CN108272790A - A kind of medicinal usage of xanthones compound A - Google Patents

Abstract

The invention discloses a pharmaceutical application of a xanthenone compound A, and the xanthenone compound A has a chemical structure shown in formula I: The chemical name is: 12R-isobractatin; the pharmaceutical use refers to the xanthanone compound A or its hydrate, pharmaceutically acceptable salt, tautomer, stereoisomer, precursor compound At least one of them is used as an active ingredient in the preparation of antitumor drugs. Experiments show that the xanthanone compound A of the present invention can significantly inhibit the proliferation of human prostate cancer cell PC-3, human cervical cancer cell HeLa, human non-small cell lung cancer cell A549 and human colon cancer cell HT-29, and has The effect of inhibiting the proliferation of tumor cells is expected to be used as an active ingredient in the preparation of antitumor drugs, and has medicinal prospects.

Description

A kind of pharmaceutical use of xanthones compound A

technical field

The invention relates to a pharmaceutical use of a xanthone compound A, belonging to the technical field of medicine.

Background technique

Malignant tumor is one of the main fatal diseases in the world, which seriously threatens human health and life. Chinese Academy of Medical Sciences Cancer Hospital, Academician Hejie of the National Cancer Center, and Professor Chen Wanqing, director of the National Cancer Registry Center, published the 2015 Chinese cancer statistics in the CA:ACancer Journal for Clinicians, showing that there were about 4.292 million new tumors in China in 2015 cases and 2.814 million deaths. With the intensification of population aging, the incidence of cancer in my country is on the rise, which has caused a huge burden on my country's health resources and national economy. Cancer control has become the focus of my country's health strategy. The National Science and Technology Major Project "Major New Drug Creation" project listed it as one of the 10 major diseases that urgently need drug research and development in clinical practice.

Tumor treatments include surgery, radiation and chemotherapy. The outstanding problems of the current clinical application of anti-tumor drugs are low effective rate, poor selectivity (high toxicity) and insensitivity to drug-resistant tumors. In addition, recurrence and metastasis are also difficulties in tumor treatment. Therefore, it is a major research topic for pharmaceutical workers to research anti-tumor drugs with good selectivity, high efficacy, clear targets, and no side effects on non-target organs.

Natural products are of great significance for the discovery, design and synthesis of new drugs, and are also an important source of biologically active substances and innovative drugs. According to literature reports: Since 1940, 42% of the anticancer chemotherapy drugs marketed in the world are directly derived from natural products, such as: Taxol, Docetaxel, Vinorelbine, Hydroxycamptothecin (camptothecin), artemisinin (Artemisinin), etc., 30% are derivatives or analogs of natural products. The advantage of developing natural products as drug sources is that biosynthesized natural product small molecule SMNPs have better biocompatibility with enzymes and receptors, and are more likely or suitable to become lead compounds for therapeutic drugs. my country's traditional Chinese medicine has a long history and is extensive and profound. It is a representative of Chinese traditional culture and a treasure of traditional culture. Therefore, searching for lead compounds with anti-tumor activity from traditional Chinese medicines and medicinal plants, studying their mechanism of action, and developing anti-tumor therapeutic drugs are current research hotspots.

Garcinia bracteata (Garcinia bracteata) is a plant of the genus Garcinia of the family Garcinia, and it is mainly distributed in the south of Yunnan to the southeast and south of Guangxi at present. At present, domestic research on the chemical components and pharmacological effects of Garcinia bracteata has been reported at home and abroad. not much.

Contents of the invention

In view of the above-mentioned problems in the prior art, the object of the present invention is to provide an anti-tumor pharmaceutical application of xanthenone compound A extracted and separated from Garcinia saccharomyces.

Xanthanone compound A of the present invention has a chemical structure shown in formula I: The chemical name is: 12R-isobractatin; the pharmaceutical use refers to the xanthanone compound A or its hydrate, pharmaceutically acceptable salt, tautomer, stereoisomer, precursor At least one of the compounds is used as an active ingredient to prepare antitumor drugs.

As a preferred solution, the antitumor drug is a drug for treating any one or two or more tumors in cervical cancer, prostate cancer, lung adenocarcinoma, and intestinal cancer.

The xanthenone compound A can be obtained by chemical synthesis or extracted from plants; as a preferred embodiment, the xanthenone compound A is obtained by extracting and separating from Gambogia spp.

In one embodiment, the method for extracting and isolating the described xanthones compound A from Garcinia macrophylla comprises the following steps:

a) Percolating and extracting the leaves of Garcinia cambogia with 95wt% ethanol until the percolating liquid is basically colorless, concentrating the percolating liquid under reduced pressure to obtain the extract;

b) suspending the extract obtained in step a) in water, and extracting with petroleum ether, to obtain the petroleum ether extraction part of the Garcinia cambogia extract;

c) separating the petroleum ether extraction part of the Garcinia cambogia extract obtained in step b) through a silica gel column, and sequentially gradient elution with a mixed solution formed of petroleum ether and ethyl acetate according to a volume ratio of 100:0 to 20:80, After each eluent was evaporated under reduced pressure, it was analyzed by TLC thin layer, and the eluents showing the same main point on the TLC thin layer plate were combined to obtain nineteen components according to the order of elution;

d) separating the fourth component obtained in step c) by high-speed countercurrent chromatography, collecting fractions containing the target compound, and concentrating under reduced pressure to obtain a crude product;

e) Purify the obtained crude product by preparative liquid chromatography to obtain the xanthenone compound A.

As a preferred version, in step d), during high-speed countercurrent chromatography purification, use n-hexane: ethyl acetate: 95wt% ethanol: water according to (6.5-7.5): (2.5-3.5): (6.5-8.5): (1.5- 3.5) The mixed solution formed by the volume ratio is used as the mobile phase for separation, and the ultraviolet detector is monitored at a wavelength of 340nm to collect the target fraction.

As a further preferred solution, a mixed solution formed by n-hexane: ethyl acetate: 95 wt% ethanol: water in a volume ratio of 7:3:7:3 is used as the mobile phase for separation.

As a preferred version, in step e), when purifying with preparative liquid chromatography, acetonitrile and water are used as the eluent to carry out isocratic elution according to the volume ratio of 60:40 mixed solution, and the ultraviolet detector is carried out at a wavelength of 340nm Monitor and collect target fractions.

The drug of the present invention can be administered to patients by various routes of administration, including but not limited to oral, transdermal, intramuscular, subcutaneous and intravenous injection.

The dosage form of the medicine of the present invention is not limited, as long as it is the dosage form that can make the active ingredient reach the body effectively, including: tablet, sugar-coated tablet, film-coated tablet, enteric-coated tablet, capsule, hard capsule medicines, soft capsules, oral liquids, buccal preparations, granules, granules, pills, powders, ointments, elixirs, suspensions, powders, solutions, injections, suppositories, ointments, plasters, creams, sprays Agents, drops, patches, etc.; preferred oral dosage forms, such as: capsules, tablets, oral liquids, granules, pills, powders, elixirs, ointments, etc.

The medicine of the present invention can be used alone or in the form of a pharmaceutical composition. Said pharmaceutical composition refers to a medicine that, in addition to the main active ingredient, can also contain a small amount of ingredients that do not affect the active ingredient. Secondary components and/or pharmaceutically acceptable carriers and necessary adjuvants for various preparations, etc. For example, when the drug is in an oral dosage form, it may contain commonly used excipients, such as binders, fillers, diluents, tableting agents, lubricants, disintegrating agents, coloring agents, flavoring agents and wetting agents, as necessary Tablets may be coated when necessary. Suitable fillers include cellulose, mannitol, lactose and other similar fillers; suitable disintegrants include starch, polyvinylpyrrolidone and starch derivatives, such as sodium starch glycolate; suitable lubricants include, for example, starch magnesium stearate; suitable pharmaceutically acceptable wetting agents include sodium lauryl sulfate.

The effective administered dose of the active ingredients of the present invention may vary with the drug used, the mode of administration and the severity of the disease to be treated.

Definitions of terms in the present invention are as follows:

The term "pharmaceutically acceptable salt" refers to the combination of a positively charged moiety (e.g., amine group) and an oppositely charged negatively charged moiety (e.g., trifluoroacetic acid, hydrochloric acid, hydrobromic acid, phosphoric acid) of the compound. , nitric acid, sulfuric acid, formic acid, acetic acid, propionic acid, succinic acid, 1,5-naphthalene disulfonic acid, subacid, oxalic acid, tartaric acid, lactic acid, salicylic acid, benzoic acid, valeric acid, diethylacetic acid, propane Diacid, succinic acid, fumaric acid, pimelic acid, adipic acid, maleic acid, malic acid, sulfamic acid, phenylpropionic acid, gluconic acid, ascorbic acid, niacin, isonicotinic acid, methanesulfonic acid, p-toluenesulfonic acid, citric acid, amino acids), or salts formed from a negatively charged moiety (eg, carboxyl group) and a positive charge (eg, sodium, potassium, calcium, magnesium) of a compound. The term "pharmaceutically acceptable" refers to substances that are suitable for human use without excessive adverse side effects (such as toxicity, irritation and allergy), that is, substances that have a reasonable benefit/risk ratio.

The term "tautomer" refers to isomers of functional groups resulting from the rapid movement of an atom in a molecule between two positions, for example: enol and the corresponding ketone.

The term "stereoisomer" refers to isomers produced by different arrangements of atoms in the molecule in space, for example: cis-trans isomers, enantiomers, conformational isomers and the like.

The term "precursor compound" refers to a compound that is inactive in vitro, but can be converted into the active ingredient of the present invention through metabolism or chemical reaction in vivo, thereby exerting its pharmacological effect.

Compared with the prior art, the present invention has the following significant beneficial effects:

The research results of the present invention show that the xanthanone compound A of the present invention: 12R-isobractatin has the effect on human prostate cancer cell PC-3, human cervical cancer cell HeLa, human non-small cell lung cancer cell A549 and human colon cancer cell HT -29 has strong cytotoxicity, IC50 is below 10μM, can significantly inhibit the proliferation of human prostate cancer cell PC-3, human cervical cancer cell HeLa, human non-small cell lung cancer cell A549 and human colon cancer cell HT-29 , indicating that the xanthanone compound A of the present invention: 12R-isobractatin has the effect of inhibiting the proliferation of tumor cells, and is expected to be used as an active ingredient in the preparation of antitumor drugs, and has a medicinal prospect.

Detailed ways

Below in conjunction with specific embodiment, further illustrate the present invention. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. For the experimental methods without specific conditions indicated in the following examples, usually follow the conventional conditions or the conditions suggested by the manufacturer. Percentages and parts are by weight unless otherwise indicated.

In the following implementation, Garcinia spp. was collected from Guangxi Province in October 2012. The plant was identified by researcher Song Zhijun of Guangxi Institute of Medicinal Botany, and the plant sample was preserved in the Innovative Chinese Medicine Laboratory of Shanghai University of Traditional Chinese Medicine;

Silica gel column chromatography selects Qingdao Ocean Chemical Co., Ltd. column chromatography silica gel (200-300 mesh);

TLC thin-layer analysis uses Yantai Jiangyou Silica Gel Development Co., Ltd. HSGF254 thin-layer chromatography silica gel plate;

Xbridge Prep C18OBD column (19×250mm, 5μm) was selected as the preparative liquid phase column.

Embodiment 1: Preparation of 12R-isobractatin

a) 7.6kg of Garcinia cambogia leaves were extracted by percolation with 95wt% ethanol until the percolation liquid was basically colorless, and the percolation liquid was concentrated under reduced pressure to obtain the extract (2.6 kg);

b) suspending the extract obtained in step a) in water, and extracting with petroleum ether to obtain the petroleum ether extraction fraction (700 g) of the Garcinia spp. extract;

c) separating the petroleum ether extraction part of the Garcinia cambogia extract obtained in step b) through a silica gel column, and sequentially gradient elution with a mixed solution formed of petroleum ether and ethyl acetate according to a volume ratio of 100:0 to 20:80, After each eluent was evaporated under reduced pressure, it was analyzed by TLC thin layer, and the eluents showing the same main point on the TLC thin layer plate were combined to obtain nineteen components according to the order of elution;

d) The fourth component obtained in step c) is separated by high-speed countercurrent chromatography, and the mixed solution formed by n-hexane: ethyl acetate: 95wt% ethanol: water according to the volume ratio of 7:3:7:3 is used as the flow Phase separation, rotation speed: 850rpm, flow rate: 2mL/min, the ultraviolet detector is monitored at a wavelength of 340nm, the target fractions containing 12R-isobractatin are collected, combined, and concentrated under reduced pressure to obtain a crude product;

e) The obtained crude product is purified by preparative liquid chromatography, and isocratic elution is carried out with acetonitrile and water according to the volume ratio of 60:40 mixed solution as eluent, and the mixed solution may contain 0.1vol% formic acid, and the flow rate is 15mL /min, monitored by an ultraviolet detector at a wavelength of 340 nm, the xanthone compound A: 12R-isobractatin (HPLC purity 98%) was obtained.

After testing and analysis, the molecular formula is C ₂₈ H ₃₂ O ₆ ; ESI-MS: m/z[M+1] ⁺ =465; ¹ HNMR (600MHz, DMSO-d ₆ ) and ¹³ CNMR (151MHz, DMSO-d ₆ ) are shown in Table 1.

Table 1 ¹ HNMR, ¹³ CNMR data of 12R-isobractatin

Example 2: 12R-isobractatin inhibits human tumor cell proliferation test

2.1 Experimental materials

The xanthenone compound A: 12R-isobractatin is extracted from Gambogia japonica with a purity of 98%; human prostate cancer cell PC-3 and human cervical cancer cell HeLa are purchased from ATCC Company of the United States; Lung cancer cell A549 was donated by Professor Lin Ge of the Chinese University of Hong Kong; human colon cancer cell HT-29 was donated by Professor Xu Naihan of Tsinghua University; human normal prostate stromal immortalized cell WPMY-1 was purchased from the Cell Bank of the Chinese Academy of Sciences; RPMI1640 was purchased from fetal Bovine serum, penicillin and streptomycin were purchased from Invitrogen Company of the United States, Taxol was purchased from Tao Su Company of the United States; propidium iodide (PI) and RNase A were purchased from Sigma Company of the United States.

2.2 Test method

Human non-small cell lung cancer cell A549 and human cervical cancer cell HeLa, with DMEM medium containing 10% fetal bovine serum, 100U/ml penicillin and 100μg/ml streptomycin; human prostate cancer cell PC-3, human colon cancer Cell HT-29 was cultured with RPMI1640 medium containing 10% fetal bovine serum, 100 U/ml penicillin and 100 μg/ml streptomycin in an incubator at 37°C, 5% CO ₂ and saturated humidity, with 0.25% After trypsinization and passage, the cells in the logarithmic growth phase were used for experiments.

Human prostate cancer cell PC-3, human non-small cell lung cancer cell A549, human cervical cancer cell HeLa, and human colon cancer cell HT-29 (3*104 cells/ml) were inoculated in a 96-well plate. Add compound 12R-isobractatin with final concentration gradients of 1.25 μM, 2.5 μM, 5 μM and 10 μM respectively in 96-well plate, 5-FU and Etoposide as positive drug control group, set blank reagent wells, normal control group, after culturing for 72 hours, Add 10ul CCK-8 to each well and incubate in the dark for 1-4h for staining. Use a microplate reader to detect the absorbance value at 450nm, and calculate the growth inhibition rate of different concentrations, growth inhibition rate%=(average value of control group-average value of treatment group)/average value of control×100%. When the cell growth inhibition rate reaches 50%, the concentration of the compound is the IC50 value. The experimental data are represented by mean ± standard error (xˉ±s), and analyzed using SPSS16.0 statistical software.

2.3 Experimental results

Table 2 shows the IC ₅₀ of the xanthanone compound A: 12R-isobractatin acting on various human tumor cells for 72 hours.

Table 2 Xanthones Compound A: IC50/μM (xˉ±s) value of 12R-isobractatin inhibiting tumor cell growth

It can be seen from Table 2 that the xanthenone compound A of the present invention: 12R-isobractatin has strong cytotoxicity, IC50 is below 10 μM, and can obviously inhibit human prostate cancer cell PC-3 and human non-small cell lung cancer cell A549. , the proliferation of human cervical cancer cells HeLa and human colon cancer cells HT-29, it can be seen that the xanthenone compound A of the present invention: 12R-isobractatin can inhibit the proliferation of tumor cells in a concentration-dependent manner, and has potential anti-tumor effects , is expected to be used as an active ingredient in the preparation of antitumor drugs, and has medicinal prospects.

Finally, it should be pointed out that the above are only some preferred embodiments of the present invention, and should not be interpreted as limiting the protection scope of the present invention. Those skilled in the art can make some non-essential improvements and adjustments based on the above contents of the present invention. All belong to the protection scope of the present invention.

Claims (6)

1. a kind of medicinal usage of xanthones compound A, the xanthones compound A has chemical constitution shown in Formulas I：Chemistry is entitled：12R-isobractatin；It is characterized in that：The medicinal usage refers to described In xanthones compound A or its hydrate, pharmaceutically acceptable salt, tautomer, stereoisomer, precursor compound At least one be used to prepare antitumor drug as active constituent.

2. medicinal usage according to claim 1, it is characterised in that：The antitumor drug is treatment cervix cancer, preceding The drug of any one or two or more tumours in row gland cancer, adenocarcinoma of lung, intestinal cancer.

3. medicinal usage according to claim 1, it is characterised in that：The xanthones compound A is from big luxuriant rattan It is extracted in Huang isolated.

4. medicinal usage according to claim 3, it is characterised in that：The xanthones described from big luxuriant gamboge extraction separation The method of compound A includes the following steps：

A) big luxuriant gamboge leaf is extracted substantially colorless to percolate with 95wt% ethanol percolations, percolate is concentrated under reduced pressure, must extract Object；

B) extract obtained by step a) is suspended in water, with petroleum ether extraction, obtains the petroleum ether of big luxuriant Resina garciniae extract Extract position；

C) by the petroleum ether extraction position of big luxuriant Resina garciniae extract obtained by step b) by silica gel post separation, with petroleum ether and acetic acid Ethyl ester is according to 100：0~20：Gradient elution, every part of eluent pass through the mixed solution that 80 volume ratio is formed after being evaporated under reduced pressure successively TLC tlc analysis will show that the eluent of identical main point merges, 19 components obtained by eluting order on TLC lamellaes；

D) the obtained in step c) the 4th component is detached with high speed adverse current chromatogram, collects the fraction containing target compound, It is concentrated under reduced pressure, obtains crude product；

E) purify gained crude product preparative liquid chromatography to get to the xanthones compound A.

5. medicinal usage according to claim 4, it is characterised in that：In step d), when high speed adverse current chromatogram purifies, with just Hexane：Ethyl acetate：95wt% ethyl alcohol：Water is according to (6.5-7.5)：(2.5-3.5)：(6.5-8.5)：The volume of (1.5-3.5) Mixed solution than formation is that mobile phase is detached, and UV detector is monitored under 340nm wavelength, collects target stream Part.

6. medicinal usage according to claim 4, it is characterised in that：In step e), when being purified with preparative liquid chromatography, Use acetonitrile and water according to volume ratio for 60:40 mixed solution carries out isocratic elution as eluent, and UV detector exists It is monitored under 340nm wavelength, collects target fraction.