CN110713453A - Preparation method and application of compounds BIA-01 and BIA-02 with CDC25B inhibitory activity - Google Patents
Preparation method and application of compounds BIA-01 and BIA-02 with CDC25B inhibitory activity Download PDFInfo
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- CN110713453A CN110713453A CN201910934106.1A CN201910934106A CN110713453A CN 110713453 A CN110713453 A CN 110713453A CN 201910934106 A CN201910934106 A CN 201910934106A CN 110713453 A CN110713453 A CN 110713453A
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- 150000001875 compounds Chemical class 0.000 title claims abstract description 36
- 230000002401 inhibitory effect Effects 0.000 title claims abstract description 26
- 238000002360 preparation method Methods 0.000 title claims abstract description 10
- 101000624631 Homo sapiens M-phase inducer phosphatase 2 Proteins 0.000 title claims abstract 10
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- 230000000694 effects Effects 0.000 claims abstract description 14
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- LSBDFXRDZJMBSC-UHFFFAOYSA-N 2-phenylacetamide Chemical class NC(=O)CC1=CC=CC=C1 LSBDFXRDZJMBSC-UHFFFAOYSA-N 0.000 claims abstract description 8
- 239000000203 mixture Substances 0.000 claims abstract description 7
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- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 35
- 238000010438 heat treatment Methods 0.000 claims description 35
- 230000008569 process Effects 0.000 claims description 33
- 239000012043 crude product Substances 0.000 claims description 28
- 238000001816 cooling Methods 0.000 claims description 27
- NLKNQRATVPKPDG-UHFFFAOYSA-M potassium iodide Chemical compound [K+].[I-] NLKNQRATVPKPDG-UHFFFAOYSA-M 0.000 claims description 27
- HVPQMLZLINVIHW-UHFFFAOYSA-N 6-bromo-1h-indole-2,3-dione Chemical compound BrC1=CC=C2C(=O)C(=O)NC2=C1 HVPQMLZLINVIHW-UHFFFAOYSA-N 0.000 claims description 25
- WTDHULULXKLSOZ-UHFFFAOYSA-N Hydroxylamine hydrochloride Chemical compound Cl.ON WTDHULULXKLSOZ-UHFFFAOYSA-N 0.000 claims description 23
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- QAOWNCQODCNURD-UHFFFAOYSA-N sulfuric acid Substances OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 19
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- 239000012065 filter cake Substances 0.000 claims description 18
- ORVVNXAFZBVFMP-BJMVGYQFSA-N (2e)-n-(3-bromophenyl)-2-hydroxyiminoacetamide Chemical compound O\N=C\C(=O)NC1=CC=CC(Br)=C1 ORVVNXAFZBVFMP-BJMVGYQFSA-N 0.000 claims description 17
- 239000012295 chemical reaction liquid Substances 0.000 claims description 17
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- 239000005457 ice water Substances 0.000 claims description 16
- DHYHYLGCQVVLOQ-UHFFFAOYSA-N 3-bromoaniline Chemical compound NC1=CC=CC(Br)=C1 DHYHYLGCQVVLOQ-UHFFFAOYSA-N 0.000 claims description 14
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 claims description 13
- 235000011114 ammonium hydroxide Nutrition 0.000 claims description 13
- 150000001448 anilines Chemical class 0.000 claims description 13
- QNVKMXGRFVLMBM-UHFFFAOYSA-N n-chloro-n-phenylacetamide Chemical class CC(=O)N(Cl)C1=CC=CC=C1 QNVKMXGRFVLMBM-UHFFFAOYSA-N 0.000 claims description 13
- PAYRUJLWNCNPSJ-UHFFFAOYSA-N N-phenyl amine Natural products NC1=CC=CC=C1 PAYRUJLWNCNPSJ-UHFFFAOYSA-N 0.000 claims description 12
- 238000004090 dissolution Methods 0.000 claims description 12
- 239000007832 Na2SO4 Substances 0.000 claims description 11
- 229910052938 sodium sulfate Inorganic materials 0.000 claims description 11
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- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical class [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 claims description 10
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- 238000005303 weighing Methods 0.000 claims description 10
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- 239000002244 precipitate Substances 0.000 claims description 7
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Substances [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 claims description 6
- 229910000027 potassium carbonate Inorganic materials 0.000 claims description 4
- 238000004519 manufacturing process Methods 0.000 claims description 3
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- 229920006395 saturated elastomer Polymers 0.000 claims description 2
- 239000007858 starting material Substances 0.000 claims description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Natural products N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 claims 2
- 230000015572 biosynthetic process Effects 0.000 abstract description 8
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- 125000001424 substituent group Chemical group 0.000 abstract description 4
- 238000002626 targeted therapy Methods 0.000 abstract description 2
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical class CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 19
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 15
- 239000013078 crystal Substances 0.000 description 10
- -1 diketone indoline derivative Chemical class 0.000 description 8
- 238000012544 monitoring process Methods 0.000 description 8
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
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- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 2
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- KDXNYSZNOWTPLE-UHFFFAOYSA-N 3'-hydroxy-6'-methoxyspiro[2-benzofuran-3,9'-xanthene]-1-one Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(OC)=CC=C21 KDXNYSZNOWTPLE-UHFFFAOYSA-N 0.000 description 2
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- 102000004160 Phosphoric Monoester Hydrolases Human genes 0.000 description 2
- 108090000608 Phosphoric Monoester Hydrolases Proteins 0.000 description 2
- 102000004022 Protein-Tyrosine Kinases Human genes 0.000 description 2
- 108090000412 Protein-Tyrosine Kinases Proteins 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 125000003368 amide group Chemical group 0.000 description 2
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- 229940041181 antineoplastic drug Drugs 0.000 description 2
- 239000002774 b raf kinase inhibitor Substances 0.000 description 2
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- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- 229940102297 B-raf kinase inhibitor Drugs 0.000 description 1
- SIKJAQJRHWYJAI-UHFFFAOYSA-N Indole Chemical compound C1=CC=C2NC=CC2=C1 SIKJAQJRHWYJAI-UHFFFAOYSA-N 0.000 description 1
- 102000004856 Lectins Human genes 0.000 description 1
- 108090001090 Lectins Proteins 0.000 description 1
- 229910020700 Na3VO4 Inorganic materials 0.000 description 1
- 102000052575 Proto-Oncogene Human genes 0.000 description 1
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- 230000003197 catalytic effect Effects 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
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- 230000030944 contact inhibition Effects 0.000 description 1
- 230000030609 dephosphorylation Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 125000005594 diketone group Chemical group 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
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- 238000002560 therapeutic procedure Methods 0.000 description 1
- IHIXIJGXTJIKRB-UHFFFAOYSA-N trisodium vanadate Chemical compound [Na+].[Na+].[Na+].[O-][V]([O-])([O-])=O IHIXIJGXTJIKRB-UHFFFAOYSA-N 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
- C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
- C07D209/04—Indoles; Hydrogenated indoles
- C07D209/30—Indoles; Hydrogenated indoles with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, directly attached to carbon atoms of the hetero ring
- C07D209/32—Oxygen atoms
- C07D209/38—Oxygen atoms in positions 2 and 3, e.g. isatin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmacology & Pharmacy (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Indole Compounds (AREA)
Abstract
The invention relates to the field of drug synthesis, and discloses a preparation method and application of compounds BIA-01 and BIA-02 with CDC25B inhibitory activity, aiming at the problems of insufficient accuracy of drug-targeted therapy, insufficient activity of substituent groups and poor binding capacity with action sites in the prior art, wherein the preparation method comprises the following steps: i. 1a- (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide; ii. Synthesizing 2 a-6-bromoindoline-2, 3-diketone; iii, synthesizing 3a, 3 b-chloracetanilide derivatives; and (3) synthesizing the iiii, the BIA-01 and the BIA-02-6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide. The compounds BIA-01 and BIA-02 prepared by the invention have strong antitumor activity and low side effect, and the composition has stronger inhibition effect on tumors.
Description
Technical Field
The invention relates to the field of drug synthesis, in particular to a preparation method and application of compounds BIA-01 and BIA-02 with CDC25B inhibitory activity.
Background
Cancer is counted by the world health organization (WTO) as one of the major diseases causing human death, and 1300 million people are predicted to die due to cancer by 2030. Cancer cells are difficult to control effectively because of their characteristics such as unlimited proliferation, loss of contact inhibition, reduced adhesion among cancer cells, susceptibility to agglutination by lectins, and reduced wall adhesion. Currently, research into anticancer drugs has entered the molecular level. Recent researches show that cell division cycle 25 phosphatase B (CDC25B) and Protein Tyrosine Kinase (PTK) have a certain relationship and become a new target for developing antitumor drugs. The research shows that the diketone indoline and the derivative thereof have various biological activities and are an active intermediate for drug synthesis, and the diketone indoline derivative has a certain inhibition effect on tumors.
Patent No. CN201210209023.4, with the patent name of "2-substituted-5-phenyl furans compound, its preparation method, pharmaceutical composition and its use", provides a 2-substituted-5-phenyl furans compound represented by the following general formula i, or its cis-trans isomer, or its pharmaceutically acceptable salt, ester, prodrug or solvate, its preparation method, pharmaceutical composition and its use in preparing B-Raf kinase inhibitor. The compounds or the pharmaceutical compositions thereof can be used as high-efficiency B-Raf kinase inhibitors for treating tumor diseases.
The invention has the disadvantages of insufficient target treatment precision, insufficient activity of substituent groups, poor binding capacity with action parts, longer synthetic route, higher cost, high reaction control requirement in the synthetic process and more side reactions.
Disclosure of Invention
The invention is in order to overcome the problem that the precision of the prior art drug targeting therapy is insufficient, the substituent group activity is insufficient and the binding capacity with the action site is poor, provide the preparation method and application with compound BIA-01, BIA-02 of CDC25B inhibitory activity, prepare and get compound BIA-01, BIA-02 with strong antitumor activity, low side effect, this compound is a pharmaceutical synthesis active intermediate, have stronger inhibitory action to tumor; and has excellent multiple biological activities, wherein the amide group contained in the molecular formula is a dominant group, and can well interact with a treatment site in the biological macromolecule.
In order to achieve the purpose, the invention adopts the following technical scheme:
a method for preparing compounds BIA-01 and BIA-02 with CDC25B inhibitory activity, comprising the following steps:
i. 1a- (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide;
ii. Synthesizing 2 a-6-bromoindoline-2, 3-diketone;
iii, synthesizing 3a, 3 b-chloracetanilide derivatives;
synthesizing iiii, BIA-01, BIA-02-6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide;
the specific synthetic route is as follows:
preferably, in the step i, the charging ratio of the raw materials based on 5mmol of m-bromoaniline is as follows: 5mmol of m-bromoaniline; anhydrous Na2SO429.5-30.5 mmol; 15-20mL of water; chloral hydrate, 5.8-6.2 mmol; a mixed solution of concentrated hydrochloric acid and ammonia water; a solution prepared from hydroxylamine hydrochloride and distilled water;
the specific process of the step i is as follows:
weighing 29.5-30.5mmol of anhydrous Na2SO4Placing the mixture into a container, adding 15-20mL of water, stirring by magnetic force, slowly heating to 50-55 ℃, and stirring until the temperature is Na2SO4And (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide is obtained after complete dissolution, cooling to room temperature, sequentially and slowly adding 5.8-6.2mmol of chloral hydrate, 5mmol of m-bromoaniline, mixed solution of concentrated hydrochloric acid and ammonia water, hydroxylamine hydrochloride and distilled water to prepare solution, heating to 60-70 ℃ under stirring, reacting for 1.5-2h, generating beige precipitate in the heating reaction process, cooling after the reaction is finished, performing suction filtration, and performing infrared drying.
Preferably, the volume ratio of the concentrated hydrochloric acid to the ammonia water in the mixed solution of the concentrated hydrochloric acid and the ammonia water is 0.28-0.3: 2.4-2.6.
Preferably, the solution is prepared by dissolving 15.8-16.2mmol of hydroxylamine hydrochloride in 5-5.4mL of distilled water.
M-bromoaniline and chloral hydrate undergo nucleophilic addition reaction, then an intermediate is obtained, and nucleophilic addition elimination reaction is carried out on the intermediate and hydroxylamine hydrochloride, so that (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide is obtained.
Preferably, in the step ii, the raw materials are fed in the following ratio in 4mmol of (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide: concentrated sulfuric acid, 15-16 mL; (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide, 4 mmol; crushing ice; water;
the specific process of the step ii is as follows: weighing 15-16mL of concentrated sulfuric acid, placing in a container, and keeping the temperature at 45-50 ℃; heating to 60-70 ℃, slowly adding 4mmol (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide, after the addition, heating to 80-85 ℃, reacting for 10-15 min, cooling to room temperature, slowly pouring the reaction liquid into crushed ice with the volume 9-11 times that of the reaction liquid, uniformly stirring, standing for about 30-40min, performing suction filtration to obtain a filter cake, washing the filter cake with water, and drying to obtain 6-bromoindoline-2, 3-dione.
Preferably, in step iii, the charge ratio of each raw material based on 10.7mmol of substituted aniline is as follows: 48-52mL of mixed solution of glacial acetic acid and saturated sodium acetate solution; chloroacetyl chloride, 4.2-4.6 mmol; ice water; ethanol;
the specific process of the step iii is as follows: weighing 10.7mmol of substituted aniline, dissolving the substituted aniline in a mixed solution of 24-26mL of glacial acetic acid and a saturated 24-26mL of sodium acetate solution, placing the mixed solution in an ice bath for cooling, slowly dropwise adding 4.2-4.6mmol of chloroacetyl chloride, reacting for 25-35min in the ice bath, after the reaction is finished, performing suction filtration to obtain a filter cake, washing the filter cake with ice water, drying to obtain a crude product, and recrystallizing the crude product with ethanol to obtain different substituted N-chloroacetanilides.
Substituted aniline as nucleophilic reagent and chloroacetyl chloride are nucleophilic added to eliminate reaction under the action of sodium acetate to obtain N-chloroacetanilide.
Preferably, in the step iiii, the raw material feeding ratio is as follows: 6-bromoindoline-2, 3-dione; different substituted N-chloroacetanilides; DMF, 18-22 mL; anhydrous K2CO3, 3.5-3.7 mmol; potassium iodide, 2.8-3.2 mmol; distilled water; dilute hydrochloric acid, 0.8-1.2N; ethanol;
the specific process of the step iiii is as follows: under the ice bath condition, adding 18-22mL of DMF (dimethyl formamide) into 6-bromoindoline-2, 3-diketone, stirring for dissolving, adding 3.5-3.7mmol of anhydrous K2CO3, heating to room temperature, continuing stirring for 1-1.5h, then adding 10.7mmol of substituted N-chloroacetanilide and 3.5-3.7mmol of potassium iodide, heating to 75-80 ℃, cooling to room temperature after complete reaction, adding distilled water with the volume being 6-7 times of that of the reaction solution for dilution, dropwise adding 0.8-1.2N of dilute hydrochloric acid to adjust the pH value of the solution to 3-4, stirring uniformly, carrying out suction filtration, washing with ice water, drying to obtain a crude product, and recrystallizing the crude product in ethanol to obtain the finished product.
6-Bromoindoline-2, 3-dione as nucleophile in K2CO3The target compound is obtained by nucleophilic substitution reaction with N-chloroacetanilide under catalysis.
The compounds BIA-01 and BIA-02 with CDC25B inhibitory activity are applied to the preparation of tumor activity inhibition drugs.
Preferably, the medicament is an injectable medicament.
Therefore, the invention has the following beneficial effects:
(1) the prepared compounds BIA-01 and BIA-02 have strong antitumor activity and low side effect, and the compound is a drug synthesis active intermediate and has stronger inhibition effect on tumors;
(2) has excellent bioactivity, wherein the amide group contained in the molecular formula is a dominant group, and can well interact with a treatment site in the biological macromolecule
(3) The invention takes the m-bromoaniline and the substituted aniline which are easy to obtain and low in price as raw materials, and the compounds BIA-01 and BIA-02 are obtained through simple and efficient synthesis, and the medicine has the advantages of short synthetic route, low cost and high synthesis efficiency;
(4) the synthesis process has good controllability, is easy for actual large-scale industrial mass production, and the prepared product has high purity.
Detailed Description
The invention is further described with reference to specific embodiments.
Example 1
A method for preparing compounds BIA-01 and BIA-02 with CDC25B inhibitory activity, comprising the following steps:
i. 1a- (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide;
ii. Synthesizing 2 a-6-bromoindoline-2, 3-diketone;
iii, synthesizing 3a, 3 b-chloracetanilide derivatives;
synthesizing iiii, BIA-01, BIA-02-6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide;
the specific synthetic route is as follows:
the specific process of the step i is as follows:
weighing 30mmol anhydrous Na by a precision analytical balance2SO4Placing in 50mL round bottom flask, adding 15mL water, placing on constant temperature magnetic stirrer, slowly heating to 53 deg.C, stirring to Na2SO4After complete dissolution, cooling to room temperature, slowly adding a solution prepared from 6mmol of chloral hydrate, 5mmol of m-bromoaniline, 0.58mL of concentrated hydrochloric acid and 5mL of ammonia water and 16mmol of hydroxyamine hydrochloride and 5.2mL of distilled water in turn, heating to 65 ℃ under stirring for reaction for 1.8h, generating beige precipitate in the heating reaction process, and monitoring the reaction by TLC. After the reaction is finished, cooling, filtering, and infrared drying to obtain beige crystals (1 a).
The specific process of the step ii is as follows:
15.5mL of concentrated sulfuric acid was weighed into a 50mL round-bottom flask, heated to 48 ℃ and slowly added with 4mmol of (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide (1a), and the temperature was further raised and maintained at 65 ℃. After the addition, heating to 83 ℃, monitoring the reaction by TLC, reacting completely after about 13min, cooling to room temperature, slowly pouring the reaction liquid into crushed ice with the volume about 10 times of that of the reaction liquid, stirring uniformly, standing for about 35min, performing suction filtration, washing a filter cake for several times by water until almost all sulfuric acid is washed off, and drying to obtain a dark orange red crystal 6-bromoindoline-2, 3-diketone (2 a).
The specific process of the step iii is as follows:
10.7mmol of substituted aniline was measured and dissolved in a mixed solution of 25mL of glacial acetic acid and 25mL of saturated sodium acetate solution, and the solution was cooled in an ice bath, and then 4.4mmol of chloroacetyl chloride was slowly added dropwise and reacted for 30min in the ice bath. After the reaction is finished, carrying out suction filtration, washing a filter cake for a plurality of times by using ice water, drying to obtain a crude product, and recrystallizing the crude product by using ethanol to obtain the N-chloroacetanilides (3a and 3b) with different substitutions.
The specific process of the step iiii is as follows:
under ice bath conditions, 6-bromoindoline-2, 3-dione (2a) was added to a 50mL round-bottomed flask, 20mL of DMF was added, and after stirring and dissolution, 3.6mmol of anhydrous K was added2CO3And naturally heating to room temperature and then continuously stirring for 1.3 h. 10.7mmol of substituted chloroacetanilides (3a, 3b) and 3.6mmol of potassium iodide were then added, the reaction was warmed to 78 ℃ and monitored by TLC until the reaction was complete. Cooling to room temperature, adding distilled water with the volume about 6.5 times of that of the reaction liquid to dilute the reaction system, dropwise adding 1N diluted hydrochloric acid to adjust the pH value of the solution to 3.5, uniformly stirring, carrying out suction filtration, washing for a plurality of times by ice water, drying to obtain a crude product, and recrystallizing the crude product in methanol or ethanol to obtain the product 6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide derivatives (BIA-01, BIA-02).
I. The relevant physicochemical data of the product obtained from QNL-Chalcone are shown in Table 1:
TABLE 1 BIA-01, BIA-02 related physicochemical data
| Compound (I) | Substituent group | Melting range | Yield (%) | Chemical formula (II) |
| BIA-01 | 2,4-Cl2 | 246.2-247.6 | 67 | C16H9BrCl2N2O3 |
| BIA-02 | 2,6-Cl2 | 267.4-269.5 | 71 | C16H9BrCl2N2O3 |
BIA-01 spectral data
IR(KBr)cm-1:3501,1733,1649,1252.1H-NMR(CDCl3,300MHz):δ9.90(1H,s,-NH),7.00–7.51(3H,m,-C6H3),7.65–7.80(3H,m,-C6H3),4.82(2H,s,-CH2).13C-NMR(CDCl3,75MHz):183.5,170.1,156.7,149.3,143.3,141.6,137.9,134.0,133.1,132.1,130.5,126.4,125.4,120.5,114.6,48.0.MS(M+1)m/z:427,429.
BIA-02 spectral data
IR(KBr)cm-1:3499,1731,1649,1252.1H-NMR(CDCl3,300MHz):δ11.24(1H,s,-NH),6.69–7.41(3H,m,-C6H3),7.44–8.10(3H,m,-C6H3),3.39(2H,s,-CH2).13C-NMR(CDCl3,75MHz):184.5,170.0,156.6,149.2,143.3,141.6,137.9,134.1,133.1,132.2,126.5,125.3,120.4,115.1,48.2.MS(M+1)m/z:427,429.
Determination of phosphatase inhibitory Activity
1. Method for measuring biological activity of CDC25B
CDC25B has proto-oncogene properties, which are overexpressed in a variety of human tumor cells, in addition to promoting mitosis of the cells. Whereas the enzymatic activity of the CDC25B catalytic domain was determined by monitoring the ompp dephosphorylation reaction. The dephosphorylated product of OMFP, 3-O-methyl fluorescein (OMF), was detected using a multi-label microplate detector and was detectable at an excitation wavelength of 485 nm/emission wavelength of 535 nm.
Test compound and positive control drug (Na)3VO4) The solution was dissolved in DMSO, prepared into solutions of various concentrations, and added to a 96-well plate at 2 μ L per well, and Tris-HCl buffer (50mmol/L), NaCl solution (50mmol/L), OMFP (5 μmol/L), CDC25B recombinant protein (20nmol/L), 1% glycerol, and DTT (1mmol/L) were added, and the total reaction volume was 100 μ L, and pH was 8.0. The enzyme activity was continuously monitored using the time-linear region of the enzyme reaction kinetics curve to determine the rate of initiation of dephosphorylation.
2. Inhibitory Activity of Compounds
Inhibition of enzyme by test compound and positive control drug and IC thereof50The value is calculated by the following formula:
inhibition ratio (%) < 100/(1+ [ IC ]50/[I]]k)
In the formula V0The initial rate of blank group (DMSO), Va the initial rate of administration group, [ I ]]K is the Hill coefficient for inhibitor concentration. Firstly, calculating to obtain the inhibition rate of tested compound to enzyme, then using inhibition rate to make graph of compound concentration and using formula to make fitting so as to obtain IC50The value is obtained.
Results of the experiment
The results of the inhibitory activities of the compounds BIA-01 and BIA-02 on CDC25B are shown in Table 2. BIA-01 and BIA-02 have good inhibitory activity to CDC25B, and IC thereof50The values were 2.14. mu.g/mL and 2.09. mu.g/mL, respectively.
TABLE 2 inhibitory Activity of Compounds BIA-01, BIA-02 on CDC25B
| Compounds | CDC 25B[IC50(μg/mL)] |
| BIA-01 | 2.14±0.36 |
| BIA-02 | 2.09±0.59 |
| Na3VO4 | 3.03±0.41 |
And (4) supplementary notes: determination of IC by regression analysis50Values, mean. + -. standard deviation of 3 groups of data
To indicate.
Conclusion IIII
The research uses 3-bromoaniline as a starting material, adopts a new medicine design method, and obtains 6-bromo-2, 3-dioxoindoline-1-N-substituted phenyl acetamide derivatives (BIA-01 and BIA-02) through a series of reactions such as Sandmeyer reaction, ring closure reaction, nucleophilic substitution and the like; and the compounds BIA-01 and BIA-02 have better inhibitory activity to CDC 25B; IC thereof50The values are respectively 2.14 mu g/mL and 2.09 mu g/mL, and the anti-tumor targeted therapeutic property is better.
Example 2
The difference from the example 1 is that the specific process of the step i is as follows:
weighing 29.5-30.5mmol anhydrous Na by a precision analytical balance2SO4Placing in 50mL round bottom flask, adding 15mL water, placing on constant temperature magnetic stirrer, slowly heating to 50-55 deg.C, stirring to Na2SO4After complete dissolution, the mixture is cooled to room temperature, and then 5.8 to 6.2mmol of chloral hydrate, 5mmol of m-bromoaniline, 0.56mL of concentrated hydrochloric acid and 4.8mL of ammonia water are slowly added in turn, 15.8mmol of hydroxylamine hydrochloride and 5mL of distilled water are added in turnThe prepared solution is heated to 60 ℃ under stirring to react for 1.5h, beige precipitate is generated in the heating reaction process, and the reaction is monitored by TLC. After the reaction is finished, cooling, filtering, and infrared drying to obtain beige crystals (1 a).
The specific process of the step ii is as follows:
15mL of concentrated sulfuric acid was weighed into a 50mL round-bottom flask, heated to 45 ℃ and 4mmol of (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide (1a) was slowly added, and the temperature was further raised and maintained at 60 ℃. After the addition, heating to 80 ℃, monitoring the reaction by TLC, reacting completely after about 10min, cooling to room temperature, slowly pouring the reaction liquid into crushed ice with the volume about 9 times that of the reaction liquid, stirring uniformly, standing for about 30min, performing suction filtration, washing a filter cake for several times by water until almost all sulfuric acid is washed off, and drying to obtain a dark orange red crystal 6-bromoindoline-2, 3-diketone (2 a).
The specific process of the step iii is as follows:
10.7mmol of substituted aniline was measured and dissolved in a mixed solution of 24mL of glacial acetic acid and 24mL of saturated sodium acetate solution, and the solution was cooled in an ice bath, and then 4.2mmol of chloroacetyl chloride was slowly added dropwise and reacted for 25min in the ice bath. After the reaction is finished, carrying out suction filtration, washing a filter cake for a plurality of times by using ice water, drying to obtain a crude product, and recrystallizing the crude product by using ethanol to obtain the N-chloroacetanilides (3a and 3b) with different substitutions.
The specific process of the step iiii is as follows:
under ice bath conditions, 6-bromoindoline-2, 3-dione (2a) was added to a 50mL round-bottomed flask, 18mL of DMF was added, and after dissolution by stirring, 3.5mmol of anhydrous K was added2CO3And naturally heating to room temperature and then continuously stirring for 1 h. 10.7mmol of substituted chloroacetanilides (3a, 3b) and 3.5mmol of potassium iodide were then added, the reaction was warmed to 75 ℃ and monitored by TLC until the reaction was complete. Cooling to room temperature, adding distilled water with the volume about 6 times of that of the reaction liquid to dilute the reaction system, dropwise adding 0.8N diluted hydrochloric acid to adjust the pH value of the solution to 3, stirring uniformly, carrying out suction filtration, washing for a plurality of times by ice water, drying to obtain a crude product, and recrystallizing the crude product in methanol or ethanol to obtain the product 6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide derivatives (BIA-01, BIA-02).
Example 3
The difference from the example 1 is that the specific process of the step i is as follows:
weighing 30.5mmol anhydrous Na by a precision analytical balance2SO4Placing in 50mL round bottom flask, adding 15mL water, placing on constant temperature magnetic stirrer, slowly heating to 55 deg.C, stirring to Na2SO4After complete dissolution, cooling to room temperature, slowly adding a solution prepared from 6.2mmol of chloral hydrate, 5mmol of m-bromoaniline, 0.6mL of concentrated hydrochloric acid and 5.2mL of ammonia water and 16.2mmol of hydroxylamine hydrochloride and 5.4mL of distilled water in turn, heating to 70 ℃ under stirring for reaction for 2h, generating beige precipitate in the heating reaction process, and monitoring the reaction by TLC. After the reaction is finished, cooling, filtering, and infrared drying to obtain beige crystals (1 a).
The specific process of the step ii is as follows:
concentrated sulfuric acid 15-16mL was weighed into a 50mL round bottom flask, heated to 50 ℃ and 4mmol of (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide (1a) was slowly added, and the temperature was further raised and maintained at 70 ℃. After the addition, heating to 85 ℃, monitoring the reaction by TLC, reacting completely after about 15min, cooling to room temperature, slowly pouring the reaction liquid into crushed ice with the volume about 11 times of that of the reaction liquid, stirring uniformly, standing for about 40min, performing suction filtration, washing a filter cake for several times by water until almost all sulfuric acid is washed off, and drying to obtain a dark orange red crystal 6-bromoindoline-2, 3-diketone (2 a).
The specific process of the step iii is as follows:
10.7mmol of substituted aniline was measured and dissolved in a mixed solution of 26mL of glacial acetic acid and 26mL of saturated sodium acetate solution, and the solution was cooled in an ice bath, and then 4.6mmol of chloroacetyl chloride was slowly added dropwise and reacted for 35min in the ice bath. After the reaction is finished, carrying out suction filtration, washing a filter cake for a plurality of times by using ice water, drying to obtain a crude product, and recrystallizing the crude product by using ethanol to obtain the N-chloroacetanilides (3a and 3b) with different substitutions.
The specific process of the step iiii is as follows:
under ice bath conditions, 6-bromoindoline-2, 3-dione (2a) was added to a 50mL round-bottomed flask, 22mL of DMF was added, and after stirring and dissolution, 3.7mmol of anhydrous K was added2CO3Naturally raising the temperature to the roomStirring was continued for 1.5h after warming. 10.7mmol of substituted chloroacetanilides (3a, 3b) and 3.7mmol of potassium iodide were then added, the reaction was warmed to 80 ℃ and monitored by TLC until the reaction was complete. Cooling to room temperature, adding distilled water with the volume about 7 times that of the reaction liquid to dilute the reaction system, dropwise adding 1.2N diluted hydrochloric acid to adjust the pH value of the solution to 4, stirring uniformly, carrying out suction filtration, washing for a plurality of times by ice water, drying to obtain a crude product, and recrystallizing the crude product in methanol or ethanol to obtain the product 6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide derivatives (BIA-01, BIA-02).
Example 4
The difference from the example 1 is that the specific process of the step i is as follows:
29.8mmol of anhydrous Na are weighed by a precision analytical balance2SO4Placing in 50mL round bottom flask, adding 15mL water, placing on constant temperature magnetic stirrer, slowly heating to 51 deg.C, stirring to Na2SO4After complete dissolution, the mixture is cooled to room temperature, a solution prepared by sequentially and slowly adding 5.9mmol of chloral hydrate, 5mmol of m-bromoaniline, 0.57mL of concentrated hydrochloric acid and 4.9mL of ammonia water and a solution prepared by 15.9mmol of hydroxylamine hydrochloride and 5.1mL of distilled water is heated to 63 ℃ under stirring for reaction for 1.6h, beige precipitate is generated in the heating reaction process, and the reaction is monitored by TLC. After the reaction is finished, cooling, filtering, and infrared drying to obtain beige crystals (1 a).
The specific process of the step ii is as follows:
15.3mL of concentrated sulfuric acid was weighed into a 50mL round-bottom flask, heated to 46 ℃ and slowly added with 4mmol of (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide (1a), and the temperature was further raised and maintained at 63 ℃. After the addition, heating to 81 ℃, monitoring the reaction by TLC, reacting completely after about 11min, cooling to room temperature, slowly pouring the reaction liquid into crushed ice with the volume about 9.5 times of that of the reaction liquid, stirring uniformly, standing for about 33min, performing suction filtration, washing a filter cake for several times by water until almost all sulfuric acid is washed off, and drying to obtain a dark orange red crystal 6-bromoindoline-2, 3-diketone (2 a).
The specific process of the step iii is as follows:
10.7mmol of substituted aniline was measured and dissolved in a mixed solution of 24.5mL of glacial acetic acid and 24.5mL of saturated sodium acetate solution, and the solution was cooled in an ice bath, and then 4.3mmol of chloroacetyl chloride was slowly added dropwise and reacted for 28min in the ice bath. After the reaction is finished, carrying out suction filtration, washing a filter cake for a plurality of times by using ice water, drying to obtain a crude product, and recrystallizing the crude product by using ethanol to obtain the N-chloroacetanilides (3a and 3b) with different substitutions.
The specific process of the step iiii is as follows:
under ice bath conditions, 6-bromoindoline-2, 3-dione (2a) was added to a 50mL round-bottomed flask, 18.5mL of DMF was added, and after dissolution by stirring, 3.55mmol of anhydrous K was added2CO3And naturally heating to room temperature and then continuously stirring for 1.1 h. 10.7mmol of substituted chloroacetanilides (3a, 3b) and 3.55mmol of potassium iodide were then added, the reaction was warmed to 76 ℃ and monitored by TLC until the reaction was complete. Cooling to room temperature, adding distilled water with the volume about 6.3 times of that of the reaction solution to dilute the reaction system, dropwise adding 0.85N diluted hydrochloric acid to adjust the pH value of the solution to 3.3, stirring uniformly, carrying out suction filtration, washing for a plurality of times by ice water, drying to obtain a crude product, and recrystallizing the crude product in methanol or ethanol to obtain the product 6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide derivative (BIA-01, BIA-02).
Example 5
The difference from the example 1 is that the specific process of the step i is as follows:
weighing 30.2mmol anhydrous Na by a precision analytical balance2SO4Placing in 50mL round bottom flask, adding 15mL water, placing on constant temperature magnetic stirrer, slowly heating to 54 deg.C, stirring to Na2SO4After complete dissolution, the mixture is cooled to room temperature, a solution prepared by sequentially and slowly adding 6.1mmol of chloral hydrate, 5mmol of m-bromoaniline, 0.59mL of concentrated hydrochloric acid and 5.1mL of ammonia water and a solution prepared by 16.1mmol of hydroxylamine hydrochloride and 5.3mL of distilled water is heated to 68 ℃ under stirring for reaction for 1.9h, beige precipitate is generated in the heating reaction process, and the reaction is monitored by TLC. After the reaction is finished, cooling, filtering, and infrared drying to obtain beige crystals (1 a).
The specific process of the step ii is as follows:
15.8mL of concentrated sulfuric acid was weighed into a 50mL round-bottom flask, heated to 49 ℃ and 4mmol of (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide (1a) was slowly added, and the temperature was further raised and maintained at 68 ℃. After the addition, heating to 84 ℃, monitoring the reaction by TLC, reacting completely after about 14min, cooling to room temperature, slowly pouring the reaction liquid into crushed ice with the volume about 10.5 times of that of the reaction liquid, stirring uniformly, standing for about 38min, performing suction filtration, washing a filter cake for several times by water until almost all sulfuric acid is washed off, and drying to obtain a dark orange red crystal 6-bromoindoline-2, 3-diketone (2 a).
The specific process of the step iii is as follows:
10.7mmol of substituted aniline was measured and dissolved in a mixed solution of 25.5mL of glacial acetic acid and 25.5mL of saturated sodium acetate solution, and the solution was cooled in an ice bath, and then 4.5mmol of chloroacetyl chloride was slowly added dropwise and reacted for 33min in the ice bath. After the reaction is finished, carrying out suction filtration, washing a filter cake for a plurality of times by using ice water, drying to obtain a crude product, and recrystallizing the crude product by using ethanol to obtain the N-chloroacetanilides (3a and 3b) with different substitutions.
The specific process of the step iiii is as follows:
under ice bath conditions, 6-bromoindoline-2, 3-dione (2a) was added to a 50mL round-bottomed flask, 21mL of DMF was added, and after dissolution by stirring, 3.65mmol of anhydrous K was added2CO3And naturally heating to room temperature and then continuously stirring for 1.4 h. 10.7mmol of substituted chloroacetanilides (3a, 3b) and 3.65mmol of potassium iodide were then added, the reaction was warmed to 79 ℃ and monitored by TLC until the reaction was complete. Cooling to room temperature, adding distilled water with the volume about 6.8 times of that of the reaction solution to dilute the reaction system, dropwise adding 1.1N diluted hydrochloric acid to adjust the pH value of the solution to 3.8, stirring uniformly, carrying out suction filtration, washing for a plurality of times by ice water, drying to obtain a crude product, and recrystallizing the crude product in methanol or ethanol to obtain the product 6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide derivative (BIA-01, BIA-02).
Under the parameter conditions of examples 1-5, the prepared novel compounds BIA-01 and BIA-02 have better inhibitory activity to CDC25B and have more accurate targeted therapy characteristics to tumors.
From the data relating to examples 1 to 5 above, it is clear that the above requirements can be satisfied in all respects only by a protocol within the scope of the claims of the present invention, and that an optimized protocol can be obtained, and that novel compounds BIA-01, BIA-02 having CDC25B inhibitory activity with optimum activity can be obtained. The change of the mixture ratio, the replacement/addition/subtraction of raw materials or the change of the feeding sequence can bring corresponding negative effects.
The raw materials and equipment used in the invention are common raw materials and equipment in the field if not specified; the methods used in the present invention are conventional in the art unless otherwise specified.
The above description is only a preferred embodiment of the present invention, and is not intended to limit the present invention, and all simple modifications, alterations and equivalents of the above embodiments according to the technical spirit of the present invention are still within the protection scope of the technical solution of the present invention.
Claims (9)
1. A method for preparing compounds BIA-01 and BIA-02 with CDC25B inhibitory activity, which is characterized by comprising the following steps:
i. 1a- (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide;
ii. Synthesizing 2 a-6-bromoindoline-2, 3-diketone;
iii, synthesizing 3a, 3 b-chloracetanilide derivatives;
synthesizing iiii, BIA-01, BIA-02-6-bromo-2, 3-dioxoindoline-1-N-substituted phenylacetamide;
the specific synthetic route is as follows:
2. the method for preparing the compounds BIA-01 and BIA-02 having the CDC25B inhibitory activity according to claim 1, wherein in step i, the raw material charge ratio based on 5mmol of m-bromoaniline is as follows: 5mmol of m-bromoaniline; anhydrous Na2SO429.5-30.5 mmol; 15-20mL of water; chloral hydrate, 5.8-6.2 mmol; a mixed solution of concentrated hydrochloric acid and ammonia water; a solution prepared from hydroxylamine hydrochloride and distilled water;
the specific process of the step i is as follows:
weighing 29.5-30.5mmol of anhydrous Na2SO4Placing the mixture into a container, adding 15-20mL of water, stirring by magnetic force, slowly heating to 50-55 ℃, and stirring until the temperature is Na2SO4And (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide is obtained after complete dissolution, cooling to room temperature, sequentially and slowly adding 5.8-6.2mmol of chloral hydrate, 5mmol of m-bromoaniline, mixed solution of concentrated hydrochloric acid and ammonia water, hydroxylamine hydrochloride and distilled water to prepare solution, heating to 60-70 ℃ under stirring, reacting for 1.5-2h, generating beige precipitate in the heating reaction process, cooling after the reaction is finished, performing suction filtration, and performing infrared drying.
3. The method for producing the compounds BIA-01 and BIA-02 having the inhibitory activity of CDC25B as set forth in claim 2, wherein the volume ratio of the concentrated hydrochloric acid to the aqueous ammonia in the mixed solution of the concentrated hydrochloric acid and the aqueous ammonia is 0.28 to 0.3:2.4 to 2.6.
4. The method for preparing the compounds BIA-01 and BIA-02 having the CDC25B inhibitory activity according to claim 2, wherein said solution is prepared by dissolving 15.8 to 16.2mmol of hydroxylamine hydrochloride in 5 to 5.4mL of distilled water.
5. The process according to claim 1 for preparing the compounds BIA-01, BIA-02 having CDC25B inhibitory activity, wherein in step ii, (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide in 4mmol, the following ratios of the starting materials are used: concentrated sulfuric acid, 15-16 mL; (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide, 4 mmol; crushing ice; water; the specific process of the step ii is as follows: weighing 15-16mL of concentrated sulfuric acid, placing in a container, and keeping the temperature at 45-50 ℃; heating to 60-70 ℃, slowly adding 4mmol (E) -N- (3-bromophenyl) -2- (hydroxyimino) acetamide, after the addition, heating to 80-85 ℃, reacting for 10-15 min, cooling to room temperature, slowly pouring the reaction liquid into crushed ice with the volume 9-11 times that of the reaction liquid, uniformly stirring, standing for about 30-40min, performing suction filtration to obtain a filter cake, washing the filter cake with water, and drying to obtain 6-bromoindoline-2, 3-dione.
6. The process according to claim 1 for the preparation of the compounds BIA-01, BIA-02 having CDC25B inhibitory activity, wherein in step iii, the charge ratios of the substituted anilines in terms of 10.7mmol are as follows: 48-52mL of mixed solution of glacial acetic acid and saturated sodium acetate solution; chloroacetyl chloride, 4.2-4.6 mmol; ice water; ethanol;
the specific process of the step iii is as follows: weighing 10.7mmol of substituted aniline, dissolving the substituted aniline in a mixed solution of 24-26mL of glacial acetic acid and a saturated 24-26mL of sodium acetate solution, placing the mixed solution in an ice bath for cooling, slowly dropwise adding 4.2-4.6mmol of chloroacetyl chloride, reacting for 25-35min in the ice bath, after the reaction is finished, performing suction filtration to obtain a filter cake, washing the filter cake with ice water, drying to obtain a crude product, and recrystallizing the crude product with ethanol to obtain different substituted N-chloroacetanilides.
7. The process according to claim 1, wherein in step iiii, the raw material ratios of the compounds BIA-01 and BIA-02 having CDC25B inhibitory activity are as follows: 6-bromoindoline-2, 3-dione; different substituted N-chloroacetanilides; DMF, 18-22 mL; anhydrous K2CO33.5-3.7 mmol; potassium iodide, 2.8-3.2 mmol; distilled water; dilute hydrochloric acid, 0.8-1.2N; ethanol;
the specific process of the step iiii is as follows: under the ice bath condition, 18-22mL of DMF is added into 6-bromoindoline-2, 3-diketone, stirred and dissolved, and then 3.5-3.7mmol of anhydrous K is added2CO3Heating to room temperature, continuing stirring for 1-1.5h, adding 10.7mmol of substituted N-chloroacetanilide and 3.5-3.7mmol of potassium iodide, heating to 75-80 ℃, cooling to room temperature after complete reaction, adding distilled water with the volume of about 6-7 times of that of the reaction solution for dilution, dropwise adding 0.8-1.2N of dilute hydrochloric acid to adjust the pH value of the solution to 3-4, uniformly stirring, performing suction filtration, washing with ice water, drying to obtain a crude product, and recrystallizing the crude product in ethanol to obtain the finished product.
8. Use of the compounds BIA-01, BIA-02 having CDC25B inhibitory activity as defined in any one of claims 1 to 7 in the manufacture of a medicament for inhibiting tumor activity.
9. The use of claim 8, wherein the medicament is an injectable medicament.
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| CN103509009A (en) * | 2012-06-21 | 2014-01-15 | 中国科学院上海药物研究所 | 2-substituted-5-phenyl furan compound, and preparation method, pharmaceutical composition and application thereof |
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