A dehydroepiandrosterone triazole derivative and its preparation method and application

Abstract

The present invention provides a dehydroepiandrosterone triazole derivative represented by formula (I), or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, and a preparation method thereof, and use thereof in preparing a drug for treating diseases related to anti-tumor activity.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical chemistry, and specifically relates to a dehydroepiandrosterone triazole derivative, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, and a preparation method thereof, and use thereof in preparing a drug for treating diseases related to anti-tumor activity. Background Art

[0002] Natural products are an important source of drug lead compounds, and their structural diversity, novelty and biological activity have always been the focus of attention. However, due to the cumbersome separation process or challenging total synthesis route, the acquisition of novel skeleton compounds with advantageous structures has been declining year by year in recent years. In order to meet the demand for diversity of natural products and their derivatives in drug research and development, how to start from easily available natural products, how to synthesize diversity can efficiently construct a natural product library with complex and diverse chemical structures for the screening and discovery of new lead compounds has become a new response strategy.

[0003] As a branch of steroidal chemistry that is constantly developing and innovating, the development of steroid-based drugs has been widely concerned for many years. The cyclopentane polyhydrophenanthrene nucleus is a common structural feature of steroidal drugs. Studies have found that splicing different heterocyclic pharmacophores on steroidal compounds with different steroidal nucleus structures may cause significant changes in their biological activity, even if there are slight changes in the cyclopentane polyhydrophenanthrene structure skeleton of the steroid. Studies have shown that some of these compounds have good inhibitory effects on tumor cell proliferation.

[0004]

[0005] Dehydroepiandrostrone (DHEA), chemical name: 3β-3-hydroxy-5-ene-17-sterone, DHEA, is the most abundant steroid substance in human blood circulation, and its content in the human body changes with age. DHEA has a wide range of physiological activities, can prevent cardiovascular, diabetes and other diseases, and is also used in the treatment of depression, improving immunity, anti-aging and anti-tumor, and it is also an important intermediate for the synthesis of various steroid hormone drugs. In recent years, the clinical application of DHEA is currently popular.

[0006] 1,2,3-triazole derivatives have multiple nitrogen atoms, which can easily form hydrogen bonds with hydrogen bond donors, form π-π interactions with aromatic rings, and produce hydrophobic and electrostatic interactions. Therefore, the structural characteristics of 1,2,3-triazole make it easy to bind to a variety of enzymes and receptors in the body, showing a broad spectrum of biological activities, such as antibacterial, antiviral, anti-tuberculosis, inhibition of HIV protease and anti-cancer. Although 1,2,3-triazole compounds have so many properties, there are relatively few triazole compounds in nature so far, and this important compound can only be obtained through artificial synthesis. Therefore, it is particularly important to obtain highly active triazole derivatives through chemical synthesis methods and to find anticancer lead compounds.

[0007] However, dehydroepiandrosterone itself has only two reactive sites, the 3-hydroxyl group and the 17-carbonyl group. Since the 3-hydroxyl group is chiral, its modification is very difficult. Therefore, the current synthetic research is mainly to obtain new derivatives by modifying its 17-carbonyl group and its α-position, which leads to a small number of new structural types of dehydroepiandrosterone; secondly, the number of highly active lead compounds is insufficient, the research on the anticancer activity of dehydroepiandrosterone derivatives started late, and the structural design of highly active dehydroepiandrosterone derivatives lacks pertinence and systematicness. In the process of anticancer activity evaluation, fewer types of cancer cells were screened, resulting in a lack of highly active lead compounds. Existing literature reports have not disclosed the modification of dehydroepiandrosterone with a heterocyclic ring containing triazole and the provision of a lead compound with better anticancer activity. Based on the existing disclosed structure, the present invention screened a series of novel and highly active structural compounds. Summary of the invention

[0008] The object of the present invention is to provide a dehydroepiandrosterone triazole derivative as shown in the structure of formula (I), specifically a compound as shown in formula (I), or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof:

[0009]

[0010] Wherein, A, B, and C are independently selected from C or N, and at least one of A, B, and C is selected from N;

[0011] R is selected from any one of hydrogen, halogen, -OH, substituted or unsubstituted -C1-6 alkyl, substituted or unsubstituted -C2-6 alkenyl, substituted or unsubstituted -C2-6 alkynyl, substituted or unsubstituted -C1-6 alkoxy, substituted or unsubstituted -C6-C10 aryl, substituted or unsubstituted -C6-C10 heteroaryl;

[0012] Wherein, the substituted -C1-6 alkyl, substituted -C2-6 alkenyl, substituted -C2-6 alkynyl, substituted -C1-6 alkoxy, substituted -C6-C10 aryl, substituted -C6-C10 aryl, substituted -C6-C10 heteroaryl is R 1 Substitute, the R 1 Any one selected from hydrogen, -C1-C6 alkyl, halogen, -C1-C6 alkoxy.

[0013] Preferably, the R is selected from substituted or unsubstituted -C1-6 alkyl, substituted or unsubstituted -C6-C10 aryl, wherein the substituted -C1-6 alkyl, substituted -C6-C10 aryl is R 1 Substitute, the R 1 Any one selected from hydrogen, -C1-C6 alkyl, halogen, -C1-C6 alkoxy.

[0014] More preferably, the -C6-C10 aryl is phenyl, the -C1-C6 alkoxy is methoxy or ethoxy, and the halogen is fluorine, chlorine or bromine.

[0015] As a preference,

[0016] A, B, C are independently selected from N; R is selected from Any one of .

[0017] More preferably, the compound described in formula I specifically comprises any one of the following:

[0018]

[0019] The present invention also provides a method for preparing the compound of formula (I), comprising the following preparation method:

[0020]

[0021] Step a: In an organic solvent, dehydroepiandrosterone compound 1 reacts with an azide compound under the action of triphenylphosphine and azodicarbonate, after the reaction is completed, a dilute alkali solution is added for quenching, and then an organic solvent is added for extraction and separation to obtain a compound 2 in which the hydroxyl group is replaced by azide;

[0022] Step b: Compound 2 and copper salt react with an acetylene compound in a polar alkaline solvent. After the reaction is completed, an organic solvent is added for extraction and separation to obtain a compound represented by formula (I), wherein A, B, C and substituent R in formula (I) are as defined above.

[0023] Preferably, the organic solvent in step a is selected from one or two of diethyl ether, tetrahydrofuran, 1,4-dioxane, methyl tert-butyl ether, acetonitrile, toluene or xylene; the azodicarbonate is diethyl azodicarbonate (DEAD) or diisopropyl azodicarbonate (DIAD); the azide compound is bis(P-nitrophenyl) azidophosphate, 15-azido-4,7,10,13-tetraoxypentadecanoic acid-N-succinimidyl ester, diphenyl phosphate azide or azide-heptaethylene glycol-pentafluorophenyl ester one of the above; in the step b: the acetylene compound is one of hexyne, phenylacetylene, p-methoxyphenylacetylene, 2-fluorophenylacetylene, 3-chlorophenylacetylene, 4-chlorophenylacetylene, 2-bromophenylacetylene or 3-bromophenylacetylene; the polar solvent is one of water-methanol, water-ethanol, water-isopropanol, water-n-butanol or water-tert-butanol; the copper salt is one of copper sulfate, copper acetate, copper chloride, copper nitrate or copper carbonate; the base is one of sodium carbonate, sodium bicarbonate, sodium sulfite, sodium bisulfite or sodium ascorbate.

[0024] The present invention also provides a pharmaceutical composition, which is a preparation prepared by using the compound, or its stereoisomer, or its pharmaceutically acceptable salt as a pharmaceutical active ingredient and adding pharmaceutically acceptable excipients.

[0025] The present invention also provides the use of the compound represented by formula (I), or its stereoisomer, or its pharmaceutically acceptable salt, or the pharmaceutical composition in the preparation of drugs for treating anti-tumor and / or anti-cancer diseases.

[0026] As further preferred, the anticancer is selected from human cervical cancer cells, human gastric cancer cells or human renal epithelial cells.

[0027] The following is an explanation and description of the terms of the present invention:

[0028] "Cancer" or "malignancy" refers to any of a variety of diseases characterized by uncontrolled abnormal proliferation of cells, the ability of affected cells to spread locally or through the bloodstream and lymphatic system to other parts of the body (i.e., metastasis), and any of a number of characteristic structural and / or molecular features. "Cancer cells" refer to cells in the early, intermediate, or advanced stages of multi-step tumor progression. Cancers include sarcomas, breast cancer, lung cancer, brain cancer, bone cancer, liver cancer, kidney cancer, colon cancer, and prostate cancer. In some embodiments, the compounds of Formula I are used to treat cancers selected from colon cancer, brain cancer, breast cancer, fibrosarcoma, human cervical cancer cells, human gastric cancer cells, and human renal epithelial cells and squamous cell carcinoma. In some embodiments, cancer is selected from human cervical cancer cells, human gastric cancer cells, and human renal epithelial cells.

[0029] The compounds and derivatives provided in the present invention can be named according to the IUPAC (International Union of Pure and Applied Chemistry) or CAS (Chemical Abstracts Service, Columbus, OH) nomenclature system.

[0030] Definitions of terms used in the present invention: Unless otherwise stated, the initial definitions provided for groups or terms in this document apply to the groups or terms throughout the specification; for terms that are not specifically defined in this document, the meaning that a person skilled in the art can give them should be given based on the disclosure and context.

[0031] "Substitution" refers to the replacement of a hydrogen atom in a molecule by another different atom or molecule.

[0032] The minimum and maximum carbon atom content of a hydrocarbon group is indicated by a prefix, for example, the prefix C1-b alkyl indicates any alkyl group containing from "a" to "b" carbon atoms. Thus, for example, "C1-4 alkyl" refers to an alkyl group containing 1 to 4 carbon atoms.

[0033] "Alkyl" refers to a saturated hydrocarbon chain having a specified number of member atoms. For example, C1-6 alkyl refers to an alkyl group having 1 to 6 member atoms, such as 1 to 4 member atoms. The alkyl group can be straight or branched. Representative branched alkyl groups have one, two or three branches. The alkyl group may be optionally substituted with one or more substituents as defined herein. Alkyl includes methyl, ethyl, propyl (n-propyl and isopropyl), butyl (n-butyl, isobutyl and tert-butyl), pentyl (n-pentyl, isopentyl and neopentyl) and hexyl. The alkyl group may also be part of other groups, such as C1-C6 alkoxy.

[0034] "Cycloalkyl", "cycloalkane" refers to a saturated or partially saturated cyclic group having carbon atoms and no ring heteroatoms and having a single ring or multiple rings (including fused, combined and). For polycyclic systems with aromatic and non-aromatic rings without ring heteroatoms, when the point of attachment is located at a non-aromatic carbon atom, the term "cycloalkyl" (e.g., 5,6,7,8,-tetrahydronaphthalene-5-yl) is applicable. The term "cycloalkyl" includes cycloalkenyl groups, such as cyclohexenyl. Examples of cycloalkyl groups include, for example, adamantyl, cyclopropyl, cyclobutyl, cyclohexyl, cyclopentyl, cyclooctyl, cyclopentenyl and cyclohexenyl. Examples of cycloalkyl groups including polybicycloalkyl ring systems are bicyclohexyl, bicyclopentyl, bicyclooctyl, etc.

[0035] "Alkenyl" refers to a straight or branched chain hydrocarbon group having 2 to 10 carbon atoms, and in some embodiments 2 to 6 carbon atoms or 2 to 4 carbon atoms, and having at least 1 site of vinyl unsaturation (>C=C<). For example, (Ca-Cb)alkenyl refers to an alkenyl group having a to b carbon atoms and is meant to include, for example, ethenyl, propenyl, isopropenyl, 1,3-butadienyl, and the like.

[0036] "Alkynyl" refers to a straight chain monovalent hydrocarbon radical or a branched monovalent hydrocarbon radical containing at least one triple bond. The term "alkynyl" is also intended to include those hydrocarbon groups having one triple bond and one double bond. For example, (C2-C6)alkynyl is intended to include ethynyl, propynyl, and the like.

[0037] "Halogen" is fluorine, chlorine, bromine or iodine.

[0038] "Halogen alkyl" refers to an alkyl group in which the hydrogen atoms may be replaced by one or more halogen atoms. For example, C1-4 halogen alkyl refers to an alkyl group containing 1 to 4 carbon atoms in which the hydrogen atoms are replaced by one or more halogen atoms.

[0039] "Heterocycle", "heterocycloalkyl" and "heterocycloalkane" refer to a saturated ring or a non-aromatic unsaturated ring containing at least one heteroatom; wherein the heteroatom refers to a nitrogen atom, an oxygen atom, or a sulfur atom;

[0040] "Aromatic heterocycle" refers to an aromatic unsaturated ring containing at least one heteroatom; wherein the heteroatom refers to a nitrogen atom, an oxygen atom, or a sulfur atom;

[0041] The compounds of the present invention may contain asymmetric centers or chiral centers and therefore exist as different stereoisomers. All stereoisomeric forms of the compounds of the present invention, including but not limited to, diastereomers, enantiomers, atropisomers, and mixtures thereof, such as racemic mixtures, form part of the present invention. Many organic compounds exist in optically active forms, i.e., they have the ability to rotate the plane of plane polarized light. When describing optically active compounds, the prefix D, L or R, S is used to indicate the absolute configuration of the chiral center of the molecule. The chemical structure of these stereoisomers is the same, but their stereostructure is different. Specific stereoisomers can be enantiomers, and mixtures of isomers are generally called enantiomeric mixtures. A 50:50 mixture of enantiomers is called a racemic mixture or racemate, which may result in no stereoselectivity or stereospecificity during chemical reactions. The terms "racemic mixture" and "racemate" refer to an equimolar mixture of two enantiomers that lacks optical activity.

[0042] The term "pharmaceutically acceptable" means that a carrier, vehicle, diluent, excipient, and / or formed salt is generally chemically or physically compatible with the other ingredients that constitute a pharmaceutical dosage form and physiologically compatible with the receptor.

[0043] The pharmaceutical composition of the present invention can be any reusable pharmaceutical preparation form, such as oral, injection, external use, etc., and the oral dosage form includes but is not limited to tablets, capsules, oral liquids, granules, pills, suspensions, injections selected from water injections and powder injections, and external preparations selected from patches and ointments. All preparations can be prepared according to conventional pharmaceutics techniques, such as using the compound of the present invention, or any one of its stereoisomers, or its pharmaceutically acceptable salt as a pharmaceutically active ingredient, adding a pharmaceutically acceptable carrier if necessary, to prepare the above-mentioned pharmaceutical dosage form suitable for taking, wherein the pharmaceutically active ingredient per unit dose can be 0.1 mg-1000 mg, such as each tablet containing 0.1 mg-1000 mg, preferably 5-500 mg, of the pharmaceutically active ingredient.

[0044] The terms "salt" and "pharmaceutically acceptable salt" refer to acidic and / or basic salts of the above-mentioned compounds or their stereoisomers, formed with inorganic and / or organic acids and bases, and also include zwitterionic salts (inner salts), and also include quaternary ammonium salts, such as alkylammonium salts. These salts can be obtained directly in the final separation and purification of the compound. It can also be obtained by mixing the above-mentioned compound, or its stereoisomer, with a certain amount of acid or base appropriately (e.g., equivalent). These salts may form a precipitate in the solution and be collected by filtering, or be recovered after solvent evaporation, or be obtained by freeze drying after reaction in an aqueous medium. The salt described in the present invention can be a hydrochloride, sulfate, citrate, benzenesulfonate, hydrobromide, hydrofluoride, phosphate, acetate, propionate, succinate, oxalate, malate, succinate, fumarate, maleate, tartrate or trifluoroacetate of the compound.

[0045] In certain embodiments, one or more compounds of the present invention may be used in combination with each other. It is also possible to select the compounds of the present invention and any other active agent for use in combination to prepare a drug or pharmaceutical composition for regulating cell function or treating a disease. If a group of compounds is used, these compounds may be administered to a subject simultaneously, separately or sequentially.

[0046] Obviously, according to the above contents of the present invention, in accordance with common technical knowledge and customary means in the art, without departing from the above basic technical ideas of the present invention, other various forms of modification, replacement or change may be made.

[0047] Beneficial technical effects of the present invention:

[0048] 1. The present invention modifies the 3-hydroxyl group of dehydroepiandrosterone through novel chemical synthesis methods such as functional group transformation and click reaction, thereby enriching the new structural types of dehydroepiandrosterone;

[0049] 2. The preparation method of the dehydroepiandrosterone 1,2,3-triazole derivative provided by the present invention is simple, the raw materials are cheap and readily available, the reaction conditions are mild, the post-treatment is simple, the yield is high, and the solvent can be recycled, which is conducive to industrial production;

[0050] 3. The present invention utilizes the principle of drug splicing to introduce a 1,2,3-triazole heterocyclic structural unit with strong anticancer activity into the dehydroepiandrosterone molecular skeleton, evaluates the inhibitory activity of the new derivatives on human cervical cancer cells (HeLa), human gastric cancer cells (HGC-27) and human renal epithelial cells (HEK-293T), and screens out highly active lead compounds, providing a research basis and lead compounds for the development of new anticancer drugs, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Various other advantages and benefits will become apparent to those of ordinary skill in the art by reading the detailed description of the preferred embodiments below. The accompanying drawings are only for the purpose of illustrating the preferred embodiments and are not to be considered as limiting the present invention. Moreover, the same reference numerals are used throughout the accompanying drawings to represent the same components. In the accompanying drawings:

[0052] Figure 1 The H NMR spectrum of compound 2 is schematically shown;

[0053] Figure 2 The carbon NMR spectrum of compound 2 is schematically shown;

[0054] Figure 3 The H NMR spectrum of compound 3a is schematically shown;

[0055] Figure 4 The carbon NMR spectrum of compound 3a is schematically shown;

[0056] Figure 5 The H NMR spectrum of compound 3b is schematically shown;

[0057] Figure 6 The carbon NMR spectrum of compound 3b is schematically shown;

[0058] Figure 7 The H NMR spectrum of compound 3c is schematically shown;

[0059] Figure 8 The carbon NMR spectrum of compound 3c is schematically shown;

[0060] Fig. 9 The H NMR spectrum of compound 3d is schematically shown;

[0061] Fig.10 The carbon NMR spectrum of compound 3d is schematically shown;

[0062] Fig.11 The H NMR spectrum of compound 3e is schematically shown;

[0063] Fig.12 The carbon NMR spectrum of compound 3e is schematically shown;

[0064] Fig.13 The H NMR spectrum of compound 3f is schematically shown;

[0065] Fig.14 The carbon NMR spectrum of compound 3f is schematically shown;

[0066] Fig.15 The H NMR spectrum of compound 3g is schematically shown;

[0067] Fig.16 The carbon NMR spectrum of compound 3g is schematically shown. DETAILED DESCRIPTION

[0068] The exemplary embodiments of the present disclosure will be described in more detail below with reference to the accompanying drawings. Although the exemplary embodiments of the present disclosure are shown in the accompanying drawings, it should be understood that the present disclosure can be implemented in various forms and should not be limited by the embodiments described herein. On the contrary, these embodiments are provided in order to enable a more thorough understanding of the present disclosure and to fully convey the scope of the present disclosure to those skilled in the art.

[0069] The structure of the compound of the present invention is determined by nuclear magnetic resonance (NMR) and mass spectrometry (MS). The NMR shift (δ) is given in units of 10-6 (ppm). The NMR is measured using a (Bruker Avance III 400 and Bruker Avance 300) nuclear magnetic spectrometer, and the measurement solvent is deuterated dimethyl sulfoxide (DMSO-d6), deuterated chloroform (CDCl3), deuterated methanol (CD3OD), and the internal standard is tetramethylsilane (TMS). The known starting materials of the present invention can be synthesized by methods known in the art or can be purchased from commercial reagent platforms.

[0070] Preparation of compound 2

[0071]

[0072] Take a 250mL round-bottom flask, weigh triphenylphosphine (1.44g, 5.5mmol), dissolve it in 35mL of tetrahydrofuran and stir to dissolve, then add diisopropyl azodicarboxylate (1.18mL, 6mmol), stir at 0℃ for 10min, then add dehydroepiandrosterone (1.44g, 5mmol), continue stirring for ten minutes, then slowly add diphenylphosphoazide (1.29mL, 6mmol), remove the ice bath, warm to room temperature, stir for 10h, and then detect by TLC. 0.1 mol / L sodium hydroxide was added for quenching, and the reaction solution was extracted with dichloromethane. The remaining aqueous phase was extracted with dichloromethane again, and the extraction was repeated three times. The organic phases were combined, washed with saturated brine, and then dried with anhydrous sodium sulfate to obtain a crude product 2. The crude product 3 was decompressed and concentrated, and column chromatography (silica gel column 300-400 mesh, eluent volume ratio of dichloromethane: petroleum ether = 1:1 for gradient elution) was performed to obtain compound 2 (1.16 g, 74.2%). Compound 2: Yield 74.2%; Characterization spectrum as shown Figure 1 , Figure 2 As shown; 1 H NMR(400MHz, CDCl3)δ5.43(d,J=4.8Hz,1H),3.89(s,1H),2.64–2.36(m,2H),2.22(d, J=15.0Hz,1H),2.11(dd,J=18.7,9.2Hz,2H),1.95(d,J=5.7Hz,1H),1.85(d,J=13.0H z,1H),1.81–1.73(m,2H),1.73–1.62(m,3H),1.53(dd,J=17.1,7.7Hz,2H),1.49–1.3 9(m,2H),1.36–1.22(m,3H),1.17(dd,J=11.1,4.2Hz,1H),1.03(s,3H),0.89(s,3H). 13 C NMR (100MHz, CDCl3) δ138.09,122.06,57.77,51.44,49.77,47.21,36.92,35.73,35.53,33.32,31.17,31.10,30.44,25.78,21.56,19.75,18.72 ,13.24; FT-IR(KBr)ν:3432.72,3139.27,2937.52,2832.03,1738.02,16 23.67,1455.42,1246.83,1229.63,1056.05,1015.96,800.84,776.42cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 19 H28 N3O{[M+H] +}314.2253, found 314.2155.

[0073] Synthesis of compounds 3a-3h

[0074]

[0075] Take two 100mL round-bottom flasks, weigh copper acetate (0.2mmol) and sodium ascorbate (0.4mmol) in the round-bottom flask, add 2mL of water and stir to dissolve, take compound 2 (1mmol) and acetylene compound (1mmol) in another flask, add 10mL of tert-butanol and stir to dissolve, combine the two systems, and react at room temperature. After the reaction is completed by TLC detection, the organic phase is extracted with ethyl acetate, and the remaining aqueous phase is extracted with ethyl acetate again, and the extraction is repeated three times, the organic phases are combined, washed with saturated brine, and dried with anhydrous sodium sulfate to obtain a crude product 3, and the crude product 3 is decompressed and concentrated, and compound 3 is obtained by column chromatography.

[0076] Example 1: Preparation of Compound 3a

[0077]

[0078] According to the same preparation method as in Example 1, hexyne was added to prepare compound 3a. Compound 3a: yield 84.1%; characterization spectrum as shown Figure 3 , Figure 4 As shown; 1 HNMR (400MHz, CDCl3) δ7.43 (s, 1H), 5.43 (d, J = 3.4Hz, 1H), 4.75 (s, 1H), 2.87 (d, J = 15.0Hz, 1H), 2 .58(t,J=7.6Hz,2H),2.48(d,J=15.6Hz,1H),2.35(dd,J=18.9,8.7Hz,1H),2.11(s,2H),2.00–1.9 1(m,1H),1.73(d,J=12.6Hz,1H),1.67–1.60(m,2H),1.60–1.51(m,4H),1.50–1.31(m,2H),1.29– 1.20(m,5H),1.17–1.12(m,2H),1.04(s,3H),0.96(dd,J=17.0,4.5Hz,1H),0.78(d,J=9.0Hz,6H); 13C NMR (100MHz, CDCl3) δ146.45,137.85,122.63,119.57,77.32,76.68,55.52,53.07,5 1.03,49.55,46.75,36.53,35.01,32.28,34.92,30.19,28.51,26.35,24.99,21.79,2 1.18,19.35,18.70,13.48,12.90; FT-IR(KBr)ν:3450.73,3143.47,2942.10,1737.68 ,1604.45,1567.17,1470.52,1454.84,1218.64,1069.86,1042.49,784.07,756.02cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 26 H 40 N3O{[M+H] +}410.3166, found410.3166.

[0079] Example 2: Preparation of Compound 3b

[0080]

[0081] According to the same preparation method as in Example 1, phenylacetylene was added to prepare compound 3b. Compound 3b: yield 72.3%; characterization spectrum as shown Figure 5 , Figure 6 As shown; 1 HNMR(400MHz, CDCl3)δ7.90(s,1H),7.73(d,J=7.3Hz,2H),7.40–7.11(m,3H),5.50(d,J=3.4Hz,1H), 4.87(s,1H),2.93(d,J=15.3Hz,1H),2.58(d,J=15.7Hz,1H),2.44–2.33(m,1H),2.20(s,1H),2.12(dd ,J=9.7,4.3Hz,2H),2.00(dd,J=19.3,9.5Hz,1H),1.89(s,1H),1.76(d,J=12.7Hz,1H),1.67(t,J=9. 4Hz,3H),1.56(d,J=2.2Hz,1H),1.47(d,J=12.2Hz,1H),1.23–1.14(m,5H),1.07(s,3H),0.82(s,3H); 13CNMR(100MHz,CDCl3)δ149.86,149.78,138.34,130.01,129.80,126.06,125.54,1 22.33,120.20,120.15,58.03,51.69,49.99,47.47,37.17,35.99,35.79,33.53,3 1.37,31.35,30.67,26.02,21.81,19.98,18.98,13.50; FT-IR(KBr)ν:3453.68,31 57.23,2832.03,1735.17,1623.67,1406.39,1297.27,1069.86,816.69,771.62cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 27 H 34 N3O{[M+H] +}416.2652, found416.2657.

[0082] Example 3: Preparation of Compound 3c

[0083]

[0084] According to the same preparation method as in Example 1, p-methoxyphenylacetylene was added to prepare compound 3c. Compound 3c: yield 82.4%; characterization spectrum as shown Figure 7 , Figure 8 As shown; 1 HNMR (400MHz, CDCl3) δ7.80 (s, 1H), 7.65 (d, J = 8.7Hz, 2H), 6.87 (d, J = 8.7Hz, 2H), 5.49(d,J=4.5Hz,1H),4.85(s,1H),3.76(s,3H),2.92(d,J=15.6Hz,1H),2.57(d,J =15.7Hz,1H),2.37(d,J=8.6Hz,1H),2.19(s,1H),2.04(s,1H),1.75(s,1H),1.66( dd,J=13.4,6.2Hz,3H),1.48(s,2H),1.30–1.12(m,5H),1.07(s,3H),0.82(s,3H). 13C NMR (100MHz, CDCl3) δ159.43,146.67,138.45,126.92,123.69,123.45,118.29,11 4.17,77.32,77.00,76.68,56.40,55.28,51.60,50.11,47.43,37.19,35.75,35.55 ,32.78,31.31,30.85,29.64,26.95,21.78,19.93,19.31,13.51; FT-IR(KBr)ν:34 54.49,3179.33,2943.14,1737.87,1467.54,1432.51,1219.48,1079.39,770.91cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 28 H 36 N3O2{[M+H] +}446.2802, found446.2807.

[0085] Example 4: Preparation of Compound 3d

[0086]

[0087] According to the same preparation method as in Example 1, 2-fluorophenylacetylene was added to prepare compound 3d. Compound 3d: yield 86.9%; characterization spectrum as shown Fig. 9 , Fig.10 As shown; 1 HNMR(400MHz, CDCl3)δ8.23–7.99(m,2H),7.19–6.79(m,3H),5.42(s,1H),4.80(s,1H),2.83(s ,1H),2.50(d,J=15.6Hz,1H),2.26(d,J=8.6Hz,1H),2.09(s,1H),2.06–1.97(m,2H),1.93–1.86 (m,1H),1.83–1.74(m,1H),1.65(d,J=12.8Hz,1H),1.56(t,J=13.6Hz,3H),1.43(s,1H),1.38( s,1H),1.28(dd,J=19.3,8.7Hz,1H),1.19–1.03(m,3H),0.96(s,3H),0.90(s,1H),0.72(s,3H); 13C NMR (100MHz, CDCl3) δ159.76,157.30,139.43,139.40,137.57,128.46,128.37,127.02,126.9 9,124.00,123.97,123.19,122.10,121.96,118.48,118.35,115.17,114.96,55.95,50.92,49 .59,46.76,36.57,35.14,34.95,32.21,30.73,30.68,30.25,26.45,21.20,19.37,18.74,12. 93; FT-IR(KBr)ν:3415.53,2958.73,2868.35,1466.76,1261.08,1079.66,1025.27,771.25cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 27 H 33 FN3O{[M+H] +}434.2602, found434.2603.

[0088] Example 5: Preparation of Compound 3e

[0089]

[0090] According to the same preparation method as in Example 1, 3-chlorophenylacetylene was added to prepare compound 3e. Compound 3e: yield 91.8%; characterization spectrum as shown Fig.11 , Fig.12 As shown; 1 H NMR (400MHz, CDCl3) δ7.94(s,1H),7.72(d,J=8.5Hz,2H),7.36(d,J=8.5Hz,2H),5.57(d ,J=4.7Hz,1H),4.92(d,J=1.9Hz,1H),3.00(d,J=15.6Hz,1H),2.64(d,J=15.7Hz,1H),2. 45(dd,J=19.2,8.7Hz,1H),2.21(dd,J=23.2,12.5Hz,3H),2.06(dd,J=18.9,9.1Hz,1H) ,2.01–1.89(m,1H),1.87–1.37(m,8H),1.25(d,J=6.0Hz,3H),1.13(s,3H),0.88(s,3H); 13C NMR (100MHz, CDCl3) δ145.77,138.33,133.59,129.46,128.92,126.86,123 .56,119.10,56.56,51.57,50.10,47.40,37.19,35.73,35.50,32.75,31.2 8,31.23,30.83,26.89,21.89,21.77,19.92,19.28,13.49; FT-IR(KBr)ν:3 443.16,2942.51,1736.03,1496.77,1247.15,1219.64,1032.83,771.97cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 27 H 33 ClN3O{[M+H] +}450.2307, found450.2307.

[0091] Example 6: Preparation of Compound 3f

[0092]

[0093] According to the same preparation method as in Example 1, 2-chlorophenylacetylene was added to prepare compound 3f. Compound 3f: yield 89.7%; characterization spectrum as shown Fig.13 , Fig.14 As shown; 1 H NMR (400MHz, DMSO-d6) δ7.87(s,1H),7.66(d,J=8.5Hz,2H),7.30(d,J=8.5Hz,2H),5.50(d,J=4.7Hz,1 H),4.86(d,J=1.9Hz,1H),2.93(d,J=15.6Hz,1H),2.59(s,1H),2.39(dd,J=19.2,8.7Hz,1H),2.26–2. 09(m,3H),1.96(s,1H),1.93–1.84(m,1H),1.75(t,J=11.8Hz,1H),1.66(dd,J=13.2,6.3Hz,3H),1.62 –1.54(m,1H),1.52–1.33(m,2H),1.19(d,J=6.0Hz,3H),1.07(s,3H),1.03–0.95(m,1H),0.82(s,3H); 13C NMR(100MHz,DMSO-d6)δ145.45,138.01,133.27,129.14,128.60,126.54,123.25, 118.78,56.24,51.26,49.78,47.09,36.87,35.41,35.19,32.43,30.96,30.91,30. 51,29.31,26.57,21.58,21.45,19.61,18.96,13.17,0.66; FT-IR(KBr)ν:3451.26, 2936.21,1737.42,1618.02,1370.52,1262.97,1047.81,977.72,800.02,784.07cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 27 H 33 ClN3O{[M+H] +}450.2307, found450.2303.

[0094] Example 7: Preparation of Compound 3g

[0095]

[0096] According to the same preparation method as in Example 1, 2-bromophenylacetylene was added to prepare compound 3a. Compound 3g: yield 85.1%; characterization spectrum as shown Fig.15 , Fig.16 As shown; 1 H NMR (400MHz, CDCl3) δ8.54(d,J=1.9Hz,1H),8.07(dd,J=7.8,1.6Hz,1H),7.54(d,J=8.0Hz,1H),7.31(dd,J=10.8,4.2 Hz,1H),7.08(dd,J=10.5,4.7Hz,1H),5.51(s,1H),5.20(d,J=2.5Hz,1H),4.92(s,1H),4.03(d,J=7.1Hz,1H),2.93(d ,J=15.5Hz,1H),2.55(d,J=1.4Hz,1H),2.37(dd,J=19.2,8.7Hz,1H),2.20–2.07(m,3H),2.02(dd,J=17.5,7.8Hz,1H) ,1.95(d,J=2.8Hz,1H),1.74(dd,J=12.8,2.4Hz,1H),1.69–1.60(m,3H),1.21–1.12(m,5H),1.02(s,3H),0.81(s,3H);13 C NMR (100MHz, CDCl3) δ144.02,138.27,133.40,131.41,130.24,128.92,127. 60,123.78,123.02,120.89,56.33,51.61,50.13,47.31,37.09,35.63,35.48 ,32.47,31.17,30.85,27.15,21.81,21.67,19.82,19.22,14.05,13.43; FT- IR(KBr)ν:3449.27,2931.27,1736.11,1655.14,1219.35,1092.61,771.93cm -1 ;HR-MS(ESI-TOF)m / z:Calcd forC 27 H 33 BrN3O{[M+H] +}494.1802, found494.1801.

[0097] Example 8: Preparation of Compound 3h

[0098]

[0099] According to the same preparation method as in Example 1, 3-bromophenylacetylene was added to prepare compound 3h. Compound 3h: yield 96.3%; 1 H NMR (400MHz, CDCl3) δ8.03–7.77(m,2H),7.58(d,J=7.4Hz,1H),7.24(d,J=7.7Hz,1H),7.16–6.99(m,1H),5. 41(s,1H),4.73(s,1H),2.85(d,J=14.7Hz,1H),2.53(d,J=15.4Hz,1H),2.27(dd,J=18.9,7.9Hz,1H),2.15( d,J=13.5Hz,1H),2.02(dd,J=20.8,11.3Hz,2H),1.88(t,J=9.1Hz,1H),1.76(s,1H),1.64(d,J=11.8Hz,1H) ,1.55(d,J=10.8Hz,3H),1.47–1.33(m,2H),1.31–1.19(m,1H),1.15–1.00(m,4H),0.97(s,3H),0.71(s,3H); 13C NMR (100MHz, CDCl3) δ144.53,137.40,132.57,130.01,129.77,127.76,123.58,123.0 0,122.24,119.17,56.08,50.81,49.31,46.72,36.50,35.13,34.83,32.21,30.67,30 .59,30.14,26.11,21.42,21.16,19.33,18.68,12.90; FT-IR(KBr)ν:3455.42,3174.6 8,2944.62,1738.21,1486.36,1433.31,1369.86,1217.71,1071.78,817.50,760.03cm -1 ;HR-MS(ESI-TOF)m / z:Calcd for C 27 H 33 BrN3O{[M+H] +}494.1802, found494.1802.

[0100] Effect Example: In vitro antitumor activity test

[0101] The MTT method was used to test the in vitro growth and proliferation inhibitory activity of the target compound on three cell lines: human cervical cancer cells (HeLa), human gastric cancer cells (HGC-27) and human renal epithelial cells (HEK-293T). All cells were cultured in DMEM medium (added with 10% bovine serum and 0.1g / L penicillin G + 0.1g / L streptomycin sulfate) at 37°C in a humidified atmosphere of 5% CO2. The target compound was dissolved in sterile DMSO (Sigma) at a concentration of 10 mg / mL and diluted with culture medium according to the concentration used. Cell viability assessment: cells in the logarithmic growth phase were diluted at about 3×10 4 The cells were inoculated at a density of 100 μL / mL in a 96-well plate, 200 μL was inoculated in each well, and the plate was placed in a CO2 incubator for 24 hours. Then, different concentrations of the test compound were added to the test wells, 6 parallel wells were set for each concentration gradient, and an equal amount of DMSO was used as a blank control. The cells were cultured in a carbon dioxide incubator for 48 hours, the cells were washed with sterile PBS (phosphate buffer saline), and then 190 μL of culture medium and 10 μL of MTT (5 mg / mL) solution were added to each well, and the cells were incubated in a carbon dioxide incubator for 4 hours; the supernatant was discarded, 200 μL of DMSO was added, and the purple precipitate was dissolved by shaking on a vibrator for 10 minutes, and then the OD value was measured at a wavelength of 490 nm using an enzyme marker. The results are shown in Table 1:

[0102] Table 1 In vitro antiproliferative activity of dehydroepiandrosterone derivatives [IC50 / (μmol·L -1 )]

[0103]

[0104] From the data in Table 1, we can see that the inhibitory activity of the raw material drug dehydroepiandrosterone on human cervical cancer cells (HeLa) is IC 50 The value is 63.3 μmol·L -1 , inhibitory activity IC of human gastric cancer cells (HGC-27) 50 The value is 42.6 μmol·L -1 ; Inhibitory activity of cisplatin on human cervical cancer cells (HeLa) IC 50 The value is 34.6 μmol·L -1 , inhibitory activity against human gastric cancer cells (HGC-27) IC 50 The value is 10.4 μmol·L -1 Among the synthesized DHEA derivatives, except for compounds 3c, 3g and 3h, these DHEA derivatives did not show obvious inhibitory activity against the tested tumor cell lines and human normal cells 293T. However, for compound 3c, it showed good inhibitory activity against the tested tumor cell lines, such as the inhibitory activity IC of 1.347 against human cervical cancer cells (HeLa). 50 The value is only 33.6 μmol·L -1 , inhibitory activity IC of human gastric cancer cells (HGC-27) 50 The value was only 22.3 μmol·L -1 , which are superior to the raw material drug dehydroepiandrosterone (its inhibitory activity against human cervical cancer cells IC 50 The value is 63.3 μmol·L -1 , inhibitory activity IC of human gastric cancer cells (HGC-27) 50 The value is 42.6 μmol·L -1 ); however, it showed little inhibitory activity against normal human renal epithelial cells (HEK-293T) (IC 50 Value>100μmol·L -1 ); for compounds 3g and 3h, they showed certain inhibitory activity against the tested tumor cell lines, such as the inhibitory activity IC50 values ​​against human cervical cancer cells (HeLa) were 93.2 μmol·L -1 and 78.5 μmol·L -1 , inhibitory activity IC of human gastric cancer cells (HGC-27) 50 The value is only 78.5 μmol·L -1 and 74.4 μmol·L-1 However, it showed little inhibitory activity against normal human renal epithelial cells (HEK-293T) (IC 50 Value>100μmol·L -1 ); This indicates that these three compounds have certain activity in inhibiting the growth and proliferation of tumor cells, while having less damage to normal cells.

[0105] It should be understood that the terms used herein are only for the purpose of describing specific example embodiments and are not intended to be limiting. Unless the context clearly indicates otherwise, the singular forms "one", "an" and "said" as used herein may also be meant to include plural forms. The terms "include", "comprise", "contain", and "have" are inclusive, and therefore specify the existence of stated features, steps, operations, elements and / or parts, but do not exclude the existence or addition of one or more other features, steps, operations, elements, parts, and / or combinations thereof. The method steps, processes, and operations described herein are not interpreted as necessarily requiring them to be performed in the specific order described or illustrated, unless the execution order is clearly indicated. It should also be understood that additional or alternative steps may be used.

[0106] Although the terms first, second, third, etc. can be used in the text to describe multiple elements, components, regions, layers and / or sections, these elements, components, regions, layers and / or sections should not be limited by these terms. These terms can only be used to distinguish an element, component, region, layer or section from another region, layer or section. Unless the context clearly indicates, terms such as "first", "second" and other numerical terms do not imply order or sequence when used in the text. Therefore, the first element, component, region, layer or section discussed below can be referred to as the second element, component, region, layer or section without departing from the teaching of the example embodiments.

[0107] For ease of description, spatial relative terms may be used herein to describe the relationship of one element or feature relative to another element or feature as shown in the figure, such as "inside", "outside", "inner side", "outer side", "below", "below", "above", "above", etc. Such spatial relative terms are intended to include different orientations of the device in use or operation in addition to the orientation depicted in the figure. For example, if the device in the figure is turned over, then the elements described as "below other elements or features" or "below other elements or features" will subsequently be oriented as "above other elements or features" or "above other elements or features". Therefore, the example term "below..." can include both upper and lower orientations. The device can be oriented otherwise (rotated 90 degrees or in other directions) and the spatial relative descriptors used in the text are interpreted accordingly.

[0108] The above is only a preferred specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any changes or substitutions that can be easily thought of by a person skilled in the art within the technical scope disclosed by the present invention should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention shall be subject to the protection scope of the claims.

Claims

1. A compound represented by formula 3c, 3g or 3h, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof:

2. A method for preparing a compound represented by formula (I), or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, comprising the following preparation method: Step a: In an organic solvent, dehydroepiandrosterone compound 1 reacts with an azide compound under the action of triphenylphosphine and azodicarbonate, after the reaction is completed, a dilute alkali solution is added for quenching, and then an organic solvent is added for extraction and separation to obtain a compound 2 in which the hydroxyl group is replaced by azide; Step b: The compound 2 reacts with a copper salt and an acetylene compound in a polar alkaline solvent. After the reaction is completed, an organic solvent is added for extraction to separate and obtain a compound represented by formula (I). In formula (I), A, B, and C are nitrogen; The substituent R is Any of .

3. The preparation method according to claim 2, characterized in that: In the step a, the organic solvent is selected from one or two or more of diethyl ether, tetrahydrofuran, 1,4-dioxane, methyl tert-butyl ether, acetonitrile, toluene or xylene; The azodicarbonate is diethyl azodicarbonate DEAD or diisopropyl azodicarbonate DIAD; The azide compound is one of bis(P-nitrophenyl) azidophosphate, 15-azido-4,7,10,13-tetraoxypentadecanoic acid-N-succinimidyl ester, diphenyl azidophosphate or azide-heptapolyethylene glycol-pentafluorophenyl ester.

4. The preparation method according to any one of claims 2 to 3, characterized in that: In the step b, the acetylene compound is selected from any one of p-methoxyphenylacetylene, 2-bromophenylacetylene or 3-bromophenylacetylene; The polar solvent is selected from any one of water-methanol, water-ethanol, water-isopropanol, water-n-butanol or water-tert-butanol; The copper salt is selected from any one of copper sulfate, copper acetate, copper chloride, copper nitrate or copper carbonate; the base is selected from any one of sodium carbonate, sodium bicarbonate, sodium sulfite, sodium bisulfite or sodium ascorbate.

5. A pharmaceutical composition, characterized in that: A preparation prepared by using the compound according to claim 1, or its stereoisomer, or its pharmaceutically acceptable salt as a pharmaceutical active ingredient and adding pharmaceutically acceptable excipients.

6. Use of the compound according to claim 1, or its stereoisomer, or its pharmaceutically acceptable salt, or the pharmaceutical composition according to claim 5 in the preparation of drugs against human cervical cancer cells and / or human gastric cancer cells.

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