WO2019052477A1 - Tanshinone compound and use thereof for treating hemangioma - Google Patents

Abstract

Provided is the use of a class of tanshinone compounds for treating hemangioma; in particular, provided is the use of a class of compounds as shown in formula I below or a pharmaceutically acceptable salt thereof. The tanshinone compounds have a hemangioma cell inhibitory activity and can be used for preparing a pharmaceutical composition for treating hemangioma.

Description

Tanshinones and their use for treating hemangiomas Technical field

The present invention relates to the field of medicine, and in particular, the present invention provides a class of tanshinone compounds and their use for treating hemangiomas.

Background technique

Aneurysm hemangiomas (IH) is the most common benign tumor in infants and young children. It is a true hemangioma caused by abnormal proliferation of vascular endothelial cells. Its incidence rate is about 5%-10%, and the ratio of male to female is 1:3-5. The disease is difficult to be discovered at birth. A recent cohort study suggests that the peak growth period of infantile hemangiomas is mostly 5.5-7.5 weeks, with an average of about 4 weeks. Then the lesions gradually become visible and enter a period of rapid growth, which is slow. , from a few months to as many as seven to ten years. A small number of children have a large area of hemangioma, leaving pigmentation, accompanied by signs of shallow scars, skin atrophy and sagging, and even lifelong unhealed.

The pathogenesis of IH has been blurred to this day. Early theory suggests that mother's behavior during pregnancy leads to the occurrence of hemangioma or vascular malformation, and strawberry hemangioma is even thought to be caused by the mother taking red fruit. At present, the mechanism of hemangioma mainly includes the following theories: abnormal vascular development during embryogenesis, angiogenic diseases, and effects of estrogen. Recent research on hemangiomas has focused on the recruitment of progenitor cells, interstitial tumor cells, abnormalities in extracellular matrices, and factors that promote angiogenesis.

More than half of IH can resolve on its own. It is generally advocated to hold a wait-and-see attitude and palliative attitude in the early stage of hemangioma. However, the occurrence and development of hemangioma may affect the life of the child. As the age increases, the self-awareness increases, and the child often shows a deficiency. Self-confidence, anxiety and guilt, especially when the growth of hemangioma endangers the life or function of the child, should be treated promptly. Current treatments include oral or topical medications, surgical resection, cryotherapy, radionuclide therapy, and laser therapy. Hormone and propranolol are currently the primary choice for oral treatment of IH, and propranolol has become the first line of treatment for IH. The mechanism of action of propranolol and hormone on IH is still unclear. At the same time, the effect is significant, and it also faces many side effects. Such as propranolol can induce bradycardia, varying degrees of atrioventricular block, hypotension, cardiogenic shock, bronchospasm, asthma and pulmonary edema; hormones can cause fungal infections, hypertension, Cushing syndrome And gastrointestinal discomfort and so on. Other topical drugs for hemangioma are still in trial use, such as imiquimod, which induces the regression of hemangioma by producing a large number of cytokines, but the actual effect needs to be observed and summarized. Therefore, the task of developing new anti-angioma drugs and developing more effective treatments is imminent.

Summary of the invention

It is an object of the present invention to provide a novel pharmaceutical compound having an activity of inhibiting hemangioblast production and its use for treating hemangiomas.

In a first aspect of the invention, there is provided the use of a compound of formula I, or a pharmaceutically acceptable salt thereof, as defined below:

Wherein the A ring is selected from the group consisting of a substituted or unsubstituted 3-10 membered carbocyclic ring, a substituted or unsubstituted 3-10 membered heterocyclic ring, a substituted or unsubstituted C ₆ -C ₁₀ aromatic ring, substituted or unsubstituted 5-12 yuan heteroaryl ring;

R _{1 is} selected from the group consisting of H, oxygen atom (=O), O-R';

R _{2 is} selected from the group consisting of H, oxygen atom, O-R';

The R' is selected from the group consisting of H, a substituted or unsubstituted C ₂ -C ₁₀ acyl group, a fluorenylmethoxycarbonyl group (Fmoc), or a substituted or unsubstituted C ₁ -C ₁₀ alkyl group; Ground, R' is H;

R _{3 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ alkyl group, C ₁ -C ₄ haloalkyl group, substitution Or an unsubstituted C ₂ -C ₁₀ acyl group, a substituted or unsubstituted C ₂ -C ₁₀ ester group, -OC(O)-R, -NH-R; wherein R is selected from the group consisting of H, C ₁ -C ₄ acyl group, a substituted or unsubstituted C ₁ -C ₄ alkyl group;

R _{4 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ alkyl group, C ₁ -C ₄ haloalkyl group, substitution Or an unsubstituted C ₂ -C ₁₀ acyl group, a substituted or unsubstituted C ₂ -C ₁₀ ester group, -OC(O)-R, -NH-R; wherein R is selected from the group consisting of H, C ₁ -C ₄ acyl group, a substituted or unsubstituted C ₁ -C ₄ alkyl group;

Or any two of R ₁ , R ₂ , R ₃ and R ₄ together with an adjacent carbon atom constitute a group selected from the group consisting of a substituted or unsubstituted C ₃ -C ₁₀ membered carbocyclic ring, substituted or unsubstituted a substituted 3-10 membered heterocyclic ring, a substituted or unsubstituted C ₆ -C ₁₀ aromatic ring, a substituted or unsubstituted 5-12 membered heteroaryl ring;

R _{5 is} selected from the group consisting of H, halogen, cyano, hydroxy, C ₁ -C ₄ alkoxy, -COOR, substituted or unsubstituted C ₁ -C ₁₀ alkyl, substituted or unsubstituted C ₁ -C _{a 10} haloalkyl group, a substituted or unsubstituted C ₂ -C ₁₀ acyl group, a substituted or unsubstituted C ₂ -C ₁₀ ester group; wherein R is selected from the group consisting of H, C ₂ -C ₄ acyl groups, substitutions Or an unsubstituted C ₁ -C ₄ alkyl group;

R _{6 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, carboxyl group, hydroxyl group, C ₁ -C ₄ alkoxy group, -COOR, substituted or unsubstituted C ₁ -C ₁₀ alkyl group, substituted or unsubstituted C ₁ -C ₁₀ haloalkyl, substituted or unsubstituted C ₂ -C ₁₀ acyl, substituted or unsubstituted C ₂ -C ₁₀ ester group; wherein R is selected from the group consisting of H, C ₂ -C ₄ is an acyl group, a substituted or unsubstituted alkyl group of C ₁ -C _4;

Or R ₅ and R ₆ together form -R"-OR"-, wherein said R" is none or a substituted or unsubstituted C ₁ -C ₄ alkylene group;

R ₇ is a group selected from the group consisting of H, hydroxy, amino, substituted or unsubstituted C ₁ -C ₁₀ alkoxy, -COOR, substituted or unsubstituted C ₂ -C ₁₀ Acyl, substituted or unsubstituted C ₁ -C ₁₀ alkyl, substituted or unsubstituted C ₂ -C ₁₀ ester group; wherein R is selected from the group consisting of H, C ₂ -C ₄ acyl, substituted or not Substituted C ₁ -C ₄ alkyl;

n=1, 2, 3, 4, 5, 6, 7, 8, 9, or 10;

In the above formulae, one or more hydrogen atoms on the substituent group are substituted with a substituent selected from the group consisting of a carboxyl group, a phenyl group, a C ₃ -C ₆ cycloalkyl group, and a C ₂ -C ₁₀ ester. a halogen, a C ₁ -C ₁₀ alkyl-oxy group, a C ₂ -C ₁₀ acyl group, a hydroxyl group, a hydroxy-C ₁ -C ₁₀ alkylene group;

Said

For double or single button;

It is characterized in that it is used for preparation:

(a) a pharmaceutical composition for treating a hemangioma;

(b) a pharmaceutical composition for inhibiting proliferative activity of hemangio cells;

(c) a pharmaceutical composition that promotes apoptosis of hemangio cells;

(d) A pharmaceutical composition that inhibits cell formation into a tube.

In another preferred embodiment, the R _{3 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkyl group, C ₁ -C ₄ haloalkyl group;

R _{4 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkyl group, C ₁ -C ₄ haloalkyl group;

Or R ₃ and R ₄ together constitute a group selected from the group consisting of a substituted or unsubstituted 3-8 membered heterocyclic ring, a substituted or unsubstituted 5-7 membered heteroaryl ring.

In another preferred embodiment, the A ring is selected from the group consisting of a substituted or unsubstituted 5-7 membered carbocyclic ring, a substituted or unsubstituted 5-7 membered heterocyclic ring, a substituted or unsubstituted 6 membered aromatic ring. A substituted or unsubstituted 5-7 membered heteroaryl ring.

In another preferred embodiment, R ₇ is one or more groups selected from the group consisting of H, hydroxy, amino, C ₁ -C ₄ alkyl, C ₁ -C ₄ haloalkyl, C ₁ -C Alkoxy group of ₄ , -COOR; wherein R is selected from the group consisting of H, C ₁ -C ₄ acyl group, substituted or unsubstituted C ₁ -C ₄ alkyl group.

In another preferred embodiment, R ₁ is =0.

In another preferred embodiment, the R ₃ is a C ₁ -C ₄ alkyl group or a C ₁ -C ₄ haloalkyl group;

R _{4 is} selected from the group consisting of H and an oxygen atom;

Or R ₃ and R ₄ together constitute a group selected from the group consisting of a substituted or unsubstituted furan ring, a substituted or unsubstituted dihydrofuran ring, a substituted or unsubstituted tetrahydrofuran ring.

In another preferred embodiment, the compound of formula I is selected from the group consisting of:

In another preferred embodiment, the effective amount of the compound of formula I in the pharmaceutical composition is from 0.1 to 50 mg/kg body weight, preferably from 1 to 20 mg/kg body weight.

In another preferred embodiment, the pharmaceutical composition is a dosage form selected from the group consisting of an oral dosage form and an injectable dosage form.

In another preferred embodiment, the pharmaceutical composition is also for inhibiting angiogenesis of hemangioendothelial cells.

It is to be understood that within the scope of the present invention, the various technical features of the present invention and the various technical features specifically described hereinafter (as in the embodiments) may be combined with each other to form a new or preferred technical solution. Due to space limitations, we will not repeat them here.

DRAWINGS

Figure 1 - Figure 3: Dihydrotanshinone I and propranolol at a corresponding drug concentration (3uM, 50uM) and fixed time (48h), cell apoptosis in a time- and concentration-dependent manner;

Figure 4: Protein results of apoptosis of dihydrotanshinone I and propranolol;

Figure 5: Results of the effects of dihydrotanshinone I and propranolol on cell tube formation;

Figure 6: Results of the effects of dihydrotanshinone I and propranolol on hemangioma in mice.

Detailed ways

After long-term and intensive research, the present inventors have unexpectedly found that tanshinone compounds have good hemangio cell inhibitory activity and are superior in activity to the conventional general drug propranolol. The compounds inhibit proliferation of hemangio cells in vivo or in vitro in a time- and dose-dependent manner. Based on the above findings, the inventors completed the present invention.

the term

The term "C ₁ -C ₄ alkyl" or "C ₁ -C ₁₀ alkyl" as used herein refers to a straight or branched alkyl group having from 1 to 4 or 1 to 10 carbon atoms, such as methyl, Ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, or the like.

The term "C ₃ -C ₆ cycloalkyl" refers to a cycloalkyl group having 3 to 6 carbon atoms, such as cyclopropyl, cyclobutyl, methylcyclobutyl, cyclopentyl, or the like.

As used herein, the term "C ₁ -C ₁₀ acyl" or "C ₁ -C ₄ acyl" refers to a straight or branched alkyl/cycloalkyl group having the form of 0 to 9 or 0 to 3 carbon atoms. Substituents for the /aryl-carbonyl" structure, such as acetyl, propionyl, butyryl, or the like.

The term "C ₂ -C ₁₀ ester group" refers to a substituent such as a "linear or branched alkyl/cycloalkyl/aryl-carbonyl-oxygen atom having 1 to 9 carbon atoms" structure, such as an acetyl group. , propionyl, butyryl, or the like.

The term "C ₁ -C ₄ alkylene" refers to a group formed after the C1 to C4 alkyl group has lost a hydrogen atom as described above, for example, -CH ₂ -, -CH ₂ -CH ₂ -, or the like. .

The term "halogen" refers to F, Cl, Br and I.

The term "C ₆ -C ₁₀ aryl" refers to an aryl group having 6 to 10 carbon atoms, such as phenyl, naphthyl and the like.

The term "C ₁ -C ₁₀ heteroaryl" refers to a heteroaryl group having from 1 to 10 carbon atoms and one or more heteroatoms selected from O, S and/or N.

In the present invention, the terms "containing", "comprising" or "including" mean that the various ingredients may be used together in the mixture or composition of the present invention. Therefore, the terms "consisting essentially of" and "consisting of" are encompassed by the term "contains."

In the present invention, the term "pharmaceutically acceptable" ingredient means a substance which is suitable for use in humans and/or animals without excessive adverse side effects (such as toxicity, irritation, and allergic reaction), that is, a reasonable benefit/risk ratio.

In the present invention, the term "effective amount" means an amount of a therapeutic agent that treats, alleviates or prevents a target disease or condition, or an amount that exhibits a detectable therapeutic or prophylactic effect. The precise effective amount for a subject will depend on the size and health of the subject, the nature and extent of the condition, and the combination of therapeutic and/or therapeutic agents selected for administration. Therefore, it is useless to specify an accurate effective amount in advance. However, for a given condition, routine experimentation can be used to determine the effective amount that the clinician can determine.

As used herein, unless otherwise specified, the term "substituted" means that one or more hydrogen atoms on the group are substituted with a substituent selected from the group consisting of halogen, unsubstituted or halogenated C1-C6 alkyl, unsubstituted. Substituted or halogenated C ₂ -C ₆ acyl, unsubstituted or halogenated C1-C6 alkyl-hydroxy.

Unless otherwise stated, all compounds appearing in the present invention are meant to include all possible optical isomers, such as a single chiral compound, or a mixture of various chiral compounds (i.e., racemates). Among all the compounds of the present invention, each of the chiral carbon atoms may be optionally in the R configuration or the S configuration, or a mixture of the R configuration and the S configuration.

The term "compound of the invention" as used herein refers to a compound of formula I. The term also encompasses various crystalline forms, pharmaceutically acceptable salts, hydrates or solvates of the compounds of formula I.

As used herein, the term "pharmaceutically acceptable salt" refers to a salt of the compound of the invention formed with an acid or base suitable for use as a medicament. Pharmaceutically acceptable salts include inorganic and organic salts. A preferred class of salts are the salts of the compounds of the invention with acids. Suitable acids for forming salts include, but are not limited to, mineral acids such as hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, nitric acid, phosphoric acid, formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, Organic acids such as maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, methanesulfonic acid, benzoic acid, and benzenesulfonic acid; and acidic amino acids such as aspartic acid and glutamic acid.

Tanshinones and their applications

In the traditional medicine of infantile hemangiomas, it belongs to the "blood tumor" of surgery. The blood tumor grows longer than the skin, which is caused by the meridian block, the camp is not harmonious, and the blood stasis is caused. Therefore, the meridian is incomprehensible. Traditional medicine, medicine and medicine are mutually integrated, and hemangioma is often caused by blood stasis in the meridians.

Salvia miltiorrhiza is the root component of the plant of the Labiatae family. It is contained in the "Shen Nong's Herbal Classic" and is a commonly used traditional Chinese medicine for activating blood circulation, removing stasis and stasis. A series of tanshinones were isolated from the fat-soluble extract of Salvia miltiorrhiza (tanshinone extract). In vitro activity screening showed that tanshinone compounds significantly inhibited the proliferation of hemangio cells (positive control: Propranolol) Among them, Dihydrotanshinone I (DHTS) is the most effective and superior to Propranolol in inhibiting hemangio cells. The mechanism of action showed that both DHTS and Propranolol could significantly promote the apoptosis of hemangio cells. Apoptosis was time- and concentration-dependent at the corresponding drug concentration (3uM, 50uM) and fixed time (48h). Western Blot and tube formation results showed that DHTS also acted on hemangio cells through FAS/FASL and mitochondrial pathways, producing apoptosis effects and inhibiting angiogenesis of hemangioendothelial cells.

The tanshinone compound of the present invention has a structure represented by the following formula:

Wherein A, R ₁ , R ₂ , R ₃ , R ₄ , R ₅ , R ₆ and R ₇ are the corresponding groups in the specific compounds of the present invention.

Preferred tanshinone compounds of the invention include compounds selected from the group consisting of:

Pharmaceutical composition and method of administration

Since the compound of the present invention has excellent inhibitory activity against hemangio cells, the compound of the present invention and various crystal forms thereof, pharmaceutically acceptable inorganic or organic salts, hydrates or solvates, and compounds containing the present invention are mainly The pharmaceutical composition of the active ingredient can be used for the treatment, prevention, and alleviation of diseases caused by hemangio cell production, such as fatty liver and the like.

The pharmaceutical compositions of the present invention comprise a safe or effective amount of a compound of the present invention, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient or carrier. By "safe and effective amount" it is meant that the amount of the compound is sufficient to significantly improve the condition without causing serious side effects. In general, the pharmaceutical compositions contain from 1 to 3000 mg (active dose range 3-30 mg/kg) of a compound/agent of the invention, more preferably from 10 to 2000 mg of a compound/agent of the invention. Preferably, the "one dose" is a capsule or tablet.

"Pharmaceutically acceptable carrier" means: one or more compatible solid or liquid fillers or gel materials which are suitable for human use and which must be of sufficient purity and of sufficiently low toxicity. By "compatibility" it is meant herein that the components of the composition are capable of intermingling with the compounds of the invention and with each other without significantly reducing the efficacy of the compound. Examples of pharmaceutically acceptable carriers are cellulose and its derivatives (such as sodium carboxymethylcellulose, sodium ethylcellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as stearic acid). , magnesium stearate), calcium sulfate, vegetable oil (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyol (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifier (such as Tween

), a wetting agent (such as sodium lauryl sulfate), a coloring agent, a flavoring agent, a stabilizer, an antioxidant, a preservative, a pyrogen-free water, and the like.

The mode of administration of the compound or pharmaceutical composition of the present invention is not particularly limited, and representative modes of administration include, but are not limited to, oral, intratumoral, rectal, parenteral (intravenous, intramuscular or subcutaneous), and topical administration. .

Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or mixed with: (a) a filler or compatibilizer, for example, Starch, lactose, sucrose, glucose, mannitol and silicic acid; (b) binders, for example, hydroxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose and gum arabic; (c) humectants, For example, glycerin; (d) a disintegrant such as agar, calcium carbonate, potato starch or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) a slow solvent such as paraffin; (f) Absorbing accelerators, for example, quaternary amine compounds; (g) wetting agents, such as cetyl alcohol and glyceryl monostearate; (h) adsorbents, for example, kaolin; and (i) lubricants, for example, talc, hard Calcium citrate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, or a mixture thereof. In capsules, tablets and pills, the dosage form may also contain a buffer.

Solid dosage forms such as tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other materials known in the art. They may contain opacifying agents and the release of the active compound or compound in such compositions may be released in a portion of the digestive tract in a delayed manner. Examples of embedding components that can be employed are polymeric and waxy materials. If necessary, the active compound may also be in microencapsulated form with one or more of the above-mentioned excipients.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or elixirs. In addition to the active compound, the liquid dosage form may contain inert diluents conventionally employed in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1 , 3-butanediol, dimethylformamide and oils, especially cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil or a mixture of these substances.

In addition to these inert diluents, the compositions may contain adjuvants such as wetting agents, emulsifying and suspending agents, sweetening agents, flavoring agents and perfumes.

In addition to the active compound, the suspension may contain suspending agents, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar or mixtures of these and the like.

Compositions for parenteral injection may comprise a physiologically acceptable sterile aqueous or nonaqueous solution, dispersion, suspension or emulsion, and a sterile powder for reconstitution into a sterile injectable solution or dispersion. Suitable aqueous and nonaqueous vehicles, diluents, solvents or vehicles include water, ethanol, polyols, and suitable mixtures thereof.

Dosage forms for the compounds of the invention for topical administration include ointments, powders, patches, propellants and inhalants. The active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or, if necessary, propellants.

The compounds of the invention may be administered alone or in combination with other pharmaceutically acceptable compounds.

When a pharmaceutical composition is used, a safe and effective amount of a compound of the invention is administered to a mammal (e.g., a human) in need of treatment wherein the dosage is a pharmaceutically effective effective dosage, for a 60 kg body weight, The dose to be administered is usually from 1 to 2000 mg, preferably from 6 to 600 mg. Of course, specific doses should also consider factors such as the route of administration, the health of the patient, etc., which are within the skill of the skilled physician.

The main advantages of the present invention over the prior art include:

1. It was first discovered that the tanshinone compound has good hemangio cell inhibition activity, and the in vitro activity is superior to the propranolol known in the prior art;

2. The tanshinone compound of the present invention can effectively inhibit hemangio cells to promote apoptosis, and can also inhibit angiogenesis of hemangioendothelial cells;

3. The compound of the present invention is effective in vitro and in vivo, and has a low active dose, and can exhibit an antitumor effect at a dose as low as 10 mg/kg, which is far lower than the effective dose of propranolol.

The invention is further illustrated below in conjunction with specific embodiments. It is to be understood that the examples are not intended to limit the scope of the invention. The experimental methods in the following examples which do not specify the specific conditions are usually in accordance with conventional conditions or according to the conditions recommended by the manufacturer. Percentages and parts are by weight unless otherwise stated.

1, instruments and equipment

In addition, the spectrum UV: SHIMADIU UV-2550 and Beckman DU-7 UV-Vis spectrometer;

Infrared spectroscopy IR: Perkin-Elmer 577 infrared spectrophotometer;

Low resolution mass spectrometry LR-EIMS: Finnigan MAT-95;

High resolution mass spectrometry HR-EIMS: Kratos 1H spectrometer;

Nuclear magnetic resonance spectrum: Varian INOVA 600 nuclear magnetic resonance spectrometer, Bruker AM-500, AM-400, AM-300 nuclear magnetic resonance spectrometer, δ (ppm), with TMS as internal standard;

LC-MS: Agilent 1100 liquid phase coupled Bruker esquire mass spectrometer;

Analytical HPLC: Waters 2690 Separate Model, Waters PDA 996 detector coupled Alltch ELSD 2000 detector, Millennium 2000 operating system, Waters RP18 column (5.0 x 125 mm, 5 μm, Waters), flow rate 1.0 ml/min, CH3CN (Merck, Germany) , H2O (Rapex); Jasco HPLC (Chiralcel IA column, 5 μm, 150 × 4.6 mm), flow rate 0.6 ml / min, hexane / ethanol (7:3).

HPLC-MS: Waters 2695 Separate Model, Waters PDA 2998 detector coupled with Alltch ELSD 2424 detector, 3100 Ms detector, SunFireTM C-18 column (4.6 x 100 mm, 3.5 μm, Waters), flow rate 1.0 ml/min, CH3CN (Merck, Germany) , H2O (Rapex); Jasco HPLC (Chiralcel IA column, 5 μm, 150 × 4.6 mm), flow rate 0.6 ml / min, hexane / ethanol (7:3).

Preparative HPLC: Varian SD1 instrument, Varians 320 single wavelength detector, C18 column (220 x 25 nm, 10 μm, Waters), flow rate 15 ml/min, CH3CN (Merck, Germany), H2O (Rapex);

Main equipment for biological experiments: carbon dioxide cell incubator, microplate reader, centrifuge, purification bench and chemiluminometer (Thermo Scientific, USA); optical microscope and photomicrography system Leica DM 4000B microscope (Lecia, Germany); inverted Fluorescence microscopy (OLYMPUS, Japan); flow cytometry (Beckman Coulter); horizontal shaker (Beijing Dingguo Changsheng Biotechnology Co., Ltd.); electrophoresis instrument (Bio-Rad, USA); fully automatic autoclave (Tomy) Digital biology, Japan); PVDF membrane (Millipore, USA);

2, reagents and materials

Column chromatography silica gel: 100-200 mesh, 200-300 mesh silica gel and silica gel H are produced by Qingdao Ocean Chemical Plant;

TLC thin layer preparation board: HSGF254 is produced by Yantai Chemical Plant;

MCI resin: CHP20P (75-150μm) is produced by Mitsubishi Corporation;

Sephadex LH-20: Pharmacia Biotech AB, Uppsala, Sweden.

Color developing agent: 10% sulfuric acid-vanillin solution, potassium permanganate aqueous solution;

Plant material: Danshen was purchased from the Luzhou medicinal materials market and collected and identified by Associate Professor Shen Jingui from the Shanghai Institute of Materia Medica. The specimens were kept in the specimen room.

Main biological reagents: 1640 medium, fetal bovine serum (Gibco, USA); double antibody (Sigma, USA); antibody: Caspase 3, Caspase 8, Caspase 9, AIF, PARP, Bax, FADD, Cyst3 (Abclonal/China Cyclodextrin (Medchem, USA); Flow Apoptosis Fluorescent Reagent (FITC-Annexin, PI) (BD, USA); Matrigel (Corning, USA); Western Blot Glue Reagent, Secondary Antibody, CCK8 Reagent, Enhanced chemiluminescence agent, Tween-20 and crystal violet (翊圣/中国); protease inhibitor (Roche, Switzerland); experimental animals are nude mice, weighing 20±5g, purchased from Xinhua Hospital affiliated to Shanghai Jiao Tong University, animals are stored in Xinhua Hospital SPF animal room.

Example 1: Isolation and Identification of Tanshinone Compounds

30 kg of Salvia miltiorrhiza was pulverized and then immersed in 95% industrial ethanol (50 L) for 5 days at room temperature for a total of 3 times. The ethanol extracts were combined and concentrated under reduced pressure to obtain a flow extract. The extract was suspended in 10 L of distilled water and extracted with 8 L of ethyl acetate three times. The ethyl acetate extract was combined and concentrated under reduced pressure to give the total extract of tanshinone.

Take 200g of total extract of tanshinone, dilute it in dichloromethane, and purify it through silica gel column (2kg, 200-300 mesh, produced by Qingdao Ocean Chemical Industry Co., Ltd.) with petroleum ether-ethyl acetate 100:2,100::3,100: 5,100:10, 100:20, 100:50, ethyl acetate eluted with low elution, and after TLC detection, 18 fractions were combined. Fraction S3 (5.46 g) was recrystallized from petroleum ether-ethyl acetate to afford tanshinone IIA (3.2 g). The fraction S6 (4.32g) was subjected to silica gel column chromatography (petroleum ether: ethyl acetate 100:3 to 10:1) to obtain 5 fractions of S6A-S6E; fraction S6A (152mg) was subjected to Sephadex LH-20 gel column chromatography. (Chloroform: methanol 1:1) and then purified by preparative silica gel plate (dichloromethane) to obtain the compound Danshenxin (5 mg); fraction S6C (311 mg) was subjected to ODS column chromatography (methanol-water, 65%-95) %) Purified compound saponin 醌A (10 mg). The 800 mg fraction S10 (6.5 g) was separated by ODS column chromatography (methanol-water, 65%-95%), and the fraction S10C was purified by preparative silica gel plate (dichloromethane) to give the compound isopropyl orthophenanthrene.醌 (10mg). Fraction S13 (10.89 g) was recrystallized from petroleum ether-ethyl acetate to give tanshinone I (3.2 g). Fraction S14 (17.72 g) was recrystallized from petroleum ether-ethyl acetate to afford ss. Fraction S15 (3.61g) was separated by silica gel column chromatography (petroleum ether: acetone 30:1 to 0:1) to obtain 5 fractions S15A-S15E; 25mg of S15D was passed through Sephadex LH-20 (chloroform:methanol 1:1) Column gel chromatography gave the compound tanshinone methyl ester (15 mg). The fraction obtained by purifying the fraction S16 (20 g) through a silica gel column (petroleum ether: acetone 100:3 to 100:10) was subjected to MCI column chromatography (ethanol-water 80% to 95%) and Sephadex LH-20 (chloroform:methanol 1). :1) After column gel chromatography, the compound dihydrotanshinone I (1.2 g) was obtained. The fraction S18 (950 mg) was subjected to Sephadex LH-20 gel column chromatography (chloroform: methanol 1:1), and the fraction S18C obtained was subjected to ODS column chromatography (methanol-water, 65%-95%) and purified by preparative silica gel. (Dichloromethane) gave the compound Danshenol B (3 mg) and tanshinone IIB (5 mg).

Tanshinone IIA, Tanshinone IIA, cherry red needle crystal. EI MS m/z 284 [M] ⁺ ; ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.59 (d, J = 8.0 Hz, 1H, H-6), 7.10 (d, J = 8.0 Hz, 1H, H -7), 7.07 (d, J = 1.0 Hz, 1H, H-14), 3.17 (t, J = 6.4 Hz, 1H, H-1), 3.02 (m, 1H, H-15), 1.79 (m) , 2H, H-3), 1.65 (m, 2H, H-2), 1.29 (s, 3H, H-18, 19), 1.16 (d, J = 6.19 Hz, 3H, H-16, 17). ¹³ C NMR (125 MHz, CDCl ₃ ) δ 183.6, 175.8, 161.7, 150.2, 144.3, 141.3, 133.6, 127.4, 126.6, 121.1, 120.2, 38.0, 34.7, 31.9, 29.8, 19.1, 8.7.

Salvia miltiorrhiza, Rosmariquinone, fuchsia solid. EI MS m/z 284 [M] ⁺ ; ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.59 (d, J = 8.0 Hz, 1H, H-6), 7.10 (d, J = 8.0 Hz, 1H, H -7), 7.07 (d, J = 1.0 Hz, 1H, H-14), 3.17 (t, J = 6.4 Hz, 1H, H-1), 3.02 (m, 1H, H-15), 1.79 (m) , 2H, H-3), 1.65 (m, 2H, H-2), 1.29 (s, 3H, H-18, 19), 1.16 (d, J = 6.19 Hz, 3H, H-16, 17). ¹³ C NMR (125 MHz, CDCl ₃ ) δ 182.6, 181.7, 149.9, 145.2, 144.7, 140.1, 134.6, 134.0, 128.4, 128.1, 38.0, 34.7, 32.0, 30.1, 27.1, 21.7, 19.3.

轮A, Sibiriquinone, sakura red solid; EI MS m/z 284 [M] ⁺ ; ¹ H NMR (400 MHz, CDCl3) δ 7.86 (d, J = 9.8 Hz, 1H, H-1) , 7.50 (d, J = 7.8 Hz, 1H, H-6), 7.11 (d, J = 7.9 Hz, 1H, H-7), 7.08 (s, 1H), 6.33 (dt, J = 9.7, 4.6 Hz) , 1H, H-2), 3.02 (p, J = 6.9 Hz, 1H, H-15), 2.27 (dd, J = 4.6, 1.9 Hz, 2H, H-3), 1.28 (s, 6H, H-) 18,H-19), 1.16 (d, J=6.9 Hz, 6H, H-16, H-17). ¹³ C NMR (126MHz, CDCl ₃ ) δ 183.38, 181.71, 148.15, 145.13, 140.09, 137.42, 134.61, 134.41, 130.74, 129.37, 124.86, 124.84, 38.16, 34.22, 28.52, 27.06, 21.68.

Isopropyl ortho-phenanthrenequinone, Ro 09-0680, cherry red solid; EI MS m/z 264 [M] ⁺ ; ¹ H NMR (500 MHz, CDCl ₃ ) δ 9.26 (d, J = 9.0 Hz, 1H, H-6), 8.30 (d, J = 9.0 Hz, 1H, H-7), 7.00-7.70 (overlap, 3H, H-1, 2, 3), 7.14 (s, 1H, H-14), 7.10 (d, J = 8.0 Hz, 1H, H-7), 7.07 (d, J = 1.0 Hz, 1H, H-14), 3.07 (m, 1H, H-15), 2.70 (s, 3H, H-) 18), 1.23 (d, J = 6.0 Hz, 6H, H-16, 17). ¹³ C NMR (125 MHz, CDCl ₃ ) δ 182.5, 181.7, 139.6, 136.9, 136.7, 135.1, 132.9, 132.9, 130.9, 128.5, 126.7, 125.4, 125.4, 125.0, 27.3, 21.7, 21.7, 19.9.

Tanshinone I, Tanshinone I, cherry red solid, EI MS m/z 264 [M] ⁺ ; ¹ H NMR (500 MHz, CDCl ₃ ) δ 9.20 (d, J = 9.0 Hz, 1H, H-6), 8.23 ( d, J = 8.0 Hz, 1H, H-7), 7.74 (d, J = 9 Hz, 1H, H-1), 7.51 - 7.54 (m, 1H, H-2), 7.32 (d, J = 9.0 Hz) , 1H, H-3), 7.27 (d, J = 1.5 Hz, 1H, H-17), 2.66 (s, 3H, H-18), 2.27 (d, J = 1.5 Hz, 1H, H-16) ¹³ C NMR (125 MHz, CDCl ₃ ) δ 183.4, 175.6, 161.1, 142.0, 135.2, 133.6, 132.7, 130.6, 129.6, 128.3, 124.7, 123.1, 121.7, 123.1, 121.7, 120.5, 118.7, 19.8, 8.8.

Cryptotanshinone, Cryptotanshinone, cherry red needle crystal, EI MS m/z 296 [M] ⁺ ; ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.64 (d, J = 8.0 Hz, 1H, H-6), 7.51 (d, J = 8.0 Hz, 1H, H-7), 4.90 and 4.37 (dd, J = 6.0, 9.6 Hz, 2H, 17), 3.59-3.65 (m, 1H), 3.22 (m, 2H), 1.77 -1.83 (m, 2H), 1.65-1.69 (m, 2H), 1.36 (d, J = 6.0 Hz, 3H, H-16), 1.32 (s, 6H, H-18/19). ¹³ CNMR (125MHz , CDCl ₃ ) δ 184.0, 175.5, 170.8, 152.3, 143.6, 132.5, 128.8, 126.1, 122.5, 118.2, 81.5, 38.7, 34.7, 34.6, 31.7, 29.6, 19.3, 18.7.

Tanshinone methyl ester, Methyl tanshinonate, red solid, EI MS m/z 338 [M] ⁺ . ¹ H NMR (500MHz, CDCl ₃ ) δ 7.56 (d, J = 8.3 Hz, 1H, H-6), 7.48 ( d, J = 8.3 Hz, 1H, H-7), 7.23 (d, J = 1.4 Hz, H-17), 3.67 (s, 3H, -OCH ₃ ), 3.24 (m, 2H, H-1), 2.27 (d, J = 1.4 Hz, 3H, H-16), 2.26 and 1.75 (m, each 1H, H-3), 1.82 (m, 2H, H-2), 1.58 (s, 3H, H-18) ¹³ CNMR (125 MHz, CDCl ₃ ) δ 183.6, 177.3, 175.8, 161.5, 144.6, 143.3, 141.8, 135.2, 128.6, 126.7, 121.5, 120.5, 52.8, 47.4, 34.2, 29.2, 27.8, 19.4, 9.0.

15,16-dihydrotanshinone I,15,16-Dihydrotanshinone I, cherry red needle-like solid, EI MS m/z 278 [M] ⁺ . ¹ H NMR (500 MHz, CDCl ₃ ) δ 9.28 (d, J = 9.0 Hz, 1H, H-6), 8.30 (d, J = 9.0 Hz, 1H, H-7), 7.76 (d, J = 9.2 Hz, H-1), 7.57 (m, 1H, H-2), 7.40 (d, J = 7.0 Hz, 1H, H-3), 4.97 and 4.43 (t, J = 6.3, 9.5 Hz, each 1H, H-17), 3.66 (m, 1H, H-17), 2.70 ( s, 3H, H-18), 1.41 (d, J = 6.5 Hz, 1H, H-16). ¹³ C NMR (125MHz, CDCl ₃ ) δ 184.3, 175.8, 170.6, 135.0, 134.6, 132.2, 131.9, 130.4, 128.9, 128.2, 126.2, 125.0, 120.5, 118.3, 81.6, 34.7, 19.8, 18.8.

Danshenol B, Tanshinol B, cherry red solid, EI MS m/z 296 [M] ⁺ . ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.95 (d, J = 8.1 Hz, 1H, H-6), 7.61 (d, J = 8.3 Hz, 1H, H-15), 7.25 (m, H-7), 3.28 (m, 1H, H-1), 3.16 (m, 1H, H-1), 2.27 (d, J = 1.3 Hz, 3H, H-17), 2.00 (overlap, 2H), 1.83 (m, 1H), 1.56 (s, 3H, H-18). ¹³ C NMR (126 MHz, CDCl ₃ ) δ 183.35, 175. 161.39,147.29, 143.95,141.74,133.68,129.04,126.09,121.46,120.80,120.49,71.03,38.75,31.18,29.14,20.17,8.94.

Tanshinone IIB, Tanshinone IIB, cherry red solid, EI MS m/z 310 [M] ⁺ . ¹ H NMR (600 MHz, CDCl ₃ ) δ 7.65 (d, J = 8.1 Hz, 1H, H-6), 7.52 ( d, J = 8.1 Hz, 1H, H-6), 7.21 (s, 1H, H-16), 3.79 (d, J = 11.1 Hz, 1H, H-18b), 3.62 (d, J = 11.3 Hz, 1H, H-18a), 3.18 (m, 2H, H-1), 2.28 (s, 3H, H-17), 1.98 (m, 1H, H-3b), 1.88 (m, 1H, H-2b) , 1.75 (m, 1H, H-2a), 1.58 (m, 1H, H-3a), 1.29 (s, 3H, H-19). ¹³ C NMR (126MHz, CDCl ₃ ) δ 183.38, 175.57, 161.55, 146.31 , 146.08, 141.59, 133.87, 127.94, 126.67, 121.31, 120.36, 120.24, 71.66, 40.09, 32.38, 29.85, 26.81, 18.91, 8.94.

Example 2: Cell proliferation inhibitory activity of tanshinones on hemangio cells

1). Cell culture: The EOMA cell line originated from the spontaneous hemangio cells of the 129/J line. The cell culture was carried out in 1640 medium containing 10% fetal calf serum, and mixed with 100 mg/l of penicillin and 100 ug/l of streptomycin. The cells were cultured in an incubator containing 5% CO 2 at 37 °C. The medium was changed every other day and passage was carried out at a cell density of 80%.

2). Dissolution of the drug: The drug powder was dissolved in DMSO, stored at a concentration of 10 uM, and stored at -20 degrees.

3).CCK8 detection IC ₅₀ : Before dosing treatment, the cells were plated in a 96-well plate, and the cells were counted so that the cell concentration of each well reached 2000-3000/ul. After the cells were attached, the medium was aspirated and added. Different concentrations of dihydrotanshinone I. After 72 h, the medium containing the drug was aspirated, cck8 was added, and three blank control wells were set at the same time, and the absorbance at 450 wavelength was measured around 2 h. According to: Calculate the percentage of cell proliferation at different concentrations [(medication group - blank control group) * 100% / (control group - blank control group)], non-linear regression by Graphpad Prim software, measured cell proliferation rate decreased The drug concentration corresponding to half - IC ₅₀ .

The cell proliferation inhibitory activity of the tanshinone compound on hemangio cells is shown in Table 1 below. The activity test results showed that the tanshinone compound can significantly inhibit the proliferation of hemangio cells.

Table 1: Inhibitory activity of tanshinones on proliferation of hemangio cells

Compound name

IC50 (μM)

Dihydrotanshinone I	2.63
Salvia miltiorrhiza	3.17
轮叶婆婆纳对醌A	3.19
Tanshinone methyl ester	4.33
Danshen alcohol B	5.32
Isopropyl ortho-phenanthrene	7.83
Cryptotanshinone	10.48
Positive control (propranolol)	53.67

Example 3: Effect of dihydrotanshinone I on apoptosis of hemangio cells

1) Cell clone formation experiment: 1000 cells / 2 ml cells were added to a six-well plate. After the cells were attached to the cells, different drug concentrations were added for treatment. The blank group was treated with DMSO (solvent) for comparison. (24h, 48h), replaced with fresh medium, and after 10 days, the crystals were stained and photographed.

2) Flow cytometry and hochest33342 to detect the percentage and morphology of apoptotic cells: Prepare the cells, plate them in a six-well plate, and after the cells are attached, perform drug treatment. After meeting the conditions (different concentrations, different time treatment) The cells were collected, added to a centrifuge tube, centrifuged at 800 rpm for 5 min, pre-cooled by eppendorf, then washed with pbs, centrifuged in a pre-cooled centrifuge, 800 rpm, 5 min, and after pipetting, the sample was added to 200 ul of binding buffer and then added. 5ul FITC-Annexin V was resuspended, protected from light for 10 min, then added 5 ul of PI, transferred into a flow tube, 4 degrees Celsius, protected from light, and applied to the machine after one hour.

3) Hochest33258 staining: cell plating was performed in a 12-well plate. After the cells were attached to the wall, the drug was treated. After the corresponding conditions were met, the cells were fixed, then the fixative was washed away, and hoses 33258 was added for staining at 37 ° C for 30 min. . Observe the densely packed nucleus.

The test results are shown in the following figure 1-3: dihydrotanshinone I and propranolol at the corresponding drug concentration (3uM, 50uM) and fixed time (48h), cell apoptosis time and concentration-dependent, analysis of variance The results suggest that the difference is statistically significant.

4) Western blot analysis of apoptotic proteins: different protein expression levels in cells can be detected by wb. After the cells were treated with the drug (dihydrotanshinone I and propranolol) and the solvent, the pbs were washed and collected, and then the cells were lysed with RIPA for 30 min on ice, and then lysed and centrifuged at 12000 g at 4 ° C. The cells were incubated for 10 min and the cell supernatant was taken. Proteins were quantified using BCA to adjust the protein to the same concentration. Load enough in the electrophoresis solution to ensure that the sample loading is 20-30ug. Transfer the protein on the SDS-PAG gel to the PVDF membrane at 300Ma, 5% BSA for 1h, and add 1:1000. Anti-over, 4 degrees overnight, TBST wash the membrane three times, each time 10 min, then add secondary antibody at a concentration of 1:3000, incubate at room temperature for 1 h, TBST wash the membrane three times, each 10 min, add luminescent liquid, and develop.

The results of Western Blot are shown in Figure 4. DHTS can act on hemangio cells through FAS/FASL and mitochondrial pathways at the same time. The mitochondrial pathway is dominant at low concentrations, and the FASL pathway is dominant at high concentrations. PropranololZ is at low concentrations. The FASL pathway is dominant, and the mitochondrial pathway is dominant at high concentrations.

Example 4: Tube formation observes the effect of drugs on cell formation:

The unsolidified Matrigel was placed on a 96-well plate in an amount of 10 ul per well, and the 96-well plate was placed in an incubator for 1 h to completely coagulate the colloid. EOMA cells (3×10 ⁴ /well) were added to the 96-well plate, and The medium with or without drug was placed on the surface of the colloid and cells, and was placed in the incubator. After 2 hours, it was observed under a microscope and photographed and retained.

The tube-forming experiment reflects the angiogenesis of cells in vivo, including the action of vascular endothelial growth factor (VEGF), the degradation of intercellular matrix (mmp9), and the migration of endothelial cells.

The results of Tube formation and Western blot are shown in Figure 5. After the drug is applied to the cells, the endothelial growth factor is decreased, the metal matrix enzyme synthesis is reduced, and the endothelial cells are unable to migrate, resulting in the cells being unable to undergo angiogenesis, which is sufficient to demonstrate the ability of the drug to inhibit blood vessels. .

Example 5: In vivo experiments in animals to explore the effect of drugs on hemangioma

The mouse species used in the in vivo experiments were BALB/c nude mice, all of which were 6-week females, a total of 15 rats. After collecting about 1×107 cells, they were suspended in 200 ul of pbs and inoculated into the armpits of nude mice for about two weeks, after the average size of the tumors reached 100 mm 3 (volume = length × width ² /2). Randomly grouped. According to the literature, dihydrotanshinone I (10mg/kg), propranolol (40mg/kg) and solvent (β-cyclodextrin + 20% DMSO) were injected into the abdominal cavity, and the changes of the tumor were detected three times a week. The body weight of nude mice, 4 weeks later, the nude mice were sacrificed, the tumor was cut and measured, and some tissues were immunohistochemically analyzed for changes in CD31, CD34, vWF, Glut1, Caspase3, MMP9 and VEGFR2.

The test results are shown in Figure 6. Both dihydrotanshinone I and Propranolol can inhibit hemangiomas at the corresponding concentrations. The results of analysis of variance suggest that the effect of drug production is statistically significant (p=0.0008). The results of DHTS and Propranolol showed no significant difference (P=0.097).

All documents mentioned in the present application are hereby incorporated by reference in their entirety in their entireties in the the the the the the the the the In addition, it should be understood that various modifications and changes may be made by those skilled in the art in the form of the present invention.

Claims (10)

1. Use of a compound of the formula I, or a pharmaceutically acceptable salt thereof, as follows:

   

   Wherein the A ring is selected from the group consisting of a substituted or unsubstituted 3-10 membered carbocyclic ring, a substituted or unsubstituted 3-10 membered heterocyclic ring, a substituted or unsubstituted C ₆ -C ₁₀ aromatic ring, substituted or unsubstituted 5-12 yuan heteroaryl ring;

   R _{1 is} selected from the group consisting of H, oxygen atom (=O), O-R';

   R _{2 is} selected from the group consisting of H, oxygen atom, O-R';

   The R' is selected from the group consisting of H, a substituted or unsubstituted C ₂ -C ₁₀ acyl group, a fluorenylmethoxycarbonyl group (Fmoc), or a substituted or unsubstituted C ₁ -C ₁₀ alkyl group; Ground, R' is H;

   R _{3 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ alkyl group, C ₁ -C ₄ haloalkyl group, substitution Or an unsubstituted C ₂ -C ₁₀ acyl group, a substituted or unsubstituted C ₂ -C ₁₀ ester group, -OC(O)-R, -NH-R; wherein R is selected from the group consisting of H, C ₁ -C ₄ acyl group, a substituted or unsubstituted C ₁ -C ₄ alkyl group;

   R _{4 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ alkyl group, C ₁ -C ₄ haloalkyl group, substitution Or an unsubstituted C ₂ -C ₁₀ acyl group, a substituted or unsubstituted C ₂ -C ₁₀ ester group, -OC(O)-R, -NH-R; wherein R is selected from the group consisting of H, C ₁ -C ₄ acyl group, a substituted or unsubstituted C ₁ -C ₄ alkyl group;

   Or any two of R ₁ , R ₂ , R ₃ and R ₄ together with an adjacent carbon atom constitute a group selected from the group consisting of a substituted or unsubstituted C ₃ -C ₁₀ membered carbocyclic ring, substituted or unsubstituted a substituted 3-10 membered heterocyclic ring, a substituted or unsubstituted C ₆ -C ₁₀ aromatic ring, a substituted or unsubstituted 5-12 membered heteroaryl ring;

   R _{5 is} selected from the group consisting of H, halogen, cyano, hydroxy, C ₁ -C ₄ alkoxy, -COOR, substituted or unsubstituted C ₁ -C ₁₀ alkyl, substituted or unsubstituted C ₁ -C _{a 10} haloalkyl group, a substituted or unsubstituted C ₂ -C ₁₀ acyl group, a substituted or unsubstituted C ₂ -C ₁₀ ester group; wherein R is selected from the group consisting of H, C ₂ -C ₄ acyl groups, substitutions Or an unsubstituted C ₁ -C ₄ alkyl group;

   R _{6 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, carboxyl group, hydroxyl group, C ₁ -C ₄ alkoxy group, -COOR, substituted or unsubstituted C ₁ -C ₁₀ alkyl group, substituted or unsubstituted C ₁ -C ₁₀ haloalkyl, substituted or unsubstituted C ₂ -C ₁₀ acyl, substituted or unsubstituted C ₂ -C ₁₀ ester group; wherein R is selected from the group consisting of H, C ₂ -C ₄ is an acyl group, a substituted or unsubstituted alkyl group of C ₁ -C _4;

   Or R ₅ and R ₆ together form -R"-OR"-, wherein said R" is none or a substituted or unsubstituted C ₁ -C ₄ alkylene group;

   R ₇ is a group selected from the group consisting of H, hydroxy, amino, substituted or unsubstituted C ₁ -C ₁₀ alkoxy, -COOR, substituted or unsubstituted C ₂ -C ₁₀ Acyl, substituted or unsubstituted C ₁ -C ₁₀ alkyl, substituted or unsubstituted C ₂ -C ₁₀ ester group; wherein R is selected from the group consisting of H, C ₂ -C ₄ acyl, substituted or not Substituted C ₁ -C ₄ alkyl;

   n=1, 2, 3, 4, 5, 6, 7, 8, 9, or 10;

   In the above formulae, one or more hydrogen atoms on the substituent group are substituted with a substituent selected from the group consisting of a carboxyl group, a phenyl group, a C ₃ -C ₆ cycloalkyl group, and a C ₂ -C ₁₀ ester. a halogen, a C ₁ -C ₁₀ alkyl-oxy group, a C ₂ -C ₁₀ acyl group, a hydroxyl group, a hydroxy-C ₁ -C ₁₀ alkylene group;

   The "" is a double bond or a single bond;

   It is characterized in that it is used for preparation:

   (a) a pharmaceutical composition for treating a hemangioma;

   (b) a pharmaceutical composition for inhibiting proliferative activity of hemangio cells;

   (c) a pharmaceutical composition that promotes apoptosis of hemangio cells;

   (d) A pharmaceutical composition that inhibits cell formation into a tube.
2. The use according to claim 1, wherein said R _{3 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkyl group, C ₁ -C a halogenated alkyl group of ₄ ;

   R _{4 is} selected from the group consisting of H, halogen, oxygen atom, cyano group, hydroxyl group, carboxyl group, C ₁ -C ₄ alkyl group, C ₁ -C ₄ haloalkyl group;

   Or R ₃ and R ₄ together constitute a group selected from the group consisting of a substituted or unsubstituted 3-8 membered heterocyclic ring, a substituted or unsubstituted 5-7 membered heteroaryl ring.
3. The use according to claim 1, wherein said ring A is selected from the group consisting of a substituted or unsubstituted 5-7 membered carbocyclic ring, a substituted or unsubstituted 5-7 membered heterocyclic ring, substituted or unsubstituted. Substituted 6-membered aromatic ring, substituted or unsubstituted 5-7 membered heteroaryl ring.
4. The use according to claim 1, wherein R ₇ is one or more groups selected from the group consisting of H, hydroxy, amino, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkyl halide a C ₁ -C ₄ alkoxy group, -COOR; wherein R is selected from the group consisting of H, a C ₁ -C ₄ acyl group, a substituted or unsubstituted C ₁ -C ₄ alkyl group.
5. The use according to claim 1, wherein R ₁ is =0.
6. The use according to claim 1, wherein R ₃ is a C ₁ -C ₄ alkyl group or a C ₁ -C ₄ haloalkyl group;

   R _{4 is} selected from the group consisting of H and an oxygen atom;

   Or R ₃ and R ₄ together constitute a group selected from the group consisting of a substituted or unsubstituted furan ring, a substituted or unsubstituted dihydrofuran ring, a substituted or unsubstituted tetrahydrofuran ring.
7. The use according to claim 1 wherein the compound of formula I is selected from the group consisting of:

   
8. The use according to claim 1, characterized in that in the pharmaceutical composition, the effective dose of the compound of the formula I is from 0.1 to 50 mg/kg body weight, preferably from 1 to 20 mg/kg body weight.
9. The use according to claim 1, wherein the pharmaceutical composition is a dosage form selected from the group consisting of an oral dosage form and an injection dosage form.
10. The use according to claim 1, wherein said pharmaceutical composition is further for inhibiting angiogenesis of vascular endothelial cells.

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