CN109730990A - Application of the 15- benzyl subunit -14- deoxidation -11,12- andrographolide in anti-fibrosis medicine - Google Patents

Abstract

The invention belongs to pharmaceutical technology fields, disclose 15- benzyl subunit -14- deoxidation -11,12- andrographolide in preparation and prevent and treat the application in human organ or tissue fibrosis drug.The experiment proved that such compound significantly inhibits people's type Ⅱ penumonocyte A549 mesenchyma conversion of the induction of TGF-β 1；Significantly inhibit people's cortex renis proximal tubular epithelial cells HK-2 mesenchyma conversion of the induction of TGF-β 1；The primary myocardial fibrosis cell HCFB proliferation of the people of inhibition angiotensin II (Ang II) induction.The mouse pulmonary fibrosis degree of bleomycin induction is significantly reduced, kunming mice degree of myocardial fibrosis caused by the Rat renal degree of fibrosis and isoprel ISO that unilateral ostruction induces is significantly reduced.The fibrosis medicines such as anti-kidney, lung, heart are used to prepare using such compound as active ingredient, high-efficiency low-toxicity, to provide new drug approach with the treatment and prevention of fibrillation related disease, to expand the selectable range of clinical application, there is good application and development prospect.

Description

15- benzyl subunit -14- deoxidation -11,12- andrographolide is in anti-fiber Application in chemical drug object

Technical field

Application the present invention relates to andrographolidume derivative as human body organ or tissue fibrosis medicine, specifically relates to And 14- deoxidation -11,12- dehydrogenation andrographolide C15 substitutive derivative, belong to pharmaceutical technology field.

Background technique

Tissue fibrosis is a kind of chronic disease, is mainly in the positions such as liver, lung, heart, kidney.The whole world has 1/3 people dead In tissue fibrosis and resulting organ failure.After tissue is damaged, it is anti-that a series of cells occur for damage location It answers, leads to extracellular matrix over-deposit, tissue fibrosis occurs.It finally can lead to organ dysfunction, or even dead.Painstaking effort Important function has been played in the reconstruct of tubing fibrosis cardiovascular organization caused by hypertension and heart failure, while being also to draw The main reason for playing atherosclerosis.Myocardial fibrosis is characterized in that extracellular matrix protein is accumulated in cytoplasm, and Cause heart contraction and diastolic dysfunction.Myocardial fibrosis is the important symbol of decompensation myocardial hypertrophy and heart failure, Myocardial remodelling caused by participation hypertension, hypertrophic cardiomyopathy, heart failure and myocardial infarction.

Pulmonary fibrosis disease incidence and lethality are on the rise in recent years.Pulmonary fibrosis is many pulmonary disease hairs Exhibition develops, the final final result of scar, and the cause of disease is varied.Many chronic lung diseases, including it is asthma, bronchiectasis, slow Lung, pulmonary tuberculosis, lung cancer, interstitial lung disease etc. are hindered, is all changed with fibrillatable pathological.Its main pathological characteristic includes between lung tissue Mesenchymal cell proliferation, cell extracellular matrix hyperplasia deposition and reconstruct of pulmonary parenchyma etc..It is comprehensive for idiopathic pulmonary fibrosis, respiratory distress For a variety of lung diseases such as simulator sickness, eosinophilic granuloma, lung tissue fibroplasia and degree of fibrosis determine facing for the disease The rear situation of bed.These diseases develop to advanced stage, serious to interfere patient's normal work and quality of life, even result in patient because exhaling Inhale failure or heart failure and death.Between past 20 years, idiopathic pulmonary fibrosis disease incidence totality presentation rises appreciably Trend, mean survival time about 3 years after diagnosis, 5 years survival rates 30%~50% are rear very poor.

Kidney fibrosis shows as extracellular matrix and inappropriate connective tissue to be assembled in kidney, leads to Reno-colic fistula change and function Impaired pathologic process and nearly all kidney trouble proceed to the co-channel of end-stage renal failure.Many acute and chronic kidneys Disease is also all closely related with tissue fibrosis occurrence and development, and especially kidney fibrosis caused by diabetic nephropathy and hypertension changes Become.Panimmunity and autoimmune disease such as arthritis, systemic sclerosis and systemic loupus erythematosus also organized fibrosis Change.

Due to causing, the cause of disease of above-mentioned various histoorgan fibroproliferative diseases is numerous, pathogenesis is complicated, the course of disease is moved Prolong several years to decades etc., the definite pathogenesis of tissue fibrosis not yet illustrates completely at present, also not generally acknowledging, real It is capable of the drug of reversing tissue fibrosis.It is considerably less to the therapeutic agent of fibrotic disease at present, adjuvant treatment is only played mostly Effect.Though glucocorticoid and immunosuppressive drug etc. can improve the symptom of pulmonary fibrosis in terms of pulmonary fibrosis, delay disease Development, but there is strong side effect.The drug of U.S. FDA approval only by InterMune, the development and production of Inc. company The Nintedanib (Nintedanib) of Esbriet (pirfenidone, pirfenidone) and the development and production of Yin Gehan company, Germany, But the current patient of both drugs benefits unsatisfactory.

Studying at present more to tissue, the relevant signal path of organ fibrosis occurrence and development includes TGF-β₁/Smad Signal path, Wnt/ β-catenin signal path, phosphatidylinositol 3-kinase/protein kinase (PI3K/Akt) signal path etc.. For example, obakulactone lowers α-SMA and collagen by the inhibition to TGF-β signal path in mouse fibroid lung tissue Generation (patent No.: 105560255 A of CN).First Peng toad chrysanthemum lactone-sulfuric ester can significantly inhibit the human embryo lung (HEL) of TGF-β induction Fibroblastic proliferation and its hydroxyproline (HYP) content are to play the pharmacological action (patent for improving pulmonary fibrosis degree Number: 103919769 A of CN).The PI3K inhibitor in TGF-β downstream is acted on for treating the pulmonary fibrosis disease (patent No.: CN 104093408 A).There are also the one isoalantolactone analog derivative of sesquialter mushroom utilized in elecampane and its salts for treating pulmonary fibrosis The patent report of (patent No.: 106496243 A of CN).In terms of anti-kidney fibrosis, Ganolactone D can obviously inhibit TGF-β 1 The renal cells Smad3 phosphorylation of induction has the therapeutic effect (patent No.: CN for renal fibrosis 106220643 A).No.3 Hospital Attached to No.3 Military Medical Univ., P.L.A. finds that apiolin mediates Ca by TRPV4²⁺ Interior stream, activate AMPK/SITR1 signal path, inhibit renal fibrosis.

Andrographolide is in the diterpene extracted in Herba Andrographitis Andrographis paniculata (Burm.f.) Nees Ester type compound is one of main effective ingredient of Chinese medicine Herba Andrographitis.Clinically it is mainly used for treating the infection of the upper respiratory tract, bacterium Property dysentery etc..In recent years, andrographolide antitumor, Hepatoprotective cholagogue, in terms of application study deepen continuously.It is yellow It can reduce bleomycin at bright equal discovery andrographolide or bleomycin A5 cause lung fibrosis in rats pulmonary alveolitis and fibrosis journey Degree reduces lung tissue PDGF expression and reduces HYP content；The wise and able equal researchs discovery andrographolide of model can reduce bleomycin cause Lung fibrosis in rats pulmonary alveolitis and pulmonary fibrosis degree reduce lung tissue I, the expression of type III collagen mrna, reduce bronchovesicular TNF-α in irrigating solution (BALF), 1 concentration of TGF-β, and to liver, kidney without obvious toxic-side effects.The researchs such as Sun Jinghui find Herba Andrographitis Interior ester injection mitigates mice with viral myocarditis inflammation and degree of fibrosis, and mechanism may be the table that can inhibit ST2 in cardiac muscle It reaches.The researchs such as Nie Mengqi find that andrographolide has certain anti-kidney region fibrosis effect, mechanism may with it is anti-inflammatory, anti- The expression of oxidation, reduction TGF-β 1 and IGFBP-3 is related.The researchs such as Zhong Xingming demonstrate andrographolide can be by improving sodium potassium ATP enzyme and calcium and magnesium atpase activity reduce hydroxyproline content to inhibit cardiac fibrosis and play resisting cardiac hypertrophy. Researches show that andrographolides to mouse lesions of liver and kidney caused by ethyl alcohol with certain protective effect by Singha P etc..Roy DN Equal researchs have shown that andrographolide is worse in anti-fibrosis and cell than single Beracilline with Beracilline combination therapy copper poisoning Dead aspect better effect.

The present inventor (CN1978437 in early-stage study；CN100999520；CN100999535；CN101003527 it) obtains The andrographolidume derivative of a large amount of structure novels was obtained, and to partial derivatives in antitumor, anti-inflammatory, Anti-HBV activity, HCV and urgency Property liver injury protection effect etc. application applied for patent protection, the present inventor is also to 15- benzyl subunit -14- deoxidation -11,12- dehydrogenation Andrographolidume derivative prepare in anti-hepatic fibrosis medicines application protected (application number: 201710214066.4).Further by being removed to 15- benzyl subunit -14- deoxidation -11,12- andrographolide Activity test research has been carried out in terms of other tissues (organ) fibrosis other than liver, has had no that pertinent literature is reported at present.

Summary of the invention

The present inventor is on the basis of previous research, by carrying out anti-human body tissue or organ to the compound of synthesis Fibrosis screening active ingredients, discovery 15- benzyl subunit -14- deoxidation -11,12- andrographolide have significant prevention With the effect for the treatment of fibrillation related disease, high-efficiency low-toxicity, having exploitation is the potentiality of anti-fibrosis medicine.For this purpose, of the invention It is designed to provide 15- benzyl subunit -14- deoxidation -11,12- andrographolide and is preparing anti-lung, kidney and heart fibre Tie up the application in chemical drug object.

The 15- benzyl subunit -14- deoxidation -11,12- andrographolide structure is as shown in general formula 1.

General formula 1

Wherein: R₁、R₂Respectively one of monosubstituted, the polysubstituted object of hydrogen or phenyl and they；R₁、R₂Respectively ring One of the cyclic substituents such as hexyl, cyclopenta group；R₁、R₂It can be identical substituent group or different substituent groups. R₃、R₄Respectively hydrogen or CH₂CH₂COOHCH₂CH₂CH₂CH₂COOH、CH₂CHCHCH₂COOH、CH₂CH₂CH₂CH₂CH₂CH₂CH₂COOH Deng one of or COR₅, R₅For aromatic heterocycles or hexamethylenes such as substitution or unsubstituted phenyl, pyridyl group, pyrrole radicals, furyls The carbocyclic rings such as base, cyclopenta, cyclopropyl, morpholinyl, piperidyl or heterocycle structure and saturation or unsaturated C1-18 length Carbochain etc..R₃、R₄It can be identical substituent group or different substituent groups simultaneously.

It is preferred that following compound: R₁, R₂Respectively hydrogen or phenyl, 2- methoxyphenyl, 3- methoxyphenyl, 4- methoxyl group Phenyl, 2,3,5- trimethoxyphenyls, 2- hydroxy phenyl, 3- hydroxy phenyl, 4- hydroxy phenyl, 2- fluorophenyl, 2- chlorphenyl, 2- The fluoro- 3- methoxybenzene of bromophenyl, 3- fluorophenyl, 3- chlorphenyl, 3- bromophenyl, 4- fluorophenyl, 4- chlorphenyl, 4- bromophenyl, 2- Base, 3- methoxyl group -4- chlorphenyl, 2,4- difluorophenyl, 2,4- dichlorophenyl, 2,4- dibromo phenyl, the fluoro- 4- chlorphenyl of 2-, 2- The fluoro- 4- chlorphenyl of bromo- 4- chlorphenyl, 3-, the bromo- 4- chlorphenyl of 3-, 3,4- difluorophenyl, 3,4- dichlorophenyl, 3,4 dibromobenzenes The chloro- 4- fluorophenyl of base, 2-, the bromo- 4- fluorophenyl of 2-, the chloro- 4- fluorophenyl of 3-, the bromo- 4- fluorophenyl of 3-, the fluoro- 4- bromophenyl of 2-, 2- The fluoro- 4- bromophenyl of chloro- 4- bromophenyl, 3-, the chloro- 4- bromophenyl of 3-, 2,3,4- trichlorophenyls, 2- methoxyl group -4- chlorphenyl, 2- hydroxyl Base -4- chlorphenyl, 2- hydroxyl -4- methoxyphenyl, 3- amino -4- chlorphenyl, 2- amino -4- chlorphenyl, 2- nitro -4- fluorobenzene Base, 2- nitro -4- chlorphenyl, R₁, R₂Simultaneously it is identical or different but not simultaneously be hydrogen；R₃、R₄Respectively hydrogen；Or R₃、R₄Respectively CH₂CH₂COOH、CH₂CH₂CH₂CH₂COOH、CH₂CHCHCH₂COOH、CH₂CH₂CH₂CH₂CH₂CH₂CH₂One of COOH, or R₃、R₄Respectively COR₅；R₅For 3- pyridyl group or CH₂CH₂COOH、CH₂CH₂CH₂CH₂COOH、CH₂CHCHCH₂COOH、 CH₂CH₂CH₂CH₂CH₂CH₂CH₂One of COOH, R₃、R₄Choosing substituent group identical or different simultaneously.

Preferred compound are as follows: R₁, R₂Respectively hydrogen or phenyl, 2- methoxyphenyl, 3- methoxyphenyl, 4- methoxybenzene Base, 2,3,5- trimethoxyphenyls, 2- hydroxy phenyl, 3- hydroxy phenyl, 4- hydroxy phenyl, 2- fluorophenyl, 2- chlorphenyl, 2- bromine The fluoro- 3- methoxyphenyl of phenyl, 3- fluorophenyl, 3- chlorphenyl, 3- bromophenyl, 4- fluorophenyl, 4- chlorphenyl, 4- bromophenyl, 2-, 3- methoxyl group -4- chlorphenyl, 2,4- difluorophenyl, 2,4- dichlorophenyl, 2,4- dibromo phenyl, the fluoro- 4- chlorphenyl of 2-, 2- are bromo- The fluoro- 4- chlorphenyl of 4- chlorphenyl, 3-, the bromo- 4- chlorphenyl of 3-, 3,4- difluorophenyl, 3,4- dichlorophenyl, 3,4 dibromo phenyls, 2- The bromo- 4- fluorophenyl of chloro- 4- fluorophenyl, 2-, the chloro- 4- fluorophenyl of 3-, the bromo- 4- fluorophenyl of 3-, the fluoro- 4- bromophenyl of 2-, the chloro- 4- bromine of 2- The fluoro- 4- bromophenyl of phenyl, 3-, the chloro- 4- bromophenyl of 3-, 2,3,4- trichlorophenyls, 2- methoxyl group -4- chlorphenyl, 2- hydroxyl -4- chlorine Phenyl, 2- hydroxyl -4- methoxyphenyl, 3- amino -4- chlorphenyl, 2- amino -4- chlorphenyl, 2- nitro -4- fluorophenyl, 2- nitre Base -4- chlorphenyl, but R₁, R₂It is different；R₃、R₄Respectively hydrogen；Or R₃、R₄Respectively CH₂CH₂COOH、CH₂CH₂CH₂CH₂COOH、 CH₂CHCHCH₂COOH、CH₂CH₂CH₂CH₂CH₂CH₂CH₂One of COOH or R₃、R₄Respectively COR₅, R₅For 3- pyridyl group Or CH₂CH₂COOH, R₃、R₄The same substituent group of phase selection.

More preferable compound are as follows: work as R₁, R₂When one of them is hydrogen, R₁, R₂One of them selects following group: phenyl, 2- first Phenyl, 3- methoxyphenyl, 4- methoxyphenyl, 2- fluorophenyl, 2- chlorphenyl, 2- bromophenyl, 3- fluorophenyl, 3- chlorobenzene The fluoro- 3- methoxyphenyl of base, 3- bromophenyl, 4- fluorophenyl, 4- chlorphenyl, 4- bromophenyl, 2-, 3- methoxyl group -4- chlorphenyl, 2, The fluoro- 4- chlorphenyl of 4- difluorophenyl, 2,4- dichlorophenyl, 2,4- dibromo phenyl, 2-, the bromo- 4- chlorphenyl of 2-, the fluoro- 4- chlorobenzene of 3- The bromo- 4- chlorphenyl of base, 3-, 3,4- difluorophenyl, 3,4- dichlorophenyl, 3, the chloro- 4- fluorophenyl of 4 dibromo phenyls, 2-, the bromo- 4- of 2- The chloro- 4- fluorophenyl of fluorophenyl, 3-, the bromo- 4- fluorophenyl of 3-, the fluoro- 4- bromophenyl of 2-, the chloro- 4- bromophenyl of 2-, the fluoro- 4- bromophenyl of 3-, The chloro- 4- bromophenyl of 3-, 2- methoxyl group -4- chlorphenyl；R₃、R₄Respectively hydrogen；Or R₃、R₄Respectively CH₂CH₂COOH、 CH₂CH₂CH₂CH₂COOH、CH₂CHCHCH₂COOH、CH₂CH₂CH₂CH₂CH₂CH₂CH₂One of COOH or R₃、R₄Respectively COR₅, R₅For 3- pyridyl group or CH₂CH₂COOH, R₃、R₄The same substituent group of phase selection.

More preferable compound are as follows:

A:R₁=H, R₂=C₆H₅, R₃=R₄=H；

B:R₁=H, R₂=2-F-C₆H₄, R₃=R₄=H；

C:R₁=H, R₂=2-Cl-C₆H₄, R₃=R₄=H；

D:R₁=H, R₂=2-Br-C₆H₄, R₃=R₄=H；

E:R₁=H, R₂=3-F-C₆H₄, R₃=R₄=H；

F:R₁=H, R₂=3-Cl-C₆H₄, R₃=R₄=H；

G:R₁=H, R₂=3-Br-C₆H₄, R₃=R₄=H；

H:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=H；

I:R₁=H, R₂=4-F-C₆H₄, R₃=R₄=H；

J:R₁=H, R₂=4-Br-C₆H₄, R₃=R₄=H；

K:R₁=H, R₂=4-CH₃O-C₆H₄, R₃=R₄=H；

L:R₁=H, R₂=2-CH₃O-4-Cl-C₆H₃, R₃=R₄=H；

M:R₂=H, R₁=2-Br-C₆H₄, R₃=R₄=H；

N:R₂=H, R₁=3-Cl-C₆H₄, R₃=R₄=H；

O:R₂=H, R₁=2-F-4-Cl-C₆H₃, R₃=R₄=H；

P:R₂=H, R₁=2,4- dichlorophenyl, R₃=R₄=H；

Q:R₂=H, R₁=4-F-C₆H₄, R₃=R₄=H；

R:R₂=H, R₁=C₆H₅, R₃=R₄=H；

S:R₁=H, R₂=3-F-4-Cl-C₆H₃, R₃=R₄=H；

T:R₁=H, R₂=2,4- difluorophenyl, R₃=R₄=H；

U:R₁=H, R₂=3,4- dichlorophenyl, R₃=R₄=H；

V:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=COR₅, R₅=3- pyridyl group；

W:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=CH₂CH₂COOH；

X:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=COR₅, R₅=CH₂CH₂COOH；

Y:R₂=H, R₁=4-Cl-C₆H₄, R₃=R₄=H；

Z:R₂=H, R₁=4-Cl-C₆H₄, R₃=R₄=COR₅, R₅=3- pyridyl group.

Above compound preparation method proposed by the present invention discloses in patent of invention CN:200510107247.4.

The application effect in anti-lung, kidney and cardiac fibrosis drug, present invention utilization are being prepared to study the compound People's type Ⅱ penumonocyte A549 evaluates the inhibition for the A549 cell mesenchyma conversion that the compounds of this invention induces TGF-β 1 Activity evaluates the pulmonary fibrosis resistant activity of compound；The present inventionization is evaluated using people cortex renis proximal tubular epithelial cells HK-2 The inhibitory activity for closing the HK-2 cell mesenchyma conversion that object induces TGF-β 1 evaluates the anti-kidney fibrosis activity of compound；It adopts Stimulate the anti-cardiac muscle of detection cell proliferative conditions evaluation the compounds of this invention after primary popular feeling myofibroblast HCFB fine with Ang II Dimensionization effect.Further, mouse pulmonary fibrosis model, the unilateral ostruction induced rat of bleomycin induction is respectively adopted The research present invention of kunming mice myocardial fibrosis model caused by kidney fibrosis model (UUO) and isoprel ISO represents Property compound anti-lung, kidney and myocardial fibrosis effect, the internal anti-fibrosis effect of evaluation portion representative configurations.

Such compound significantly inhibits people's type Ⅱ penumonocyte A549 mesenchyma conversion of the induction of TGF-β 1；Significant suppression People's cortex renis proximal tubular epithelial cells HK-2 mesenchyma conversion that TGF-β 1 processed induces；Inhibition angiotensin II (Ang II) The primary myocardial fibrosis cell HCFB proliferation of the people of induction.The significant KM mouse lung tissue fibrosis journey for mitigating bleomycin induction Degree, the significant SD Rat renal degree of fibrosis for mitigating unilateral ostruction induction, the significant isoprel ISO that mitigates are caused Kunming mice degree of myocardial fibrosis.

Suitable, the antistructure of such compound all have anti-fibrosis activity, with containing and without the crystallization water for such compound Molecular forms, various crystal forms or the unformed all kinds of prodrug forms for effective medicinal component or such compound, individually or with Other medicines combination is assisted and/or is added with acceptable in pharmacy by current various conventional pharmaceutical methods and technique requirement After the mixing of addition part, the various kinds of drug dosage forms such as the oral type preparation for anti-fibrosis, injection-type preparation are made.It is preferred that being made The standby drug for treating or preventing all kinds of fibrotic diseases such as lung, kidney, the heart.Oral type preparation be tablet, pill, capsule, electuary or Syrup etc.；Injection-type preparation includes injection or freeze-dried powder dosage form etc..

Advantage of the present invention and innovative point: by screening active ingredients, determine that above compound has specific anti-kidney, lung, heart Organ and tissue fibrosis activity.The experiment proved that the compounds of this invention is anti-compared with parent compound andrographolide (AD) The effect of the organ fibrosis such as lung, kidney and heart significantly improves.Therefore, it is used to prepare using such compound as active ingredient anti- Kidney, lung, cardiac fibrosis drug, to provide new drug approach with the treatment and prevention of fibrosis related diseases, to expand The big selectable range of clinical application, has good application and development prospect.

Detailed description of the invention

Fig. 1 is the influence of AD and the compounds of this invention to people's type Ⅱ penumonocyte A549 vigor, and compound concentration is 30.00 μM, in figure: 1.AD；2.A；3.B；4.C；5.D；6.E；7.F；8.G；9.H；10.I；11.J；12.K；13.L；14.M； 15.N；16.O；17.P；18.Q；19.R；20.S；21.T；22.U；23.V；24.W；25.X；26.Y；27.Z；

People's type Ⅱ penumonocyte A549 mesenchyma conversion that Fig. 2 is AD and the compounds of this invention inhibits TGF-β 1 to induce It acting on (statistical result), low, the high concentration of compound AD, N, P, Q, S-W and Z are respectively 0.63 μM and 1.25 μM, compound C, Low, the high concentration of E-G, I-L, O, R, X and Y are respectively 0.31 μM and 0.63 μM, the low of compound A, B, D, H and M, high concentration point It Wei not be 0.16 μM and 0.31 μM；In figure: 1. controls；2.TGF-β1；3.TGF-β1+AD；4.TGF-β1+A；5.TGF-β1+B； 6.TGF-β1+C；7.TGF-β1+D；8.TGF-β1+E；9.TGF-β1+F；10.TGF-β1+G；11.TGF-β1+H；12.TGF-β1 +I；13.TGF-β1+J；14.TGF-β1+K；15.TGF-β1+L；16.TGF-β1+M；17.TGF-β1+N；18.TGF-β1+O； 19.TGF-β1+P；20.TGF-β1+Q；21.TGF-β1+R；22.TGF-β1+S；23.TGF-β1+T；24.TGF-β1+U； 25.TGF-β1+V；26.TGF-β1+W；27.TGF-β1+X；28.TGF-β1+Y；29.TGF-β1+Z；

Fig. 3 is the influence of AD and the compounds of this invention to people's cortex renis proximal tubular epithelial cells HK-2 vigor, compound Concentration is 30.00 μM, in figure: 1.AD；2.A；3.B；4.C；5.D；6.E；7.F；8.G；9.H；10.I；11.J；12.K；13.L； 14.M；15.N；16.O；17.P；18.Q；19.R；20.S；21.T；22.U；23.V；24.W；25.X；26.Y；27.Z；

Fig. 4 is people's cortex renis proximal tubular epithelial cells HK-2 cell that AD and the compounds of this invention inhibit TGF-β 1 to induce Mesenchyma transformation (part displaing micro picture；× 100 times), in figure: 1. controls；2.TGF-β1；3.TGF-β1+AD(0.63μ M)；4.TGF-β1+T(0.63μM)；5.TGF-β1+A(0.63μM)；6.TGF-β1+H(0.08μM)；7.TGF-β1+F(0.63μ M)；8.TGF-β1+J(0.63μM)；9.TGF-β1+K(0.63μM)；10.TGF-β1+Z(0.63μM)；11.TGF-β1+Y(0.16 μM)。

Fig. 5 is AD and representation compound H of the present invention (0.3,3.0 and 15.0 μM) to primary popular feeling myofibroblast HCFB The influence of proliferation function.

Fig. 6 is AD (0.63 μM) and (0.08 μM, 0.16 μM, 0.31 μM and 0.63 μM) of representation compound H of the present invention inhibition The primary popular feeling myofibroblast HCFB proliferation function of angiotensinⅡ (Ang II) induction, and AngII group ratio,^*P < 0.05,^**P < 0.01；With AD group ratio,^#P < 0.05,^##P < 0.01.

Fig. 7 is the influence for the KM mouse lung tissue degree of fibrosis that representative compound H of the present invention induces bleomycin (area of collagen/%), and model group ratio,^*P < 0.05,^**P < 0.01；With AD group ratio,^##P < 0.01；

Fig. 8 is the influence for the KM mouse lung tissue degree of fibrosis that representative compound H of the present invention induces bleomycin (part Masson dyes picture；× 100 times)；

Fig. 9 is the shadow for the SD Rat renal degree of fibrosis that representative compound H of the present invention induces unilateral ostruction It rings (opposite area of collagen/%), and model group ratio,^*P < 0.05,^**P < 0.01；With AD group ratio,^##P < 0.01；

Figure 10 is the shadow for the SD Rat renal degree of fibrosis that representative compound H of the present invention induces unilateral ostruction Ring (part Masson dyeing picture；× 100 times).

Figure 11 is representative compound H of the present invention KM mouse cardiac muscle degree of fibrosis caused by isoprel ISO Influence (area of collagen/%), in figure: 1. normal controls；2. model group；3.AD(15mg/kg)；4.AD(30mg/kg)；5.H (3.75mg/kg)；6.H(7.5mg/kg)；7.H(15mg/kg)；With model group ratio,^*P < 0.05,^**P < 0.01；With AD group Than,^##P < 0.01；

Figure 12 is representative compound H of the present invention KM mouse cardiac muscle degree of fibrosis caused by isoprel ISO Influence (part sirius red stains picture；× 100 times), in figure: 1. normal controls；2. model group；3.AD(15mg/kg)； 4.AD(30mg/kg)；5.H(3.75mg/kg)；6.H(7.5mg/kg).

Specific embodiment

The present invention is illustrated below with reference to specific embodiment.It should be understood that these embodiments be merely to illustrate the present invention and It is not used in and limits the scope of the invention.Compound of the present invention is not limited to representative configurations used in embodiment, can be with The different substituents of replacement 15, obtaining has the active compound of anti-fibrosis；Various the reason of leading to fibrosis can be used Show that the compounds of this invention has anti-fibrosis effect as research object；Also other various in vivo and in vitro moulds be can use Type (method) come obtain the compounds of this invention have anti-fibrosis effect.

1 the compounds of this invention of embodiment inhibits the conversion of people's typeⅡalveolarcells A549 mesenchyma

Stimulation of the typeⅡalveolarcells by cell factors such as inflammatory mediator, growth factors being present in alveolar, Cellular morphology becomes fusiform from cobblestone-appearance, completes epithelium mesenchyma conversion process (EMT), is provided with the function of interstitial cell, And then collagenous fibres are synthesized, a large amount of Collagen fiber depositions can aggravate the course of disease of pulmonary interstitial fibrosis.Therefore, with andrographolide ratio Compared with using people typeⅡalveolarcells A549, using morphological observation method and cell scratch (migration) experimental evaluation present invention The effect of Compound ira vitro pulmonary fibrosis resistant.

1) cell culture

By A549 cell culture containing 10% (V/V) fetal calf serum, 100 μ g/mL streptomysins, 100IU/mL penicillin In RPMI1640 culture solution, volume fraction 5%CO is set₂In saturated humidity, 37 DEG C of cultures in incubator.

2) drug and reagent

By the production of Jin Xin Biotechnology Co., Ltd, ShenFang,SiChuan city, (lot number: 120822), purity is greater than andrographolide 99%；The compounds of this invention laboratory where the present inventor synthesizes, and purity is greater than 99%, similarly hereinafter.

3) mtt assay measures cell toxicant

By after 0.25% (W/V) trypsin digestion of A549 cell for growing logarithmic phase, with containing 10% (V/V) tire ox blood Clear RPMI1640 culture medium is diluted to 5 × 10⁴/ mL cell suspension, is laid in 96 orifice plates, 200 holes μ L/, and in 37 DEG C, volume is divided Number 5%CO₂It is cultivated in incubator for 24 hours, pastille culture medium is added, drug final concentration is up to 30.00 μm of ol/L, each processing 4 Hole repeats, and continues to cultivate 48h, be added MTT (5mg/mL), 20 holes μ L/, cultivate 4h, abandons supernatant, and 150 μ L DMSO, concussion is added L0min measures light absorption value with microplate reader.Measurement wavelength is 570nm, reference wavelength 450nm.It is thin after calculating compound effects Born of the same parents' survival rate, survival rate (%)=medicine group OD_570-450Value/control group OD_570-450Value × 100%, results are averaged, sees attached Fig. 1.

4) influence of the morphological observation method detection drug to A549 cell EMT

By after 0.25% (W/V) trypsin digestion of A549 cell for growing logarithmic phase, with containing 10% (V/V) tire ox blood Clear RPMI1640 culture medium is diluted to 3 × 10⁴/ mL cell suspension, is laid in 96 orifice plates, every 200 μ L of hole.After culture for 24 hours When attached cell is fused to 80%~90%, former culture medium is discarded, serum free medium synchronizing culture for 24 hours, discards culture Base is washed twice with PBS, while the RPMI1640 that 200 μ L contain TGF-β 1 (5ng/mL) and various concentration untested compound is added It takes pictures under the microscope immediately after culture medium (100 ×).If 3 holes repeat and control are arranged.Respectively in microscope after culture 48h Under take pictures.Every kind of compound same concentrations choose 5 visuals field, and measurement is greater than 100 cells.Utilize photoshop CS6 software Picture is handled, and calculates its circularity (formula e=4 π × S/C2, wherein e represents circularity, and S represents area, and C is represented Perimeter).Results are averaged, sees attached drawing 2.

5) experimental result

Attached drawing 1 the result shows that, compared with AD, under 30.00 μM of concentration, cell toxicant of the compounds of this invention to A549 cell Activity has no enhancing.

Attached drawing 2 the result shows that: the compounds of this invention under non-toxic concn, can significantly inhibit A549 cell epithelia mesenchyma turn Change, and with AD ratio, to people's typeⅡalveolarcells mesenchyma conversion inhibiting effect it is stronger, safety index is higher.

People's cortex renis proximal tubular epithelial cells HK-2 mesenchyma that 2 the compounds of this invention of embodiment inhibits TGF-β 1 to induce Transformation

Early stage research discovery renal cells to fibroblast transdifferentiation and can express its marker protein into fibre It ties up specific proteins (FSP1, fibroblast-specific protein 1), renal cells-mesenchymal cell turns Differentiation is one of important pathogenesis of renal interstitial fibrosis.Therefore, close using people's cortex renis compared with andrographolide AD Distal convoluted tubule epithelial cell HK-2 (being provided by China typical culture collection center), morphological observation method after being stimulated using TGF-β 1 With the anti-kidney fibrosis effect in vitro of scratch experiment research the compounds of this invention.

1) cell culture

By HK-2 cell culture containing 10% fetal calf serum (V/V), 100 μ g/mL streptomysins, 100IU/mL penicillin In DMEM-F12 culture solution, 5%CO containing volume fraction is set₂In incubator, in saturated humidity, 37 DEG C of cultures.

2) mtt assay measures cell toxicant

After the HK-2 cell for growing logarithmic phase is digested with 0.25% (W/V) trypsase+0.02%EDTA (W/V), use DMEM-F12 culture medium containing 10% (V/V) fetal calf serum is diluted to 7.0 × 10⁴/ mL cell suspension, is laid in 96 orifice plates, 200 The hole μ L/, in 37 DEG C, volume fraction 5%CO₂It is cultivated in incubator for 24 hours, is changed to pharmaceutical culture medium containing various concentration, drug is most Final concentration of 30.00 μM high, 4 holes of each processing repeat, and continue to cultivate 48h.Other are the same as embodiment 1.Results are averaged, such as attached Shown in Fig. 3.

3) influence of the observation drug to HK-2 cellular morphology after TGF-β 1 stimulates

It will grow to after the HK-2 cell of logarithmic phase digests with 0.25% (W/V) trypsase+0.02%EDTA, with containing The DMEM-F12 culture medium of 10% (V/V) fetal calf serum is diluted to 5.0 × 10⁴/ mL cell suspension, is laid in 96 orifice plates, every hole 200μL.Cell grows up to single layer after culture for 24 hours, discards former culture medium, is cleaned twice with 0.01MPBS, replaces free serum culture Base is to synchronize, after being further cultured for for 24 hours, inhale abandon serum free medium, be added 200 μ L untested compounds containing various concentration with stimulate because The DMEM-F12 culture medium of sub- TGF-β 1 (5ng/mL).If 3 holes repeat and control are arranged.Respectively in microscope after culture 48h Under photograph to record.Morphological change after the compounds of this invention function cells of part is shown in attached drawing 4.

4) experimental result

Attached drawing 3 the result shows that, compared with AD, under 30.00 μM of concentration, suppression of the compounds of this invention to HK-2 cell Proliferation Production significant decrease.

Table 1 and attached drawing 3,4 the result shows that: the compounds of this invention under non-toxic concn, can significantly inhibit TGF-β 1 induction Human colon carcinoma cell line HK-2 is converted to mesenchymal cell, and with AD ratio, the suppression converted to HK-2 cell to mesenchyma With stronger, safety index is higher for production.

1 the compounds of this invention of table inhibits the human colon carcinoma cell line HK-2 of TGF-β induction to turn to mesenchymal cell Change effect

Note: test concentrations are 0.08-1.25 μM；Control: having interaction between epithelial cell, institutional framework is close, Cell is in typical paving stone shape；The processing of TGF-β 1: epithelial cell loses its typicalness, interacts and disappears between cell, Institutional framework relative loose, cell density become smaller, cube in paving stone columnar epithelium cells switch be fusiform fibrocyte shape State；Extremely strong (inhibiting effect): cell is almost no different with compareing, and fusiform is seldom seen under the visual field, and intercellular restores interaction, form Restore its typical paving stone shape；By force (inhibiting effect): inhibiting the invasion of cell, cell consolidation, cell state is almost complete It is complete to restore, rare spindle fibrous cell；In strong (inhibiting effect): cell density becomes larger, and most cells are still in a cube state.

The primary myocardial fibrosis cell of people of 3 the compounds of this invention inhibition angiotensin II (Ang II) of embodiment induction HCFB proliferation function

Some researches show that Cardiac Fibroblasts are the main effects cells of myocardial fibrosis, by II isoreactivity of Ang Quantity proliferation can occur after material incentive, Phenotypic Change is the myofibroblast for having secretion extracellular matrix function.Therefore, it adopts The cyto-inhibition after primary popular feeling myofibroblast HCFB is stimulated to evaluate the compounds of this invention H with mtt assay detection Ang II Function of resisting myocardial fibrillation.

1) cell culture

It (is provided), and is worn by store Bei Na Chuan Lian Biotechnology Co., Ltd using primary popular feeling myofibroblast HCFB Heart lotus lactone compares, and studies the external function of resisting myocardial fibrillation of the compounds of this invention H.HCFB cell culture is being contained into H-DMEM In the culture bottle of culture solution, 10% fetal calf serum of volume fraction (U.S. GIBCO, article No.: 302220F), 100 μ are contained in culture solution G/mL streptomysin, 100IU/mL penicillin, being placed in volume fraction is 5%CO₂In saturation in incubator (German Binder company) Humidity, 37 DEG C of cultures.

2) mtt assay measures cell toxicant

After the HCFB cell for growing logarithmic phase is digested with 0.25% trypsase+0.02%EDTA, with containing volume fraction The H-DMEM culture medium of 10% fetal calf serum is diluted to 5.0 × 10⁴/ mL cell suspension, being laid on 96 orifice plates, (U.S. Costar is public Department) in, 200 holes μ L/, in 37 DEG C, volume fraction 5%CO₂, cultivate in the incubator of saturated humidity for 24 hours, be added and contain various concentration The culture medium of compound AD or H continue to cultivate 48h, be added MTT (5mg/mL), 20 holes μ L/, cultivate 4h, abandon supernatant, is added 150 μ L DMSO shakes l0min, measures light absorption value with microplate reader.Measurement wavelength is 570nm, reference wavelength 450nm.Calculate chemical combination Cell survival rate after object effect, survival rate (%)=medicine group A value/cell controls group A value × 100%, the results are shown in attached figure 5. Data are handled and are analyzed using 17.0 statistics software of SPSS.

3) inhibiting effect of the mtt assay detection drug to the Cardiac Fibroblasts HCFB proliferative capacity stimulated of Ang II

After the HCFB cell of logarithmic growth phase is digested with 0.25% trypsase+0.02%EDTA, with containing volume fraction The H-DMEM culture medium of 10% fetal calf serum is diluted to 5.0 × 10⁴/ mL cell suspension, is laid in 96 orifice plates, every 200 μ L of hole.Training It supports and grows up to single layer to cell for 24 hours, discard former culture medium, cleaned twice with 0.01M PBS, replace serum free medium with synchronization Change, after continuing culture for 24 hours, inhales and abandon serum free medium, 200 μ L untested compounds containing various concentration and stimulating factor Ang is added Ⅱ(10^-7Mol/L H-DMEM culture medium).If 3 holes repeat, using the H-DMEM culture medium containing 0.5%DMSO as negative control, contain Stimulating factor Ang II (10^-7Mol/L) and the H-DMEM culture medium of 0.5%DMSO is positive control.Mtt assay detects after cultivating 48h Cell survival rate.The results are shown in attached figure 6.Data are handled and are analyzed using 17.0 statistics software of SPSS.Data are all made of flat Means standard deviationTo indicate；0.05 group difference of P ﹤ has significant.

4) experimental result

Attached drawing 5 the result shows that, the compounds of this invention H is under 15.00 μM of concentration, Cardiac Fibroblasts HCFB primary to people Cell Proliferation does not show apparent inhibiting effect.

Attached drawing 6 the result shows that: the compounds of this invention H can significantly inhibit AngII and make to the proliferation of HCFB under non-toxic concn With, and with AD ratio, stronger to people's HCFB inhibited proliferation, safety index is higher.

4 the compounds of this invention of embodiment significantly reduces the KM mouse pulmonary fibrosis degree of bleomycin induction

Pulmonary fibrosis is the lung injury as caused by many reasons, and the pathomechanism that pulmonary fibrosis is formed is complicated, different Virulence factor starts inflammation, immune response, and being related to various kinds of cell includes vascular endothelial cell, alveolar epithelial cells, at fiber finer The interaction of born of the same parents and macrophage etc., cytokine profiles and inflammatory mediator.Bleomycin belongs to alkaline glycopeptide class anticancer antibiosis Element, the serious malicious adverse reaction of the medicine first is that causing pulmonary fibrosis, in animal experiments it has proven convenient that lung caused by bleomycin is fine The change of dimensionization Histopathology is closely similar with mankind's pulmonary fibrosis, by the model generally as research pulmonary fibrosis.

1 materials and methods

1) experimental animal

Cleaning grade KM mouse, male, 20 ± 2g of weight are purchased from experimental animal center of henan province.Credit number: SCXK (Henan) 2015-0004。

2) drug, reagent and its preparation

Bleocin HC1 vial produces (lot number: YBH15562005 traditional Chinese medicines standard by the pfizer inc Hai Zheng Word: H20055883)；Prednisone acetate piece produced by Zhejiang Province XianJu Pharmacy stock Co., Ltd (lot number: 170410, traditional Chinese medicines are quasi- Word, 33021207).Other are for reagent object, compound with embodiment 1；Drug is made into 0.5% sodium carboxymethylcellulose (CMC- Na) suspension.The oligomeric sodium carboxymethylcellulose of pharmaceutical grade (CMC-Na) is produced by Anhui Shanhe Medical Accessary Material Co., Ltd. and (is criticized Number: 131114).

3) experimental method

It after KM mouse adaptable fed 3d, is grouped at random according to weight: Sham-operated control group, model group, AD control group (250mg/kg), Bo Nisong control group (5mg/kg), H compound administration group (62.5mg/kg) and H compound administration group (250.mg/kg), totally 6 groups, every group 15.Be injected intraperitoneally mass percent 4% yellow Jackets (2ml/kg) to mouse into Row anesthesia, the fixed mouse of dorsal position shave off neck hair, after iodine disinfection skin, along neck to downlink 1cm or so notch, trip From bronchus, bleomycin (2mg/mL) 50 μ L is injected, injects 150 μ L air immediately, quick rotation mouse divides medical fluid uniformly Cloth, then with No. 4/0 silk suture notch, after iodophor disinfection, sterile gauze is bound up a wound, and finally send postoperative mouse to warm Place is until revival, wherein the physiological saline of sham-operation group injection equal volume.After modeling terminates for 24 hours, starts right place and determine people Gastric infusion, wherein sham-operation group gives the CMC-Na that equal volume ratio refers to, 1 time a day, terminates experiment after 28d is administered. Mouse 12h before last stomach-filling replaces padding, is strictly deprived of food but not water.It is complete using eyeball method collection mouse is plucked after 0.5h after administration Blood, cervical dislocation puts to death mouse after having acquired blood, acquires lungs, and weighing observes and records pulmonary lesion.By lung after taking pictures It is fixed in 4% paraformaldehyde fixer.Fixed sufficiently rear progress pathological section, contaminates observation hepatic fibrosis-renal tubular ectasia syndrome through tri- color of Masson Degree, Image-Pro Plus software carry out fibrosed tissue semi-quantitative analysis to the result of taking pictures of Masson stained slice. The opposite area of collagen of calculating: (administration group average area-normally organize average area)/(normally group is average for model group average area- Area) × 100%, the results are shown in attached figure 7,8.

2 experimental results

Attached drawing 7 and 8 the result shows that: bleomycin induction mouse pulmonary fibrosis model on, the compounds of this invention H is significant Pulmonary fibrosis KM mouse lung tissue fibrosis area is reduced, and effect is significantly stronger than AD.

5 the compounds of this invention of embodiment significantly reduces the SD Rat renal degree of fibrosis of unilateral ostruction induction

Unilateral ostruction induced rat kidney fibrosis model (UUO) is one of research kidney fibrosis classical model, should The aspect of model is cell component accumulation in renal tubular interstitium, fibroblast differentiation/proliferation, ECM deposition increases and renal tubule withers Contracting etc., it is similar to the occurrence and development process of clinical renal disease, extremely facilitate various tubulointerstitial injury disease prevention and cure Research, and modeling method is easy, at mould rate 100%, lesion is uniform, there is preferable repeatability, can cause fiber in a short time Change, is a kind of more quick, reliable animal model for studying the occurrence regularity and Mechanism Discussion of kidney region fibrosis. Therefore, UUO model is widely used in kidney region fibrosis Mechanism Study and improves the evaluation of the therapeutic effect of kidney region fibrosis Deng.

1 materials and methods

1) experimental animal

Cleaning grade SD rat, male, 200 ± 20g of weight are purchased from experimental animal center of henan province.Credit number: SCXK (Henan) 2015-0004.

2) drug, reagent and its preparation

Andrographolide, the source of sodium carboxymethylcellulose and this experimental compound and preparation are the same as embodiment 1 or 4.

3) experimental method

It after SD rat adaptable fed 3d, is grouped at random according to weight: Sham-operated control group, model group, AD control group (0.15mmol/kg), H compound administration group (0.06mmol/kg), H compound administration group (0.10mmol/kg), H compound are given Totally 6 groups of medicine group (0.15mmol/kg), every group 4.Preoperative Method and anesthesia are with embodiment 4, and after anesthesia, left lateral position is fixed big Mouse shaves off breastbone lower edge to hind leg and sees hair, in paving operation cloth and the skin for shaving off hair, after iodine disinfection skin, along breastbone To downlink 2cm or so notch at lower edge about 0.2cm, kidney, Ureter dissection, apart from about 1/3 urine output length of tube of bladder are squeezed out Place abdomen after kidney sends abdominal cavity back to, is closed with No. 4/0 silk thread holostrome, Iodophor disappears with No. 4/0 dual ligation of silk thread and detachment ureter After poison, sterile gauze is bound up a wound, and is finally sent postoperative rat to warm place until revival, wherein sham-operation group is only dissociated defeated Urinary catheter does not ligature, not from section.After modeling terminates for 24 hours, starts right place and determine people's gastric infusion, medication is given with embodiment 4 Terminate experiment after medicine 14d.Rat 12h before last stomach-filling replaces padding, is strictly deprived of food but not water.0.5h is injected intraperitoneally after administration Yellow Jackets (2mL/kg) anesthetic of mass percent 3% completely cuts open rapidly from left kidney after blood sampling, weighs kidney Weight measures kidney size, is fixed in 4% paraformaldehyde fixer after taking pictures.Serum preparation and Masson colouring method and statistics Processing is the same as embodiment 2.Masson coloration result and opposite area of collagen statistical framework such as attached drawing 9 and 10.

2 experimental results

In conjunction with attached drawing 9 and 10, the results showed that on UUO model, representative compound H of the present invention significantly reduces kidney fiber Change degree improves lesion nephridial tissue structure, and effect is significantly stronger than AD.

6 the compounds of this invention of embodiment significantly mitigates KM mouse cardiac muscle fibrosis journey caused by isoprel (ISO) Degree

1 materials and methods

Experimental animal: SPF grades of kunming mices, male, 23 scholar 2g of weight are purchased from experimental animal center of henan province.It is permitted Card number is SCXK (Henan) 2017-0001

Isoprel ISO:sigma: product；Test compound is made into 0.5% sodium carboxymethylcellulose (CMC- Na) suspension.Andrographolide, the source of sodium carboxymethylcellulose and this experimental compound and preparation are the same as embodiment 1 or 4.

Test method: after KM mouse adaptable fed 3d, be grouped at random according to weight: saline control group, model group, 15mg/kg the and 30mg/kg dosage group of AD, the 3.75mg/kg of the compounds of this invention H, 7.5mg/kg, 15mg/kg dosage group, often Group 9.Model group and remaining each administration group are with 20mg/kg subcutaneous injection ISO, and continuous 7 days, equivalent was subcutaneously injected in physiological saline group Physiological saline.After drug 2h is subcutaneously injected, 0.5%CMC-Na is given in physiological saline group and model group stomach-filling, remaining each administration group The 0.5%CMC-Na relative medicine of suspension is given, administration 2w terminates.Mouse 12h before last stomach-filling replaces padding, stringent fasting It can't help water.1h after administration, eyeball blood sampling are completely cutd open rapidly later and are centrifuged dirty and take pictures.After centrifugal blood, supernatant is taken, is saved It is spare.It takes mouse heart tissue fixed in 10 times of volume 4% (w/v) paraformaldehyde fixers, updates fixer afterwards for 24 hours.Gu Fixed sufficiently rear progress pathological section, observes cardiac fibrosis degree through sirius red stains, Image-Pro Plus software is to day The result of taking pictures of wolf star stained slice carries out fibrosed tissue semi-quantitative analysis.Opposite area of collagen is calculated, as a result sees attached drawing 11 and Figure 12.Data are handled and are analyzed using 17. statistics software of SPSS.Data are all made of average value scholar's standard deviation (X Scholar S) it indicates: the group difference of P < 0.05 has significant.

2 experimental results

In conjunction with Figure 11, Figure 12, the results showed that caused on myocardial fibrosis model in isoprel, the compounds of this invention Effect of anti hepatic fibrosis is significantly stronger than AD drug-treated group.

Claims (6)

1. structure 15- benzyl subunit -14- deoxidation -11,12- andrographolide as shown in general formula 1 is preparing drug In application, which is characterized in that be used to prepare anti-kidney, lung or myocardial fibrosis drug as active ingredient；

Wherein: R₁, R₂Respectively hydrogen or phenyl, 2- methoxyphenyl, 3- methoxyphenyl, 4- methoxyphenyl, 2,3,5- front threes Phenyl, 2- hydroxy phenyl, 3- hydroxy phenyl, 4- hydroxy phenyl, 2- fluorophenyl, 2- chlorphenyl, 2- bromophenyl, 3- fluorobenzene The fluoro- 3- methoxyphenyl of base, 3- chlorphenyl, 3- bromophenyl, 4- fluorophenyl, 4- chlorphenyl, 4- bromophenyl, 2-, 3- methoxyl group -4- The fluoro- 4- chlorphenyl of chlorphenyl, 2,4- difluorophenyl, 2,4- dichlorophenyl, 2,4- dibromo phenyl, 2-, the bromo- 4- chlorphenyl of 2-, 3- The bromo- 4- chlorphenyl of fluoro- 4- chlorphenyl, 3-, 3,4- difluorophenyl, 3,4- dichlorophenyl, 3, the chloro- 4- fluorobenzene of 4 dibromo phenyls, 2- The bromo- 4- fluorophenyl of base, 2-, the chloro- 4- fluorophenyl of 3-, the bromo- 4- fluorophenyl of 3-, the fluoro- 4- bromophenyl of 2-, the chloro- 4- bromophenyl of 2-, 3- The chloro- 4- bromophenyl of fluoro- 4- bromophenyl, 3-, 2,3,4- trichlorophenyls, 2- methoxyl group -4- chlorphenyl, 2- hydroxyl -4- chlorphenyl, 2- Hydroxyl -4- methoxyphenyl, 3- amino -4- chlorphenyl, 2- amino -4- chlorphenyl, 2- nitro -4- fluorophenyl, 2- nitro -4- chlorine Phenyl, R₁, R₂Simultaneously it is identical or different but not simultaneously be hydrogen；R₃、R₄Respectively hydrogen；Or R₃、R₄Respectively CH₂CH₂COOH or CH₂CH₂CH₂CH₂COOH、CH₂CHCHCH₂COOH、CH₂CH₂CH₂CH₂CH₂CH₂CH₂One of COOH or R₃、R₄Respectively COR₅, R₅For substitution or unsubstituted phenyl, pyridyl group, pyrrole radicals, furyl or cyclohexyl, cyclopenta, cyclopropyl, morpholine The carbochain of base, piperidyl or saturation or unsaturated C1-18 length；R₃、R₄Select identical or different substituent group.

2. 15- benzyl subunit -14- deoxidation -11,12- andrographolide as described in claim 1 is preparing drug In application, which is characterized in that R₁, R₂Respectively hydrogen or phenyl, 2- methoxyphenyl, 3- methoxyphenyl, 4- methoxybenzene Base, 2,3,5- trimethoxyphenyls, 2- hydroxy phenyl, 3- hydroxy phenyl, 4- hydroxy phenyl, 2- fluorophenyl, 2- chlorphenyl, 2- bromine The fluoro- 3- methoxyphenyl of phenyl, 3- fluorophenyl, 3- chlorphenyl, 3- bromophenyl, 4- fluorophenyl, 4- chlorphenyl, 4- bromophenyl, 2-, 3- methoxyl group -4- chlorphenyl, 2,4- difluorophenyl, 2,4- dichlorophenyl, 2,4- dibromo phenyl, the fluoro- 4- chlorphenyl of 2-, 2- are bromo- The fluoro- 4- chlorphenyl of 4- chlorphenyl, 3-, the bromo- 4- chlorphenyl of 3-, 3,4- difluorophenyl, 3,4- dichlorophenyl, 3,4 dibromo phenyls, 2- The bromo- 4- fluorophenyl of chloro- 4- fluorophenyl, 2-, the chloro- 4- fluorophenyl of 3-, the bromo- 4- fluorophenyl of 3-, the fluoro- 4- bromophenyl of 2-, the chloro- 4- bromine of 2- The fluoro- 4- bromophenyl of phenyl, 3-, the chloro- 4- bromophenyl of 3-, 2,3,4- trichlorophenyls, 2- methoxyl group -4- chlorphenyl, 2- hydroxyl -4- chlorine Phenyl, 2- hydroxyl -4- methoxyphenyl, 3- amino -4- chlorphenyl, 2- amino -4- chlorphenyl, 2- nitro -4- fluorophenyl, 2- nitre Base -4- chlorphenyl, but R₁, R₂It is different；R₃、R₄Respectively hydrogen；Or R₃、R₄Respectively CH₂CH₂COOH、CH₂CH₂CH₂CH₂COOH、 CH₂CHCHCH₂COOH、CH₂CH₂CH₂CH₂CH₂CH₂CH₂One of COOH or R₃、R₄Respectively COR₅, R₅For 3- pyridyl group Or CH₂CH₂COOH, R₃、R₄The same substituent group of phase selection.

3. 15- benzyl subunit -14- deoxidation -11,12- andrographolide as described in claim 1 is preparing drug In application, which is characterized in that work as R₁, R₂When one of them is hydrogen, R₁, R₂One of them selects following group: phenyl, 2- methoxy Base phenyl, 3- methoxyphenyl, 4- methoxyphenyl, 2- fluorophenyl, 2- chlorphenyl, 2- bromophenyl, 3- fluorophenyl, 3- chlorobenzene The fluoro- 3- methoxyphenyl of base, 3- bromophenyl, 4- fluorophenyl, 4- chlorphenyl, 4- bromophenyl, 2-, 3- methoxyl group -4- chlorphenyl, 2, The fluoro- 4- chlorphenyl of 4- difluorophenyl, 2,4- dichlorophenyl, 2,4- dibromo phenyl, 2-, the bromo- 4- chlorphenyl of 2-, the fluoro- 4- chlorobenzene of 3- The bromo- 4- chlorphenyl of base, 3-, 3,4- difluorophenyl, 3,4- dichlorophenyl, 3, the chloro- 4- fluorophenyl of 4 dibromo phenyls, 2-, the bromo- 4- of 2- The chloro- 4- fluorophenyl of fluorophenyl, 3-, the bromo- 4- fluorophenyl of 3-, the fluoro- 4- bromophenyl of 2-, the chloro- 4- bromophenyl of 2-, the fluoro- 4- bromophenyl of 3-, The chloro- 4- bromophenyl of 3-, 2- methoxyl group -4- chlorphenyl；R₃、R₄Respectively hydrogen；Or R₃、R₄Respectively CH₂CH₂COOH、 CH₂CH₂CH₂CH₂COOH、CH₂CHCHCH₂COOH、CH₂CH₂CH₂CH₂CH₂CH₂CH₂One of COOH or R₃、R₄Respectively COR₅, R₅For 3- pyridyl group or CH₂CH₂COOH, R₃、R₄The same substituent group of phase selection.

4. 15- benzyl subunit -14- deoxidation -11,12- andrographolide as described in claim 1 is preparing drug In application, which is characterized in that preferred following compound:

A:R₁=H, R₂=C₆H₅, R₃=R₄=H；

B:R₁=H, R₂=2-F-C₆H₄, R₃=R₄=H；

C:R₁=H, R₂=2-Cl-C₆H₄, R₃=R₄=H；

D:R₁=H, R₂=2-Br-C₆H₄, R₃=R₄=H；

E:R₁=H, R₂=3-F-C₆H₄, R₃=R₄=H；

F:R₁=H, R₂=3-Cl-C₆H₄, R₃=R₄=H；

G:R₁=H, R₂=3-Br-C₆H₄, R₃=R₄=H；

H:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=H；

I:R₁=H, R₂=4-F-C₆H₄, R₃=R₄=H；

J:R₁=H, R₂=4-Br-C₆H₄, R₃=R₄=H；

K:R₁=H, R₂=4-CH₃O-C₆H₄, R₃=R₄=H；

L:R₁=H, R₂=2-CH₃O-4-Cl-C₆H₃, R₃=R₄=H；

M:R₂=H, R₁=2-Br-C₆H₄, R₃=R₄=H；

N:R₂=H, R₁=3-Cl-C₆H₄, R₃=R₄=H；

O:R₂=H, R₁=2-F-4-Cl-C₆H₃, R₃=R₄=H；

P:R₂=H, R₁=2,4- dichlorophenyl, R₃=R₄=H；

Q:R₂=H, R₁=4-F-C₆H₄, R₃=R₄=H；

R:R₂=H, R₁=C₆H₅, R₃=R₄=H；

S:R₁=H, R₂=3-F-4-Cl-C₆H₃, R₃=R₄=H；

T:R₁=H, R₂=2,4- difluorophenyl, R₃=R₄=H；

U:R₁=H, R₂=3,4- dichlorophenyl, R₃=R₄=H；

V:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=COR₅, R₅=3- pyridyl group；

W:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=CH₂CH₂COOH；

X:R₁=H, R₂=4-Cl-C₆H₄, R₃=R₄=COR₅, R₅=CH₂CH₂COOH；

Y:R₂=H, R₁=4-Cl-C₆H₄, R₃=R₄=H；

Z:R₂=H, R₁=4-Cl-C₆H₄, R₃=R₄=COR₅, R₅=3- pyridyl group.

5. the 15- benzyl subunit -14- deoxidation -11,12- andrographolide as described in one of claim 1-4 Application in medicine preparation, pharmaceutical methods and technique requirement routinely, is made oral type preparation or injection-type preparation.

6. 15- benzyl subunit -14- deoxidation -11,12- andrographolide as claimed in claim 5 is preparing drug In application, which is characterized in that oral type preparation be tablet, pill, capsule, electuary or syrup；Injection-type preparation is injection Or freeze-dried powder dosage form.