Diaryl heptanone alkyne compound as well as preparation method and application thereof

Abstract

The invention relates to the technical field of pharmaceutical chemicals, in particular to a diaryl heptanone alkyne compound as well as a preparation method and application thereof. The compound has a structure shown in the specification, has good anti-tumor pharmacological activity, and is particularly suitable for high-flux target screening, biological imaging and anti-cancer drugs of the anti-cancer effect of the diaryl heptanone compound and development as a functional unit. Compared with the prior art, the invention has obvious advantages in the aspects of synthesis method, drug activity, target confirmation and biological imaging. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceutical chemistry, and particularly relates to a diarylheptanone alkyne compound, a preparation method thereof, and an application thereof. Background Art

[0002] Diarylheptanone compounds are a class of structures of natural products and their derivatives with significant anti-tumor activity. Among them, 1,7-di-(4-hydroxyphenyl)-heptane-1,4-diene-3-one (hereinafter referred to as "DHDK") has been proven to inhibit the proliferation of various tumor cells (such as breast cancer MCF-7, lung cancer A549, etc.), and has an inhibitory effect on multiple stages of tumorigenesis and development. Although this class of compounds has good safety and biological activity, their application is limited by bottleneck problems such as poor water solubility, low metabolic stability, insufficient targeting, limited structural modification, and lack of multifunctionality. To overcome the above problems, introducing an alkyne group has become an effective strategy: Alkyne modification can significantly improve metabolic stability and targeting, and achieve multifunctional design through click chemistry reactions (such as coupling with water-soluble groups, organelle targeting molecules, imaging probes, or target recognition molecules), and at the same time, based on the characteristics of bioorthogonal reactions, it provides an innovative tool for the efficient fishing of drug target proteins and the analysis of action mechanisms, meeting the requirements of targeting, visualization, and mechanism traceability in drug development under the background of precision medicine. It can not only effectively solve the inherent defects of diarylheptanone compounds, but also provide a new direction for their multifunctional development and mechanism research.

[0003] Click chemistry (proposed by K. Barry Sharpless et al.) has become a core tool for drug target research due to its high efficiency and strong specificity. Its advantages in drug target recognition lie in its high efficiency, selectivity, flexibility, and bioorthogonality, which can quickly and accurately label and identify targets, promoting drug discovery and development. However, it also has limitations in terms of molecular stability, reaction efficiency, specificity, etc. There are still the following limitations in click chemistry for the modification of diarylheptanone compounds or other natural products:

[0004] Insufficient ester bond stability: Existing methods often introduce alkynes into phenolic hydroxyl-containing compounds (such as diarylheptanone compounds) through ester bonds, such as patents CN 117924174 A and CN 116669713 A, but the ester bond is prone to hydrolysis under physiological conditions, resulting in molecular inactivation;

[0005] Difficulty in selectively modifying asymmetric molecules: For natural products with asymmetric structures (such as DHDK), polyphenolic hydroxyls may be contained on different benzene rings, and the site of alkyne introduction needs to be precisely controlled. Existing methods usually introduce alkynes at the final product stage, making it difficult to achieve selective modification.

[0006] Weak reactivity: The nucleophilicity of phenolic hydroxyl is weaker than that of aliphatic alcohols, and the reactivity of alkynes with protecting groups (such as TMS-propynyl bromide) is further reduced, making it difficult to react under ordinary alkaline conditions (such as K2CO3), and there is no relevant report on this yet.

[0007] Harsh conditions: Natural products often contain sensitive groups such as unsaturated bonds and have poor tolerance to reaction conditions (such as high temperature, strong acid / alkali). There is an urgent need to develop mild, rapid and simple reaction conditions.

[0008] Reduced probe activity: After introducing alkynes, the pharmacological activity of some compounds may decrease. Therefore, the alkyne structure should be as small as possible to minimize the impact on the activity of the compounds.

[0009] In summary, there is an urgent need to develop an efficient, mild and highly selective synthesis method that can stably introduce alkynyl groups while retaining the functions of compounds, and achieve high-throughput screening, bioimaging and drug development of targets. Summary of the Invention

[0010] The object of the present invention is to provide a diarylheptanone alkyne compound, a preparation method thereof and an application thereof.

[0011] A diarylheptanone alkyne compound, the compound has the following structure:

[0012]

[0013] In the formula,

[0014] R1 and R2 can be the same or different and are selected from H, C3-C7 alkynyl groups which are unsubstituted or substituted by at least one R5, and at least one of R1 and R2 is selected from C3-C7 alkynyl groups which are unsubstituted or substituted by at least one R5;

[0015] R3 and R4 can be the same or different and are selected from H, halogen, C1-C4 alkyl, C1-C4 alkanoyl, C1-C4 alkoxy or hydroxyl;

[0016] R5 is selected from halogen, C1-C4 alkyl, C1-C4 alkanoyl, C1-C4 alkoxy or hydroxyl;

[0017] x and n can be the same or different and are selected from 1, 2, 3 or 4, and x + a is less than or equal to 5;

[0018] a and b can be the same or different and are selected from 1, 2, 3 or 4, and n + b is less than or equal to 5.

[0019] Preferably, in the structural formula of the compound,

[0020] R1 and R2 are independently selected from H and alkynyl groups having 3 to 7 carbon atoms, and at least one of R1 and R2 is an alkynyl group having 3 to 7 carbon atoms;

[0021] R3 and R4 are independently selected from H, halogen, alkyl groups having 1 to 4 carbon atoms, alkanoyl groups having 1 to 4 carbon atoms, alkoxy groups having 1 to 4 carbon atoms, or hydroxyl;

[0022] x and n are independently selected from 1, 2, 3, or 4, and x + a ≤ 5;

[0023] a and b are independently selected from 1, 2, 3, or 4, and n + b ≤ 5.

[0024] More preferably, in the chemical structure formula of the compound,

[0025] R1 and R2 are independently selected from H and alkynyl groups having 3 to 7 carbon atoms, and at least one of R1 and R2 is an alkynyl group having 3 to 7 carbon atoms;

[0026] x and n are independently selected from 1, 2, 3, 4, or 5.

[0027] Even more preferably, the compound is

[0028]

[0029] A method for preparing the diarylheptanone alkyne compound as described above:

[0030] (1) Esterify the hydroxycinnamic acid with a side chain, and then perform nucleophilic substitution, reduction, and oxidation reactions to obtain an aldehyde product;

[0031] (2) Subject the hydroxybenzaldehyde with a side chain to a nucleophilic reaction and aldol condensation to obtain a product;

[0032] (3) Perform aldol condensation on the products obtained in steps (1) and (2), and remove the protecting group to obtain a terminal alkyne structure, thereby obtaining the diarylheptanone alkyne compound;

[0033] In the above steps (1) and (2), the nucleophilic substitution is to protect the phenolic hydroxyl group or introduce a silicon-protected alkynyl halide; and in at least one step, a silicon-protected alkynyl halide is introduced.

[0034] In step (2), the hydroxybenzaldehyde with a side chain undergoes protection of the phenolic hydroxyl group through a nucleophilic reaction, and then aldol condensation to obtain a product; or, after deprotecting the product obtained by aldol condensation, a nucleophilic reaction is performed to introduce a terminal alkyne, thereby obtaining a product.

[0035] In steps (1) and (2), the nucleophilic substitution reaction is the protection of the phenolic hydroxyl group and the introduction of a silicon-protected alkynyl halide.

[0036] When protecting the phenolic hydroxyl group, the esterified product of hydroxycinnamic acid with a side chain or benzaldehyde with a side chain is used as a raw material, and in an alkaline environment, it is respectively subjected to a nucleophilic substitution reaction with a phenolic hydroxyl group protecting group at 4 °C - 100 °C for 1 h - 10 h. Among them, the molar ratio of the raw material to the base substance providing the alkaline environment is 1:0.1 eq - 1:10 eq; the molar ratio of the raw material to the phenolic hydroxyl group protecting group is 1:1 eq - 1:10 eq;

[0037] When introducing a terminal alkyne, the esterified product of hydroxycinnamic acid with a side chain, hydroxybenzaldehyde with a side chain, or the product obtained after phenolic hydroxyl group protection, aldol condensation, and then deprotection of hydroxybenzaldehyde with a side chain is used as a raw material, and in an alkaline environment, it is subjected to a nucleophilic substitution reaction with a silicon-protected haloalkyne at 4 °C - 100 °C for 0.16 h - 2 h to introduce a terminal alkyne. Among them, the molar ratio of the raw material to the base substance providing the alkaline environment is 1:0.1 eq - 1:10 eq; the molar ratio of the raw material to the silicon-protected haloalkyne is 1:1 eq - 1:10 eq; the base substance is an inorganic base and an organic base, and the ratio of the two is 1:0.1 eq - 10:1 eq.

[0038] Among them, the inorganic base is potassium carbonate, sodium carbonate, sodium bicarbonate, potassium hydroxide, sodium hydroxide, and the organic base is triethylamine, N,N-diisopropylethylamine (DIPEA), pyridine.

[0039] Specifically, taking the obtained compound with alkynes at both ends as an example, step (1) taking the preparation of an alkyne-substituted aldehyde as an example, includes the following steps:

[0040] ① Add hydroxycinnamic acid with a side chain to an alcohol solution at 10 - 40 °C, stir to dissolve, add sulfuric acid for reaction, extract, dry, and rotary evaporate to obtain intermediate 1;

[0041] Among them, the weight ratio of hydroxycinnamic acid with a side chain to sulfuric acid is 1:0.01 - 1:1;

[0042] ② Dissolve intermediate 1 in a CH2Cl2 solution, stir evenly, carry out a nucleophilic substitution reaction under alkaline conditions, react at 4 - 100 °C, after the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 2;

[0043] Among them, the nucleophilic substitution is to introduce a silicon-protected haloalkyne.

[0044] The silicon protecting group of the silicon-protected haloalkane is TMS (trimethylsilyl), TES (triethylsilyl), TBS (tert-butyldimethylsilyl), triisopropylsilyl (TIPS)

[0045] The halo reagent of the haloalkane with a protecting group is bromoalkyne (R-C≡C-Br), iodoalkyne (R-C≡C-I), chloroalkyne (R-C≡C-Cl)

[0046] Among them, the weight - volume ratio (g / mL) of intermediate 1, the basic substance and the silicon - protected halogenated hydrocarbon reagent is 1:0.1:1 - 1:10:20;

[0047] The basic condition is controlled by two of N,N - diisopropylethylamine, triethylamine, DBU, potassium carbonate, potassium hydroxide, sodium carbonate, etc.; the types of the two bases are one organic base and one inorganic base used in combination. The mass - volume ratio (g / mL) of the two is 1:0.1 - 10:1.

[0048] ③ Dissolve intermediate 2 in an anhydrous DMF or THF solution, and slowly add a reducing agent to the reaction solution under an ice - bath condition. After the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 3;

[0049] The reducing agent is controlled by one of lithium aluminum hydride, sodium borohydride, and tetraethylboron.

[0050] Among them, the molar ratio of intermediate 2 to the reducing agent is 1:1eq - 1:4eq;

[0051] ④ Dissolve intermediate 3 in a CH2Cl2 solution, react with sodium bicarbonate for 30 min under an ice - bath condition, add an oxidizing agent, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 4.

[0052] The oxidizing agent is controlled by one of Dess - Martin periodinane or Swern oxidation (Swern oxidation contains DMSO, oxalyl chloride and triethylamine).

[0053] Among them, the weight ratio of intermediate 3 to sodium bicarbonate is 1:1 - 1:20, and the molar ratio of intermediate 3 to the oxidizing agent is 1:0.2eq - 1:20eq;

[0054] Specifically, taking the preparation of an alkyne - substituted ketone as an example, step (2) includes the following steps:

[0055] ① Dissolve the hydroxyl - benzaldehyde with a side chain in a CH2Cl2 solution, carry out a nucleophilic substitution reaction under the conditions of - 10 - 70 °C and basic conditions. After the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 5;

[0056] The nucleophilic substitution reaction is the protection of the phenolic hydroxyl group;

[0057] The phenolic hydroxyl - protecting group reagent is protected by ethers or silyl ethers.

[0058] The ethers are those with protecting groups such as methoxymethyl (MOM) and tetrahydropyranyl (THP);

[0059] The silyl - ether protecting groups are TMS, TBS, and TES.

[0060] Among them, the mass-volume ratio (g / mL) of the hydroxybenzaldehyde with a side chain, the basic substance, and the phenolic hydroxyl protecting group reagent is 1:0.1:1 - 1:10:20.

[0061] The basic condition is controlled by one of N,N-diisopropylethylamine, triethylamine, DBU, potassium carbonate, potassium hydroxide, sodium carbonate, etc.

[0062] ② Add intermediate 5 to acetone, add a base and react at -10 - 20 °C. After the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 6;

[0063] Among them, the base is sodium hydroxide or potassium hydroxide; the weight-volume ratio (g / mL) of intermediate 5 to the base is 10:1 - 1:1;

[0064] ③ Add intermediate 6 to methanol, add an acid and react at 4 - 80 °C. After the reaction is completed, extract, dry, rotary evaporate to obtain intermediate 7;

[0065] Among them, the weight-volume ratio (g / mL) of intermediate 6 to the acid is 1:1 - 1:5; the acid is hydrochloric acid or sulfuric acid;

[0066] ④ Add intermediate 7 to acetone, and react with a silicon-based protected haloalkyne at 4 - 100 °C under basic conditions. After the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 8;

[0067] The basic condition and the condition of the silicon-based protected haloalkyne are the same as those in ② of step (one).

[0068] Specifically, step (3) includes the following steps:

[0069] ① Add intermediate 8 to anhydrous THF, and slowly add compound 4 thereto in the presence of a basic substance at -80 - 4 °C. After the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 9;

[0070] The basic substance is LDA, LiHMDS, NaHMDS, KHMDS, NaH, LiTMP;

[0071] Among them, the molar ratio of intermediate 8 to the basic substance is 1:0.5 - 1:50 eq.

[0072] ② At 4 - 100 °C, add intermediate 9 to anhydrous THF, add a dehaloalkyne protecting group reagent, react for 2 - 4 h, then add sulfuric acid or hydrochloric acid. After the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain the diarylheptanone alkyne compound.

[0073] The protecting group reagent for dehaloalkynes is a fluorinating reagent, specifically tetraethylammonium fluoride, tetrabutylammonium fluoride, hydrofluoric acid, and trifluoroacetic acid.

[0074] Among them, the molar ratio of intermediate 9 to the desilyl protecting group reagent is 1:0.1eq - 1:0.5eq; the weight ratio (g / mL) of intermediate 9 to sulfuric acid or hydrochloric acid is 1:0.01 - 1:5.

[0075] Specifically, taking the compound with an alkyne group at the ketone end as an example, step (1) takes the preparation of an aldehyde protected by a phenolic hydroxyl group as an example, and includes the following steps:

[0076] ① Add hydroxycinnamic acid with a side chain to an alcohol solution at 10 - 40 °C, stir to dissolve, add sulfuric acid for reaction, extract, dry, rotary evaporate to obtain intermediate 1;

[0077] Among them, the weight - volume ratio (g / mL) of hydroxycinnamic acid with a side chain to sulfuric acid is 1:0.01 - 1:1;

[0078] ② Dissolve intermediate 1 in a CH2Cl2 solution, stir evenly, carry out a nucleophilic substitution reaction under alkaline conditions, react at 4 - 100 °C, after the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 10;

[0079] The said nucleophilic substitution reaction is for phenolic hydroxyl protection, and the alkaline conditions, phenolic hydroxyl protecting group, and steps are the same as those in (2)① for the preparation of the compound with alkynes at both ends;

[0080] ③ Dissolve intermediate 10 in an anhydrous DMF or THF solution, slowly add a reducing agent to the reaction solution under ice - bath conditions, after the reaction is completed, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 11;

[0081] The type, conditions, and steps of the said reducing agent are the same as those in (1)③ for the preparation of the compound with alkynes at both ends;

[0082] ④ Dissolve intermediate 11 in a CH2Cl2 solution, react with sodium bicarbonate for 30 min under ice - bath conditions, add an oxidizing agent, extract, dry, rotary evaporate, and purify by column chromatography to obtain intermediate 12.

[0083] The type, conditions, and steps of the said oxidizing agent are the same as those in (1)④ for the preparation of the compound with alkynes at both ends;

[0084] Specifically, for step (2) to prepare a ketone substituted by an alkyne, the conditions and steps are the same as those in (2) for the preparation of the compound with alkynes at both ends, and intermediates 5, 6, 7, and 8 are obtained

[0085] Specifically, step (3) is the same as that for preparing alkynes at both ends to obtain intermediate 13 and diarylheptanone compounds, including the following steps:

[0086] ① React intermediate 8 and intermediate 12 under the same reaction conditions and steps as in (3)① for preparing compounds with alkynes at both ends to obtain intermediate 13;

[0087] ② React intermediate 13 and a deprotection reagent under the same reaction conditions and steps as in (3)② for preparing compounds with alkynes at both ends to obtain diarylheptanone compounds.

[0088] Specifically, taking the obtained compound with an aldehyde - terminal alkyne as an example, step (1) takes the preparation of an alkyne - substituted ester as an example, and is the same as the conditions and steps of (1)①② for preparing compounds with alkynes at both ends to obtain intermediates 1 and 2.

[0089] Specifically, taking the preparation of a phenol - hydroxyl - protected ketone as an example, step (2) is the same as the conditions and steps of (2)①② for preparing compounds with alkynes at both ends to obtain intermediates 5 and 6.

[0090] Specifically, step (3) is the same as that for preparing compounds with alkynes at both ends to obtain intermediate 14 and diarylheptanone compounds, including the following steps:

[0091] ① React intermediate 2 and intermediate 6 under the same reaction conditions and steps as in (3)① for preparing compounds with alkynes at both ends to obtain intermediate 14;

[0092] ② React intermediate 14 and a deprotection reagent under the same reaction conditions and steps as in (3)② for preparing compounds with alkynes at both ends to obtain diarylheptanone compounds.

[0093] All solvents in the text are common solvents. Without special instructions, other reagents can be used instead.

[0094] An application of the described diarylheptanone alkyne compound, the application of the diarylheptanone alkyne compound in the preparation of anti - tumor drugs.

[0095] An application of the described diarylheptanone alkyne compound, the application of the diarylheptanone alkyne compound in biological imaging.

[0096] The application of the diarylheptanone alkyne compound as a probe for diarylheptanone anti - tumor compounds.

[0097] The application of the diarylheptanone alkyne compound as a probe for identifying anti - tumor protein targets that interact with it in vivo or in vitro.

[0098] The present invention has the following advantages:

[0099] The present invention achieves high selectivity through a stepwise protection strategy, is compatible with symmetric / asymmetric molecules and complex skeletons containing saturated / unsaturated bonds, has mild conditions and simple operations. The probe has good anti-tumor pharmacological activity and is particularly suitable for high-throughput target screening, bioimaging of the anti-cancer effects of diarylheptanone compounds, anti-cancer drugs, and development as a functional unit. Compared with the prior art, the present invention has significant advantages in terms of synthesis methods, drug activity, target confirmation, and bioimaging; furthermore,

[0100] 1. It can improve the properties of the parent nucleus: Introduce alkynes as functional units in diarylheptanone compounds. By constructing terminal alkyne structures in diarylheptanone compounds, target labeling or bioimaging can be achieved through click chemistry, which can be used to improve the water solubility and metabolic stability of the parent nucleus structure, providing diverse molecular tools for subsequent structural modification and drug development.

[0101] 2. The first dual-base strategy: Adopt the "dual-base strategy" of weak cooperation between inorganic base and organic base. The synergistic effect of inorganic base and organic base is used to solve the problem of low reaction activity between phenolic hydroxyl groups and protected haloalkynes, and realize the efficient etherification reaction between phenolic hydroxyl groups and haloalkynes with silicon protecting groups (such as TMS-propargyl bromide). The reaction conditions are mild, avoiding the destruction of sensitive groups (such as unsaturated bonds), and the introduction site of alkynes in asymmetric molecules can be accurately controlled. This method has mild conditions, high selectivity, and does not damage sensitive groups such as unsaturated bonds, meeting the synthesis requirements of complex molecules.

[0102] 3. Replace the ester bond with an ether bond: Connect alkynes with an ether bond instead of the traditional ester bond. Utilize the high chemical stability and bioorthogonality of the ether bond to avoid the problem of molecular inactivation caused by the hydrolysis of the ester bond under physiological conditions, and at the same time avoid the steric effect caused by the introduction of large-volume groups, retaining the pharmacological activity of the original compound to the greatest extent.

[0103] 4. Selective introduction of alkynes: Introduce alkynes with silicon protecting groups (such as TMS-propargyl bromide) during the synthesis process. Through a stepwise protection strategy, alkynes can be selectively introduced into compounds containing multiple phenolic hydroxyl groups and asymmetric structures, while protecting sensitive groups (such as unsaturated bonds)

[0104] 5. High activity and rapid labeling: The probe (such as DHDK-P) has anti-tumor activity comparable to that of the parent nucleus compound (DHDK), and target in-situ labeling is achieved through click chemistry, with a short labeling time and significantly improved detection efficiency.

[0105] 6. Multi-dimensional target verification: The probe can directly trace target proteins through Western blot and bioimaging (azide fluorescein labeling) techniques, combined with multi-path verification of click chemical reactions, and the results are reliable.

[0106] In summary, compared with the prior art, the present invention for the first time develops a method for synthesizing diarylheptanone alkynyl probes based on the "dual-base strategy", stably introduces terminal alkynes through ether bonds, and is compatible with asymmetric molecules and complex skeletons containing sensitive groups. Taking DHDK-P as an example, this probe has both anti-tumor activity (IC 50 = 3.2 μM) and target labeling function (labeling efficiency > 90%), and can be widely used in target discovery, mechanism of action research and drug development. This method can be extended to the alkynylation modification of other phenol hydroxyl-containing molecules (such as curcumin, resveratrol), providing a general technical platform for the development of natural product drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0107] Figure 1 It is the reaction formula of the compound provided by the embodiment of the present invention.

[0108] Figure 2 It is the cell imaging diagram of the cell target competition experiment after the compound obtained by the present invention acts on cells

[0109] Figure 3 It is the fluorescence co-localization diagram of the target point and the protein PRDX1 after the compound obtained by the present invention acts on cells;

[0110] Figure 4 : The gel imaging diagram after different concentrations of DHDK-P act on cells;

[0111] Figure 5 : The proteomics heat map;

[0112] Figure 6 : The results diagram of the pull down and magnetic bead experiment; DETAILED DESCRIPTION OF THE INVENTION

[0113] The following further elaborates on the concept and technical effects generated by the present invention in combination with specific embodiments to fully understand the purpose, features and effects of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative efforts all fall within the scope of protection of the present invention. The methods are all conventional methods unless otherwise specified. The materials can all be obtained from public commercial channels unless otherwise specified.

[0114] Example 1: Preparation method of diarylheptanone alkynyl compounds with alkynes at both ends.

[0115] 1. Preparation of 4-(methoxymethoxy)-benzaldehyde

[0116] To a 500 mL round-bottom flask, 10 g (0.082 mol) of 4-hydroxybenzaldehyde, 50 mL of CH2Cl2 were added successively. After stirring well, 20 mL (0.115 mol) of DIPEA was added under ice bath conditions. After reacting for 15 min, about 20 mL (0.24 mol) of bromomethyl methyl ether was added to the bottom of the liquid surface, and the reaction was carried out under ice bath conditions. The reaction process was monitored by TLC. When the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl, extracted with EtOAc three times, 50 mL each time, dried over Na2SO4, filtered by suction, the solvent was rotary evaporated, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 5:1, v:v) to obtain the compound 4-(methoxymethoxy)-benzaldehyde.

[0117] 2. Preparation of 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one

[0118] To a 500 mL round-bottom flask, 10 g (0.06 mol) of 4-(methoxymethoxy)-benzaldehyde and 50 mL of acetone were added. 5 mL of 10% NaOH was added under ice bath conditions. The reaction process was monitored by TLC. After the raw materials reacted completely, the pH was adjusted to 6, extracted with EtOAc three times, 50 mL each time, dried over Na2SO4, filtered by suction, the solvent was rotary evaporated, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 10:1, v:v) to obtain the compound 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one. 1 1H NMR (600 MHz, MeOD) δ 9.16 (s, 1H), 7.42 (d, J = 7.5 Hz, 2H), 7.36 (dd, J = 12.0, 4.8 Hz, 4H), 7.30 (d, J = 7.4 Hz, 1H), 6.95 (d, J = 8.7 Hz, 2H), 6.70 (d, J = 16.3 Hz, 1H), 6.25 (d, J = 16.3 Hz, 1H), 5.07 (d, J = 6.4 Hz, 2H), 1.90 (s, 3H), 1.52 (s, 3H).

[0119] 3. Preparation of 4-(4-hydroxyphenyl)but-3-en-2-one

[0120] To a 500 mL round-bottom flask, 10 g (0.049 mol) of 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one was added, 50 mL of methanol and 5 ml of 20% HCl were added, and the mixture was refluxed at 60 °C. The reaction process was monitored by TLC. After the raw materials reacted completely, the pH was adjusted to 6, extracted with EtOAc three times, 50 mL each time, dried over Na2SO4, filtered by suction, the solvent was rotary evaporated, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 2:1, v:v) to obtain the compound 4-(4-hydroxyphenyl)but-3-en-2-one. 1HNMR (DMSO-d6, 400 MHz) δ: 10.01 (1H, brs, 4'-OH), 7.55 (2H, d, J = 8.6 Hz, H-2',6'), 7.53 (1H, d, J = 16.3 Hz, H-1), 6.81 (2H, d, J = 8.6 Hz, H-3',5'), 6.60 (1H, d, J = 16.3 Hz, H-2), 2.28 (3H, s, 4-CH3). 13 CNMR (400 MHz, DMSO-d6): δ 143.677, 124.1105, 197.862, 27.1989, 125.3452, 130.4682, 115.947, 160.0666。

[0121] 4. Preparation of 4-(4-(Trimethylsilylpropynyloxy)phenyl)but-3-en-2-one

[0122] To a 500 mL round-bottom flask, 0.50 g (3.08 mmol) of 4-(4-hydroxyphenyl)but-3-en-2-one, 1.54 g of K2CO3, 1.9 mL (6.31 mmol) of N,N-diisopropylethylamine, and 50 mL of acetone were added successively. After stirring well, 0.7 mL (4.39 mmol) of propargyl bromide was added under ice bath conditions. The reaction was carried out at 60 °C. When the production of the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl, and extracted with EtOAc three times, 50 mL each time. The solvent was rotary evaporated, dried over Na2SO4, filtered by suction, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 10:1, v:v) to obtain the compound 4-(4-(trimethylsilylpropynyloxy)phenyl)but-3-en-2-one. 13 C NMR (400 MHz, DMSO-d6): δ 142.8889, 125.3946, 197.8869, 27.1897, 127.5724, 130.0504, 115.216, 159.1119, 56.3346, 100.9602, 92.1371, -0.384。

[0123] 5. Preparation of methyl 3-(4-hydroxyphenyl)propionate

[0124] To a 500 mL round-bottom flask, 10 g (0.072 mol) of 4-hydroxybenzoic acid and 150 mL of methanol were added successively, and stirred at 35 °C. 5 drops of concentrated H2SO4 were added to the reaction solution. The reaction progress was monitored by TLC. After the raw materials reacted completely, 100 mL of water was added to quench the reaction, and the pH was adjusted to 6 with saturated NaHCO3. The methanol was rotary evaporated, extracted with EtOAc three times, 10 mL each time, dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated to obtain the compound methyl 3-(4-hydroxyphenyl)propionate.

[0125] 6. Preparation of methyl 3-(4-(trimethylsilylpropynyloxy)phenyl)propionate

[0126] To a 500 mL round-bottom flask, 0.50 g (2.77 mmol) of methyl 3-(4-hydroxyphenyl)propionate, 1.54 g of K2CO3, 1.9 mL (6.31 mmol) of N,N-diisopropylethylamine, and 50 mL of acetone were added successively. After stirring well, 0.7 mL (4.39 mmol) of propargyl bromide silane was added under ice bath conditions. The reaction was carried out at 60 °C. When the production of the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl, and extracted with EtOAc three times, 50 mL each time. The solvent was rotary evaporated, dried over Na2SO4, filtered by suction, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 10:1, v:v) to obtain the compound methyl 3-(4-(trimethylsilylpropynyloxy)phenyl)propionate.

[0127] 7. Preparation of 3-(4-(trimethylsilylpropynyloxy)phenyl)propan-1-ol

[0128] To a 500 mL round-bottom flask, 4 g (13.8 mmol) of methyl 3-(4-(trimethylsilylpropynyloxy)phenyl)propionate and 100 mL of THF were added successively. Stirring was carried out under ice bath. 1.5 g of lithium aluminum hydride was slowly added to THF to form a suspension, and the lithium aluminum hydride suspension was slowly added to the reaction solution. The reaction process was monitored by TLC. After the raw materials reacted completely, water was slowly added dropwise to the reaction solution to quench the reaction. The mixture was extracted with EtOAc three times, 50 mL each time, dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated to obtain the compound 3-(4-(trimethylsilylpropynyloxy)phenyl)propan-1-ol.

[0129] 8. Preparation of 3-(4-(trimethylsilylpropynyloxy)phenyl)propanal

[0130] To a 500 mL round-bottom flask, 4 g (15.2 mmol) of 3-(4-(trimethylsilylpropynyloxy)phenyl)propan-1-ol, 100 mL of CH2Cl2, and 11 g of NaHCO3 were added successively. The reaction was carried out at ice bath conditions for 30 min, and 20 g of Dess-Martin periodinane (47.2 mmol) was added. The reaction process was monitored by TLC. After the raw materials reacted completely, the reaction was quenched with saturated Na2S2O3. The mixture was extracted with EtOAc three times, 100 mL each time, dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated. The crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 20:1, v:v) to obtain the compound 3-(4-(trimethylsilylpropynyloxy)phenyl)propanal.

[0131] 9. Preparation of 1,7 - bis(4 - (trimethylsilylpropynyloxy)phenyl)hepta - 1,4 - diene - 3 - one

[0132] Add 2 g (0.0073 mol) of 4 - (4 - (trimethylsilylpropynyloxy)phenyl)but - 3 - en - 2 - one, 30 mL of anhydrous THF to a 100 - mL round - bottom flask in sequence. Under N₂ protection, cool the temperature to - 78 °C, slowly add 7.5 mL of LiHMDS (0.0075 mol), stir for 30 min, then slowly add 1.9 g of 3 - (4 - (trimethylsilylpropynyloxy)phenyl)propanal (0.0073 mol). Monitor the reaction process by TLC. When the product no longer increases, pour the reaction solution into saturated NH₄Cl, extract with CH₂Cl₂ three times, 20 mL each time, dry over Na₂SO₄, filter by suction, rotary evaporate the solvent, and separate and purify the crude product by silica gel column chromatography (PE:EtOAc = 5:1, v:v) to obtain the compound 1,7 - bis(4 - (trimethylsilylpropynyloxy)phenyl) - hepta - 1,4 - diene - 3 - one.

[0133] 10. Preparation of 1,7 - bis(4 - (propynyloxy)phenyl)hepta - 1,4 - diene - 3 - one

[0134] Add 0.5 g (0.89 mmol) of the compound 1,7 - bis(4 - (trimethylsilylpropynyloxy)phenyl) - hepta - 1,4 - diene - 3 - one, 10 mL of THF, and 0.1 g of tetraethylammonium fluoride to a 50 - mL round - bottom flask. Monitor the reaction process by TLC. When the raw materials react completely, add 1 mL of 20% sulfuric acid to the reaction solution and react at room temperature. Monitor the reaction process by TLC. After most of the raw materials are consumed, add 15 mL of saturated NaCl, extract with EtOAc three times, 20 mL each time, dry over Na₂SO₄, filter by suction, rotary evaporate the solvent, and separate and purify the crude product by ODS column chromatography (MeOH:H₂O = 55:1 → 70:1, v:v) to obtain the compound 1,7 - bis(4 - (propynyloxy)phenyl)hepta - 1,4 - diene - 3 - one.

[0135]

[0136] Example 2: Preparation method of diarylheptanone - type alkyne compounds with an alkyne at the ketone end

[0137] 1. Preparation of methyl 3 - (4 - hydroxyphenyl)propionate

[0138] 10 g (0.072 mol) of 4-hydroxybenzoic acid and 150 mL of methanol were successively added to a 500 mL round-bottom flask, stirred at 35 °C, and 5 drops of concentrated H2SO4 were added to the reaction solution. The reaction progress was monitored by TLC. After the raw materials were completely reacted, 100 mL of water was added to quench the reaction, and the pH was adjusted to 6 with saturated NaHCO3. Methanol was removed by rotary evaporation, and the mixture was extracted 3 times with 10 mL of EtOAc each time, dried over Na2SO4, filtered by suction, and the solvent was removed by rotary evaporation to obtain methyl 3-(4-hydroxyphenyl)propionate.

[0139] 2. Preparation of methyl 3-(4-(methoxymethoxy)phenyl)propionate

[0140] 10 g (0.055 mol) of methyl 3-(4-hydroxyphenyl)propionate and 250 mL of CH2Cl2 were successively added to a 500 mL round-bottom flask. After sufficient stirring, 20 mL (0.115 mol) of N,N-diisopropylethylamine was added under ice bath conditions. After reacting for 15 min, 20 mL (0.24 mol) of bromomethyl methyl ether was added under ice bath conditions. The reaction progress was monitored by TLC. When the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl, extracted 3 times with 50 mL of EtOAc each time, dried over Na2SO4, filtered by suction, and the solvent was removed by rotary evaporation. The crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 5:1, v:v) to obtain methyl 3-(4-(methoxymethoxy)phenyl)propionate. 1 1H NMR (600 MHz, CD3OD) δ 7.11 (d, J = 8.5 Hz, 2H), 6.93 (d, J = 8.6 Hz, 2H), 5.13 (s, 2H), 3.63 (s, 3H), 3.43 (s, 3H), 2.85 (t, J = 7.6 Hz, 2H), 2.59 (t, J = 7.6 Hz, 2H).

[0141] 3. Preparation of 3-(4-(methoxymethoxy)phenyl)propan-1-ol

[0142] 4 g (0.016 mol) of methyl 3-(4-(methoxymethoxy)phenyl)propionate and 100 mL of THF were successively added to a 500 mL round-bottom flask and stirred under ice bath. 1.5 g of lithium aluminum hydride was slowly added to THF to form a suspension, and the lithium aluminum hydride suspension was slowly added to the reaction solution. The reaction progress was monitored by TLC. After the raw materials were completely reacted, water was slowly added dropwise to the reaction solution to quench the reaction, extracted 3 times with 50 mL of EtOAc each time, dried over Na2SO4, filtered by suction, and the solvent was removed by rotary evaporation to obtain 3-(4-(methoxymethoxy)phenyl)propan-1-ol. 11H NMR (600 MHz, CDCl3): δ 7.11 (d, J = 8.6 Hz, 2H), 6.96 (d, J = 8.6 Hz, 2H), 5.15 (s, 2H), 3.66 (t, J = 6.4 Hz, 2H), 3.48 (s, 3H), 2.67 - 2.63 (m, 2H), 1.89 - 1.83 (m, 2H).

[0143] 4. Preparation of 3-(4-(methoxymethoxy)phenyl)propanal

[0144] To a 500 mL round-bottom flask, 4 g (0.019 mol) of 3-(4-(methoxymethoxy)phenyl)propan-1-ol, 100 mL of CH2Cl2, and 11 g of NaHCO3 were added successively. The reaction was carried out for 30 min under ice bath conditions. Then 20 g of Dess-Martin periodinane (0.0472 mol) was added. The reaction progress was monitored by TLC. After the raw materials were completely reacted, the reaction was quenched with saturated Na2S2O3. The mixture was extracted with EtOAc three times, 100 mL each time, dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated. The crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 20:1, v:v) to obtain 3-(4-(methoxymethoxy)phenyl)propanal.

[0145] 1. Preparation of 4-(methoxymethoxy)-benzaldehyde

[0146] To a 500 mL round-bottom flask, 10 g (0.082 mol) of p-hydroxybenzaldehyde and 50 mL of CH2Cl2 were added successively. After stirring well, 20 mL (0.115 mol) of DIPEA was added under ice bath conditions. After reacting for 15 min, about 20 mL (0.24 mol) of bromomethyl methyl ether was added to the bottom of the liquid surface, and the reaction was carried out under ice bath conditions. The reaction progress was monitored by TLC. When the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl. The mixture was extracted with EtOAc three times, 50 mL each time, dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated. The crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 5:1, v:v) to obtain 4-(methoxymethoxy)-benzaldehyde.

[0147] 2. Preparation of 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one

[0148] Add 10 g (0.06 mol) of 4-(methoxymethoxy)-benzaldehyde and 50 mL of acetone to a 500 mL round-bottom flask. Add 5 mL of 10% NaOH under an ice bath. Monitor the reaction progress by TLC. After the raw materials have completely reacted, adjust the pH to 6. Extract with EtOAc three times, 50 mL each time. Dry over Na2SO4, filter by suction, and rotary evaporate the solvent. The crude product is separated and purified by silica gel column chromatography (PE:EtOAc = 10:1, v:v) to obtain the compound 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one. 1 H NMR (600 MHz, MeOD) δ 9.16 (s, 1H), 7.42 (d, J = 7.5 Hz, 2H), 7.36 (dd, J = 12.0, 4.8 Hz, 4H), 7.30 (d, J = 7.4 Hz, 1H), 6.95 (d, J = 8.7 Hz, 2H), 6.70 (d, J = 16.3 Hz, 1H), 6.25 (d, J = 16.3 Hz, 1H), 5.07 (d, J = 6.4 Hz, 2H), 1.90 (s, 3H), 1.52 (s, 3H).

[0149] 3. Preparation of 4-(4-hydroxyphenyl)but-3-en-2-one

[0150] Add 10 g (0.049 mol) of 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one to a 500 mL round-bottom flask. Add 50 mL of methanol and 5 mL of 20% HCl. Reflux at 60 °C. Monitor the reaction progress by TLC. After the raw materials have completely reacted, adjust the pH to 6. Extract with EtOAc three times, 50 mL each time. Dry over Na2SO4, filter by suction, and rotary evaporate the solvent. The crude product is separated and purified by silica gel column chromatography (PE:EtOAc = 2:1, v:v) to obtain the compound 4-(4-hydroxyphenyl)but-3-en-2-one. 1 HNMR (DMSO-d6, 400 MHz) δ: 10.01 (1H, brs, 4'-OH), 7.55 (2H, d, J = 8.6 Hz, H-2',6'), 7.53 (1H, d, J = 16.3 Hz, H-1), 6.81 (2H, d, J = 8.6 Hz, H-3',5'), 6.60 (1H, d, J = 16.3 Hz, H-2), 2.28 (3H, s, 4-CH3). 13 CNMR (400 MHz, DMSO-d6): δ 143.677, 124.1105, 197.862, 27.1989, 125.3452, 130.4682, 115.947, 160.0666.

[0151] 4. Preparation of 4-(4-(trimethylsilylpropynyloxy)phenyl)but-3-en-2-one

[0152] To a 500 mL round-bottom flask, 0.50 g (3.08 mmol) of 4-(4-hydroxyphenyl)but-3-en-2-one, 1.54 g of K2CO3, 1.9 mL (6.31 mmol) of N,N-diisopropylethylamine, and 50 mL of acetone were successively added. After stirring well, 0.7 mL (4.39 mmol) of propargyl bromide silane was added under ice bath conditions. The reaction was carried out at 60 °C. When the production of the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl, and extracted with EtOAc three times, 50 mL each time. The solvent was rotary evaporated, dried over Na2SO4, filtered by suction, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 10:1, v:v) to obtain the compound 4-(4-(trimethylsilylpropargyloxy)phenyl)but-3-en-2-one. 13 13C NMR (400 MHz, DMSO-d6): δ 142.8889, 125.3946, 197.8869, 27.1897, 127.5724, 130.0504, 115.216, 159.1119, 56.3346, 100.9602, 92.1371, -0.384.

[0153] 9. Preparation of 1-(4-(trimethylsilylpropargyloxy)phenyl)-7-(4-(methoxymethoxy)phenyl)hept-1,4-dien-3-one

[0154] To a 500 mL round-bottom flask, 2 g (7.34 mmol) of 4-(4-(trimethylsilylpropargyloxy)phenyl)but-3-en-2-one and 30 mL of anhydrous THF were successively added. Under N2 protection, the temperature was lowered to -78 °C, and 7.5 mL of LiHMDS (7.5 mmol) was slowly added. After stirring for 30 min, 1.43 g of 3-(4-(methoxymethoxy)phenyl)propanal (7.5 mmol) was slowly added. The reaction process was monitored by TLC. When the production of the product no longer increased, the reaction solution was poured into saturated NH4Cl, and extracted with CH2Cl2 three times, 20 mL each time. It was dried over Na2SO4, filtered by suction, the solvent was rotary evaporated, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 5:1, v:v) to obtain the compound 1-(4-(trimethylsilylpropargyloxy)phenyl)-7-(4-(methoxymethoxy)phenyl)hept-1,4-dien-3-one. 1313C NMR (400 MHz, DMSO-d6): δ 141.8878, 125.2282, 198.9147, 48.0338, 66.4986, 39.3024, 30.4486, 127.6591, 130.0702, 116.0783, 159.0908, 135.3759, 129.1618, 115.2071, 154.8109, 56.3365, 100.9693, 92.1341, 93.9323, 55.4008, -0.3816.

[0155] 10. Preparation of 1-(4-(Propargyloxy)phenyl)-7-(4-hydroxyphenyl)-hepta-1,4-dien-3-one

[0156] To a 500 mL round-bottom flask, 0.5 g (0.001 mol) of 1-(4-(Trimethylsilylpropargyloxy)phenyl)-7-(4-(methoxymethoxy)phenyl)-hepta-1,4-dien-3-one was added successively. 10 mL of THF and 0.1 g of tetraethylammonium fluoride were added. The reaction progress was monitored by TLC. After the raw materials reacted completely, 1 mL of 20% sulfuric acid was added to the reaction solution and reacted at room temperature. The reaction progress was monitored by TLC. After most of the raw materials were consumed, 15 mL of saturated NaCl was added, and extracted with EtOAc three times, 20 mL each time. Dried over Na2SO4, filtered by suction, the solvent was rotary evaporated, and the crude product was separated and purified by ODS column chromatography (MeOH:H2O = 55:1 → 70:1, v:v) to obtain the compound 1-(4-(Propargyloxy)phenyl)-7-(4-hydroxyphenyl)-hepta-1,4-dien-3-one. 13 13C NMR (400 MHz, DMSO-d6): δ 142.2744, 123.1602, 188.3628, 129.5167, 147.0739, 32.9478, 34.1417, 127.9181, 130.27, 115.3062, 159.0627, 131.0757, 129.208, 115.1189, 155.4989, 55.5935, 78.9606, 78.5612.

[0157]

[0158] Example 3: Preparation method of diarylheptanone alkynes compounds with aldehyde end carrying alkyne.

[0159] 1. Preparation of 4-(Methoxymethoxy)-benzaldehyde

[0160] 10 g (0.082 mol) of 4-hydroxybenzaldehyde, 50 mL of CH2Cl2 were successively added to a 500 mL round-bottomed flask. After stirring well, 20 mL (0.115 mol) of DIPEA was added under ice bath conditions. After reacting for 15 min, about 20 mL (0.24 mol) of bromomethyl methyl ether was added to the bottom of the liquid surface, and the reaction was carried out under ice bath conditions. The reaction process was monitored by TLC. When the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl, extracted 3 times with 50 mL of EtOAc each time, dried over Na2SO4, filtered by suction, the solvent was rotary evaporated, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 5:1, v:v) to obtain compound 4-(methoxymethoxy)-benzaldehyde.

[0161] 2. Preparation of 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one

[0162] 10 g (0.06 mol) of 4-(methoxymethoxy)-benzaldehyde and 50 mL of acetone were added to a 500 mL round-bottomed flask. 5 mL of 10% NaOH was added under ice bath conditions. The reaction process was monitored by TLC. After the raw materials reacted completely, the pH was adjusted to 6, extracted 3 times with 50 mL of EtOAc each time, dried over Na2SO4, filtered by suction, the solvent was rotary evaporated, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 10:1, v:v) to obtain compound 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one. 1 1H NMR (600 MHz, MeOD) δ 9.16 (s, 1H), 7.42 (d, J = 7.5 Hz, 2H), 7.36 (dd, J = 12.0, 4.8 Hz, 4H), 7.30 (d, J = 7.4 Hz, 1H), 6.95 (d, J = 8.7 Hz, 2H), 6.70 (d, J = 16.3 Hz, 1H), 6.25 (d, J = 16.3 Hz, 1H), 5.07 (d, J = 6.4 Hz, 2H), 1.90 (s, 3H), 1.52 (s, 3H).

[0163] 5. Preparation of methyl 3-(4-hydroxyphenyl)propionate

[0164] 10 g (0.072 mol) of 4-hydroxybenzoic acid and 150 mL of methanol were successively added to a 500 mL round-bottomed flask, stirred at 35 °C, and 5 drops of concentrated H2SO4 were added to the reaction solution. The reaction process was monitored by TLC. After the raw materials reacted completely, 100 mL of water was added to quench the reaction, and the pH was adjusted to 6 with saturated NaHCO3. Methanol was removed by rotary evaporation, extracted 3 times with 10 mL of EtOAc each time, dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated to obtain compound methyl 3-(4-hydroxyphenyl)propionate.

[0165] 6. Preparation of methyl 3-(4-(trimethylsilylpropynyloxy)phenyl)propionate

[0166] To a 500 mL round-bottom flask, 0.50 g (2.77 mmol) of methyl 3-(4-hydroxyphenyl)propionate, 1.54 g of K2CO3, 1.9 mL (6.31 mmol) of N,N-diisopropylethylamine, and 50 mL of acetone were added successively. After stirring well, 0.7 mL (4.39 mmol) of propargyl bromide was added under ice bath conditions. The reaction was carried out at 60 °C. When the production of the product no longer increased, the reaction solution was poured into a large amount of saturated NH4Cl, and extracted with EtOAc three times, 50 mL each time. The solvent was rotary evaporated, dried over Na2SO4, filtered by suction, and the crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 10:1, v:v) to obtain methyl 3-(4-(trimethylsilylpropynyloxy)phenyl)propionate.

[0167] 7. Preparation of 3-(4-(trimethylsilylpropynyloxy)phenyl)propan-1-ol

[0168] To a 500 mL round-bottom flask, 4 g (13.8 mmol) of methyl 3-(4-(trimethylsilylpropynyloxy)phenyl)propionate and 100 mL of THF were added successively. Stirring was carried out under ice bath. 1.5 g of lithium aluminum hydride was slowly added to THF to form a suspension, and the lithium aluminum hydride suspension was slowly added to the reaction solution. The reaction progress was monitored by TLC. After the raw materials reacted completely, water was slowly added dropwise to the reaction solution to quench the reaction, and extracted with EtOAc three times, 50 mL each time. Dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated to obtain 3-(4-(trimethylsilylpropynyloxy)phenyl)propan-1-ol.

[0169] 8. Preparation of 3-(4-(trimethylsilylpropynyloxy)phenyl)propanal

[0170] To a 500 mL round-bottom flask, 4 g (15.2 mmol) of 3-(4-(trimethylsilylpropynyloxy)phenyl)propan-1-ol, 100 mL of CH2Cl2, and 11 g of NaHCO3 were added successively. The reaction was carried out under ice bath conditions for 30 min, and 20 g of Dess-Martin periodinane (47.2 mmol) was added. The reaction progress was monitored by TLC. After the raw materials reacted completely, the reaction was quenched with saturated Na2S2O3, extracted with EtOAc three times, 100 mL each time. Dried over Na2SO4, filtered by suction, and the solvent was rotary evaporated. The crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 20:1, v:v) to obtain 3-(4-(trimethylsilylpropynyloxy)phenyl)propanal.

[0171] 9. Preparation of 1-(4-(methoxymethoxy)phenyl)-7-(4-(trimethylsilylpropynyloxy)phenyl)hepta-1,4-dien-3-one

[0172] To a 500 mL round-bottom flask, 2 g (9.7 mmol) of 4-(4-(methoxymethoxy)phenyl)but-3-en-2-one, 30 mL of anhydrous THF were added successively under N₂ protection. The temperature was lowered to -78 °C, and 9.7 mL of LiHMDS (9.7 mmol) was slowly added. After stirring for 30 min, 2.5 g of 3-(4-(trimethylsilylpropynyloxy)phenyl)propanal (9.7 mmol) was slowly added. The reaction process was monitored by TLC. When the product no longer increased, the reaction solution was poured into saturated NH₄Cl, extracted 3 times with 20 mL of CH₂Cl₂ each time, dried over Na₂SO₄, filtered by suction, and the solvent was rotary evaporated. The crude product was separated and purified by silica gel column chromatography (PE:EtOAc = 5:1, v:v) to obtain the compound 1-(4-(methoxymethoxy)phenyl)-7-(4-(trimethylsilylpropynyloxy)phenyl)hepta-1,4-dien-3-one.

[0173] 10. Preparation of 1-(4-hydroxyphenyl)-7-(4-(propynyloxy)phenyl)hepta-1,4-dien-3-one

[0174] To a 500 mL round-bottom flask, 2 g (0.0041 mol) of the compound 1-(4-(methoxymethoxy)phenyl)-7-(4-(trimethylsilylpropynyloxy)phenyl)hepta-1,4-dien-3-one, 10 mL of THF, and 0.1 g of tetraethylammonium fluoride were added. The reaction process was monitored by TLC. After the raw materials reacted completely, 1 mL of 20% sulfuric acid was added to the reaction solution, and the reaction was carried out at room temperature. The reaction process was monitored by TLC. After most of the raw materials were consumed, 15 mL of saturated NaCl was added, and the mixture was extracted 3 times with 20 mL of EtOAc each time, dried over Na₂SO₄, filtered by suction, and the solvent was rotary evaporated. The crude product was separated and purified by ODS column chromatography (MeOH:H₂O = 55:1 → 70:1, v:v) to obtain the compound 1-(4-hydroxyphenyl)-7-(4-(propynyloxy)phenyl)hepta-1,4-dien-3-one.

[0175]

[0176] Comparative Example 1

[0177] 1. Methyl 3-(4-(trimethylsilylpropynyloxy)phenyl)propionate

[0178] Dissolve p - hydroxybenzoate in CH₂Cl₂ solution, stir evenly, add N,N - diisopropylethylamine at room temperature, add TMS - propargyl bromide, and conduct a catalytic reaction at room temperature - 70 °C. Monitor the reaction by TLC, and no target product appears. As time increases, the TMS group of TMS - propargyl bromide gradually drops off. Try changing the base to inorganic base potassium carbonate, monitor the reaction by TLC, and still no target product appears. As time increases, the TMS group of TMS - propargyl bromide gradually drops off. Therefore, the subsequent steps cannot be carried out.

[0179] With the double - base strategy in the above - mentioned embodiment, by providing an alkaline environment synergistically with potassium carbonate and N,N - diisopropylethylamine, adding TMS - propargyl bromide, the reaction can occur rapidly at 40 - 60 °C, and the reaction can reach the maximum within 1 h, with a yield of about 50%. Within this time, the TMS group of TMS - propargyl bromide exists almost stably.

[0180] 2. 4 - (4 - (Trimethylsilylpropynyloxy)phenyl)but - 3 - en - 2 - one

[0181] Add 0.50 g (3.08 mmol) of 4 - (4 - hydroxyphenyl)but - 3 - en - 2 - one, 1.54 g (0.011 mmol) of K₂CO₃, and 20 mL of acetone to a 500 - mL round - bottom flask in sequence. After stirring well, add 0.7 mL of TMS - propargyl bromide under ice - bath conditions, and try to catalyze the reaction at different temperatures (4 °C - 60 °C). Monitor the reaction by TLC, and no target product appears. As time increases, the TMS group of TMS - propargyl bromide gradually drops off. Try changing the base to inorganic base potassium carbonate, monitor the reaction by TLC, and still no target product appears. As time increases, the TMS group of TMS - propargyl bromide gradually drops off.

[0182] In summary, the method for preparing diarylheptanone alkynes compounds of the present invention is simple in operation, high in purity and yield, filling the blank in the synthesis of this compound.

[0183] Application Example 1: Comparison of Compound Activities

[0184] 1. Cell culture and drug administration. After counting KMS - 11, A549, and HepG2 cells respectively, seed about 10000 cells per well in a 96 - well plate, incubate at 37 °C until the cells adhere. Set different concentrations of the compound obtained in Example 1 above (dissolved in DMSO) and DHDK (dissolved in DMSO). Discard the blank medium in the original 96 - well plate, and then add 200 μL of drug - containing medium and incubate for 24 h.

[0185] 2. Cell viability detection. The CCK-8 method was used for cell viability detection. 20 μL of CCK-8 solution was added to each well and incubated at 37 °C for 2 h, and the absorbance was measured at 450 nm.

[0186] 3. Calculation of IC 50 . The cell survival rate was calculated using the absorbance values and imported into data processing and graphing software such as GraphPad to calculate the corresponding IC 50 value (see Table 1).

[0187] Table 1

[0188]

[0189] As shown in Table 1, the IC 50 values of the compound obtained in Example 1 above for these three cells were all lower than those of DHDK. The IC 50 value is the half-maximal inhibitory concentration, which is used to measure the effect of a drug in inhibiting the growth of tumor cells. The lower the IC 50 value, the lower the concentration of the drug required to inhibit the growth of 50% of tumor cells, and the better the drug efficacy. Therefore, compared with the diarylheptanone compound DHDK, the diarylheptanone alkyne compound has similar pharmacological activities and better anti-tumor effects, and can be further developed as a drug nucleus click chemical probe.

[0190] Application Example 2: Drug target competition cell imaging experiment

[0191] 1. Cell culture.

[0192] Add 14 mm sterile coverslips into a 6-well plate, add 2 mL of complete medium containing 500,000 PC3 cells, mix well, and culture in an incubator for 24 h. When the cells are adherent and in good growth condition, divide the PC3 cells into a DMSO group, the compound group obtained in Example 1 above, and a competition group (the competition group is DHDK and the compound obtained in Example 1) to start the experiment. Discard the medium in the 6-well plate, add PBS and rinse 3 times. Add 2 mL of complete medium (containing 5 μL of DMSO) to the cells in the DMSO group, add 2 mL of complete medium containing 6 μM of the compound obtained in Example 1 above to the dosing group, and add 10-fold amount of DHDK of the compound obtained in Example 1 to the competition group in advance. After incubating for 1 h, add the compound obtained in Example 1 and incubate for 1 h. Discard the medium, wash once with PBS, add 1 mL of 4% paraformaldehyde, fix for 10 min, wash 3 times with PBS, 5 min each time. Dropwise add 100 μL of 0.1% Triton-X100 and incubate at room temperature for 10 min. Wash 3 times with PBS, 5 min each time. Add 500 μL of click reaction mixture (containing 50 μM azide Cy5 aqueous solution, 50 μM BTTAA aqueous solution, 100 μM TCEP aqueous solution and 200 μM CuSO4 aqueous solution) and 2.5 mM sodium ascorbate to each well, and react at room temperature in the dark for 2 h. After the reaction is completed, wash three times with PBS, add 100 μL of DAPI working solution, act in the dark for 2 min, discard, wash 3 times with PBS, 5 min each time. Take out the coverslips and place them on glass slides, seal with a mounting medium, and wait for measurement.

[0193] 2. Observation with a confocal microscope

[0194] It can be seen from Figure 2 that the compound obtained in Example 1 above can enter cells and has the ability of intracellular fluorescence imaging, showing the localization and distribution of the compound obtained in Example 1 in cells. Among them, blue is the cell nucleus, red is the probe target, and the Merge figure is the co-localization figure of the two, indicating that the two are in the same sample. Horizontally, the three figures from left to right are the fluorescence figures of the cell nucleus, the probe target, and the co-localization of the target and the cell nucleus of a sample respectively. And vertically, the three groups of figures are the fluorescence figures of three samples, namely DMSO, the probe group, and the competition group. From Figure 2It was observed that in the above Example 1, for the compound obtained and the DMSO group, the probe group had obvious red color while DMSO had no red color, indicating that the compound obtained in the above Example 1 could specifically bind to the intracellular target; when both the compound obtained in the above Example 1 and the diarylheptanone anti-tumor compound DHDK were present in the system, the red color of the compound obtained in the above Example 1 significantly decreased, indicating that in the competition group, the compound DHDK would significantly occupy the binding site of the compound obtained in Example 1, resulting in a significant reduction in its binding to the target, indicating that the compound obtained in the above Example 1 had the same action target as this drug, and the diarylheptanone alkyne compound could be used to conduct "target fishing" research on the diarylheptanone anti-tumor compound. Application Example 3: Immunofluorescence co-localization experiment

[0195] 1. Cell culture.

[0196] Add 14 mm sterile coverslips into a 6-well plate, add 2 ml of complete medium containing 500,000 PC3 cells, mix well, and culture in an incubator for 24 h. When the cells are adherent and in good growth state, divide the PC3 cells into the DMSO group and the compound group obtained in the above Example 1 to start the experiment. Discard the medium in the 6-well plate, add PBS and rinse 3 times. Add 2 mL of complete medium (containing 5 μL of DMSO) to the cells in the DMSO group, and add 2 mL of complete medium containing 6 μM of the compound obtained in the above Example 1 to the dosing group. Incubate for 1 h, discard the medium, wash once with PBS, add 1 mL of 4% paraformaldehyde, and fix for 10 min. Wash 3 times with PBS, 5 min each time. Dropwise add 100 μl of 0.1% Triton-X100 and treat at room temperature for 10 min. Wash 3 times with PBS, 5 min each time. Add 500 μL of click reaction mixture (containing 50 μM of azide Cy5 aqueous solution, 50 μM of BTTAA aqueous solution, 100 μM of TCEP aqueous solution and 200 μM of CuSO4 aqueous solution) and 2.5 mM of sodium ascorbate to each well, and react at room temperature in the dark for 2 h. After the reaction, wash three times with PBS, add 100 μL of DAPI working solution, react in the dark for 2 min, discard, wash 3 times with PBS, 5 min each time. Take out the coverslips and place them on glass slides, seal with a mounting medium, and wait for measurement.

[0197] 2. Observation under a confocal microscope

[0198] From Figure 3It can be seen that the compound obtained in the above Example 1 can enter cells, has the ability of intracellular fluorescence imaging, and is in the same position as the protein PRDX1 in cells, enabling co-localization. Among them, blue represents the cell nucleus, red represents the probe target, green represents the antibody of PRDX1, and the Merge image is the co-localization image of the three, indicating that the three are in the same sample. The three horizontal images from left to right are the fluorescence co-localization images of the cell nucleus, probe target, PRDX1 protein, and the Merge of the three in a cell sample. The two vertical groups of images are the fluorescence images of two samples, DMSO and the probe. The DMSO group does not have red, but has blue and green, indicating that there are cell nuclei and PRDX1 protein in this cell sample, but there is no case of target binding between DMSO and this cell sample, excluding the influence of the reagent. The compound obtained in the above Example 1 has blue, red, and green, indicating that the compound obtained in Example 1 above can fish out the target in this cell sample, and at the same time the target can achieve co-localization with the PRDX1 protein, that is, it indicates that PRDX1 is the target protein of DHDK, but not limited to this.

[0199] Application Example 4: Gel Imaging

[0200] 1. Cell culture. One day in advance, seed KMS-11 cells in a 6-cm cell culture dish containing 6 mL of culture medium at an inoculation amount of 5 million cells per well. Discard the culture medium when the cells adhere to the dish and reach a density of 80% the next day. Set up groups including the DMSO group and the compound group obtained in the above Example 1. Among them, the low concentration of the compound component is 6 μM and the high concentration is 18 μM. After each group is washed 3 times with PBS, add 6 mL of complete medium containing 5 μL of DMSO to the cells in the DMSO group, add 6 mL of complete medium containing 6 μM of this compound to the low-concentration compound group obtained in the above Example 1, and add 6 mL of complete medium containing 18 μM of this compound to the high-concentration compound group obtained in the above Example 1. Incubate in a 37 °C cell incubator for 2 h.

[0201] 2. Protein extraction and quantification. Blow the cells off the culture dish, centrifuge, discard the supernatant, add an appropriate amount of PBS to wash the cells, repeat the washing twice, then add an appropriate amount of PBS solution and protease inhibitor. After lysing the cells with a cell ultrasonic disruptor, centrifuge in a 4 °C centrifuge (12,000 r, 30 min), take the supernatant, and perform protein quantification by the BCA method.

[0202] 3. Click Chemistry. Preparation of reaction samples: Take 1000 μg of the protein obtained from the above extraction and make up the volume to 1 mL with PBS solution pre-cooled at 4°C. Add 50 μL of the click reaction mixture to each sample to make the final concentration of the click reaction solution contain azide Cy5 aqueous solution (50 μM), BTTAA aqueous solution (50 μM), TCEP aqueous solution (100 μM), CuSO4 aqueous solution (200 μM), and 2.5 mM sodium ascorbate. Place it on a shaker and react for 2 h, during which it should be protected from light.

[0203] 4. Protein precipitation and dissolution: After the reaction is completed, transfer the reaction solution to a 10 mL centrifuge tube, add 7 mL of acetone pre-cooled at -20°C, and place it in a -20°C refrigerator overnight to precipitate the protein for 12 h, during which it should also be protected from light. After precipitation, centrifuge at 3500 rpm for 5 min, discard the supernatant, collect the precipitate into a 1.5 mL EP tube, wash it twice with acetone pre-cooled at -20°C, centrifuge at 4°C and 4000 g for 10 min, discard the supernatant, and dry the precipitate. After drying, add 20 μL of 1.2% SDS solution to dissolve the protein.

[0204] 5. Gel electrophoresis: Add buffer to the dissolved protein according to the ratio of protein SDS solution: Loading buffer = 4:1, denature at 90°C for 5 min, then load the sample and perform electrophoresis.

[0205] 6. Development: After the electrophoresis is completed, remove the gel, perform gel imaging, and observe and record the red fluorescence corresponding to different probe concentrations.

[0206] It can be seen from Figure 4 that when performing development imaging under a gel imaging system, the amount of protein that can be labeled increases with the increase in the concentration of the compound obtained from the above Example 1, and in situ protein labeling of cells is carried out in a dose-dependent manner, indicating that considering various factors comprehensively, an appropriate probe concentration is selected for subsequent experiments.

[0207] Application Example 5: Chemical Proteomics

[0208] 1. Cell culture. 10 million KMS-11 cells were seeded in a 10-cm cell culture dish and cultured with 10 mL of RPMI 1640 complete medium. The cells were cultured at 37 °C in a CO2 cell incubator and used for the experiment when they grew to more than 80% and were in good condition. The DMSO group, the compound obtained in Example 1 above, and the competition group (DHDK containing 10-fold amount of the compound obtained in Example 1 and the compound obtained in Example 1) were set up to start the experiment. The medium in the 10-cm dish was discarded and the cells were rinsed 3 times with PBS: (1) 10 mL of RPMI 1640 complete medium (containing 5 μL of DMSO) was added to the cells in the DMSO group; (2) 10 mL of RPMI 1640 complete medium containing 6 μM of the compound in Example 1 above was added to the dosing group; (3) 10-fold amount of DHDK obtained in Example 1 was added to the competition group in advance, and after incubation for 1 h, the compound obtained in Example 1 was added and incubated for 1 h.

[0209] 2. Protein extraction and quantification. After discarding the medium, the cells were washed once with PBS, then the cells were detached from the culture dish by pipetting, centrifuged, and the supernatant was discarded. The cells were transferred to a centrifuge tube, washed with an appropriate amount of PBS twice, then an appropriate amount of PBS solution and protease inhibitor were added, and the cells were lysed with a cell sonicator. The lysate was centrifuged at 12,000 r in a 4 °C centrifuge for 30 min, and the supernatant was taken for protein quantification by the BCA method.

[0210] 3. Click chemistry. Preparation of reaction samples: 1000 μg of the protein obtained above was diluted to 1 mL with 4 °C pre-cooled PBS solution. 50 μL of click reaction mixture was added to each sample to make the final concentration of the click reaction solution contain azide Cy5 aqueous solution (50 μM), BTTAA aqueous solution (50 μM), TCEP aqueous solution (100 μM), CuSO4 aqueous solution (200 μM), and sodium ascorbate (2.5 mM). The reaction was carried out on a shaker for 2 h in the dark. After the reaction was completed, the sample was passed through a protein desalting column to remove excess biotin.

[0211] 4. Protein precipitation and dissolution. Cold acetone was added to the sample and precipitated overnight. The supernatant was discarded after centrifugation. The protein precipitate was washed twice with cold methanol. 1.2% SDS was added to the protein precipitate to dissolve it, and then PBS was added to make the volume up to 1 mL.

[0212] 5. Interaction with streptavidin. The dissolved protein was added to the washed magnetic beads and mixed at room temperature for 1 h, then adsorbed with a magnetic stand and the supernatant was aspirated. After washing the magnetic beads again, the disulfide bonds were reduced with DTT and the thiols were alkylated with IAA. Buffer and trypsin were added to the magnetic beads, adsorbed with a magnetic stand, and the supernatant was taken, desalted, concentrated, and re-dissolved. The peptide fragments were analyzed by LC-MS in combination with computer software to infer the target protein (see Figure 5 ).

[0213] The results are as Figure 5 shown in the heat map. The above-mentioned protein did not pull out the above-mentioned target in the DMSO group, indicating that there was no non-specific adsorption of the above-mentioned protein in the DMSO group; it could be pulled out in the probe group, indicating that the probe could specifically pull out the relevant protein; the protein content in the competition group mostly decreased significantly, indicating that the ability of the probe to capture the target could be blocked by DHDK, indicating that the proteins pulled out by the probe were the target proteins of DHDK.

[0214] Application Example 6: Magnetic bead pull-down experiment 1. Cell culture. 10 million KMS-11 cells were seeded in a 10-cm cell culture dish and cultured with 10 mL of RPMI 1640 complete medium. The cells were cultured at 37 °C in a CO2 cell incubator. When they grew to more than 80% and were in good condition, they were used for this experiment. Set up the DMSO group, the compound obtained in Example 1 above, and the competition group to start the experiment. Discard the medium in the 10-cm dish and add PBS to wash it 3 times: (1) Add 10 mL of RPMI 1640 complete medium (containing 5 μL of DMSO) to the cells in the DMSO group, (2) Add 10 mL of RPMI 1640 complete medium containing 6 μM of the compound obtained in Example 1 above to the drug administration group, (3) Add 10-fold amount of DHDK obtained in Example 1 to the competition group in advance. After incubating for 1 h, then add the compound obtained in Example 1 and incubate for 1 h;

[0215] 2. Protein extraction and quantification. Blow the cells off the culture dish, centrifuge, discard the supernatant, transfer to a centrifuge tube, add an appropriate amount of PBS to wash the cells, repeat twice, then add an appropriate amount of PBS solution and protease inhibitor, break the cells with a cell ultrasonic disruptor, centrifuge in a 4 °C centrifuge (12,000 r, 30 min), take the supernatant, and perform protein quantification by the BCA method.

[0216] 3. Click chemistry. Sample preparation: Take 1000 μg of the protein obtained by the above extraction, make up the volume to 1 mL with pre-cooled PBS solution at 4 °C, add 50 μL of click reaction mixture to each sample, so that the final concentration of the click reaction solution contains azide Cy5 aqueous solution (50 μM), BTTAA aqueous solution (50 μM), TCEP aqueous solution (100 μM), CuSO4 aqueous solution (200 μM), and 2.5 mM sodium ascorbate, and place it on a shaker to react for 2 h, during which it should be protected from light. After the reaction is completed, pass the sample through a protein desalting column to remove excess biotin.

[0217] 4. Protein precipitation and dissolution. Add cold acetone to the sample, precipitate overnight, centrifuge and discard the supernatant, and wash the protein precipitate twice with methanol. Add 1.2% SDS to the protein precipitate to dissolve the protein precipitate.

[0218] 5. Pull-down experiment. Take 150 μl of magnetic beads and put them into a 2-ml EP tube. Add 1.5 ml of PBS, adsorb with a magnetic stand, discard the supernatant, and repeat this process 5 times. Add the dissolved protein to the magnetic beads, mix at room temperature for 2 h, adsorb with a magnetic stand, and discard the supernatant. Add PBS solution with different concentrations of SDS to the magnetic beads, wash the magnetic beads, and then discard the supernatant.

[0219] 6. Treat the magnetic beads. Resuspend the enriched magnetic beads with 0.5 mL of PBS, adsorb with a magnetic stand, collect the supernatant and label it. Then resuspend the magnetic beads with 0.5 mL of PBS again, heat in a 90°C water bath for 5 min, collect the supernatant and label it.

[0220] 7. Gel preparation and electrophoresis. Prepare a 10% polyacrylamide gel. After heating the two batches of supernatant at 90°C in a water bath for 5 min, load the sample according to the ratio of loading buffer: protein solution (4:1). Start electrophoresis at a voltage of 70 V until the protein sample enters the separating gel, then change to 120 V until the blue band formed by bromophenol blue enters the bottom of the gel, and then stop electrophoresis.

[0221] 8. Electrotransfer. After electrophoresis, cut a PVDF membrane of appropriate size, activate it in methanol, and perform membrane transfer according to the "sandwich structure", and transfer the membrane at a constant current of 250 mA according to the molecular weight of the target protein.

[0222] 9. Blocking and incubation. After membrane transfer, take out the PVDF membrane, block it in skim milk powder for 1.5 h, wash it 3 times with TBST, 10 min each time, add the primary antibody, incubate overnight at 4°C, wash it 3 times with TBST, 10 min each time, add the secondary antibody, incubate at room temperature for 2 h, and wash it 3 times with TBST, 10 min each time.

[0223] 10. Exposure. Prepare an appropriate volume of ECL developing solution (the volume ratio of solution A and solution B is 1:1) according to the size of the PVDF membrane. Place the PVDF membrane on the platform of the exposure machine, add the developing solution that completely covers the PVDF membrane above the membrane, and perform exposure.

[0224] The experimental results are as Figure 6As shown, the target protein was not detected in the DMSO group, indicating that PRDX1 protein could not be retained on the magnetic beads and there was no non-specific adsorption; the probe group could pull out PRDX1 protein in cells, indicating that the probe could covalently bind to PRDX1, and PRDX1 was the potential target of the probe; the amount of PRDX1 pulled out by the probe in the competition group was significantly reduced, indicating that DHDK (the parent compound of the probe) could effectively reduce the ability of the probe to pull out PRDX1, indicating that DHDK would occupy the site where the probe binds to PRDX1, that is, PRDX1 was the target of DHDK, and this probe could be used for DHDK target fishing. The above experiments corroborated each other and jointly proved that the diarylheptanone alkynyl probe prepared by the present invention could be used for the target confirmation of this anti-tumor compound, enhancing the reliability of the results and laying a foundation for the study of the action mechanism of this anti-tumor compound.

[0225] In summary, the compounds of the present invention have anti-tumor effects and have good pharmacological effects on solid tumors such as colon cancer, skin cancer, prostate cancer, liver cancer, neuroblastoma, breast cancer, etc. and hematological tumors such as multiple myeloma and monocytic macrophage leukemia. In addition, it can also be used as a probe for detecting the target protein of anti-tumor compounds to complete the study of the anti-cancer mechanism of this anti-tumor compound. The described target protein detection methods include gel imaging, immunoblotting, proteomics, pull down experiments and magnetic bead detection.

[0226] The above embodiments only represent several embodiments of the present invention, and the description is relatively specific and detailed, but it should not be construed as a limitation to the scope of the patent application.

Claims

1. A diarylheptanone alkyne compound, characterized in that, The compound has the structure shown below: In the formula, R1 and R2 can be the same or different and are independently selected from H, C3-C7 alkynyl which is unsubstituted or substituted by at least one R5, and at least one of R1 and R2 is selected from C3-C7 alkynyl which is unsubstituted or substituted by at least one R5; R3 and R4 can be the same or different and are independently selected from H, halogen, C1-C4 alkyl, C1-C4 alkanoyl, C1-C4 alkoxy or hydroxy; R5 is selected from halogen, C1-C4 alkyl, C1-C4 alkanoyl, C1-C4 alkoxy or hydroxy; x and n can be the same or different and are independently selected from 1, 2, 3 or 4, and x + a ≤ 5; a and b can be the same or different and are independently selected from 1, 2, 3 or 4, and n + b ≤ 5.

2. The diarylheptanone alkyne compound according to claim 1, wherein In the structural formula of the said compound, R1 and R2 can be the same or different and are independently selected from H, C3-C7 alkynyl, and at least one of R1 and R2 is selected from C3-C7 alkynyl; R3 and R4 can be the same or different and are independently selected from H, halogen, C1-C4 alkyl, C1-C4 alkanoyl, C1-C4 alkoxy or hydroxy; x and n can be the same or different and are independently selected from 1, 2, 3 or 4, and x + a ≤ 5; a and b can be the same or different and are independently selected from 1, 2, 3 or 4, and n + b ≤ 5.

3. The diarylheptanone alkyne compound according to claim 2, characterized in that, In the structural formula of the said compound, R1 and R2 can be the same or different and are independently selected from H, C3-C7 alkynyl, and at least one of R1 and R2 is selected from C3-C7 alkynyl; x and n can be the same or different and are independently selected from 1, 2, 3, 4 or 5.

4. The diarylheptanone alkyne compound according to claim 3, wherein The said compound is 5. A method for preparing the diarylheptanone alkyne compound according to claim 1, characterized in that: (1) Esterify the hydroxycinnamic acid with a side chain, and then perform nucleophilic substitution, reduction and oxidation reactions to obtain an aldehyde product; (2) Subject the hydroxybenzaldehyde with a side chain to a nucleophilic reaction and aldol condensation to obtain a product; (3) Perform aldol condensation and deprotection on the products obtained in steps (1) and (2) to obtain a terminal alkyne structure, namely the diarylheptanone alkyne compound; In the above steps (1) and (2), the nucleophilic substitution is to protect the phenolic hydroxyl group or introduce a silicon-protected haloalkyne; and in at least one step, a silicon-protected haloalkyne is introduced.

6. The preparation method of the diarylheptanone alkyne compound according to claim 5, characterized in that: In step (2), the hydroxybenzaldehyde with a side chain undergoes phenolic hydroxyl group protection through a nucleophilic reaction, and then aldol condensation to obtain a product; or, the obtained product is deprotected, and then a silicon-protected haloalkyne is introduced through a nucleophilic reaction to obtain a product.

7. The preparation method of the diarylheptanone alkyne compound according to claim 5 or 6, characterized in that: The nucleophilic substitution reaction in steps (1) and (2) is When protecting the phenolic hydroxyl group, the esterified product of the hydroxycinnamic acid with a side chain or the benzaldehyde with a side chain is used as a raw material, and in an alkaline environment, it is respectively subjected to a nucleophilic substitution reaction with a phenolic hydroxyl group protecting group at 4°C - 100°C for 1h - 10h, wherein the molar ratio of the raw material to the base substance providing the alkaline environment is 1:0.1eq - 1:10eq; the molar ratio of the raw material to the phenolic hydroxyl group protecting group is 1:1eq - 1:10eq; When introducing a terminal alkyne, the esterified product of hydroxycinnamic acid with a side chain, hydroxybenzaldehyde with a side chain, or the product after phenolic hydroxyl protection, aldol condensation, and then deprotection of hydroxybenzaldehyde with a side chain is used as a raw material to carry out a nucleophilic substitution reaction with a silicon-protected haloalkyne at 4 °C - 100 °C for 0.16 h - 2 h to introduce the terminal alkyne. Among them, the molar ratio of the raw material to the base substance providing the alkaline environment is 1:0.1 eq - 1:10 eq; the molar ratio of the raw material to the silicon-protected haloalkyne is 1:1 eq - 1:10 eq.

8. Use of the diarylheptanone alkyne compound according to claim 1, characterized in that: Use of the diarylheptanone alkyne compound in the preparation of anti-tumor drugs.

9. Use of the diarylheptanone alkyne compound according to claim 1, characterized in that: Use of the diarylheptanone alkyne compound in biological imaging.

10. A probe for identifying protein targets for anti-tumor, characterized in that: The probe is the compound described in claim 1.

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