阅读说明：本技术 包含作为有效成分的靛玉红衍生物的药物组合物 (Pharmaceutical composition comprising the derivatives of indirubin as effective component ) 是由 崔康叶 韩均和 黄政涵 崔涩和 吴昌墨 于 2018-04-10 设计创作，主要内容包括：根据本发明,用于促进长骨生长的组合物促进软骨细胞的增殖和成骨细胞的分化,并且具有同时提高长骨长度和厚度(骨密度)的效果,并且因此具有可广泛针对多个年龄组开具处方的优点,从而可用作促进长骨长度和厚度生长的药物组合物。与常规的基于激素的治疗剂不同,根据本发明的靛玉红衍生物通过经口施用具有优异的预防或治疗效果,并且具有优异的有竞争力的价格。另外,与常规治疗剂不同,本发明的用于预防或治疗骨疾病的药物组合物是稳定的化合物,已知其对人体基本上无细胞毒性,并且是尚未确定有副作用的材料,并且其对骨疾病和成骨不全具有优异的治疗效果。(According to the present invention, for promoting the composition of long bone uptake to promote the proliferation of cartilage cell and the differentiation of osteoblast, and have the effect of while improving long bone length and thickness (bone density), and therefore has the advantages that multiple age group prescriptions can be directed to extensively, thus can be used as the pharmaceutical composition for promoting long bone length and grown in thickness.Different from the conventional therapeutic agent based on hormone, derivatives of indirubin according to the present invention has excellent prevention or therapeutic effect by oral administration, and has excellent competitive price.In addition, different from conventional therapy agent, of the invention is stable compound for preventing or treating the pharmaceutical composition of bone disease, it is known that it is to human body substantially no cytotoxicity, it and is the not yet determining material for having side effect, and it has excellent therapeutic effect to bone disease and osteogenesis imperfecta.)

1. it includes by formula 1 for promoting composition any or more in improving selected from bone length growth and bone thickness Or the derivatives of indirubin that formula 2 indicates is as active constituent

Formula 1

[formula 2]

2. it is according to claim 1 for promotes to grow selected from bone length and the raising of bone thickness in it is any or more Composition promotes chondrocyte proliferation, osteoblast differentiation, and activates beta-catenin stable and the nuclear translocation of beta-catenin.

3. it is according to claim 1 for promotes to grow selected from bone length and the raising of bone thickness in it is any or more Composition, wherein the bone length is the length of long bone, and the bone thickness is bone density.

4. for preventing or treating the pharmaceutical composition selected from one of bone uptake obstacle and bone disease or more disease, Comprising the derivatives of indirubin that is indicated by the following Expression 1 or formula 2 as active constituent

Formula 1

[formula 2]

5. according to claim 4 for preventing or treating selected from one of bone uptake obstacle and bone disease or more The composition of disease, wherein the bone uptake obstacle is at least one in of short and small stature, dwarf, cretinism and sex premature Kind, and the bone disease be selected from osteoporosis, malacosteon, osteomyelitis, epithelial proliferation and Bezzet disease at least It is a kind of.

6. for preventing or improving the food compositions selected from least one of bone length growth disorder and bone disease, it includes The derivatives of indirubin indicated by formula 1 or formula 2 is as active constituent

Formula 1

[formula 2]

Technical field

The present invention relates to for promoting, bone length is grown and/or what bone thickness improved is used as activity comprising derivatives of indirubin The composition of ingredient, it is used to prevent or treat bone length growth disorder and/or disease relevant to bone density, the compositions Comprising derivatives of indirubin as active constituent.

Background technique

Bone is to constitute the most hard tissue of people's bone bone and have a large amount of bone matrixs (bone matrix), is present in complete Body, and play key effect.From infancy to manhood, the length growth of the long bone (long bone) of arm and leg is constituted, And determine the height of our bodies.The process is caused by the cartilage cell (chondrocyte) in bone growth plate, and because It is limited open so it is extremely important in adolescence growth for growth plate.In entire puberty, growth plate is gradually closed It closes, and with adult, no longer generation bone uptake, but bone maintains bone thickness or bone density still through lasting metabolic process To support body.In this process, bone is respectively by osteoblast (osteoblast) and osteoclast (osteoclast) shape At and decompose, and the balance between both cells for maintain bone density and prevention fracture it is extremely important.

In this way, bone takes part in the whole development of our bodies, and the form of body is maintained in structure, is protected Internal organs are physiologically maintaining its blood level and provide new blood as the deposit of the minerals such as such as calcium and phosphorus Liquid.Therefore, osteodysplasty or metabolic disorder can lead to a variety of diseases, such as sex premature, nanism, osteoporosis, rickets Disease, malacosteon, osteomyelitis, epithelial proliferation and Bezett disease.It must be it is to be understood that bon e formation process be divided into: 1) thin by cartilage Length caused by born of the same parents improves and 2) improves bone density by balance osteoblast and osteoclast.The two processes are different , and the growth of length determines the height of people, and the raising of bone density is related with many metabolic bone diseases.

Currently, these most of treatments for bone disease are limited to hormonal medicaments.Height growth relevant to bone length promotees It is the growth hormone clinically uniquely used into agent, and the treatment of osteoporosis relevant to bone density uniquely uses Parathyroid hormone.However, Crescormon is not suitable for the normal children of hormone secretion, even more there is such as thyroid gland The adverse reaction of dysfunction and hyperplasia, and it is expensive and not easy to use.In addition, clinically obtainable be used for osteoporosis Therapeutic agent be unique recombined parathyroid hormone drug and the like for obtaining FDA approval, be also limited to serious ridge The patient of column osteoporosis.Further, since worrying the generation of hypercalcinemia, parathyroid hormone has 2 years maximum therapies The limitation of phase, because it has a significant impact to the metabolism of calcium in body.

Therefore, it is necessary to develop new treatment come the shortcomings that overcoming existing bone disorder treatment, and therefore, it is also desirable to develop base In the drug of small molecule compound.Since conventional hormone preparation or synthesis peptide formulations cannot orally be taken, with small molecule Compound is compared, pretty troublesome and expensive by injection application.Due to can be applied to bone related disease and abnormal small molecule Compound oral administration and can have cost competitiveness, therefore it can become treatment promising novel drugs in the market.

Summary of the invention

Technical problem

The present invention has been made to solve the above problems.The object of the present invention is to provide show that excellent bone length is raw Long facilitation and include derivatives of indirubin as active constituent while in vivo almost without the composition of toxicity.

Furthermore it is also an object that provide show excellent bone disease prevention or therapeutic effect and including Derivatives of indirubin is as active constituent simultaneously in vivo almost without toxicity for preventing or treating the medicine group of bone disease Close object.

Technical solution

To achieve the goals above, the present invention provides for promoting the combination that bone length is grown and/or bone thickness improves Object, it includes the derivatives of indirubin indicated by following formula (1) or (2) as active constituent.

Formula 1

[formula 2]

Composition is characterized in that by swashing via the stabilization and nuclear location of beta-catenin in osteoblast (catenin) Living Wnt/ beta-catenin signal transduction promotes the proliferation of cartilage cell, the differentiation of osteoblast.

Bone length is the length of long bone, and bone thickness is bone density.

In order to realize another above-mentioned purpose, the present invention provides for prevent or treat bone length growth disorder and/or The pharmaceutical composition of bone disease, it includes the derivatives of indirubin indicated by following formula (1) or (2) as active constituent.

Formula 1

Formula 2

Bone length growth disorder is selected from any one of nanism and sex premature or more, and bone disease is selected from sclerotin Any one of osteoporosis, rickets, malacosteon, osteomyelitis, epithelial proliferation and Bezzet disease or more.The present invention provides For preventing or improving the pharmaceutical composition of bone length growth disorder and/or bone disease, it includes by following formula (1) or (2) table The derivatives of indirubin shown is as active constituent, to realize another above-mentioned purpose.

Formula 1

[formula 2]

The advantageous effects of invention

Composition according to the present invention for promoting bone length to grow is to the proliferation of cartilage cell and point of osteoblast Change all has facilitation, to improve both bone length and bone thickness (bone density).Therefore, because it has for multiple The advantages of extensive prescription of age group, therefore its pharmaceutical composition that can effectively serve as can promote bone length and bone grown in thickness Object.

Different from the conventional therapeutic agent based on hormone, derivatives of indirubin according to the present invention has by oral administration Excellent prevention or therapeutic effect, and there is competitiveness in price.

In addition, different from conventional therapeutic agent, of the invention is steady for preventing or treating the pharmaceutical composition of bone disease Fixed compound finds that the compound almost without toxicity, and the ill-effect that does not determine and has excellent human body Treatment effect.

Detailed description of the invention

Fig. 1 is handled when in vitro culture and with the derivatives of indirubin in embodiment 1,2 and comparative example 1,8,9,10 to 12 The photo of duration bone uptake.

Fig. 2 is shown in the figure and table of the length variation of the long bone measured in Fig. 1.The derivatives of indirubin of processing is respectively etc. In 0.5 μM, handle 6 days in the medium.

Fig. 3 A is when in vitro culture and with the 6- bromo indigo red-3 ' in comparative example 6-oxime handling duration bone uptake photo The figure (right side) of (left side) and this growth of quantitative display.

Fig. 3 B is to carry out MTT analysis to cartilage cell according to different bromo indigo reds -3 '-oxime concentration to verify with comparing Whether reducing for the long bone length of 6- bromo indigo red-3 ' in example 6-oxime processing has with the influence of the viability to cartilage cell The figure of pass.

Fig. 4 is 5,6- dichloro- indigo red-the 3 '-first oxime shown in embodiment 1, the 5- methoxyl group indigo jade of embodiment 2 The active figure of ALP and table in the cell of derivatives of indirubin processing in red -3 '-oxime, comparative example 1 to 5 and 12.

Fig. 5 is to carry out western engram analysis to the protein extract obtained from MC3T3-E1 cell (osteoblast) As a result, to check the 5- methoxyl group in 5, the 6- dichloro- indigo red-3 ' in embodiment 1-first oxime (A3051) and embodiment 2 Indigo red -3 '-oxime (A3334) effect stable to beta-catenin.

Fig. 6 is to determine beta-catenin with 5, the 6- dichloro- indigo red -3 '-in embodiment 1 by immunocytochemistry Expression in 5- methoxyl group indigo red-3 ' in first oxime (A3051) and embodiment 2-oxime (A3334) processing ATDC5 cell and The photo of positioning.

Preferred embodiment of the present invention

Hereinafter, many aspects and embodiment of the invention will be described in more detail.

One aspect of the present invention is related to the pharmaceutical composition for promoting bone uptake, and it includes what is indicated by formula 1 or formula 2 Derivatives of indirubin is as active constituent.

Formula 1

[formula 2]

Indigo red is a kind of indigo compound, is taken on a red color, chemical structure with it is indigo closely similar.In general, using indigo plant (polygonum tinctorium) and woaded blue (isatis tinctoria) can produce during producing indigo (blue dyes) Raw a small amount of byproduct indigo red.Danggui longhui pills, danggui longhui wan (Danggui Longhui Wan) is prescriptions of traditional Chinese medicine, is had been used for by 11 kinds The drug composition for treating chronic leukemia.Wherein, discovery indigo red be effective drug, and recently, it is known that its be as with In treatment chronic leukemia and neurodegenerative disease such as Alzheimer disease (Bri J.Haemato, 130:..681-690, 2005Nature Cell Biol, 1:.60-67,1999) cell cycle inhibitor the drug with great potential.Indigo is beautiful It is red to be used as precursor that form a variety of derivatives, such as Lndirubin palate derivative, indigo red hydazone derivative, indigo red N- acetyl group spread out Biology and indigo red amine derivative.

In the present invention, as 5, the 6- dichloro- indigo red -3 ' of effective component-first oxime (formula 1) in derivatives of indirubin Make to minimize by applying the side effect that conventional hormone preparation generates with 5- methoxyl group indigo red -3 '-oxime (formula 2), at Osteocyte and cartilage cell are non-toxic, have and improve long bone length to the significant effect of about twice (180%) relative to control group Fruit, and by promoting the differentiation of the proliferation and osteoblast of cell to improve bone density, i.e. bone thickness.

In addition, according to the present invention, in isolated culture system, compared with SB415286 (Sigma aldrich), by formula 1 Or the derivatives of indirubin that formula 2 indicates improves the length of long bone greater than 37 and 3/4 times.

Therefore, different from other drugs, 5- methoxyl group indigo red-the 3 '-oxime according to the present invention indicated by formula (1) can answer For the patient of all ages and classes, because it may have effect to bone uptake, bone thickness or both, and these effects are than other The effect of derivatives of indirubin is more significantly improved greater than 30%.

Specifically, as described in experimental example below, 5,6- dichloro- 3 '-first of indigo red oximes or 5- methoxyl group indigo red- 3 '-oximes promote the stabilization of beta-catenin in the proliferation, osteoblast differentiation and osteoblast of chondroblast.

Specifically, derivatives of indirubin (5,6- dichloro- indigo reds -3 '-first oxime or the 5- of the formula 1 in the present invention or formula 2 Methoxyl group indigo red -3 '-oxime) shin bone (tibia) length in isolated culture system is promoted as new small molecule compound It is the important symbol that bone density is improved by osteoblast differentiation system with ALP activity.In addition, it is determined that thin for cartilage Critically important beta-catenin (key component of the Wnt/ beta-catenin signal transducting system) In of bon e formation in born of the same parents and osteoblast With these drug-treateds later by stabilization, activation and nuclear location.

Therefore, the composition of this research can be used as pharmaceutical composition, show for the bone disease being made up of excellent Different effect: osteoporosis, myopathyRickets, malacosteon, osteomyelitis, epithelial proliferation and Bezzet disease, And bone length growth disorder (such as of short and small stature, dwarf, cretinism and sex premature).

When the indigo that will the derivatives of indirubin that indicated by formula 1 (5,6- dichloro- indigo reds -3 '-first oxime) and be indicated by formula 2 It is considered safe when rubine derivative (5- methoxyl group indigo red -3 '-oxime) is administered orally in mouse for toxicity research Substance, because its 50% lethal dose (LD 50) is at least 1,000mg/kg.

Bone length means the length of long bone, and bone thickness means bone density.

The derivatives of indirubin of general formula (1) or general formula (2) in the present invention is the composition that can be used for a long time, because its is several There is no toxicity and ill-effect.

Relative to the total weight of composition, composition of the invention preferably comprise 0.1 to 50 parts by weight by formula 1 or formula 2 The derivatives of indirubin of expression, but it is not limited to these.

Another aspect of the present invention is related to the derivatives of indirubin comprising being indicated by formula 1 or formula 2 as active constituent For preventing or treating the pharmaceutical composition of bone length growth disorder and bone disease.

Formula 1

[formula 2]

In the description of the derivatives of indirubin of formula (1) or (2), the description being overlapped with foregoing description will be omitted.

Term used in the present invention " prevention or treatment bone disease " includes real by applying composition according to the present invention Existing prevention and bone disease is treated completely or partially.It further includes reducing or improving bone disease symptom, mitigate pain, reduce bone The incidence of disease and improve any variation in the patients for the treatment of results.

Bone length growth disorder can be selected from any one of nanism and sex premature or more.

Bone disease can be selected from osteoporosis disease, rickets, malacosteon, osteomyelitis, epithelial proliferation and Bezzet Any one of disease or more.

Pharmaceutical composition of the invention is the pharmaceutical composition for preventing or treating bone length growth disorder and/or bone disease Object.5,6- dichloro- indigo red-the 3 '-first oxime indicated by formula 1 and 5- methoxyl group indigo red-the 3 '-oxime indicated by formula 2 are by swashing The stabilization of beta-catenin and nuclear location promote the proliferation of cartilage cell and the differentiation of osteoblast, and needle in osteoblast living There is excellent prevention or therapeutic effect to bone length growth disorder or bone disease or both.

In other words, can be applied to the patient at multiple ages because its may to bone length grow or bone thickness improve or The two all has an impact, and has determined that compared with other derivatives of indirubin, these, which are acted in significant numberical range, has Excellent effect.

Pharmaceutical composition of the invention also may include the suitable carrier for being commonly used for preparing pharmaceutical composition, excipient and Diluent.The pharmaceutical dosage form of the derivatives of indirubin of formula 1 or formula 2 of the invention can also be used in the form of its officinal salt.

The derivatives of indirubin of formula (1) or formula (2) of the invention is the composition that can be used for a long time because its almost without Side effect.Relative to the total weight of composition, pharmaceutical composition of the invention preferably comprise 0.1 to 50 parts by weight by The derivatives of indirubin that formula 1 or formula 2 indicate, but it is not limited to these.

Pharmaceutical composition of the invention can be according to conventional methods respectively with oral, external preparation, suppository and sterile note Penetrate solution, for example, powder, granule, tablet, capsule, suspension, emulsion, syrup and aerosol form come using.It can Include carrier, excipient and diluent in the composition of the derivatives of indirubin containing formula 1 may include lactose, dextrose, Sucrose, D-sorbite, mannitol, xylitol, antierythrite, maltose, starch, acacia rubber, alginates, gelatin, phosphorus Sour calcium, calcium silicates, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methyl hydroxybenzoate, hydroxyl Yl benzoic acid propyl ester, talcum powder, magnesium stearate and mineral oil.When prepare when, using diluent or excipient, for example, commonly at Layer agent, incremental agent, binder, wetting agent, disintegrating agent and surfactant.It include tablet, ball for oral administration solid pharmaceutical preparation Agent, powder, granule, capsule etc., and such solid pharmaceutical preparation includes at least one excipient, such as starch, calcium carbonate Or the derivatives of indirubin of above-mentioned formula 1 or formula 2.It is prepared by mixing (calcium carbonate), sucrose or lactose, gelatin etc..In addition to Except simple excipient, lubricant, such as magnesium stearate and talcum powder are also used.Liquid preparation for orally using includes Suspension, solution, emulsion and syrup, and other than usually used simple diluent atoleine, it may also include A variety of excipient, such as wetting agent, sweetener, fragrance and preservative.Formulation for parenteral administration include aseptic aqueous solution, Non-aqueous solvent, suspension, emulsion, lyophilized preparation and suppository.As non-aqueous solvent or suspension, propylene glycol can be used, gather Ethylene glycol, vegetable oil (such as olive oil), injectable esters (such as ethyl oleate) etc..As the matrix of suppository, can be used Witepsol, Macrogol, Tween61, cocoa butter, laurel tallow, glycerin gelatine etc..

The preferred dose of pharmaceutical composition of the invention is according to the situation and weight, the degree of disease, the shape of drug of patient Formula, administration method and duration and change, and can be properly selected by those skilled in the technology concerned.However, being Acquisition desired effect, composition of the invention with daily 0.0001 to 2000mg/kg, preferably with daily 0.001 to 1000mg/kg application.The concentration of the derivatives of indirubin of formula 1 or formula 2 be 1nM to 1M or 1 μM to 1mM or 0.1mM extremely 0.2mM or 0.05 μM to 5 μM, but not limited to this.Drug can be administered once a day or for several times.Above-mentioned dosage is equal in any way It does not limit the scope of the invention.

Pharmaceutical composition of the invention can be applied to mammal, such as rat, mouse, domestic animal, people through a variety of ways. All method of administration are all expectable, such as pass through oral, per rectum or intravenous, intramuscular, subcutaneous, intrauterine, dura mater or brain Indoor (intracerebroventricular) injection.

Another aspect of the present invention is related to preservative or for preventing or improving bone length growth disorder and/or bone disease Improvement food compositions, which includes the derivatives of indirubin that is indicated by formula 1 or formula 2 as active constituent.

Term used in the present invention " improvement " means to reduce parameter level relevant to illness to be treated, such as symptom journey Any movement of degree.

Food compositions of the invention can in many ways, such as treat bone length growth disorder or bone disease or The drug, food and beverage of the two use.The food that the compounds of this invention can be added includes, for example, numerous food, beverage, glue, Tea, vitamin complex, health care replenisher etc., and can be used in the form of powder, particle, tablet, capsule or beverage.

The amount of compound is usually added to food by based on total food weight 0.01 to 15% in food or beverage of the invention In product composition, and it is 0.02 to 5g that healthy beverage composition, which is based on 100ml, and being preferably based on 100ml is 0.3 to 1g.

If beverage is made in food compositions of the invention, it is necessary for shown ratio, and to liquid composition (example Such as common beverages) it is not particularly limited, and may include several corrigents or natural carbohydrate as supplementary element.It is above-mentioned The example of natural carbohydrate is monosaccharide (such as glucose, fructose etc.), disaccharides (such as maltose, sucrose etc.) and polysaccharide (such as dextrin and cyclodextrin and common sugar, such as xylitol, D-sorbite and antierythrite).It is rectified as in addition to above-mentioned Corrigent other than taste agent, it may be advantageous to use natural flavorant (such as taurine, stevia extract, such as the A based on steam And glycyrrhizin, and synthesis corrigent, such as Sa Kalin, Aspartame).The ratio of above-mentioned natural carbohydrate, relatively In 100ml composition of the invention, typically about 1 to 20g, preferably from about 5 to 12g.

In addition to that mentioned above, composition of the invention includes various nutrients, vitamin, minerals (electrolysis property), example Such as the corrigent of synthetic perfume and natural perfume material, colorant and neutralizer (such as cheese, chocolate), pectic acid and its salt, algae Acid and its salt, protective colloid thickener, PH regulator, stabilizer, preservative, glycerol, alcohol, make in soda organic acid Carbonating agent etc..

Composition of the invention also may include pulp to produce fruit juice, fruit drink and vegetable beverage.These components It may be used alone or in combination use.The ratio of such additive is not very crucial, but with respect to combination of the invention 100 parts by weight of object are ordinarily selected to 0 to about 0.1 parts by weight.