JP2013241355A - Tie2 activator, angiogenesis inhibitor, blood vessel maturing agent, blood vessel normalizing agent, blood vessel stabilizer and pharmaceutical composition - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a Tie2 activator, an angiogenesis inhibitor, a blood vessel maturing agent, a blood vessel normalizing agent, a blood vessel stabilizer and a pharmaceutical composition which have excellent Tie2 activating action, angiogenesis inhibitory action, blood vessel maturing action, blood vessel normalizing action and blood vessel stabilizing action and high safety.SOLUTION: A Tie2 activator, an angiogenesis inhibitor, a blood vessel maturing agent, a blood vessel normalizing agent, a blood vessel stabilizer and a pharmaceutical composition include an extract of Astragalus complanatus as an active ingredient.

Description

  The present invention relates to a Tie2 activator, an angiogenesis inhibitor, a vascular maturation agent, a vascular normalizer and a vascular stabilizer, and a pharmaceutical composition.

  A blood vessel has a structure in which vascular endothelial cells and vascular wall cells (vascular smooth muscle cells and pericytes) are indirectly or directly bonded via an extracellular matrix, and oxygen and nutrients are absorbed by living bodies. It has a function of supplying waste tissue to remove waste from living tissue.

  In general, the formation of blood vessels includes angiogenesis in which new blood vessels are formed, and angiogenesis in which new blood vessels are formed by the elongation and branching of existing blood vessels. And divided into two stages. In the former, vascular endothelial growth factor (VEGF) acts and is involved in a very wide range of angiogenesis from the initial development of blood vessels called angiogenesis to the subsequent angiogenesis, and the latter is angiopoietin. (Ang) acts and participates in the control of adhesion between vascular endothelial cells and vascular wall cells and the structural stabilization of blood vessels.

  Under normal oxygen conditions, vascular endothelial cells and the vascular wall cells lining them are firmly adhered to each other, and the vascular structure is kept stable. However, when hypoxia occurs in tissues, vascular wall cells May detach from vascular endothelial cells and increase the growth of disordered blood vessels. Such a phenomenon is called angiogenesis and is often observed in diseases mainly consisting of vascular lesions such as tumors, rheumatoid arthritis, diabetic retinopathy, hyperlipidemia, and hypertension.

  These angiogenesis are known to be suppressed by activating the receptor tyrosine kinase Tie2 (Tyrosine kinase with Ig and EGF homology domain 2) expressed in vascular endothelial cells (see, for example, Patent Document 1). ). It has been reported that an ischemic disease caused by suppression of narrowing of a blood vessel or expansion of a blood vessel results in enlargement of a blood vessel cavity due to activation of Tie2 (for example, see Non-Patent Document 1). In addition, it has been reported that Tie2 activation suppresses cell death of vascular endothelial cells (see, for example, Non-Patent Document 2).

Thus, activation of Tie2 is known not only to suppress angiogenesis but also to maturate, normalize, and stabilize blood vessels.
For example, in vascular regenerative medicine, it is known that the activation of Tie2 induces adhesion between vascular endothelial cells and vascular wall cells in a blood vessel to mature the blood vessel.
For example, in diseases where vascular wall cells observed in tumors, diabetic retinopathy, etc. do not adhere to vascular endothelial cells, disordered blood vessels grow, and by activating Tie2, vascular wall cells adhere to endothelial cells, It is known to normalize blood vessels.
For example, with respect to environmental factors that disrupt various intracellular and external vascular structures, activation of Tie2 is known to suppress vascular destabilization and stabilize blood vessels.

  On the other hand, an extract of cinnamon has been proposed as a naturally derived substance that suppresses angiogenesis through activation of Tie2 (see, for example, Patent Document 1). However, this proposal has a problem that the activity is insufficient. Moreover, although suramin is known as a substance which suppresses angiogenesis (for example, refer patent document 2), there exists a problem that it is not excellent in safety.

  Accordingly, there is a strong demand for rapid development of highly safe substances having excellent Tie2 activation action, angiogenesis inhibition action, blood vessel maturation action, blood vessel normalization action, and blood vessel stabilization action. is the current situation.

JP 2009-263358 A JP-T 9-503488

Science. 1999 Dec. 24; 286 (5449): 2511-4. P. N. A. S. 2004 Apr. 13; 101 (15): 5553-8.

An object of the present invention is to solve the conventional problems and achieve the following objects. That is, an object of the present invention is to provide a Tie2 activator having an excellent Tie2 activation action and high safety.
Another object of the present invention is to provide a highly safe angiogenesis inhibitor having an excellent angiogenesis inhibitory action.
In addition, the present invention has an excellent vascular maturation effect, vascular normalization effect or vascular stabilization effect, and is a highly safe vascular maturation agent, vascular normalization agent or vascular stabilization agent. The purpose is to provide.
In addition, the present invention has an excellent Tie2 activation action, angiogenesis inhibition action, vascular maturation action, vascular normalization action, and vascular stabilization action, and is highly safe. The object is to provide a pharmaceutical composition.

  As a result of intensive studies by the inventor in order to solve the above-mentioned problems, the Turrengue extract has an excellent Tie2 activation action, angiogenesis inhibition action, blood vessel maturation action, blood vessel normalization action, and blood vessel stabilization. The present invention has been completed by knowing that it has a crystallization effect.

This invention is based on the said knowledge by this inventor, and as a means for solving the said subject, it is as follows. That is,
<1> A Tie2 activator characterized by containing a vine extract as an active ingredient.
<2> An angiogenesis inhibitor characterized by containing a vine extract as an active ingredient.
<3> A vascular maturation agent, a vascular normalization agent, or a vascular stabilization agent characterized by containing a vine extract as an active ingredient.
<4> The Tie2 activator according to <1> above, the angiogenesis inhibitor according to <2> above, and the vascular maturation agent, blood vessel normalizing agent or blood vessel stability according to <3> above It is a pharmaceutical composition characterized by containing at least one of the agent.

According to the Tie2 activator of the present invention, it is possible to solve the conventional problems and to provide a highly safe Tie2 activator having an excellent Tie2 activation action.
According to the angiogenesis inhibitor of the present invention, the conventional problems can be solved, and an angiogenesis inhibitor having an excellent angiogenesis inhibitory action and high safety can be provided.
According to the blood vessel maturation agent, blood vessel normalization agent, or blood vessel stabilization agent of the present invention, the above-mentioned conventional problems are solved, and excellent blood vessel maturation action, blood vessel normalization action or blood vessel stabilization. It is possible to provide a highly safe vascular maturation agent, vascular normalization agent, or vascular stabilization agent that has an action.
According to the pharmaceutical composition of the present invention, the conventional problems are solved, and at least an excellent Tie2 activation action, angiogenesis inhibition action, blood vessel maturation action, blood vessel normalization action, and blood vessel stabilization action are provided. A pharmaceutical composition having any action and high safety can be provided.

(Tie2 activator, angiogenesis inhibitor, vascular maturation agent, vascular normalization agent, and vascular stabilization agent)
The Tie2 activator, the angiogenesis inhibitor, the vascular maturation agent, the vascular normalization agent, and the vascular stabilization agent of the present invention contain a turrengue extract as an active ingredient, and if necessary, other Contains ingredients.

  The Tie2 activator has a Tie2 activating action of phosphorylating Tie2 to convert it into its active form (phosphorylated Tie2). The activation of Tie2 causes autophosphorylation of the intracellular tyrosine kinase domain and induces adhesion between vascular endothelial cells and vascular wall cells. In an ischemic disease caused by suppression of vascular narrowing or vasodilation, the vascular cavity is enlarged by activation of Tie2. In addition, activation of Tie2 can suppress cell death of vascular endothelial cells.

  The angiogenesis inhibitor has an angiogenesis inhibitory action that suppresses a network of new blood vessels formed from existing blood vessels. In the hypoxic state, the activation of Tie2 is temporarily suppressed, the adhesion between the vascular endothelial cell and the vascular wall cell is dissociated, and a new blood vessel network is formed from the vascular endothelial cell from which the adhesion is dissociated. The angiogenesis inhibitor can suppress the disordered growth of blood vessels caused by the adhesion of such vascular wall cells to endothelial cells.

  The vascular maturation agent induces adhesion between vascular endothelial cells and vascular wall cells, and between vascular endothelial cells such that intravascular environmental factors (cells and humoral factors) do not easily leak out of the blood vessel. It has a maturation effect to form adhesion spots. Further, in vascular regenerative medicine, activation of Tie2 can induce adhesion between vascular endothelial cells and vascular wall cells in blood vessels, thereby allowing blood vessels to mature.

  The blood vessel normalizing agent promotes the lining of vascular wall cells to vascular endothelial cells by enhancing adhesion between vascular endothelial cells, thereby causing the growth of vascular permeability disrupted blood vessels and disordered blood vessels. It has a normalizing action to make abnormal blood vessels normal. In addition, in diseases where vascular wall cells observed in tumors, diabetic retinopathy, etc. do not adhere to vascular endothelial cells, diseased blood vessels grow, and by activating Tie2, vascular wall cells adhere to endothelial cells, It can normalize blood vessels.

  The blood vessel stabilizer suppresses cell death of vascular endothelial cells by suppressing damage to existing blood vessels, dissociation between vascular endothelial cells, and dissociation between vascular endothelial cells and vascular wall cells, Have In addition, with respect to environmental factors that break down various vascular structures inside and outside cells, activation of Tie2 can suppress vascular instability and stabilize blood vessels.

<Village extract>
The Astragalus complanatas R. Br. Is a perennial plant belonging to the genus Leguminosae, and is also referred to as a vine. The vines grow in mountain areas such as Liaoning, Jilin, Hebei, Shaanxi, Gansu, Shanxi, Inner Mongolia, etc. in China and are easily available from these areas. The vines have a plant height of 1 m or more, are covered with short bristles, and have stems that are slightly flat and grow on the ground. The seeds of vines are rounded kidneys, which are also referred to as shaenshi, and are said to have medicinal effects such as supplementing liver and kidney functions.

  The extract of vines can be easily obtained by using a method generally used for plant extraction. It should be noted that the extract of the vine genus includes an extract of vine jellyfish, a diluted or concentrated solution of the extract, a dried product of the extract, or a crudely purified product or a purified product thereof.

  There is no restriction | limiting in particular as an extraction site | part of the said vine rengue, According to the objective, it can select suitably, For example, a leaf part, a flower part, a stem part, a fruit, a seed, a root part, etc. are mentioned. Among these, seeds are preferable.

  As a method for preparing the extraction site of the vine, it can be obtained by drying each site and then pulverizing it as it is or using a crusher and subjecting it to solvent extraction. The drying may be performed in the sun, or may be performed using a commonly used dryer.

  The method for extracting the turrengue is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include a method for extracting using any extraction device at room temperature or under reflux heating. Specifically, in the treatment tank filled with the extraction solvent, the turrengue as an extraction raw material is added, and further, with occasional stirring as necessary, it is allowed to stand for 30 minutes to 2 hours to elute soluble components, followed by filtration. Then, a method of extracting the solid by removing the solid solvent, distilling the extraction solvent from the obtained extract and drying it may be mentioned.

  There is no restriction | limiting in particular as a solvent used for the extraction of the said turrengue, According to the objective, it can select suitably, For example, water, a hydrophilic organic solvent, or these mixed solvents etc. can be used.

  The water that can be used as the extraction solvent for the vines is not particularly limited and can be appropriately selected according to the purpose. For example, pure water, tap water, well water, mineral spring water, mineral water, hot spring water, spring water, In addition to fresh water, etc., those subjected to various treatments are included. Examples of the treatment applied to water include purification, heating, sterilization, filtration, ion exchange, adjustment of osmotic pressure, buffering, and the like. The water that can be used as the extraction solvent includes purified water, hot water, ion exchange water, physiological saline, phosphate buffer, phosphate buffered saline, and the like.

  There is no restriction | limiting in particular as a hydrophilic organic solvent which can be used as an extraction solvent of the said turrengue, According to the objective, it can select suitably, For example, C1-C5, such as methanol, ethanol, propyl alcohol, isopropyl alcohol, etc. Lower aliphatic ketones such as acetone and methyl ethyl ketone; polyhydric alcohols having 2 to 5 carbon atoms such as 1,3-butylene glycol, propylene glycol, and glycerin, and the like. A mixed solvent or the like can also be used. In addition, when using the mixed solvent of the said water and the said hydrophilic organic solvent, in the case of a lower alcohol, 1 mass part-90 mass parts with respect to 10 mass parts of water, and the water 10 in the case of a lower aliphatic ketone. It is preferable to use what mixed 1 mass part-40 mass parts with respect to the mass part. Moreover, in the case of polyhydric alcohol, it is preferable to use what mixed 1 mass part-90 mass parts with respect to 10 mass parts of water.

  There are no particular limitations on the extraction conditions for the vine rengue, and it can be appropriately selected according to the purpose. The extraction solvent amount is preferably 5 to 15 times (mass ratio) the extraction raw material, and the extraction solvent When water is used, extraction is preferably performed at 50 ° C. to 95 ° C. for about 1 hour to 4 hours. When a mixed solvent of water and ethanol is used as an extraction solvent, 40 ° C. to 90 ° C. It is preferable to extract in about 30 minutes to 4 hours.

  There is no restriction | limiting in particular as a purification method of the said turlengue extract, According to the objective, it can select suitably, For example, purification methods, such as liquid-liquid partition extraction, various chromatography, and membrane separation, are mentioned.

  The above-mentioned extract of turrengue has an excellent Tie2 activation action, angiogenesis inhibition action, blood vessel maturation action, blood vessel normalization action, and blood vessel stabilization action. It can be used as an active ingredient of the inventive Tie2 activator, angiogenesis inhibitor, vascular maturation agent, vascular normalization agent, and vascular stabilization agent.

<Other ingredients>
The other components are not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include excipients, moisture-proofing agents, preservatives, reinforcing agents, thickeners, emulsifiers, antioxidants, and sweeteners. , Sour, seasoning, coloring, flavoring, whitening agent, moisturizer, oil component, UV absorber, surfactant, thickener, alcohol, powder component, colorant, aqueous component, water, skin nutrition Agents and the like.
Further, specific examples of the other components are not particularly limited and can be appropriately selected depending on the purpose. For example, hyaluronic acid, hyaluronic acid hydrolyzate, collagen, collagen hydrolyzate, ascorbic acid, ascorbine Acid derivatives, ascorbic acid glycosides, coenzyme Q10, propolis, royal jelly, royal jelly proteolysate, fucoidan, aloe powder, aloe extract, blueberry powder, blueberry extract, isoflavone, noni powder, noni extract, garlic powder, garlic Extract, turmeric powder, turmeric extract, chitosan, glucosamine, chlorella powder, chlorella extract, carnitine, maca powder, maca extract, cassis powder, cassis extract, hanabiratake powder, hanabiratake extract, and other beauty Plant powder Beauty / or extract and the like.

<Application>
The Tie2 activator, angiogenesis inhibitor, blood vessel maturation agent, blood vessel normalizing agent, and blood vessel stabilizer of the present invention have excellent Tie2 activation action, angiogenesis inhibition action, and blood vessel maturation action. Since it has a blood vessel normalizing action and blood vessel stabilizing action, it is a disease mainly composed of vascular lesions such as tumor, rheumatoid arthritis, diabetic retinopathy, hyperlipidemia, hypertension, atopic dermatitis, and pollen. It can be suitably used as a pharmaceutical agent for allergic diseases such as infectious diseases and a safe preventive agent for these diseases.
In addition, it has been confirmed that the Tie2 activator, angiogenesis inhibitor, vascular maturation agent, vascular normalization agent, and vascular stabilizer of the present invention are not digested in the digestive tract. Therefore, it can be widely used as food and drink such as beauty food and drink and health food and drink.

  There is no particular limitation on the content of the Tsurengage extract in the Tie2 activator, angiogenesis inhibitor, vascular maturation agent, vascular normalization agent, and vascular stabilization agent of the present invention, depending on the purpose. The vine extract itself may be used as the Tie2 activator, angiogenesis inhibitor, vascular maturation agent, vascular normalization agent, and vascular stabilization agent of the present invention. However, it is preferably 1 μg or more, preferably 1 μg or more, and preferably 10 μg or more and 1,000 μg or less per mL of Tie2 activator, angiogenesis inhibitor, blood vessel maturation agent, blood vessel normalizing agent, and blood vessel stabilizer. More preferably, it is 80 μg or more and 820 μg or less, more preferably 380 μg or more and 820 μg or less.

  The administration form of the Tie2 activator, vascular maturation agent, vascular normalization agent, vascular stabilization agent, and angiogenesis inhibitor of the present invention is not particularly limited and should be appropriately selected according to the purpose. Examples thereof include oral, parenteral, and external use.

  The dosage forms of the Tie2 activator, angiogenesis inhibitor, blood vessel maturation agent, blood vessel normalizing agent, and blood vessel stabilizer of the present invention are not particularly limited and should be appropriately selected according to the purpose. For example, tablets, powders, capsules, granules, extracts, syrups and other oral administration agents; injections, infusions, suppositories, and other parenteral administration agents; ointments, creams, emulsions, lotions, packs And external preparations such as bath preparations and hair cosmetics.

(Pharmaceutical composition)
The pharmaceutical composition of the present invention contains at least one of the above-described Tie2 activator, angiogenesis inhibitor, vascular maturation agent, vascular normalization agent, and vascular stabilization agent of the present invention, You may contain the additive normally used for a pharmaceutical as needed.

Since the pharmaceutical composition of the present invention has at least one of the excellent Tie2 activation action, angiogenesis inhibition action, blood vessel maturation action, blood vessel normalization action, and blood vessel stabilization action, As a drug for allergic diseases such as rheumatoid arthritis, diabetic retinopathy, hyperlipidemia, hypertension and other vascular lesions, atopic dermatitis, and hay fever, and as a safe preventive drug for these diseases It can be used suitably.
Moreover, since it has been confirmed that the pharmaceutical composition of the present invention is not digested in the digestive tract, it can be widely used as a food or drink such as a beauty food or a health food.

  The content of the vine extract in the pharmaceutical composition of the present invention is not particularly limited and may be appropriately selected depending on the purpose. The vine extract itself may be used as the pharmaceutical composition of the present invention. However, it is preferably 1 μg or more, preferably 10 μg or more and 1,000 μg or less, more preferably 80 μg or more and 820 μg or less, and particularly preferably 380 μg or more and 820 μg or less per 1 mL of the pharmaceutical composition.

  There is no restriction | limiting in particular as an administration form of the pharmaceutical composition of this invention, According to the objective, it can select suitably, For example, oral, parenteral, external use, etc. are mentioned.

  There is no restriction | limiting in particular as a dosage form of the pharmaceutical composition of this invention, According to the objective, it can select suitably, For example, oral administration agents, such as a tablet, a powder agent, a capsule, a granule, an extract, a syrup agent, etc. ; Parenteral administration agents such as injections, instillations, suppositories; and external preparations such as ointments.

  Examples of the present invention will be described below, but the present invention is not limited to these examples.

[Test Example 1: Tie2 activation action (immunoassay) test]
(Example 1: Turrengue extract)
Normal human umbilical vein endothelial cells (HUVEC) cultured to confluence were seeded in a 96-well plate at 2.0 × 10 ⁴ cells / 0.1 mL / well, and a medium for low serum vascular endothelial cell proliferation (Kurashiki Spinning) The culture was carried out overnight using Humedia-EG2). Next, the HUVEC after overnight culture was replaced with 0.1 mL of vascular endothelial cell basal medium (Humdia-EB2 manufactured by Kurashiki Boseki Co., Ltd.) 3 hours before cell stimulation (addition of test sample) and cultured again. Went. Thereafter, 0.1 mL of a turrengue extract (Turengage dry extract F, manufactured by Maruzen Pharmaceutical Co., Ltd.) prepared as a test sample to the concentration shown in Table 1 was added to the well as a test sample, and incubated for 10 minutes. Went. After incubation, the amount of phosphorylated Tie2 and the total amount of Tie2 in the cells were measured according to the protocol using an immunoassay kit (manufactured by R & D Systems, Human Phospho-Tie2 (Y992) Immunoassay). The ratio was calculated.
For dimethyl sulfoxide (DMSO) used as a negative control, the ratio of phosphorylated Tie2 to total Tie2 was calculated in the same manner.
The Tie2 activation rate was calculated using the following formula (1). And phosphorylation action was evaluated based on the obtained activation rate. The results are shown in Table 1.

(Reference Example 1: Positive control)
In Example 1, the vine extract was changed to Angiopoietin-1 (manufactured by R & D system) as a positive control, and tested in the same manner as in Example 9, except that the concentrations shown in Table 1 were used. The results are shown in Table 1.

[Test Example 1: Test results]
The test result of Tie2 activation action (immunoassay) will be described.
When the Tie2 activation action was evaluated using the immunoassay kit, it was confirmed that Tie2 was phosphorylated and activated by the Turrengue extract. In the system to which DMSO as a negative control was added, no significant phosphorylation of Tie2 was observed. Moreover, in the system which added Angiopoietin-1 which is a positive control of Reference Example 1, it was confirmed that Tie2 was phosphorylated and activated. Here, it is known that phosphorylation of Tie2 leads to vascular maturation, vascular normalization, and vascular stabilization, and angiogenesis is suppressed.
From the above, it was suggested that the extracts used in the examples had Tie2 phosphorylation effects, which resulted in vascular maturation, vascular normalization, and vascular stabilization, and angiogenesis was suppressed.

The Tie2 activator, angiogenesis inhibitor, blood vessel maturation agent, blood vessel normalizing agent, blood vessel stabilizer, and pharmaceutical composition of the present invention have excellent Tie2 activation action, angiogenesis inhibition action, Mainly used for vascular lesions such as tumor, rheumatoid arthritis, diabetic retinopathy, hyperlipidemia, and hypertension because it has at least one of vascular maturation, vascular normalization, and vascular stabilization. It can be widely used as a pharmaceutical product relating to the above diseases and a safe preventive agent relating to these diseases.
Further, the Tie2 activator, angiogenesis inhibitor, blood vessel maturation agent, blood vessel normalizing agent, blood vessel stabilizer, and pharmaceutical composition of the present invention are not as digestible in the digestive tract. Therefore, it can be widely used as food and drink such as beauty food and drink and health food and drink.

Claims (4)

  A Tie2 activator characterized by containing a vine extract as an active ingredient.   An angiogenesis inhibitor characterized by containing a vine renage extract as an active ingredient.   A vascular maturation agent, a vascular normalization agent, or a vascular stabilization agent, characterized by containing a turlengue extract as an active ingredient.   The Tie2 activator according to claim 1, the angiogenesis inhibitor according to claim 2, and the vascular maturation agent, vascular normalization agent or vascular stabilization agent according to claim 3. A pharmaceutical composition comprising: