It is considered useful.

  Furthermore, DDPIV is known to be involved in the metabolism of several neuropeptides and growth hormones; invasion, metastasis, angiogenesis, etc. in cancer; HIV infection of lymphocytes, and the like. Therefore, by inhibiting the activity of DDPIV, neuropathy such as pain, neurodegenerative diseases and neuropsychiatric diseases (eg sciatica, Alzheimer's disease, depression, etc.); growth hormone deficiency and growth hormone are used for treatment Diseases; cancers (eg, T-cell lymphoma, acute lymphoblastic leukemia, thyroid cancer, basal cell cancer, breast cancer, etc.); diseases such as HIV infection (AIDS) may be treated.

  Conventionally, fluoropyrrolidine (Patent Document 1), proline derivative (Patent Document 2) and the like have been proposed as compounds having a DDPIV inhibitory action, but these are synthetic compounds, which are sufficient in terms of safety. Absent. Therefore, it is desired to provide a component derived from a natural product having a DDPIV inhibitory action and high safety.

JP 2007-63286 A JP 2007-537231 A

Journal of the Japanese Pharmacological Society, 2005, Vol. 125, p. 379-384

  An object of the present invention is to find a dipeptidyl peptidase IV inhibitory action among highly safe natural products, and to provide a dipeptidyl peptidase IV inhibitor containing the same as an active ingredient.

  In order to solve the above-mentioned problem, the dipeptidyl peptidase IV inhibitor of the present invention is one selected from the group consisting of a grapevine extract, a sesame peptide, a lemon balm extract, a ginkgo extract, a yacon extract and a bodaige extract Alternatively, it contains two or more kinds of natural product-derived components as active ingredients.

  According to the present invention, a highly safe dipeptidyl peptidase IV inhibitor having an excellent dipeptidyl peptidase IV inhibitory action can be provided.

Embodiments of the present invention will be described below.
The dipeptidyl peptidase IV inhibitor of this embodiment is one or more natural products selected from the group consisting of grapevine extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract and bodaige extract A derived component is contained as an active ingredient.

  Here, in the present embodiment, the “extract” refers to an extract obtained using the above plant as an extraction raw material, a diluted or concentrated solution of the extract, a dried product obtained by drying the extract, or these Either a crude product or a purified product is included.

[Manufacture of vine grape extract, lemon balm extract, ginkgo biloba extract, yacon extract, body extract]
In this embodiment, the raw materials used to produce the grape extract, lemon balm extract, ginkgo biloba extract, yacon extract and bodaiju extract are grapevine (scientific name: Ampelopsis brevipedunculata Trautv.), Lemon balm (scientific name: Melissa). officinalis L.), Ginkgo (scientific name: Ginkgo biloba L.), Yacon (scientific name: Smallanthus sonchifolia), and Bodaige (scientific name: Tilia cordata).

  Grapevine (Ampelopsis brevipedunculata Trautv.), Also known as gabion, is a deciduous vine plant that grows naturally in Hokkaido from Okinawa, China, and the Korean Peninsula, and is easily available from these areas. Examples of the constituent parts of grape vine that can be used as an extraction raw material include, for example, stem parts, vine parts, leaf parts, fruit parts, flower parts, etc. Among them, in particular, one kind of stem parts, vine parts and leaf parts Or it is preferable to use 2 or more types.

  Lemon balm (Melissa officinalis L.) is a perennial herb of Lamiaceae distributed throughout Europe, Central Asia, North America, etc., and can be easily obtained from these regions. Examples of the constituent parts of lemon balm that can be used as an extraction raw material include the above-ground parts such as leaves, stems, and flower parts, roots, or a mixture of these parts, but preferably the leaves and / or stems. It is.

  Ginkgo biloba L. is a ginkgo deciduous tree that grows naturally in Hokkaido, Honshu, Shikoku, Kyushu, etc., and can be easily obtained from these areas. Examples of constituent parts of ginkgo that can be used as an extraction raw material include, for example, leaf parts, branch parts, bark parts, trunk parts, stem parts, flower parts, seed parts, fruit parts, etc., root parts, or a mixture of these parts. The leaf portion is preferred.

  Yacon (Smallanthus sonchifolia) is an Asteraceae plant native to the Andean Plateau of South America and is cultivated in various parts of the world and can be easily obtained from these regions. Examples of the constituent parts of yacon that can be used as the raw material for extraction include above-ground parts such as leaf parts, stem parts, flower parts, underground parts such as tuberous root parts, root parts, or a mixture thereof. It is.

  Bodaiju (Tilia cordata), also known as linden, is a deciduous tree of the linden family that is widely distributed in Europe and is readily available from these regions. Examples of the constituent parts of the body that can be used as the raw material for extraction include the above-ground parts such as leaves, branches, stems, flower parts, buds, fruit parts, and skin parts, or a mixture thereof. Hanabe.

  The extract from the plant can be obtained by drying the extraction raw material, pulverizing it as it is or using a crusher, and subjecting it to extraction with an extraction solvent. Drying may be performed in the sun or using a commonly used dryer. Moreover, after performing pretreatment, such as degreasing, with a nonpolar solvent such as hexane, it may be used as an extraction raw material. By performing pretreatment such as degreasing, extraction treatment with the polar solvent of the plant can be efficiently performed.

  As the extraction solvent, it is preferable to use a polar solvent, and examples thereof include water and hydrophilic organic solvents. These are used alone or in combination of two or more at room temperature or a temperature below the boiling point of the solvent. It is preferable.

  Examples of water that can be used as the extraction solvent include pure water, tap water, well water, mineral spring water, mineral water, hot spring water, spring water, fresh water, and the like, and those subjected to various treatments. Examples of the treatment applied to water include purification, heating, sterilization, filtration, ion exchange, osmotic pressure adjustment, buffering, and the like. Therefore, the water that can be used as the extraction solvent in this embodiment includes purified water, hot water, ion exchange water, physiological saline, phosphate buffer, phosphate buffered saline, and the like.

  Examples of hydrophilic organic solvents that can be used as extraction solvents include lower aliphatic alcohols having 1 to 5 carbon atoms such as methanol, ethanol, propyl alcohol, and isopropyl alcohol; lower aliphatic ketones such as acetone and methyl ethyl ketone; 1,3-butylene. Examples thereof include polyhydric alcohols having 2 to 5 carbon atoms such as glycol, propylene glycol and glycerin.

  When using the liquid mixture of 2 or more types of polar solvents as an extraction solvent, the mixing ratio can be adjusted suitably. For example, when using a mixed solution of water and a lower aliphatic alcohol, it is preferable to mix 1 to 90 parts by volume of a lower aliphatic alcohol with respect to 10 parts by volume of water. When using a mixed solution, it is preferable to mix 1 to 40 parts by volume of a lower aliphatic ketone with 10 parts by volume of water, and when using a mixed solution of water and a polyhydric alcohol, water 10 It is preferable to mix 10 to 90 parts by volume of a polyhydric alcohol with respect to the volume part.

  The extraction treatment is not particularly limited as long as the soluble component contained in the extraction raw material can be eluted in the extraction solvent, and can be performed according to a conventional method. For example, the extraction raw material is immersed in an extraction solvent 5 to 15 times (mass ratio) of the extraction raw material, the soluble components are extracted at room temperature or under reflux, and then filtered to remove the extraction residue. A liquid can be obtained. When the solvent is distilled off from the obtained extract, a paste-like concentrate is obtained, and when this concentrate is further dried, a dried product is obtained.

  In addition, although the extract obtained as mentioned above can be used as an active ingredient of a dipeptidyl peptidase IV inhibitor as it is, a concentrated solution or a dried product is easier to use.

  In addition, since the extract from the plant has a peculiar odor, it can be purified for the purpose of decolorization, deodorization and the like within a range not causing a decrease in its physiological activity. Since it is not used in a large amount when it is blended, etc., there is no practical problem even if it is not purified.

[Production of sesame peptide]
The sesame peptide in this embodiment is obtained by enzymatic treatment of sesame protein obtained by extracting sesame (scientific name: Sesamum indicum) with a protease or the like.

  Sesame (Sesamum indicum) is an annual plant of the sesame family that originates in Africa or India and is cultivated in various parts of the world, and can be easily obtained from these regions. The constituent part of sesame that can be used as an extraction raw material is not particularly limited as long as it is a part that contains abundant proteins, but is preferably a seed part.

  The extraction process is not particularly limited, and can be performed by a known method. For example, after the extraction raw material is dried, it may be pulverized as it is or using a crusher, etc., and used for extraction with an extraction solvent. Moreover, you may use as a raw material for extraction, after giving pretreatments, such as degreasing by oil extraction etc. and degreasing using nonpolar solvents, such as hexane.

  Next, the extraction raw material is put into 5 to 50 times the amount of extraction solvent. As the extraction solvent, it is preferable to use a polar solvent, and it is particularly preferable to use an alkaline solvent such as an aqueous sodium hydroxide solution. Sesame protein can be efficiently dissolved by using an alkaline solvent.

  The obtained sesame protein is dissolved and suspended in a reaction solvent, adjusted to an optimum pH and an optimum temperature as necessary, and subjected to an enzyme treatment with a protease or the like. Thereafter, the enzyme is deactivated by adding acid or alkali, heating, or performing these treatments together. A powdered sesame peptide can be obtained by filtering, concentrating, drying and pulverizing the resulting reaction solution.

[Dipeptidyl peptidase IV inhibitor]
The grapevine extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract and bodaiju extract obtained as described above have an excellent dipeptidyl peptidase IV inhibitory action, so dipeptidyl peptidase IV It can be used as an active ingredient of an inhibitor. The dipeptidyl peptidase IV inhibitor of this embodiment can be used for a wide range of uses such as pharmaceuticals, quasi drugs, and cosmetics.

  In this embodiment, any one of the grape extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract or bodaiju extract may be used as the active ingredient, or these may be used. You may mix and use as said active ingredient. When grape extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract or bodaige extract is used as the above active ingredient, the compounding ratio is as follows: grape extract, sesame peptide, lemon balm extract, ginkgo What is necessary is just to adjust suitably according to the grade of the dipeptidyl peptidase IV inhibitory action, etc. which an extract, a yacon extract, or a Bodaiju extract has.

  The dipeptidyl peptidase IV inhibitor of the present embodiment may be composed only of no grape extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract, bodaige extract or a mixture thereof, A sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract, bodaige extract or a mixture thereof may be formulated.

  The dipeptidyl peptidase IV inhibitor of the present embodiment is in the form of a powder, granule, tablet, liquid, etc. according to a conventional method using a pharmaceutically acceptable carrier such as dextrin or cyclodextrin or any other auxiliary agent. It can be formulated into any dosage form. In this case, as an auxiliary agent, for example, an excipient, a binder, a disintegrant, a lubricant, a stabilizer, a flavoring / flavoring agent, and the like can be used. The dipeptidyl peptidase IV inhibitor can be used by blending it with other compositions (for example, external preparations for skin, cosmetic foods, etc.), and can be used as an ointment, liquid for external use, patch, etc. it can.

  When the dipeptidyl peptidase IV inhibitor of the present embodiment is formulated, the content of the grape extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract, bodaiju extract or a mixture thereof is particularly limited. It can be set as appropriate according to the purpose.

  In addition, the dipeptidyl peptidase IV inhibitor of the present embodiment, if necessary, other natural extracts having a dipeptidyl peptidase IV inhibitory action, grape extract, sesame peptide, lemon balm extract, ginkgo biloba extract, It can be blended with a yacon extract, a bodaige extract or a mixture thereof and used as an active ingredient.

  Examples of the administration method of the dipeptidyl peptidase IV inhibitor of the present embodiment to a patient include transdermal administration, oral administration, and the like. Depending on the type of the disease, a suitable method for its prevention / treatment can be appropriately selected. That's fine.

  In addition, the dose of the dipeptidyl peptidase IV inhibitor of the present embodiment may be appropriately increased or decreased depending on the disease type, severity, patient differences, administration method, administration period, and the like.

  The dipeptidyl peptidase IV inhibitor of this embodiment is a type 2 diabetes, obesity, through the dipeptidyl peptidase IV inhibitory action possessed by the grapevine extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract or bodaiju extract. It can prevent and treat hypertension, insulin resistance and the like, and can also prevent and treat autoimmune diseases such as rheumatoid arthritis and transplant rejection. In addition, the dipeptidyl peptidase IV inhibitor of the present embodiment is capable of pain, neurodegeneration through the dipeptidyl peptidase IV inhibitory action of the grapevine extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract or bodaiju extract. Diseases, and neurological disorders such as neuropsychiatric disorders (eg sciatica, Alzheimer's disease, depression, etc.); growth hormone deficiency and diseases in which growth hormone is used for treatment; cancer (eg T-cell lymphoma, acute lymphoblasts) Leukemia, thyroid cancer, basal cell cancer, breast cancer, etc.); diseases such as HIV infection (AIDS) can be prevented and treated. However, the dipeptidyl peptidase IV inhibitor of the present embodiment can be used for all purposes that are meaningful for exerting a dipeptidyl peptidase IV inhibitory action in addition to these uses.

  Moreover, since the dipeptidyl peptidase IV inhibitor of this embodiment has the outstanding dipeptidyl peptidase IV inhibitory effect, it is suitable for mix | blending with a skin external preparation or food-drinks, for example. In this case, the grape extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract, bodaige extract or a mixture thereof may be blended as they are, or grape extract, sesame peptide, lemon balm extract, A dipeptidyl peptidase IV inhibitor formulated from ginkgo biloba extract, yacon extract, bodaige extract or mixtures thereof may be incorporated.

  Here, the topical skin preparation is not limited in its category, and includes a wide range of skin cosmetics, quasi-drugs, pharmaceuticals, and the like used transdermally. Creams, milky lotions, beauty essences, lotions, packs, foundations, lip balms, bath salts, hair nicks, hair lotions, soaps, body shampoos and the like.

  There is no restriction on the category of food and drink, and it includes a wide range of foods such as general foods, health foods, and functional health foods that are taken orally.

  Further, since the dipeptidyl peptidase IV inhibitor of the present embodiment has an excellent dipeptidyl peptidase IV inhibitory action, it can be suitably used as a reagent for research related to the activity control mechanism of dipeptidyl peptidase IV. it can.

  The dipeptidyl peptidase IV inhibitor of the present embodiment is suitably applied to humans, but as long as the respective effects are exhibited, animals other than humans (for example, mice, rats, hamsters) , Dogs, cats, cows, pigs, monkeys, etc.).

  Hereinafter, although a manufacture example and a test example are shown and this invention is demonstrated concretely, this invention is not restrict | limited to each following example at all. In this test example, lyophilized products of the products shown in Table 1 were used as samples (samples 1 to 6).

Test Example 1 DPPIV Activity Inhibitory Action Test Each of the natural product-derived components (samples 1 to 6) was tested for dipeptidyl peptidase IV (DPPIV) inhibitory action as follows.

  In a 96-well plate, 25 μL of a solution of a test sample (samples 1-6, see Table 2 below for the final concentration) prepared with 25 mM Tris-HCl buffer (pH 8.0) or the same buffer without addition of the sample Then, 0.4 μg / mL DDPIV (rhCD26, manufactured by R & D System) solution prepared in the above buffer solution or 25 μL of the same buffer solution with no enzyme added thereto was mixed and preincubated at 37 ° C. for 5 minutes. Thereafter, 50 μL of a substrate solution (0.5 mM Gly-Pro-p-NA • Tos, manufactured by Peptide Laboratories) prepared in the above buffer was added, and reacted at 37 ° C. for 90 minutes. After completion of the reaction, absorbance at a wavelength of 415 nm was measured. From the obtained results, the DPPIV inhibition rate (%) was calculated by the following formula.

DPPIV inhibition rate (%) = {1− (CD) / (A−B)} × 100
In the formula, A represents “absorbance at a wavelength of 415 nm when no sample is added and an enzyme is added”, B represents “absorbance at a wavelength of 415 nm when no sample is added and an enzyme is not added”, and C is “addition of test sample, enzyme D represents the “absorbance at a wavelength of 415 nm when a test sample was added and no enzyme was added”.
The results are shown in Table 2.

  As shown in Table 2, all of Sample 1 to Sample 6 exhibited a concentration-dependent DPPIV inhibitory action. Therefore, it was revealed that the grapevine extract, sesame peptide, lemon balm extract, ginkgo biloba extract, yacon extract and bodaige extract have excellent DPPIV inhibitory action.

  The dipeptidyl peptidase IV inhibitor of the present invention can greatly contribute to the treatment of diseases such as type 2 diabetes, obesity, hypertension, insulin resistance and the like; autoimmune diseases and transplant rejection.

Claims (1)

A pharmaceutical composition for inhibiting dipeptidyl peptidase IV containing, as an active ingredient, a grapevine extract using one or more selected from the group consisting of stem, vine and leaf of grapevine as an extraction ingredient (rheumatoid arthritis) , Except for the prevention or improvement of diabetes and obesity) .