JP2021001158A - Synthesis method of methylene ether/urushiol/hydroxamic acid derivative having hdac inhibitory activity - Google Patents

Abstract

To provide a methylene urushiol hydroxamic acid derivative having HDAC inhibitory activity.SOLUTION: A methylene ether urushiol hydroxamic acid derivative contains 5-(10-(benzo[d][1,3]dioxolane-4-yl)decane-2-en-1-yl)-N-hydroxyl-2-methylcyclohexane-3-en-1-formamide, 5-(10-(benzo[d][1,3]dioxolane-4-yl)-3-hydroxyl-decane)-N-hydroxyl-2-methylcyclohexane-3-en-1-formamide and 5-(10-(7-nitrobenzo[d][1,3]dioxolane-4-yl)decane-2-en-1-yl)-N-hydroxyl-2-methylcyclohexane-3-en-1-formamide.SELECTED DRAWING: None

Description

The present invention belongs to the field of drug synthesis technology, and relates to a method for synthesizing a methylene ether / urushiol / hydroxamic acid derivative and its HDAC inhibitory activity.

Urushiol is a naturally active ingredient in the secretion lacquer of poison ivy (Toxicodendron verniciflum (Stokes) FA Barkl.), An important forest product in China, and 85% of the lacquer in the world is produced from China. Urushiol is an alkylphenolic compound with a pyrocatechol structure, the side chain of which is a C15 alkane with different saturation. Ursiol has excellent antitumor bioactivity and has an inhibitory effect on 9 types of organs and 29 types of tumor cells in the human body. The mechanism of action is tumor apoptosis induction, tumor cell growth inhibition, and tumor angiogenesis. Includes inhibition, inhibition of nuclear transcription factors, tumor cell poisoning, etc. Urushiol has a history of thousands of years in Japan as a Chinese herbal medicine for tumor adjuvant treatment. Therefore, although it is highly expected that urushiol will be developed as an anticancer agent, unsaturated urushiol has an unstable chemical structure, is easily oxidized and polymerized, and greatly reduces its antitumor therapeutic effect. It limits the development and application of urushiol as antitumor.

Histone deacetylase (HDAC) is an important target for cancer treatment recognized at home and abroad. The imbalance of histone acetylation due to HDAC abnormalities is closely related to tumor development and development, and overexpression of HDAC has already been found in a large number of tumor cells. By altering intracellular acetylation levels, HDAC inhibitors generate antitumor activity in terms of regulating gene translation processes, suppressing cell growth, inducing apoptosis or differentiation, and suppressing tumor cell vascular regeneration. Therefore, the design and development of low-toxicity, high-efficiency HDAC inhibitor-based analogs has already become a hotspot for antitumor targeting drug research in Japan and overseas. HDACs are classified into four types, such as hydroxamic acid type, benzamide type, electrophilic ketone type and cyclopeptide type, depending on their structural characteristics. Among them, hydroxamic acid-based compounds have advantages such as strong specificity, good antitumor activity, and small toxic side effects, and are currently the most extensive and deeply studied HDAC inhibitors, and are typical hydroxamic acids. The structure of the system inhibitor is divided into three parts, such as a surface recognition area, a spacer arm area, and a zinc ion bonding area, according to function. The surface recognition area consists mainly of hydrophobic segments, usually benzene ring derivatives, the sparcer arm area is a fatty chain, and the functional group of the zinc ion binding area is hydroxamic acid. Studies show that hydroxamic acid functional groups are HD
Key point of AC inhibitor It is a medicinal functional group, and the activity of HDAC inhibitor can be effectively suppressed by directly binding to the Zn2 + structure of HDAC enzyme. Currently, Mer
Vorinostat (SAHA), a hydroxamic acid-based HDAC inhibitor from ck, has already been approved by the US FDA as a therapeutic agent for lymphoma, and several other hydroxamic acid-based HDAC inhibitors are in the clinical research stage.

Studies have shown that unsaturated urushiol has a certain HDAC inhibitory activity and its structure is FDA.
It is similar in structure to the HDAC inhibitor SAHA, which has been approved for use by HDAC, but lacks the zinc ion binding area, which is a key point structural unit for HDAC inhibition. Therefore, in the present invention, unsaturated urushiol is used as a raw material, and urushiol is oxidized through an etherification reaction.
Hydroxamic acid groups are introduced into the tail of the side chain of urciol through reactions such as Diels-Alder, hydrolysis and aminohydroxylation by blocking polymerization, and nitro groups and hydroxyl groups are introduced into the benzene ring or alkyl group chain. Three types of methylene ether ursiol-hydroxamic acid derivatives were synthesized by introducing groups with different efficacy, but all three types of compounds were able to bind well to the active pocket of HDAC, and stable hydrogen bonds with their residues. Forming an interaction, forming a stable chelate with Zn2 + at the bottom of the active pocket, excellent HD
Has AC inhibitory activity and is half the inhibitory concentration of compounds 1, 2 and 3 with respect to HDAC2 (IC50)
) Are 0.27, 0.25 and 0.22 ug / mL, respectively, and the half-suppressed concentration (IC50) for HDAC8 is 0.29, 0.26 and 0.24 ug / mL, respectively, permitted by the FDA. It corresponds to the IC50 value of the HDAC inhibitor SAHA, which can be used in clinical antitumor agents, has high added value, and is clinically a new type of urushiol-based HDA.
It can be used as a new technology for developing C inhibitors.

An object of the present invention is to provide a method for synthesizing a methylene ether urushiol hydroxamic acid derivative having HDAC inhibitory activity. The method synthesized three methylene ether urushiol hydroxamic derivatives. All three compounds can bind well to the active pocket of HDAC, and the half-suppressed concentration (IC50) for HDAC2 and HDAC8 is FDA.
Equivalent to the IC50 value of the HDAC inhibitor SAHA approved by HDAC and excellent HDAC
It has inhibitory activity, and a new type of urushiol-based HDAC inhibitor can be developed and used in antitumor agents, and has extremely high added value.

1. 1. It contains compounds 1, 2 and 3, and the chemical names are 5-(10- (benzo [d] [1,3], respectively.
] Dioxolane -4-yl) Decane -2 -En -1 -Il) -N-Hydroxy-2 -Methylcyclohexane -3-En -1 -Formamide 5- (10-(Benzo [d] [1,
3] Dioxolane -4 -Il) -3 -Hydroxy-Decan) -N -Hydroxy -2-
Methylcyclohexane -3-ene -1-formamide and 5-(10-(7-nitrobenzo [d] [1,3] dioxolane -4-yl) decane-2-ene -1-yl) -N-hydroxyl -2 -Methylcyclohexane -3-ene -1-formamide, the chemical formula of which is
Compound 1
Compound 2
Compound 3
A methylene ether urushiol hydroxamic acid derivative having HDAC inhibitory activity.

2. 2. The method for synthesizing the compound 1 includes the following steps.
(1) Using compound I urushiol as a raw material, take a certain amount of urushiol and take DMF and DC.
Dissolve in M, add a certain amount of NaH, reflux under protection of argon gas for 12 hours, cool to room temperature, slowly add a certain amount of water, and extract 3 times with DCM. The organic layer is combined, washed with saturated brine, dried over anhydrous sodium sulfate, and then decompressed and evaporated.
Perform column chromatography, elute with PE, and reduce and evaporate the eluate to obtain compound II.
(2) Take a certain amount of Compound II, dissolve it in toluene, add a certain amount of acrylic acid ester, heat and reflux for 36 hours under the protection of argon gas, and reduce and evaporate the solvent.
Add a certain amount of ethanol and KOH, heat and reflux for 10 minutes, reduce the pressure and elute the solvent, add a certain amount of water, extract 3 times with EA, combine the extract and dry with anhydrous sodium sulfate. After that, the mixture is vacuumed and dried, column chromatographed, eluted with PE-EA, and the eluate is vacuumed and evaporated to obtain Compound III.
(3) Take a certain amount of Compound III, dissolve it in anhydrous DMF, add a certain amount of DIPEA, silicon-protected hydroxylamine and HATU, spend 12 hours while reacting and stirring, and reduce the pressure and elute the solvent. Perform column chromatography directly, elute with dichloromethane-methanol, reduce the pressure and evaporate the eluate, then put the product in a certain amount of TBAF-THF solution, stir at room temperature for 40 minutes, reduce the pressure and evaporate. , Direct column chromatography, elution with DCM-MeOH, decompression and evaporation of the eluate, and further purification by HPLC to give the target compound 1.
The urushiol in step (1) is derived from lacquer and is obtained by extracting with methanol using the lacquer as a raw material, and the purity of the obtained urushiol is 90% or more.
The HPLC purification method in step (3) has a chromatography column of a C18 preparation column, a fluid phase of acetonitrile-water, a ratio of 75:25 and a measurement wavelength of 210 nm.
The synthetic route for compound 1 is as follows:

3. 3. The method for synthesizing the compound 2 includes the following steps.
(1) Using compound III as a raw material, a certain amount of compound III was dissolved in dichloromethane, a certain amount of mCPBA was added, the mixture was spent for 12 hours with stirring at room temperature, and washed with saturated sodium bicarbonate three times. Dry with anhydrous sodium sulfate, reduce pressure and evaporate to obtain a colorless oil.
All of them are placed in anhydrous tetrahydrofuran, and a constant amount of 100 mg / mL lithium aluminum hydride is slowly dropped into the turbid solution, and then the temperature is raised to 70 ° C. for 2 hours to react, and then cooled to room temperature. Slowly add a certain amount of water, add a certain amount of 15% sodium hydroxide solution again, then add a certain amount of water again, filter after 30 minutes, wash with tetrahydrofuran, and reduce the pressure of the filtrate. Remove tetrahydrofuran, add a certain amount of water,
Extract 4 times with ethyl acetate, combine the extracts, wash with saturated brine, dry with anhydrous magnesium sulfate, reduce pressure and evaporate, perform silica gale column chromatography, and perform PE-E.
Elute at A and vacuum and evaporate the eluate to give compound IV.
(2) Compound IV is dissolved in a certain amount of acetone, the temperature is lowered in an ice-water bath, and a certain amount of Jones reagent (Jones reagent, chromium anhydride used as an oxidizing agent (chromium oxide (VI)).
) Is a concentrated sulfuric acid solution. Jones oxidation is a chemical reaction in which a primary or secondary alcohol is oxidized to a carboxylic acid or a ketone using chromic acid. ) And 30 at room temperature
After reacting for 1 minute, add a certain amount of isopropanol, stir for 20 minutes, add a certain amount of saturated sodium chloride solution, extract 5 times with EA, combine the extracts, and dry with anhydrous sodium sulfate. , Reduce and evaporate, perform silica gale column chromatography, PE-
Elute with EA and vacuum and evaporate the eluate to give compound V.
(3) Dissolve compound V in a certain amount of methanol, add a certain amount of NaBH ₄ , react at room temperature for 30 minutes, reduce the pressure and evaporate, then dissolve in a certain amount of EA, saturated saline solution. Wash twice with EA, extract twice with EA, combine with the extract, dry with anhydrous sodium sulfate, vacuum and evaporate the solvent, dry for 2 hours under vacuum conditions, and a certain amount of DCM, NH ₂ OTHP,
EDC. Add Cl and TEA, spend 12 hours while reacting, wash twice with 5% aqueous citric acid solution, extract twice with DCM, combine with organic layer, reduce and dry the solvent, and methanol. Add the solution, lower the temperature in an ice-water bath, add a certain amount of 10% HCl solution, react for 1 hour, then reduce and evaporate the solvent, dissolve in methanol, and filter with a micropore filtration membrane. , Further purified with HCl to obtain target compound 2.
In the HPLC purification method in step (3), the chromatography column is C.
It is an 18-prepared column, the fluid phase is acetonitrile-water, the ratio is 68:32, and the measurement wavelength is 210 nm.
The synthetic route for compound 2 is as follows:

4. The method for synthesizing the compound 3 includes the following steps.
(1) Using compound III as a raw material, a certain amount of compound III is put into newly evaporated acetic anhydride, the temperature is lowered to 0 ° C., a certain amount of anhydrous copper nitrate is added with vigorous stirring, and a certain amount is added after 2 hours. Add anhydrous copper nitrate, add a certain amount of water after 3 hours, stir for 20 minutes, and then add 2 with ethyl acetate.
Extraction is performed twice, the organic layer is combined, washed twice with saturated sodium bicarbonate solution, dried over anhydrous magnesium sulfate, reduced pressure and evaporated to obtain a pale yellow oily substance.
(2) The pale yellow oil obtained from step 1 is dissolved in methanol, a certain amount of water and potassium hydroxide are added, and the mixture is heated and refluxed for 3 hours, and then the solvent is reduced pressure and evaporated to obtain a certain amount. Water is added, and the mixture is extracted three times with ethyl acetate, combined with the extract, washed with saturated brine, dried over anhydrous magnesium sulfate, reduced pressure and evaporated to obtain a pale yellow oil.
(3) The pale yellow oil obtained from step 2 was dried under a vacuum of 45 ° C., dissolved in anhydrous dichloromethane after 5 hours, and a certain amount of NH ₂ OTHP, TBTU and DIPEA was added.
After reacting at room temperature for 6 hours, it was washed twice with a 10% aqueous citric acid solution, further washed twice with a saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, and the solvent was depressurized and evaporated.
Silica gale column chromatography is performed, the eluate is eluted with PE-EA, and the eluate is reduced in pressure and evaporated to obtain a colorless oil.
(4) Dissolve the colorless oil obtained from step 3 in methanol, add a certain amount of 6M hydrochloric acid solution, stir at room temperature for 2 hours, reduce the pressure and evaporate the solvent, and add a certain amount of water. , Extract 3 times with ethyl acetate, combine the extracts, wash with saturated sodium bicarbonate solution, wash with saturated saline, dry with anhydrous sodium sulfate, reduce the solvent and evaporate to pale yellow oil. The product is obtained, which is dissolved in chromatogram methanol, filtered through a micropore filtration membrane, and further purified by HPLC to obtain the target compound 3.
In the HPLC purification method in step (3), the chromatography column is C.
It is an 18-prepared column, the fluid phase is acetonitrile-water, the ratio is 70:30, and the measurement wavelength is 210 nm.
The synthetic route for compound 3 is as follows:

In the present invention, using toluenealkyl urushiol as a starting material, an etherification reaction is carried out with dichloromethane and NaH to produce methylene ether urushiol, and urushiol is oxidized.
The polymerization can be effectively blocked, and the conjugated double bond of the alkyl side chain of urushiol is used to carry out a Diels-Alder reaction with an acrylic acid ester, and further through hydrolysis, cyclohexa-1-enecarboxylic acid. Obtain intermediate compound III with structure, but compound III
Is reacted with silicon group-protected hydroxylamine and HATU to obtain target compound 1 having a xamate group at the tail of the side chain. Target compounds 2 and 3 were synthesized to discuss the effect of introducing functional groups on the benzene ring and fatty chains of urushiol on HDAC inhibitory activity. Using intermediate compound III as a raw material, it is first reacted with mCPBA to oxidize the double bond on the fat chain to ketone and further reduce the aluminum lithium hydroxyl group hydride, but also the carbonyl group at the side chain tail of compound III. Since it is reduced to a hydroxyl group, an oxidation reaction is subsequently carried out with a Jones reagent, and a reduction reaction is carried out with NaBH ₄ to obtain an intermediate compound IV. Compound IV is reacted with NH ₂ OTPH to cause hydroxylation on a fat chain. A target compound 2 having a basic structure is obtained. Using the intermediate compound III as a raw material, the reaction is first carried out with anhydrous copper nitrate, a nitro group is introduced onto the benzene ring, and then the reaction is carried out with NH ₂ OTPH to obtain the target compound 3 having a nitro group structure on the benzene ring. .. The present invention is 1H-NMR and 13C-NMR.
, ESI-MS, IR, and other technical means are used to confirm the structure of the three target compounds.

In the present invention, each of the three compound molecules is HDAC using molecular simulation software.
Molecular docking studies were conducted on 2 and HDAC8 crystals. According to the Glide scoring results, the docking scores of Compounds 1, 2 and 3 and HDAC2 are -7.724 and-, respectively.
It is 7.914 and -7.964, and the corresponding points with HDAC8 are -9.23, respectively.
0, -9.835 and -9.835, and all three compounds are HDAC2 and HDA.
It can bind well to the active pocket of C8, and all of its xamate groups can form a stable chelate with Zn2 + at the bottom of the pocket, and the three compounds are H HDAC2 enzyme H.
Stable hydrogen bond interactions can be formed with residues such as is143, Tyr308, Glu103 and Asp104, and the HDAC8 enzymes Gly140, His143, Ly
Stable hydrogen bond interactions could be formed with residues such as s33, Ala32, His142 and Tyr154. Compounds 2 and 3 among the three compounds showed relatively high Glide scores, which were due to the introduction of an electron-donating group carbonyl group into the alkyl group side chain of ursiol or the substitution of a nitro group on the benzene ring. It will be explained that the introduction of the group significantly improves the binding compatibility between HDAC2 and HDAC8. This is because they form stronger hydrogen bond interactions with the residues.

In the present invention, the inhibitory activity of the three compounds on HDAC2 and HDAC8 was measured using the HDAC test reagent kit. Compounds 1, 2 and 3 both have an excellent inhibitory effect on HDAC2 and HDAC8, and the inhibition rate tends to increase as the concentration increases, and when the concentration is 20 ug / mL, compound 1, The inhibition rates of 2 and 3 for HDAC2 were 95.6%, 96.5% and 97.9%, respectively, and the inhibition rates for HDAC8 were 94.
It was 8%, 95.4% and 96.9%, and the half-suppressed concentrations (I50) of Compounds 1, 2 and 3 with respect to HDAC2 were 0.27, 0.25 and 0.22 ug / mL and HDAC8, respectively.
Half-suppressing concentration (I50) was 0.29, 0.26 and 0.24 ug / mL, respectively.
The IC50 values of the three compounds corresponded to the IC50 values of the positive drug SAHA.

(1) The present invention synthesizes an HDAC inhibitor using natural urushiol as a raw material. Urushiol is a secretion of urushiol, and it is easy to obtain because of its many sources. It is environmentally friendly, safe and can be recycled. Not only is urushiol has excellent antitumor activity, and its structure is similar to that of the FDA-approved HDAC inhibitor SAHA, so it is a natural target antitumor developed from urushiol. The drug has an excellent application foreground.
(2) The present invention uses urushiol as a raw material, blocks the oxidation and polymerization of urushiol through an etherification reaction, stabilizes the chemical structure of urushiol, and through reactions such as Diels-Alder, hydrolysis and hydroxylamine formation. Introducing xamic acid, which is a key medicinal group for suppressing HDAC, into the tail of the urushiol side chain, and acting its target on the pocket site of HDAC enzyme to effectively chelate with zinc ions in HDAC enzyme. , The effect of selectively suppressing HDAC enzymes can be achieved, and the biocompatibility of unsaturated urushiol can be improved, and medicinal groups such as nitro group and hydroxyl group are introduced into the benzene ring or alkyl chain of urushiol. By doing so, the HDAC inhibitory activity is enhanced and HDAC
It can improve the recognition and binding force to enzymes.
(3) The three types of methylene ether, ursiol, and xamic acid derivatives synthesized by the present invention can bind well to the active pocket of HDAC and form a stable hydrogen bond interaction with the residue. At the same time, it can form a stable chelate with Zn2 + at the bottom of the active pocket, has excellent HDAC inhibitory activity, and the half inhibitory concentrations (IC50) of the three compounds with respect to HDAC2 are 0.27, 0, and 25, respectively. And 0.22 ug / mL, H
Half-suppressed concentrations (IC50) for DAC8 are 0.29, 0.26 and 0.2, respectively.
Since it was 4 ug / mL and corresponded to the IC50 value of the FDA-approved HDAC inhibitor SAHA, the methylene ether urushiol xamic acid derivative synthesized by the present invention was developed as a new natural HDAC inhibitor. It is very likely that it can be applied in targeted antitumor agents to effectively increase the added value of natural urushiol.

Hereinafter, the present invention will be described in more detail, and the present invention is not limited thereto.
Example 1
Synthesis of Target Compound 1 (1) Dissolve 3 g of Compound I Urushiol in 50 mL of DMF and 50 mL of DCM.
Add 0.8 g of NaH, reflux under protection of argon gas, spend 12 hours, cool to room temperature, slowly add 100 mL of water, extract 3 times with DCM and use each time. Make 60 mL, combine with the extract, wash twice with saturated saline, and use 90 mL each time.
After drying over anhydrous sodium sulfate, the mixture is evaporated under reduced pressure, column chromatography is performed, the eluate is eluted with PE, and the eluate is reduced under reduced pressure and evaporated to obtain Compound II.
(2) 1.2 g of Compound II is dissolved in 2 mL of toluene, 2 mL of acrylic acid ester is added, and the mixture is refluxed for 36 hours under the protection of argon gas to reduce the pressure and evaporate the solvent, and 10
Add mL of ethanol and 1.2 g of KOH, heat and reflux for 10 minutes, and reduce the solvent.
Evaporate, add 10 mL of water, extract 3 times with EA to make the amount used each time 15 mL, combine the extracts, dry with anhydrous sodium sulfate, reduce the pressure and elute, and perform column chromatography. Then, the mixture is eluted with PE-EA, PE: EA = 100: 1-3: 1, and the eluate is reduced in pressure and evaporated to obtain Compound III.
(3) 30 mg of Compound III was dissolved in 1 mL of anhydrous DMF, and 30 uL of DIPEA, 110 mg.
Add 57 mg of HATU with silicon group-protected hydroxylamine, react and spend 12 hours with stirring, reduce pressure and evaporate, perform direct chromatography, elute with dichloromethane-methanol, and set dichloromethane: methanol = 200: 1. After vacuuming and evaporating the eluate, the product is dissolved in 2 mL of TBAF-THF solution, stirred at room temperature for 40 minutes, vacuumed and evaporated, and then directly subjected to column chromatography to perform DCM-MeO.
Elute with H, make DCM: MeOH = 50: 1, reduce the pressure and evaporate the eluate, and make the product HPL.
Purified with C, the chromatography column is a C18 preparation column, the flow phase is acetonitrile-water, the ratio is 75:25, the measurement wavelength is 210 nm, the target components are collected, vacuumed and evaporated. Obtain 11.6 mg of target compound 1.

Synthesis of Target Compound 2 (1) Dissolve 415 g of Compound III in 5 mL of dichloromethane, add 250 mg of mCPBA, spend 12 hours with stirring at room temperature, wash 3 times with saturated sodium bicarbonate, and wash.
The amount used each time is 5 mL, dried over anhydrous sodium sulfate, vacuumed and evaporated to obtain a colorless oil. Put all of it in 2 mL of anhydrous tetrahydrofuran and slowly 5 mL.
Drip into a turbid solution containing 500 mg lithium aluminum hydride, then set the temperature to 70
Raise to ° C and react for 2 hours, cool to room temperature, slowly add 0.5 mL of water, add 0.5 mL of 15% sodium hydroxide solution again, then add 1.5 mL of water again. Add, filter after 30 minutes, wash with 5 mL of tetrahydrofuran, reduce the pressure of the filtrate to remove tetrahydrofuran, add 3 mL of water, extract 4 times with ethyl acetate, and use 3 each time.
The mixture was made into mL, the extract was combined, washed with saturated brine, dried over anhydrous magnesium sulfate, depressurized and evaporated, silica gale column chromatography was performed, and the mixture was eluted with PE-EA and P.
E: EA = 3: 1 to 2: 1 and the eluate is vacuumed and evaporated to obtain compound IV.
(2) Dissolve 59 mg of compound IV in 0.5 mL of acetone, lower the temperature in an ice-water bath, add 200 uL of Jones reagent, react at room temperature for 30 minutes, and then add 100 uL of isopropanol. Stir for 20 minutes, add 2 mL of saturated sodium chloride solution, extract 5 times with EA, use 2 mL each time, combine with the extract, dry with anhydrous sodium sulfate, reduce pressure and evaporate, silica. Perform Gale column chromatography and PE-
Elute with EA, set PE: EA = 1: 1, and reduce the pressure and evaporate the eluate to obtain compound V.
(3) Dissolve 55 mg of compound V in 0.5 mL of methanol and dissolve 10 mg of NaBH ₄
, React at room temperature for 30 minutes, reduce pressure and evaporate, dissolve in 3 mL of EA, wash twice with saturated brine, extract twice with EA, and use 3 mL each time. The extracts were combined and dried over anhydrous sodium sulfate, the solvent was vacuumed and evaporated, dried under vacuum for 2 hours, 1 mL DCM, 13 mg NH ₂ OTHP, 15 mg EDC. Cl and 20
Add uL TEA, spend 12 hours while reacting, wash twice with 2 mL of 5% aqueous citric acid solution, extract twice with DCM, use 3 mL each time, and combine with the organic layer. Then, reduce the pressure and dry the solvent, add 0.8 mL of methanol solution, lower the temperature in an ice-water bath, add 1 mL of 10% HCl solution, react for 1 hour, and then reduce and evaporate the solvent. Dissolved in 2, 2 mL of methanol, filtered through a micropore filter membrane, and further purified with HCl, the chromatography column was a C18 preparation column, the fluid phase was acetonitrile-water, and the ratio was 68:32. The measurement wavelength is 210 nm, and the target component is collected to reduce the pressure.
Evaporate to give 30 mg of target compound 2.

Synthesis of Target Compound 3 (1) Put 620 mg of Compound III in 5 mL of freshly evaporated acetic anhydride and set the temperature to 0 ° C.
Add 300 mg of anhydrous copper nitrate while stirring vigorously, add 295 mg of anhydrous copper nitrate after 2 hours, add 15 mL of water after 3 hours, stir for 20 minutes, and then extract twice with ethyl acetate, each time. The amount used is 10 mL, the organic layer is combined, washed twice with saturated sodium bicarbonate solution, the amount used each time is 20 mL, dried with anhydrous magnesium sulfate, reduced pressure and evaporated, and a pale yellow oily substance is used. To get.
(2) The pale yellow oil obtained from step 1 was dissolved in 10 mL of methanol and 0.5 m.
Add L of water and 1.1 g of potassium hydroxide, heat and reflux for 3 hours, then reduce the pressure of the solvent.
Evaporate, add 5 mL of water, extract 3 times with ethyl acetate, use 5 mL each time, combine the extracts, wash with 10 mL of saturated brine, dry with anhydrous magnesium sulfate, and reduce the pressure. -Evaporate to obtain a pale yellow oil.
(3) The pale yellow oil obtained from step 2 is dried under a vacuum of 45 ° C., and after 5 hours, 5 mL
Dissolved in anhydrous dichloromethane, 160 mg NH ₂ OTHP, 400 mg TBTU
And 0.6 mL of DIPEA was added and reacted at room temperature for 6 hours, and then washed twice with a 10% aqueous citric acid solution to make the amount used 5 mL each time, and further washed twice with a saturated sodium bicarbonate solution. , The amount used each time is 10 mL, dried over anhydrous sodium sulfate, the solvent is depressurized and evaporated, silica gale column chromatography is performed, and the mixture is eluted with PE-EA.
EA = 3: 1 to 2: 1 and the eluate is vacuumed and evaporated to obtain a colorless oil.
(4) Dissolve the colorless oil obtained in step 3 in 3 mL of methanol, add 0.5 mL of 6M hydrochloric acid solution, stir at room temperature for 2 hours, reduce the pressure and evaporate the solvent, and add 3 mL of water. Put and extract 3 times with ethyl acetate, each time the amount used is 3 mL, and the extract is mixed and 5 mL
Wash with saturated sodium bicarbonate solution, further wash with 5 mL saturated saline, dry with anhydrous sodium sulfate, and reduce and elute the solvent to give a pale yellow oil, which is in 1 mL chromatogram methanol. The chromatography column is a C18 preparation column, and the flow phase is acetonitrile-, which is dissolved in, filtered through a micropore filtration membrane, and further purified by HPLC.
It is water, the ratio is 70:30, the measurement wavelength is 210 nm, the target components are collected, vacuumed and evaporated to obtain 200 mg of the target compound 3.

Example 2
Synthesis of Target Compound 1 (1) Dissolve 4 g of Compound I Urushiol in 60 mL of DMF and 60 mL of DCM.
Add 0.85 g of NaH, reflux under protection of argon gas, spend 12 hours, cool to room temperature, slowly add 100 mL of water, extract 3 times with DCM and use each time. Make 60 mL, combine with the extract, wash twice with saturated saline, and use 100 each time.
The mixture is made into mL, dried over anhydrous sodium sulfate, evaporated under reduced pressure, column chromatographically performed, eluted with PE, and the eluate is reduced under pressure and evaporated to obtain Compound II.
(2) 1.5 g of Compound II was dissolved in 3 mL of toluene, 3 mL of acrylic ester was added, and the mixture was refluxed for 36 hours under the protection of argon gas to reduce the pressure and evaporate the solvent.
Add mL of ethanol and 1.5 g of KOH, heat and reflux for 10 minutes, and reduce the solvent.
Evaporate, add 15 mL of water, extract 3 times with EA to make the amount used each time 15 mL, combine the extracts, dry with anhydrous sodium sulfate, reduce the pressure and elute, and perform column chromatography. Then, the mixture is eluted with PE-EA, PE: EA = 100: 1-3: 1, and the eluate is reduced in pressure and evaporated to obtain Compound III.
(3) 40 mg of Compound III was dissolved in 2 mL of anhydrous DMF, and 40 uL of DIPEA, 120 mg.
Add silicon group-protected hydroxylamine and 60 mg of HATU, react and spend 12 hours with stirring, reduce pressure and evaporate, perform direct chromatography, elute with dichloromethane-methanol, and set dichloromethane: methanol = 200: 1. After vacuuming and evaporating the eluate, the product is dissolved in 3 mL of TBAF-THF solution, stirred at room temperature for 40 minutes, vacuumed and evaporated, and then directly subjected to column chromatography to perform DCM-MeO.
Elute with H, make DCM: MeOH = 50: 1, reduce the pressure and evaporate the eluate, and make the product HPL.
Purified with C, the chromatography column is a C18 preparation column, the flow phase is acetonitrile-water, the ratio is 75:25, the measurement wavelength is 210 nm, the target components are collected, vacuumed and evaporated. Obtain 12.5 mg of target compound 1.

Synthesis of Target Compound 2 (1) Dissolve 450 g of Compound III in 6 mL of dichloromethane, add 300 mg of mCPBA, spend 12 hours with stirring at room temperature, wash 3 times with saturated sodium bicarbonate, and wash.
The amount used each time is 6 mL, dried over anhydrous sodium sulfate, vacuumed and evaporated to obtain a colorless oil. Put all of it in 3 mL of anhydrous tetrahydrofuran and slowly 6 mL.
Drip into a turbid solution containing 600 mg lithium aluminum hydride, then set the temperature to 70
Raise to ° C for 2 hours, allow to cool to room temperature, slowly add 0.8 mL of water, add 0.8 mL of 15% sodium hydroxide solution again, then add 2.0 mL of water again. Add, filter after 30 minutes, wash with 5 mL of tetrahydrofuran, reduce the pressure of the filtrate to remove tetrahydrofuran, add 4 mL of water, extract 4 times with ethyl acetate, and use 4 times each time.
The mixture was made into mL, the extract was combined, washed with saturated brine, dried over anhydrous magnesium sulfate, depressurized and evaporated, silica gale column chromatography was performed, and the mixture was eluted with PE-EA and P.
E: EA = 3: 1 to 2: 1 and the eluate is vacuumed and evaporated to obtain compound IV.
(2) Dissolve 60 mg of compound IV in 0.5 mL of acetone, lower the temperature in an ice-water bath, add 200 uL of Jones reagent, react at room temperature for 30 minutes, and then add 100 uL of isopropanol. Stir for 20 minutes, add 3 mL of saturated sodium chloride solution, extract 5 times with EA, use 3 mL each time, combine with the extract, dry with anhydrous sodium sulfate, reduce pressure and evaporate, silica. Perform Gale column chromatography and PE-
Elute with EA, set PE: EA = 1: 1, and reduce the pressure and evaporate the eluate to obtain compound V.
(3) Dissolve 60 mg of compound V in 0.6 mL of methanol and dissolve 10 mg of NaBH ₄
, React at room temperature for 30 minutes, reduce pressure and evaporate, dissolve in 4 mL of EA, wash twice with saturated brine, extract twice with EA, and use 4 mL each time. The extracts were combined and dried over anhydrous sodium sulfate, the solvent was vacuumed and evaporated, dried under vacuum for 2 hours, 1 mL DCM, 15 mg NH ₂ OTHP, 15 mg EDC. Cl and 20
Add uL TEA, spend 12 hours while reacting, wash twice with 3 mL of 5% aqueous citric acid solution, extract twice with DCM, use 4 mL each time, and combine with the organic layer. Then, reduce the pressure and dry the solvent, add 0.8 mL of methanol solution, lower the temperature in an ice-water bath, add 1 mL of 10% HCl solution, react for 1 hour, and then reduce and evaporate the solvent. Dissolved in 3, mL of methanol, filtered through a micropore filter membrane, and further purified with HCl, the chromatography column was a C18 preparation column, the fluid phase was acetonitrile-water, and the ratio was 68:32. The measurement wavelength is 210 nm, and the target component is collected to reduce the pressure.
Evaporate to give 32 mg of target compound 2.

Synthesis of Target Compound 3 (1) Put 700 mg of Compound III in 6 mL of freshly evaporated acetic anhydride and set the temperature to 0 ° C.
Add 350 mg of anhydrous copper nitrate while stirring vigorously, add 300 mg of anhydrous copper nitrate after 2 hours, add 20 mL of water after 3 hours, stir for 20 minutes, and then add 2 with ethyl acetate.
Extract once, use 15 mL each time, combine organic layers, wash twice with saturated sodium bicarbonate solution, use 20 mL each time, dry with anhydrous magnesium sulfate, reduce pressure.
Evaporate to give a pale yellow oil.
(2) The pale yellow oil obtained from step 1 was dissolved in 10 mL of methanol and 0.6 m.
Add L of water and 1.2 g of potassium hydroxide, heat and reflux for 3 hours, then reduce the pressure of the solvent.
Evaporate, add 6 mL of water, extract 3 times with ethyl acetate, use 6 mL each time, combine with the extract, wash with 15 mL saturated brine, dry with anhydrous magnesium sulfate, and reduce pressure. -Evaporate to obtain a pale yellow oil.
(3) The pale yellow oil obtained from step 2 is dried under a vacuum of 45 ° C., and after 5 hours, 6 mL
Dissolved in anhydrous dichloromethane, 160 mg NH ₂ OTHP, 400 mg TBTU
And 0.6 mL of DIPEA was added and reacted at room temperature for 6 hours, and then washed twice with a 10% aqueous citric acid solution to make the amount used each time 6 mL, and further washed twice with a saturated sodium bicarbonate solution. , The amount used each time is 10 mL, dried over anhydrous sodium sulfate, the solvent is depressurized and evaporated, silica gale column chromatography is performed, and the mixture is eluted with PE-EA.
EA = 3: 1 to 2: 1 and the eluate is vacuumed and evaporated to obtain a colorless oil.
(4) Dissolve the colorless oil obtained from step 3 in 4 mL of methanol, add 0.6 mL of 6M hydrochloric acid solution, stir at room temperature for 2 hours, reduce the pressure and evaporate the solvent, and add 5 mL of water. Put in and extract 3 times with ethyl acetate, each time the amount used is 5 mL, and the extract is combined to make 5 mL.
Wash with saturated sodium bicarbonate solution, further wash with 5 mL saturated saline, dry with anhydrous sodium sulfate, and reduce and elute the solvent to give a pale yellow oil, which is in 1 mL chromatogram methanol. The chromatography column is a C18 preparation column, and the flow phase is acetonitrile-, which is dissolved in, filtered through a micropore filtration membrane, and further purified by HPLC.
It is water, the ratio is 70:30, the measurement wavelength is 210 nm, the target components are collected, vacuumed and evaporated to obtain 210 mg of the target compound 3.

Example 3
Synthesis of Target Compound 1 (1) Dissolve 5 g of Compound I Urushiol in 90 mL of DMF and 90 mL of DCM.
Add 1.2 g of NaH, reflux under protection of argon gas, spend 12 hours, cool to room temperature, slowly add 150 mL of water, extract 3 times with DCM and use each time. Make 100 mL, combine with the extract, wash twice with saturated brine, and use 100 each time.
The mixture is made into mL, dried over anhydrous sodium sulfate, evaporated under reduced pressure, column chromatographically performed, eluted with PE, and the eluate is reduced under pressure and evaporated to obtain Compound II.
(2) 2.0 g of Compound II was dissolved in 4 mL of toluene, 4 mL of acrylic acid ester was added, and the mixture was refluxed for 36 hours under the protection of argon gas to reduce the pressure and evaporate the solvent.
Add mL of ethanol and 1.9 g of KOH, heat and reflux for 10 minutes, and reduce the solvent.
Evaporate, add 15 mL of water, extract 3 times with EA to make the amount used each time 15 mL, combine the extracts, dry with anhydrous sodium sulfate, reduce the pressure and elute, and perform column chromatography. Then, the mixture is eluted with PE-EA, PE: EA = 100: 1-3: 1, and the eluate is reduced in pressure and evaporated to obtain Compound III.
(3) 50 mg of Compound III was dissolved in 2 mL of anhydrous DMF and 48 uL of DIPEA, 170 mg.
Add silicon group-protected hydroxylamine and 90 mg of HATU, react and spend 12 hours with stirring, reduce pressure and evaporate, perform direct chromatography, elute with dichloromethane-methanol, and set dichloromethane: methanol = 200: 1. After depressurizing and evaporating the eluate, the product was dissolved in 3.5 mL of TBAF-THF solution and 40 at room temperature.
Stir for minutes, reduce pressure and evaporate, then perform direct column chromatography, DCM-M
Elute with eOH, make DCM: MeOH = 50: 1, reduce the pressure and evaporate the eluate, and make the product H.
Purified with PLC, the chromatography column is a C18 preparation column, the flow phase is acetonitrile-water, the ratio is 75:25, the measurement wavelength is 210 nm, the target components are collected, depressurized and evaporated. Obtain 19 mg of target compound 1.

Synthesis of Target Compound 2 (1) Dissolve 650 g of Compound III in 9 mL of dichloromethane, add 400 mg of mCPBA, spend 12 hours with stirring at room temperature, wash 3 times with saturated sodium bicarbonate, and wash.
The amount used each time is 8 mL, dried over anhydrous sodium sulfate, vacuumed and evaporated to obtain a colorless oil. Put all of it in 3 mL anhydrous tetrahydrofuran and slowly 8 mL.
Drip into a turbid solution containing 800 mg lithium aluminum hydride, then temperature 70
Raise to ° C for 2 hours, allow to cool to room temperature, slowly add 0.8 mL of water, add 0.8 mL of 15% sodium hydroxide solution again, then add 2.5 mL of water again. Add, filter after 30 minutes, wash with 8 mL of tetrahydrofuran, reduce the pressure of the filtrate to remove tetrahydrofuran, add 5 mL of water, extract 4 times with ethyl acetate, and use 5 each time.
The mixture was made into mL, the extract was combined, washed with saturated brine, dried over anhydrous magnesium sulfate, depressurized and evaporated, silica gale column chromatography was performed, and the mixture was eluted with PE-EA and P.
E: EA = 3: 1 to 2: 1 and the eluate is vacuumed and evaporated to obtain compound IV.
(2) Dissolve 95 mg of compound IV in 0.8 mL of acetone, lower the temperature in an ice-water bath, add 300 uL of Jones reagent, react at room temperature for 30 minutes, and then add 160 uL of isopropanol. Stir for 20 minutes, add 3 mL of saturated sodium chloride solution, extract 5 times with EA, use 3 mL each time, combine with the extract, dry with anhydrous sodium sulfate, reduce pressure and evaporate, silica. Perform Gale column chromatography and PE-
Elute with EA, set PE: EA = 1: 1, and reduce the pressure and evaporate the eluate to obtain compound V.
(3) Dissolve 85 mg of compound V in 0.8 mL of methanol and 16 mg of NaBH ₄
, React at room temperature for 30 minutes, reduce pressure and evaporate, dissolve in 5 mL of EA, wash twice with saturated brine, extract twice with EA, and use 5 mL each time. The extracts were combined and dried over anhydrous sodium sulfate, the solvent was vacuumed and evaporated, dried under vacuum for 2 hours, 1.5 mL DCM, 20 mg NH ₂ OTHP, 20 mg EDC. Add Cl and 30 uL of TEA, spend 12 hours while reacting, wash twice with 3 mL of 5% aqueous citric acid solution, extract twice with DCM, and use 5 mL each time to prepare the organic layer. After merging, reduce the pressure and dry the solvent, add 1.2 mL of methanol solution, lower the temperature in an ice-water bath, add 1.5 mL of 10% HCl solution, react for 1 hour, and then add the solvent. Decompressed and evaporated, dissolved in 3 mL of methanol, filtered through a microporous filtration membrane and further purified with HCl, the chromatography column was a C18 preparation column, the fluid phase was acetonitrile-water, the ratio was 68: It is 32, the measurement wavelength is 210 nm, and the target component is collected and reduced in pressure and evaporated to obtain 47 mg of the target compound 2.

Synthesis of Target Compound 3 (1) Put 950 mg of Compound III in 8 mL of freshly evaporated acetic anhydride and set the temperature to 0 ° C.
Add 500 mg of anhydrous copper nitrate while stirring vigorously, add 450 mg of anhydrous copper nitrate after 2 hours, add 25 mL of water after 3 hours, stir for 20 minutes, and then extract twice with ethyl acetate, each time. The amount used was 15 mL, the organic layer was combined, washed twice with saturated sodium bicarbonate solution, the amount used each time was 30 mL, dried with anhydrous magnesium sulfate, reduced pressure and evaporated, and a pale yellow oily substance was used. To get.
(2) The pale yellow oil obtained from step 1 was dissolved in 16 mL of methanol and 0.8 m.
Add L of water and 1.6 g of potassium hydroxide, heat and reflux for 3 hours, then reduce the pressure of the solvent.
Evaporate, add 8 mL of water, extract 3 times with ethyl acetate, use 8 mL each time, combine with the extract, wash with 15 mL of saturated brine, dry with anhydrous magnesium sulfate, and reduce the pressure. -Evaporate to obtain a pale yellow oil.
(3) The pale yellow oil obtained from step 2 is dried under a vacuum of 45 ° C., and after 5 hours, 8 mL
Dissolved in anhydrous dichloromethane, 250 mg NH ₂ OTHP, 650 mg TBTU
And 0.9 mL of DIPEA was added and reacted at room temperature for 6 hours, and then washed twice with a 10% aqueous citric acid solution to make the amount used each time 8 mL, and further washed twice with a saturated sodium bicarbonate solution. , The amount used each time is 15 mL, dried over anhydrous sodium sulfate, the solvent is reduced under reduced pressure and evaporated, silica gale column chromatography is performed, and the mixture is eluted with PE-EA.
EA = 3: 1 to 2: 1 and the eluate is vacuumed and evaporated to obtain a colorless oil.
(4) Dissolve the colorless oil obtained in step 3 in 5 mL of methanol, add 0.8 mL of 6M hydrochloric acid solution, stir at room temperature for 2 hours, reduce the pressure and evaporate the solvent, and add 5 mL of water. Add and extract 3 times with ethyl acetate, each time the amount used is 5 mL, and the extract is combined to make 8 mL.
Wash with saturated sodium bicarbonate solution, further wash with 8 mL saturated saline, dry with anhydrous sodium sulfate, reduce the solvent and elute to give a pale yellow oil, which is in 2 mL chromatogram methanol. The chromatography column is a C18 preparation column, and the flow phase is acetonitrile-, which is dissolved in, filtered through a micropore filtration membrane, and further purified by HPLC.
It is water, the ratio is 70:30, the measurement wavelength is 210 nm, and the target components are collected and vacuumed and evaporated to obtain 315 mg of the target compound 3.

Example 4
Structural assessment of the three compounds 1 Using means such as 1 H-NMR, 13C-NMR, ESI-MS, and IR, confirm the assessment of the chemical structures of the synthesized compounds 1, 2 and 3, and the three compounds See Table 1 for physicochemical properties and spectrum data.
Table 1 Physicochemical constants and spectrum data of compounds 1, 2 and 3

Example 5
Molecular docking of three compounds with HDAC2 and HDAC8 The present invention uses the GLIDE program to attach the molecules of compounds 1, 2 and 3 to HDAC, respectively.
Crystal structure of 2 (PDB ID: 4LXZ) and crystal structure of HDAC8 (PDB ID)
: 3SFF) and using the Grid scoring function, compound and HDA
The docking effect with C2 and HDAC8 was evaluated. As a result, compounds 1, 2 and 3 and H
The docking points with DAC2 are -7.724, -7.914 and -7.946, respectively, and the docking points with HDAC8 are -9.230, -9.583 and -9, respectively.
835, indicating that all three compounds can bind well to the active pockets of HDAC2 and HDAC8, the fat chain portion of the three compounds occupies an elongated tube site in the pocket, and all of the xynamate groups are present. Located at the bottom of the tube, any xosate group can form a stable chelate with Zn2 + at the bottom of the pocket, and the hydroxyl and carbonyl in the xosate group are stable hydrogen bonds with the His145 and Try308 residues of the HDAC2 enzyme. An interaction can be formed, a stable hydrogen bond interaction can be formed with Gly140 and His143 residues of the HDAC8 enzyme, and a carbonyl group on the fatty chain of compound 2 and a benzene ring of compound 3 can be formed. The nitro group can form a stable hydrogen bond interaction with residues such as Glu103 and Asp104 of the HDAC2 enzyme, and HDAC8
Stable hydrogen bond interactions can be formed with residues such as the enzymes Lys33, Ala32, His142 and Tyr154. Compounds 2 and 3 among the three compounds are relatively high Gli
The d-point was shown, which was achieved by introducing an electron-donating group carbonyl group into the alkyl group side chain of ursiol, or by introducing a substituent such as a nitro group on the benzene ring.
It is expressed that the binding compatibility between DAC2 and HDAC8 is significantly improved. This is because they form stronger hydrogen bond interactions with the residues.

Example 6
Evaluation of inhibitory activity of the three compounds against HDAC2 and HDAC8 In the present invention, the inhibitory activity of the three compounds against HDAC2 and HDAC8 was measured using the HDAC test reagent kit. Compounds 1, 2 and 3 all had an excellent inhibitory effect on HDAC2 and HDAC8, and the inhibition rate tended to increase as the concentration increased, showing a clear concentration requestability. HD of compounds 1, 2 and 3 when the concentration is 20 ug / mL
The inhibition rates for AC2 were 95.6%, 96.5% and 97.9%, respectively, and the inhibition rates for HDAC8 were 94.8%, 95.4% and 96.9%, respectively, and compounds 1, 2 and 3 The half-suppressed concentrations (I50) of HDAC2 were 0.27, 0.25 and 0, respectively.
.. Half-suppressed concentration (I50) for 22 ug / mL and HDAC8 is 0.29 and 0, respectively.
.. It is 26 and 0.24 ug / mL, and the IC50 value of the three compounds is the IC of the positive drug SAHA.
It was equivalent to 50 values. In addition, the inhibitory effect of Compound 2 and Compound 3 on HDAC2 and HDAC8 is superior to that of Compound 1, and the inhibitory results of the three compounds on HDAC2 and HDAC8 are in agreement with the molecular docking results, which is the alkyl of urciol. It is expressed that the inhibitory activity against HDAC2 and HDAC8 can be remarkably improved by introducing an electron donating group carbonyl group into the group side chain or introducing a substituent such as a nitro group on the benzene ring.

Table 2 Effects of different concentration compounds on HDAC2 and HDAC8 inhibition rates

Claims (8)

Contains compounds 1, 2 and 3
The chemical names of the compounds 1, 2 and 3 are 5-(10-(benzo [d] [1,3]], respectively.
Dioxolane -4-yl) Decane -2-en -1-yl) -N-Hydroxy-2-methylcyclohexane -3-ene -1 -formamide 5-(10-(benzo [d] [1,3]
] Dioxolane -4-yl) -3-hydroxyl-decane) -N-hydroxyl-2 -methylcyclohexane -3-ene -1-formamide and 5-(10-(7-nitrobenzo)
[d] [1,3] Dioxolane -4-yl) Decane -2-ene -1-yl) -N-Hydroxy-2-methylcyclohexane -3-ene -1 -formamide,
The chemical formulas of the compounds 1, 2 and 3 are, respectively.
Compound 1
Compound 2
Compound 3
A methylene ether urushiol hydroxamic acid inducer characterized by having HDAC inhibitory activity. The method for synthesizing the compound 1 is



(1) Using compound I urushiol as a raw material, take a certain amount of urushiol and take DMF and DC.
Dissolve in M, add a certain amount of NaH, reflux under protection of argon gas for 12 hours, cool to room temperature, slowly add a certain amount of water, and extract 3 times with DCM. The organic layer is combined, washed with saturated brine, dried over anhydrous sodium sulfate, and then decompressed and evaporated.
A step of performing column chromatography, eluting with PE, and decompressing and evaporating the eluate to obtain Compound II.
(2) Take a certain amount of Compound II, dissolve it in toluene, add a certain amount of acrylic acid ester, heat and reflux for 36 hours under the protection of argon gas, and reduce and evaporate the solvent.
Add a certain amount of ethanol and KOH, heat and reflux for 10 minutes, reduce the pressure and elute the solvent, add a certain amount of water, extract 3 times with EA, combine the extract and dry with anhydrous sodium sulfate. After that, depressurize and dry, perform column chromatography, elute with PE-EA, and depressurize and evaporate the eluate to obtain compound III.
(3) Take a certain amount of Compound III, dissolve it in anhydrous DMF, add a certain amount of DIPEA, silicon-protected hydroxylamine and HATU, spend 12 hours while reacting and stirring, and reduce the pressure and elute the solvent. Perform column chromatography directly, elute with dichloromethane-methanol, reduce the pressure and elute the eluate, then put the product in a certain amount of TBAF-THF solution, stir at room temperature for 40 minutes, reduce the pressure and evaporate. , Direct column chromatography, elution with DCM-MeOH, decompression and evaporation of the eluate, and further purification of the product by HPLC to obtain the target compound 1.
The method for synthesizing a methylene ether urushiol hydroxamic acid derivative having an HDAC inhibitory activity according to claim 1, wherein the method comprises. The method for synthesizing the compound 2 is

(1) Using compound III as a raw material, a certain amount of compound III was dissolved in dichloromethane, a certain amount of mCPBA was added, the mixture was spent for 12 hours with stirring at room temperature, and washed with saturated sodium bicarbonate three times. Dry with anhydrous sodium sulfate, reduce pressure and evaporate to obtain a colorless oil.
Put all of them in anhydrous tetrahydrofuran, slowly drip a certain amount of 100 mg / mL lithium aluminum hydride into the turbid solution, then raise the temperature to 70 ° C. for 2 hours reaction, cool to room temperature, and then slowly. Add a certain amount of water, add a certain amount of 15% sodium hydroxide solution again, then add a certain amount of water again, filter after 30 minutes, wash with tetrahydrofuran, and reduce the pressure of the filtrate. Remove tetrahydrofuran, add a certain amount of water,
Extract 4 times with ethyl acetate, combine the extracts, wash with saturated brine, dry with anhydrous magnesium sulfate, reduce pressure and evaporate, perform silica gale column chromatography, and perform PE-E.
The step of eluting with A and decompressing and evaporating the eluate to obtain compound IV,
(2) Dissolve Compound IV in a certain amount of acetone, lower the temperature in an ice-water bath, add a certain amount of Jones reagent, react at room temperature for 30 minutes, and then add a certain amount of isopropanol. Stir for 20 minutes, add a certain amount of saturated sodium chloride solution, extract 5 times with EA,
The steps of combining the extracts, drying over anhydrous sodium sulfate, depressurizing and evaporating, performing silica gale column chromatography, eluting with PE-EA, depressurizing and evaporating the eluate to obtain compound V, and
(3) Dissolve compound V in a certain amount of methanol, add a certain amount of NaBH ₄ , react at room temperature for 30 minutes, reduce the pressure and evaporate, then dissolve in a certain amount of EA, saturated saline solution. Wash twice with EA, extract twice with EA, combine with the extract, dry with anhydrous sodium sulfate, vacuum and evaporate the solvent, dry for 2 hours under vacuum conditions, and a certain amount of DCM, NH ₂ OTHP,
EDC. Add Cl and TEA, spend 12 hours while reacting, wash twice with 5% aqueous citric acid solution, extract twice with DCM, combine with organic layer, reduce the pressure and dry the solvent, and methanol. Add the solution, lower the temperature in an ice-water bath, add a certain amount of 10% HCl solution, react for 1 hour, then reduce the pressure and evaporate the solvent, dissolve in methanol, and filter with a micropore filtration membrane. , Further purification with HCl to obtain target compound 2 and
The method for synthesizing a methylene ether urushiol hydroxamic acid derivative having an HDAC inhibitory activity according to claim 1, wherein the method comprises. The method for synthesizing the compound 3 is
(1) Using compound III as a raw material, a certain amount of compound III is put into newly evaporated acetic anhydride, the temperature is lowered to 0 ° C., a certain amount of anhydrous copper nitrate is added with vigorous stirring, and a certain amount is added after 2 hours. Add anhydrous copper nitrate, add a certain amount of water after 3 hours, stir for 20 minutes, and then add 2 with ethyl acetate.
The step of extracting twice, merging the organic layer, washing twice with saturated sodium bicarbonate solution, drying with anhydrous magnesium sulfate, depressurizing and evaporating to obtain a pale yellow oily substance,
(2) The pale yellow oil obtained from step 1 is dissolved in methanol, a certain amount of water and potassium hydroxide are added, and the mixture is heated and refluxed for 3 hours, and then the solvent is reduced pressure and evaporated to obtain a certain amount. Add water, extract 3 times with ethyl acetate, combine the extracts, wash with saturated brine, dry with anhydrous magnesium sulfate, reduce pressure and evaporate to obtain a pale yellow oil.
(3) The pale yellow oil obtained from step 2 was dried under a vacuum of 45 ° C., dissolved in anhydrous dichloromethane after 5 hours, and a certain amount of NH ₂ OTHP, TBTU and DIPEA was added.
After reacting at room temperature for 6 hours, it was washed twice with a 10% aqueous citric acid solution, further washed twice with a saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, and the solvent was depressurized and evaporated.
A step of performing silica gale column chromatography, eluting with PE-EA, and decompressing and evaporating the eluate to obtain a colorless oil.
(4) Dissolve the colorless oil obtained in step 3 in methanol, add a certain amount of 6M hydrochloric acid solution, stir at room temperature for 2 hours, reduce the pressure and evaporate the solvent, and add a certain amount of water. , Extracted 3 times with ethyl acetate, combined with the extract, washed with saturated sodium bicarbonate solution, further washed with saturated saline, dried with anhydrous sodium sulfate, and the solvent was depressurized and evaporated to give a pale yellow oil. A step of obtaining a target compound 3 by dissolving it in chromatogram methanol, filtering it with a micropore filtration membrane, and further purifying it by HPLC.
The method for synthesizing a methylene ether urushiol hydroxamic acid derivative having an HDAC inhibitory activity according to claim 1, wherein the method comprises. The second aspect of claim 2 is that the urushiol in step (1) is derived from lacquer and is obtained by extracting the lacquer with methanol as a raw material, and the purity of the urushiol is 90% or more. A method for synthesizing a methylene ether / urushiol / hydroxamic acid derivative, which comprises the above-mentioned HDAC inhibitory activity. In the HPLC purification method in step (3), the chromatography column is a C18 preparation column, the fluid phase is acetonitrile-water, the ratio is 75:25, and the measurement wavelength is 2.
The method for synthesizing a methylene ether urushiol hydroxamic acid derivative, which has the HDAC inhibitory activity according to claim 2, which is 10 nm. In the HPLC purification method in step (3), the chromatography column is a C18 preparation column, the fluid phase is acetonitrile-water, the ratio is 68:32, and the measurement wavelength is 2.
The method for synthesizing a methylene ether urushiol hydroxamic acid derivative having an HDAC inhibitory activity according to claim 3, wherein the diameter is 10 nm. The HPLC purification method in step (3) is characterized in that the chromatography column is a C18 preparation column, the fluid phase is acetonitrile-water, the ratio is 70:30, and the measurement wavelength is 210 nm. The method for synthesizing a methylene ether urushiol hydroxamic acid derivative having an HDAC inhibitory activity according to claim 4.