CN101781353A

CN101781353A - 20(S)-ginsenoside Rh2 derivatives for regulating and controlling ER Alpha/ER Beta-TNF Alpha channel as well as preparation and anti-tumor application thereof

Info

Publication number: CN101781353A
Application number: CN 201010118460
Authority: CN
Inventors: 罗志勇; 刘苏友; 黄景嘉; 张建亭
Original assignee: Central South University
Current assignee: Central South University
Priority date: 2010-03-05
Filing date: 2010-03-05
Publication date: 2010-07-21
Anticipated expiration: 2030-03-05
Also published as: CN101781353B

Abstract

The invention relates to an ER Alpha/ER Beta-TNF Alpha channel which regulates and controls formation, growth and apoptosis processes of tumor cells and can be taken as a signal probe and a new medicine target spot. The invention relates to the preparation and the application of 20(S)-ginsenoside Rh2 derivatives with anti-tumor activity. Medicines containing the 20(S)-ginsenoside Rh2 derivatives and salts thereof, which is provided in the invention, are mainly used for treating tumors and other diseases; and compared with 20(S)-ginsenoside-Rh2 and 20(S)-protopanoxadiol, the medicines have stronger anti-tumor activity. A structural general formula (I) and a structural general formula (II) are as shown in the specification.

Description

20(S) -ginsenoside Rh2 derivative for regulating ER alpha/ER beta-TNF alpha pathway, preparation and anti-tumor application

The technical field is as follows:

the invention relates to 20(S) -ginsenoside Rh2 derivatives.

The invention also relates to a preparation method of the derivative;

the invention also relates to the application of the compound in preparing anti-tumor drugs.

Background art:

a medicinal plant ginseng Panax ginseng C.A. Meyer root is a traditional famous and precious Chinese medicinal material, and a tetracyclic triterpene dammarane micromolecule compound which is composed of ginsenoside (G) isoprene unit (five carbons) and is a main medicinal effective component of the ginseng, and consists of panaxadiol and panaxatriol saponin. At present, panaxadiol saponins mainly comprise G-Rb1, G-Rb2, G-Rc, G-Rd, G-Rg3, G-Rh2 and G-Rh 3. The panaxatriol saponin mainly comprises G-Re, G-Rf, G-Rg1, G-Rg2 and G-Rh 1. Wherein, the ginsenosides Rh2, Rg3, Compound K, 20(S) -protopanoxadiol and 20(R) -protopanaxadiol which belong to the panaxadiol group have very obvious antitumor activity after being proved by in vivo and in vitro experiments. Compared with the traditional chemotherapy drugs, the compound has wide and complex tumor inhibition effect and presents the characteristics of multiple parts, multiple links and multiple targets: can directly act on cancer cells, inhibit the growth of tumors by inducing the apoptosis of the cancer cells or induce the differentiation of the cancer cells to reverse the growth of the tumors; can inhibit tumor metastasis through acting on multiple links of tumor invasion; affecting the activity of cellular telomerase; can reverse the drug resistance of the tumor and improve the tumor inhibition activity of the chemotherapeutic drugs; can also enhance the resistance of the organism to diseases by influencing metabolism and regulating immunologic function, thereby inhibiting the growth of tumors; it also can affect cell junction communication or inhibit the activity of enzyme to antagonize the action of carcinogen, and has chemical anticancer effect. The comparison of the tumor inhibition effect of various ginsenosides shows that the ginsenoside Rh2 has the strongest capability of inhibiting the proliferation of cancer cells, and the ginsenoside Rg3 times.

A large number of in vivo and in vitro experiments prove that the ginsenoside Rh2(ginsenoside Rh2, G-Rh2) has extremely strong antitumor pharmacological activity, and the antitumor activity of the 20(S) -ginsenoside Rh2 is stronger than that of the 20(R) -ginsenoside Rh 2.

Chemical structure of 20(S) ginsenoside Rh2

G-Rh2 shows obvious anticancer activity on the aspects of inhibiting the proliferation, inducing the differentiation and/or the apoptosis of tumor cells of a plurality of tumor tissue cells such as breast cancer, ovarian cancer, cervical cancer, leukemia, lung cancer, liver cancer, nerve cells, glioma, prostatic cancer, melanoma and the like. The results show that G-Rh2 has a non-organ-specific broad-spectrum anti-tumor effect. Because G-Rh2 is the natural active component of ginseng, the toxicity is low, the molecular weight is low, and the synergistic effect exists with other anti-tumor chemotherapeutic drugs. Therefore, the compound is expected to become a clinical broad-spectrum anti-tumor therapeutic drug.

The antitumor effect of G-Rh2 is finally manifested by cell cycle arrest and apoptosis. It has been found that positive and negative regulators involved in cell cycle regulation intervene. In most tumor cells treated with G-Rh2, expression of cyclinD1, cyclinD3, cyclinE and cdk6 was down-regulated, activity of cyclinE/cdk2 was reduced, and cyclin P27^kip、P21^WAF1/CIP1、P16^INK4aExpression is up-regulated. The G-Rh 2-induced apoptosis was independent of bcl-2 and involved in the execution of apoptosis, Caspase 3. Caspase3 was produced by cleaving PRPP, P21^WAF1/CIP1And may mediate apoptosis of cells in the presence of PKC δ. When G-Rh2 induces apoptosis of lung adenocarcinoma A549 cells, expression of DR4 is up-regulated.

At present, the research on the antitumor effect of G-Rh2 is advanced, but the research on the antitumor mechanism of G-Rh2 is mostly limited to the study on the relationship between a certain known molecule and/or signal transduction pathway and the anticancer effect in the downstream, mainly stays on the observation of the phenomenon and the detection of individual indexes, but lacks of comprehensive understanding on the anticancer regulation network of G-Rh 2.

Estrogen (estrogen) is a key regulator that regulates the growth and differentiation of a wide range of target tissues, such as reproductive, mammary, central nervous and skeletal systems. The dammarane type ginsenoside structure including G-Rh2 is similar to steroid, and has a steroid skeleton composed of four trans-rings, and C-20 is connected with a longer side chain. Because the ginsenoside has a structure similar to that of the steroid, the ginsenoside also shows the activity of some steroids. Ginsenoside has structural commonality of steroid skeleton and C-20 side chain, and different ginsenosides have different structural individuality. Kovalchuk confirms G-Rh2 utensilHas weak phytoestrogen activity, can be combined with estrogen receptor (ER alpha), and has activity close to that of estradiol (17 beta-estradiol, E2) IC₅₀30% (Kovalchuk SN, Kozhemyako VB, Atopkina LN, et al, Escherichia coli activity of tripentene glycosides in yeast two-hybrid assay, journal of Steroid biochemistry)&Molecular Biology，2006；101：226-231)。

Estrogen signaling is mediated through Estrogen Receptors (ERs). Members of the human estrogen receptor superfamily (also found in part of the plasma membrane of tissues) include two subtypes, era and ER β, which consist of 595 and 530 amino acids encoded by two different genes located on chromosomes 6 and 14, respectively. ERs comprise 6 domains (A-F) and constitute 4 major functional domains. Transcriptional activation of ERs is mediated primarily by two activation domains, AF1 and AF2, including ligand-induced allosteric AF2 domain (AF1 is ligand-independent) recruitment of coactivators (cocavastors)

With co-repressors (corepressors) and interacting therewith, the activated ERs can bind directly to the ERE element of the target gene, or can bind to the Sp1 and AP1DNA binding sites via the ERs-Sp1 and ERs-AP1 complexes, thereby activating or inhibiting the expression of the target gene. Recent studies have shown that ER α and ER β have their unique adverse effects in estrogen-dependent tumors. ER alpha can promote tumor proliferation, while ER beta has the effect of inhibiting tumor proliferation. For example, in breast cancer, ER β expression is lost in most breast cancers due to the high methylation of the ER β gene promoter. ER β is thus considered to be a potential tumor suppressor gene (Heldring N, Pike A, Andersson S, equivalent. expression receptors: how do the signal and what are the target of our microorganism. physiol Rev.2007 Jul; 87 (3): 905-31; Safe S, Kim K. Non-classical genetic Estimator (ER)/specific genetic protein and ER/activating protein-1 signaling pathway. J. Mol Endocrinol.2008 Nov; 41 (5): 263-75).

TNF α is a pleiotropic cytokine that plays an important role in immunity and inflammation, as well as in controlling cell proliferation, differentiation, and apoptosis. TNF alpha with high concentration is a tumor inhibition and apoptosis inducing factor, and can activate three signal paths of Caspase protease, JNK and transcription factor NF-kB by combining with a cell membrane specific receptor TNFR, so as to realize the biological functions of apoptosis, cytotoxicity, antivirus, immunoregulation and the like (Aggarwal BB. signalling pathways of the TNFaperfemiance: a double-injected sweet. Nature reviews Immunology, 2003; 3: 745-75).

Ginsenoside consists of two parts of lipophilic steroid group and hydrophilic glycosyl group. Because of the difference of the number, the variety and the connection mode of the glycosyl, a plurality of ginsenosides with different chemical structures and biological activities can be formed. Researches find that the tumor inhibiting activity of the ginsenoside is influenced by mother nucleus and glycosyl, and the relationship rule between the tumor inhibiting activity structure and the effect is as follows: the law that the antitumor activity is influenced by glycosyl is as follows: monoglycoside (such as G-Rh2) > diglycoside > triglycoside > tetraglucoside; the strength rule that the tumor inhibition activity is influenced by C-20 configuration is as follows: 20(S) -ginsenoside Rh2(20(S) -G-Rh2) > 20(R) -ginsenoside Rh2(20(R) -G-Rh2), 20(S) -protopanaxadiol > 20(R) -protopanaxadiol.

Protopanoxadiol is the main pharmacophore and mother nucleus structure with bioactivity of the compound. However, G-Rh2 and protopanaxadiol have poor solubility and low bioavailability. Based on the structure of protopanaxadiol, Mashuanggang (CN1704427A, CN1778810A), Huimengzheng (CN1931177A), Loujin (CN1651451A) and the like disclose the preparation and the anti-tumor effect of the derivatives thereof.

Disclosure of Invention

The invention aims to provide a 20(S) -ginsenoside Rh2 derivative, the structure of which is a compound with the following general formula (I) and general formula (II)

General structural formula (I)

General structural formula (II)

Wherein:

r1 and R2 can be hydrogen, alkyl of 1-5 carbons;

the combination NR1R2 may also be a heterocyclic ring containing one nitrogen atom, such as tetrahydropyrrole, piperidine;

the combination NR1R2 can also be a heterocyclic ring containing more than one heteroatom, such as nitrogen-containing heterocyclic groups of piperazine, morpholine and the like;

a is CH2, N, aryl;

b is-C ═ O, -CH 2-;

a and B are combined to form-CH 2CH2-, -CH2C (O) -, -NC (O) -, -PhC (O) -, -PhNC (O) -,

q is- (CH2) n-, n is 0-3;

r3 is hydrogen, alkyl, alkanoyl, aroyl, -BAQNR1R2, etc.;

w is alkylene, aryl, or alkylene;

m is hydrogen atom, metal cation, amine cation, organic base, etc

The invention comprises 20(S) -G-Rh2 and pharmaceutically acceptable salts in a structural general formula (I) and a structural general formula (II), wherein:

the derivative with the structural general formula (I) can form pharmaceutically acceptable acid addition salts with inorganic acids or organic acids, wherein the salts are inorganic acids such as hydrochloride, sulfate, phosphate, acetate, nitrate, sulfonate, sulfite and the like and other organic acid salts capable of forming salts.

The derivative of the general structural formula (II) provided by the invention can form pharmaceutically acceptable base addition salts with inorganic bases or organic bases, wherein the inorganic bases comprise potassium hydroxide, potassium hydroxide and ammonia water, and the organic bases comprise aliphatic amines (such as triethylamine), alcamines (such as ethanolamine), amino acids (such as histidine) and amino sugars (such as glucose).

In the general structural formula (I) of the present invention, -BAQNR1R2, the following formula is preferred, but not limited thereto,

the H OS (O)2-M-CO in the general structural formula (II) of the invention is preferably, but not limited to, the following groups and sulfonate thereof

The invention relates to a method for preparing compounds in structural general formula (I) and structural general formula (II) and intermediates thereof in a second aspect, the invention provides a method for preparing compounds in general formula (I), which comprises the following synthetic routes and steps:

synthesis scheme 1

Synthesis scheme 2

Synthesis scheme 3

Synthesis scheme 4

20(S) -ginsenoside Rh2 carboxylate containing amine side chain can be prepared by reacting haloacyl chloride or haloanhydride with 20(S) -protopanaxadiol or its derivative in suitable organic solvent to obtain intermediate halocarboxylate, and reacting the halocarboxylate with amine under alkaline condition (see synthetic scheme 1 and synthetic scheme 2). The organic solvent can be chloroform, tetrahydrofuran, pyridine, dimethylformyl, benzene, toluene, dimethyl sulfoxide, dichloromethane, dioxane, etc., and the esterification reaction can be carried out in the presence of a suitable catalyst, such as triethylamine and dimethylaminopyridine. The reaction temperature may be 20-150 ℃.

The amine side chain-containing 20(S) -ginsenoside Rh2 ether derivative is a halogenated intermediate obtained by alkylation reaction of corresponding dihaloalkane and 20(S) -protopanoxadiol under the catalysis of alkali, wherein the halogenated intermediate is obtained by alcohol oxyalkylation at 12-position or substitution of 3, 12-position dialkanol oxyalkyl, and the halogenated intermediate is reacted with amine to obtain the amine side chain-containing 20(S) -ginsenoside Rh2 ether derivative (see synthetic scheme 3). The base can be sodium hydride, sodium amide, potassium tert-butoxide, sodium isopropoxide, sodium ethoxide, sodium methoxide, etc., the solvent for reaction can be dimethylformamide, benzene, toluene, dichloromethane, dimethyl sulfoxide, etc., and the reaction temperature can be 0-150'. The ether derivatives can also be prepared by phase transfer catalysis, such as tetrabutylammonium bromide, triethylbenzylammonium chloride, and the like.

The di (sulfo) carboxylic acid derivative of 20(S) -ginsenoside Rh2 is prepared by reacting the corresponding acid ester with 20(S) -protopanaxadiol in an organic solvent (see scheme 4). The reaction solvent may be anhydrous solvent, anhydrous dichloromethane, anhydrous tetrahydrofuran, anhydrous dimethylformamide, dimethyl sulfoxide, chloroform, etc. The reaction temperature is 0-150 deg.C, preferably 50-100 deg.C. The reaction can be carried out in the presence of a suitable catalyst, such as dimethylaminopyridine and triethylamine.

The salt of dicarboxylic (sulfonic) acid and inorganic acid derivative of 20(S) -ginsenoside Rh2 is obtained by adjusting pH with aqueous solution of hydroxide of corresponding ion, and precipitating with organic solvent. For example, the pH is adjusted to neutral with sodium hydroxide, ammonia and then precipitated with acetone to give the corresponding salt.

The structural general formula (I) and the structural general formula (II) of the invention are that the parent structure of protopanaxadiol is modified, some new groups are introduced, especially side chains containing water-soluble polar groups such as amino substitution or side chains containing sulfonate are introduced to synthesize the 20(S) -G-Rh2 derivative, so that the solubility and the bioavailability of the 20(S) -G-Rh2 derivative are improved, and the antitumor activity of the compound is enhanced.

The invention discovers that the tumor necrosis factor TNF alpha is a key molecule which mediates the broad-spectrum anti-tumor effect of the 20(S) -G-Rh2 derivative with the structural general formula (I) and the structural general formula (II); the 20(S) -G-Rh2 derivative has the structural general formula (I) and the structural general formula (II) and realizes the broad-spectrum antitumor effect of the structural general formula (I) and the structural general formula (II) of the G-Rh2 and the derivative thereof by targeting and selectively acting estrogen receptor ER alpha/ER beta, not only antagonizing ER alpha expression, but also activating ER beta expression, and further activating and up-regulating a molecular regulation network for inducing tumor cell apoptosis by TNF alpha. The invention also relates to a synthesis of the derivative of the 20(S) -G-Rh2 derivative based on the ER alpha/ER beta-TNF alpha pathway and the structural general formula (I) and the structural general formula (II) and an application thereof in the anti-tumor field.

The compound with the structural general formula (I) and the structural general formula (II) is used as a small molecular compound probe of ER alpha/ER beta-TNF alpha pathway for tumor formation, growth and apoptosis, and the anti-tumor activity of the compound with the structural general formula (I) and the structural general formula (II) is

The invention relates to a modern biological method such as applying transcriptome and proteome high-throughput screening, and finds that tumor necrosis factor TNF alpha is a key molecule for mediating 20(S) -ginsenoside-Rh 2(20(S) -G-Rh2) and derivatives thereof to play a broad-spectrum anti-tumor role in a structural general formula (I) and a structural general formula (II); the 20(S) -G-Rh2 and the structural general formula (I) and the structural general formula (II) of the derivatives thereof are found to be a molecular regulation network which can antagonize ER alpha expression and activate ER beta expression by targeting selective action of estrogen receptors ER alpha/ER beta, further activate and up-regulate TNF alpha to induce tumor cell apoptosis, and realize broad-spectrum antitumor action of the G-Rh2 derivatives in the structural general formula (I) and the structural general formula (II). The invention also relates to a synthesis method of the small molecular probe 20(S) -G-Rh2 based on the ER alpha/ER beta-TNF alpha pathway and a derivative thereof, a general structural formula (I) and a structural formula (II) and application thereof in the anti-tumor field.

The invention takes a compound with a structural general formula (I) of a 20(S) -G-Rh2 derivative as an example, and deeply explains an antitumor molecular mechanism of the compound with the structural general formula (I) of the ER alpha/ER beta mediated 20(S) -G-Rh2 derivative for activating TNF alpha to induce apoptosis (see examples 4-5 for details).

The invention provides application of compounds with a structural general formula (I) and a structural general formula (II) in preparation of an anti-tumor agent.

The present invention provides a medicament containing the above 20(S) -G-Rh2 derivative and a salt thereof: the above derivatives and their salts are mixed with conventional pharmaceutical adjuvants to make into preparations. The preparation can be granule, capsule, tablet, injection, infusion or suppository. The medicine of the present invention may be used in treating tumor.

The compounds described in the general structural formula (I) and the general structural formula (II) can be used for treating the growth and the metastasis of various tumors, wherein the tumors mainly comprise: lung cancer, non-small cell lung cancer, liver cancer, pancreatic cancer, stomach cancer, bone cancer, esophageal cancer, breast cancer, prostate cancer, testicular cancer, colon cancer, ovarian cancer, skin cancer, cervical cancer, melanoma, squamous cell cancer, basal cell cancer, adenocarcinoma, sweat gland cancer, sebaceous gland cancer, papillary adenocarcinoma, cystic cancer, medullary cancer, bronchial cancer, osteocytic cancer, epithelial cancer, bile duct cancer, choriocarcinoma, embryonal cancer, seminoma, Wilms' cancer, glial cell cancer, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, vocal cord neuroma, meningioma, neuroblastoma, retinoblastoma, neurofibroma, fibrosarcoma, fibroadenoma, fibrochondroma, fibrocystic tumor, polycystic carcinoma, melanoma, cervical cancer, melanoma, cervical cancer, Fibroma, fibrosarcoma, myxosarcoma, mucinocytoma, mucinochondrocoma, mucinochondrocarcoma, mucinadenoma, mucinoblastoma, liposarcoma, lipoma, lipoadenoma, lipoblastoma, lipochondrosaoma, lipofibroma, lipohemangioma, mucinochinoma, chondrosarcoma, chondroma, chondromas, chordoma, chorioadenoma, chorioepithelioma, chorioblastoma, osteosarcoma, osteoblastoma, osteochondroma, angiosarcoma, angiolipoma, angiofibroma, angiolipoma, angioglioma, hemangioblastoma, angiofibroma, Angiolymphomas, angiolipoleiomyomas, angiomyolipomas, angiomyoneuromas, angiomyxomas, angioreticular endotheliomas, lymphangiosarcomas, lymphogranulomas, lymphangiomas, lymphomas, lymphomyxomas, lymphosarcomas, lymphangiofibromas, lymphoblastomas, lymphoepitheliomas, lymphoblastomas, endotheliomas, endothelioblastomas, synoviomas, synovial sarcomas, mesotheliomas, connective tissue tumors, ewing's tumors, leiomyomas, leiomyosarcomas, rhabdomyosarcomas, rhabdomyomyxomas, acute lymphoid leukemias, acute myeloid leukemias, chronic diseased cells, polycythemia, lymphomas, multiple myeloma.

The compounds described in general formula (I) and general formula (II) are useful in the treatment of various related diseases caused by the modulation of the era/ER β -TNF α pathway, mainly: tumor, rheumatic arthritis, angioneurotic senile dementia, etc.

The main advantages of the invention are:

introducing alkoxy, ester group, carbamate group and other groups containing substituted amino side chain into 3-position or 12-position of natural product 20(S) -protopanoxadiol to synthesize 20(S) -G-Rh2 derivative with high antitumor activity.

The 3-position or 12-position of the natural product 20(S) -protopanaxadiol is introduced with dibasic acid and pharmaceutically acceptable salt formed by neutralizing with alkali, and the synthesized 20(S) -G-Rh2 derivative has improved water solubility.

The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. The experimental procedures, in which specific conditions are not noted in the following examples, are generally carried out according to conventional conditions or according to conditions recommended by the manufacturers.

Drawings

FIG. 1 is a dose-response curve of 20(S) -protopanaxadiol-12 β -2' -N, N-dimethylacetate compound acting on breast cancer MCF-7 cell line;

FIG. 2 is a dose-response curve of the 20(S) -G-Rh2 action breast cancer cell line MCF-7 and the normal breast epithelial cell line MCF-10A 1;

FIG. 3 shows that 20(S) G-Rh2 derivative compound 2 and 20(S) -G-Rh2 modulate the ER α/ER β -TNF α signaling pathway.

Detailed Description

Example 1

Preparation of 20(S) -protopanaxadiol-12 beta-2' -chloroacetate.

Compound 1

100mg (0.0434mmol) of protopanaxadiol was placed in a 25ml dry single-necked flask, 10ml of anhydrous chloroform was added, and stirred until completely dissolved. To the above solution was added 70. mu.l (0.20mmol) of triethylamine, and after stirring to uniformity 25. mu.l (0.13mmol) of chloroacetyl chloride was added, a large amount of white mist was immediately generated. The intermediate solution is supplemented with 35X10-3ml of triethylamine and 15 mu l of chloroacetyl chloride. The reaction produced a small polar spot, and the reaction was the same as in the first experiment. After reacting for 48h at room temperature, the raw materials basically disappear. After the reaction was terminated, 10ml of water was added, and after stirring for 10min, 20ml of chloroform was added and extracted 3 times, the chloroform layers were combined, dried over anhydrous sodium sulfate, the solvent was removed by evaporation under reduced pressure, and the mixture was separated by column chromatography using dichloromethane to methanol at a ratio of 25: 1, whereby 95mg of 20(S) -G-Rh2 derivative 20(S) -protopanaxadiol-12 β -2' -chloroacetate was obtained. Delta (ppm, CHCl)₃)：5.15(t，1H，H-24)，4.95(m，1H，H-12)，4.07(d，2H，ClCH₂CO-)，3.21(dd，1H，H-3)，2.37(s，2H)，2.16(m，1H)，1.91-2.10(m，4H)，1.71(s，3H)，1.60-1.70(m，4H)，1.44-1.60(m，6H)，1.20-1.44(m，5H)，1.14(s，3H)，1.04-1.12(m，1H)，1.01(s，3H)，0.98(s，3H)，0.95(s，3H)，0.84-0.90(m，4H)，0.78(s，3H)，0.73(d，1H，H-5)

Example 2

Preparation of 20(S) -protopanaxadiol-12 beta-2' -N, N-dimethyl acetate

Compound 2

10mg (0.0186mmol) of the product of example 1 are placed in a 10ml dry single-neck flask, 2ml of anhydrous acetonitrile are added and stirred until completely dissolved. To the above solution was added anhydrous potassium carbonate (about 7mg, 0.0480mmol), and after stirring for 5min, dimethylamine hydrochloride (2 mg, 0.025 mmol) was addedl). After stirring at room temperature for 10min, the mixture was transferred to an oil bath and heated to 50 ℃. The reaction produced a large polar spot, which was visible after iodine fumigation. The raw materials have no hydrolysis phenomenon. The starting material substantially disappeared after about 2.5d of reaction. The reaction was stopped, cooled to room temperature and the solvent was removed by evaporation under reduced pressure. Adding 2ml water, dissolving the obtained solid, adding 5ml ethyl acetate, extracting for 3 times, sucking organic layers by a suction pipe, combining ethyl acetate layers, drying by anhydrous sodium sulfate, removing the solvent by decompression, and separating by a column chromatography with ethyl acetate and methanol being 8: 1 to obtain 27mg of 20(S) -G-Rh2 derivative 20(S) -protopanaxadiol-12 beta-2' -N, N-dimethyl acetate. 1H-NMR: delta (ppm, CHCl)₃)：5.15(t，1H，H-24)，4.78(m，1H，H-12)，4.24(d，2H，NCH₂CO-)，3.19(dd，1H，H-3)，2.34(s，6H，

)，2.15-2.25(m，1H)，1.95-2.15(m，4H)，1.81-1.95(m，2H)，1.72(s，3H)，1.60-1.69(m，4H)，1.38-1.60(m，6H)，1.16-1.38(m，5H)，1.13(s，3H)，1.04-1.10(m，1H)，1.01(s，3H)，0.98(s，3H)，0.95(s，3H)，0.86-0.94(m，1H)，0.85(s，3H)，0.78(s，3H)，0.73(d，1H，H-5)

Example 3

Preparation of 20(S) -protopanaxadiol-12 beta-o-sulfonic acid benzoate and ammonium salt thereof

Compound 3

100mg (0.0434mmol) of protopanaxadiol was placed in a 25ml dry single-necked flask, 10ml of anhydrous chloroform was added, and stirred until completely dissolved. Adding 360mg (0.20mmol) of 2-sulfobenzoic anhydride into the solution, reacting overnight at 50 ℃ under the protection of nitrogen, cooling to 0 ℃ by using an ice salt bath, adding a methanol ammonia water solution, stirring for 2 hours, evaporating the solvent, and performing column chromatography to obtain 6mg of 20(S) -G-Rh2 derivative 20(S) -protopanoxadiol-12 beta-o-sulfonate benzoate. 1H-NMR: δ (ppm, DMSO-d 6): 5.15(t, 1H, H-24), 4.85(m, 1H, H-12), 7.20-7.8(m, 4H, H-Ar)

Example 4

MTT (methyl thiazolyl tetrazolium) experiments show that 20(S) G-Rh2 derivative 20(S) -protopanaxadiol-12 beta-2' -N, N-dimethyl acetate and 20(S) G-Rh2 can inhibit proliferation of various tumor cell lines, such as breast cancer cell lines (MCF-7 and MDAMB-231), human acute promyelocytic leukemia cell line (HL-60), chronic myelocytic leukemia cell line (K562), lung cancer cell line (A549) and cervical cancer cell line (HeLa), have broad-spectrum antitumor effect, and IC is determined₅₀The value is obtained.

As shown in FIG. 1, 20(S) -G-Rh2 derivative 20(S) -protopanaxadiol-12 β -2' -N, N-dimethylacetate acted on IC of breast cancer cell line MCF-748 hr₅₀17.48. mu.M significantly below 20(S) -G-Rh2 IC₅₀44.61 μ M, i.e. 20(S) -protopanaxadiol-12 β -2' -N, N-dimethylacetate, showed significantly stronger antitumor activity than 20(S) -G-Rh2, which was 2.5 times as high as 20(S) -G-Rh2 (fig. 1 and 2). Meanwhile, the 20(S) -G-Rh2 derivative compound shown in the general formula (I) is found to have the characteristic of selectively killing tumor cells, and has lower cytotoxicity on normal cells such as a mammary epithelial cell line MCF-10A1 (figure 1 and figure 2).

Example 5

Western blot analysis showed 18. mu.M (IC)₅₀) The 20(S) G-Rh2 derivative 20(S) -protopanaxadiol-12 beta-2' -N, N-dimethyl acetate (FIG. 3A) and 20(S) G-Rh2 (FIG. 3B) down-regulate ER alpha expression of breast cancer MCF-7 cells in a concentration-dependent manner, up-regulate ER beta expression, and further up-regulate TNF alpha expression. The mechanism of 20(S) -protopanaxadiol-12 beta-2' -N, N-dimethylacetate consistent with the mechanism of 20(S) -G-Rh2 in regulating ER alpha/ER beta-TNF alpha signaling pathway to exert antitumor effect (FIG. 3A and FIG. 3B).

Claims

1. 20(S) -ginsenoside Rh2 derivatives with the following structural general formula (I) and structural general formula (II) and salts thereof form pharmaceutically acceptable acid addition salts with inorganic acids or organic acids, or form pharmaceutically acceptable base addition salts with inorganic bases or organic bases;

general structural formula (I)

General structural formula (II)

Wherein,

r1 and R2 are selected from hydrogen, 1-5 carbon alkyl;

or NR1R2 in combination is a heterocyclic ring containing one nitrogen atom,

or NR1R2 is in combination a nitrogen-containing heterocyclic group containing more than one heteroatom;

a is CH2, N, aryl;

b is-C ═ O, -CH 2-;

q is- (CH2) n-, n is 0-3;

r3 is hydrogen, alkyl, alkanoyl, aroyl, -BAQNR1R2, etc.;

w is alkylene, aryl, or alkylene;

m is hydrogen atom, metal cation, amine cation, organic base, etc.

2. In the general structural formula (I) according to claim 1, -BAQNR1R2 is selected from the following structures:

。

3. the MOS (O)2-M-CO of the general structural formula (II) according to claim 1, which is selected from the group consisting of the following groups and their corresponding sulfonates

。

4. A process for the preparation of the compounds of claims 1 and 2 which comprises

(1) Halogenated acyl chloride or halogenated anhydride reacts with 20(S) -protopanaxadiol in an organic solvent to obtain intermediate halogenated carboxylic ester, and then the halogenated carboxylic ester reacts with amine under the alkaline condition to obtain the intermediate halogenated carboxylic ester; the organic solvent is selected from chloroform, tetrahydrofuran, pyridine, dimethylformyl, benzene, toluene, dimethyl sulfoxide, dichloromethane and dioxane, and the catalyst for esterification is selected from triethylamine and dimethylaminopyridine; the reaction temperature is 20-150 ℃;

(2) the dihaloalkane and 20(S) -protopanaxadiol are subjected to alkylation reaction under the catalysis of alkali to obtain 12-position alcohol oxyalkylation or 3, 12-position dialkoxyl substituted halogenated intermediate, the halogenated intermediate is reacted with amine to obtain 20(S) -ginsenoside Rh2 ether derivatives of amino side chains, the alkali is selected from sodium hydride, sodium amide, potassium tert-butoxide, sodium isopropoxide, sodium ethoxide and sodium methoxide, the reaction solvent is selected from dimethylformamide, benzene, toluene, dichloromethane, dimethyl sulfoxide and the like, and the reaction temperature can be 0-150 ℃; (3) the carboxylic ester of 20(S) -protopanaxadiol containing sulfonic group is obtained by reacting dicarboxylic acid (anhydride) with 20(S) -protopanaxadiol in organic solvent, and the corresponding sulfonic acid salt can be obtained by neutralizing with organic or inorganic base.

5. Use of a compound according to claims 1 and 2 for the preparation of an antitumor medicament.